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Simple Summary: Dozens of species of small exotic mammals, such as chinchillas, golden hamsters, Java squirrels, Mongolian gerbils, mice, rats, African hedgehogs, and sugar gliders, are kept as pets globally. These exotic companion mammals are potential reservoirs for maintaining and transmitting zoonotic pathogens. Using molecular assays, our study aimed to investigate the occurrence of some potential zoonotic viruses in various small exotic mammals kept as pets in Italy. Our findings show that aichivirus (AiV) was further characterized as murine kobuvirus-1 in a rat (Rattus norvegicus). To the best of our knowledge, this is the first study reporting the detection of AiV in rodents in Italy. Our results also revealed the absence of other viruses investigated while highlighting the importance of ongoing monitoring of infectious agents in these increasingly common pets to prevent the spread of new potential zoonotic pathogens. We investigated the occurrence of eight potential zoonotic viruses in 91 exotic companion mammals from pet shops in southern Italy via real-time PCR and end-point PCR. The animals were screened for aichivirus, sapovirus, astrovirus, hepatitis A, noroviruses (GI and GII), rotavirus, circovirus, and SARS-CoV-2. Among the nine species of exotic pets studied, only one rat tested positive for aichivirus. The high sequence similarity to a murine kobuvirus-1 strain previously identified in China suggests that the virus may have been introduced into Italy through the importation of animals from Asia. Since exotic companion mammals live in close contact with humans, continuous sanitary monitoring is crucial to prevent the spread of new pathogens among domestic animals and humans. Further investigations on detecting and typing zoonotic viruses are needed to identify emerging and re-emerging viruses to safeguard public health. [ABSTRACT FROM AUTHOR]

This article aims at the presenting of emerging picornaviruses, namely the genus Kobuvirus. These have been reported to be associated with clinical problems including diarrhea, vomiting, fever, purulent conjunctivitis, and respiratory symptoms in humans. The molecular methods reveal that kobuviruses (the genus Kobuvirus (AiV, Aichivirus) Picornaviridae family), may be the causative agents of gastroenteritis in a variety of domestic and wild animal species in several regions of the world. They were isolated from a wide range of hosts: ferrets, mice, foxes, bats, dogs, cats, cattle, sheep, pigs, and goats. Kobuviruses are frequently present in mixed infections with other pathogens. Currently, there are six species within the genus Kobuvirus: Aichivirus A (humans, dogs, cats, rats, mice, and the European roller), Aichivirus B (cattle, ferrets, and sheep), Aichivirus C (pigs and goats), Aichivirus D (cattle), Aichivirus E (rabbits) and Aichivirus F (bats). AiVs stability allows persistence in the environment for long periods, as well as its transmission to humans with contaminated water and molluscs. Although until now AiVs are responsible for a low percentage of reported gastroenteritis outbreaks, the high seroprevalence shown in human populations, indicates their role as emerging enteric agents. Kobuviruses are difficult to isolate and the diagnosis is mainly based on molecular methods. Currently, there are no vaccines available. [ABSTRACT FROM AUTHOR]

We investigated the occurrence of eight potential zoonotic viruses in 91 exotic companion mammals from pet shops in southern Italy via real-time PCR and end-point PCR. The animals were screened for aichivirus, sapovirus, astrovirus, hepatitis A, noroviruses (GI and GII), rotavirus, circovirus, and SARS-CoV-2. Among the nine species of exotic pets studied, only one rat tested positive for aichivirus. The high sequence similarity to a murine kobuvirus-1 strain previously identified in China suggests that the virus may have been introduced into Italy through the importation of animals from Asia. Since exotic companion mammals live in close contact with humans, continuous sanitary monitoring is crucial to prevent the spread of new pathogens among domestic animals and humans. Further investigations on detecting and typing zoonotic viruses are needed to identify emerging and re-emerging viruses to safeguard public health.

Calf diarrhea results in significant economic loss and is caused by a variety of pathogens, including enteric viruses. Many of these viruses, including bovine norovirus (BNoV), bovine torovirus (BToV), and bovine kobuvirus (BKoV), are recognized as the causative agents of diarrhea; however, they remain understudied as major pathogens. We developed a multiplex reverse-transcription quantitative real-time PCR (RT-qPCR) assay for rapid and simple detection of BNoV, BToV, and BKoV. Our method had high sensitivity and specificity, with detection limits of 1 × 102 copies/μL for BNoV, BToV, and BKoV, which is a lower detection limit than conventional RT-PCR for BNoV and BKoV and identical for BToV. We tested fecal samples from 167 diarrheic calves with our multiplex RT-qPCR method. Viral detection was superior to conventional RT-PCR methods in all samples. The diagnostic sensitivity of the multiplex RT-qPCR method (100%) is higher than that of the conventional RT-PCR methods (87%). Our assay can detect BNoV, BToV, and BKoV in calf feces rapidly and with high sensitivity and specificity. [ABSTRACT FROM AUTHOR]

