CORD-19-Sentences  

CORD-19:29b6131c0d572d383cbc6774b12e4471aa612b07 JSONTXT 8 Projects

Annnotations TAB TSV DIC JSON TextAE-old TextAE

Id Subject Object Predicate Lexical cue
TextSentencer_T1 0-116 Sentence denotes Vaccine Efficacy in Senescent Mice Challenged with Recombinant SARS-CoV Bearing Epidemic and Zoonotic Spike Variants
T34712 0-116 Sentence denotes Vaccine Efficacy in Senescent Mice Challenged with Recombinant SARS-CoV Bearing Epidemic and Zoonotic Spike Variants
TextSentencer_T2 118-126 Sentence denotes Abstract
T13845 118-126 Sentence denotes Abstract
TextSentencer_T3 127-136 Sentence denotes Citation:
T98034 127-136 Sentence denotes Citation:
TextSentencer_T4 137-316 Sentence denotes Deming D, Sheahan T, Heise M, Yount B, Davis N, et al. (2006) Vaccine efficacy in senescent mice challenged with recombinant SARS-CoV bearing epidemic and zoonotic spike variants.
T3662 137-316 Sentence denotes Deming D, Sheahan T, Heise M, Yount B, Davis N, et al. (2006) Vaccine efficacy in senescent mice challenged with recombinant SARS-CoV bearing epidemic and zoonotic spike variants.
TextSentencer_T5 317-338 Sentence denotes PLoS Med 3(12): e525.
T23294 317-338 Sentence denotes PLoS Med 3(12): e525.
TextSentencer_T6 340-560 Sentence denotes In 2003, severe acute respiratory syndrome coronavirus (SARS-CoV) was identified as the etiological agent of severe acute respiratory syndrome, a disease characterized by severe pneumonia that sometimes results in death.
T7513 340-560 Sentence denotes In 2003, severe acute respiratory syndrome coronavirus (SARS-CoV) was identified as the etiological agent of severe acute respiratory syndrome, a disease characterized by severe pneumonia that sometimes results in death.
TextSentencer_T7 561-746 Sentence denotes SARS-CoV is a zoonotic virus that crossed the species barrier, most likely originating from bats or from other species including civets, raccoon dogs, domestic cats, swine, and rodents.
T91237 561-746 Sentence denotes SARS-CoV is a zoonotic virus that crossed the species barrier, most likely originating from bats or from other species including civets, raccoon dogs, domestic cats, swine, and rodents.
TextSentencer_T8 747-953 Sentence denotes A SARS-CoV vaccine should confer long-term protection, especially in vulnerable senescent populations, against both the 2003 epidemic strains and zoonotic strains that may yet emerge from animal reservoirs.
T39669 747-953 Sentence denotes A SARS-CoV vaccine should confer long-term protection, especially in vulnerable senescent populations, against both the 2003 epidemic strains and zoonotic strains that may yet emerge from animal reservoirs.
TextSentencer_T9 954-1097 Sentence denotes We report the comprehensive investigation of SARS vaccine efficacy in young and senescent mice following homologous and heterologous challenge.
T65628 954-1097 Sentence denotes We report the comprehensive investigation of SARS vaccine efficacy in young and senescent mice following homologous and heterologous challenge.
TextSentencer_T10 1098-1473 Sentence denotes Using Venezuelan equine encephalitis virus replicon particles (VRP) expressing the 2003 epidemic Urbani SARS-CoV strain spike (S) glycoprotein (VRP-S) or the nucleocapsid (N) protein from the same strain (VRP-N), we demonstrate that VRP-S, but not VRP-N vaccines provide complete short-and long-term protection against homologous strain challenge in young and senescent mice.
T122 1098-1473 Sentence denotes Using Venezuelan equine encephalitis virus replicon particles (VRP) expressing the 2003 epidemic Urbani SARS-CoV strain spike (S) glycoprotein (VRP-S) or the nucleocapsid (N) protein from the same strain (VRP-N), we demonstrate that VRP-S, but not VRP-N vaccines provide complete short-and long-term protection against homologous strain challenge in young and senescent mice.
TextSentencer_T11 1474-1990 Sentence denotes To test VRP vaccine efficacy against a heterologous SARS-CoV, we used phylogenetic analyses, synthetic biology, and reverse genetics to construct a chimeric virus (icGDO3-S) encoding a synthetic S glycoprotein gene of the most genetically divergent human strain, GDO3, which clusters among the zoonotic SARS-CoV. icGD03-S replicated efficiently in human airway epithelial cells and in the lungs of young and senescent mice, and was highly resistant to neutralization with antisera directed against the Urbani strain.
T9683 1474-1990 Sentence denotes To test VRP vaccine efficacy against a heterologous SARS-CoV, we used phylogenetic analyses, synthetic biology, and reverse genetics to construct a chimeric virus (icGDO3-S) encoding a synthetic S glycoprotein gene of the most genetically divergent human strain, GDO3, which clusters among the zoonotic SARS-CoV. icGD03-S replicated efficiently in human airway epithelial cells and in the lungs of young and senescent mice, and was highly resistant to neutralization with antisera directed against the Urbani strain.
TextSentencer_T12 1991-2175 Sentence denotes Although VRP-S vaccines provided complete short-term protection against heterologous icGD03-S challenge in young mice, only limited protection was seen in vaccinated senescent animals.
T90393 1991-2175 Sentence denotes Although VRP-S vaccines provided complete short-term protection against heterologous icGD03-S challenge in young mice, only limited protection was seen in vaccinated senescent animals.
TextSentencer_T13 2176-2376 Sentence denotes VRP-N vaccines not only failed to protect from homologous or heterologous challenge, but resulted in enhanced immunopathology with eosinophilic infiltrates within the lungs of SARS-CoVchallenged mice.
T63041 2176-2376 Sentence denotes VRP-N vaccines not only failed to protect from homologous or heterologous challenge, but resulted in enhanced immunopathology with eosinophilic infiltrates within the lungs of SARS-CoVchallenged mice.
TextSentencer_T14 2377-2525 Sentence denotes VRP-N-induced pathology presented at day 4, peaked around day 7, and persisted through day 14, and was likely mediated by cellular immune responses.
T87181 2377-2525 Sentence denotes VRP-N-induced pathology presented at day 4, peaked around day 7, and persisted through day 14, and was likely mediated by cellular immune responses.
TextSentencer_T15 2526-2677 Sentence denotes Severe acute respiratory syndrome coronavirus (SARS-CoV) infection results in severe acute respiratory disease, pneumonia, and sometimes death [1, 2] .
T76270 2526-2677 Sentence denotes Severe acute respiratory syndrome coronavirus (SARS-CoV) infection results in severe acute respiratory disease, pneumonia, and sometimes death [1, 2] .
TextSentencer_T16 2678-2796 Sentence denotes The disease was reported in Guangdong Province, China, in 2002 and spread to more than 30 nations within a few months.
T82049 2678-2796 Sentence denotes The disease was reported in Guangdong Province, China, in 2002 and spread to more than 30 nations within a few months.
TextSentencer_T17 2797-2946 Sentence denotes Disease severity was linked to age and other comorbidities, with mortality rates increasing with age and exceeding 50% in individuals over 65 y [3] .
T16174 2797-2946 Sentence denotes Disease severity was linked to age and other comorbidities, with mortality rates increasing with age and exceeding 50% in individuals over 65 y [3] .
TextSentencer_T18 2947-3149 Sentence denotes SARS-CoV is a zoonotic virus that crossed the species barrier, most likely originating from bats [4] [5] [6] or from other species including civets, raccoon dogs, domestic cats, swine, and rodents [7] .
T40250 2947-3149 Sentence denotes SARS-CoV is a zoonotic virus that crossed the species barrier, most likely originating from bats [4] [5] [6] or from other species including civets, raccoon dogs, domestic cats, swine, and rodents [7] .
TextSentencer_T19 3150-3325 Sentence denotes New zoonotic variants may emerge as evidenced by sporadic cases of human disease in late 2003 and early 2004, which arose from strains distinct from that of the epidemic [8] .
T83592 3150-3325 Sentence denotes New zoonotic variants may emerge as evidenced by sporadic cases of human disease in late 2003 and early 2004, which arose from strains distinct from that of the epidemic [8] .
TextSentencer_T20 3326-3448 Sentence denotes In 2004, several laboratory-acquired infections were reported, including secondary spread resulting in fatal disease [9] .
T98588 3326-3448 Sentence denotes In 2004, several laboratory-acquired infections were reported, including secondary spread resulting in fatal disease [9] .
TextSentencer_T21 3449-3639 Sentence denotes Given the significant health and economic impact, the development of an effective vaccine strategy that is protective against both epidemic and zoonotic SARS-CoV strains is highly desirable.
T69111 3449-3639 Sentence denotes Given the significant health and economic impact, the development of an effective vaccine strategy that is protective against both epidemic and zoonotic SARS-CoV strains is highly desirable.
TextSentencer_T22 3640-3999 Sentence denotes Attenuated and killed SARS-CoV, DNA, and viral vectored vaccines are being evaluated in a number of animal models including mouse, ferret, hamster, and primate [10] [11] [12] [13] [14] [15] [16] [17] [18] [19] [20] [21] [22] [23] , and have demonstrated that the SARS-CoV spike (S) glycoprotein is the principal component of protective immunity [15, 24, 25] .
T43468 3640-3999 Sentence denotes Attenuated and killed SARS-CoV, DNA, and viral vectored vaccines are being evaluated in a number of animal models including mouse, ferret, hamster, and primate [10] [11] [12] [13] [14] [15] [16] [17] [18] [19] [20] [21] [22] [23] , and have demonstrated that the SARS-CoV spike (S) glycoprotein is the principal component of protective immunity [15, 24, 25] .
TextSentencer_T23 4000-4268 Sentence denotes Although strong immune responses are elicited against both S glycoprotein and nucleocapsid (N) protein [10, 15, 26, 27] , passive transfer studies have illustrated that only anti-S antibody confers protection from SARS-CoV replication in the mouse model [16, 17, 28] .
T86413 4000-4268 Sentence denotes Although strong immune responses are elicited against both S glycoprotein and nucleocapsid (N) protein [10, 15, 26, 27] , passive transfer studies have illustrated that only anti-S antibody confers protection from SARS-CoV replication in the mouse model [16, 17, 28] .
TextSentencer_T24 4269-4591 Sentence denotes Vaccine development faces a series of potential concerns including reversion or recombination repair of attenuated vaccine strains, induction of immune-mediated enhancement of pathology, waning immune protection, lack of cross-protection for heterologous strains, and limited vaccine efficacy within senescent populations.
T6818 4269-4591 Sentence denotes Vaccine development faces a series of potential concerns including reversion or recombination repair of attenuated vaccine strains, induction of immune-mediated enhancement of pathology, waning immune protection, lack of cross-protection for heterologous strains, and limited vaccine efficacy within senescent populations.
TextSentencer_T25 4592-4873 Sentence denotes Furthermore, immune enhancement has been demonstrated with another coronavirus, feline infectious peritonitis coronavirus [29] , and more recently with a modified vaccinia vector expressing SARS-S that exacerbated hepatitis in ferrets while failing to protect from infection [30] .
T78132 4592-4873 Sentence denotes Furthermore, immune enhancement has been demonstrated with another coronavirus, feline infectious peritonitis coronavirus [29] , and more recently with a modified vaccinia vector expressing SARS-S that exacerbated hepatitis in ferrets while failing to protect from infection [30] .
TextSentencer_T26 4874-5153 Sentence denotes Notably, some antibodies against the epidemic Urbani strain increased the infectivity of lentiviruses pseudotyped with an animal SARS-S glycoprotein in an in vitro model, raising the specter of vaccine-mediated immune enhancement of disease following heterotypic challenge [31] .
T17464 4874-5153 Sentence denotes Notably, some antibodies against the epidemic Urbani strain increased the infectivity of lentiviruses pseudotyped with an animal SARS-S glycoprotein in an in vitro model, raising the specter of vaccine-mediated immune enhancement of disease following heterotypic challenge [31] .
TextSentencer_T27 5154-5297 Sentence denotes Another potential problem is that SARS vaccines might fail to induce antibodies that protect from infection with divergent strains of SARS-CoV.
T73762 5154-5297 Sentence denotes Another potential problem is that SARS vaccines might fail to induce antibodies that protect from infection with divergent strains of SARS-CoV.
TextSentencer_T28 5298-5494 Sentence denotes The S glycoprotein of SARS-CoV contains about 2%-20% amino acid variation between zoonotic and the 2003 epidemic strains [8, 31] , possibly limiting the effectiveness of monotypic SARS-S vaccines.
T14273 5298-5494 Sentence denotes The S glycoprotein of SARS-CoV contains about 2%-20% amino acid variation between zoonotic and the 2003 epidemic strains [8, 31] , possibly limiting the effectiveness of monotypic SARS-S vaccines.
TextSentencer_T29 5495-5651 Sentence denotes Finally, studies measuring the duration of protective immunity or vaccine efficacy in animals greater than 4 mo post-boost have not yet been reported [17] .
T98876 5495-5651 Sentence denotes Finally, studies measuring the duration of protective immunity or vaccine efficacy in animals greater than 4 mo post-boost have not yet been reported [17] .
TextSentencer_T30 5652-5931 Sentence denotes In this report, the efficacy of Venezuelan equine encephalitis virus replicon particle (VRP) vaccines expressing the Urbani SARS-CoV S glycoprotein (VRP-S) and N protein (VRP-N), either individually or in combination (VRP-SþN), are determined in young and senescent mouse models.
T63859 5652-5931 Sentence denotes In this report, the efficacy of Venezuelan equine encephalitis virus replicon particle (VRP) vaccines expressing the Urbani SARS-CoV S glycoprotein (VRP-S) and N protein (VRP-N), either individually or in combination (VRP-SþN), are determined in young and senescent mouse models.
TextSentencer_T31 5932-6238 Sentence denotes We tested whether the senescent mouse model, which exhibits an age-related susceptibility to SARS-CoV similar to that seen in the human disease [32] , will provide a sensitive measure of vaccine efficacy and reveal potential complications in SARS-CoV vaccine development for vulnerable elderly populations.
T83633 5932-6238 Sentence denotes We tested whether the senescent mouse model, which exhibits an age-related susceptibility to SARS-CoV similar to that seen in the human disease [32] , will provide a sensitive measure of vaccine efficacy and reveal potential complications in SARS-CoV vaccine development for vulnerable elderly populations.
TextSentencer_T32 6239-6412 Sentence denotes We evaluated the duration of protective immunity following homologous and heterologous SARS virus challenge, examining the impact of waning immunity on long-term protection.
T37596 6239-6412 Sentence denotes We evaluated the duration of protective immunity following homologous and heterologous SARS virus challenge, examining the impact of waning immunity on long-term protection.
TextSentencer_T33 6413-6729 Sentence denotes Through the use of publicly available SARS-CoV sequence databases, bioinformatics approaches, synthetic biology, and reverse genetics, we constructed a viable heterologous challenge virus to test the ability of current vaccine regimens to protect against zoonotic strains; the likely source of future epidemics [8] .
T33042 6413-6729 Sentence denotes Through the use of publicly available SARS-CoV sequence databases, bioinformatics approaches, synthetic biology, and reverse genetics, we constructed a viable heterologous challenge virus to test the ability of current vaccine regimens to protect against zoonotic strains; the likely source of future epidemics [8] .
TextSentencer_T34 6730-7047 Sentence denotes The Urbani, Tor-2, recombinant Urbani (icSARS), and a recombinant chimeric virus encoding the S gene of GDO3 SARS-CoV (icGD03-S), strains were propagated on VeroE6 cells in Eagle's MEM supplemented with 10% fetal calf serum, kanamycin (0.25 lg/ml), and gentamycin (0.05 lg/ml) at 37 8C in a humidified CO 2 incubator.
T95812 6730-7047 Sentence denotes The Urbani, Tor-2, recombinant Urbani (icSARS), and a recombinant chimeric virus encoding the S gene of GDO3 SARS-CoV (icGD03-S), strains were propagated on VeroE6 cells in Eagle's MEM supplemented with 10% fetal calf serum, kanamycin (0.25 lg/ml), and gentamycin (0.05 lg/ml) at 37 8C in a humidified CO 2 incubator.
TextSentencer_T35 7048-7231 Sentence denotes For virus growth, cultures of VeroE6 cells were infected at a multiplicity of infection (MOI) of 1 for 1 h, the monolayer washed twice with 2 ml of PBS and overlaid with complete MEM.
T71397 7048-7231 Sentence denotes For virus growth, cultures of VeroE6 cells were infected at a multiplicity of infection (MOI) of 1 for 1 h, the monolayer washed twice with 2 ml of PBS and overlaid with complete MEM.
TextSentencer_T36 7232-7323 Sentence denotes Virus samples were harvested at different times post-infection and titered by plaque assay.
T2067 7232-7323 Sentence denotes Virus samples were harvested at different times post-infection and titered by plaque assay.
TextSentencer_T37 7324-7389 Sentence denotes Plaques were visualized by neutral red staining and then counted.
T21909 7324-7389 Sentence denotes Plaques were visualized by neutral red staining and then counted.
TextSentencer_T38 7390-7673 Sentence denotes Human nasal and tracheobronchial epithelial cells were obtained from airway specimens rejected from patients undergoing elective surgery under University of North Carolina (UNC) Institutional Review Board-approved protocols by the UNC Cystic Fibrosis (CF) Center Tissue Culture Core.
T64057 7390-7673 Sentence denotes Human nasal and tracheobronchial epithelial cells were obtained from airway specimens rejected from patients undergoing elective surgery under University of North Carolina (UNC) Institutional Review Board-approved protocols by the UNC Cystic Fibrosis (CF) Center Tissue Culture Core.
TextSentencer_T39 7674-7894 Sentence denotes Briefly, primary cells were expanded on plastic to generate passage 1 cells and plated at a density of 250,000 cells per well on permeable Transwell-Col (T-Col, 12-mm diameter; Corning [http://www.corning.com]) supports.
T79329 7674-7894 Sentence denotes Briefly, primary cells were expanded on plastic to generate passage 1 cells and plated at a density of 250,000 cells per well on permeable Transwell-Col (T-Col, 12-mm diameter; Corning [http://www.corning.com]) supports.
TextSentencer_T40 7895-8212 Sentence denotes Human airway epithelium (HAE) cultures were generated by provision of an air-liquid interface for 4-6 wk to form welldifferentiated, polarized cultures that resemble in vivo pseudo-stratified mucosciliary epithelium, and infected with wild-type or recombinant SARS-CoV as previously described by our laboratory [33] .
T81240 7895-8212 Sentence denotes Human airway epithelium (HAE) cultures were generated by provision of an air-liquid interface for 4-6 wk to form welldifferentiated, polarized cultures that resemble in vivo pseudo-stratified mucosciliary epithelium, and infected with wild-type or recombinant SARS-CoV as previously described by our laboratory [33] .
