Detection and Genetic Characterization of Astroviruses in Brain Tissues of Wild Raccoon Dogs.

Astroviruses (AstVs) have been detected in a wide range of animal species, including mammals and birds. Recently, a novel AstV associated with neurological symptoms has been detected in the brains of some mammals. Raccoon dog AstV has been reported recently in China. However, there have been no reports in South Korea. Therefore, the present study aimed to detect and genetically characterize AstVs in the intestine and brain tissues of 133 wild raccoon dogs collected in Korea between 2017 and 2019. Of the seven raccoon dogs, AstVs were detected in six intestinal tissues and four brain tissues. Analysis of the capsid protein amino acid sequences of raccoon dog AstVs detected in Korea revealed a high similarity to canine AstVs, suggesting possible interspecies transmission between raccoon dogs and dogs. Phylogenetic and capsid protein amino acid sequence analysis of raccoon dog AstVs detected in the brain the 17-148B strain belonging to the HMO clade and exhibiting conserved sequences found in neurotropic AstVs (NT-AstVs), indicating their potential as NT-AstVs. However, the pathogenicity and transmission routes of the raccoon dog AstV detected in Korea have not yet been elucidated, so further research and continued surveillance for AstV in wild raccoon dogs are needed.

Genetic Characteristics of Canine Adenovirus Type 2 Detected in Wild Raccoon Dogs (Nyctereutes procyonoides) in Korea (2017-2020).

Adenovirus has been detected in a wide range of hosts like dogs, foxes, horses, bats, avian animals, and raccoon dogs. Canine adenoviruses with two serotypes host mammals and are members of the mastadenovirus family. Canine adenovirus type 1 (CAdV-1) and canine adenovirus type 2 (CAdV-2) cause infectious canine hepatitis and infectious bronchial disease, respectively. In this study, we investigated the prevalence of CAdV-1 and 2 in wild Nyctereutes procyonoides in Korea in 2017-2020 from 414 tissue samples, including the liver, kidney, lung, and intestine, collected from 105 raccoon dog carcasses. Only CAdV-2 was detected in two raccoon dogs, whereas CAdV-1 was not detected. Tissue samples from raccoon dogs were screened for CAdV-1 and CAdV-2 using conventional PCR. Adenovirus was successfully isolated from PCR positive samples using the Vero cell line, and the full-length gene sequence of the isolated viruses was obtained through 5' and 3' rapid amplification of cDNA ends (RACE). The major genes of the isolated CAdV-2/18Ra54 and CAdV-2/18Ra-65 strains showed the closest relationship with that of the CAdV-2 Toronto A26/61 strain isolated from Canada in 1976. There is no large mutation between CAdV-2, which is prevalent worldwide, and CAdV-2, which is prevalent in wild animals in Korea. In addition, it is still spreading and causing infections. The Toronto A26/61 strain, which showed the most similarity to CAdV-2/18Ra-54, was likely transmitted to wild animals through vaccinated companion animals, suggesting that further research is needed on safety measures surrounding animal vaccination. This study provides information on the genetic characteristics and prevalence of canine adenovirus in domestic wild animals and provides a better understanding of canine adenovirus.

First report of structural characteristics and polymorphisms of the prion protein gene in raccoon dogs: The possibility of prion disease-resistance.

