3.4. Influenza Viruses in Swine in Europe

3.4.1. Belgium
The H1N1 virus was identified in swine lung tissues or trachea of two of the deceased sows after an influenza-like disease erupted at two swine farms in January 1979. Interestingly, it was also reported that the identical virus was detected in wild ducks in Germany [179]. Since it was already established that H1N1 from wild ducks can successfully infect swine if inoculated via intranasal route [179] hence this observation suggested the transmission of H1N1 from wild ducks to the swine [180]. A second investigation isolated three avian-like H1N1, two H1N2 and twelve human-like H3N2 viruses from eight commercial swine farms in March 1999 [181].

3.4.2. Denmark
Denmark has been running a passive surveillance program for IAV detection in swine since 2011. The H1N2 virus having the H1 gene which evolved from H1N1 avian-like viruses and N2 gene which evolved from human H3N2 viruses was reported in swine during 2011–2013 [182]. This was an example of how IAV can evolve through reassortment and may emerge into a new IAV strain.
The other investigation included swine sampling at different time intervals to assess the persistence of IAV shedding in Danish swine which detected one avian-like H1N1 and 107 reassortant H1N2 viruses. This study observed that most of the swine were shedding IAV right before achieving six weeks of age. Surprisingly, a piglet as young as just three days was found infected with IAV [183].
Two H3N2 isolates having H3 genes from seasonal human influenza along with internal genes that originated from A(H1N1)pdm09 virus and NA genes from contemporary N2 swine influenza viruses that have been in circulation in Denmark were retrieved from young piglets at two locations during 2011–2014 [184]. H3N2 virus was also detected from piglets having respiratory illness and from sows with reproductive problems in commercial piggeries in 2014 [184].

3.4.3. United Kingdom
The H3N2 virus antibodies were first detected in English swine in 1973 which revealed the past exposure of swine to H3N2 virus [185]. Later, the antibodies for H1N1 and H3N2 viruses were detected in swine at a slaughterhouse in England during 1991–1992 [186]. Interestingly, this serological investigation also reported the antibodies for IBV in eight and for ICV in 198 swine [186].
A molecular investigation identified a novel H1N7 virus in swine in England which had six of its RNA segments closely related to those of human viruses while two RNA segments were identical to those of equine viruses which concluded that the H1N7 strain may have evolved due to reassortment between human H1 and equine H7N7 viruses [187,188].
The first report of A(H1N1)pdm09 virus in English swine appeared in September 2009 when histology and immunofluorescence assays followed by molecular diagnostics and sequencing confirmed four A(H1N1)pdm09 virus infected swine in the Northern Ireland [189]. After this, 17 more A(H1N1)pdm09 virus isolates were reported in swine in England during September 2009–October 2010 which revealed that A(H1N1)pdm09 virus was in circulation in English swine during the 2009 flu pandemic [190]. The same year, four H1N2 virus isolates were reported in English swine which had six internal genes of A(H1N1)pdm09 virus along with HA and NA genes of H1N2 virus hence were identified as the novel reassortant H1N2 strains [191]. In a more recent study, two more IAV positive swine were identified in the United Kingdom in 2016 [192].

3.4.4. Finland
However the first report of seroprevalence of H1N1 virus in Finnish swine appeared in 2008 during an investigation which detected H1N1 virus antibodies in swine at seven swine farms which further increased to 24 swine farms in 2009 [193] but the first isolate of avian-like swine H1N1 virus (indicative of active infection) was detected from the lung tissues of a swine in February 2009. Later, the first A(H1N1)pdm09 virus in Finnish swine was detected in November 2009 [193]. Three more swine were identified with IAV antibodies during May 2011–January 2014 which was due to a past exposure to IAV [194].

3.4.5. France
The H1N1 viruses in turkey and swine were identified after the swine influenza outbreak hit the turkey population in Brittany, France in February 1983 which suggested that IAV transmission happened from swine to turkey [195]. Later two strains of H1N2 virus were isolated from six swine exhibiting influenza-like illness in Brittany during 1987–1988 [196]. Another investigation reported H1N1, H1N2, and H3N2 viruses in swine herds experiencing respiratory disease in Brittany region [197].
A negative prevalence of IAV was reported in wild boars in Camargue during September 2009–November 2010 given that all the 315 nasal swabs obtained from either hunted or trapped wild boars along with all the sera samples were negative for IAV [198].
A more recent investigation reported the zoonotic transmission of A(H1N1)pdm09 virus from swine to a farmer in January 2018. This farmer along with a veterinarian collected nasal swab samples from three pregnant sows exhibiting influenza-like illness on the swine farm and submitted to a local diagnostic laboratory for analysis which, as a result, were found IAV positive. Few days later, the farmer and the veterinarian both developed the influenza-like symptoms. The farmer was later diagnosed with A(H1N1)pdm09 virus [199].