TextSentencer_T41 8213-8380 Sentence denotes All virus work was performed in a biological safety cabinet (BSC cabinet) in a biosafety level three (BSL3) laboratory containing redundant HEPA-filtered exhaust fans.
T62973 8213-8380 Sentence denotes All virus work was performed in a biological safety cabinet (BSC cabinet) in a biosafety level three (BSL3) laboratory containing redundant HEPA-filtered exhaust fans.
TextSentencer_T42 8381-8519 Sentence denotes Personnel were double gloved and wore Tyvek suits with hoods supplied with HEPA-filtered air by a powered air-purifying respirator (PAPR).
T14968 8381-8519 Sentence denotes Personnel were double gloved and wore Tyvek suits with hoods supplied with HEPA-filtered air by a powered air-purifying respirator (PAPR).
TextSentencer_T43 8520-8571 Sentence denotes The GD03-S glycoprotein sequence has been reported.
T7721 8520-8571 Sentence denotes The GD03-S glycoprotein sequence has been reported.
TextSentencer_T44 8572-8735 Sentence denotes A synthetic DNA containing the 59-most GD03 mutations was purchased (Blue Heron Biotechnology [http//www. blueheronbio.com]) and inserted into the SARS-E fragment.
T11906 8572-8735 Sentence denotes A synthetic DNA containing the 59-most GD03 mutations was purchased (Blue Heron Biotechnology [http//www. blueheronbio.com]) and inserted into the SARS-E fragment.
TextSentencer_T45 8736-8842 Sentence denotes The plasmid clone (SARS-E GD03) was fully sequenced and shown to contain all of the appropriate mutations.
T50876 8736-8842 Sentence denotes The plasmid clone (SARS-E GD03) was fully sequenced and shown to contain all of the appropriate mutations.
TextSentencer_T46 8843-8922 Sentence denotes The remaining GDO3 mutation was incorporated by PCR mutagenesis (59 amplicon A:
T52736 8843-8922 Sentence denotes The remaining GDO3 mutation was incorporated by PCR mutagenesis (59 amplicon A:
TextSentencer_T47 8923-8964 Sentence denotes 59-CTGTTTTCCCTGGGATCGC-39; 39 amplicon A:
T49807 8923-8964 Sentence denotes 59-CTGTTTTCCCTGGGATCGC-39; 39 amplicon A:
TextSentencer_T48 8965-9021 Sentence denotes 59-NNNNNNCACCTGCTTTTGGGCAACTC-CAATGCC-39; 59 amplicon B:
T62413 8965-9021 Sentence denotes 59-NNNNNNCACCTGCTTTTGGGCAACTC-CAATGCC-39; 59 amplicon B:
TextSentencer_T49 9022-9116 Sentence denotes 59-NNNNNNCACCTG-C A G T T G C C C A A A A T G T T C T C T A T G A G A A C -3 9; 39 amplicon B:
T84406 9022-9116 Sentence denotes 59-NNNNNNCACCTG-C A G T T G C C C A A A A T G T T C T C T A T G A G A A C -3 9; 39 amplicon B:
TextSentencer_T50 9117-9246 Sentence denotes 59-CATAAATTGGATCCATTGCTGG), followed by seamless ligation of the amplicons as previously described [34] into the SARS-F subclone.
T75393 9117-9246 Sentence denotes 59-CATAAATTGGATCCATTGCTGG), followed by seamless ligation of the amplicons as previously described [34] into the SARS-F subclone.
TextSentencer_T51 9247-9373 Sentence denotes The final construct (SARS-F GD03) was fully sequenced and found to contain the appropriate set of GD03-S glycoprotein alleles.
T11966 9247-9373 Sentence denotes The final construct (SARS-F GD03) was fully sequenced and found to contain the appropriate set of GD03-S glycoprotein alleles.
TextSentencer_T52 9374-9425 Sentence denotes The icGDO3-S was generated as previously described.
T5964 9374-9425 Sentence denotes The icGDO3-S was generated as previously described.
TextSentencer_T53 9426-9605 Sentence denotes Infectious clone fragment plasmid DNA was prepared in Escherichia coli (TOP-10, Invitrogen [http://www.invitrogen. com]), isolated, and purified (Qiagen [http://www.qiagen. com]).
T80703 9426-9605 Sentence denotes Infectious clone fragment plasmid DNA was prepared in Escherichia coli (TOP-10, Invitrogen [http://www.invitrogen. com]), isolated, and purified (Qiagen [http://www.qiagen. com]).
TextSentencer_T54 9606-9672 Sentence denotes Infectious clone fragments B, C, D, and E were digested with BglI.
T74614 9606-9672 Sentence denotes Infectious clone fragments B, C, D, and E were digested with BglI.
TextSentencer_T55 9673-9756 Sentence denotes Infectious clone fragments A and F were digested with EcoRI and NotI, respectively.
T9234 9673-9756 Sentence denotes Infectious clone fragments A and F were digested with EcoRI and NotI, respectively.
TextSentencer_T56 9757-9847 Sentence denotes Infectious clone fragments A and F were then dephosphorylated and then digested with BglI.
T56516 9757-9847 Sentence denotes Infectious clone fragments A and F were then dephosphorylated and then digested with BglI.
TextSentencer_T57 9848-10031 Sentence denotes Individual cDNA fragments were gel purified (Qiagen) and ligated (Roche [http://www.roche.com]) to form a full-length genomic cDNA and then chloroform extracted and EtOH precipitated.
T77246 9848-10031 Sentence denotes Individual cDNA fragments were gel purified (Qiagen) and ligated (Roche [http://www.roche.com]) to form a full-length genomic cDNA and then chloroform extracted and EtOH precipitated.
TextSentencer_T58 10032-10172 Sentence denotes N cDNA and full-length viral genomic cDNA were then used as templates for in vitro transcription reactions (Ambion [http://www.ambion.com]).
T48877 10032-10172 Sentence denotes N cDNA and full-length viral genomic cDNA were then used as templates for in vitro transcription reactions (Ambion [http://www.ambion.com]).
TextSentencer_T59 10173-10258 Sentence denotes N and full-length viral genomic transcripts were then electroporated into Vero cells.
T30190 10173-10258 Sentence denotes N and full-length viral genomic transcripts were then electroporated into Vero cells.
TextSentencer_T60 10259-10335 Sentence denotes Cell culture media containing virus was harvested 48 h post-electroporation.
T75521 10259-10335 Sentence denotes Cell culture media containing virus was harvested 48 h post-electroporation.
TextSentencer_T61 10336-10400 Sentence denotes Virus was plaque purified and then passaged twice in Vero cells.
T46601 10336-10400 Sentence denotes Virus was plaque purified and then passaged twice in Vero cells.
TextSentencer_T62 10401-10468 Sentence denotes The resultant stock was plaque titered and cyropreserved at À80 8C.
T27144 10401-10468 Sentence denotes The resultant stock was plaque titered and cyropreserved at À80 8C.
TextSentencer_T63 10469-10826 Sentence denotes Twelve hours post-infection, Urbani-, icSARS-CoV-, SARS-CoV-, Tor-2-, and icGD03-S-infected cells were washed in 1X PBS, lysed in buffer containing 20 mM Tris-HCl (pH 7.6), 150 mM NaCl, 0.5% deoxycholine, 1% Nonidet-p-40, 0.1% SDS, and post-nuclear supernatants added to an equal volume of 5 mM EDTA/0.9% SDS, resulting in a final SDS concentration of 0.5%.
T68675 10469-10826 Sentence denotes Twelve hours post-infection, Urbani-, icSARS-CoV-, SARS-CoV-, Tor-2-, and icGD03-S-infected cells were washed in 1X PBS, lysed in buffer containing 20 mM Tris-HCl (pH 7.6), 150 mM NaCl, 0.5% deoxycholine, 1% Nonidet-p-40, 0.1% SDS, and post-nuclear supernatants added to an equal volume of 5 mM EDTA/0.9% SDS, resulting in a final SDS concentration of 0.5%.
TextSentencer_T64 10827-10910 Sentence denotes Samples were then heat inactivated for 30 min at 90 8C in the BL3 prior to removal.
T68027 10827-10910 Sentence denotes Samples were then heat inactivated for 30 min at 90 8C in the BL3 prior to removal.
TextSentencer_T65 10911-10986 Sentence denotes At BL2, samples were again heat inactivated for 30 min at 90 8C before use.
T48973 10911-10986 Sentence denotes At BL2, samples were again heat inactivated for 30 min at 90 8C before use.
TextSentencer_T66 10987-11145 Sentence denotes Equivalent sample volumes were loaded onto 4% to 20% Criterion gradient gels (BioRad [http://www.bio-rad.com]) and then transferred to PVDF membrane (BioRad).
T72217 10987-11145 Sentence denotes Equivalent sample volumes were loaded onto 4% to 20% Criterion gradient gels (BioRad [http://www.bio-rad.com]) and then transferred to PVDF membrane (BioRad).
TextSentencer_T67 11146-11430 Sentence denotes Blots were probed with polyclonal mouse antisera directed against the Urbani-S glycoprotein diluted 1:200 or convalescent human sera 1128 diluted 1:400 and developed using electrogenerated chemiluminescence (ECL) reagents (Amersham Biosciences [http://www5. amershambiosciences.com]).
T48429 11146-11430 Sentence denotes Blots were probed with polyclonal mouse antisera directed against the Urbani-S glycoprotein diluted 1:200 or convalescent human sera 1128 diluted 1:400 and developed using electrogenerated chemiluminescence (ECL) reagents (Amersham Biosciences [http://www5. amershambiosciences.com]).
TextSentencer_T68 11431-11512 Sentence denotes Patient #1128 was infected during the second disease outbreak in Toronto, Canada.
T232 11431-11512 Sentence denotes Patient #1128 was infected during the second disease outbreak in Toronto, Canada.
TextSentencer_T69 11513-11786 Sentence denotes One hundred plaque-forming units (pfu) of either icSARS-CoV or icGDO3-S were treated with heat-inactivated serum diluted to final concentrations of 1:100, 1:200, 1:400, 1:800, or 1:1,600 and incubated at 37 8C for 30 min, and the resulting titer determined by plaque assay.
T60233 11513-11786 Sentence denotes One hundred plaque-forming units (pfu) of either icSARS-CoV or icGDO3-S were treated with heat-inactivated serum diluted to final concentrations of 1:100, 1:200, 1:400, 1:800, or 1:1,600 and incubated at 37 8C for 30 min, and the resulting titer determined by plaque assay.
TextSentencer_T70 11787-12127 Sentence denotes Plaque numbers formed by virus treated with each dilution of sera from individual mice vaccinated with VRP-S or VRP-SþN were compared to the average number of plaques formed after treatment with a given dilution of sera from VRP expressing the influenza A HA protein (VRP-HA)-or PBS-vaccinated mice and expressed as the relative percentage.
T21141 11787-12127 Sentence denotes Plaque numbers formed by virus treated with each dilution of sera from individual mice vaccinated with VRP-S or VRP-SþN were compared to the average number of plaques formed after treatment with a given dilution of sera from VRP expressing the influenza A HA protein (VRP-HA)-or PBS-vaccinated mice and expressed as the relative percentage.
TextSentencer_T71 12128-12240 Sentence denotes The dilution at which 80% of plaques were neutralized was determined for each VRP-Sor VRP-SþN-vaccinated animal.
T56095 12128-12240 Sentence denotes The dilution at which 80% of plaques were neutralized was determined for each VRP-Sor VRP-SþN-vaccinated animal.
TextSentencer_T72 12241-12452 Sentence denotes Female BALB/c mice (Charles River Laboratories [http:// www.criver.com]) were anesthetized with a ketamine (1.3 mg/ mouse) and xylazine (0.38 mg/mouse) mixture administered intraperitoneally with a 50-ll volume.
T28051 12241-12452 Sentence denotes Female BALB/c mice (Charles River Laboratories [http:// www.criver.com]) were anesthetized with a ketamine (1.3 mg/ mouse) and xylazine (0.38 mg/mouse) mixture administered intraperitoneally with a 50-ll volume.
TextSentencer_T73 12453-12566 Sentence denotes Each mouse was intranasally (i.n.) inoculated with 50 ll of virus at a concentration of 2 3 10 6 pfu/ml of virus.
T36119 12453-12566 Sentence denotes Each mouse was intranasally (i.n.) inoculated with 50 ll of virus at a concentration of 2 3 10 6 pfu/ml of virus.
TextSentencer_T74 12567-12693 Sentence denotes Four days postinfection, the right lung was removed and frozen at À70 8C for later plaque assay determination of viral titers.
T69978 12567-12693 Sentence denotes Four days postinfection, the right lung was removed and frozen at À70 8C for later plaque assay determination of viral titers.
TextSentencer_T75 12694-12779 Sentence denotes Half of the left lung was placed into Trizol Reagent (Invitrogen) for RNA extraction.
T54160 12694-12779 Sentence denotes Half of the left lung was placed into Trizol Reagent (Invitrogen) for RNA extraction.
TextSentencer_T76 12780-12940 Sentence denotes The second half of the left lung was fixed in 4% PFA in PBS (pH 7.4) for at least 7 d prior to paraffin imbedding and sectioning for histopathological analysis.
T52459 12780-12940 Sentence denotes The second half of the left lung was fixed in 4% PFA in PBS (pH 7.4) for at least 7 d prior to paraffin imbedding and sectioning for histopathological analysis.
TextSentencer_T77 12941-13083 Sentence denotes All mice were housed under sterile conditions, and sentinel mice were used to verify that the colony was mouse hepatitis virus (MHV) negative.
T33406 12941-13083 Sentence denotes All mice were housed under sterile conditions, and sentinel mice were used to verify that the colony was mouse hepatitis virus (MHV) negative.
TextSentencer_T78 13084-13204 Sentence denotes Experimental protocols were reviewed and approved by the Institutional Animal Care and Use Committee at UNC Chapel Hill.
T71599 13084-13204 Sentence denotes Experimental protocols were reviewed and approved by the Institutional Animal Care and Use Committee at UNC Chapel Hill.
TextSentencer_T79 13205-13393 Sentence denotes Young mice refer to those challenged with SARS-CoV at ages equal or less than 5 mo old, whereas old or senescent mice are those animals with ages greater than 1 y at the time of challenge.
T1548 13205-13393 Sentence denotes Young mice refer to those challenged with SARS-CoV at ages equal or less than 5 mo old, whereas old or senescent mice are those animals with ages greater than 1 y at the time of challenge.
TextSentencer_T80 13394-13490 Sentence denotes Lungs were weighed and homogenized in four equivalent volumes of PBS to generate a 20% solution.
T22945 13394-13490 Sentence denotes Lungs were weighed and homogenized in four equivalent volumes of PBS to generate a 20% solution.
TextSentencer_T81 13491-13710 Sentence denotes The solution was centrifuged at 13,000 rpm on a tabletop centrifuge for 5 min, the clarified supernatant serially diluted in PBS, and 200-ll volumes of the dilutions placed onto monolayers of Vero cells in 60-mm dishes.
T15216 13491-13710 Sentence denotes The solution was centrifuged at 13,000 rpm on a tabletop centrifuge for 5 min, the clarified supernatant serially diluted in PBS, and 200-ll volumes of the dilutions placed onto monolayers of Vero cells in 60-mm dishes.
TextSentencer_T82 13711-13802 Sentence denotes Following a 1-h incubation at 37 8C, cells were overlaid with 1% agarose-containing medium.
T42581 13711-13802 Sentence denotes Following a 1-h incubation at 37 8C, cells were overlaid with 1% agarose-containing medium.
TextSentencer_T83 13803-13881 Sentence denotes Two days later, plates were stained with neutral red and then plaques counted.
T39633 13803-13881 Sentence denotes Two days later, plates were stained with neutral red and then plaques counted.
TextSentencer_T84 13882-14030 Sentence denotes The VRP constructs were made in two rounds of PCR, the first to generate two amplicons, and a second round of overlapping PCR to fuse them together.
T21833 13882-14030 Sentence denotes The VRP constructs were made in two rounds of PCR, the first to generate two amplicons, and a second round of overlapping PCR to fuse them together.
TextSentencer_T85 14031-14132 Sentence denotes The fused DNA was digested with ApaI and AscI, and ligated into the similarly digested pVR21 plasmid.
T2533 14031-14132 Sentence denotes The fused DNA was digested with ApaI and AscI, and ligated into the similarly digested pVR21 plasmid.
TextSentencer_T86 14133-14333 Sentence denotes PCR reactions were performed with Expand Long Taq (Roche Molecular Biochemicals http://www.roche-applied-science.com) in 30 cycles of 94 8C for 30 s, 55 8C for 30 s, and extensions at 68 8C for 1 min.
T19727 14133-14333 Sentence denotes PCR reactions were performed with Expand Long Taq (Roche Molecular Biochemicals http://www.roche-applied-science.com) in 30 cycles of 94 8C for 30 s, 55 8C for 30 s, and extensions at 68 8C for 1 min.
TextSentencer_T87 14334-14561 Sentence denotes The first amplicon, which was used in the construction of both VRP-S and VRP-N, was generated with primers 59nsp4Sw (59-GATTGAGGCGGCTTTCGGCG) and 3926S (59-TTAATTAAGTCAATCGGCGCGCCCTTGGCG-GACTAGACTATGTC) using pVR21 as template.
T50709 14334-14561 Sentence denotes The first amplicon, which was used in the construction of both VRP-S and VRP-N, was generated with primers 59nsp4Sw (59-GATTGAGGCGGCTTTCGGCG) and 3926S (59-TTAATTAAGTCAATCGGCGCGCCCTTGGCG-GACTAGACTATGTC) using pVR21 as template.
TextSentencer_T88 14562-14758 Sentence denotes The Ngene-specific amplicon was produced using primers V59SARNg (59-AGTCTAGTCCGCCAAGATGTCTGA-TAATGGACCCCAATC) and 39SARSNg (59-NNNNTTAAT-TAATTATGCCTGAGTTGAATCAGC) with SARS-F plasmid for template.
T62665 14562-14758 Sentence denotes The Ngene-specific amplicon was produced using primers V59SARNg (59-AGTCTAGTCCGCCAAGATGTCTGA-TAATGGACCCCAATC) and 39SARSNg (59-NNNNTTAAT-TAATTATGCCTGAGTTGAATCAGC) with SARS-F plasmid for template.
TextSentencer_T89 14759-14963 Sentence denotes The S-gene-containing amplicon was made with V59SARSg (59-AGTCTAGTCCGCCAAGATGTT-TATTTTCTTATTATTTCTTACTCTCAC) and SARS39Sg (59-NNNNTTAATTAATTATGTGTAATGTAATTTGA-CACCC) using ligated SARS-E and -F fragments.