Prion diseases are fatal degenerative encephalopathies caused by misfolded prion protein (PrPSc) converted from normal prion protein (PrPC). Previous studies have reported that genetic polymorphisms of the prion protein gene (PRNP) play a critical role in susceptibility to prion diseases. In addition, prion disease-resistant animals showed unique structural features of prion protein (PrP) related to species-specific amino acids. However, investigations of genetic polymorphisms of the PRNP gene and structural characteristics of PrP have not been performed in raccoon dogs thus far. We investigated genetic polymorphisms of PRNP in 87 raccoon dogs using amplicon sequencing and analyzed the genotype, allele, haplotype frequencies, and linkage disequilibrium (LD) using Haploview version 4.2. In addition, we performed phylogenetic analysis and multiple sequence alignment (MSA) using MEGA X version 10.1.8 and Clustal X version 2.1, respectively. We estimated the impact of raccoon dog and Canidae family-specific amino acids using PolyPhen-2, PROVEAN, and AMYCO. Furthermore, we analyzed the effect of raccoon dog and Canidae family-specific amino acids using the AlphaFold2 and Swiss-PdbViewer programs. We found 4 novel single nucleotide polymorphisms (SNPs) of the raccoon dog PRNP gene. In addition, the raccoon dog PrP showed 99.61% identity and the closest genetic distance to dog PrP. Among the substitutions of Canidae-specific amino acids with interspecific amino acids, D163N showed increased amyloidogenic propensity, and R181H showed alterations of hydrogen bonds. Furthermore, electrostatic potentials were changed according to the substitutions of D163N and R181H. By comparing PrP between raccoon dogs and raccoons, R168K and K224R were found to be related to changes in hydrogen bonds, and K224R altered the electrostatic potential of raccoon dog PrP. In the present study, we first reported 4 novel synonymous SNPs of the raccoon dog PRNP gene. We also identified that the PrP of raccoon dog has high homology (99.61%) with PrP of dog, which is a prion-resistant animal. In addition, raccoon dog PrP-specific amino acids are related to low amyloid propensity and inherent characteristics of 3D structure of raccoon dog PrP compared to the PrP of prion-susceptible species.

Evidence of Exposure to SARS-CoV-2 in Dogs and Cats from Households and Animal Shelters in Korea.

The COVID-19 pandemic was caused by the zoonotic SARS-CoV-2. A variety of animals involved in human life worldwide have been investigated for infection. As the degree of infection increased, extensive monitoring in animals became necessary to determine the degree of infection in animals. The study was conducted on a sample of dogs and cats, which were randomly sampled according to the number of confirmed cases in the region. Animals from both COVID-19-confirmed households and generally disease-negative families and animal shelters were included. Tests included real-time qPCR tests for SARS-CoV-2 antigens, ELISA for antibodies, and plaque reduction neutralization tests (PRNT) for neutralizing antibodies. As a result, SARS-CoV-2 viral RNA was detected in 2 cats out of 1018 pets (672 dogs and 346 cats). A total of 16 dogs (2.38%) and 18 cats (5.20%) tested positive using ELISA, and 14 dogs (2.08%) and 17 cats (4.91%) tested positive using PRNT. Antigens of- and/or antibodies to SARS-CoV-2 were detected in the animals regardless of whether the companion family was infected; this was the case even in animal shelters, which have been regarded as relatively safe from transmission. In conclusion, continuous viral circulation between humans and animals is inevitable; therefore, continuous monitoring in animals is required.

First Isolation of Pseudogymnoascus destructans, the Fungal Causative Agent of White-Nose Syndrome, in Korean Bats (Myotis petax)

White-nose syndrome (WNS), caused by Pseudogymnoascus destructans (Pd), is a lethal fungal disease that affects hibernating bats in North America. Recently, the presence of Pd was reported in countries neighboring Korea. However, Pd has not been investigated in Korea. Therefore, this study aimed to identify the presence of Pd in Korean bats. Altogether, wings from 241 bats were collected from 13 cities and cultured. A total of 79 fungal colonies were isolated, and two isolates were identified as Pd using polymerase chain reaction. Of the nine bat species captured in 13 cities, Pd was isolated only from Myotis petax in Goryeong. Atypical, curved conidia were observed in two isolated fungal colonies. Although histological lesions were not observed by hematoxylin and eosin or periodic acid−Schiff staining, fungal invasion was observed in the tissue sections. Taken together, these results confirmed the presence of Pd in Korean bats and suggest the possibility of WNS outbreaks in Korean bats. This is the first report of the isolation and molecular analysis of Pd from Korean bats.

