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Recent epidemiological studies have discovered that a lot of cases of porcine epidemic diarrhea virus (PEDV) infection are frequently accompanied by porcine kobuvirus (PKV) infection, suggesting a potential relationship between the two viruses in the development of diarrhea. To investigate the impact of PKV on PEDV pathogenicity and the number of intestinal lymphocytes, piglets were infected with PKV or PEDV or co-infected with both viruses. Our findings demonstrate that co-infected piglets exhibit more severe symptoms, acute gastroenteritis, and higher PEDV replication compared to those infected with PEDV alone. Notably, PKV alone does not cause significant intestinal damage but enhances PEDV's pathogenicity and alters the number of intestinal lymphocytes. These results underscore the complexity of viral interactions in swine diseases and highlight the need for comprehensive diagnostic and treatment strategies addressing co-infections., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Porcine epidemic diarrhea virus (PEDV) is a virus that can cause diarrhea in pigs, resulting in significant economic losses to the pig industry. The mutation of the virus and its co-infection with other enteroviruses leads to poor control of PEDV infection. In this study, we found that the diarrhea outbreak in a pig farm in Shandong Province was mainly caused by PEDV infection. Through high-throughput sequencing, we also detected one other diarrhea-related virus (porcine kobuvirus). In the phylogenetic analysis and molecular characterization of the detected PEDV S gene and PKV, it was found that the S gene of the PEDV strain detected in this study (named SD22-2) had more mutations than the CV777 strain. The highest homology between PKV (named SD/2022/China) detected in this study and other strains was only 89.66%. Based on polyprotein, we divided SD/2022/China strains into a new grouping (designated group 4) and detected recombination signals. In summary, SD22-2 detected in this study is a new PEDV variant strain, and SD/2022/China strain might be a novel PKV strain. We also found the co-infection of the new PEDV variant and the novel PKV isolated from piglets with diarrhea. Our data suggested the importance of continuous surveillance of PEDV and PKV., (© 2023. The Author(s) under exclusive licence to Sociedade Brasileira de Microbiologia.)

The newly identified porcine Kobuvirus (PKV) has raised concerns owing to its association with diarrheal symptom in pigs worldwide. The process involving the emergence and global spread of PKV remains largely unknown. Here, the origin, genetic diversity, and geographic distribution of PKV were determined based on the available PKV sequence information. PKV might be derived from the rabbit Kobuvirus and sheep were an important intermediate host. The most recent ancestor of PKV could be traced back to 1975. Two major clades are identified, PKVa and PKVb, and recombination events increase PKV genetic diversity. Cross-species transmission of PKV might be linked to interspecies conserved amino acids at 13-17 and 25-40 residue motifs of Kobuvirus VP1 proteins. Phylogeographic analysis showed that Spain was the most likely location of PKV origin, which then spread to pig-rearing countries in Asia, Africa, and Europe. Within China, the Hubei province was identified as a primary hub of PKV, transmitting to the east, southwest, and northeast regions of the country. Taken together, our findings have important implications for understanding the evolutionary origin, genetic recombination, and geographic distribution of PKV thereby facilitating the design of preventive and containment measures to combat PKV infection.

Porcine kobuvirus (PKV) infection is very common in both healthy pigs and diarrhea pigs throughout the world. However, there is no proof that it causes diarrhea, and little is known about its role in diarrhea. There are only a few reports concerning porcine kobuvirus separation at present, which makes investigating its invasion and pathogenesis mechanisms difficult. This study sequenced the entire genome of a porcine kobuvirus strain termed "Wuhan2020" after it was isolated from intestinal tissue samples of healthy piglets. The analysis results revealed that it shared the most resemblance with the WUH1 strain (89.5%) and belonged to the same evolutionary branch as the Hungarian strain S-1-SUN. The PKV was located using the in situ hybridization (ISH) approach, which revealed that it was colonized in intestinal villus epithelial cells and lymphocytes in the Peyer's patch. In general, we analyzed the genetic evolution of PKV, discovered PKV susceptible cells and determined PKV localization in the intestine of infected pigs, providing a reference for future research., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