T55551 14759-14963 Sentence denotes The S-gene-containing amplicon was made with V59SARSg (59-AGTCTAGTCCGCCAAGATGTT-TATTTTCTTATTATTTCTTACTCTCAC) and SARS39Sg (59-NNNNTTAATTAATTATGTGTAATGTAATTTGA-CACCC) using ligated SARS-E and -F fragments.
TextSentencer_T90 14964-15093 Sentence denotes The VRP-S and VRP-N cDNA templates were sequenced for verification and replicon particles produced as previously described [35] .
T91413 14964-15093 Sentence denotes The VRP-S and VRP-N cDNA templates were sequenced for verification and replicon particles produced as previously described [35] .
TextSentencer_T91 15094-15197 Sentence denotes Mice were vaccinated with 10 6 infectious units (IU) of VRP in a 10-ll volume in the left rear footpad.
T5449 15094-15197 Sentence denotes Mice were vaccinated with 10 6 infectious units (IU) of VRP in a 10-ll volume in the left rear footpad.
TextSentencer_T92 15198-15407 Sentence denotes Lungs were fixed in 4% PFA in PBS for 7 d before being submitted to the Histopathology Core Facility (UNC, Chapel Hill) for paraffin imbedding, sectioning at 5-lm thickness, and hematoxylin and eosin staining.
T78397 15198-15407 Sentence denotes Lungs were fixed in 4% PFA in PBS for 7 d before being submitted to the Histopathology Core Facility (UNC, Chapel Hill) for paraffin imbedding, sectioning at 5-lm thickness, and hematoxylin and eosin staining.
TextSentencer_T93 15408-15579 Sentence denotes Approximately onequarter of the total lungs were sectioned, with four to six sections mounted from cuts taken at five different depths within the paraffin-imbedded tissue.
T41702 15408-15579 Sentence denotes Approximately onequarter of the total lungs were sectioned, with four to six sections mounted from cuts taken at five different depths within the paraffin-imbedded tissue.
TextSentencer_T94 15580-15719 Sentence denotes Lung pathology was scored in a blinded manner, in which six to ten sections per animal were evaluated and scored using the following scale.
T6210 15580-15719 Sentence denotes Lung pathology was scored in a blinded manner, in which six to ten sections per animal were evaluated and scored using the following scale.
TextSentencer_T95 15720-15965 Sentence denotes 1.0 to 2.0 ¼ no to mild inflammation, 2.0 to 3.0 ¼ mild to moderate inflammation, 3.0 to 4.0 ¼ moderate to severe inflammation in less than half of the tissue section, and 4.0 to 5.0 ¼ severe inflammation in more than half of the tissue section.
T5349 15720-15965 Sentence denotes 1.0 to 2.0 ¼ no to mild inflammation, 2.0 to 3.0 ¼ mild to moderate inflammation, 3.0 to 4.0 ¼ moderate to severe inflammation in less than half of the tissue section, and 4.0 to 5.0 ¼ severe inflammation in more than half of the tissue section.
TextSentencer_T96 15966-16066 Sentence denotes The same sets of tissues were also evaluated qualitatively by a respiratory pathologist (author JH).
T1116 15966-16066 Sentence denotes The same sets of tissues were also evaluated qualitatively by a respiratory pathologist (author JH).
TextSentencer_T97 16067-16314 Sentence denotes The 5 lm-thick paraffin-embedded sections were probed with 35 S UTP-labeled riboprobes complementary to the N gene of SARS-CoV (Urbani) or the HA gene of the A/PR8 strain of influenza as a negative control using previously described methods [36] .
T7364 16067-16314 Sentence denotes The 5 lm-thick paraffin-embedded sections were probed with 35 S UTP-labeled riboprobes complementary to the N gene of SARS-CoV (Urbani) or the HA gene of the A/PR8 strain of influenza as a negative control using previously described methods [36] .
TextSentencer_T98 16315-16490 Sentence denotes In brief, following treatment to prevent nonspecific probe binding, the tissues were incubated overnight with either probe at 5 3 10 4 cpm/ll in hybridization buffer at 42 8C.
T2507 16315-16490 Sentence denotes In brief, following treatment to prevent nonspecific probe binding, the tissues were incubated overnight with either probe at 5 3 10 4 cpm/ll in hybridization buffer at 42 8C.
TextSentencer_T99 16491-16649 Sentence denotes The slides were then washed, dehydrated, and coated with NBT emulsion (Kodak [http:// www.kodak.com]), and incubated at À80 8C. for 1 wk prior to development.
T18733 16491-16649 Sentence denotes The slides were then washed, dehydrated, and coated with NBT emulsion (Kodak [http:// www.kodak.com]), and incubated at À80 8C. for 1 wk prior to development.
TextSentencer_T100 16650-16728 Sentence denotes Positive signal, as determined by silver grain deposition, was then evaluated.
T79309 16650-16728 Sentence denotes Positive signal, as determined by silver grain deposition, was then evaluated.
TextSentencer_T101 16729-16824 Sentence denotes Antibody titers were determined by standard indirect enzyme-linked immunosorbent assay (ELISA).
T25696 16729-16824 Sentence denotes Antibody titers were determined by standard indirect enzyme-linked immunosorbent assay (ELISA).
TextSentencer_T102 16825-17089 Sentence denotes High-binding 96-well round-bottom plates (Corning [http://www.corning. com]) were coated with 10 lg/ml of SARS-S, SARS-N, or inactivated influenza A diluted in carbonate buffer containing 32 mM sodium carbonate, 68 mM sodium bicarbonate (pH 9.6) at 4 8C overnight.
T92190 16825-17089 Sentence denotes High-binding 96-well round-bottom plates (Corning [http://www.corning. com]) were coated with 10 lg/ml of SARS-S, SARS-N, or inactivated influenza A diluted in carbonate buffer containing 32 mM sodium carbonate, 68 mM sodium bicarbonate (pH 9.6) at 4 8C overnight.
TextSentencer_T103 17090-17299 Sentence denotes Mouse sera, diluted 1:100 in casein blocking buffer (Sigma [http://www.sigmaaldrich.com]), were added to wells in duplicate, and 2-fold serial dilutions were performed, followed by incubation for 2 h at 37 8C.
T66377 17090-17299 Sentence denotes Mouse sera, diluted 1:100 in casein blocking buffer (Sigma [http://www.sigmaaldrich.com]), were added to wells in duplicate, and 2-fold serial dilutions were performed, followed by incubation for 2 h at 37 8C.
TextSentencer_T104 17300-17554 Sentence denotes Plates were then incubated for 1 h with goat anti-mouse IgG with alkaline phosphatase (AP) conjugate (Sigma), developed with p-nitrophenyl phosphate (pNPP; Sigma), and the optical density (OD) at 405 nm was measured (Bio-Rad Model 680 microplate reader).
T54529 17300-17554 Sentence denotes Plates were then incubated for 1 h with goat anti-mouse IgG with alkaline phosphatase (AP) conjugate (Sigma), developed with p-nitrophenyl phosphate (pNPP; Sigma), and the optical density (OD) at 405 nm was measured (Bio-Rad Model 680 microplate reader).
TextSentencer_T105 17555-17879 Sentence denotes Log 10 half-maximum ELISA titers were calculated with Sigmaplot (Systat [http://www.systat.com]) using the solution of the sigmoidal line of the plot of the log of the reciprocal dilutions of mouse sera and the resulting absorbances to determine the log of the reciprocal dilution at which an absorbance of 2.1 was achieved.
T77055 17555-17879 Sentence denotes Log 10 half-maximum ELISA titers were calculated with Sigmaplot (Systat [http://www.systat.com]) using the solution of the sigmoidal line of the plot of the log of the reciprocal dilutions of mouse sera and the resulting absorbances to determine the log of the reciprocal dilution at which an absorbance of 2.1 was achieved.
TextSentencer_T106 17880-18016 Sentence denotes Since very low amounts of antibody were being measured in the passive transfer experiment, log 10 OD ¼ 0.2 ELISA titers were calculated.
T55291 17880-18016 Sentence denotes Since very low amounts of antibody were being measured in the passive transfer experiment, log 10 OD ¼ 0.2 ELISA titers were calculated.
TextSentencer_T107 18017-18171 Sentence denotes Mice were inoculated with 10 6 IU of VRP-HA, VRP-S, or VRP-N at 7 wk of age, boosted 4 wk later, and terminally bled via cardiac puncture 3 wk post-boost.
T32218 18017-18171 Sentence denotes Mice were inoculated with 10 6 IU of VRP-HA, VRP-S, or VRP-N at 7 wk of age, boosted 4 wk later, and terminally bled via cardiac puncture 3 wk post-boost.
TextSentencer_T108 18172-18284 Sentence denotes The sera of each group were pooled and 150 ll transferred by tail vein injection into mice at 7 or 43 wk of age.
T51845 18172-18284 Sentence denotes The sera of each group were pooled and 150 ll transferred by tail vein injection into mice at 7 or 43 wk of age.
TextSentencer_T109 18285-18359 Sentence denotes Mice receiving sera were bled and i.n. challenged with 10 5 pfu of icSARS.
T77175 18285-18359 Sentence denotes Mice receiving sera were bled and i.n. challenged with 10 5 pfu of icSARS.
TextSentencer_T110 18360-18452 Sentence denotes Unless otherwise noted, two-tailed Mann-Whitney tests were used for statistical comparisons.
T67734 18360-18452 Sentence denotes Unless otherwise noted, two-tailed Mann-Whitney tests were used for statistical comparisons.
TextSentencer_T111 18453-18682 Sentence denotes The Fisher exact tests were completed by comparing the number of animals positive for viral replication within the lungs of a group of animals vaccinated with VRP-S or VRP-SþN to that of the negative control group, VRP-HA or PBS.
T9455 18453-18682 Sentence denotes The Fisher exact tests were completed by comparing the number of animals positive for viral replication within the lungs of a group of animals vaccinated with VRP-S or VRP-SþN to that of the negative control group, VRP-HA or PBS.
TextSentencer_T112 18683-18792 Sentence denotes Values outside the limit of detection were assigned a value equal to the limit of detection for any analysis.
T63008 18683-18792 Sentence denotes Values outside the limit of detection were assigned a value equal to the limit of detection for any analysis.
TextSentencer_T113 18793-18861 Sentence denotes The plus or minus (6) symbol is used to refer to standard deviation.
T18924 18793-18861 Sentence denotes The plus or minus (6) symbol is used to refer to standard deviation.
TextSentencer_T114 18862-19192 Sentence denotes An amino acid multiple alignment was generated for the entire S gene of viral sequences representing early, middle, and late phases of the SARS epidemic in humans, as well as animal strains of SARS-CoV isolated from civets and racoon dogs found in Chinese live animal markets or housed on farms in China that supplied the markets.
T82319 18862-19192 Sentence denotes An amino acid multiple alignment was generated for the entire S gene of viral sequences representing early, middle, and late phases of the SARS epidemic in humans, as well as animal strains of SARS-CoV isolated from civets and racoon dogs found in Chinese live animal markets or housed on farms in China that supplied the markets.
TextSentencer_T115 19193-19268 Sentence denotes The sequences were aligned using ClustalX 1.83 with default settings [37] .
T30661 19193-19268 Sentence denotes The sequences were aligned using ClustalX 1.83 with default settings [37] .
TextSentencer_T116 19269-19379 Sentence denotes A phylogenetic tree was generated using Bayesian inference as implemented in the program MrBayes v3.0b4 [38] .
T44186 19269-19379 Sentence denotes A phylogenetic tree was generated using Bayesian inference as implemented in the program MrBayes v3.0b4 [38] .
TextSentencer_T117 19380-19691 Sentence denotes Briefly, the alignment was exported in the nexus format, the amino acid substitution model was set to JTT [39] using the lset command, and Markov chain Monte Carlo simulation [38] was used to approximate the posterior probabilities of trees with sampling conducted on four chains over 500,000 generations [40] .
T93574 19380-19691 Sentence denotes Briefly, the alignment was exported in the nexus format, the amino acid substitution model was set to JTT [39] using the lset command, and Markov chain Monte Carlo simulation [38] was used to approximate the posterior probabilities of trees with sampling conducted on four chains over 500,000 generations [40] .
TextSentencer_T118 19692-19895 Sentence denotes Trees were sampled every 100 generations, and the 5,001 trees collected were summarized with the sumt command set to a burnin of 1,000, which generated a consensus tree using the 50% majority rule [40] .
T79754 19692-19895 Sentence denotes Trees were sampled every 100 generations, and the 5,001 trees collected were summarized with the sumt command set to a burnin of 1,000, which generated a consensus tree using the 50% majority rule [40] .
TextSentencer_T119 19896-20142 Sentence denotes The burnin value was determined using the sump command with an arbitrary burnin of 250, which demonstrated that stationarity occurred prior to the 100,000th generation, indicating that a burnin of 1,000 was appropriate for the sumt command [40] .
T95869 19896-20142 Sentence denotes The burnin value was determined using the sump command with an arbitrary burnin of 250, which demonstrated that stationarity occurred prior to the 100,000th generation, indicating that a burnin of 1,000 was appropriate for the sumt command [40] .
TextSentencer_T120 20143-20285 Sentence denotes The SARS-CoV S glycoprotein gene and N protein gene were PCR cloned, sequence verified, and inserted into Venezuelan equine encephalitis VRPs.
T80663 20143-20285 Sentence denotes The SARS-CoV S glycoprotein gene and N protein gene were PCR cloned, sequence verified, and inserted into Venezuelan equine encephalitis VRPs.
TextSentencer_T121 20286-20435 Sentence denotes VRP-S and VRP-N constructs were packaged to give titers greater than 10 9 IU per ml and shown to express antigenically relevant recombinant proteins.
T1183 20286-20435 Sentence denotes VRP-S and VRP-N constructs were packaged to give titers greater than 10 9 IU per ml and shown to express antigenically relevant recombinant proteins.
TextSentencer_T122 20436-20542 Sentence denotes VRP-infected cell lysates were probed with antiserum 1128, derived from a convalescent human SARS patient.
T43714 20436-20542 Sentence denotes VRP-infected cell lysates were probed with antiserum 1128, derived from a convalescent human SARS patient.
TextSentencer_T123 20543-20816 Sentence denotes Western blot analysis of VRP-S-infected lysates revealed the expected S glycoprotein doublet of approximately 180-210 kDa, whereas that of VRP-N-infected lysates revealed a major product of less than 50 kDa, the expected sizes for SARS-S and -N, respectively ( Figure 1A ).
T64866 20543-20816 Sentence denotes Western blot analysis of VRP-S-infected lysates revealed the expected S glycoprotein doublet of approximately 180-210 kDa, whereas that of VRP-N-infected lysates revealed a major product of less than 50 kDa, the expected sizes for SARS-S and -N, respectively ( Figure 1A ).
TextSentencer_T124 20817-20918 Sentence denotes VRP-S was inoculated into BALB/c mice and tested for its ability to induce antigen-specific antibody.
T91397 20817-20918 Sentence denotes VRP-S was inoculated into BALB/c mice and tested for its ability to induce antigen-specific antibody.
TextSentencer_T125 20919-21149 Sentence denotes Western blots were performed with Vero cell lysates infected with Urbani, SARS-CoV Tor-2, icSARS-CoV (the Urbani recombinant virus), and icGD03-S, a chimeric SARS-CoV expressing the S glycopro-tein of the heterologous GDO3 strain.
T58658 20919-21149 Sentence denotes Western blots were performed with Vero cell lysates infected with Urbani, SARS-CoV Tor-2, icSARS-CoV (the Urbani recombinant virus), and icGD03-S, a chimeric SARS-CoV expressing the S glycopro-tein of the heterologous GDO3 strain.
TextSentencer_T126 21150-21259 Sentence denotes Blots were probed with anti-VRP-S mouse serum, 1128 human convalescent serum, or with anti-VRP-N mouse serum.
T33106 21150-21259 Sentence denotes Blots were probed with anti-VRP-S mouse serum, 1128 human convalescent serum, or with anti-VRP-N mouse serum.
TextSentencer_T127 21260-21497 Sentence denotes The Western blots demonstrated that probing with human serum resolved bands corresponding to the major SARS antigens S (a doublet at ;180-210 kDa) and N (triplet at ,50 kDa), as well as other unidentified SARS-CoV proteins ( Figure 1B ).
T45813 21260-21497 Sentence denotes The Western blots demonstrated that probing with human serum resolved bands corresponding to the major SARS antigens S (a doublet at ;180-210 kDa) and N (triplet at ,50 kDa), as well as other unidentified SARS-CoV proteins ( Figure 1B ).
TextSentencer_T128 21498-21723 Sentence denotes Serum from mice vaccinated with VRP-S only identified SARS-S ( Figure 1C ), whereas serum from mice vaccinated with VRP-N recognized SARS-N in addition to another SARS-CoV protein that is probably a dimer of N ( Figure 1D ).
T49058 21498-21723 Sentence denotes Serum from mice vaccinated with VRP-S only identified SARS-S ( Figure 1C ), whereas serum from mice vaccinated with VRP-N recognized SARS-N in addition to another SARS-CoV protein that is probably a dimer of N ( Figure 1D ).
TextSentencer_T129 21724-21989 Sentence denotes As a general measure of vaccine efficacy (Table 1 , experiment 1), six 4-wk-old BALB/c mice were vaccinated with either 10 5 IU of VRP-S or VRP-HA, boosted 4 wk later with an equal amount of VRP, and then i.n. challenged with 10 5 pfu of icSARS-CoV 8 wk post-boost.
T20022 21724-21989 Sentence denotes As a general measure of vaccine efficacy (Table 1 , experiment 1), six 4-wk-old BALB/c mice were vaccinated with either 10 5 IU of VRP-S or VRP-HA, boosted 4 wk later with an equal amount of VRP, and then i.n. challenged with 10 5 pfu of icSARS-CoV 8 wk post-boost.
TextSentencer_T130 21990-22211 Sentence denotes Consistent with other studies that made use of vectored SARS-S-expressing vaccines to induce protective responses [16, 18, 41, 42] , vaccination with VRP-S also prevented the replication of icSARS-CoV following challenge.
T35595 21990-22211 Sentence denotes Consistent with other studies that made use of vectored SARS-S-expressing vaccines to induce protective responses [16, 18, 41, 42] , vaccination with VRP-S also prevented the replication of icSARS-CoV following challenge.
TextSentencer_T131 22212-22444 Sentence denotes No virus was detected by plaque assay (250 pfu/ g limit of detection) in the lungs of VRP-S-vaccinated animals at 2 d post-infection, whereas the VRP-HA-vaccinated mouse lung had a mean titer of 6.7 6 0.5 log 10 pfu/g ( Figure 2A ).