Simple Summary: In 2008, researchers described a new virus, porcine kobuvirus (PKV). Some viruses are very harmful, while others infect animals without having any significance for their health. Some researchers have suspected that porcine kobuvirus causes gastrointestinal disease (e.g., diarrhea) in pigs. Therefore, this paper tried to answer the question: Is porcine kobuvirus a cause of gastrointestinal disease in young pigs? A systematic literature review was conducted, meaning that a database was searched for all reports of research studies investigating kobuvirus in pigs. In general, there was not much research of good quality that could possibly answer the question. Therefore, the study concluded that there is a lack of good evidence supporting the idea that PKV causes gastrointestinal disease. The absence of such documentation does not mean that we can conclude the opposite: that PKV is not causing gastrointestinal disease. That said, the sparse research available did indicate that PKV has a limited ability to cause diarrhea. Since porcine kobuvirus (PKV) was first described in 2008, researchers have speculated whether the virus is of clinical importance. This systematic literature review answers the question: Is porcine kobuvirus a cause of gastrointestinal disease in young pigs? A case-control study showed that PKV was not associated with neonatal diarrhea. A cohort study suffered from a very small sample size (n = 5), and in an experimental trial, the effect of PKV inoculation could not be separated from the effect of being inoculated with porcine epidemic diarrhea virus. In 13 poorly defined observational studies, more than 4000 young pigs had been assigned a diarrhea status and their feces analyzed for PKV. Unfortunately, the studies lacked well-characterized unbiased samples, and thus the strongest possible inference from these studies was that a very strong association between PKV and diarrhea is unlikely. PKV was commonly detected in non-diarrheic pigs, and this could indicate that PKV is not a sufficient cause in itself or that reinfection of individuals with some immunological protection due to previous infections is common. Conclusively, there is a lack of good evidence of PKV being a cause of gastrointestinal disease, but the sparse available evidence suggests that PKV is of limited clinical importance. [ABSTRACT FROM AUTHOR]

Enteric diseases are considered an important factor aff ecting the livestock industry; however, there are limited data on viruses that cause enteric diseases in small ruminants in our country. In this study, we focused on enterovirus (EV), hunnivirus (HuV), and kobuvirus (KoV), which are members of the family of Picornaviridae. In order to provide further information on the prevalence and the molecular epidemiology of these viruses, a total of 73 fecal samples or gut contents from diarrheic sheep and goats were screened for EV, HuV, and KoV. For this purpose, RT-PCRs were performed by using the specific primers for each virus. Th e prevalence rate determined in the sampled population was 2.7% (2/73) for each virus. In detail, EVs (2/60, 3.3%) and HuVs (2/60, 3.3%) were each detected in only sheep samples while KoVs were identified in a goat sample (1/13, 7.6%) and a sheep sample (1/60, 1.6%). Th ere was no evidence of coinfection with these viruses in the tested animals. According to the results of the molecular analyzes, our EVs were clustered in caprine/ovinespecific EV-G and HuV strains retrieved in this study were grouped along with the other caprine/ovine origin sequences in Hunnivirus A2 genotype. Moreover, it was observed that the detected KoVs clustered in distinct species: Aichivirus B and Aichivirus C. In conclusion, this study, which reported the detection of EVs and HuVs from sheep as well as KoVs from sheep and goats in our country, provides valuable data on the epidemiology and molecular characteristics of these viruses.

Eastern roe deer (Capreolus pygargus) is a small ruminant and is widespread across China. This creature plays an important role in our ecological system. Although a few studies have been conducted to investigate pathogens harbored by this species, our knowledge of the virus diversity is still very sparse. In this study, we conducted the whole virome profiling of a rescue-failed roe deer, which revealed a kobuvirus (KoV), a bocaparvovirus (BoV), and multiple circular single-stranded viruses. These viruses were mainly recovered from the rectum, but PCR detection showed systematic infection of the KoV. Particularly, the KoV and BoV exhibited closely genetic relationships with bovine and canine viruses, respectively, highly suggesting the spillover of viruses from domestic animals to wildlife. Although these viruses were unlikely to have been responsible for the death of the animal, they provide additional data to understand the virus spectrum harbored by roe deer. The transmission of viruses between domestic animals and wildlife highlights the need for extensive investigation of wildlife viruses. [ABSTRACT FROM AUTHOR]

Abstract Backround Astrovirus, Norovirus and Sapovirus exhibit a wide distribution in swine pig herds worldwide. However, the association of porcine Astrovirus (PAstV), porcine Norovirus (PoNoV) and porcine Sapovirus (PoSaV) with disease in pigs remains uncertain. In this study, we investigated the prevalence of PAstV, PoNoV and PoSaV in Greek pig farms using both conventional RT-PCR and SYBR-Green Real-time RT-PCR in an effort to compare the sensitivity of the two methods. We examined 1400 stool samples of asymptomatic pigs originating from 28 swine farms throughout Greece in pools of five. Results PAstV was detected in all 28 swine farms examined, with an overall prevalence of 267/280 positive pools (95.4%). Porcine Caliciviruses prevalence was found at 36 and 57 out of the 280 examined samples, by the conventional and SYBR-Green Real time RT-PCR, respectively. Sequencing and phylogenetic analysis of the positive samples revealed that the detected PAstV sequences are clustered within PAstV1, 3 and 4 lineages, with PAstV3 being the predominant haplotype (91.2%). Interestingly, sequencing of the Calicivirus positive samples demonstrated the presence of non-target viruses, i.e. Sapovirus, Kobuvirus and Sapelovirus sequences and one sequence highly similar to bat Astrovirus, while no Norovirus sequence was detected. Conclusions The high prevalence of PAstV in Greek pig farms poses a necessity for further investigation of the pathogenicity of this virus and its inclusion in surveillance programs in case that it proves to be important. To our knowledge, this is the first epidemiological study of these viruses in pig farms in Greece.

