CORD-19-Sentences  

CORD-19:a71e4139dc4ba7ca03da5d99cac0781f092d5223 JSONTXT 8 Projects

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Id Subject Object Predicate Lexical cue
TextSentencer_T1 0-157 Sentence denotes Open Access TECHNICAL ADVANCE Rapid semi-automated quantitative multiplex tandem PCR (MT-PCR) assays for the differential diagnosis of influenza-like illness
TextSentencer_T1 0-157 Sentence denotes Open Access TECHNICAL ADVANCE Rapid semi-automated quantitative multiplex tandem PCR (MT-PCR) assays for the differential diagnosis of influenza-like illness
TextSentencer_T2 159-167 Sentence denotes Abstract
TextSentencer_T2 159-167 Sentence denotes Abstract
TextSentencer_T3 168-179 Sentence denotes Background:
TextSentencer_T3 168-179 Sentence denotes Background:
TextSentencer_T4 180-290 Sentence denotes Influenza A, including avian influenza, is a major public health threat in developed and developing countries.
TextSentencer_T4 180-290 Sentence denotes Influenza A, including avian influenza, is a major public health threat in developed and developing countries.
TextSentencer_T5 291-528 Sentence denotes Rapid and accurate detection is a key component of strategies to contain spread of infection, and the efficient diagnosis of influenza-like-illness is essential to protect health infrastructure in the event of a major influenza outbreak.
TextSentencer_T5 291-528 Sentence denotes Rapid and accurate detection is a key component of strategies to contain spread of infection, and the efficient diagnosis of influenza-like-illness is essential to protect health infrastructure in the event of a major influenza outbreak.
TextSentencer_T6 529-750 Sentence denotes We developed a multiplexed PCR (MT-PCR) assay for the simultaneous diagnosis of respiratory viruses causing influenza-like illness, including the specific recognition of influenza A haemagglutinin subtypes H1, H3, and H5.
TextSentencer_T6 529-750 Sentence denotes We developed a multiplexed PCR (MT-PCR) assay for the simultaneous diagnosis of respiratory viruses causing influenza-like illness, including the specific recognition of influenza A haemagglutinin subtypes H1, H3, and H5.
TextSentencer_T7 751-887 Sentence denotes We tested several hundred clinical specimens in two diagnostic reference laboratories and compared the results with standard techniques.
TextSentencer_T7 751-887 Sentence denotes We tested several hundred clinical specimens in two diagnostic reference laboratories and compared the results with standard techniques.
TextSentencer_T8 888-1100 Sentence denotes The sensitivity and specificity of these assays was higher than individual assays based on direct antigen detection and standard PCR against a range of control templates and in several hundred clinical specimens.
TextSentencer_T8 888-1100 Sentence denotes The sensitivity and specificity of these assays was higher than individual assays based on direct antigen detection and standard PCR against a range of control templates and in several hundred clinical specimens.
TextSentencer_T9 1101-1223 Sentence denotes The MT-PCR assays provided differential diagnoses as well as potentially useful quantitation of virus in clinical samples.
TextSentencer_T9 1101-1223 Sentence denotes The MT-PCR assays provided differential diagnoses as well as potentially useful quantitation of virus in clinical samples.
TextSentencer_T10 1224-1357 Sentence denotes MT-PCR is a potentially powerful tool for the differential diagnosis of influenza-like illness in the clinical diagnostic laboratory.
TextSentencer_T10 1224-1357 Sentence denotes MT-PCR is a potentially powerful tool for the differential diagnosis of influenza-like illness in the clinical diagnostic laboratory.
TextSentencer_T11 1359-1486 Sentence denotes Approximately one billion cases of influenza occur around the world every year, despite the availability of effective vaccines.
TextSentencer_T11 1359-1486 Sentence denotes Approximately one billion cases of influenza occur around the world every year, despite the availability of effective vaccines.
TextSentencer_T12 1487-1671 Sentence denotes These are associated with an estimated 3-500 000 deaths and 3-5 million hospitalizations annually and impose a significant economic burden on even the most developed countries [1, 2] .
TextSentencer_T12 1487-1671 Sentence denotes These are associated with an estimated 3-500 000 deaths and 3-5 million hospitalizations annually and impose a significant economic burden on even the most developed countries [1, 2] .
TextSentencer_T13 1672-1852 Sentence denotes In an epidemic or pandemic setting, the rapid differentiation of influenza A from other influenza-like illness (ILI) is essential for infection control and patient management [3] .
TextSentencer_T13 1672-1852 Sentence denotes In an epidemic or pandemic setting, the rapid differentiation of influenza A from other influenza-like illness (ILI) is essential for infection control and patient management [3] .
TextSentencer_T14 1853-2193 Sentence denotes However, a relatively non-specific presentation means that influenza cannot be reliably distinguished from other ILI presenting in the autumn and winter seasons, and currently available diagnostic systems lack both the sensitivity and specificity required for efficient differ-entiation of ILI at the health facility or triage station [4] .
TextSentencer_T14 1853-2193 Sentence denotes However, a relatively non-specific presentation means that influenza cannot be reliably distinguished from other ILI presenting in the autumn and winter seasons, and currently available diagnostic systems lack both the sensitivity and specificity required for efficient differ-entiation of ILI at the health facility or triage station [4] .
TextSentencer_T15 2194-2368 Sentence denotes A variety of different diagnostic tests are available to detect most common respiratory viruses, but there is still an unmet need for rapid, sensitive and accurate diagnosis.
TextSentencer_T15 2194-2368 Sentence denotes A variety of different diagnostic tests are available to detect most common respiratory viruses, but there is still an unmet need for rapid, sensitive and accurate diagnosis.
TextSentencer_T16 2369-2621 Sentence denotes Commonly used conventional methods of virus culture, serological testing or antigen detection by direct immunofluorescence are reasonably sensitive, but relatively complex and labour-intensive, and are generally too slow to be clinically relevant [5] .
TextSentencer_T16 2369-2621 Sentence denotes Commonly used conventional methods of virus culture, serological testing or antigen detection by direct immunofluorescence are reasonably sensitive, but relatively complex and labour-intensive, and are generally too slow to be clinically relevant [5] .
TextSentencer_T17 2622-2833 Sentence denotes Rapid antigen or "point-ofcare" tests are widely available for influenza and RSV [6] [7] [8] and take only about 15-30 minutes to perform, but their sensitivity is lower than nucleic acid testing (NAT) [9, 10] .
TextSentencer_T17 2622-2833 Sentence denotes Rapid antigen or "point-ofcare" tests are widely available for influenza and RSV [6] [7] [8] and take only about 15-30 minutes to perform, but their sensitivity is lower than nucleic acid testing (NAT) [9, 10] .
TextSentencer_T18 2834-3128 Sentence denotes Predictable oseltamivir resistance occurring within outbreaks of certain influenza subtypes and the presence of several co-circulating influenza A viruses of different epidemic potential also makes subtyping particularly valuable and this is not generally available in any of the rapid methods.