T13047 22212-22444 Sentence denotes No virus was detected by plaque assay (250 pfu/ g limit of detection) in the lungs of VRP-S-vaccinated animals at 2 d post-infection, whereas the VRP-HA-vaccinated mouse lung had a mean titer of 6.7 6 0.5 log 10 pfu/g ( Figure 2A ).
TextSentencer_T132 22445-22612 Sentence denotes Vaccination with VRP-S demonstrated significant protection at the time of peak lung titer relative to VRP-HA-vaccinated control animals (p ¼ 0.007, Fisher exact test).
T89783 22445-22612 Sentence denotes Vaccination with VRP-S demonstrated significant protection at the time of peak lung titer relative to VRP-HA-vaccinated control animals (p ¼ 0.007, Fisher exact test).
TextSentencer_T133 22613-22692 Sentence denotes A second vaccine experiment was completed to evaluate long-term VRP protection.
T37458 22613-22692 Sentence denotes A second vaccine experiment was completed to evaluate long-term VRP protection.
TextSentencer_T134 22693-22843 Sentence denotes Five-week-old BALB/c mice were vaccinated with 10 5 IU of VRP-HA, VRP-S, VRP-N, or a combination of VRP-S and VRP-N (VRP-SþN), and boosted 5 wk later.
T25300 22693-22843 Sentence denotes Five-week-old BALB/c mice were vaccinated with 10 5 IU of VRP-HA, VRP-S, VRP-N, or a combination of VRP-S and VRP-N (VRP-SþN), and boosted 5 wk later.
TextSentencer_T135 22844-23003 Sentence denotes Fifty-four weeks post-boost, mice were i.n. challenged with 10 5 pfu of icSARS-CoV and lungs removed 4 d post-infection (summarized in Table 1 , experiment 2).
T59229 22844-23003 Sentence denotes Fifty-four weeks post-boost, mice were i.n. challenged with 10 5 pfu of icSARS-CoV and lungs removed 4 d post-infection (summarized in Table 1 , experiment 2).
TextSentencer_T136 23004-23198 Sentence denotes Although day 2 post-challenge demonstrates peak viral titers, day 4 was chosen to harvest lungs because it is the time at which the highest level of pathology is evident in senescent mice [32] .
T62813 23004-23198 Sentence denotes Although day 2 post-challenge demonstrates peak viral titers, day 4 was chosen to harvest lungs because it is the time at which the highest level of pathology is evident in senescent mice [32] .
TextSentencer_T137 23199-23342 Sentence denotes Titers in the lungs ( Figure 2B ) of animals vaccinated with VRP-S or the combination of VRP-SþN were below the limit of detection (250 pfu/g).
T38901 23199-23342 Sentence denotes Titers in the lungs ( Figure 2B ) of animals vaccinated with VRP-S or the combination of VRP-SþN were below the limit of detection (250 pfu/g).
TextSentencer_T138 23343-23515 Sentence denotes In contrast, the lung titers of VRP-HA-vaccinated animals were 5.8 6 0.6 log 10 pfu/g, comparable to the VRP-N-vaccinated animal titers of 5.3 6 0.6 log 10 pfu/g (p ¼ 0.2).
T44796 23343-23515 Sentence denotes In contrast, the lung titers of VRP-HA-vaccinated animals were 5.8 6 0.6 log 10 pfu/g, comparable to the VRP-N-vaccinated animal titers of 5.3 6 0.6 log 10 pfu/g (p ¼ 0.2).
TextSentencer_T139 23516-23651 Sentence denotes These plaque assay results were confirmed by SARS-CoV-specific in situ hybridization on lung tissues from the infected mice ( Figure 2C
T91075 23516-23651 Sentence denotes These plaque assay results were confirmed by SARS-CoV-specific in situ hybridization on lung tissues from the infected mice ( Figure 2C
TextSentencer_T140 23653-23753 Sentence denotes To perform cross-protection efficacy studies, it was necessary to construct a heterologous SARS-CoV.
T26594 23653-23753 Sentence denotes To perform cross-protection efficacy studies, it was necessary to construct a heterologous SARS-CoV.
TextSentencer_T141 23754-23910 Sentence denotes Selection of a likely candidate strain was made after Bayesian analysis of the SARS-CoV S glycoprotein, which demonstrated three main phylogenetic branches.
T68434 23754-23910 Sentence denotes Selection of a likely candidate strain was made after Bayesian analysis of the SARS-CoV S glycoprotein, which demonstrated three main phylogenetic branches.
TextSentencer_T142 23911-24249 Sentence denotes Two of the branches include viruses isolated from animals, such as the palm civet and raccoon dog, and low pathogenic viruses sporadically isolated from humans, such as GDO3 and GZ0401.Viruses representing the 2003 early, middle, and late phases of the epidemic strains form the third branch in the SARS-S phylogenetic tree ( Figure 3A) .
T22396 23911-24249 Sentence denotes Two of the branches include viruses isolated from animals, such as the palm civet and raccoon dog, and low pathogenic viruses sporadically isolated from humans, such as GDO3 and GZ0401.Viruses representing the 2003 early, middle, and late phases of the epidemic strains form the third branch in the SARS-S phylogenetic tree ( Figure 3A) .
TextSentencer_T143 24250-24358 Sentence denotes We resurrected the S glycoprotein of GDO3, a virus reported from a sporadic SARS case on December 22, 2003 .
T2144 24250-24358 Sentence denotes We resurrected the S glycoprotein of GDO3, a virus reported from a sporadic SARS case on December 22, 2003 .
TextSentencer_T144 24359-24450 Sentence denotes Although GDO3 was not successfully isolated, its S glycoprotein was sequenced and reported.
T56773 24359-24450 Sentence denotes Although GDO3 was not successfully isolated, its S glycoprotein was sequenced and reported.
TextSentencer_T145 24451-24656 Sentence denotes Compared to epidemic strains, GDO3 likely represented an independent introduction, was reported to be less pathogenic, and its S glycoprotein sequence is among the most divergent of all human strains [8] .
T18120 24451-24656 Sentence denotes Compared to epidemic strains, GDO3 likely represented an independent introduction, was reported to be less pathogenic, and its S glycoprotein sequence is among the most divergent of all human strains [8] .
TextSentencer_T146 24657-24924 Sentence denotes The GDO3-S glycoprotein contains 17 amino acid changes relative to Urbani-S ( Figure 3B ), many of which map within neutralizing epitopes between amino acids 130-150 and 318-510, part of the receptor binding domain (RBD) [41, [43] [44] [45] [46] [47] [48] [49] [50] .
T67481 24657-24924 Sentence denotes The GDO3-S glycoprotein contains 17 amino acid changes relative to Urbani-S ( Figure 3B ), many of which map within neutralizing epitopes between amino acids 130-150 and 318-510, part of the receptor binding domain (RBD) [41, [43] [44] [45] [46] [47] [48] [49] [50] .
TextSentencer_T147 24925-25122 Sentence denotes Importantly, polyclonal antibody directed against the late-phase Urbani strain was less effective at neutralizing pseudotyped viruses bearing GDO3-S glycoproteins than those bearing Urbani-S [31] .
T97355 24925-25122 Sentence denotes Importantly, polyclonal antibody directed against the late-phase Urbani strain was less effective at neutralizing pseudotyped viruses bearing GDO3-S glycoproteins than those bearing Urbani-S [31] .
TextSentencer_T148 25123-25302 Sentence denotes The Urbani-S glycoprotein was removed from the SARS-CoV molecular clone, replaced with a synthetic cDNA encoding the GD03-S sequence, and used to generate recombinant virus [34] .
T33244 25123-25302 Sentence denotes The Urbani-S glycoprotein was removed from the SARS-CoV molecular clone, replaced with a synthetic cDNA encoding the GD03-S sequence, and used to generate recombinant virus [34] .
TextSentencer_T149 25303-25545 Sentence denotes Sequence analysis of plaque-purified icGDO3-S recombinant virus confirmed the presence of the GDO3-S glycoprotein and two additional changes in the S gene relative to Urbani-S (F7L and D613G), which likely arose as tissue-culture adaptations.
T46881 25303-25545 Sentence denotes Sequence analysis of plaque-purified icGDO3-S recombinant virus confirmed the presence of the GDO3-S glycoprotein and two additional changes in the S gene relative to Urbani-S (F7L and D613G), which likely arose as tissue-culture adaptations.
TextSentencer_T150 25546-25899 Sentence denotes The chimeric icGDO3-S, which only differs from Urbani SARS-CoV in its S glycoprotein, and wild-type icSARS-CoV recombinant viruses replicated in Vero cells to comparable titers that approached 10 7 PFU/ml within 24 h (unpublished data) and their proteins were both detected in Western blots with human antiserum from convalescent patients ( Figure 1B) .
T2908 25546-25899 Sentence denotes The chimeric icGDO3-S, which only differs from Urbani SARS-CoV in its S glycoprotein, and wild-type icSARS-CoV recombinant viruses replicated in Vero cells to comparable titers that approached 10 7 PFU/ml within 24 h (unpublished data) and their proteins were both detected in Western blots with human antiserum from convalescent patients ( Figure 1B) .
TextSentencer_T151 25900-26385 Sentence denotes Given the reduced amount of N present in the lysate of icGDO3-S-infected cells, the reduced intensity of the GDO3-S band probed with either anti-VRP-S mouse sera or the convalescent human serum is most likely due to the presence of lower GDO3-S protein rather than a marked difference in antibody specificity between GDO3-S and Urbani-S. icGD03-S replicated efficiently in HAE cells, although its maximum titer was approximately 1 log lower than that of icSARS or Urbani ( Figure 3C ).
T63745 25900-26385 Sentence denotes Given the reduced amount of N present in the lysate of icGDO3-S-infected cells, the reduced intensity of the GDO3-S band probed with either anti-VRP-S mouse sera or the convalescent human serum is most likely due to the presence of lower GDO3-S protein rather than a marked difference in antibody specificity between GDO3-S and Urbani-S. icGD03-S replicated efficiently in HAE cells, although its maximum titer was approximately 1 log lower than that of icSARS or Urbani ( Figure 3C ).
TextSentencer_T152 26386-26526 Sentence denotes To compare the growth of icSARS and icGDO3-S in animals, 6-wk-old BALB/c mice were i.n. infected with 10 5 pfu of either icSARS or icGDO3-S.
T88553 26386-26526 Sentence denotes To compare the growth of icSARS and icGDO3-S in animals, 6-wk-old BALB/c mice were i.n. infected with 10 5 pfu of either icSARS or icGDO3-S.
TextSentencer_T153 26527-26677 Sentence denotes At 2 d post-infection, icSARS-CoV mean lung titer was 6.8 6 0.5 log 10 pfu/g, whereas icGDO3-S titers were lower at 6.3 6 0.2 log 10 pfu/g (p ¼ 0.04).
T28850 26527-26677 Sentence denotes At 2 d post-infection, icSARS-CoV mean lung titer was 6.8 6 0.5 log 10 pfu/g, whereas icGDO3-S titers were lower at 6.3 6 0.2 log 10 pfu/g (p ¼ 0.04).
TextSentencer_T154 26678-26814 Sentence denotes The mean lung titer of icSARS-CoV-infected mice on day 4 was 4.5 6 0.5 log 10 pfu/g compared to icGDO3-S titers of 3.7 6 0.3 (p ¼ 0.04).
T57306 26678-26814 Sentence denotes The mean lung titer of icSARS-CoV-infected mice on day 4 was 4.5 6 0.5 log 10 pfu/g compared to icGDO3-S titers of 3.7 6 0.3 (p ¼ 0.04).
TextSentencer_T155 26815-27003 Sentence denotes By the seventh day, virus replication in the lungs of three of five mice infected with icSARS-CoV and four of five mice infected with icGDO3-S fell below the limit of detection (50 pfu/g).
T26257 26815-27003 Sentence denotes By the seventh day, virus replication in the lungs of three of five mice infected with icSARS-CoV and four of five mice infected with icGDO3-S fell below the limit of detection (50 pfu/g).
TextSentencer_T156 27004-27158 Sentence denotes Average icSARS-CoV and icGDO3-S titers were similar on day 7 with 1.7 6 0.1 log 10 pfu/g and 1.8 6 0.1 log 10 pfu/g (p ¼ 0.9), respectively ( Figure 3D ).
T66519 27004-27158 Sentence denotes Average icSARS-CoV and icGDO3-S titers were similar on day 7 with 1.7 6 0.1 log 10 pfu/g and 1.8 6 0.1 log 10 pfu/g (p ¼ 0.9), respectively ( Figure 3D ).
TextSentencer_T157 27159-27459 Sentence denotes To evaluate VRP protection against short-term heterologous challenge, groups of eight animals were primed at 7 wk of age with 10 6 IU of VRP-S, VRP-N, VRP-SþN, or VRP-HA, boosted 3 wk later, and then challenged 7 wk post-boost with 10 5 pfu of icGD03-S (summarized in Table 1 Table 1 , experiment 4).
T30002 27159-27459 Sentence denotes To evaluate VRP protection against short-term heterologous challenge, groups of eight animals were primed at 7 wk of age with 10 6 IU of VRP-S, VRP-N, VRP-SþN, or VRP-HA, boosted 3 wk later, and then challenged 7 wk post-boost with 10 5 pfu of icGD03-S (summarized in Table 1 Table 1 , experiment 4).
TextSentencer_T158 27460-27640 Sentence denotes At 4 d post-infection, mean titers in the lungs of animals vaccinated with VRP-N and PBS were similar at 4.4 6 0.5 and 4.7 6 0.6 log 10 pfu/g (p ¼ 0.2), respectively ( Figure 4B ).
T55396 27460-27640 Sentence denotes At 4 d post-infection, mean titers in the lungs of animals vaccinated with VRP-N and PBS were similar at 4.4 6 0.5 and 4.7 6 0.6 log 10 pfu/g (p ¼ 0.2), respectively ( Figure 4B ).
TextSentencer_T159 27641-27872 Sentence denotes VRP-S vaccination provided partial protection when compared to the PBS control group (p ¼ 0.026, Fisher exact test), with the lungs of three of eight animals positive for viral replication at a mean titer of 2.9 6 1.8 log 10 pfu/g.
T27745 27641-27872 Sentence denotes VRP-S vaccination provided partial protection when compared to the PBS control group (p ¼ 0.026, Fisher exact test), with the lungs of three of eight animals positive for viral replication at a mean titer of 2.9 6 1.8 log 10 pfu/g.
TextSentencer_T160 27873-28147 Sentence denotes All eight of the lungs harvested from the VRP-SþN-vaccinated animals were positive for viral replication, although a mean titer of 3.5 6 1.2 log 10 pfu/g was comparable to the mean titer for VRP-S-vaccinated mice (p ¼ 0.4) and reduced relative to the PBS control (p ¼ 0.02).
T84032 27873-28147 Sentence denotes All eight of the lungs harvested from the VRP-SþN-vaccinated animals were positive for viral replication, although a mean titer of 3.5 6 1.2 log 10 pfu/g was comparable to the mean titer for VRP-S-vaccinated mice (p ¼ 0.4) and reduced relative to the PBS control (p ¼ 0.02).
TextSentencer_T161 28148-28285 Sentence denotes The presence of SARS-CoV replication in the lungs of control and vaccinated animals was confirmed by in situ hybridization ( Figure 4C ).
T44697 28148-28285 Sentence denotes The presence of SARS-CoV replication in the lungs of control and vaccinated animals was confirmed by in situ hybridization ( Figure 4C ).
TextSentencer_T162 28286-28427 Sentence denotes SARS-CoV N-specific riboprobe was hybridized to lung sections of mice from PBS-, VRP-N-, VRP-S-, and VRP-SþN-vaccinated groups ( Figure 4C ).
T61111 28286-28427 Sentence denotes SARS-CoV N-specific riboprobe was hybridized to lung sections of mice from PBS-, VRP-N-, VRP-S-, and VRP-SþN-vaccinated groups ( Figure 4C ).
TextSentencer_T163 28428-28683 Sentence denotes All tested lung sections from PBS mocks (unpublished data) and VRP-N-vaccinated ( Figure 4C , image a) animals exhibited in situ signal (arrows), although the signal did not appear to be as intense as that of the icSARS-CoV-infected animals ( Figure 2C ).
T21547 28428-28683 Sentence denotes All tested lung sections from PBS mocks (unpublished data) and VRP-N-vaccinated ( Figure 4C , image a) animals exhibited in situ signal (arrows), although the signal did not appear to be as intense as that of the icSARS-CoV-infected animals ( Figure 2C ).
TextSentencer_T164 28684-28815 Sentence denotes Lungs of the VRP-S-vaccinated animals ( Figure 4C , image b) had two of five slides exhibiting SARS-CoV-specific signal above back-
T76469 28684-28815 Sentence denotes Lungs of the VRP-S-vaccinated animals ( Figure 4C , image b) had two of five slides exhibiting SARS-CoV-specific signal above back-
TextSentencer_T165 28817-29087 Sentence denotes Because neutralizing antibody has been reported to confer protection from SARS-CoV replication within the lungs of mice [13, 16, 28] , it was of interest to determine whether the VRP-S vaccine established high neutralizing antibody levels that persisted until challenge.
T68330 28817-29087 Sentence denotes Because neutralizing antibody has been reported to confer protection from SARS-CoV replication within the lungs of mice [13, 16, 28] , it was of interest to determine whether the VRP-S vaccine established high neutralizing antibody levels that persisted until challenge.
TextSentencer_T166 29088-29351 Sentence denotes Plaque reduction neutralization titer (PRNT) of serum samples harvested prior to vaccination showed no neutralization of icSARS-CoV; similar results were noted from serum collected from mice vaccinated with the negative controls, VRP-HA or PBS (unpublished data).
T26996 29088-29351 Sentence denotes Plaque reduction neutralization titer (PRNT) of serum samples harvested prior to vaccination showed no neutralization of icSARS-CoV; similar results were noted from serum collected from mice vaccinated with the negative controls, VRP-HA or PBS (unpublished data).
TextSentencer_T167 29352-29637 Sentence denotes The 80% PRNT values (PRNT 80 ), the dilution of serum at which plaque numbers are reduced by 80% relative to virus treated with control sera, for VRP-S-and VRP-SþNvaccinated animals at 5 and 53 wk post-boost against both icSARS-CoV and icGDO3-S were compared ( Figure 5A ; Table S1 ).
T53506 29352-29637 Sentence denotes The 80% PRNT values (PRNT 80 ), the dilution of serum at which plaque numbers are reduced by 80% relative to virus treated with control sera, for VRP-S-and VRP-SþNvaccinated animals at 5 and 53 wk post-boost against both icSARS-CoV and icGDO3-S were compared ( Figure 5A ; Table S1 ).
TextSentencer_T168 29638-29789 Sentence denotes The mean reciprocal dilutions for the PRNT 80 of VRP-S and VRP-SþN against icSARS were measured at 796 6 307 at 5 wk post-boost and 628 6 363 at 53 wk.