Enteric diseases are considered an important factor affecting the livestock industry; however, there are limited data on viruses that cause enteric diseases in small ruminants in our country. In this study, we focused on enterovirus (EV), hunnivirus (HuV), and kobuvirus (KoV), which are members of the family of Picornaviridae. In order to provide further information on the prevalence and the molecular epidemiology of these viruses, a total of 73 fecal samples or gut contents from diarrheic sheep and goats were screened for EV, HuV, and KoV. For this purpose, RT-PCRs were performed by using the specific primers for each virus. The prevalence rate determined in the sampled population was 2.7% (2/73) for each virus. In detail, EVs (2/60, 3.3%) and HuVs (2/60, 3.3%) were each detected in only sheep samples while KoVs were identified in a goat sample (1/13, 7.6%) and a sheep sample (1/60, 1.6%). There was no evidence of coinfection with these viruses in the tested animals. According to the results of the molecular analyzes, our EVs were clustered in caprine/ovinespecific EV-G and HuV strains retrieved in this study were grouped along with the other caprine/ovine origin sequences in Hunnivirus A2 genotype. Moreover, it was observed that the detected KoVs clustered in distinct species: Aichivirus B and Aichivirus C. In conclusion, this study, which reported the detection of EVs and HuVs from sheep as well as KoVs from sheep and goats in our country, provides valuable data on the epidemiology and molecular characteristics of these viruses. [ABSTRACT FROM AUTHOR]

Since porcine kobuvirus (PKV) was first described in 2008, researchers have speculated whether the virus is of clinical importance. This systematic literature review answers the question: Is porcine kobuvirus a cause of gastrointestinal disease in young pigs? A case-control study showed that PKV was not associated with neonatal diarrhea. A cohort study suffered from a very small sample size (n = 5), and in an experimental trial, the effect of PKV inoculation could not be separated from the effect of being inoculated with porcine epidemic diarrhea virus. In 13 poorly defined observational studies, more than 4000 young pigs had been assigned a diarrhea status and their feces analyzed for PKV. Unfortunately, the studies lacked well-characterized unbiased samples, and thus the strongest possible inference from these studies was that a very strong association between PKV and diarrhea is unlikely. PKV was commonly detected in non-diarrheic pigs, and this could indicate that PKV is not a sufficient cause in itself or that reinfection of individuals with some immunological protection due to previous infections is common. Conclusively, there is a lack of good evidence of PKV being a cause of gastrointestinal disease, but the sparse available evidence suggests that PKV is of limited clinical importance.

Kobuviruses are an unusual and poorly characterized genus within the picornavirus family and can cause gastrointestinal enteric disease in humans, livestock, and pets. The human kobuvirus Aichi virus (AiV) can cause severe gastroenteritis and deaths in children below the age of 5 years; however, this is a very rare occurrence. During the assembly of most picornaviruses (e.g., poliovirus, rhinovirus, and foot-and-mouth disease virus), the capsid precursor protein VP0 is cleaved into VP4 and VP2. However, kobuviruses retain an uncleaved VP0. From studies with other picornaviruses, it is known that VP4 performs the essential function of pore formation in membranes, which facilitates transfer of the viral genome across the endosomal membrane and into the cytoplasm for replication. Here, we employ genome exposure and membrane interaction assays to demonstrate that pH plays a critical role in AiV uncoating and membrane interactions. We demonstrate that incubation at low pH alters the exposure of hydrophobic residues within the capsid, enhances genome exposure, and enhances permeabilization of model membranes. Furthermore, using peptides we demonstrate that the N terminus of VP0 mediates membrane pore formation in model membranes, indicating that this plays an analogous function to VP4. IMPORTANCE To initiate infection, viruses must enter a host cell and deliver their genome into the appropriate location. The picornavirus family of small nonenveloped RNA viruses includes significant human and animal pathogens and is also a model to understand the process of cell entry. Most picornavirus capsids contain the internal protein VP4, generated from cleavage of a VP0 precursor. During entry, VP4 is released from the capsid. In enteroviruses this forms a membrane pore, which facilitates genome release into the cytoplasm. Due to high levels of sequence similarity, it is expected to play the same role for other picornaviruses. Some picornaviruses, such as Aichi virus, retain an intact VP0, and it is unknown how these viruses rearrange their capsids and induce membrane permeability in the absence of VP4. Here, we have used Aichi virus as a model VP0 virus to test for conservation of function between VP0 and VP4. This could enhance understanding of pore function and lead to development of novel therapeutic agents that block entry. [ABSTRACT FROM AUTHOR]

Despite significant economic and public health implications, swine enteric viruses that do not manifest clinical symptoms are often overlooked, and data on their epidemiology and pathogenesis are still scarce. Here, an epidemiological study was carried out by using reverse transcription-polymerase chain reaction (RT-PCR) and sequence analysis in order to better understand the distribution and genetic diversity of porcine astrovirus (PAstV), porcine encephalomyocarditis virus (EMCV), porcine kobuvirus (PKV), and porcine sapovirus (PSaV) in healthy pigs reared under specific pathogen-free (SPF) or conventional farms. PKV was the most prevalent virus (51.1%, 247/483), followed by PAstV (35.4%, 171/483), then PSaV (18.4%, 89/483), and EMCV (8.7%, 42/483). Overall, at least one viral agent was detected in 300 out of 483 samples. Out of the 300 samples, 54.0% (162/300), 13.0% (39/300), or 1.0% (3/300) were found coinfected by two, three, or four viruses, respectively. To our knowledge, this is the first report of EMCV detection from porcine fecal samples in China. Phylogenetic analysis revealed genetically diverse strains of PAstV, PKV, and PSaV circulating in conventional and SPF farms. Detection of swine enteric viruses with a high coinfection rate in healthy pigs highlights the importance of continuous viral surveillance to minimize future economic and public health risks. [ABSTRACT FROM AUTHOR]