Porcine epidemic diarrhea virus (PEDV) frequently causes diarrhea outbreaks. However, whether newly discovered enteric viruses such as porcine kobuvirus (PKV) and porcine astroviruses (PAstVs) are also correlated with diarrhea is still unclear. Diarrhea outbreaks were reported in a PEDV-vaccinated pig farm in Xinjiang Uygur Autonomous Region of China from 2019 to 2020. PEDV was a common pathogen detected in fecal samples by routine RT-PCR assays. The PEDV positive fecal sample was used for pathogenic analysis due to the failure isolation of PEDV. The challenged neonatal piglets appeared watery diarrhea within one day post infection (dpi) and all died within 6 dpi. Histopathological and immunohistochemical examinations supported that PEDV is a major pathogen causing intestinal lesions. To further explore enteric viruses associated with neonatal piglet diarrhea, metagenomics sequencing was performed for the diarrheic piglets. Remarkably, PKV was the most abundant virus (58.33%) followed by PEDV (34.45%) and PAstVs (7.22%), which were also confirmed by real-time RT-PCR assays. Significant in vivo replications of PEDV and PKV could only be observed in challenged piglets whilst PAstVs maintained similar virus loads in both challenged and mock infected piglets. Overall, this study provides first pathogenic and metagenomic evidence that significant proliferations of PEDV and PKV are closely associated with severe diarrhea in neonatal piglets, while PAstVs likely play limited roles in neonatal piglet diarrhea., Competing Interests: Declaration of competing interest The authors declare no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Many enteric viruses are found in pig farms around the world and can cause death of animals or important production losses for breeders. Among the wide spectrum of enteric viral species, porcine Sapelovirus (PSV), porcine Kobuvirus (PKoV) and porcine Astrovirus (PAstV) are frequently found in pig feces. In this study we investigated sixteen pig farms in Corsica, France, to evaluate the circulation of three enteric viruses (PKoV, PAstV-1 and PSV). In addition to the three viruses studied by RT-qPCR (908 pig feces samples), 26 stool samples were tested using the Next Generation Sequencing method (NGS). Our results showed viral RNA detection rates (i) of 62.0% [58.7-65.1] (n = 563/908) for PSV, (ii) of 44.8% [41.5-48.1] (n = 407/908) for PKoV and (iii) of 8.6% [6.8-10.6] (n = 78/908) for PAstV-1. Significant differences were observed for all three viruses according to age (P-value = 2.4e-13 for PAstV-1; 2.4e-12 for PKoV and 0.005 for PSV). The type of breeding was significantly associated with RNA detection only for PAstV-1 (P-value = 9.6e-6). Among the 26 samples tested with NGS method, consensus sequences corresponding to 10 different species of virus were detected. This study provides first insight on the presence of three common porcine enteric viruses in France. We also showed that they are frequently encountered in pigs born and bred in Corsica, which demonstrates endemic local circulation., Competing Interests: The authors have declared that no competing interests exist.

Porcine Kobuvirus (PKV) infection is very common in pigs throughout the world. Since it has never been investigated in Serbia, to contribute to the knowledge of Porcine Kobuvirus, its role, and distribution, we tested 200 samples from domestic pigs and wild boars. From domestic pigs, 10 fecal, 22 spleen and 68 serum samples, and 100 spleen samples from wild boars were tested. The virus prevalence determined by real-time RT-PCR in domestic pigs was 22% and in wild boars 6%. The phylogenetic analysis of 3D region revealed that Serbian strains are closest related to the Hungarian strain from wild boar from 2011. This is the first report on PKV in Serbia in domestic pigs and wild boars, implying its wide circulation. Although the infection could not be directly related to any clinical manifestation, the frequency of virus found in feces suggests viral affinity to the gastrointestinal tract. However, due to the rather ubiquitous presence of PKV, the clinical and pathological assessment have to be considered when PKV infection is diagnosed., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The major enteric RNA viruses in pigs include porcine epidemic diarrhoea virus (PEDV), transmissible gastroenteritis virus (TGEV), porcine rotavirus A (PRV-A), porcine kobuvirus (PKV), porcine sapovirus (PSaV) and porcine deltacoronavirus (PDCoV). For differential diagnosis, a multiplex RT-PCR method was established on the basis of the N genes of TGEV, PEDV and PDCoV, the VP7 gene of PRV-A, and the polyprotein genes of PKV and PSaV. This multiplex RT-PCR could specifically detect TGEV, PEDV, PDCoV, PRV-A, PKV and PSaV without cross-reaction to any other major viruses circulating in Chinese pig farms. The limit of detection of this method was as low as 10 0 -10 1  ng cDNA of each virus. A total of 398 swine faecal samples collected from nine provinces of China between October 2015 and April 2017 were analysed by this established multiplex RT-PCR. The results demonstrated that PDCoV (144/398), PSaV (114/398), PEDV (78/398) and PRV-A (70/398) were the main pathogens, but TGEV was not found in the pig herds in China. In addition, dual infections, for example, PDCoV + PSaV, PDCoV + PRV-A, PRA-V + PSaV and PEDV + PDCoV, and triple infections, for example, PDCoV + PRV-A + PSaV and PEDV + PDCoV + PKV, were found among the collected samples. The multiplex RT-PCR provided a valuable tool for the differential diagnosis of swine enteric viruses circulating in Chinese pig farms and will facilitate the prevention and control of swine diarrhoea in China., (© 2019 Blackwell Verlag GmbH.)