T94052 29638-29789 Sentence denotes The mean reciprocal dilutions for the PRNT 80 of VRP-S and VRP-SþN against icSARS were measured at 796 6 307 at 5 wk post-boost and 628 6 363 at 53 wk.
TextSentencer_T169 29790-29934 Sentence denotes Sera from mice vaccinated with the combination of VRP-SþN had mean reciprocal PRNT 80 of 1,091 6 361 and 370 6 179 at 5 and 53 wk, respectively.
T25805 29790-29934 Sentence denotes Sera from mice vaccinated with the combination of VRP-SþN had mean reciprocal PRNT 80 of 1,091 6 361 and 370 6 179 at 5 and 53 wk, respectively.
TextSentencer_T170 29935-30339 Sentence denotes The initial neutralizing response in animals vaccinated with VRP-S and VRP-SþN was similar (p ¼ 0.2 at 5 wk post-boost), and although there was not significant waning of the icSARS-neutralizing activity over the 48-wk period in the VRP-S-vaccinated animals (p ¼ 0.3 Wilcoxin matched pairs signed-rank test), VRP-SþN serum was diminished by about 3-fold (p ¼ 0.03 Wilcoxin matched pairs signed-rank test).
T38724 29935-30339 Sentence denotes The initial neutralizing response in animals vaccinated with VRP-S and VRP-SþN was similar (p ¼ 0.2 at 5 wk post-boost), and although there was not significant waning of the icSARS-neutralizing activity over the 48-wk period in the VRP-S-vaccinated animals (p ¼ 0.3 Wilcoxin matched pairs signed-rank test), VRP-SþN serum was diminished by about 3-fold (p ¼ 0.03 Wilcoxin matched pairs signed-rank test).
TextSentencer_T171 30340-30512 Sentence denotes All tested sera remained above the lower limit of detection (1:100) and were sufficient to prevent icSARS replication within the lungs of challenged animals ( Figure 2B ).
T22602 30340-30512 Sentence denotes All tested sera remained above the lower limit of detection (1:100) and were sufficient to prevent icSARS replication within the lungs of challenged animals ( Figure 2B ).
TextSentencer_T172 30513-30710 Sentence denotes The neutralizing activity of sera from the vaccinated animals was more effective against the vaccine strain than against heterologous icGDO3-S virus for both sera harvests and vaccine combinations.
T12228 30513-30710 Sentence denotes The neutralizing activity of sera from the vaccinated animals was more effective against the vaccine strain than against heterologous icGDO3-S virus for both sera harvests and vaccine combinations.
TextSentencer_T173 30711-30941 Sentence denotes The reciprocal dilutions for the PRNT 80 of the VRP-S samples at 5 wk post-boost were below the limit of detection, whereas two samples of the week 53 bleed were measured above the limit of detection with a mean value of 112 6 28.
T16021 30711-30941 Sentence denotes The reciprocal dilutions for the PRNT 80 of the VRP-S samples at 5 wk post-boost were below the limit of detection, whereas two samples of the week 53 bleed were measured above the limit of detection with a mean value of 112 6 28.
TextSentencer_T174 30942-31253 Sentence denotes The icGDO3-S PRNT 80 measurements for VRP-SþN at weeks 5 and 53 post-boost were below the limit of detection with one exception for each time point: one mouse was measured to have a PRNT 80 of 124 at 5 wk, for an average titer of 103 6 8, and another a PRNT 80 of 107 at 53 wk post-boost, an average of 101 6 3.
T11185 30942-31253 Sentence denotes The icGDO3-S PRNT 80 measurements for VRP-SþN at weeks 5 and 53 post-boost were below the limit of detection with one exception for each time point: one mouse was measured to have a PRNT 80 of 124 at 5 wk, for an average titer of 103 6 8, and another a PRNT 80 of 107 at 53 wk post-boost, an average of 101 6 3.
TextSentencer_T175 31254-31688 Sentence denotes Given that the VRP-S vaccine's ability to provide long-term protection was likely due to the strong SARS-CoV-neutralizing response induced in vaccinated mice, we measured the PRNT 80 of the VRP-S immune sera from mice vaccinated when old (Table 1 , experiment 4) to determine if the incomplete protection seen in that study could be linked to a reduced neutralizing antibody response in the senescent animals ( Figure 5B ; Table S1 ).
T81307 31254-31688 Sentence denotes Given that the VRP-S vaccine's ability to provide long-term protection was likely due to the strong SARS-CoV-neutralizing response induced in vaccinated mice, we measured the PRNT 80 of the VRP-S immune sera from mice vaccinated when old (Table 1 , experiment 4) to determine if the incomplete protection seen in that study could be linked to a reduced neutralizing antibody response in the senescent animals ( Figure 5B ; Table S1 ).
TextSentencer_T176 31689-31948 Sentence denotes Against the vaccine strain, the reciprocal dilutions of the mean PRNT 80 were 170 6 82 with two of six samples falling below the limit of detection for VRP-S mice at 12 wk and 142 6 65 at 29 wk post-boost with four of six falling below the limit of detection.
T4470 31689-31948 Sentence denotes Against the vaccine strain, the reciprocal dilutions of the mean PRNT 80 were 170 6 82 with two of six samples falling below the limit of detection for VRP-S mice at 12 wk and 142 6 65 at 29 wk post-boost with four of six falling below the limit of detection.
TextSentencer_T177 31949-32172 Sentence denotes Sera from the VRP-SþN-inoculated mice had PRNT 80 values falling near or below the limit of detection with only one measurable sample at 114, for an average of 103 6 6 and no icSARS-neutralizing ability detected at week 29.
T21017 31949-32172 Sentence denotes Sera from the VRP-SþN-inoculated mice had PRNT 80 values falling near or below the limit of detection with only one measurable sample at 114, for an average of 103 6 6 and no icSARS-neutralizing ability detected at week 29.
TextSentencer_T178 32173-32412 Sentence denotes Against icGDO3-S, the VRP-S PRNT 80 values were below the limit of detection with the exception of a single VRP-S-vaccinated animal showing a neutralizing titer of 179 at 12 wk post-boost, for an average dilution of 116 6 35 for the group.
T43561 32173-32412 Sentence denotes Against icGDO3-S, the VRP-S PRNT 80 values were below the limit of detection with the exception of a single VRP-S-vaccinated animal showing a neutralizing titer of 179 at 12 wk post-boost, for an average dilution of 116 6 35 for the group.
TextSentencer_T179 32413-32717 Sentence denotes Sera harvested from these animals exhibited a marked reduction in neutralizing ability when compared to the response in animals vaccinated when young, even against the vaccine strain (p ¼ 0.008 for VRP-S week 5 versus VPP-S week 12 post-boost; p ¼ 0.006 VRP-SþN week 5 versus VRP-SþN week 12 post-boost).
T35330 32413-32717 Sentence denotes Sera harvested from these animals exhibited a marked reduction in neutralizing ability when compared to the response in animals vaccinated when young, even against the vaccine strain (p ¼ 0.008 for VRP-S week 5 versus VPP-S week 12 post-boost; p ¼ 0.006 VRP-SþN week 5 versus VRP-SþN week 12 post-boost).
TextSentencer_T180 32718-32835 Sentence denotes A strong anti-SARS-CoV neutralizing response was not induced by the VRP vaccines when administered to senescent mice.
T16482 32718-32835 Sentence denotes A strong anti-SARS-CoV neutralizing response was not induced by the VRP vaccines when administered to senescent mice.
TextSentencer_T181 32836-33026 Sentence denotes ELISAs for total IgG specific for SARS-S and influenza-HA were performed to compare the VRP vaccines' ability to induce antibody to those antigens in mice vaccinated when young or senescent.
T38387 32836-33026 Sentence denotes ELISAs for total IgG specific for SARS-S and influenza-HA were performed to compare the VRP vaccines' ability to induce antibody to those antigens in mice vaccinated when young or senescent.
TextSentencer_T182 33027-33078 Sentence denotes ELISA for SARS-S was performed on sera Figure 6A ).
T87901 33027-33078 Sentence denotes ELISA for SARS-S was performed on sera Figure 6A ).
TextSentencer_T183 33079-33323 Sentence denotes Mice vaccinated young with VRP-S (experiment 2) had an average log 10 half-maximum ELISA titer of 2.6 6 0.6 at 53 wk postboost, whereas that of the senescent animals was approximately a log lower at 1.5 6 0.9 at 29 weeks post-boost (p ¼ 0.007).
T98007 33079-33323 Sentence denotes Mice vaccinated young with VRP-S (experiment 2) had an average log 10 half-maximum ELISA titer of 2.6 6 0.6 at 53 wk postboost, whereas that of the senescent animals was approximately a log lower at 1.5 6 0.9 at 29 weeks post-boost (p ¼ 0.007).
TextSentencer_T184 33324-33457 Sentence denotes The difference between animals of the two age groups was even more striking when anti-S IgG levels were compared in the VRP-SþN mice.
T92920 33324-33457 Sentence denotes The difference between animals of the two age groups was even more striking when anti-S IgG levels were compared in the VRP-SþN mice.
TextSentencer_T185 33458-33647 Sentence denotes The animals vaccinated young with VRP-SþN had an average titer of 2.6 6 0.3, whereas the average for senescent animals was at the limit of detection of 0.02 log 10 half-maximum ELISA titer.
T9137 33458-33647 Sentence denotes The animals vaccinated young with VRP-SþN had an average titer of 2.6 6 0.3, whereas the average for senescent animals was at the limit of detection of 0.02 log 10 half-maximum ELISA titer.
TextSentencer_T186 33648-33848 Sentence denotes To verify that the reduced ability of the senescent animals to mount specific antibody responses was not limited to the SARS-S antigen; anti-HA IgG titers were compared in mice vaccinated with VRP-HA.
T51135 33648-33848 Sentence denotes To verify that the reduced ability of the senescent animals to mount specific antibody responses was not limited to the SARS-S antigen; anti-HA IgG titers were compared in mice vaccinated with VRP-HA.
TextSentencer_T187 33849-34054 Sentence denotes The average anti-HA titer of animals vaccinated when young was 4.7 6 0.2, whereas the titer of the vaccinated senescent mice was approximately one-tenth the size with a mean titer of 3.7 6 0.8 (p , 0.001).
T36595 33849-34054 Sentence denotes The average anti-HA titer of animals vaccinated when young was 4.7 6 0.2, whereas the titer of the vaccinated senescent mice was approximately one-tenth the size with a mean titer of 3.7 6 0.8 (p , 0.001).
TextSentencer_T188 34055-34210 Sentence denotes The reduced ability of sera harvested from senescent animals to neutralize virus correlates to a general reduction in antigen-specific antibody production.
T46573 34055-34210 Sentence denotes The reduced ability of sera harvested from senescent animals to neutralize virus correlates to a general reduction in antigen-specific antibody production.
TextSentencer_T189 34211-34376 Sentence denotes Lungs from the vaccinated senescent mice challenge studies (Table 1 , experiments 2 and 4) were sectioned, hematoxylin and eosin stained, and analyzed for pathology.
T91234 34211-34376 Sentence denotes Lungs from the vaccinated senescent mice challenge studies (Table 1 , experiments 2 and 4) were sectioned, hematoxylin and eosin stained, and analyzed for pathology.
TextSentencer_T190 34377-34582 Sentence denotes Though there was some animal-to-animal variation, in general only minor inflammatory changes were observed in the senescent control mice challenged with either icSARS or icGDO3-S (Figures 7C, 8A, and 8B) .
T49315 34377-34582 Sentence denotes Though there was some animal-to-animal variation, in general only minor inflammatory changes were observed in the senescent control mice challenged with either icSARS or icGDO3-S (Figures 7C, 8A, and 8B) .
TextSentencer_T191 34583-34991 Sentence denotes However, following SARS-CoV challenge in senescent mice, the N-vaccinated groups exhibited more marked bronchiolitis and alveolitis, as well as a conspicuous perivascular and peribronchiolar interstitial accumulation of numerous mononuclear leukocytes (mainly lymphocytes and plasma cells; i.e., lymphoplasmacytic cuffing) and increased numbers of widely scattered eosinophils ( Figures 7B, 7C, 8C, and 8D) .
T62349 34583-34991 Sentence denotes However, following SARS-CoV challenge in senescent mice, the N-vaccinated groups exhibited more marked bronchiolitis and alveolitis, as well as a conspicuous perivascular and peribronchiolar interstitial accumulation of numerous mononuclear leukocytes (mainly lymphocytes and plasma cells; i.e., lymphoplasmacytic cuffing) and increased numbers of widely scattered eosinophils ( Figures 7B, 7C, 8C, and 8D) .
TextSentencer_T192 34992-35294 Sentence denotes Upon SARS-CoV challenge, the animals vaccinated with SþN also exhibited a similar, but less severe, lymphoplasmacytic infiltration around pulmonary vessels and bronchiolar airways, although alveolitis and eosinophil infiltration were not a prominent feature in these animals ( Figures 7F, 8G, and 8H ).
T66974 34992-35294 Sentence denotes Upon SARS-CoV challenge, the animals vaccinated with SþN also exhibited a similar, but less severe, lymphoplasmacytic infiltration around pulmonary vessels and bronchiolar airways, although alveolitis and eosinophil infiltration were not a prominent feature in these animals ( Figures 7F, 8G, and 8H ).
TextSentencer_T193 35295-35467 Sentence denotes The lungs of animals vaccinated with VRP-S were similar to VRP-HAor PBS mock-vaccinated animals with minimal lymphoplasmacytic cell accumulations (Figures 7E, 8E, and 8F ).
T12467 35295-35467 Sentence denotes The lungs of animals vaccinated with VRP-S were similar to VRP-HAor PBS mock-vaccinated animals with minimal lymphoplasmacytic cell accumulations (Figures 7E, 8E, and 8F ).
TextSentencer_T194 35468-35683 Sentence denotes Therefore, not only did N vaccination fail to control SARS-CoV replication within the lungs, but N vaccination also resulted in an enhanced immunopathology in the lungs of the senescent animals upon viral challenge.
T32023 35468-35683 Sentence denotes Therefore, not only did N vaccination fail to control SARS-CoV replication within the lungs, but N vaccination also resulted in an enhanced immunopathology in the lungs of the senescent animals upon viral challenge.
TextSentencer_T195 35684-35930 Sentence denotes In order to determine the kinetics of the VRP-N-associated immunopathology and whether this effect was age dependent, young and old VRP-N-or VRP-HA-vaccinated mice were challenged with icSARS and sacrificed on days 2, 4, 7, and 14 post-challenge.
T97558 35684-35930 Sentence denotes In order to determine the kinetics of the VRP-N-associated immunopathology and whether this effect was age dependent, young and old VRP-N-or VRP-HA-vaccinated mice were challenged with icSARS and sacrificed on days 2, 4, 7, and 14 post-challenge.
TextSentencer_T196 35931-36132 Sentence denotes Young (8 wk of age) or senescent (53 wk of age) female BALB/c mice were vaccinated with 10 6 IU of VRP-N or VRP-HA, boosted 7 wk later, then i.n. challenged with 10 5 pfu of icSARS-CoV 4 wk post-boost.
T40641 35931-36132 Sentence denotes Young (8 wk of age) or senescent (53 wk of age) female BALB/c mice were vaccinated with 10 6 IU of VRP-N or VRP-HA, boosted 7 wk later, then i.n. challenged with 10 5 pfu of icSARS-CoV 4 wk post-boost.
TextSentencer_T197 36133-36262 Sentence denotes Lungs were harvested on days 2, 4, 7, and 14, titered, and processed for histology (Summarized in Table 1 , experiments 5 and 6).
T27962 36133-36262 Sentence denotes Lungs were harvested on days 2, 4, 7, and 14, titered, and processed for histology (Summarized in Table 1 , experiments 5 and 6).
TextSentencer_T198 36263-36417 Sentence denotes In young mice, day 2 average lung titers were 7.5 6 0.2 log 10 pfu/ g and 8.1 6 0.1 log 10 pfu/g for the VRP-N-and VRP-HAvaccinated animals, respectively.
T67365 36263-36417 Sentence denotes In young mice, day 2 average lung titers were 7.5 6 0.2 log 10 pfu/ g and 8.1 6 0.1 log 10 pfu/g for the VRP-N-and VRP-HAvaccinated animals, respectively.
TextSentencer_T199 36418-36542 Sentence denotes Day 4 average titers were 5.5 6 0.3 log 10 pfu/g for VRP-N-vaccinated and 5.7 6 0.1 log 10 pfu/g for VRP-HA-vaccinated mice.
T97292 36418-36542 Sentence denotes Day 4 average titers were 5.5 6 0.3 log 10 pfu/g for VRP-N-vaccinated and 5.7 6 0.1 log 10 pfu/g for VRP-HA-vaccinated mice.
TextSentencer_T200 36543-36756 Sentence denotes SARS titers in the VRP-N-vaccinated mice lungs had dropped to the limit of detection by day 7, with two of the three lungs from VRP-HA-vaccinated mice showing measurable titers at a mean of 3.2 6 0.3 log 10 pfu/g.
T18021 36543-36756 Sentence denotes SARS titers in the VRP-N-vaccinated mice lungs had dropped to the limit of detection by day 7, with two of the three lungs from VRP-HA-vaccinated mice showing measurable titers at a mean of 3.2 6 0.3 log 10 pfu/g.
TextSentencer_T201 36757-36807 Sentence denotes By day 14, virus was undetectable in either group.
T73989 36757-36807 Sentence denotes By day 14, virus was undetectable in either group.
TextSentencer_T202 36808-36959 Sentence denotes In senescent animals, the day 2 average lung titers were 8.2 6 0.2 and 8.5 6 0.1 log 10 pfu/g for the VRP-Nand VRP-HA-vaccinated animals, respectively.
T5431 36808-36959 Sentence denotes In senescent animals, the day 2 average lung titers were 8.2 6 0.2 and 8.5 6 0.1 log 10 pfu/g for the VRP-Nand VRP-HA-vaccinated animals, respectively.
TextSentencer_T203 36960-37103 Sentence denotes By day 4, the mean titers had dropped to 5.4 6 0.6 log 10 pfu/g for VRP-Nvaccinated animals and 5.7 6 0.5 log 10 pfu/g for the VRP-HA controls.
T87110 36960-37103 Sentence denotes By day 4, the mean titers had dropped to 5.4 6 0.6 log 10 pfu/g for VRP-Nvaccinated animals and 5.7 6 0.5 log 10 pfu/g for the VRP-HA controls.
TextSentencer_T204 37104-37302 Sentence denotes On day 7, one VRP-N-vaccinated mouse had a detectable titer of 3.7 log 10 pfu/g, whereas the lungs of two VRP-HA-vaccinated mice were positive for replication with a titer of 3.7 6 0.5 log 10 pfu/g.