BackgroundMurine Kobuvirus (MuKV) is a novel picornavirus of the genus Kobuvirus, and was first identified in the feces of murine rodents in the USA in 2011. There is limited information on the transmission route of MuKV. Thus, we conducted a study to investigate virus detection rates in fecal, serum, throat, and lung tissue samples from murine rodents.ResultsA total of 413 fecal samples, 385 lung samples, 269 throat swab samples, and 183 serum samples were collected from 413 murine rodents (Rattus norvegicus, Rattus tanezumi, and Rattus rattus) captured in urban Shenzhen. Kobuviruses were detected via RT-PCR. Only fecal samples were positive, with prevalence rates of 34.9% in Rattus norvegicus and 29.4% in Rattus tanezumi. Phylogenetic analysis based on partial 3D and complete VP1 sequence regions indicated that all of the MuKV sequences obtained belonged to Aichivirus A, and were genetically closely related to other MuKVs reported in China, Hungary, and the USA. Twenty-eight full-length MuKV sequences were acquired. Phylogenetic analysis of two sequences randomly selected from the two species (SZ59 and SZ171) indicated that they shared very high nucleotide and amino acid identity with one another (94.0 and 99.3%, respectively), and comparison with human Kobuvirus revealed amino acid identity values of ~80%. Additionally, a sewage-derived sequence shared high similarity with the rat-derived sequences identified in this study, with respective nucleotide and amino acid identity values from 86.5 and 90.7% to 87.2 and 91.1%.ConclusionThe results of the current study provide evidence that murine Kobuvirus is transmitted via the fecal-oral route.

Backround: Astrovirus, Norovirus and Sapovirus exhibit a wide distribution in swine pig herds worldwide. However, the association of porcine Astrovirus (PAstV), porcine Norovirus (PoNoV) and porcine Sapovirus (PoSaV) with disease in pigs remains uncertain. In this study, we investigated the prevalence of PAstV, PoNoV and PoSaV in Greek pig farms using both conventional RT-PCR and SYBR-Green Real-time RT-PCR in an effort to compare the sensitivity of the two methods. We examined 1400 stool samples of asymptomatic pigs originating from 28 swine farms throughout Greece in pools of five. Results: PAstV was detected in all 28 swine farms examined, with an overall prevalence of 267/280 positive pools (95.4%). Porcine Caliciviruses prevalence was found at 36 and 57 out of the 280 examined samples, by the conventional and SYBR-Green Real time RT-PCR, respectively. Sequencing and phylogenetic analysis of the positive samples revealed that the detected PAstV sequences are clustered within PAstV1, 3 and 4 lineages, with PAstV3 being the predominant haplotype (91.2%). Interestingly, sequencing of the Calicivirus positive samples demonstrated the presence of non-target viruses, i.e. Sapovirus, Kobuvirus and Sapelovirus sequences and one sequence highly similar to bat Astrovirus, while no Norovirus sequence was detected. Conclusions: The high prevalence of PAstV in Greek pig farms poses a necessity for further investigation of the pathogenicity of this virus and its inclusion in surveillance programs in case that it proves to be important. To our knowledge, this is the first epidemiological study of these viruses in pig farms in Greece. [ABSTRACT FROM AUTHOR]

Abstract Background Murine kobuviruses (MuKV) are newly recognized picornaviruses first detected in murine rodents in the USA in 2011. Little information on MuKV epidemiology in murine rodents is available. Therefore, we conducted a survey of the prevalence and genomic characteristics of rat kobuvirus in Guangdong, China. Results Fecal samples from 223 rats (Rattus norvegicus) were collected from Guangdong and kobuviruses were detected in 12.6% (28) of samples. Phylogenetic analysis based on partial 3D and complete VP1 sequence regions showed that rat kobuvirus obtained in this study were genetically closely related to those of rat/mouse kobuvirus reported in other geographical areas. Two near full-length rat kobuvirus genomes (MM33, GZ85) were acquired and phylogenetic analysis of these revealed that they shared very high nucleotide/amino acids identity with one another (95.4%/99.4%) and a sewage-derived sequence (86.9%/93.5% and 87.5%/93.7%, respectively). Comparison with original Aichivirus A strains, such human kobuvirus, revealed amino acid identity values of approximately 80%. Conclusion Our findings indicate that rat kobuvirus have distinctive genetic characteristics from other Aichivirus A viruses. Additionally, rat kobuvirus may spread via sewage.

Eastern roe deer (Capreolus pygargus) is a small ruminant and is widespread across China. This creature plays an important role in our ecological system. Although a few studies have been conducted to investigate pathogens harbored by this species, our knowledge of the virus diversity is still very sparse. In this study, we conducted the whole virome profiling of a rescue-failed roe deer, which revealed a kobuvirus (KoV), a bocaparvovirus (BoV), and multiple circular single-stranded viruses. These viruses were mainly recovered from the rectum, but PCR detection showed systematic infection of the KoV. Particularly, the KoV and BoV exhibited closely genetic relationships with bovine and canine viruses, respectively, highly suggesting the spillover of viruses from domestic animals to wildlife. Although these viruses were unlikely to have been responsible for the death of the animal, they provide additional data to understand the virus spectrum harbored by roe deer. The transmission of viruses between domestic animals and wildlife highlights the need for extensive investigation of wildlife viruses.