Enteric diseases in swine are often caused by different pathogens and thus metagenomics are a useful tool for diagnostics. The capacities of nanopore sequencing for viral diagnostics were investigated here. First, cell culture-grown porcine epidemic diarrhea virus and rotavirus A were pooled and sequenced on a MinION. Reads were already detected at 7 seconds after start of sequencing, resulting in high sequencing depths (19.2 to 103.5X) after 3 h. Next, diarrheic feces of a one-week-old piglet was analyzed. Almost all reads (99%) belonged to bacteriophages, which may have reshaped the piglet's microbiome. Contigs matched Bacteroides, Escherichia and Enterococcus phages. Moreover, porcine kobuvirus was discovered in the feces for the first time in Belgium. Suckling piglets shed kobuvirus from one week of age, but an association between peak of viral shedding (10 6.42 -10 7.01 copies/swab) and diarrheic signs was not observed during a follow-up study. Retrospective analysis showed the widespread (n = 25, 56.8% positive) of genetically moderately related kobuviruses among Belgian diarrheic piglets. MinION enables rapid detection of enteric viruses. Such new methodologies will change diagnostics, but more extensive validations should be conducted. The true enteric pathogenicity of porcine kobuvirus should be questioned, while its subclinical importance cannot be excluded.

In this study, a multiplex RT-PCR method was developed for detection of seven diarrhea-associated porcine viruses, including porcine teschovirus (PTV), porcine sapovirus (PSV), porcine deltacornavirus (PDCoV), porcine kobuvirus (PKV), porcine sapovirus (PSaV), porcine astrovirus (PAstV) and porcine torovirus (PToV). A total of 419 samples were screened by this method and results showed that PKV had the highest positive rate of 26.98%?45.79% and its mixed infection rate reached 9.52%-18.54%. On account of high positive rate of PKV and its important role in diarrhea disease, complete genomic sequences of three PKV positive samples were further sequenced. Three PKV labeled as PD-PKV, JS-PKV and CM-PKV were classified into porcine kobuvirus genus and had far genetic distance with other kobuviruses. The complete genome homologies among them were 88.1%-89.1%. CM-PKV had the highest identity with the Chinese strain JS-02a-CHN/2013 reported in 2013 while JS-PKV and PD-PKV were most closed to the K-30-HUN/2008/HUN strain reported in Hungary in 2008. This illustrates the significant genetic differences of the different PKV isolates in Shanghai while its relationship with the viral pathogenicity still needs to be explored. This research provides references for further understanding the prevalence of PKV and its role in swine diarrhea.

Porcine kobuviruses (PoKoVs) are ubiquitously distributed in pig populations worldwide and are thought to be enteric viruses in swine. Although PoKoVs have been detected in pigs in Japan, no complete genome data for Japanese PoKoVs are available. In the present study, 24 nearly complete or complete sequences of the PoKoV genome obtained from 10 diarrheic feces and 14 non-diarrheic feces of Japanese pigs were analyzed using a metagenomics approach. Japanese PoKoVs shared 85.2-100% identity with the complete coding nucleotide (nt) sequences and the closest relationship of 85.1-98.3% with PoKoVs from other countries. Twenty of 24 Japanese PoKoVs carried a deletion of 90 nt in the 2B coding region. Phylogenetic tree analyses revealed that PoKoVs were not grouped according to their geographical region of origin and the phylogenetic trees of the L, P1, P2, and P3 genetic regions showed topologies different from each other. Similarity plot analysis using strains from a single farm revealed partially different similarity patterns among strains from identical farm origins, suggesting that recombination events had occurred. These results indicate that various PoKoV strains are prevalent and not restricted geographically on pig farms worldwide and the coexistence of multiple strains leads to recombination events of PoKoVs and contributes to the genetic diversity and evolution of PoKoVs.

Since the first report of porcine kobuvirus (PKV) in 2008, it has been confirmed that PKV is widely present in several countries and plays an important role in diarrhoea outbreak in pigs. Studies have shown that the biggest difference among PKVs is the presence or absence of a specific 30-amino acid (aa) sequence in the 2B region of the polyprotein gene. Based on this unique feature, most PKV sequences could be divided into two groups (Group 1 without deletion and Group 2 with deletion), but a few sequences did not follow this rule due to possible recombination. In this study, two PKV genome sequences, designated JXAT2015 (8,123 nucleotide) and JXJC2015 (8,120 nucleotide), were identified on two different commercial swine farms with the severe diarrhoea outbreak accompanying with highly PKV infection (90%, 18/20) and moderate infection (40%, 8/20) of porcine bocavirus 1 (PBoV1) in Jiangxi province of China. Sequence analysis based on the polyprotein gene showed that they shared low nucleotide similarity (86.3%-88.1%) with other known PKVs. Although both possessed the 30-aa deletion in the 2B region, phylogenetic analysis showed that JXJC2015 was distinct from Group 1 and even Group 2, and formed a new Group (designated Group 3). The findings of this study further revealed genetic diversity and the possible pathogenic role of PKV in conjunction with other pathogens in piglets., (© 2017 Blackwell Verlag GmbH.)