T41975 37104-37302 Sentence denotes On day 7, one VRP-N-vaccinated mouse had a detectable titer of 3.7 log 10 pfu/g, whereas the lungs of two VRP-HA-vaccinated mice were positive for replication with a titer of 3.7 6 0.5 log 10 pfu/g.
TextSentencer_T205 37303-37366 Sentence denotes No virus was detected in either group on day 14 post-challenge.
T6951 37303-37366 Sentence denotes No virus was detected in either group on day 14 post-challenge.
TextSentencer_T206 37367-37708 Sentence denotes Though enhanced inflammation was observed in a subset of the VRP-N-vaccinated animals at day 2 post-infection ( Figure 9B ), the inflammatory infiltrates were readily apparent in both young and senescent animals at day 4 postinfection ( Figure 9D and unpublished data) and were maintained at days 7 and 14 post-infection ( Figure 9F and 9H).
T85394 37367-37708 Sentence denotes Though enhanced inflammation was observed in a subset of the VRP-N-vaccinated animals at day 2 post-infection ( Figure 9B ), the inflammatory infiltrates were readily apparent in both young and senescent animals at day 4 postinfection ( Figure 9D and unpublished data) and were maintained at days 7 and 14 post-infection ( Figure 9F and 9H).
TextSentencer_T207 37709-37832 Sentence denotes As had been noted previously, VRP-HA-and VRP-Nvaccinated animals also differed by the presence of eosinophils (Figure 10 ).
T85006 37709-37832 Sentence denotes As had been noted previously, VRP-HA-and VRP-Nvaccinated animals also differed by the presence of eosinophils (Figure 10 ).
TextSentencer_T208 37833-37966 Sentence denotes At day 2 post-infection, eosinophils were rarely seen in either VRP-HA-or VRP-N-vaccinated mice ( Figure 10A and 10B, respectively) .
T27528 37833-37966 Sentence denotes At day 2 post-infection, eosinophils were rarely seen in either VRP-HA-or VRP-N-vaccinated mice ( Figure 10A and 10B, respectively) .
TextSentencer_T209 37967-38159 Sentence denotes In VRP-N-vaccinated animals, but not HA-vaccinated animals, eosinophils were widespread on days 4 and 7 ( Figure 10D and 10F) , but had largely cleared from the lungs by day 14 ( Figure 10H ).
T54054 37967-38159 Sentence denotes In VRP-N-vaccinated animals, but not HA-vaccinated animals, eosinophils were widespread on days 4 and 7 ( Figure 10D and 10F) , but had largely cleared from the lungs by day 14 ( Figure 10H ).
TextSentencer_T210 38160-38369 Sentence denotes There was no apparent difference in severity between the young and senescent animals, suggesting that the immune pathology was specific to pre-existing N immunity, but was not age dependent (unpublished data).
T78306 38160-38369 Sentence denotes There was no apparent difference in severity between the young and senescent animals, suggesting that the immune pathology was specific to pre-existing N immunity, but was not age dependent (unpublished data).
TextSentencer_T211 38370-38641 Sentence denotes A passive transfer of anti-N, anti-HA, or anti-S sera into naive mice was performed to determine if the increased inflammatory response could be attributed to N-specific antibody, and to confirm that protection in S-vaccinated animals was mediated by S-specific antibody.
T57333 38370-38641 Sentence denotes A passive transfer of anti-N, anti-HA, or anti-S sera into naive mice was performed to determine if the increased inflammatory response could be attributed to N-specific antibody, and to confirm that protection in S-vaccinated animals was mediated by S-specific antibody.
TextSentencer_T212 38642-38932 Sentence denotes Hyperimmune sera against VRP-HA (5.26 log 10 OD ¼ 0.2 ELISA titer), VRP-N (5.6 log 10 OD ¼ 0.2 ELISA titer), or VRP-S (5.5 OD ¼ 0.2 ELISA titer) were intravenously transferred in 150-ll volumes to groups of 8-wk-old or 43-wk-old naive BALB/c mice prior to challenge with 10 5 pfu of icSARS.
T90732 38642-38932 Sentence denotes Hyperimmune sera against VRP-HA (5.26 log 10 OD ¼ 0.2 ELISA titer), VRP-N (5.6 log 10 OD ¼ 0.2 ELISA titer), or VRP-S (5.5 OD ¼ 0.2 ELISA titer) were intravenously transferred in 150-ll volumes to groups of 8-wk-old or 43-wk-old naive BALB/c mice prior to challenge with 10 5 pfu of icSARS.
TextSentencer_T213 38933-39046 Sentence denotes Prior to challenge, serum was collected from mice and antigen-specific IgG titered to verify successful transfer.
T73160 38933-39046 Sentence denotes Prior to challenge, serum was collected from mice and antigen-specific IgG titered to verify successful transfer.
TextSentencer_T214 39047-39222 Sentence denotes Young mice receiving anti-VRP-HA (n ¼ 4) had a mean ELISA serum titer of 2.8 6 2.4 log 10 OD ¼ 0.2, whereas senescent mice (n ¼ 4) had a 2.9 6 1.8 log 10 OD ¼ 0.2 ELISA titer.
T76314 39047-39222 Sentence denotes Young mice receiving anti-VRP-HA (n ¼ 4) had a mean ELISA serum titer of 2.8 6 2.4 log 10 OD ¼ 0.2, whereas senescent mice (n ¼ 4) had a 2.9 6 1.8 log 10 OD ¼ 0.2 ELISA titer.
TextSentencer_T215 39223-39413 Sentence denotes The average serum titers for mice injected with anti-VRP-N were 2.7 6 2.4 log 10 OD ¼ 0.2 ELISA titer in young (n ¼ 4) and 2.3 6 1.9 log 10 OD ¼ 0.2 ELISA titer in senescent animals (n ¼ 4).
T21646 39223-39413 Sentence denotes The average serum titers for mice injected with anti-VRP-N were 2.7 6 2.4 log 10 OD ¼ 0.2 ELISA titer in young (n ¼ 4) and 2.3 6 1.9 log 10 OD ¼ 0.2 ELISA titer in senescent animals (n ¼ 4).
TextSentencer_T216 39414-39538 Sentence denotes The anti-VRP-S log 10 OD ¼ 0.2 ELISA titers were 3.1 6 3.1 in young animals (n ¼ 3) and 2.9 6 2.3 in senescent mice (n ¼ 3).
T38623 39414-39538 Sentence denotes The anti-VRP-S log 10 OD ¼ 0.2 ELISA titers were 3.1 6 3.1 in young animals (n ¼ 3) and 2.9 6 2.3 in senescent mice (n ¼ 3).
TextSentencer_T217 39539-39644 Sentence denotes Lungs were harvested 4 d postchallenge, virus titers determined, and processed for histological analysis.
T26098 39539-39644 Sentence denotes Lungs were harvested 4 d postchallenge, virus titers determined, and processed for histological analysis.
TextSentencer_T218 39645-39857 Sentence denotes Virus titers in the young mice were 5.0 6 0.1 log 10 pfu/g in the anti-VRP-HA group, 5.2 6 0.1 in the anti VRP-N group, and below the limit of detection (2.4 log 10 pfu/g) in animals injected with the anti-VRP-S.
T37345 39645-39857 Sentence denotes Virus titers in the young mice were 5.0 6 0.1 log 10 pfu/g in the anti-VRP-HA group, 5.2 6 0.1 in the anti VRP-N group, and below the limit of detection (2.4 log 10 pfu/g) in animals injected with the anti-VRP-S.
TextSentencer_T219 39858-40056 Sentence denotes Titers in the senescent mice were 5.3 6 0.3 log 10 pfu/g in the anti-VRP-HA group, 5.6 6 0.6 in animals receiving the anti-VRP-N, and below the limit of detection in mice inoculated with anti-VRP-S.
T892 39858-40056 Sentence denotes Titers in the senescent mice were 5.3 6 0.3 log 10 pfu/g in the anti-VRP-HA group, 5.6 6 0.6 in animals receiving the anti-VRP-N, and below the limit of detection in mice inoculated with anti-VRP-S.
TextSentencer_T220 40057-40262 Sentence denotes None of the mice displayed the enhanced inflammation noted in VRP-N-vaccinated animals (unpublished data), indicating that the observed immunopathology was not the result of antibody-dependent enhancement.
T69595 40057-40262 Sentence denotes None of the mice displayed the enhanced inflammation noted in VRP-N-vaccinated animals (unpublished data), indicating that the observed immunopathology was not the result of antibody-dependent enhancement.
TextSentencer_T221 40263-40510 Sentence denotes VRP vaccine vectors induce robust mucosal and cellular immune responses against a large number of foreign antigens [35, 51] and were evaluated as candidate vaccines against homologous and zoonotic SARS-CoV challenge in young and senescent animals.
T53649 40263-40510 Sentence denotes VRP vaccine vectors induce robust mucosal and cellular immune responses against a large number of foreign antigens [35, 51] and were evaluated as candidate vaccines against homologous and zoonotic SARS-CoV challenge in young and senescent animals.
TextSentencer_T222 40511-40664 Sentence denotes Inoculation of mice with VRP-S induced antibody that recognized the epidemic SARS-CoV S glycoprotein as well as the S of a highly divergent strain, GDO3.
T30763 40511-40664 Sentence denotes Inoculation of mice with VRP-S induced antibody that recognized the epidemic SARS-CoV S glycoprotein as well as the S of a highly divergent strain, GDO3.
TextSentencer_T223 40665-40894 Sentence denotes The VRP-S vaccine induced long-term protection against challenge with the vaccine strain, complete short-term protection against icGDO3-S challenge, and partial protection against the divergent virus in the senescent mouse model.
T77791 40665-40894 Sentence denotes The VRP-S vaccine induced long-term protection against challenge with the vaccine strain, complete short-term protection against icGDO3-S challenge, and partial protection against the divergent virus in the senescent mouse model.
TextSentencer_T224 40895-41154 Sentence denotes In contrast, vaccination with VRP-N failed to inhibit viral replication within the lungs of either young or senescent animals, resulted in enhanced immunopathology following viral challenge, and did not provide any measurable benefit when combined with VRP-S.
T54359 40895-41154 Sentence denotes In contrast, vaccination with VRP-N failed to inhibit viral replication within the lungs of either young or senescent animals, resulted in enhanced immunopathology following viral challenge, and did not provide any measurable benefit when combined with VRP-S.
TextSentencer_T225 41155-41573 Sentence denotes The data suggest that vaccine regimens eliciting complete protection against antigenically heterologous forms of SARS-CoV in healthy individuals may not be sufficient for higher risk groups, including vulnerable (Table 1 , experiment 2) taken from an untreated control mouse (A), a VRP-Nvaccinated mouse (B) and (D), a VRP-HA-vaccinated mouse (C), a VRP-Svaccinated mouse (E), and a VRP-S-and VRP-N-treated mouse (F).
T49486 41155-41573 Sentence denotes The data suggest that vaccine regimens eliciting complete protection against antigenically heterologous forms of SARS-CoV in healthy individuals may not be sufficient for higher risk groups, including vulnerable (Table 1 , experiment 2) taken from an untreated control mouse (A), a VRP-Nvaccinated mouse (B) and (D), a VRP-HA-vaccinated mouse (C), a VRP-Svaccinated mouse (E), and a VRP-S-and VRP-N-treated mouse (F).
TextSentencer_T226 41574-41611 Sentence denotes No histopathology was evident in (A).
T25309 41574-41611 Sentence denotes No histopathology was evident in (A).
TextSentencer_T227 41612-41840 Sentence denotes A marked mixed inflammatory infiltrate composed mainly of mononuclear leukocytes (lymphocytes and plasma cells) and widely scattered eosinophils are evident in the perivascular and peribronchiolar interstitium (asterisk) in (B).
T42303 41612-41840 Sentence denotes A marked mixed inflammatory infiltrate composed mainly of mononuclear leukocytes (lymphocytes and plasma cells) and widely scattered eosinophils are evident in the perivascular and peribronchiolar interstitium (asterisk) in (B).
TextSentencer_T228 41841-42001 Sentence denotes Similar inflammatory cells are also present in bronchiolar (br) airways and alveolar airspaces along with enlarged and vacuolated alveolar macrophages (arrows).
T85114 41841-42001 Sentence denotes Similar inflammatory cells are also present in bronchiolar (br) airways and alveolar airspaces along with enlarged and vacuolated alveolar macrophages (arrows).
TextSentencer_T229 42002-42224 Sentence denotes The box in (B) denotes the site of the light photomicrograph (D) that was taken at a higher magnification to better illustrate the lymphoplasmacytic inflammatory cell infiltrate with lesser numbers of eosinophils (arrows).
T91479 42002-42224 Sentence denotes The box in (B) denotes the site of the light photomicrograph (D) that was taken at a higher magnification to better illustrate the lymphoplasmacytic inflammatory cell infiltrate with lesser numbers of eosinophils (arrows).
TextSentencer_T230 42225-42370 Sentence denotes Similar, but slightly less severe, perivascular inflammatory infiltrates (asterisk) are also present in (F), but without accompanying alveolitis.
T9415 42225-42370 Sentence denotes Similar, but slightly less severe, perivascular inflammatory infiltrates (asterisk) are also present in (F), but without accompanying alveolitis.
TextSentencer_T231 42371-42478 Sentence denotes Minimal lymphoplasmacytic cell accumulations around the pulmonary arteriole (a) are evident in (C) and (E).
T21628 42371-42478 Sentence denotes Minimal lymphoplasmacytic cell accumulations around the pulmonary arteriole (a) are evident in (C) and (E).
TextSentencer_T232 42479-42531 Sentence denotes All tissues were stained with hematoxylin and eosin.
T82075 42479-42531 Sentence denotes All tissues were stained with hematoxylin and eosin.
TextSentencer_T233 42532-42846 Sentence denotes Bars denote the scale of the magnification. a, pulmonary arteriole; ap, alveolar parenchyma; br, bronchiolar lumen; e, surface epithelium of the bronchiole. doi:10.1371/journal.pmed.0030525.g007 elderly populations, and that there is a need for further testing of candidate vaccines that induce an anti-N response.
T24999 42532-42846 Sentence denotes Bars denote the scale of the magnification. a, pulmonary arteriole; ap, alveolar parenchyma; br, bronchiolar lumen; e, surface epithelium of the bronchiole. doi:10.1371/journal.pmed.0030525.g007 elderly populations, and that there is a need for further testing of candidate vaccines that induce an anti-N response.
TextSentencer_T234 42847-42925 Sentence denotes VRP-S vaccination generated a strong neutralizing antibody response (PRNT 80 .
T8984 42847-42925 Sentence denotes VRP-S vaccination generated a strong neutralizing antibody response (PRNT 80 .
TextSentencer_T235 42926-43039 Sentence denotes 1:600) that persisted for over a year and provided complete protection against challenge with the vaccine strain.
T59206 42926-43039 Sentence denotes 1:600) that persisted for over a year and provided complete protection against challenge with the vaccine strain.
TextSentencer_T236 43040-43153 Sentence denotes In humans, neutralization titers have been measured from 1:12 to 1:512 with a geometric mean titer of 1:61 [52] .
T57531 43040-43153 Sentence denotes In humans, neutralization titers have been measured from 1:12 to 1:512 with a geometric mean titer of 1:61 [52] .
TextSentencer_T237 43154-43310 Sentence denotes In a study evaluating an inactivated virus vaccine, neutralizing antibody titers greater than 1:114 resulted in complete protection against challenge [21] .
T44485 43154-43310 Sentence denotes In a study evaluating an inactivated virus vaccine, neutralizing antibody titers greater than 1:114 resulted in complete protection against challenge [21] .
TextSentencer_T238 43311-43539 Sentence denotes Mice inoculated with vesicular stomatitis virus vectors expressing SARS-S developed lower average neutralizing titers of 1:32, which were nevertheless protective against SARS-CoV infection for up to 4 mo after vaccination [17] .
T65085 43311-43539 Sentence denotes Mice inoculated with vesicular stomatitis virus vectors expressing SARS-S developed lower average neutralizing titers of 1:32, which were nevertheless protective against SARS-CoV infection for up to 4 mo after vaccination [17] .
TextSentencer_T239 43540-43585 Sentence denotes We followed animals for over 1 y after boost.
T4296 43540-43585 Sentence denotes We followed animals for over 1 y after boost.
TextSentencer_T240 43586-43700 Sentence denotes To our knowledge, these are the first assays illustrating waning immune responses to a SARS-CoV candidate vaccine.
T89507 43586-43700 Sentence denotes To our knowledge, these are the first assays illustrating waning immune responses to a SARS-CoV candidate vaccine.
TextSentencer_T241 43701-43957 Sentence denotes On average, mice vaccinated when young with VRP-S did not show a significant reduction in neutralizing titers up to 53 wk post-boost, whereas mice vaccinated with the combination of VRP-SþN experienced about a 3-fold reduction over the same period of time.
T30405 43701-43957 Sentence denotes On average, mice vaccinated when young with VRP-S did not show a significant reduction in neutralizing titers up to 53 wk post-boost, whereas mice vaccinated with the combination of VRP-SþN experienced about a 3-fold reduction over the same period of time.
TextSentencer_T242 43958-44296 Sentence denotes Although it is problematic to compare our neutralizing antibody titers to those induced by other SARS vaccines due to the use of different assays, we demonstrate protection from challenge with either vaccine or heterologous challenge virus strains in animals with an icSARS PRNT 80 greater than 1:114, near the assay's limit of detection.
T54936 43958-44296 Sentence denotes Although it is problematic to compare our neutralizing antibody titers to those induced by other SARS vaccines due to the use of different assays, we demonstrate protection from challenge with either vaccine or heterologous challenge virus strains in animals with an icSARS PRNT 80 greater than 1:114, near the assay's limit of detection.
TextSentencer_T243 44297-44473 Sentence denotes Vaccines that induce robust neutralizing titers against the homologous strain will likely confer protection against zoonotic reintroductions, especially in younger populations.
T30170 44297-44473 Sentence denotes Vaccines that induce robust neutralizing titers against the homologous strain will likely confer protection against zoonotic reintroductions, especially in younger populations.
TextSentencer_T244 44474-44824 Sentence denotes As reported with other SARS-N-expressing DNA and vectored vaccines [15, 24] , VRP-N did not protect mice from SARS-CoV replication, and no benefit to vaccination with a cocktail of both VRP-S and VRP-N was observed, although an approximately half-log reduction in viral titers within the lungs of some VRP-N-vaccinated mice was occasionally observed.
T37236 44474-44824 Sentence denotes As reported with other SARS-N-expressing DNA and vectored vaccines [15, 24] , VRP-N did not protect mice from SARS-CoV replication, and no benefit to vaccination with a cocktail of both VRP-S and VRP-N was observed, although an approximately half-log reduction in viral titers within the lungs of some VRP-N-vaccinated mice was occasionally observed.