3.4.6. Germany
Sixty-five IAV positive wild boars were identified across five German states during 1997–2006. Cloning and sequencing identified H1N1 and H3N2 viruses in these wild boars [200]. Later thirteen H1N1, three reassorted A(H1N1)pdm09 and four H1N2 viruses were detected in swine during 2009–2010. Interestingly, the A(H1N1)pdm09 virus isolates had high similarity with the A(H1N1)pdm09 viruses reported earlier in humans within Germany which suggested a reverse zoonotic transmission of the A(H1N1)pdm09 virus [201].
A nationwide sero-surveillance identified 12,585 swine with H1N1, 9,566 swine with human-like H1N2, 12,220 swine with human-like H3N2 and 11,086 swine with A(H1N1)pdm09 virus antibodies during June 2009–December 2012 which reflected a high seroprevalence of influenza viruses in German swine population [202].
Later 273 IAV positive swine exhibiting influenza-like illness were detected between January 2010–December 2013. Subtyping successfully distinguished 198 of 273 samples into H1N1, H1N2, H3N2 and A(H1N1)pdm09 viruses. The H1N1 virus was the most widely occurring in German swine while A(H1N1)pdm09 virus had the lowest prevalence [203].

3.4.7. Greece
The H1N1, H1N2, H3N2, and A(H1N1)pdm09 viruses were detected in swine sera samples collected from apparently healthy swine at 42 swine farms during 2002–2004 and from 46 swine farms during 2010–2012 [204].

3.4.8. Italy
The seropositivity of Italian swine to H3N2 virus was first reported during December 1976–November 1977 when 24 swine were detected with H3N2 antibodies [205]. The first report of H1N1 active infection in Italian swine appeared during an influenza disease outbreak between 1977 to 1986 which identified 63 H1N1 viruses [206]. Further, four H3N2 viruses were detected in swine nasal swabs originated from three swine farms and an abattoir during 1981–1982 [207].
Later 47 H1N1 and 37 H3N2 viruses were detected in swine during 1992–1995. Interestingly, four human sera samples were also positive for H1N1 and 77 samples were positive for H3N2 viruses which might be due to the transmission between human and swine [208]. Further IAV seroprevalence was detected in the age group of three-month to four-year old swine during 2002–2004 [209].
The first report of A(H1N1)pdm09 virus in Italian swine appeared after a respiratory disease outbreak in piggeries in Lombardia region of Northern Italy in November 2009. Piglets experienced diarrhea and weight loss while the sows experienced reduction in reproduction rate [210]. Two more A(H1N1)pdm09 virus isolates were reported in female swine in Sicily in December 2009 [211] while five isolates of A(H1N1)pdm09 virus were identified in swine at three different locations during 2011–2012 [212].
A novel strain of reassorted H1N2 virus having 99–100% identity through six gene segments with A(H1N1)pdm09 virus along with HA and NA genes similar to H1N2 virus was reported in swine in Mantua Province [213]. Reassorted H1N2 viruses were again detected in 34 piglets during 2013–2014 [214].
Seroprevalence of Italian wild boars with one H1N1, ten H1N2, and one H3N2 viruses at two different locations was reported during 2012. On the contrary, active infection was found only in three wild boars whose nasal swabs were positive for the IAV [215]. One more investigation reported active infection of IAV in 12 wild boars while 78 wild boars had IAV antibodies during July–December 2012 [216]. Further molecular and serological investigations detected avian-like H1N1 viruses in Italian wild boars [216].
The first complete genome sequence of IDV in Italian swine was retrieved from a symptomatic sow in 2015 which was identified to be closely related to the IDV sequence reported in Oklahoma swine in 2011 [217]. Further IDV prevalence in Italian swine was reported when 14, three and four swine were found positive for IDV antibodies in Veneto, Emilia Romagna and Lombardia regions, respectively during June 2015–May 2016. As a result, swine clinical samples collected during 2013–2014 were investigated retrospectively for IDV prevalence but were reported negative. An extended serological investigation detected IDV antibodies in 364 swine sera samples collected during 2015. These findings suggested that IDV was in circulation in Italian swine population only after 2014 [45].