Effect of feeding raw potato starch on the composition dynamics of the piglet intestinal microbiome.

OBJECTIVE: Raw potato starch (RPS) is resistant to digestion, escapes absorption, and is metabolized by intestinal microflora in the large intestine and acts as their energy source. In this study, we compared the effect of different concentrations of RPS on the intestinal bacterial community of weaned piglets. METHODS: Male weaned piglets (25-days-old, 7.03±0.49 kg) were either fed a corn/soybean-based control diet (CON, n = 6) or two treatment diets supplemented with 5% RPS (RPS5, n = 4) or 10% RPS (RPS10, n = 4) for 20 days and their fecal samples were collected. The day 0 and 20 samples were analyzed using a 16S rRNA gene sequencing technology, followed by total genomic DNA extraction, library construction, and high-throughput sequencing. After statistical analysis, five phyla and 45 genera accounting for over 0.5% of the reads in any of the three groups were further analyzed. Furthermore, short-chain fatty acids (SCFAs) in the day 20 fecal samples were analyzed using gas chromatography. RESULTS: Significant changes were not observed in the bacterial composition at the phylum level even after 20 d post feeding (dpf); however, the abundance of Intestinimonas and Barnesiella decreased in both RPS treatment groups compared to the CON group. Consumption of 5% RPS increased the abundance of Roseburia (p<0.05) and decreased the abundance of Clostridium (p<0.01) and Mediterraneibacter (p< 0.05). In contrast, consumption of 10% RPS increased the abundance of Olsenella (p<0.05) and decreased the abundance of Campylobacter (p<0.05), Kineothrix (p<0.05), Paraprevotella (p<0.05), and Vallitalea (p<0.05). Additionally, acetate (p<0.01), butyrate (p<0.05), valerate (p = 0.01), and total SCFAs (p = 0.01) were upregulated in the RPS5 treatment group. CONCLUSION: Feeding 5% RPS altered bacterial community composition and promoted gut health in weaned piglets. Thus, resistant starch as a feed additive may prevent diarrhea in piglets during weaning.

Experimental infection of dogs with severe fever with thrombocytopenia syndrome virus: Pathogenicity and potential for intraspecies transmission.

Severe fever with thrombocytopenia syndrome (SFTS) is caused by infection with Dabie bandavirus [formerly SFTS virus (SFTSV)] and is an emerging zoonotic disease. Dogs can be infected with SFTSV, but its pathogenicity and transmissibility have not been fully elucidated. In experiment 1, immunocompetent dogs were intramuscularly inoculated with SFTSV. In experiment 2, immunosuppressed dogs (immunosuppressed group; oral azathioprine 5 mg/kg/day for 30 days) were intramuscularly inoculated with SFTSV. Both immunosuppressed and immunocompetent contact dogs were co-housed with the SFTSV-inoculated dogs that had been immunosuppressed. Immunocompetent SFTSV-infected dogs did not show any clinical symptom. However, immunosuppressed SFTSV-infected dogs showed high fever and weight loss without lethality. In all SFTSV-infected dogs, viral RNA could be measured in the serum only after 3 days post infection (DPI) and neutralizing antibodies were detected in the serum beginning 9 DPI. SFTSV shedding in the urine and faeces of some infected dogs occurred between 4 and 6 DPI. The immunocompromised SFTSV-infected dogs showed thrombocytopenia beginning 3 DPI to the end of the experiment (24 DPI). We confirmed SFTSV transmission to one of three immunocompetent co-housed dogs. This dog showed a high fever, weight loss, and shed viral RNA by urine. Viral RNA and neutralizing antibodies were also detected in the serum. These results demonstrated that intramuscular inoculation with SFTSV induced minor clinical symptoms in dogs, and intraspecies SFTSV transmission in dogs can occur by contact.