A total of 636 fecal samples collected from piglets with and without diarrhea during 2009 to 2013 were tested for porcine kobuvirus by RT-PCR. From a total of 528 fecal samples collected from piglets with diarrhea and 108 from healthy controls, 505 (95.6%) and 104 (96.3%) were positive for porcine kobuvirus, respectively. The detection rates of porcine kobuvirus were remarkable equally high in both diarrheic and healthy piglets. Phylogenetic analysis revealed that porcine kobuvirus strains detected in both symptomatic and asymptomatic piglets were genetically closely related to each other and also to other porcine kobuviruses reported worldwide. It was interesting to point out that one of the porcine kobuvirus strains isolated from piglet in our study was similar to a porcine-like bovine kobuvirus reference strain isolated previously in South Korea. This finding provided the evidence to support the interspecies transmission of kobuviruses between cattle and swine.

Kobuviruses have been detected in a wide range of mammals including cats, dogs, pigs, cattle, goats, sheep and bats. Kobuviruses have been detected in symptomatic and asymptomatic animals; however, the clinical significance of infection in animals is still unclear. To date, there is no information regarding kobuvirus prevalence in livestock in Ireland. This study reports the first detection of kobuviruses in pigs, bovines and ovines using quantitative PCR. In this study, mesenteric lymph node was collected from cattle (n = 57), pigs (n = 53) and sheep (n = 50) from farms in Northern Ireland and the Republic of Ireland, from animals which had been submitted by private veterinary practitioners from 2009 to 2011 for routine post mortem and clinico-pathological examination. Kobuviruses were detected in 14 cows (24.5%), 5 pigs (9.4%) and 1 sheep (2%). Phylogenetic analysis of Irish kobuviruses from cattle and pigs revealed that the isolates clustered according to their host species. Interestingly, the sheep kobuvirus clustered with bovine kobuviruses detected in this study and other published kobuvirus strains. The data presented in this study contributes to the understanding of the epidemiology of these viruses in animals and to the genetic diversity that these viruses possess.

The porcine kobuvirus 1 (PKV-1) is believed to be an enteric virus. To investigate the prevalence of PKV-1 in pigs, virus was detected by RT-PCR in rectal swabs originating from 414 healthy and diarrheic pigs of different age categories on farms in Slovakia. Among all ages of animals, PKV-1 was detected equally in diarrheic (63.8%) and clinically healthy (62.9%) pigs. PKV-1 was more often detected in diarrheic (74.6%) than in healthy (64.4%) suckling piglets (<28days) but data was not statistically significant. Results in weaned (28-70days) and fattening (>70days) of both healthy and diarrheic pigs were inconsistent ranging in interval 56.2% to 67.9%. This study did not confirm a clear relationship of PKV-1 infection with diarrhea in pigs. Rotavirus A infection was detected among the same animals in 39% diarrheic and 9.2% healthy suckling piglets (p<0.001) confirming rotavirus as a causative agent of diarrhea in this age group. The difference was not significant in older pigs with both diarrheic and healthy pigs being infected within a range of 0% to 12.2%. Co-infection with PKV-1 and rotavirus A was detected overall in 5.6% of healthy and in 13.5% of diarrheic pigs and was highest in suckling piglets (33.9%). The PKV-1sequences from pigs in Slovakia were analyzed at the genetic level in the partial 3D gene region for the first time. The viral sequences were grouped in phylogenetic clusters according to their farm of origin. When compared with 157 nucleotide sequences originating from pig samples of different countries around the world Slovakian PKV-1 sequences were clustered in the phylogenetic tree with Asian sequences but not with nucleotide sequences from the neighbouring countries of Czech Republic or Hungary., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

Porcine kobuvirus (PKV) is a newly emerging virus that has been detected in diarrheic pigs. Presently, reverse transcription-polymerase chain reaction (RT-PCR) and RT-loop-mediated amplification are the only methods that can be used to detect PKV. To develop a TaqMan real-time RT-PCR for the rapid detection and quantitation of PKV nucleic acid in fecal samples, a pair of primers and a probe were designed to amplify the conserved 3D region of the PKV genome. After optimization, the TaqMan real-time RT-PCR was highly specific and ∼1000 times more sensitive than conventional RT-PCR, and the detection limit was as low as 30 DNA copies. Among the 148 intestinal samples from piglets with diarrhea, 136 and 118 were positive based on the TaqMan and conventional RT-PCR methods, respectively, indicating that the TaqMan RT-PCR was more sensitive than conventional RT-PCR, and the total concordance of the two methods was approximately 87.84%. Thus, the TaqMan real-time RT-PCR should be a useful tool for the early detection and quantitation of PKV., (Copyright © 2016. Published by Elsevier B.V.)