TextSentencer_T18 2834-3128 Sentence denotes Predictable oseltamivir resistance occurring within outbreaks of certain influenza subtypes and the presence of several co-circulating influenza A viruses of different epidemic potential also makes subtyping particularly valuable and this is not generally available in any of the rapid methods.
TextSentencer_T19 3129-3361 Sentence denotes Here, we used the recently described MT-PCR method [11] to develop quantitative assays for the most important causes of ILI and the identification of Influenza A subtypes, and evaluate their performance in two clinical laboratories.
TextSentencer_T19 3129-3361 Sentence denotes Here, we used the recently described MT-PCR method [11] to develop quantitative assays for the most important causes of ILI and the identification of Influenza A subtypes, and evaluate their performance in two clinical laboratories.
TextSentencer_T20 3362-3656 Sentence denotes Combined nose and throat swabs collected at the Centre for Infectious Diseases and Microbiology (CIDM, at Westmead Hospital in Sydney) in the southern hemisphere winter of 2006 and 2007 were used for initial assay validation at the Centre For Immunology at St Vincents Hospital in Sydney (CFI).
TextSentencer_T20 3362-3656 Sentence denotes Combined nose and throat swabs collected at the Centre for Infectious Diseases and Microbiology (CIDM, at Westmead Hospital in Sydney) in the southern hemisphere winter of 2006 and 2007 were used for initial assay validation at the Centre For Immunology at St Vincents Hospital in Sydney (CFI).
TextSentencer_T21 3657-3981 Sentence denotes Gamma-irradiated MDCK-tissue culture extracts with known amounts of Indonesian (Indo 05, Indonesia "clade 2") and Vietnamese (HN 3028, Vietnam "clade 1") strains of H5N1 influenza A virus were provided Australia-wide as control templates [12] , and we used these along with locally cultivated tissue culture extracts of H3N2
TextSentencer_T21 3657-3981 Sentence denotes Gamma-irradiated MDCK-tissue culture extracts with known amounts of Indonesian (Indo 05, Indonesia "clade 2") and Vietnamese (HN 3028, Vietnam "clade 1") strains of H5N1 influenza A virus were provided Australia-wide as control templates [12] , and we used these along with locally cultivated tissue culture extracts of H3N2
TextSentencer_T22 3983-4272 Sentence denotes This was performed using fluorescein-conjugated monoclonal antibodies (Chemicon International, Temecula, CA, USA) against influenza A and B, RSV, adenoviruses, and hPIV1-3 on acetone-fixed smears of material from respiratory tract specimens, as indicated and as previously described [13] .
TextSentencer_T22 3983-4272 Sentence denotes This was performed using fluorescein-conjugated monoclonal antibodies (Chemicon International, Temecula, CA, USA) against influenza A and B, RSV, adenoviruses, and hPIV1-3 on acetone-fixed smears of material from respiratory tract specimens, as indicated and as previously described [13] .
TextSentencer_T23 4273-4492 Sentence denotes Nucleic acid was extracted from clinical specimens and from tissue culture using the High Pure Viral RNA Extraction kit (Roche Diagnostics GmbH, Mannheim, Germany), eluting 200 μl of sample into 50 μl of elution buffer.
TextSentencer_T23 4273-4492 Sentence denotes Nucleic acid was extracted from clinical specimens and from tissue culture using the High Pure Viral RNA Extraction kit (Roche Diagnostics GmbH, Mannheim, Germany), eluting 200 μl of sample into 50 μl of elution buffer.
TextSentencer_T24 4493-4543 Sentence denotes A negative control was extracted with every batch.
TextSentencer_T24 4493-4543 Sentence denotes A negative control was extracted with every batch.
TextSentencer_T25 4544-4823 Sentence denotes MT-PCR primers were designed with a modified version of Primer 3 software [14] , and were based on alignments of available sequences deposited in GenBank (Table 1) and of cDNA amplicons obtained from known virus isolates at CIDM cloned into pGEM-T (Promega) as positive controls.
TextSentencer_T25 4544-4823 Sentence denotes MT-PCR primers were designed with a modified version of Primer 3 software [14] , and were based on alignments of available sequences deposited in GenBank (Table 1) and of cDNA amplicons obtained from known virus isolates at CIDM cloned into pGEM-T (Promega) as positive controls.
TextSentencer_T26 4824-4996 Sentence denotes A nested reverse transcriptase polymerase chain reaction (RT-PCR) was used to detect influenza A and B and other respiratory viruses, as previously described [13, 15, 16] .
TextSentencer_T26 4824-4996 Sentence denotes A nested reverse transcriptase polymerase chain reaction (RT-PCR) was used to detect influenza A and B and other respiratory viruses, as previously described [13, 15, 16] .
TextSentencer_T27 4997-5184 Sentence denotes DNA sequences of all PCR amplicons were determined as needed by cloning into pGEM-T for M13 dye primer sequencing (Promega) and were compared with known sequences using EclustalW (ANGIS).
TextSentencer_T27 4997-5184 Sentence denotes DNA sequences of all PCR amplicons were determined as needed by cloning into pGEM-T for M13 dye primer sequencing (Promega) and were compared with known sequences using EclustalW (ANGIS).
TextSentencer_T28 5185-5319 Sentence denotes The melt temperature, with a range of 1.5°C either side of predicted Tm, was deemed acceptable if the melt curve was normal sigmoidal.
TextSentencer_T28 5185-5319 Sentence denotes The melt temperature, with a range of 1.5°C either side of predicted Tm, was deemed acceptable if the melt curve was normal sigmoidal.
TextSentencer_T29 5320-5542 Sentence denotes Verification of correctsized discrete amplicons derived in step 2 MT-PCR assays was performed using a Bioanalyzer DNA separation chip (Agilent Technologies, Forest Hill, Victoria, Australia), as previously described [11] .
TextSentencer_T29 5320-5542 Sentence denotes Verification of correctsized discrete amplicons derived in step 2 MT-PCR assays was performed using a Bioanalyzer DNA separation chip (Agilent Technologies, Forest Hill, Victoria, Australia), as previously described [11] .
TextSentencer_T30 5543-5716 Sentence denotes This is a two-step assay using nested primer pairs in which the first step involves a highly multiplexed reaction to pre-amplify multiple targets for (typically, 15) cycles.
TextSentencer_T30 5543-5716 Sentence denotes This is a two-step assay using nested primer pairs in which the first step involves a highly multiplexed reaction to pre-amplify multiple targets for (typically, 15) cycles.
TextSentencer_T31 5717-5974 Sentence denotes These are then aliquoted into individual reaction tubes containing nested specific PCR primers as template for the second step reaction [11] which was performed using a liquid handling robotics system provided by AusDiagnostics Pty Ltd. (Sydney, Australia).