3.4.9. Spain
Isolation and characterization of 12 H3N2, nine H1N1 and one H1N2 viruses reported the prevalence of influenza viruses for the first time in Spanish swine herds experiencing the respiratory illness and pneumonia during November 2001–April 2004 [218]. More strains of H1N1, H1N2 and H3N2 viruses were isolated, sequenced and characterized in Spanish swine during 2006–2011 [219,220,221]. Interestingly, five H1N1, three H1N2, and four H3N2 virus isolates retrieved between January 2010 and August 2011 had significant similarities with other European isolates which was an evidence of continent-wide transmission of these swine influenza viruses [220].

3.4.10. Luxembourg
A molecular investigation reported a negative prevalence of IDV in swine in Luxembourg during 2009 but later successfully detected three IDV positive swine during 2014–2015. Further, the serological investigation confirmed that swine in Luxembourg were free from IDV during 2012 but interestingly, IDV antibodies were detected in 17 swine samples collected during 2014–2015. These observations suggested that IDV was not in circulation in swine in Luxembourg during 2009–2012 but became prevalent at a low frequency later during 2014–2015 [46] which was almost the same time IDV was reported in Italian swine populations [45].

3.4.11. The Netherlands
A serological investigation of swine in the Netherlands identified 601 H1N1, 584 H1N2, and 229 H3N2 virus antibodies in 29 swine herds during January–May 1999 [222] with no further evidence of IAV in swine in the country after that.

3.4.12. Norway
After the 18 swine which were experiencing influenza-like illness were found infected with A(H1N1)pdm09 virus on a Norwegian swine farm in October 2009 the surveillance was expanded to the 39 nearby swine farms which determined that 23 of these farms were positive for the A(H1N1)pdm09 virus. Intriguingly, one human subject at the index farm who had influenza-like symptoms was also found positive for A(H1N1)pdm09 virus. This study suggested that the symptoms first appeared in the human subject at the index farm and later the disease got transmitted to the swine. Hence the findings of this study suggested the reverse zoonosis of the influenza virus from human to pig [223].
Further molecular and serological investigations identified 48 more swine herds that were positive for IAV during September 2009–October 2010 [224]. A more comprehensive nation-wide surveillance in Norwegian swine identified 16 A(H1N1)pdm09 virus positive swine herds during 2009 which later increased to 190 swine herds in 2010 [225]. Later 194 more swine were found infected with A(H1N1)pdm09 virus in Norway between April and July 2011 and reported that the IAV infected swine took longer to weigh 100 kg body mass [226].

3.4.13. Poland
The first active IAV infection in swine in Poland was reported in 2010 when 21 oral fluid samples collected from three swine farms detected IAV [227]. Soon after, five avian-like H1N1 viruses were reported from the swine lung tissues during 2011–2013 [228]. Later a serological surveillance identified 1212 H1N1, 851 H1N2, 1012 H3N2, and 572 A(H1N1)pdm09 virus antibodies in swine during March 2011–February 2015 [229]. Surprisingly, 34 of these swine had antibodies against all four IAV subtypes i.e., H1N1, H1N2, H3N2, and A(H1N1)pdm09 viruses [229] suggesting the past co-infections.

3.4.14. Czechoslovakia
The human-like H3N2 virus was isolated from a swine in Czechoslovakia during 1969–1972 [230]; however, no other reports ever appeared from the country in later years.

3.4.15. Hungary
Complete genome of an H1N1 virus was reported from a Hungarian swine having fever and conjunctivitis in May 2011 [231]. This was the only report of H1N1 virus in the swine in Hungary.

3.4.16. Multi-National Surveillances in European Countries
A large-scale investigation across seven European countries reported a high seroprevalence (˃62%) of IAV antibodies in swine populations of Belgium, Germany, Spain, Italy while a relatively lower (˂21.25%) seroprevalence was observed in swine populations of Czech Republic, Poland and Ireland during 2002–2003. Antibodies against H1N1, H1N2, and H3N2 viruses were reported in swine from the European countries under surveillance except Poland where swine had antibodies against only H1N1 virus [232].
A virological surveillance across five European countries including Belgium, United Kingdom, Italy, France and Spain reported 169 IAV positive swine during 2006–2008. The H1N1, H1N2, and H3N2 viruses were detected in swine from Belgium, Italy, and Spain while the samples from United Kingdom and France were found infected with H1N1 and H1N2 viruses [233].
Briefly, the virological and/or serological prevalence of influenza viruses in European countries (Figure 4D) identified the strains of H1N1, H1N2, H3N2, and A(H1N1)pdm09 viruses in swine populations of the United Kingdom, Ireland, Italy, Germany, France, Norway, Finland, Denmark, Belgium, Spain, Poland, Greece, Hungary, Netherlands, Czech Republic, and Czechoslovakia while the swine in Luxembourg and Italy were found infected with IDV.