Piglet diarrhea epidemics result in major economic losses for the swine industry. Four viruses are closely linked to porcine diarrhea: porcine kobuvirus (PKV), porcine epidemic diarrhea virus (PEDV), porcine transmissible gastroenteritis virus (TGEV), and porcine rotavirus (PRoV). We have conducted an epidemiology study to determine the frequency of infection and co-infection with these viruses in China, and characterized the genetic variation of the isolated PEDV and PKV strains. Stool and intestinal samples (n = 314) were collected from piglets with diarrhea in China from years 2012 to 2014. RT-PCR was used to detect PKV, PEDV, TGEV, and PRoV. Phylogenetic relationships between reference strains and the isolated PEDV and PKV strains were determined based on the M and 3D gene sequence. The rates of infection with PKV, PEDV, TGEV and PRoV were 29.9%, 24.2%, 1.91%, and 0.31%, respectively. Co-infections with PKV and the other three viruses were very common. Co-infection of PKV and PEDV was detected in 15.0% (47/314) of the samples. Phylogenetic analysis of the PKV 3D gene indicated that there were some phylogenetic differences in the PKV strains across regions within China. However, according to the PEDV M gene, strains clustered into three groups and the primary group was distinct from the vaccine strain CV777. This study provides insights in to the prevalence of diarrhea viruses and their prevention and control in China.

A recent survey of pigs in Dong Thap province, Vietnam identified a high frequency of enterovirus species G (EV-G) infection (144/198; 72.7%). Amongst these was a plethora of EV-G types (EV-G1, EV-G6 and four new types EV-G8-EV-G11). To better characterize the genetic diversity of EV-G and investigate the possible existence of further circulating types, we performed a larger-scale study on 484 pig and 45 farm-bred boar faecal samples collected in 2012 and 2014, respectively. All samples from the previous and current studies were also screened for kobuviruses. The overall EV infection frequency remained extremely high (395/484; 81.6%), but with comparable detection rates and viral loads between healthy and diarrhoeic pigs; this contrasted with less frequent detection of EV-G in boars (4/45; 8.9%). EV was most frequently detected in pigs ≤ 14 weeks old (∼ 95%) and declined in older pigs. Infections with EV-G1 and EV-G6 were most frequent, whilst less commonly detected types included EV-G3, EV-G4 and EV-G8-EV-G11, and five new types (EV-G12-EV-G16). In contrast, kobuvirus infection frequency was significantly higher in diarrhoeic pigs (40.9 versus 27.6%; P = 0.01). Kobuviruses also showed contrasting epizootiologies and age associations; a higher prevalence was found in boars (42%) compared with domestic pigs (29%), with the highest infection frequency amongst pigs >52 weeks old. Although genetically diverse, all kobuviruses identified belonged to the species Aichivirus C. In summary, this study confirms infection with EV-G was endemic in Vietnamese domestic pigs and exhibits high genetic diversity and extensive inter-type recombination.

Two-hundred eight swine fecal samples from six Italian farms were tested using a kobuvirus-specific RT-PCR with primers that amplify a region within the 3D gene. All farms were kobuvirus positive, with prevalence rates ranging between 24 % and 84 %. Overall, 57.5 % of asymptomatic pigs and 49.7 % of animals with diarrhea were positive for kobuvirus. Sequence analysis showed a different predominant strain circulating on each farm and indicated that the strains detected were related to both European and Asiatic strains. A possible pathogenic role of kobuvirus should be investigated further, since infections with this virus occur frequently in pigs of different ages.

In this study, 396 samples from diarrheic pigs on 46 pig farms in Jiangsu Province, China, were analyzed by RT-PCR. One-hundred eighty-one pigs from 37 farms tested positive for porcine kobuvirus (PKV). Phylogenetic analysis of the 3D gene from 19 isolates showed sequence homology of 88.0 %-100 % and 69.4 %-100 % for nucleotides and amino acids, respectively, while similarity to isolates of other kobuviruses was 69.6 %-78.8 % and 27.8 %-56.9 %, respectively. One-hundred eighty-five samples contained two or more pathogens, and 31/68 PKV-positive samples tested positive for other diarrheic pathogens, confirming the existence of PKV infection and coinfection.