Genetic diversity of bat coronaviruses and comparative genetic analysis of MERS-related coronaviruses in South Korea.

Bats have been identified as a natural reservoir of several potentially zoonotic viruses, including Lyssavirus, Ebola virus, Marburg virus, Hendra virus, Nipah virus, as well as severe acute respiratory syndrome and Middle East respiratory syndrome coronavirus (CoV). Here, we performed a molecular epidemiological investigation of South Korean bat viruses. Genetic comparative analysis was performed on the spike glycoprotein gene of the detected MERS-related CoVs. Among 1640 samples (348 oral swabs, 1199 faecal samples, 83 urine samples and 10 bat carcass) collected across 24 South Korean provinces during 2017-2019, CoV was detected in 82 samples (75 faeces and seven oral swab samples) from 11 provinces. Surveillance over the 3 years during which samples were collected revealed significantly higher CoV detection rates between spring and autumn, and a high detection rate in Vespertillionidae and Rhinolophidae bats. Our phylogenetic analysis shows that Korean bat CoVs are genetically diverse regardless of their spatiotemporal distribution and their host species, and that the discovered bat CoVs belong to various subgenera within the Alpha- and Betacoronavirus genera. Twenty detected MERS-related CoVs belonging to the genus Betacoronavirus were similar to the Ia io bat CoV NL140422 and NL13845 strains. A comprehensive genetic analysis of two Korean bat MERS-related CoV spike receptor binding domain (RBDs) (176 and 267 strains) showed that the 18 critical residues that are involved in interactions with the human DPP4 receptor are most similar to the NL13845 strain, which is known to not bind with hDPP4. A deeper analysis of the interfacing residues in the Korean bat MERS-related CoVs RBD-hDPP4 complexes showed that the Korean bat CoVs has fewer polar contacts than the NL13845 strain. Although further study will be needed, these results suggest that Korean bat MERS-related CoVs are unlikely to bind with hDPP4. Nevertheless, these findings highlight the need for continuous monitoring to identifying the origin of new infectious diseases, specifically mutant CoV.

Bacterial Profiles of Brain in Downer Cattle with Unknown Etiology.

Downer cow can be caused by muscular paralysis, neurological damage, metabolic disorder, and/or the complication of microbial infection. However, downer cow with unknown etiology is issued because of the non-detection of its bacterial etiological agent. In this study, differences in the bacterial community in brain tissues between downer cattle with unknown etiology and healthy slaughtered cattle are investigated. Bacterial diversity and representative genera between downer and normal cattle were significantly different (p < 0.05). There are significant differences in representative genera of downer and normal cattle, especially the significance, fold change, and area under the receiver operating characteristic curve score (p < 0.05). Furthermore, the prediction of functional genes in brain microbiota between the downer and normal cattle revealed differences in the cluster of orthologous gene categories, such as lipid transport and metabolism, secondary metabolite biosynthesis, and signal transduction (p < 0.05). This study revealed a significant difference in microbiota between the downer and normal cattle. Thus, we demonstrate that representative genera from downer cattle through 16S rRNA gene amplicon sequencing and microbiota analysis have the potential as candidates for bacterial etiological agents for downer cow.

Astrovirus Infection in Cattle with Nonsuppurative Meningoencephalitis in South Korea.

Neurological diseases in cattle can be caused by several infectious agents. Astroviruses are increasingly recognized as the causative agent of encephalitis in various animals, including humans. In this study, a neuroinvasive astrovirus (BoAstV 20B05) was discovered in the brain tissues of an 81-month-old Korean native cattle with neurological symptoms. Lymphocyte infiltration and multifocal perivascular cuffing were observed in the cerebrum and brain stem, and viral antigens were also detected in the meninges. In particular, the concentration of the astroviral genome was high in the brain tissues. Korean BoAstV 20B05 was classified into the CH13/NeuroS1 clade and was closely related to the Neuro-Uy and KagoshimaSR28-462 strains. Our evolutionary analysis showed that Korean BoAstV 20B05 belongs to the sub-lineage NeuroS1 and evolved independently of BoAstV KagoshimaSR28-462. These results suggest that neuroinvasive astroviruses were first introduced in Korea. However, analysis is limited by the lack of reference astrovirus sequences reported in various countries within Asia, and further analysis should be performed using more strains. In this study, we identified a neuroinvasive astrovirus infection with neurological symptoms for the first time in South Korea and confirmed that BoAstV 20B05 may have been introduced in South Korea a long time ago.