TextSentencer_T31 5717-5974 Sentence denotes These are then aliquoted into individual reaction tubes containing nested specific PCR primers as template for the second step reaction [11] which was performed using a liquid handling robotics system provided by AusDiagnostics Pty Ltd. (Sydney, Australia).
TextSentencer_T32 5975-6111 Sentence denotes Primer and artificial internal control sequences are not shown due to commercial confidentiality agreements with AusDiagnostics Pty Ltd.
TextSentencer_T32 5975-6111 Sentence denotes Primer and artificial internal control sequences are not shown due to commercial confidentiality agreements with AusDiagnostics Pty Ltd.
TextSentencer_T33 6112-6181 Sentence denotes The target regions are specified in Table 1 , with example sequences.
TextSentencer_T33 6112-6181 Sentence denotes The target regions are specified in Table 1 , with example sequences.
TextSentencer_T34 6182-6258 Sentence denotes The internal control is a contrived sequence that does not appear in nature.
TextSentencer_T34 6182-6258 Sentence denotes The internal control is a contrived sequence that does not appear in nature.
TextSentencer_T35 6259-6330 Sentence denotes No base pairing redundancies were specified in any of the primers used.
TextSentencer_T35 6259-6330 Sentence denotes No base pairing redundancies were specified in any of the primers used.
TextSentencer_T36 6331-6659 Sentence denotes Lyophilized primers and reagent kits were prepared at CFI and sent to CIDM, and to the Institute of Medical and Veterinary Science (IMVS, at Royal Adelaide Hospital in Adelaide), for further testing of clinical samples collected during routine investigation of ILI outbreaks and sporadic infection at these two referral centres.
TextSentencer_T36 6331-6659 Sentence denotes Lyophilized primers and reagent kits were prepared at CFI and sent to CIDM, and to the Institute of Medical and Veterinary Science (IMVS, at Royal Adelaide Hospital in Adelaide), for further testing of clinical samples collected during routine investigation of ILI outbreaks and sporadic infection at these two referral centres.
TextSentencer_T37 6660-6884 Sentence denotes For MT-PCR performance, a strip tube containing Step 1 multiplexed primers was placed in a thermal cycler, along with a Gene-Disc containing lyophilised Step 2 reagents and oil (for covering PCR reactions) on the robot deck.
TextSentencer_T37 6660-6884 Sentence denotes For MT-PCR performance, a strip tube containing Step 1 multiplexed primers was placed in a thermal cycler, along with a Gene-Disc containing lyophilised Step 2 reagents and oil (for covering PCR reactions) on the robot deck.
TextSentencer_T38 6885-6957 Sentence denotes The samples were directly added to the strip tube in the thermal cycler.
TextSentencer_T38 6885-6957 Sentence denotes The samples were directly added to the strip tube in the thermal cycler.
TextSentencer_T39 6958-7189 Sentence denotes A software template for the reaction was then selected and all operations for performing the Step 1 multiplexed preamplification, dilution and aliquoting into the Step 2 reaction tubes in the Gene-Disc were performed automatically.
TextSentencer_T39 6958-7189 Sentence denotes A software template for the reaction was then selected and all operations for performing the Step 1 multiplexed preamplification, dilution and aliquoting into the Step 2 reaction tubes in the Gene-Disc were performed automatically.
TextSentencer_T40 7190-7325 Sentence denotes The Gene-Disc was then hermetically sealed in a heat-sealer and Step 2 amplification carried out in a Rotor-Gene RG6000 thermal cycler.
TextSentencer_T40 7190-7325 Sentence denotes The Gene-Disc was then hermetically sealed in a heat-sealer and Step 2 amplification carried out in a Rotor-Gene RG6000 thermal cycler.
TextSentencer_T41 7326-7642 Sentence denotes At the end of Step 2 the presence or absence of each target was automatically called using a software routine (AusDiagnostics Pty Ltd.) that compared melt temperature of the product with expected values and checked the purity and quantity against predetermined threshold values, and these were all manually verified.
TextSentencer_T41 7326-7642 Sentence denotes At the end of Step 2 the presence or absence of each target was automatically called using a software routine (AusDiagnostics Pty Ltd.) that compared melt temperature of the product with expected values and checked the purity and quantity against predetermined threshold values, and these were all manually verified.
TextSentencer_T42 7643-7756 Sentence denotes The cycle threshold (Ct) is the number of cycles at the takeoff point of the amplification curve (see Figure 1 ).
TextSentencer_T42 7643-7756 Sentence denotes The cycle threshold (Ct) is the number of cycles at the takeoff point of the amplification curve (see Figure 1 ).
TextSentencer_T43 7757-7942 Sentence denotes Quantitation was by comparison with the internal control, which was assigned an arbitrary value of 10 000, chosen so that the lowest concentration target detected had a value above "1".
TextSentencer_T43 7757-7942 Sentence denotes Quantitation was by comparison with the internal control, which was assigned an arbitrary value of 10 000, chosen so that the lowest concentration target detected had a value above "1".
TextSentencer_T44 7943-8053 Sentence denotes Concentration of the final step 2 product was expressed relative to the control as previously described [11] .
TextSentencer_T44 7943-8053 Sentence denotes Concentration of the final step 2 product was expressed relative to the control as previously described [11] .
TextSentencer_T45 8054-8511 Sentence denotes Assays were developed to detect Influenza viruses A and B (INF-A and INF-B), the Respiratory Syncytial Virus (RSV), Rhinovirus (RV), and human Parainfluenza virus type 3 (hPIV-3), and to identify Influenza A haemagglutinin gene types H1 (not H1N1/09), H3 and H5 within a multiplexed tandem PCR (MT-PCR) assay profile [11] , using sequences available from strains in our laboratory and lodged in GenBank, including the reference sequences listed in Table 1 .
TextSentencer_T45 8054-8511 Sentence denotes Assays were developed to detect Influenza viruses A and B (INF-A and INF-B), the Respiratory Syncytial Virus (RSV), Rhinovirus (RV), and human Parainfluenza virus type 3 (hPIV-3), and to identify Influenza A haemagglutinin gene types H1 (not H1N1/09), H3 and H5 within a multiplexed tandem PCR (MT-PCR) assay profile [11] , using sequences available from strains in our laboratory and lodged in GenBank, including the reference sequences listed in Table 1 .
TextSentencer_T46 8512-8705 Sentence denotes Multiplexed assays are generally configured so as to divide neatly into 72 (the number of positions available in a standard step 2 template in the RG6000 cycler), with no empty wells remaining.
TextSentencer_T46 8512-8705 Sentence denotes Multiplexed assays are generally configured so as to divide neatly into 72 (the number of positions available in a standard step 2 template in the RG6000 cycler), with no empty wells remaining.
TextSentencer_T47 8706-8924 Sentence denotes Primers targeting the Influenza A haemagglutinin H5 gene (in "Influenza profile 1") were replaced by primers targeting the nucleoprotein gene of human Parainfluenza virus type 3 (hPIV-3) in another assay ("profile 3").