Porcine kobuvirus, a member of the genus Kobuvirus that is associated with diarrhea, has been reported in many countries. We determined the complete genome sequence and investigated the genetic evolution of the kobuvirus strain swKoV CH441, which was detected in the highland of Gansu province in Northwest China. The viral genome is 8149 nucleotides (nt) long, including a 29-nt poly(A) tail of the 3' end, and is 90 nt shorter in the 2B coding region than those of other kobuvirus strains whose sequences are available in the GenBank database. Phylogenetic analysis showed that swKoV CH441 was most closely related to porcine kobuvirus CH/HNXX-4 but more distantly related to other strains, including the strains GS-1/2012/CH and GS-2/2012/CH, which were detected in Gansu province, indicating that porcine kobuvirus may have geographic and host differences in evolution.

Aichivirus C is the third species in the genus Kobuvirus, family Picornaviridae, and the virus is circulating in pigs worldwide. Aichivirus A in humans and Aichivirus B in cows have been shown to associate with diarrheal diseases, however, the pathogenesis of Aichivirus C has not been demonstrated clearly. In this study, the full genome nucleotide sequence of the Thai strain, CMP06/2007/THA collected from stool sample of a diarrheal piglet was analyzed and identified as a variant type with a 90-nt deletion in the 2B-coding region. In addition, molecular characterization of nucleotide sequences of the 2B-coding region of Aichivirus C strains from six diarrheal and six healthy piglets in Thailand, and four strains from healthy pigs in Japan revealed that all of the strains in this study were variant types. These findings indicate that variant strains of Aichivirus C are circulating in Asian countries such as China, Thailand and Japan, and deletion of tandem repeat of 2B-region is unlikely to associate with the pathogenesis of the virus., (Copyright © 2014 Elsevier B.V. All rights reserved.)

In this study, we report the complete nucleotide sequence of porcine kobuvirus CH/HZ/2011 (CH/HZ) isolated from infected piglets in China. Dendrograms indicated that CH/HZ is closely related to other porcine kobuviruses. The complete genome sequence of CH/HZ has an adenine insertion at position 124, an 18-nt deletion in the 5'UTR, and a 90-nt deletion in the 2B coding region. A neighbor-joining tree based on the VP1 gene indicated that 31 porcine kobuvirus strains are divided into four clusters (Thai-Japanese group, Hungarian group, Chinese group I and Chinese group II). The strain CH/HZ was included in Chinese group II.

The aim of this study was to evaluate the frequency of Aichivirus C infection in the three major industrial pig-producing regions of Brazil. This retrospective study evaluated 63 faecal samples that were collected between 2004 and 2011 from suckling piglets (1 to 3 weeks old) belonging to 46 pig herds located in the South, Southeast and Central-west regions of Brazil. The presence of Aichivirus C in the samples was evaluated by reverse transcription-polymerase chain reaction assay. A 216-bp fragment of the aichivirus RNA-dependent RNA polymerase gene was amplified from 48 out of 63 (76.2 %) faecal samples. Aichivirus C was identified in all the evaluated regions, and variations in the frequency of virus detection among the different Brazilian regions were observed. Aichivirus C was detected at significantly higher rates in 2-week-old (18/20; 90.0 %) and 3-week-old (19/22; 86.4 %) piglets than in 1-week-old piglets (11/21; 52.4 %) (P < 0.05). The nucleotide sequences of three amplicons clustered together with other sequences previously isolated in Brazil, revealing that there were no phylogenetic differences in the strains of Aichivirus C by region or age. This study demonstrated that Aichivirus C has been present in Brazilian pig herds for years and has spread to the major pig-producing regions of the country.

Kobuviruses comprise three species, the Aichivirus A, Aichivirus B, and Aichivirus C (porcine kobuvirus). Porcine kobuvirus is endemic to pig farms and is not restricted geographically but, rather, is distributed worldwide. The complete genomic sequences of four porcine kobuvirus strains isolated during a diarrhea outbreak in piglets in the Gansu province of China were determined. Two of these strains exhibited variations relative to the traditional strains. The potential 3C/3D cleavage sites of the variant strains were Q/C, which differed from the Q/S in the traditional porcine kobuvirus genome. A 90-nucleotide deletion in the 2B protein and a single nucleotide insertion in the 3'UTR were found in the variant strains. The VP1 regions of all four porcine kobuviruses in our study were highly variable (81%-86%). Ten common amino acid mutations were found specifically at certain positions within the VP1 region. Significant recombination sites were identified using SimPlot scans of whole genome sequences. Porcine kobuviruses were also detected in pig serum, indicating that the virus can escape the gastrointestinal tract and travel to the circulatory system. These findings suggest that mutations and recombination events may have contributed to the high level of genetic diversity of porcine kobuviruses and serve as a driving force in its evolution.