Genetic Characterization and Pathogenicity of H7N7 and H7N9 Avian Influenza Viruses Isolated from South Korea.

H7 low pathogenic avian influenza viruses (LPAIVs) can mutate into highly pathogenic avian influenza viruses (HPAIVs). In addition to avian species, H7 avian influenza viruses (AIVs) also infect humans. In this study, two AIVs, H7N9 (20X-20) and H7N7 (34X-2), isolated from the feces of wild birds in South Korea in 2021, were genetically analyzed. The HA cleavage site of the two H7 Korean viruses was confirmed to be ELPKGR/GLF, indicating they are LPAIVs. There were no amino acid substitutions at the receptor-binding site of the HA gene of two H7 Korean viruses compared to that of A/Anhui/1/2013 (H7N9), which prefer human receptors. In the phylogenetic tree analysis, the HA gene of the two H7 Korean viruses shared the highest nucleotide similarity with the Korean H7 subtype AIVs. In addition, the HA gene of the two H7 Korean viruses showed high nucleotide similarity to that of the A/Jiangsu/1/2018(H7N4) virus, which is a human influenza virus originating from avian influenza virus. Most internal genes (PB2, PB1, PA, NP, NA, M, and NS) of the two H7 Korean viruses belonged to the Eurasian lineage, except for the M gene of 34X-2. This result suggests that active reassortment occurred among AIVs. In pathogenicity studies of mice, the two H7 Korean viruses replicated in the lungs of mice. In addition, the body weight of mice infected with 34X-2 decreased 7 days post-infection (dpi) and inflammation was observed in the peribronchiolar and perivascular regions of the lungs of mice. These results suggest that mammals can be infected with the two H7 Korean AIVs. Our data showed that even low pathogenic H7 AIVs may infect mammals, including humans, as confirmed by the A/Jiangsu/1/2018(H7N4) virus. Therefore, continuous monitoring and pathogenicity assessment of AIVs, even of LPAIVs, are required.

Relationship among bats, parasitic bat flies, and associated pathogens in Korea.

BACKGROUND: Bats are hosts for many ectoparasites and act as reservoirs for several infectious agents, some of which exhibit zoonotic potential. Here, species of bats and bat flies were identified and screened for microorganisms that could be mediated by bat flies. METHODS: Bat species were identified on the basis of their morphological characteristics. Bat flies associated with bat species were initially morphologically identified and further identified at the genus level by analyzing the cytochrome c oxidase subunit I gene. Different vector-borne pathogens and endosymbionts were screened using PCR to assess all possible relationships among bats, parasitic bat flies, and their associated organisms. RESULTS: Seventy-four bat flies were collected from 198 bats; 66 of these belonged to Nycteribiidae and eight to Streblidae families. All Streblidae bat flies were hosted by Rhinolophus ferrumequinum, known as the most common Korean bat. Among the 74 tested bat flies, PCR and nucleotide sequencing data showed that 35 (47.3%) and 20 (27.0%) carried Wolbachia and Bartonella bacteria, respectively, whereas tests for Anaplasma, Borrelia, Hepatozoon, Babesia, Theileria, and Coxiella were negative. Phylogenetic analysis revealed that Wolbachia endosymbionts belonged to two different supergroups, A and F. One sequence of Bartonella was identical to that of Bartonella isolated from Taiwanese bats. CONCLUSIONS: The vectorial role of bat flies should be checked by testing the same pathogen and bacterial organisms by collecting blood from host bats. This study is of great interest in the fields of disease ecology and public health owing to the bats' potential to transmit pathogens to humans and/or livestock.