TextSentencer_T47 8706-8924 Sentence denotes Primers targeting the Influenza A haemagglutinin H5 gene (in "Influenza profile 1") were replaced by primers targeting the nucleoprotein gene of human Parainfluenza virus type 3 (hPIV-3) in another assay ("profile 3").
TextSentencer_T48 8925-9109 Sentence denotes All data were deidentified; additional testing of laboratory samples was approved as a Quality Assurance activity by the Sydney West Area Health Service Human Research Ethics Committee
TextSentencer_T48 8925-9109 Sentence denotes All data were deidentified; additional testing of laboratory samples was approved as a Quality Assurance activity by the Sydney West Area Health Service Human Research Ethics Committee
TextSentencer_T49 9110-9454 Sentence denotes Thirty-nine viral cell-culture extracts from stored clinical isolates (including isolates identified as INF-A n = 5, INF-B n = 2, RSV n = 1, RV n = 7, along with undifferentiated picornavirus n = 4, and one each of hPIV-1, hPIV-3, coronavirus 229E, coronavirus OC43, hMPV, enterovirus EV-68, adenovirus type 3 and adenovirus type 4) were tested
TextSentencer_T49 9110-9454 Sentence denotes Thirty-nine viral cell-culture extracts from stored clinical isolates (including isolates identified as INF-A n = 5, INF-B n = 2, RSV n = 1, RV n = 7, along with undifferentiated picornavirus n = 4, and one each of hPIV-1, hPIV-3, coronavirus 229E, coronavirus OC43, hMPV, enterovirus EV-68, adenovirus type 3 and adenovirus type 4) were tested
TextSentencer_T50 9456-9778 Sentence denotes To specifically examine INF-A subtype detection, we tested serially diluted irradiated whole extracts and cell culture supernatants of reference strains of H1N1, H3N2, and Indonesian and Vietnamese H5N1 strains with Influenza Profile 1 (Table 1) , with the most diluted samples containing only 1-10 copies of RNA virus/μl.
TextSentencer_T50 9456-9778 Sentence denotes To specifically examine INF-A subtype detection, we tested serially diluted irradiated whole extracts and cell culture supernatants of reference strains of H1N1, H3N2, and Indonesian and Vietnamese H5N1 strains with Influenza Profile 1 (Table 1) , with the most diluted samples containing only 1-10 copies of RNA virus/μl.
TextSentencer_T51 9779-9954 Sentence denotes Where one haemagglutinin type gave a greater than 100-fold fluorescence signal than another after normalization against the internal control, it was deemed the correct result.
TextSentencer_T51 9779-9954 Sentence denotes Where one haemagglutinin type gave a greater than 100-fold fluorescence signal than another after normalization against the internal control, it was deemed the correct result.
TextSentencer_T52 9955-10239 Sentence denotes Due to limited availability of control samples, only 2 μl of eluate (equivalent to the same concentration of virus if extraction efficiency was 100%) was used for analysis (5 μl is recommended for extracted nasal pharyngeal samples) and all assays in Table 2 were performed only once.
TextSentencer_T52 9955-10239 Sentence denotes Due to limited availability of control samples, only 2 μl of eluate (equivalent to the same concentration of virus if extraction efficiency was 100%) was used for analysis (5 μl is recommended for extracted nasal pharyngeal samples) and all assays in Table 2 were performed only once.
TextSentencer_T53 10240-10429 Sentence denotes Serial ten-fold dilutions of H1 whole cell culture extracts from 10 -1 down to 10 -4 yielded specific signals for H1 (63904 down to 188 normalised units) and INF-A (67191 down to 231) only.
TextSentencer_T53 10240-10429 Sentence denotes Serial ten-fold dilutions of H1 whole cell culture extracts from 10 -1 down to 10 -4 yielded specific signals for H1 (63904 down to 188 normalised units) and INF-A (67191 down to 231) only.
TextSentencer_T54 10430-10734 Sentence denotes Similarly, serial dilutions of H3 whole cell culture extracts from 10 -1 down to 10 -4 yielded specific signals for H3 (33064 to 295) and INF-A (4734 to 15), with only very low cross-reactivity with H1 in the least diluted samples but with a 500-fold differential (e.g. H3 = 33064 and H1 = 66 at 10 -1 ).
TextSentencer_T54 10430-10734 Sentence denotes Similarly, serial dilutions of H3 whole cell culture extracts from 10 -1 down to 10 -4 yielded specific signals for H3 (33064 to 295) and INF-A (4734 to 15), with only very low cross-reactivity with H1 in the least diluted samples but with a 500-fold differential (e.g. H3 = 33064 and H1 = 66 at 10 -1 ).
TextSentencer_T55 10735-10884 Sentence denotes H5-specific signal was obtained from all but the highest dilutions (10 -6 ) of whole cell culture extracts, in which INF-A was detected at low level.
TextSentencer_T55 10735-10884 Sentence denotes H5-specific signal was obtained from all but the highest dilutions (10 -6 ) of whole cell culture extracts, in which INF-A was detected at low level.
TextSentencer_T56 10885-10978 Sentence denotes Low-level cross-reactivity (less than 1:200 relative to H5) was observed with H1 ( Table 2 ).
TextSentencer_T56 10885-10978 Sentence denotes Low-level cross-reactivity (less than 1:200 relative to H5) was observed with H1 ( Table 2 ).
TextSentencer_T57 10979-11196 Sentence denotes The concentration of virus in H5N1 samples (shown in parentheses in Table 2 ) had been previously determined [12] and were detected at levels as low as a single copy of virus/μl in the original sample in both strains.
TextSentencer_T57 10979-11196 Sentence denotes The concentration of virus in H5N1 samples (shown in parentheses in Table 2 ) had been previously determined [12] and were detected at levels as low as a single copy of virus/μl in the original sample in both strains.
TextSentencer_T58 11197-11460 Sentence denotes We next tested the same set of assays (Influenza Profile 1) at CIDM on 5 ul of the nucleic acid extracts obtained from forty-five clinical specimens retrieved from storage IF results on original samples were all congruent where both had been performed (Table 3) .
TextSentencer_T58 11197-11460 Sentence denotes We next tested the same set of assays (Influenza Profile 1) at CIDM on 5 ul of the nucleic acid extracts obtained from forty-five clinical specimens retrieved from storage IF results on original samples were all congruent where both had been performed (Table 3) .
TextSentencer_T59 11461-11567 Sentence denotes However, six of the sixteen samples reported negative by IF for INF-A were positive by MT-PCR (not shown).
TextSentencer_T59 11461-11567 Sentence denotes However, six of the sixteen samples reported negative by IF for INF-A were positive by MT-PCR (not shown).
TextSentencer_T60 11568-11771 Sentence denotes Of these six, one was also weakly positive by RT-PCR for INF-A (but had been reported as a negative result) and earlier specimens from the same patients had been positive for INF-A in another four cases.