In this study, fecal specimens (n=260) collected from Italian healthy pigs aged between 6 and 10 months were screened by RT-PCR using generic kobuvirus primers. Porcine kobuviruses (PKVs) were detected in 3.85% of the samples tested. Based on the analysis of the partial 3D gene, the Italian sequences identified here were more closely related to PKVs previously identified in Japan, Thailand, Korea, China and Brazil than to the European PKVs recently detected in Hungary and in the Netherlands., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Background: Porcine kobuvirus (PKoV) is a member of the Kobuvirus genus within the Picornaviridae family. PKoV is distributed worldwide with high prevalence in clinically healthy pigs and those with diarrhea., Methods: Fecal and intestinal samples (n = 163) from pig farms in Sichuan Province, China were obtained to determine the presence of PKoV using reverse transcription polymerase chain reaction assays. Specific primers were used for the amplification of the gene encoding the PKoV VP1 protein sequence. Sequence and phylogenetic analyses were conducted to clarify evolutionary relationships with other PKoV strains., Results: Approximately 53% (87/163) of pigs tested positive for PKoV. PKoV was widespread in asymptomatic pigs and those with diarrhea. A high prevalence of PKoV was observed in pigs younger than 4 weeks and in pigs with diarrhea. Phylogenetic analysis of 36 PKoV VP1 protein sequences showed that Sichuan PKoV strains formed four distinct clusters. Two pigs with diarrhea were found to be co-infected with multiple PKoV strains. Sequence and phylogenetic analyses revealed diversity within the same host and between different hosts. Significant recombination breakpoints were observed between the CHN/SC/31-A1 and CHN/SC/31-A3 strains in the VP1 region, which were isolated from the same sample., Conclusion: PKoV was endemic in Sichuan Province regardless of whether pigs were healthy or suffering from diarrhea. Based on our statistical analyses, we suggest that PKoV was the likely causative agent of high-mortality diarrhea in China from 2010. For the first time, we provide evidence for the co-existence of multiple PKoV strains in one pig, and possible recombination events in the VP1 region. Our findings provide further insights into the molecular properties of PKoV, along with its epidemiology.

The prevalence of porcine sapoviruses (SaVs) and noroviruses (NoVs) in nursing piglets on three pig farms in Ohio was studied. Fecal samples (n = 139) were collected from individual pigs and screened for caliciviruses by RT-PCR. Phylogenetic analysis was conducted using partial sequences of the RNA polymerase region. Three different SaV genogroups, including a newly emerging one (DO19 Korea-like) were detected. No NoVs were detected. Kobuviruses, emerging members of the family Picornaviridae, were detected by primers designed for SaV. To our knowledge, this is the first report of porcine DO19 Korea-like SaV and kobuvirus in the United States.

Small, non-enveloped RNA viruses belonging to the genera Sapovirus, Kobuvirus, and Mamastrovirus are usually associated with gastroenteritis in humans and animals. These enteric pathogens are considered potential zoonotic agents. In this study, the prevalence and genetic diversity of sapoviruses (SaVs), kobuviruses (KoVs), and astroviruses (AstVs) in asymptomatic pigs were investigated using a PCR approach. KoV was found to be the most prevalent virus (87.3 %), followed by AstV (34.2 %) and SaV (10.2 %). Interestingly, the intra- and inter-cluster distances between porcine SaV capsid sequences revealed one strain (P38/11/CZ) that formed a new genotype within genogroup III of porcine SaVs, and it is tentatively called "P38/11-like" genotype. Moreover, this is the first report of porcine kobuvirus detection on Czech pig farms. The high prevalence rate of gastroenteritis-producing viruses in clinically healthy pigs represents a continuous source of infection of pigs, and possibly to humans.

Fecal samples (N = 10) from 6- to 8-week-old wild boar piglets (Sus scrofa), collected from an animal park in Hungary in April 2011, were analyzed using viral metagenomics and complete genome sequencing. Kobuvirus (genus Kobuvirus, family Picornaviridae) was detected in all (100 %) specimens, with the closest nucleotide (89 %) and amino acid (94 %) sequence identity of the strain wild boar/WB1-HUN/2011/HUN (JX177612) to the prototype porcine kobuvirus S-1-HUN (EU787450). This study suggests that genetically highly similar (practically the same geno-/serotype) porcine kobuvirus circulate in wild boars, the wildlife counterparts of domestic pigs. Wild boars could be an important host and reservoir for kobuvirus.

Porcine kobuvirus, an emerging virus, was first identified in Hungary in 2007. We report here the complete genome sequence of porcine kobuvirus strain WUH1 isolated from piglets with severe diarrhea, which will help toward understanding the molecular and evolutionary characteristics of the porcine kobuvirus.