Intergenic recombination in feline calicivirus associated with a hemorrhagic-like disease in the Republic of Korea.

Feline calicivirus (FCV) is a common cause of upper respiratory tract disease in cats. In this study, the complete genome sequence of FCV 14Q315, which was detected from a dead domestic cat with a hemorrhagic-like disease, was analyzed to identify the genetic characteristics. The FCV 14Q315 genome was 7,684 bp. Phylogenetic analyses based on the ORF1, ORF2, and ORF3 sequences indicated that FCV 14Q315 is more closely related to FCV 15D022 than to other FCV strains. ORF1 of FCV 14Q315 shared high sequence similarity with ORF1 of FCVs 15D022 and UTCVM-H1. We further evaluated genetic recombination in ORF1 of FCV 14Q315 and detected intergenic recombination between p30 and the ORF1/ORF2 junction with high significance. Particularly, the non-recombination region in ORF1 of FCV 14Q315 showed high sequence similarity with FCVs GX2019, CH-JL2, and 15D022. The recombination region in ORF1 of FCV 14Q315 showed the highest similarity with FCV UTCVM-H1, which is associated with a hemorrhagic-like disease. The results suggest that the UTCVM-H1-like FCV was introduced into the Republic of Korea and presumably recombined with Korean FCVs by occasional mixed infections. In addition, the Korean FCV strains were located in several phylogenetic clusters with marked genetic diversity in the ORF2 region. These results imply that Korean FCVs possess high genetic diversity owing to mutations and recombination. Furthermore, it is possible that certain FCVs caused cyclical infections in the Korean cat population based on a phylogenetic analysis of FCVs isolated at different time points. Keywords: calicivirus; virulent systemic feline calicivirus; recombination; hemorrhagic-like disease.

Distribution and genotypic analysis of Enterocytozoon bieneusi from wild boars in Korea.

Enterocytozoon bieneusi, an important microsporidian fungus, causes chronic diarrhea in humans and animals worldwide. Out of the 502 fecal samples from wild boars, 13 were positive for the E. bieneusi internal transcribed spacer region, with a prevalence of 2.6%. Six E. bieneusi genotypes, D, EbpC, and four novel KWB1-KWB4, were identified with zoonotic potential. Genotypes D (subgroup 1a) and EbpC (subgroup 1d) were first reported in Korean swine and Korea, respectively; KWB1-KWB4 (subgroup 1e) were most prevalent in this study. Because zoonotic genotypes have been identified, E. bieneusi transmission through wild boars must be closely monitored for proper prevention and treatment, despite their low prevalence. LAY SUMMARY: Enterocytozoon bieneusi is an important microsporidian fungus. Its sequences from wild boars were identified with zoonotic potential. Genotypes D and EbpC were first reported in Korean swine and Korea, respectively. E. bieneusi should be closely monitored to properly prevent and treat animals.

Genetic Characteristics of Avian Influenza Virus Isolated from Wild Birds in South Korea, 2019-2020.