TextSentencer_T60 11568-11771 Sentence denotes Of these six, one was also weakly positive by RT-PCR for INF-A (but had been reported as a negative result) and earlier specimens from the same patients had been positive for INF-A in another four cases.
TextSentencer_T61 11772-12066 Sentence denotes These additional INF-A detections were all low-level (normalised result < 100) but all had correctsized discrete second-stage amplicons as determined on Bioanalyzer (Agilent, SantaClara, CA) with sigmoidal cycling curves and single melt peaks and are thought to represent true positive results.
TextSentencer_T61 11772-12066 Sentence denotes These additional INF-A detections were all low-level (normalised result < 100) but all had correctsized discrete second-stage amplicons as determined on Bioanalyzer (Agilent, SantaClara, CA) with sigmoidal cycling curves and single melt peaks and are thought to represent true positive results.
TextSentencer_T62 12067-12154 Sentence denotes Sequencing was performed on all six of the unexpected positives to confirm the results.
TextSentencer_T62 12067-12154 Sentence denotes Sequencing was performed on all six of the unexpected positives to confirm the results.
TextSentencer_T63 12155-12305 Sentence denotes Of those that tested positive by MT-PCR, IF had detected a significant pathogen in 14/22 (64%) and inhouse RT-PCR had previously detected 23/28 (82%).
TextSentencer_T63 12155-12305 Sentence denotes Of those that tested positive by MT-PCR, IF had detected a significant pathogen in 14/22 (64%) and inhouse RT-PCR had previously detected 23/28 (82%).
TextSentencer_T64 12306-12578 Sentence denotes For greater clinical utility in the Australian influenza season, we then replaced the H5 assay with an hPIV-3 assay (Influenza profile 3: INF-A, INF-B, H1, H3, RSV, RV, hPIV-3) and tested this profile independently in two clinical laboratories (CIDM and IMVS) ( Table 4) .
TextSentencer_T64 12306-12578 Sentence denotes For greater clinical utility in the Australian influenza season, we then replaced the H5 assay with an hPIV-3 assay (Influenza profile 3: INF-A, INF-B, H1, H3, RSV, RV, hPIV-3) and tested this profile independently in two clinical laboratories (CIDM and IMVS) ( Table 4) .
TextSentencer_T65 12579-12798 Sentence denotes Two hundred and seventy-six clinical specimens tested by IF at the CIDM were tested by MT-PCR; congruent results were obtained in 53/56 IF-positive specimens; three of the 38 INF-A specimens were not detected by MT-PCR.
TextSentencer_T65 12579-12798 Sentence denotes Two hundred and seventy-six clinical specimens tested by IF at the CIDM were tested by MT-PCR; congruent results were obtained in 53/56 IF-positive specimens; three of the 38 INF-A specimens were not detected by MT-PCR.
TextSentencer_T66 12799-12936 Sentence denotes Two of these three could not be confirmed by RT-PCR for INF-A either, suggesting a false-positive IF result (no inhibition was detected).
TextSentencer_T66 12799-12936 Sentence denotes Two of these three could not be confirmed by RT-PCR for INF-A either, suggesting a false-positive IF result (no inhibition was detected).
TextSentencer_T67 12937-12982 Sentence denotes 220 additional specimens were negative by IF:
TextSentencer_T67 12937-12982 Sentence denotes 220 additional specimens were negative by IF:
TextSentencer_T68 12983-13183 Sentence denotes 14 of these samples were also positive by MT-PCR for INF-A (and/or H3), one for INF-B, and five for RSV (these five all coming from symptomatic patients taken during investigation of an RSV outbreak).
TextSentencer_T68 12983-13183 Sentence denotes 14 of these samples were also positive by MT-PCR for INF-A (and/or H3), one for INF-B, and five for RSV (these five all coming from symptomatic patients taken during investigation of an RSV outbreak).
TextSentencer_T69 13184-13274 Sentence denotes All of these 14 additional positive MT-PCR results were low-level (< 50 normalised units).
TextSentencer_T69 13184-13274 Sentence denotes All of these 14 additional positive MT-PCR results were low-level (< 50 normalised units).
TextSentencer_T70 13275-13425 Sentence denotes Two additional hPIV-3 and ten RSV were also detected in samples that had only been tested for INF-A and INF-B, but these are not included in Table 4 .
TextSentencer_T70 13275-13425 Sentence denotes Two additional hPIV-3 and ten RSV were also detected in samples that had only been tested for INF-A and INF-B, but these are not included in Table 4 .
TextSentencer_T71 13427-13570 Sentence denotes One hundred and seventy-six additional clinical specimens, completely unrelated to those tested above, were tested by RT-PCR at the IMVS [16] :
TextSentencer_T71 13427-13570 Sentence denotes One hundred and seventy-six additional clinical specimens, completely unrelated to those tested above, were tested by RT-PCR at the IMVS [16] :
TextSentencer_T72 13571-13658 Sentence denotes 87 were reported positive and MT-PCR results were congruent in all of these (Table 4 ).
TextSentencer_T72 13571-13658 Sentence denotes 87 were reported positive and MT-PCR results were congruent in all of these (Table 4 ).
TextSentencer_T73 13659-13800 Sentence denotes Of the 89 negative results, five tested positive by MT-PCR (profile 3): one was a very low-level INF-A, while the other four were RSV and RV.
TextSentencer_T73 13659-13800 Sentence denotes Of the 89 negative results, five tested positive by MT-PCR (profile 3): one was a very low-level INF-A, while the other four were RSV and RV.
TextSentencer_T74 13801-13920 Sentence denotes These specimens had not previously been tested for these pathogens and the MT-PCR results are not included in Table 4 .
TextSentencer_T74 13801-13920 Sentence denotes These specimens had not previously been tested for these pathogens and the MT-PCR results are not included in Table 4 .
TextSentencer_T75 13921-14060 Sentence denotes An INF-A/RSV co-infection which had been already identified by RT-PCR was identified as an INF-A H3/RSV co-infection by MT-PCR (not shown).
TextSentencer_T75 13921-14060 Sentence denotes An INF-A/RSV co-infection which had been already identified by RT-PCR was identified as an INF-A H3/RSV co-infection by MT-PCR (not shown).
TextSentencer_T76 14061-14075 Sentence denotes The Table 5 ).
TextSentencer_T76 14061-14075 Sentence denotes The Table 5 ).
TextSentencer_T77 14076-14431 Sentence denotes The set of INF-A/NONO ratios in Table 5 were derived from assays of serial samples of patients treated with oseltamivir for influenza A during the course of management of an institutional outbreak, demonstrating the potential for quantitation as a marker of sample quality (normalised NONO value) and perhaps of the course of infection (INF-A/NONO ratio).
TextSentencer_T77 14076-14431 Sentence denotes The set of INF-A/NONO ratios in Table 5 were derived from assays of serial samples of patients treated with oseltamivir for influenza A during the course of management of an institutional outbreak, demonstrating the potential for quantitation as a marker of sample quality (normalised NONO value) and perhaps of the course of infection (INF-A/NONO ratio).