Livestock products supply about 13 percent of energy and 28 percent of protein in diets consumed worldwide. Diarrhea is a leading cause of sickness and death of beef and dairy calves in their first month of life and also affecting adult cattle, resulting in large economic losses and a negative impact on animal welfare. Despite the usual multifactorial origin, viruses are generally involved, being among the most important causes of diarrhea. There are several viruses that have been confirmed as etiological agents (i.e., rotavirus and coronavirus), and some viruses that are not yet confirmed as etiological agents. This review summarizes the viruses that have been detected in the enteric tract of cattle and tries to deepen and gather knowledge about them. [ABSTRACT FROM AUTHOR]

Since the first identification in 1989 in humans, kobuviruses (KoVs) have been identified from a wide range of animal species including carnivores, rodents, birds, ungulates, rabbits, and bats. Several studies have described the identification of genetically related KoVs in the fecal virome of domestic and wild animals suggesting a mutual exchange of viruses. By screening a total of 231 fecal samples from wild and domestic ungulates, KoVs RNA was detected in wild boars (3.2%; 2/63), chamois (4.6%; 2/43), and goats (2.6%; 2/77). On phylogenetic analysis of the partial RdRp sequence, the wild boar strains clustered within the species Aichivirus C whilst the strains identified in domestic and wild ruminants grouped into the species Aichivirus B. The complete VP1 gene was obtained for chamois and goat KoVs. Interestingly, upon phylogenetic analysis the strains grouped together with a KoV of ovine origin within a distinct genetic type (B3) of the species Aichivirus B.

Abstract Background Canine Kobuvirus (CaKoV) has been detected both in healthy and diarrheic dogs and in asymptomatic wild carnivores. In this study, we conducted a survey of CaKoV at small animal hospitals in Bangkok and vicinity of Thailand during September 2016 to September 2018. Results Three hundred and seven rectal swab samples were collected from healthy dogs (n = 55) and dogs with gastroenteritis symptoms (n = 252). Of 307 swab samples tested by using one-step RT-PCR specific to 3D gene, we found CaKoV positivity at 17.59% (54/307). CaKoVs could be detected in both sick (19.44%) and healthy (9.09%) animals. In relation to age group, CaKoV could be frequently detected in younger dogs (25.45%). Our result showed no seasonal pattern of CaKoV infection in domestic dogs. In this study, we characterized CaKoVs by whole genome sequencing (n = 4) or 3D and VP1 gene sequencing (n = 8). Genetic and phylogenetic analyses showed that whole genomes of Thai CaKoVs were closely related to Chinese CaKoVs with highest 99.5% amino acid identity suggesting possible origin of CaKoVs in Thailand. Conclusions In conclusion, this study was the first to report the detection and genetic characteristics of CaKoVs in domestic dogs in Thailand. CaKoVs could be detected in both sick and healthy dogs. The virus is frequently detected in younger dogs. Thai CaKoVs were genetically closely related and grouped with Chinese CaKoVs. Our result raises the concerns to vet practitioners that diarrhea in dogs due to canine Kobuvirus infection should not be ignored.

Despite significant economic and public health implications, swine enteric viruses that do not manifest clinical symptoms are often overlooked, and data on their epidemiology and pathogenesis are still scarce. Here, an epidemiological study was carried out by using reverse transcription-polymerase chain reaction (RT-PCR) and sequence analysis in order to better understand the distribution and genetic diversity of porcine astrovirus (PAstV), porcine encephalomyocarditis virus (EMCV), porcine kobuvirus (PKV), and porcine sapovirus (PSaV) in healthy pigs reared under specific pathogen-free (SPF) or conventional farms. PKV was the most prevalent virus (51.1%, 247/483), followed by PAstV (35.4%, 171/483), then PSaV (18.4%, 89/483), and EMCV (8.7%, 42/483). Overall, at least one viral agent was detected in 300 out of 483 samples. Out of the 300 samples, 54.0% (162/300), 13.0% (39/300), or 1.0% (3/300) were found coinfected by two, three, or four viruses, respectively. To our knowledge, this is the first report of EMCV detection from porcine fecal samples in China. Phylogenetic analysis revealed genetically diverse strains of PAstV, PKV, and PSaV circulating in conventional and SPF farms. Detection of swine enteric viruses with a high coinfection rate in healthy pigs highlights the importance of continuous viral surveillance to minimize future economic and public health risks.

UnlabelledDespite wide sequence divergence, multiple picornaviruses use the Golgi adaptor acyl coenzyme A (acyl-CoA) binding domain protein 3 (ACBD3/GCP60) to recruit phosphatidylinositol 4-kinase class III beta (PI4KIIIβ/PI4KB), a factor required for viral replication. The molecular basis of this convergent interaction and the cellular function of ACBD3 are not fully understood. Using affinity purification-mass spectrometry, we identified the putative Rab33 GTPase-activating proteins TBC1D22A and TBC1D22B as ACBD3-interacting factors. Fine-scale mapping of binding determinants within ACBD3 revealed that the interaction domains for TBC1D22A/B and PI4KB are identical. Affinity purification confirmed that PI4KB and TBC1D22A/B interactions with ACBD3 are mutually exclusive, suggesting a possible regulatory mechanism for recruitment of PI4KB. The C-terminal Golgi dynamics (GOLD) domain of ACBD3 has been previously shown to bind the 3A replication protein from Aichi virus. We find that the 3A proteins from several additional picornaviruses, including hepatitis A virus, human parechovirus 1, and human klassevirus, demonstrate an interaction with ACBD3 by mammalian two-hybrid assay; however, we also find that the enterovirus and kobuvirus 3A interactions with ACBD3 are functionally distinct with respect to TBC1D22A/B and PI4KB recruitment. These data reinforce the notion that ACBD3 organizes numerous cellular functionalities and that RNA virus replication proteins likely modulate these interactions by more than one mechanism.ImportanceMultiple viruses use the same Golgi protein (ACBD3) to recruit the lipid kinase phosphatidylinositol 4-kinase class III beta (PI4KB) in order to replicate. We identify a new binding partner of ACBD3 in the evolutionarily conserved Rab GTPase-activating proteins (RabGAPs) TBC1D22A and -B. Interestingly, TBC1D22A directly competes with PI4KB for binding to the same location of ACBD3 by utilizing a similar binding domain. Different viruses are able to influence this interaction through distinct mechanisms to promote the association of PI4KB with ACBD3. This work informs our knowledge of both the physical interactions of the proteins that help maintain metazoan Golgi structure and how viruses subvert these evolutionarily conserved interactions for their own purposes.