Wild aquatic birds, a natural reservoir of avian influenza viruses (AIVs), transmit AIVs to poultry farms, causing huge economic losses. Therefore, the prevalence and genetic characteristics of AIVs isolated from wild birds in South Korea from October 2019 to March 2020 were investigated and analyzed. Fresh avian fecal samples (3256) were collected by active monitoring of 11 wild bird habitats. Twenty-eight AIVs were isolated. Seven HA and eight NA subtypes were identified. All AIV hosts were Anseriformes species. The HA cleavage site of 20 representative AIVs was encoded by non-multi-basic amino acid sequences. Phylogenetic analysis of the eight segment genes of the AIVs showed that most genes clustered within the Eurasian lineage. However, the HA gene of H10 viruses and NS gene of four viruses clustered within the American lineage, indicating intercontinental reassortment of AIVs. Representative viruses likely to infect mammals were selected and evaluated for pathogenicity in mice. JB21-58 (H5N3), JB42-93 (H9N2), and JB32-81 (H11N2) were isolated from the lungs, but JB31-69 (H11N9) was not isolated from the lungs until the end of the experiment at 14 dpi. None of infected mice showed clinical sign and histopathological change in the lung. In addition, viral antigens were not detected in lungs of all mice at 14 dpi. These data suggest that LPAIVs derived from wild birds are unlikely to be transmitted to mammals. However, because LPAIVs can reportedly infect mammals, including humans, continuous surveillance and monitoring of AIVs are necessary, despite their low pathogenicity.

Comparative genetic analyses of Korean bat coronaviruses with SARS-CoV and the newly emerged SARS-CoV-2.

BACKGROUND: Bats have been considered natural reservoirs for several pathogenic human coronaviruses (CoVs) in the last two decades. Recently, a bat CoV was detected in the Republic of Korea; its entire genome was sequenced and reported to be genetically similar to that of the severe acute respiratory syndrome CoV (SARS-CoV). OBJECTIVES: The objective of this study was to compare the genetic sequences of SARS-CoV, SARS-CoV-2, and the two Korean bat CoV strains 16BO133 and B15-21, to estimate the likelihood of an interaction between the Korean bat CoVs and the human angiotensin-converting enzyme 2 (ACE2) receptor. METHODS: The phylogenetic analysis was conducted with the maximum-likelihood (ML) method using MEGA 7 software. The Korean bat CoVs receptor binding domain (RBD) of the spike protein was analyzed by comparative homology modeling using the SWISS-MODEL server. The binding energies of the complexes were calculated using PRODIGY and MM/GBGA. RESULTS: Phylogenetic analyses of the entire RNA-dependent RNA polymerase, spike regions, and the complete genome revealed that the Korean CoVs, along with SARS-CoV and SARS-CoV-2, belong to the subgenus Sarbecovirus, within BetaCoVs. However, the two Korean CoVs were distinct from SARS-CoV-2. Specifically, the spike gene of the Korean CoVs, which is involved in host infection, differed from that of SARS-CoV-2, showing only 66.8%-67.0% nucleotide homology and presented deletions within the RBD, particularly within regions critical for cross-species transmission and that mediate interaction with ACE2. Binding free energy calculation revealed that the binding affinity of Korean bat CoV RBD to hACE2 was drastically lower than that of SARS-CoV and SARS-CoV-2. CONCLUSIONS: These results suggest that Korean bat CoVs are unlikely to bind to the human ACE2 receptor.

Genetic Diversity and Zoonotic Potential of Blastocystis in Korean Water Deer, Hydropotes inermis argyropus.

Blastocystis is a protozoan parasite commonly detected in the intestinal tract of humans and animals. It has been actively studied worldwide; however, information on Blastocystis is limited in Korea. Because there is an increasing concern about the contact between wildlife and domestic animals or humans, we assessed the infection status and zoonotic potential of Blastocystis in Korean water deer (KWD, Hydropotes inermis argyropus) using genotyping and phylogenetic analysis. A total of 125 fresh fecal samples were collected from KWD which were killed by vehicles on highways or roadsides in this study. Among the 125 samples, 51 (40.8%) were PCR positive. We performed nucleotide sequencing and phylogenetic analysis of 26 of the 51 PCR-positive samples. By analyzing Blastocystis 18S rRNA, two subtypes (ST4 and ST14) were identified in this study. Of the 26 samples analyzed, 25 were identified as ST14 and one as ST4. Infection of ST14 in humans has not been reported. Although only one ST4 sample was detected in this study, ST4 has zoonotic potential without showing ruminant specificity. Thus, continuous attention should be provided to the potential of transmission between wildlife and domestic animals and humans.