TextSentencer_T78 14433-14589 Sentence denotes Detection methods are available for the major ILI pathogens (Table 1) , as well as for the 15 known H and 9 known N variants [17] [18] [19] [20] [21] [22] .
TextSentencer_T78 14433-14589 Sentence denotes Detection methods are available for the major ILI pathogens (Table 1) , as well as for the 15 known H and 9 known N variants [17] [18] [19] [20] [21] [22] .
TextSentencer_T79 14590-14819 Sentence denotes Antigen-based rapid assays are generally quite specific but range from 60-70% in sensitivity [6, 8] ; PCR methods appear to be the most sensitive and specific, with detection limits down to < 0.1 TCID 50 /mL [23] [24] [25] [26] .
TextSentencer_T79 14590-14819 Sentence denotes Antigen-based rapid assays are generally quite specific but range from 60-70% in sensitivity [6, 8] ; PCR methods appear to be the most sensitive and specific, with detection limits down to < 0.1 TCID 50 /mL [23] [24] [25] [26] .
TextSentencer_T80 14820-14955 Sentence denotes Multiplex PCR must deal with the competition and interference that arises as a result of using large numbers of primers simultaneously.
TextSentencer_T80 14820-14955 Sentence denotes Multiplex PCR must deal with the competition and interference that arises as a result of using large numbers of primers simultaneously.
TextSentencer_T81 14956-15054 Sentence denotes DNA hybridisation after multiplexed PCR is one solution but is slow and labourintensive [27, 28] .
TextSentencer_T81 14956-15054 Sentence denotes DNA hybridisation after multiplexed PCR is one solution but is slow and labourintensive [27, 28] .
TextSentencer_T82 15055-15284 Sentence denotes MT-PCR deals with the problem by stopping the multiplexed first round of amplification before competition develops, then using the amplified products as template for a second round of individual specific 'nested' reactions [11] .
TextSentencer_T82 15055-15284 Sentence denotes MT-PCR deals with the problem by stopping the multiplexed first round of amplification before competition develops, then using the amplified products as template for a second round of individual specific 'nested' reactions [11] .
TextSentencer_T83 15285-15454 Sentence denotes This evaluation does not control for variation in specimen quality or operator performance or any of a number of other variables applicable in real working laboratories.
TextSentencer_T83 15285-15454 Sentence denotes This evaluation does not control for variation in specimen quality or operator performance or any of a number of other variables applicable in real working laboratories.
TextSentencer_T84 15455-15598 Sentence denotes We used an automated liquid-handling system and robotic cycler that operates on a preset algorithm for both cycling and for calling of results.
TextSentencer_T84 15455-15598 Sentence denotes We used an automated liquid-handling system and robotic cycler that operates on a preset algorithm for both cycling and for calling of results.
TextSentencer_T85 15599-15701 Sentence denotes All results were manually verified in this study but required operator expertise is otherwise minimal.
TextSentencer_T85 15599-15701 Sentence denotes All results were manually verified in this study but required operator expertise is otherwise minimal.
TextSentencer_T86 15702-15766 Sentence denotes Second-stage amplified product is sealed and discarded unopened.
TextSentencer_T86 15702-15766 Sentence denotes Second-stage amplified product is sealed and discarded unopened.
TextSentencer_T87 15767-15875 Sentence denotes Every secondstep 72-well gene disc has 12 wells for each target of a 6target assay, all separated spatially.
TextSentencer_T87 15767-15875 Sentence denotes Every secondstep 72-well gene disc has 12 wells for each target of a 6target assay, all separated spatially.
TextSentencer_T88 15876-16037 Sentence denotes The additional positives we describe came on different days, all with negative controls and with several other negatives in the runs in which they were detected.
TextSentencer_T88 15876-16037 Sentence denotes The additional positives we describe came on different days, all with negative controls and with several other negatives in the runs in which they were detected.
TextSentencer_T89 16038-16178 Sentence denotes All MT-PCR reactions yielded normal melt curves and all reactions giving unexpected results also contained discrete correct-sized amplicons.
TextSentencer_T89 16038-16178 Sentence denotes All MT-PCR reactions yielded normal melt curves and all reactions giving unexpected results also contained discrete correct-sized amplicons.
TextSentencer_T90 16179-16268 Sentence denotes No multiple or unexpected-size amplicons or discrepant melt temperatures were identified.
TextSentencer_T90 16179-16268 Sentence denotes No multiple or unexpected-size amplicons or discrepant melt temperatures were identified.
TextSentencer_T91 16269-16582 Sentence denotes Nevertheless, sequencing of clinical isolates was not performed after initial test validation and no additional alternative independent testing was performed to specifically verify second-step results, so contamination of the second-step reaction with unrelated first-step amplicons cannot be completely excluded.
TextSentencer_T91 16269-16582 Sentence denotes Nevertheless, sequencing of clinical isolates was not performed after initial test validation and no additional alternative independent testing was performed to specifically verify second-step results, so contamination of the second-step reaction with unrelated first-step amplicons cannot be completely excluded.
TextSentencer_T92 16583-16750 Sentence denotes The MT-PCR method is essentially a nested RT-PCR and would therefore be expected to be more sensitive than a conventional PCR or direct antigen detection system [29] .
TextSentencer_T92 16583-16750 Sentence denotes The MT-PCR method is essentially a nested RT-PCR and would therefore be expected to be more sensitive than a conventional PCR or direct antigen detection system [29] .
TextSentencer_T93 16751-16883 Sentence denotes Specificity may thus be underestimated if MT-PCR detects target sequences in material below levels detected by conventional methods.
TextSentencer_T93 16751-16883 Sentence denotes Specificity may thus be underestimated if MT-PCR detects target sequences in material below levels detected by conventional methods.
TextSentencer_T94 16884-17141 Sentence denotes Testing of control templates gave "clean" results with no evident cross-reaction (Figures 1, 2, 3, 4 ) and the unexpected positive results obtained in "negative" clinical specimens were almost uniformly at low levels, consistent with this as an explanation.
TextSentencer_T94 16884-17141 Sentence denotes Testing of control templates gave "clean" results with no evident cross-reaction (Figures 1, 2, 3, 4 ) and the unexpected positive results obtained in "negative" clinical specimens were almost uniformly at low levels, consistent with this as an explanation.
TextSentencer_T95 17142-17295 Sentence denotes Also consistent, unexpected additional positive results from MT-PCR were much more common in specimens deemed negative by IF than by PCR (see Table 4 ).
TextSentencer_T95 17142-17295 Sentence denotes Also consistent, unexpected additional positive results from MT-PCR were much more common in specimens deemed negative by IF than by PCR (see Table 4 ).
TextSentencer_T96 17296-17621 Sentence denotes For example, the additional MT-PCR-positive IF-negative samples at CIDM (n = 14) for INF-A were collected from symptomatic patients during an influenza outbreak and several were either confirmed by RT-PCR or were associated with seroconversion (greater than four-fold increase) or initial high (> = 32) titres to Influenza A.