Kobuvirus infections are common among humans, rodents, carnivores, pigs, and ruminants. We report herein the complete genome sequence of a novel caprine kobuvirus (MN604700) from diarrheic kids in Minnesota. Whole-genome sequencing revealed a kobuvirus genome of 8,139 nt with a single ORF region encoding a polyprotein of 2,480 amino acids. Further analysis revealed nt substitutions along the genome compared with that of the caprine kobuvirus reference strain, with 93% identity. Phylogenetic analysis indicated that the clade of the caprine kobuvirus was most closely related to porcine kobuviruses rather than bovine or ovine kobuviruses. Using primers designed from this genome, caprine kobuvirus was identified in the stools of other goats. Sanger sequencing of PCR products indicated 3D and VP1 gene nucleotides of this latter strain were 95% and 91% identical with those of MN604700, respectively. There were 35 and 101 nt substitutions in 3D and VP1 genes, respectively. Findings of kobuvirus over a 2-y period may indicate an endemic state, which needs further research. In addition, screening for kobuviruses over large geographic areas is needed to identify the evolutionary connections among different strains. [ABSTRACT FROM AUTHOR]

Background: Murine kobuviruses (MuKV) are newly recognized picornaviruses first detected in murine rodents in the USA in 2011. Little information on MuKV epidemiology in murine rodents is available. Therefore, we conducted a survey of the prevalence and genomic characteristics of rat kobuvirus in Guangdong, China. Results: Fecal samples from 223 rats (Rattus norvegicus) were collected from Guangdong and kobuviruses were detected in 12.6% (28) of samples. Phylogenetic analysis based on partial 3D and complete VP1 sequence regions showed that rat kobuvirus obtained in this study were genetically closely related to those of rat/mouse kobuvirus reported in other geographical areas. Two near full-length rat kobuvirus genomes (MM33, GZ85) were acquired and phylogenetic analysis of these revealed that they shared very high nucleotide/amino acids identity with one another (95.4%/99.4%) and a sewage-derived sequence (86.9%/93.5% and 87.5%/93.7%, respectively). Comparison with original Aichivirus A strains, such human kobuvirus, revealed amino acid identity values of approximately 80%. Conclusion: Our findings indicate that rat kobuvirus have distinctive genetic characteristics from other Aichivirus A viruses. Additionally, rat kobuvirus may spread via sewage. [ABSTRACT FROM AUTHOR]

Livestock plays a crucial role in ensuring food security and driving the global economy. However, viral infections can have far-reaching consequences beyond economic productivity, affecting the health of cattle, as well as posing risks to human health and other animals. Identifying viruses present in fecal samples, a primary route of pathogen transmission, is essential for developing effective prevention, control, and surveillance strategies. Viral metagenomic approaches offer a broader perspective and hold great potential for detecting previously unknown viruses or uncovering previously undescribed agents. Ubaté Province is Colombia's dairy capital and a key center for livestock production in the country. Therefore, the purpose of this study was to characterize viral communities in fecal samples from cattle in this region. A total of 42 samples were collected from three municipalities in Ubaté Province, located in central Colombia, using a convenient non-probabilistic sampling method. We utilized metagenomic sequencing with Oxford Nanopore Technologies (ONT), combined with diversity and phylogenetic analysis. The findings revealed a consistent and stable viral composition across the municipalities, primarily comprising members of the Picornaviridae family. At the species level, the most frequent viruses were Enterovirus E (EVE) and Bovine Astrovirus (BoAstV). Significantly, this study reported, for the first time in Colombia, the presence of viruses with veterinary importance occurring at notable frequencies: EVE (59%), Bovine Kobuvirus (BKV) (52%), and BoAstV (19%). Additionally, the study confirmed the existence of Circular replicase-encoding single-stranded (CRESS) Virus in animal feces. These sequences were phylogenetically grouped with samples obtained from Asia and Latin America, underscoring the importance of having adequate representation across the continent. The virome of bovine feces in Ubaté Province is characterized by the predominance of potentially pathogenic viruses such as BoAstV and EVE that have been reported with substantial frequency and quantities. Several of these viruses were identified in Colombia for the first time. This study showcases the utility of using metagenomic sequencing techniques in epidemiological surveillance. It also paves the way for further research on the influence of these agents on bovine health and their frecuency across the country., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Juan David Ramirez reports financial support was provided by University of Rosario., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Swine enteric viral infections are responsible for substantial economic losses in the pork industry worldwide. Porcine epidemic diarrhea (PEDV) is one of the main causative agents of diarrhea in lactating pigs, and reports of PEDV coinfection with other enteric viruses highlight the importance of viral interactions for disease presentation and outcomes. Using next-generation sequencing (NGS) and sequence analyses from samples taken from piglets with acute diarrhea, we explored the possible interactions between PEDV and other less reported pathogens. PEDV coinfection with porcine kobuvirus (PKV) was detected in 36.4% (27/74) of samples. Full genomes from porcine coronavirus and kobuvirus were obtained, as was a partial porcine sapovirus genome (PSaV). The phylogenetic results show the clustering of these strains corresponding to the geographical relationship. To our knowledge, this is the first full genome and isolation report for porcine kobuvirus in México, as well as the first phylogenetic analysis for porcine sapovirus in the country. The NGS approach provides a better perspective of circulating viruses and other pathogens in affected production units.