Porcine kobuvirus is a new candidate species of the genus Kobuvirus in the family Picornaviridae, and information is still limited. The identification of porcine kobuvirus has been performed by the sequence analyses of the 3D region of the viruses. Therefore, the purpose of this study was to characterize the molecular properties of VP1 nucleotide sequences of the porcine kobuviruses isolated from porcine stool samples in Japan during 2009 and Thailand between 2006 and 2008. In addition, previous identification of a unique porcine kobuvirus; Japanese H023/2009/JP, which is a bovine kobuvirus-like strain based on sequence analysis of the 3D region, was also included in this study. All of the strains were amplified by the VP1-specific primer pair: the amplicons were subjected to direct sequencing and compared with the VP1 nucleotide sequences of reference strains. The VP1 sequences of strains from the GenBank database revealed high nucleotide sequence identity at 84.3-100%. On the other hand, the nucleotide identities among the 15 porcine kobuvirus strains analyzed in this study ranged from 78.8 to 99.8%. The results revealed that diversity of the strains in this study were higher than those of the strains in previous studies. Furthermore, it was found that the VP1 region of the bovine kobuvirus-like strain, H023/2009/JP, clustered with nine porcine kobuvirus strains that were isolated in Thailand and Japan. Since this strain was previously found to be closely related to bovine kobuviruses in the 3D gene region, it may be a natural recombinant.

Porcine kobuvirus was first identified in 2007 in Hungary. The virus has been detected in several Asian countries. In our study, the complete genome of the recently identified porcine kobuvirus strain SH-W-CHN was amplified by RT-PCR and was sequenced. Dendrograms indicated that SH-W-CHN is closely related to other porcine kobuviruses. To identify sites of possible recombination within the genome of the SH-W-CHN strain, the SimPlot program was used to perform recombination analysis. The results showed that no significant recombination event between strain S-1-HUN and Y-1-CHI had occurred. However, certain possible recombination signals were identified, indicating that some early recombination events may have contributed to the genome of SH-W-CHN. This study further confirmed the existence of multiple lineages of porcine kobuvirus and indicated that homologous recombination may be a driving force in its evolution.

In this study, a total of 116 stool specimens were collected from pigs of different ages in three pig farms of Shanghai in China during 2010. Forty-five (38.8%) stool specimens were positive for porcine kobuvirus using reverse transcriptase-polymerase chain reaction (RT-PCR). The prevalence rate of porcine kobuvirus in the three pig farms which are located in Minhang District, Qingpu District and Fengxian District was 46.7%(21/45), 35.1%(13/37), and 32.4%(11/34), respectively. We demonstrated that porcine kobuvirus infections are existent in certain domestic pigs in Shanghai and high prevalence of this virus was found in the piglets under the age of 6 weeks and in pigs with diarrhea in Shanghai. Twenty-seven representative strains of porcine kobuvirus detected in this study were randomly selected based on the equal distribution of the sampling date throughout the study and analyzed for their phylogenetic relationships with other kobuvirus reference strains. The phylogenetic analysis suggested the presence of multiple porcine kobuvirus lineages in Shanghai, China.

To survey for porcine kobuvirus infection, a total of 119 pig fecal samples in Korea were collected from three pig farms with good breeding facilities in three provinces. Forty-three (36.1%) of the fecal samples tested were positive for porcine kobuvirus. In addition, porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) infections were not identified in any of the fecal samples, but porcine group A rotavirus (GAR) infection was identified in 11.8% (14/119) of the samples. Nucleotide sequence analysis showed sharp divergence between the five major Korean lineages by the Neighbor-joining method, and each of these lineages had high levels of bootstrap support (99 or 100%). Porcine kobuvirus is widespread in Korea regardless of the clinical condition (diarrhea or non-diarrhea). Piglets under the age of 3 weeks were the main age group infected with porcine kobuvirus, which is the general case for other species of kobuviruses.

Kobuviruses are small, non-enveloped viruses with a single-stranded, positive-sense genomic RNA, belonging to the family Picornaviridae, a highly diverse family of important pathogens of human and other animals. Porcine kobuvirus has been found recently, and consequently, information about the virus is lacking. In this study, we identified porcine kobuviruses from pigs in Korea by RT-PCR, cloning and sequencing, and we showed the existence of genetic diversity among geographically separated porcine kobuviruses through genetic and phylogenetic analysis. Epidemiological studies of porcine kobuvirus linked to diarrhea indicated that porcine kobuvirus infections are endemic in diarrheic pigs in Korea. Statistical analysis of the porcine kobuvirus positive rate between diarrheic and healthy pigs as well as a survey for other enteric pathogens in diarrheic pigs suggests that porcine kobuvirus may play a role as a causative agent of gastroenteritis in pigs.

Porcine kobuvirus was first identified in early 2007 in Hungary. Originally thought to be confined to the intestine, almost 2 years later the virus was found in the blood of clinically healthy pigs on the same farm. Porcine kobuvirus may be widely distributed on pig farms worldwide.