TextSentencer_T96 17296-17621 Sentence denotes For example, the additional MT-PCR-positive IF-negative samples at CIDM (n = 14) for INF-A were collected from symptomatic patients during an influenza outbreak and several were either confirmed by RT-PCR or were associated with seroconversion (greater than four-fold increase) or initial high (> = 32) titres to Influenza A.
TextSentencer_T97 17622-17750 Sentence denotes This level of additional positive results (~10%) is very similar to that seen in previous reports for assays of this type [30] .
TextSentencer_T97 17622-17750 Sentence denotes This level of additional positive results (~10%) is very similar to that seen in previous reports for assays of this type [30] .
TextSentencer_T98 17751-17921 Sentence denotes Taken all together and including assays tested within both profiles, MT-PCR correctly identified 141 (97.9%) of 144 clinical specimens found positive by standard methods.
TextSentencer_T98 17751-17921 Sentence denotes Taken all together and including assays tested within both profiles, MT-PCR correctly identified 141 (97.9%) of 144 clinical specimens found positive by standard methods.
TextSentencer_T99 17922-18114 Sentence denotes Conversely, taking into account all of those specimens found to be positive by MT-PCR in this study, IF detected a significant pathogen in 56/73 (77%) and inhouse RT-PCRs detected 87/88 (99%).
TextSentencer_T99 17922-18114 Sentence denotes Conversely, taking into account all of those specimens found to be positive by MT-PCR in this study, IF detected a significant pathogen in 56/73 (77%) and inhouse RT-PCRs detected 87/88 (99%).
TextSentencer_T100 18115-18168 Sentence denotes For influenza A Table 2 included, as 188/189 (99.5%).
TextSentencer_T100 18115-18168 Sentence denotes For influenza A Table 2 included, as 188/189 (99.5%).
TextSentencer_T101 18169-18347 Sentence denotes Conversely, the IF detected 70/95 (74%) and RT-PCR detected 110/116 (95%) of those that were MT-PCR positive for any pathogen, which included many additional low-level positives.
TextSentencer_T101 18169-18347 Sentence denotes Conversely, the IF detected 70/95 (74%) and RT-PCR detected 110/116 (95%) of those that were MT-PCR positive for any pathogen, which included many additional low-level positives.
TextSentencer_T102 18348-18472 Sentence denotes For INF-A, the relative sensitivity overall of IF relative to MT-PCR was 44/63 (70%) and of in-house RT-PCR was 58/64 (91%).
TextSentencer_T102 18348-18472 Sentence denotes For INF-A, the relative sensitivity overall of IF relative to MT-PCR was 44/63 (70%) and of in-house RT-PCR was 58/64 (91%).
TextSentencer_T103 18473-18661 Sentence denotes Human Influenza virus replicates throughout the respiratory tract and viral loads normally peak (10 3 to 10 7 TCID 50 /mL of nasopharyngeal samples) at 24-72 hours after onset of symptoms.
TextSentencer_T103 18473-18661 Sentence denotes Human Influenza virus replicates throughout the respiratory tract and viral loads normally peak (10 3 to 10 7 TCID 50 /mL of nasopharyngeal samples) at 24-72 hours after onset of symptoms.
TextSentencer_T104 18662-18834 Sentence denotes Determining the infectious period and its variability is crucial for individual and public health planning but are likely to be both host-and strain-specific [24, 31, 32] .
TextSentencer_T104 18662-18834 Sentence denotes Determining the infectious period and its variability is crucial for individual and public health planning but are likely to be both host-and strain-specific [24, 31, 32] .
TextSentencer_T105 18835-19068 Sentence denotes In previous testing of RNA targets using MT-PCR, the correlation of quantitation over two logs of template concentration was 0.99 and the coefficient of variation over ten independent cycle threshold (Ct) measurements was 0.03 [11] .
TextSentencer_T105 18835-19068 Sentence denotes In previous testing of RNA targets using MT-PCR, the correlation of quantitation over two logs of template concentration was 0.99 and the coefficient of variation over ten independent cycle threshold (Ct) measurements was 0.03 [11] .
TextSentencer_T106 19069-19184 Sentence denotes Here, we also used NONO as a marker of human epithelial cells in the specimen to normalise viral titres (Table 5 ).
TextSentencer_T106 19069-19184 Sentence denotes Here, we also used NONO as a marker of human epithelial cells in the specimen to normalise viral titres (Table 5 ).
TextSentencer_T107 19185-19351 Sentence denotes For well-collected specimens with significant amounts of human epithelial cells present, the INF-A/NONO ratio may be useful to monitor viral shedding and infectivity.
TextSentencer_T107 19185-19351 Sentence denotes For well-collected specimens with significant amounts of human epithelial cells present, the INF-A/NONO ratio may be useful to monitor viral shedding and infectivity.
TextSentencer_T108 19352-19664 Sentence denotes However, actual primer sequences may fail to recognise new viral variants and so a lack of transparency by manufacturers in releasing such information requires evidence that the very latest sequences available are completely homologous with the primers included, if clinical laboratories are to trust the assays.
TextSentencer_T108 19352-19664 Sentence denotes However, actual primer sequences may fail to recognise new viral variants and so a lack of transparency by manufacturers in releasing such information requires evidence that the very latest sequences available are completely homologous with the primers included, if clinical laboratories are to trust the assays.
TextSentencer_T109 19665-19791 Sentence denotes Given the ready availability of influenza A sequences (eg. via PubMed), it is reasonable to expect this as a minimum standard.
TextSentencer_T109 19665-19791 Sentence denotes Given the ready availability of influenza A sequences (eg. via PubMed), it is reasonable to expect this as a minimum standard.
TextSentencer_T110 19792-19931 Sentence denotes Our testing of fully-characterised strains included only those listed in Table 2 , and the reference sequences used are listed in Table 1 .
TextSentencer_T110 19792-19931 Sentence denotes Our testing of fully-characterised strains included only those listed in Table 2 , and the reference sequences used are listed in Table 1 .
TextSentencer_T111 19932-20161 Sentence denotes In this study, the isolates tested in the clinical laboratories came only from two Australian population centres and different regions may encounter strains which are significantly different from those listed in Tables 1 and 2 .
TextSentencer_T111 19932-20161 Sentence denotes In this study, the isolates tested in the clinical laboratories came only from two Australian population centres and different regions may encounter strains which are significantly different from those listed in Tables 1 and 2 .
TextSentencer_T112 20162-20590 Sentence denotes The initial H5 target is a consensus for clades 1-3 of the highly pathogenic H5N1 strains, consistent with WHO guidelines http://www.who.int/csr/disease/avian_influenza/guidelines/RecAIlabtestsAug07.pdf, but the region targeted by our consensus INF-A primers varies significantly from that of the highly pathogenic avian strains of Influenza A, and this was reflected in a relatively low sensitivity in the INF-A assay for H5N1.