• Picornaviral RNA genome serves as messenger RNA and is directly transcribed into a single polyprotein chain. • The RNA-dependent RNA polymerase 3Dpol, arises from a stable 3CDpro precursor that has high proteolytical but no polymerase activity. • Several mechanisms of how cleavage of the 3CD precursor activates the polymerase activity were suggested. • We have solved two crystal structures of atypical picornaviral polymerases that have a serine at the first position where other picornaviruses have a glycine residue. • The structures reveal a common mechanism of 3Dpol activation for all picornaviral polymerases that lies in the stabilization of the α10 helix. RNA-dependent RNA polymerase 3Dpol is a key enzyme for the replication of picornaviruses. The viral genome is translated into a single polyprotein that is subsequently proteolytically processed into matured products. The 3Dpol enzyme arises from a stable 3CD precursor that has high proteolytic activity but no polymerase activity. Upon cleavage of the precursor the newly established N-terminus of 3Dpol is liberated and inserts itself into a pocket on the surface of the 3Dpol enzyme. The essential residue for this mechanism is the very first glycine that is conserved among almost all picornaviruses. However, kobuviruses and siciniviruses have a serine residue instead. Intrigued by this anomaly we sought to solve the crystal structure of these 3Dpol enzymes. The structures revealed a unique fold of the 3Dpol N-termini but the very first serine residues were inserted into a charged pocket in a similar manner as the glycine residue in other picornaviruses. These structures revealed a common underlying mechanism of 3Dpol activation that lies in activation of the α10 helix containing a key catalytical residue Asp238 that forms a hydrogen bond with the 2′ hydroxyl group of the incoming NTP nucleotide. [ABSTRACT FROM AUTHOR]

• Pigs can shed Kobuvirus at all life stages. • More piglets with diarrhea shed Kobuvirus in late nursing than healthy pigs do. • The nursery life stage is associated with a Kobuvirus shedding peak. • Similar Kobuvirus strains were found throughout a pig's life. Porcine kobuviruses are widely distributed in swine, but the clinical significance of these viruses remains unclear, since they have been associated with both diarrheic and healthy pigs. In addition, there is a paucity of data on Kobuvirus prevalence in Canadian pig herds. In this study, a total of 181 diarrheic and healthy piglets were monitored and sampled on four occasions, intended to represent the different stages of production. The piglets were sampled at the nursing farms (birth to weaning stage), at the nursery farms (post-weaning stage), and at finishing farms (at the beginning and the end of the fattening stage). Fecal and environmental samples were collected during each life stage. Following viral extraction, Kobuvirus detection by RT-PCR was conducted, and positive samples were sequenced. During the late-nursing stage (6–21 days old), piglets with diarrhea shed more Kobuvirus than healthy individuals. Piglets shed more Kobuvirus during the post-weaning stage (nursery farms) than during any of the other life stages. This was evidenced in individual samples as well as in environmental samples. Over 97% of the sampled piglets shed Kobuvirus at least once in their lifetime. All piglets shedding a Kobuvirus strain or mix of strains at the nursing stage did not appear to shed another porcine kobuvirus strain at a later life stage. Overall, our findings throw light on Kobuvirus shedding dynamics and their potential role in neonatal diarrhea at the nursing stage, which appears to be the point of entry for kobuviruses into swine production systems. [ABSTRACT FROM AUTHOR]

Abstract Feline kobuvirus (FeKoV), a novel picornavirus of the genus kobuvirus , was initially identified in the feces of cats with diarrhea in South Korea in 2013. To date, there is only one report of the circulation of kobuvirus in cats in southern China. To investigate the presence and genetic variability of FeKoV in northeast China, 197 fecal samples were collected from 105 cats with obvious diarrhea and 92 asymptomatic cats in Shenyang, Jinzhou, Changchun, Jilin and Harbin regions, Northeast China, and viruses were detected by RT-PCR with universal primers targeting all kobuviruses. Kobuvirus was identified in 28 fecal samples with an overall prevalence of 14.2% (28/197) of which 20 samples were co-infected with feline parvovirus (FPV) and/or feline bocavirus (FBoV). Diarrhoeic cats had a higher kobuvirus prevalence (19.1%, 20/105) than asymptomatic cats (8.7%, 8/92). By genetic analysis based on partial 3D gene, all kobuvirus-positive samples were more closely related to previous FeKoV strains with high identities of 90.5%–97.8% and 96.6%–100% at the nucleotide and amino acid levels. Additionally, phylogenetic analysis based on the complete VP1 gene indicated that all FeKoV strains identified in this study were placed into a cluster, which separated from other reference strains previously reported, and three identical amino acid substitutions were present at the C-terminal of the VP1 protein for these FeKoV strains. Furthermore, two complete FeKoV polyprotein genomes were successfully obtained from two positive samples and designated 16JZ0605 and 17CC0811, respectively. The two strains shared 92.9%–94.9% nucleotide identities and 96.8%–98.4% amino acid identities to FeKoV prototype strains. Phylogenetic analysis indicated that FeKoVs were clustered according to their geographical regions, albeit with limited sequences support. This study provides the first molecular evidence that FeKoV circulates in cats in northeast China, and these FeKoVs exhibit genetic diversity and unique evolutionary trend. Highlights • This study first provides the molecular evidence for the circulation of feline kobuvirus in cats in northeast China. • Feline kobuvirus infection is closely related to viral diarrhea in cats. • All the strains identified in this study displayed genetic diversity and unique evolutionary trend. • Feline kobuvirus strains were clustered according to their geographical regions. [ABSTRACT FROM AUTHOR]