TextSentencer_T245 44825-45440 Sentence denotes Although any reduction in SARS-CoV titer can be interpreted as a positive aspect of a potential vaccine, given the relationship between viral titer and SARS disease severity [53, 54] , the increased number of lymphocytic and eosinophilic inflammatory infiltrates, which are also characteristic of the immune pathology observed with respiratory syncytial virus (RSV) infection following vaccination with formalininactivated RSV [55, 56] , raises concerns that vaccination with N alone will not only fail to effectively protect against SARS-CoV replication, but may result in vaccine-enhanced pulmonary disease [57] .
T85894 44825-45440 Sentence denotes Although any reduction in SARS-CoV titer can be interpreted as a positive aspect of a potential vaccine, given the relationship between viral titer and SARS disease severity [53, 54] , the increased number of lymphocytic and eosinophilic inflammatory infiltrates, which are also characteristic of the immune pathology observed with respiratory syncytial virus (RSV) infection following vaccination with formalininactivated RSV [55, 56] , raises concerns that vaccination with N alone will not only fail to effectively protect against SARS-CoV replication, but may result in vaccine-enhanced pulmonary disease [57] .
TextSentencer_T246 45441-45632 Sentence denotes N-induced pathology has not been previously reported, probably because most studies examined young mice at 2-3 d post-infection, prior to the infiltration of inflammatory cells into the lung.
T31490 45441-45632 Sentence denotes N-induced pathology has not been previously reported, probably because most studies examined young mice at 2-3 d post-infection, prior to the infiltration of inflammatory cells into the lung.
TextSentencer_T247 45633-45831 Sentence denotes VRP-N-induced pathology was clearly evident by day 4 and persisted for 1-2 wk following wild-type virus challenge, suggesting the potential for serious complications in lung physiology and function.
T68364 45633-45831 Sentence denotes VRP-N-induced pathology was clearly evident by day 4 and persisted for 1-2 wk following wild-type virus challenge, suggesting the potential for serious complications in lung physiology and function.
TextSentencer_T248 45832-46080 Sentence denotes This finding has particular significance for SARS-N and inactivated SARS-CoV vaccines currently under development that also induce anti-N antibody and T cell responses [12, 19, 21, 26, [58] [59] [60] [61] , because they may lead to adverse effects.
T38043 45832-46080 Sentence denotes This finding has particular significance for SARS-N and inactivated SARS-CoV vaccines currently under development that also induce anti-N antibody and T cell responses [12, 19, 21, 26, [58] [59] [60] [61] , because they may lead to adverse effects.
TextSentencer_T249 46081-46190 Sentence denotes Therefore, caution is merited with respect to the inclusion of SARS-CoV N protein in any vaccine formulation.
T39339 46081-46190 Sentence denotes Therefore, caution is merited with respect to the inclusion of SARS-CoV N protein in any vaccine formulation.
TextSentencer_T250 46191-46444 Sentence denotes The passive transfer of anti-N antibody did not contribute to inflammation and leads us to hypothesize that it is the activity of SARS-N-specific T cells in the absence of effective neutralizing anti-SARS-CoV antibody that mediates the adverse response.
T18467 46191-46444 Sentence denotes The passive transfer of anti-N antibody did not contribute to inflammation and leads us to hypothesize that it is the activity of SARS-N-specific T cells in the absence of effective neutralizing anti-SARS-CoV antibody that mediates the adverse response.
TextSentencer_T251 46445-46652 Sentence denotes It is interesting that a T h 2-skewed cytokine profile is a hallmark of the RSV vaccine-enhanced disease, which raises the possibility that the N-specific immune response is skewed in a similar manner [62] .
T67765 46445-46652 Sentence denotes It is interesting that a T h 2-skewed cytokine profile is a hallmark of the RSV vaccine-enhanced disease, which raises the possibility that the N-specific immune response is skewed in a similar manner [62] .
TextSentencer_T252 46653-46790 Sentence denotes SARS-CoV strain diversity was mostly confined to China where many human and animal isolates were not successfully cultured in vitro [8] .
T67657 46653-46790 Sentence denotes SARS-CoV strain diversity was mostly confined to China where many human and animal isolates were not successfully cultured in vitro [8] .
TextSentencer_T253 46791-46923 Sentence denotes Consequently, most available experimental strains, like Urbani, are nearly identical and do not reflect natural diversity [63, 64] .
T79462 46791-46923 Sentence denotes Consequently, most available experimental strains, like Urbani, are nearly identical and do not reflect natural diversity [63, 64] .
TextSentencer_T254 46924-47167 Sentence denotes Recent advances in synthetic biology used to reconstruct extinct viruses, or specific genes of those viruses, de novo from their nucleotide sequences [65] [66] [67] provide the means for expanding the number of available SARS-CoV test strains.
T41512 46924-47167 Sentence denotes Recent advances in synthetic biology used to reconstruct extinct viruses, or specific genes of those viruses, de novo from their nucleotide sequences [65] [66] [67] provide the means for expanding the number of available SARS-CoV test strains.
TextSentencer_T255 47168-47306 Sentence denotes Using a comprehensive SARS-CoV genetic database [8, 31] , we resurrected the divergent GDO3-S glycoprotein in the Urbani genetic backbone.
T62352 47168-47306 Sentence denotes Using a comprehensive SARS-CoV genetic database [8, 31] , we resurrected the divergent GDO3-S glycoprotein in the Urbani genetic backbone.
TextSentencer_T256 47307-47660 Sentence denotes The icGD03-S recombinant virus was identical to the molecular clone except for the presence of two mutations in S that likely evolved after transfection of full-length RNA and virus passage in Vero cells, similar to the cell culture adaptations reported in S for other SARS-CoV strains isolated from human clinical specimens and passaged in vitro [68] .
T43384 47307-47660 Sentence denotes The icGD03-S recombinant virus was identical to the molecular clone except for the presence of two mutations in S that likely evolved after transfection of full-length RNA and virus passage in Vero cells, similar to the cell culture adaptations reported in S for other SARS-CoV strains isolated from human clinical specimens and passaged in vitro [68] .
TextSentencer_T257 47661-47894 Sentence denotes The icGDO3-S CoV's sequence divergence from Urbani, efficient in vitro replication in HAE and Vero cultures, and robust in vivo replication in the mouse model, make it an excellent heterologous challenge inoculum for vaccine studies.
T1935 47661-47894 Sentence denotes The icGDO3-S CoV's sequence divergence from Urbani, efficient in vitro replication in HAE and Vero cultures, and robust in vivo replication in the mouse model, make it an excellent heterologous challenge inoculum for vaccine studies.
TextSentencer_T258 47895-48039 Sentence denotes The GD03 RBD is also present in many zoonotic isolates described in civets and raccoon dogs, supporting its use as a zoonotic model strain [8] .
T60527 47895-48039 Sentence denotes The GD03 RBD is also present in many zoonotic isolates described in civets and raccoon dogs, supporting its use as a zoonotic model strain [8] .
TextSentencer_T259 48040-48214 Sentence denotes Furthermore, the reduced replication in HAE cultures of icGDO3-S compared to Urbani-CoV is consistent with the reduced pathogenesis noted in the GDO3 human case [8, 53, 54] .
T71652 48040-48214 Sentence denotes Furthermore, the reduced replication in HAE cultures of icGDO3-S compared to Urbani-CoV is consistent with the reduced pathogenesis noted in the GDO3 human case [8, 53, 54] .
TextSentencer_T260 48215-48501 Sentence denotes Consistent with previous work comparing the susceptibility of pseudotyped lentiviruses bearing the S glycoproteins of various SARS strains to neutralization by anti-S (Urbani) IgG [31] , anti-VRP-S antibody demonstrated reduced neutralization of icGDO3-S relative to the vaccine strain.
T7237 48215-48501 Sentence denotes Consistent with previous work comparing the susceptibility of pseudotyped lentiviruses bearing the S glycoproteins of various SARS strains to neutralization by anti-S (Urbani) IgG [31] , anti-VRP-S antibody demonstrated reduced neutralization of icGDO3-S relative to the vaccine strain.
TextSentencer_T261 48502-48610 Sentence denotes In spite of this, the VRP-S vaccine successfully provided short-term protection against the divergent virus.
T38975 48502-48610 Sentence denotes In spite of this, the VRP-S vaccine successfully provided short-term protection against the divergent virus.
TextSentencer_T262 48611-48806 Sentence denotes Vaccination of senescent animals produced significantly reduced antibody responses compared with younger mice, and when challenged with the heterologous icGDO3-S virus, protection was incomplete.
T4339 48611-48806 Sentence denotes Vaccination of senescent animals produced significantly reduced antibody responses compared with younger mice, and when challenged with the heterologous icGDO3-S virus, protection was incomplete.
TextSentencer_T263 48807-48989 Sentence denotes However, any animal with a PRNT 80 value above 1:114 against icSARS showed reduced viral replication within the lungs following challenge with either the vaccine or icGDO3-S strains.
T21095 48807-48989 Sentence denotes However, any animal with a PRNT 80 value above 1:114 against icSARS showed reduced viral replication within the lungs following challenge with either the vaccine or icGDO3-S strains.
TextSentencer_T264 48990-49326 Sentence denotes As noted for the homologous challenge studies, the combination of VRP-SþN did not enhance protection from heterologous challenge, but may actually have weakened it, with senescent animals showing even lower anti-S antibody responses and an even higher rate of viral replication, albeit with reduced titers, and increased lung pathology.
T21913 48990-49326 Sentence denotes As noted for the homologous challenge studies, the combination of VRP-SþN did not enhance protection from heterologous challenge, but may actually have weakened it, with senescent animals showing even lower anti-S antibody responses and an even higher rate of viral replication, albeit with reduced titers, and increased lung pathology.
TextSentencer_T265 49327-49450 Sentence denotes One possible cause for vaccine failure is the emergence of an escape mutant in an environment of suboptimal neutralization.
T78970 49327-49450 Sentence denotes One possible cause for vaccine failure is the emergence of an escape mutant in an environment of suboptimal neutralization.
TextSentencer_T266 49451-49618 Sentence denotes However, initial data comparing the neutralization susceptibility of viruses isolated from these mice to the challenge stock refute this conjecture (unpublished data).
T70181 49451-49618 Sentence denotes However, initial data comparing the neutralization susceptibility of viruses isolated from these mice to the challenge stock refute this conjecture (unpublished data).
TextSentencer_T267 49619-49830 Sentence denotes Incomplete protection by a vaccine in immunosenescent animals and humans is well documented and is more likely the result of an age-related compromise in one or more stages of the immune response to the vaccine.
T10563 49619-49830 Sentence denotes Incomplete protection by a vaccine in immunosenescent animals and humans is well documented and is more likely the result of an age-related compromise in one or more stages of the immune response to the vaccine.
TextSentencer_T268 49831-50077 Sentence denotes For instance, antibody responses in the immunosenescent tend to offer less protection with limited switching to secondary isotypes, lower antibody levels in general, and production of antibody with lower affinities [69] [70] [71] [72] [73] [74] .
T63653 49831-50077 Sentence denotes For instance, antibody responses in the immunosenescent tend to offer less protection with limited switching to secondary isotypes, lower antibody levels in general, and production of antibody with lower affinities [69] [70] [71] [72] [73] [74] .
TextSentencer_T269 50078-50248 Sentence denotes Although we have not tested single-vaccine dose regimens, previous studies have demonstrated that these are efficacious against SARS-CoV challenge in young animals [17] .
T50357 50078-50248 Sentence denotes Although we have not tested single-vaccine dose regimens, previous studies have demonstrated that these are efficacious against SARS-CoV challenge in young animals [17] .
TextSentencer_T270 50249-50384 Sentence denotes Given the low antibody titers following boost in senescent populations, single-vaccine dose formulations will likely prove ineffective.
T99603 50249-50384 Sentence denotes Given the low antibody titers following boost in senescent populations, single-vaccine dose formulations will likely prove ineffective.
TextSentencer_T271 50385-50538 Sentence denotes Rather, improving the VRP-S efficacy in older vaccinees may require additional vaccine boosts, the use of adjuvants, or other additional therapies [75] .
T72593 50385-50538 Sentence denotes Rather, improving the VRP-S efficacy in older vaccinees may require additional vaccine boosts, the use of adjuvants, or other additional therapies [75] .
TextSentencer_T272 50539-50702 Sentence denotes Another likely contributing factor to vaccine failure in older animals was the resistance of icGDO3-S to neutralization relative to the vaccine strain, icSARS-CoV.
T97913 50539-50702 Sentence denotes Another likely contributing factor to vaccine failure in older animals was the resistance of icGDO3-S to neutralization relative to the vaccine strain, icSARS-CoV.
TextSentencer_T273 50703-50922 Sentence denotes At least three neutralizing sites have been identified in the SARS-CoV S glycoprotein, two of which map at the Nterminus and in the RBD of the S glycoprotein, and one to a weak third site near the carboxy-terminus of S.
T87977 50703-50922 Sentence denotes At least three neutralizing sites have been identified in the SARS-CoV S glycoprotein, two of which map at the Nterminus and in the RBD of the S glycoprotein, and one to a weak third site near the carboxy-terminus of S.
TextSentencer_T274 50923-51205 Sentence denotes Given that most of the GD03 mutations map in and around the Nterminus and RBD in S1 [31] , it is possible that either one or both of these critical epitopes are significantly different in icGD03-S, and likely explains the resistance to neutralization with antisera against Urbani-S.
T27238 50923-51205 Sentence denotes Given that most of the GD03 mutations map in and around the Nterminus and RBD in S1 [31] , it is possible that either one or both of these critical epitopes are significantly different in icGD03-S, and likely explains the resistance to neutralization with antisera against Urbani-S.
TextSentencer_T275 51206-51437 Sentence denotes These data suggest that robust neutralizing titers should be induced by candidate vaccines to provide long-term protection from SARS-CoV infection, especially in the vulnerable senescent population and against heterologous strains.
T86347 51206-51437 Sentence denotes These data suggest that robust neutralizing titers should be induced by candidate vaccines to provide long-term protection from SARS-CoV infection, especially in the vulnerable senescent population and against heterologous strains.
TextSentencer_T276 51438-51748 Sentence denotes Earlier work had indicated that antibodies to the Urbani strain of SARS-CoV enhanced the in vitro infectivity of pseudotyped viruses bearing the S glycoprotein of zoonotic strains, primarily with strains SZ16 and SZ3, and raised the specter of S-vaccine-induced complications with newly emergent strains [31] .
T51976 51438-51748 Sentence denotes Earlier work had indicated that antibodies to the Urbani strain of SARS-CoV enhanced the in vitro infectivity of pseudotyped viruses bearing the S glycoprotein of zoonotic strains, primarily with strains SZ16 and SZ3, and raised the specter of S-vaccine-induced complications with newly emergent strains [31] .
TextSentencer_T277 51749-52000 Sentence denotes In contrast, it was shown that monoclonal, but not polyclonal, antibodies that neutralized the epidemic strain may enhance the infectivity of pseudotyped viruses bearing GD03-S glycoproteins, although the enhanced infection was marginal at best [31] .
T96649 51749-52000 Sentence denotes In contrast, it was shown that monoclonal, but not polyclonal, antibodies that neutralized the epidemic strain may enhance the infectivity of pseudotyped viruses bearing GD03-S glycoproteins, although the enhanced infection was marginal at best [31] .
TextSentencer_T278 52001-52231 Sentence denotes Our research with antibody directed against Urbani-S indicated that the polyclonal antibody neutralized icGD03-S on Vero cells, although less efficiently than the vaccine strain, which is consistent with the previous report [31] .
T73087 52001-52231 Sentence denotes Our research with antibody directed against Urbani-S indicated that the polyclonal antibody neutralized icGD03-S on Vero cells, although less efficiently than the vaccine strain, which is consistent with the previous report [31] .
TextSentencer_T279 52232-52467 Sentence denotes Moreover, in the young and senescent mouse models, VRP-S-vaccinated animals challenged with homologous or heterologous icGD03-S recombinant viruses did not display vaccinemediated enhancement of virus replication or enhanced pathology.
T30216 52232-52467 Sentence denotes Moreover, in the young and senescent mouse models, VRP-S-vaccinated animals challenged with homologous or heterologous icGD03-S recombinant viruses did not display vaccinemediated enhancement of virus replication or enhanced pathology.
TextSentencer_T280 52468-52697 Sentence denotes Because VRP-S vaccines induce broad neutralizing antibody responses that likely target multiple epitopes across the S glycoprotein, it is possible that the noted enhancement of infectivity with monoclonal antibodies is nullified.
T76351 52468-52697 Sentence denotes Because VRP-S vaccines induce broad neutralizing antibody responses that likely target multiple epitopes across the S glycoprotein, it is possible that the noted enhancement of infectivity with monoclonal antibodies is nullified.
TextSentencer_T281 52698-53081 Sentence denotes Indeed, recent work showed that antibody specific for the RBD of Tor-2-S, GDO3-S, and SZ3-S glycoproteins did not reproduce enhanced infectivity in pseudotyped viruses bearing SZ3-S and identified conserved epitopes that allowed all three strains to be effectively neutralized, raising hope that a single vaccine could be effective against widely divergent strains of SARS-CoV [76] .
T24439 52698-53081 Sentence denotes Indeed, recent work showed that antibody specific for the RBD of Tor-2-S, GDO3-S, and SZ3-S glycoproteins did not reproduce enhanced infectivity in pseudotyped viruses bearing SZ3-S and identified conserved epitopes that allowed all three strains to be effectively neutralized, raising hope that a single vaccine could be effective against widely divergent strains of SARS-CoV [76] .
TextSentencer_T282 53082-53280 Sentence denotes Clearly, additional studies are needed with more heterologous strains in alternative animal models before the possibility of vaccine-induced enhancement of infection and pathology can be discounted.
T48166 53082-53280 Sentence denotes Clearly, additional studies are needed with more heterologous strains in alternative animal models before the possibility of vaccine-induced enhancement of infection and pathology can be discounted.
TextSentencer_T283 53281-53479 Sentence denotes To our knowledge, this study is the first to demonstrate that a SARS-CoV vaccine conferred long-term protection into the period in which a host is most susceptible to SARS-CoV pathology: senescence.
T72463 53281-53479 Sentence denotes To our knowledge, this study is the first to demonstrate that a SARS-CoV vaccine conferred long-term protection into the period in which a host is most susceptible to SARS-CoV pathology: senescence.
TextSentencer_T284 53480-53717 Sentence denotes Furthermore, the VRP-S vaccination of young animals protected against challenge with a divergent strain of SARS-CoV, indicating that current vaccines may also provide protection from many zoonotic strains that might emerge in the future.