TextSentencer_T112 20162-20590 Sentence denotes The initial H5 target is a consensus for clades 1-3 of the highly pathogenic H5N1 strains, consistent with WHO guidelines http://www.who.int/csr/disease/avian_influenza/guidelines/RecAIlabtestsAug07.pdf, but the region targeted by our consensus INF-A primers varies significantly from that of the highly pathogenic avian strains of Influenza A, and this was reflected in a relatively low sensitivity in the INF-A assay for H5N1.
TextSentencer_T113 20591-20896 Sentence denotes Since this assay was developed, we have also experienced the influenza A/ H1N1/09 pandemic, which caused a significant number of intensive care admissions and deaths in this country (33) and required modifications to include the NP gene of A/H1N1/09 and an M gene consensus (H1N1; A/H1N1/ 09; H3N2; H5N1).
TextSentencer_T113 20591-20896 Sentence denotes Since this assay was developed, we have also experienced the influenza A/ H1N1/09 pandemic, which caused a significant number of intensive care admissions and deaths in this country (33) and required modifications to include the NP gene of A/H1N1/09 and an M gene consensus (H1N1; A/H1N1/ 09; H3N2; H5N1).
TextSentencer_T114 20897-21038 Sentence denotes Performance in the Australian 2009 influenza season appears comparable to a nested RT-PCR specifically optimised for A/H1N1/09 only [9, 10] .
TextSentencer_T114 20897-21038 Sentence denotes Performance in the Australian 2009 influenza season appears comparable to a nested RT-PCR specifically optimised for A/H1N1/09 only [9, 10] .
TextSentencer_T115 21039-21143 Sentence denotes The diagnosis of co-infections (e.g. RSV and INF-A) and other causes of ILI may be clinically important.
TextSentencer_T115 21039-21143 Sentence denotes The diagnosis of co-infections (e.g. RSV and INF-A) and other causes of ILI may be clinically important.
TextSentencer_T116 21144-21439 Sentence denotes Pandemic influenza dramatically increases requirements for high-level Intensive Care [33] and the ability to efficiently distinguish true INF-A from ILI of a less threatening nature in the context of an outbreak or epidemic may be vital to minimise the burden on the health infrastructure [34] .
TextSentencer_T116 21144-21439 Sentence denotes Pandemic influenza dramatically increases requirements for high-level Intensive Care [33] and the ability to efficiently distinguish true INF-A from ILI of a less threatening nature in the context of an outbreak or epidemic may be vital to minimise the burden on the health infrastructure [34] .
TextSentencer_T117 21440-21735 Sentence denotes Respiratory syncytial virus (RSV) is the single most important cause of acute lower respiratory tract diseases including bronchiolitis and pneumonia in infants and young children and may contribute to hundreds of thousands of deaths annually among the elderly and immunocompromised [1, 35, 36] .
TextSentencer_T117 21440-21735 Sentence denotes Respiratory syncytial virus (RSV) is the single most important cause of acute lower respiratory tract diseases including bronchiolitis and pneumonia in infants and young children and may contribute to hundreds of thousands of deaths annually among the elderly and immunocompromised [1, 35, 36] .
TextSentencer_T118 21736-21991 Sentence denotes Human Parainfluenza virus type 3 (hPIV-3) is associated with pharyngitis, bronchiolitis and pneumonia in children, and respiratory infections in adults and is the second most significant acute viral respiratory tract infection in young children after RSV.
TextSentencer_T118 21736-21991 Sentence denotes Human Parainfluenza virus type 3 (hPIV-3) is associated with pharyngitis, bronchiolitis and pneumonia in children, and respiratory infections in adults and is the second most significant acute viral respiratory tract infection in young children after RSV.
TextSentencer_T119 21992-22095 Sentence denotes Both are leading causes of hospitalization in adults with acute community-acquired respiratory disease.
TextSentencer_T119 21992-22095 Sentence denotes Both are leading causes of hospitalization in adults with acute community-acquired respiratory disease.
TextSentencer_T120 22096-22269 Sentence denotes The rhinoviruses (RV), of which there are more than 100 serotypes, act either as single pathogens or in mixed infections, but all of these may present as a non-specific ILI.
TextSentencer_T120 22096-22269 Sentence denotes The rhinoviruses (RV), of which there are more than 100 serotypes, act either as single pathogens or in mixed infections, but all of these may present as a non-specific ILI.
TextSentencer_T121 22270-22586 Sentence denotes We have tested several hundred clinical samples in two reference laboratories and found multiplexed tandem (MT)-PCR to be comparable to target-specific RT-PCR and a great deal more sensitive than direct immunofluorescence for the detection of several of the most important causative agents of influenza-like illness.
TextSentencer_T121 22270-22586 Sentence denotes We have tested several hundred clinical samples in two reference laboratories and found multiplexed tandem (MT)-PCR to be comparable to target-specific RT-PCR and a great deal more sensitive than direct immunofluorescence for the detection of several of the most important causative agents of influenza-like illness.
TextSentencer_T122 22587-22770 Sentence denotes Importantly, this method simultaneously subtypes Influenza A, identifies multiple pathogens and co-infections in a single specimen and provides potentially valuable quantitative data.
TextSentencer_T122 22587-22770 Sentence denotes Importantly, this method simultaneously subtypes Influenza A, identifies multiple pathogens and co-infections in a single specimen and provides potentially valuable quantitative data.
TextSentencer_T123 22771-22921 Sentence denotes A multiplexed method also facilitates recognition of co-circulating viruses in outbreak situations in which they might otherwise be assumed identical.
TextSentencer_T123 22771-22921 Sentence denotes A multiplexed method also facilitates recognition of co-circulating viruses in outbreak situations in which they might otherwise be assumed identical.
TextSentencer_T124 22922-23182 Sentence denotes It is clear that, as for influenza vaccine strategies, primer designs need to be reviewed regularly in light of prevalent sequence variations and the configuration of the multiplexed assay itself needs to be informed by the relevant epidemiology of the region.
TextSentencer_T124 22922-23182 Sentence denotes It is clear that, as for influenza vaccine strategies, primer designs need to be reviewed regularly in light of prevalent sequence variations and the configuration of the multiplexed assay itself needs to be informed by the relevant epidemiology of the region.
TextSentencer_T125 23183-23278 Sentence denotes The assay we describe presently costs around AUD$15 (USD$12) for 6 targets (~USD$2 per target).
TextSentencer_T125 23183-23278 Sentence denotes The assay we describe presently costs around AUD$15 (USD$12) for 6 targets (~USD$2 per target).
TextSentencer_T126 23279-23392 Sentence denotes Even with additional purification and labour costs, this is potentially cost-effective in the routine laboratory.
TextSentencer_T126 23279-23392 Sentence denotes Even with additional purification and labour costs, this is potentially cost-effective in the routine laboratory.