Aichi virus 1 (AiV-1) has a worldwide distribution and is associated with gastroenteritis in humans. In this study, raw sewage and mussel samples were analyzed for the presence of AiV-1 using reverse transcription-PCR (RT-PCR). Amplification and sequencing of the 3CD and VP1 genomic regions followed by phylogenetic analysis using selected genome sequences revealed the presence of AiV-1, genotype B. The results highlight the importance of further screening to evaluate the prevalence and epidemiology of this clinically important virus in South Africa. [ABSTRACT FROM AUTHOR]

Aichi virus 1 (AiV-1), belonging to the genus Kobuvirus in the family Picornaviridae, has been proposed as a causative agent of human gastroenteritis potentially transmitted by fecal-oral routes through contaminated food or water. AiV-1 is globally distributed and has been detected in various types of environmental samples, such as sewage, river water, groundwater, and shellfish. Recent environmental studies revealed that this virus could be detected in higher frequency and greater abundance than other human enteric viruses. These findings suggest that AiV-1 could potentially be an appropriate indicator of viral contamination in the environment because of its high prevalence in water environments as well as structural and genetic similarity with some of the other important enteric viruses. Further studies on the occurrence and fate of AiV-1 in environments, even in combination with clinical studies of many regions, are needed for a better understanding of their epidemiology, temporal and geographical distribution, environmental stability, and potential health risks to humans.

The fecal virome has been investigated in humans and various animal species using next generation sequencing. However, limited information is available about the fecal virome of dogs with chronic enteropathy (CE). We aimed to characterize the canine fecal virome of dogs with CE and compare it with the virome of previously analyzed healthy dogs.A total of 16 adult dogs; 8 healthy dogs (data from a parallel study) and 8 dogs with CE had fecal samples assessed by viral shotgun sequencing. Fecal samples were subjected to enrichment of viral nucleic acids prior to sequencing and metagenomic analyses. Characterization of the complete genome of a canine kobuvirus was performed by Sanger sequencing. An additional 21 healthy dogs and 14 dogs with CE were further analyzed for the prevalence of canine kobuvirus.Three fecal samples from dogs with CE contained in total 3 eukaryotic viral families. In contrast, 4/8 fecal samples previously identified from healthy dogs, contained 5 eukaryotic viral families with 2 families exclusive to this group. Bacteriophages were identified in all fecal samples from CE and healthy dogs. Canine kobuvirus was identified in one dog with CE, by shotgun sequencing, and the complete genome was then characterized. This kobuvirus was classified within canine kobuvirus group, being similar to strains from Korea and China. The larger prevalence study did not detect additional samples positive for canine kobuvirus. The fecal virome of dogs with CE differs in number and type of viral families from healthy dogs. The first Australian canine kobuvirus sequence was identified and characterized from a dog with CE. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

An 11-d-old Holstein bull calf was presented to the Veterinary Medical Teaching Hospital at the University of Wisconsin–Madison because of a 4-d history of diarrhea and persistent low-grade fever. Initial diagnosis was enteritis caused by Cryptosporidium and rotavirus. During hospitalization, the calf became stuporous and was only responsive to noxious stimuli, with hypotonia of all 4 limbs, tail, head, and neck. A cerebrospinal fluid analysis revealed xanthochromia, with marked lymphocytic pleocytosis, which was suggestive of viral meningitis and/or encephalitis. Aichivirus B, which belongs to the Kobuvirus genus, was tentatively identified in spinal fluid by next-generation DNA sequencing. This virus can affect a multitude of species, including humans and cattle, and has been isolated from both healthy and diarrheic individuals. However, to date, a possible connection with neurologic disease has not been described, to our knowledge. [ABSTRACT FROM AUTHOR]

Summary Picornaviruses are small positive-sense single-stranded RNA viruses that include many important human pathogens. Within the host cell, they replicate at specific replication sites called replication organelles. To create this membrane platform, they hijack several host factors including the acyl-CoA-binding domain-containing protein-3 (ACBD3). Here, we present a structural characterization of the molecular complexes formed by the non-structural 3A proteins from two species of the Kobuvirus genus of the Picornaviridae family and the 3A-binding domain of the host ACBD3 protein. Specifically, we present a series of crystal structures as well as a molecular dynamics simulation of the 3A:ACBD3 complex at the membrane, which reveals that the viral 3A proteins act as molecular harnesses to enslave the ACBD3 protein leading to its stabilization at target membranes. Our data provide a structural rationale for understanding how these viral-host protein complexes assemble at the atomic level and identify new potential targets for antiviral therapies. [ABSTRACT FROM AUTHOR]

Detection of Aichi virus in humans was initially reported in Japan in 1989. To establish a timeline for the prevalence of Aichi virus infection among humans in the Netherlands, we conducted molecular analysis of archival water samples from 1987–2000 and 2009–2012. Aichi virus RNA was detected in 100% (8/8) of sewage samples and 100% (7/7) of surface water samples collected during 1987–2000 and 100% (8/8) of sewage samples and 71% (5/7) of surface water samples collected during 2009–2012. Several genotype A and B Aichi virus lineages were observed over the 25-year period studied, but the time course of viral genetic diversity showed recent expansion of the genotype B population over genotype A. Our results show that Aichi virus has been circulating among the human population in the Netherlands since before its initial detection in humans was reported and that genotype B now predominates in this country.