T47883 53480-53717 Sentence denotes Furthermore, the VRP-S vaccination of young animals protected against challenge with a divergent strain of SARS-CoV, indicating that current vaccines may also provide protection from many zoonotic strains that might emerge in the future.
TextSentencer_T285 53718-53870 Sentence denotes Such cross protection has been observed among other vaccines, such as HA formulations for influenza virus [77] and VRP vaccines against norovirus [78] .
T85868 53718-53870 Sentence denotes Such cross protection has been observed among other vaccines, such as HA formulations for influenza virus [77] and VRP vaccines against norovirus [78] .
TextSentencer_T286 53871-54081 Sentence denotes Inducing robust immune responses in older animals is more challenging, but VRP-S vectors provided some protection from icGDO3-S challenge, and did so without the enhanced pathology induced by the VRP-N vaccine.
T91146 53871-54081 Sentence denotes Inducing robust immune responses in older animals is more challenging, but VRP-S vectors provided some protection from icGDO3-S challenge, and did so without the enhanced pathology induced by the VRP-N vaccine.
TextSentencer_T287 54082-54211 Sentence denotes Because human infections have not been reported since 2004, animal models are essential for the development of SARS-CoV vaccines.
T27645 54082-54211 Sentence denotes Because human infections have not been reported since 2004, animal models are essential for the development of SARS-CoV vaccines.
TextSentencer_T288 54212-54359 Sentence denotes The young mouse model provides readily available animals on a homogenous genetic background and efficient replication within the respiratory tract.
T45269 54212-54359 Sentence denotes The young mouse model provides readily available animals on a homogenous genetic background and efficient replication within the respiratory tract.
TextSentencer_T289 54360-54492 Sentence denotes The senescent mouse model adds the benefit of enhanced pathogenesis and mimicks the age-related susceptibility seen in humans [32] .
T33447 54360-54492 Sentence denotes The senescent mouse model adds the benefit of enhanced pathogenesis and mimicks the age-related susceptibility seen in humans [32] .
TextSentencer_T290 54493-54663 Sentence denotes Although aging decreases B and T cell immunity and innate immune function in humans and mice, characterization of these immune deficiencies is incomplete in both species.
T25119 54493-54663 Sentence denotes Although aging decreases B and T cell immunity and innate immune function in humans and mice, characterization of these immune deficiencies is incomplete in both species.
TextSentencer_T291 54664-54949 Sentence denotes SARS-CoV infection in senescent mice provides a key model to evaluate the mechanisms by which aging deters immune responsiveness to highly pathogenic emerging viruses like SARS-CoV and influenza virus, and develop key intervention approaches to enhance vaccine efficacy in the elderly.
T33383 54664-54949 Sentence denotes SARS-CoV infection in senescent mice provides a key model to evaluate the mechanisms by which aging deters immune responsiveness to highly pathogenic emerging viruses like SARS-CoV and influenza virus, and develop key intervention approaches to enhance vaccine efficacy in the elderly.
TextSentencer_T292 54950-55047 Sentence denotes The expense and limited availability of other senescent species makes the mouse model invaluable.
T13191 54950-55047 Sentence denotes The expense and limited availability of other senescent species makes the mouse model invaluable.
TextSentencer_T293 55048-55112 Sentence denotes Important caveats must be considered while evaluating this work.
T16817 55048-55112 Sentence denotes Important caveats must be considered while evaluating this work.
TextSentencer_T294 55113-55160 Sentence denotes Murine models of SARS disease have limitations.
T66764 55113-55160 Sentence denotes Murine models of SARS disease have limitations.
TextSentencer_T295 55161-55389 Sentence denotes The disease progression in mice is faster than in humans, rodents and humans do not share the same symptoms, and virus infection is less severe; limitations that are also evident in the hamster, primate, and ferret models [79] .
T31097 55161-55389 Sentence denotes The disease progression in mice is faster than in humans, rodents and humans do not share the same symptoms, and virus infection is less severe; limitations that are also evident in the hamster, primate, and ferret models [79] .
TextSentencer_T296 55390-55606 Sentence denotes These shortcomings necessitate that vaccine candidates be tested in other animal systems and underscore the critical need for the development of highly pathogenic challenge models for vaccine and therapeutic testing.
T49978 55390-55606 Sentence denotes These shortcomings necessitate that vaccine candidates be tested in other animal systems and underscore the critical need for the development of highly pathogenic challenge models for vaccine and therapeutic testing.
TextSentencer_T297 55607-55752 Sentence denotes This report does not provide a mechanism for the VRP-N-induced pathology nor provide solutions for minimizing potential risks associated with it.
T70375 55607-55752 Sentence denotes This report does not provide a mechanism for the VRP-N-induced pathology nor provide solutions for minimizing potential risks associated with it.
TextSentencer_T298 55753-56035 Sentence denotes Although the passive transfer of anti-SARS-N serum did not reproduce the inflammation seen in VRP-N-vaccinated animals, these results must be interpreted with caution because the anti-N antibody levels in the recipient mice were lower than those of mice directly immunized by VRP-N.
T11081 55753-56035 Sentence denotes Although the passive transfer of anti-SARS-N serum did not reproduce the inflammation seen in VRP-N-vaccinated animals, these results must be interpreted with caution because the anti-N antibody levels in the recipient mice were lower than those of mice directly immunized by VRP-N.
TextSentencer_T299 56036-56268 Sentence denotes The data presented in this manuscript do reveal critical needs and potential complications in vaccine design, laying the foundation for continuing and future studies to improve the quality, safety, and efficacy of SARS-CoV vaccines.
T79257 56036-56268 Sentence denotes The data presented in this manuscript do reveal critical needs and potential complications in vaccine design, laying the foundation for continuing and future studies to improve the quality, safety, and efficacy of SARS-CoV vaccines.
TextSentencer_T300 56269-56551 Sentence denotes Our model systems provide a means for identifying the host factors that contribute to immune senescence and will allow us to evaluate whether changes in vaccine design or regimen (Table 1 , experiment 6), showed results indistinguishable from those of young mice (unpublished data).
T33187 56269-56551 Sentence denotes Our model systems provide a means for identifying the host factors that contribute to immune senescence and will allow us to evaluate whether changes in vaccine design or regimen (Table 1 , experiment 6), showed results indistinguishable from those of young mice (unpublished data).
TextSentencer_T301 56552-56694 Sentence denotes All tissues were stained with hematoxylin and eosin. doi:10.1371/journal.pmed.0030525.g010 will improve vaccine efficacy in senescent animals.
T56915 56552-56694 Sentence denotes All tissues were stained with hematoxylin and eosin. doi:10.1371/journal.pmed.0030525.g010 will improve vaccine efficacy in senescent animals.
TextSentencer_T302 56695-56899 Sentence denotes The models provide clear rational to test candidate SARS vaccines in the hamster, ferret, and primate models in which pathology and clinical disease are more prominent following wild-type virus challenge.
T85693 56695-56899 Sentence denotes The models provide clear rational to test candidate SARS vaccines in the hamster, ferret, and primate models in which pathology and clinical disease are more prominent following wild-type virus challenge.
TextSentencer_T303 56900-57181 Sentence denotes Our research provides a model for future experiments designed to characterize the components and inducers of the VRP-N-enhanced pulmonary inflammation, and suggests that vaccine regimens that contain N protein should be used with caution in human populations until further testing.
T56887 56900-57181 Sentence denotes Our research provides a model for future experiments designed to characterize the components and inducers of the VRP-N-enhanced pulmonary inflammation, and suggests that vaccine regimens that contain N protein should be used with caution in human populations until further testing.
TextSentencer_T304 57182-57562 Sentence denotes The successful resurrection of a novel recombinant SARS challenge virus bearing zoonotic S glycoproteins suggests that it might be feasible to reconstruct other rare zoonotic SARS-CoVs that have never been successfully cultured, providing novel challenge viruses for vaccine and therapeutic drug testing against potential future zoonotic SARS introductions into human populations.
T35808 57182-57562 Sentence denotes The successful resurrection of a novel recombinant SARS challenge virus bearing zoonotic S glycoproteins suggests that it might be feasible to reconstruct other rare zoonotic SARS-CoVs that have never been successfully cultured, providing novel challenge viruses for vaccine and therapeutic drug testing against potential future zoonotic SARS introductions into human populations.
TextSentencer_T305 57563-57831 Sentence denotes Finally, these studies should encourage the development of senescent animal models of human disease and encourage vaccine testing and design against influenza, West Nile virus, and other pathogens that produce disproportionate disease burdens in the elderly [80, 81] .
T15470 57563-57831 Sentence denotes Finally, these studies should encourage the development of senescent animal models of human disease and encourage vaccine testing and design against influenza, West Nile virus, and other pathogens that produce disproportionate disease burdens in the elderly [80, 81] .
TextSentencer_T306 57833-57949 Sentence denotes The GenBank (http://www.ncbi.nlm.nih.gov/Genbank) accession number for the GD03-S glycoprotein sequence is AY525636.
T11837 57833-57949 Sentence denotes The GenBank (http://www.ncbi.nlm.nih.gov/Genbank) accession number for the GD03-S glycoprotein sequence is AY525636.
TextSentencer_T307 57950-57961 Sentence denotes Background.
T18875 57950-57961 Sentence denotes Background.
TextSentencer_T308 57962-58124 Sentence denotes Severe acute respiratory syndrome (SARS) is a flu-like illness and was first recognized in China in 2002, after which the disease rapidly spread around the world.
T50485 57962-58124 Sentence denotes Severe acute respiratory syndrome (SARS) is a flu-like illness and was first recognized in China in 2002, after which the disease rapidly spread around the world.
TextSentencer_T309 58125-58199 Sentence denotes SARS was associated with high death rates, much higher than those for flu.
T24605 58125-58199 Sentence denotes SARS was associated with high death rates, much higher than those for flu.
TextSentencer_T310 58200-58321 Sentence denotes Around 10% of people recognized as being infected with SARS died, and the death rate approached 50% among elderly people.
T46922 58200-58321 Sentence denotes Around 10% of people recognized as being infected with SARS died, and the death rate approached 50% among elderly people.
TextSentencer_T311 58322-58500 Sentence denotes The virus causing SARS was identified as a member of the coronavirus family; it is generally thought that this virus ''jumped'' to humans from bats, which harbor related viruses.
T86796 58322-58500 Sentence denotes The virus causing SARS was identified as a member of the coronavirus family; it is generally thought that this virus ''jumped'' to humans from bats, which harbor related viruses.
TextSentencer_T312 58501-58731 Sentence denotes Although SARS was declared eradicated by the World Health Organization in May 2005, there is still the possibility that similar viruses will again cross the species barrier and infect humans, with potentially serious consequences.
T11597 58501-58731 Sentence denotes Although SARS was declared eradicated by the World Health Organization in May 2005, there is still the possibility that similar viruses will again cross the species barrier and infect humans, with potentially serious consequences.
TextSentencer_T313 58732-58830 Sentence denotes As a result, many groups are working to develop vaccines that will protect against SARS infection.
T66368 58732-58830 Sentence denotes As a result, many groups are working to develop vaccines that will protect against SARS infection.
TextSentencer_T314 58831-58855 Sentence denotes Why Was This Study Done?
T84262 58831-58855 Sentence denotes Why Was This Study Done?
TextSentencer_T315 58856-59008 Sentence denotes A SARS vaccine should be effective in people of all ages, including the elderly who are more likely to get seriously ill or die if they become infected.
T22111 58856-59008 Sentence denotes A SARS vaccine should be effective in people of all ages, including the elderly who are more likely to get seriously ill or die if they become infected.
TextSentencer_T316 59009-59216 Sentence denotes In addition, potential vaccines should protect against different variants of the virus, because there are different types of the virus that could potentially cross the species barrier from animals to humans.
T12294 59009-59216 Sentence denotes In addition, potential vaccines should protect against different variants of the virus, because there are different types of the virus that could potentially cross the species barrier from animals to humans.
TextSentencer_T317 59217-59393 Sentence denotes Of the different proteins that make up the SARS coronavirus, the spike glycoprotein is thought to elicit an immune response in humans that can protect against future infection.
T11225 59217-59393 Sentence denotes Of the different proteins that make up the SARS coronavirus, the spike glycoprotein is thought to elicit an immune response in humans that can protect against future infection.
TextSentencer_T318 59394-59553 Sentence denotes The researchers therefore examined vaccine candidates based on this particular protein (termed SARS-CoV S), as well as a second one called SARS-CoV N, in mice.
T37968 59394-59553 Sentence denotes The researchers therefore examined vaccine candidates based on this particular protein (termed SARS-CoV S), as well as a second one called SARS-CoV N, in mice.
TextSentencer_T319 59554-59756 Sentence denotes Specifically, they tested whether the vaccines would protect against SARS infection in both young and older mice, and whether they would protect against infection by different strains of the SARS virus.
T43422 59554-59756 Sentence denotes Specifically, they tested whether the vaccines would protect against SARS infection in both young and older mice, and whether they would protect against infection by different strains of the SARS virus.
TextSentencer_T320 59757-59794 Sentence denotes What Did the Researchers Do and Find?
T71024 59757-59794 Sentence denotes What Did the Researchers Do and Find?
TextSentencer_T321 59795-59997 Sentence denotes The researchers created vaccines based on SARS-CoV S and SARS-CoV N by taking the genes coding for those proteins and inserting them into another type of virus particle that acted as a delivery vehicle.
T75421 59795-59997 Sentence denotes The researchers created vaccines based on SARS-CoV S and SARS-CoV N by taking the genes coding for those proteins and inserting them into another type of virus particle that acted as a delivery vehicle.
TextSentencer_T322 59998-60150 Sentence denotes They injected mice with these vaccines and then tested whether the mice generated an immune response against the specific SARS proteins, which they did.
T7922 59998-60150 Sentence denotes They injected mice with these vaccines and then tested whether the mice generated an immune response against the specific SARS proteins, which they did.
TextSentencer_T323 60151-60278 Sentence denotes The next step was to work out whether mice injected with the vaccines would be protected against later infection with SARS-CoV.
T8675 60151-60278 Sentence denotes The next step was to work out whether mice injected with the vaccines would be protected against later infection with SARS-CoV.
TextSentencer_T324 60279-60527 Sentence denotes The researchers found that mice injected with vaccine based on SARS-CoV S were protected against later infection with a standard SARS-CoV strain, both in the short term (eight weeks after vaccination) and the long term (54 weeks after vaccination).
T58079 60279-60527 Sentence denotes The researchers found that mice injected with vaccine based on SARS-CoV S were protected against later infection with a standard SARS-CoV strain, both in the short term (eight weeks after vaccination) and the long term (54 weeks after vaccination).
TextSentencer_T325 60528-60699 Sentence denotes However, the vaccine based on SARS-CoV N did not seem to result in protection, and, worryingly, caused pathological changes in the lungs of mice following virus challenge.
T18441 60528-60699 Sentence denotes However, the vaccine based on SARS-CoV N did not seem to result in protection, and, worryingly, caused pathological changes in the lungs of mice following virus challenge.
TextSentencer_T326 60700-60923 Sentence denotes To find out if their candidate vaccines would protect against different strains of SARS, the researchers made a synthetic test virus that contained a mixture of genetic material from different natural variants of the virus.
T72707 60700-60923 Sentence denotes To find out if their candidate vaccines would protect against different strains of SARS, the researchers made a synthetic test virus that contained a mixture of genetic material from different natural variants of the virus.
TextSentencer_T327 60924-61027 Sentence denotes This test virus was used to ''challenge'' mice that had been immunized with the two different vaccines.
T40655 60924-61027 Sentence denotes This test virus was used to ''challenge'' mice that had been immunized with the two different vaccines.
TextSentencer_T328 61028-61235 Sentence denotes The researchers found that the vaccine based on SARS-CoV S protected against infection by the test virus when mice were vaccinated young, but it failed to efficiently protect when administered to older mice.
T54943 61028-61235 Sentence denotes The researchers found that the vaccine based on SARS-CoV S protected against infection by the test virus when mice were vaccinated young, but it failed to efficiently protect when administered to older mice.
TextSentencer_T329 61236-61264 Sentence denotes What Do These Findings Mean?
T1748 61236-61264 Sentence denotes What Do These Findings Mean?
TextSentencer_T330 61265-61400 Sentence denotes The findings confirm others suggesting that vaccines based on the SARS-CoV S protein are more effective than those based on SARS-CoV N.
T47066 61265-61400 Sentence denotes The findings confirm others suggesting that vaccines based on the SARS-CoV S protein are more effective than those based on SARS-CoV N.
TextSentencer_T331 61401-61528 Sentence denotes They also suggest that the former can provide long-term protection in animals vaccinated young against closely related viruses.
T7049 61401-61528 Sentence denotes They also suggest that the former can provide long-term protection in animals vaccinated young against closely related viruses.
TextSentencer_T332 61529-61651 Sentence denotes However, protection against more distantly related viruses remains a challenge, especially when vaccinating older animals.
T32552 61529-61651 Sentence denotes However, protection against more distantly related viruses remains a challenge, especially when vaccinating older animals.
TextSentencer_T333 61652-61893 Sentence denotes The differences seen between young and older mice suggest that older mice might provide a useful model for animal testing of candidate vaccines for diseases like SARS, flu, and West Nile virus that pose a particular threat to elderly people.
T216 61652-61893 Sentence denotes The differences seen between young and older mice suggest that older mice might provide a useful model for animal testing of candidate vaccines for diseases like SARS, flu, and West Nile virus that pose a particular threat to elderly people.
TextSentencer_T334 61894-61990 Sentence denotes Overall, these results provide useful lessons toward future SARS vaccine development in animals.
T40345 61894-61990 Sentence denotes Overall, these results provide useful lessons toward future SARS vaccine development in animals.
TextSentencer_T335 61991-62108 Sentence denotes The synthetic virus strain generated here, and others like it, are likely to be useful tools for such future studies.
T15622 61991-62108 Sentence denotes The synthetic virus strain generated here, and others like it, are likely to be useful tools for such future studies.
TextSentencer_T336 62109-62132 Sentence denotes Additional Information.
T79949 62109-62132 Sentence denotes Additional Information.
TextSentencer_T337 62133-62244 Sentence denotes Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.
T87281 62133-62244 Sentence denotes Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.
TextSentencer_T338 62245-62253 Sentence denotes 0030525.
T73457 62245-62253 Sentence denotes 0030525.
TextSentencer_T339 62254-62534 Sentence denotes The World Health Organization provides guidance, archives, and other information resources on SARS Information from the US Centers for Disease Control on SARS Wikipedia (an internet encyclopedia anyone can edit) has an entry on SARS Collected resources from MedLinePlus about SARS
T65858 62254-62534 Sentence denotes The World Health Organization provides guidance, archives, and other information resources on SARS Information from the US Centers for Disease Control on SARS Wikipedia (an internet encyclopedia anyone can edit) has an entry on SARS Collected resources from MedLinePlus about SARS