Novel viruses detected in bats in the Republic of Korea.

Bats are natural reservoirs for potential zoonotic viruses. In this study, next-generation sequencing was performed to obtain entire genome sequences of picornavirus from a picornavirus-positive bat feces sample (16BF77) and to explore novel viruses in a pooled bat sample (16BP) from samples collected in South Korea, 2016. Fourteen mammalian viral sequences were identified from 16BF77 and 29 from 16BP, and verified by RT-PCR. The most abundant virus in 16BF77 was picornavirus. Highly variable picornavirus sequences encoding 3Dpol were classified into genera Kobuvirus, Shanbavirus, and an unassigned group within the family Picornaviridae. Amino acid differences between these partial 3Dpol sequences were ≥ 65.7%. Results showed that one bat was co-infected by picornaviruses of more than two genera. Retrovirus, coronavirus, and rotavirus A sequences also were found in the BP sample. The retrovirus and coronavirus genomes were identified in nine and eight bats, respectively. Korean bat retroviruses and coronavirus demonstrated strong genetic relationships with a Chinese bat retrovirus (RfRV) and coronavirus (HKU5-1), respectively. A co-infection was identified in one bat with a retrovirus and a coronavirus. Our results indicate that Korean bats were multiply infected by several mammal viruses.

Pathogenicity of severe fever with thrombocytopenia syndrome virus in mice regulated in type I interferon signaling: Severe fever with thrombocytopenia and type I interferon.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging zoonotic disease, which causes high fever, thrombocytopenia, and death in humans and animals in East Asian countries. The pathogenicity of SFTS virus (SFTSV) remains unclear. We intraperitoneally infected three groups of mice: wild-type (WT), mice treated with blocking anti-type I interferon (IFN)-α receptor antibody (IFNAR Ab), and IFNAR knockout (IFNAR-/-) mice, with four doses of SFTSV (KH1, 5 × 105 to 5 × 102 FAID50). The WT mice survived all SFTSV infective doses. The IFNAR Ab mice died within 7 days post-infection (dpi) with all doses of SFTSV except that the mice were infected with 5 × 102 FAID50 SFTSV. The IFNAR-/- mice died after infection with all doses of SFTSV within four dpi. No SFTSV infection caused hyperthermia in any mice, whereas all the dead mice showed hypothermia and weight loss. In the WT mice, SFTSV RNA was detected in the eyes, oral swabs, urine, and feces at 5 dpi. Similar patterns were observed in the IFNAR Ab and IFNAR-/- mice after 3 dpi, but not in feces. The IFNAR Ab mice showed viral shedding until 7 dpi. The SFTSV RNA loads were higher in organs of the IFNAR-/- mice compared to the other groups. Histopathologically, coagulation necrosis and mononuclear inflammatory cell infiltration in the liver and white pulp atrophy in the spleen were seen as the main lesions in the IFN signaling lacking mice. Immunohistochemically, SFTSV antigens were mainly detected in the marginal zone of the white pulp of the spleen in all groups of mice, but more viral antigens were observed in the spleen of the IFNAR-/- mice. Collectively, the IFN signaling-deficient mice were highly susceptible to SFTSV and more viral burden could be demonstrated in various excreta and organs of the mice when IFN signaling was inhibited.

Molecular Detection and Subtyping of Blastocystis Detected in Wild Boars (Sus scrofa) in South Korea.

Blastocystis is a genus of parasitic protozoans that live in humans, mammals, and birds and which has been widely studied due to its low host specificity. Limited data are available, however, regarding its presence in wildlife, particularly in South Korea. Contact between wild boars (Sus scrofa) and livestock or humans has steadily increased as wild boars venture down from the mountains to farms and residential areas. In this study, we examined the status and subtypes (STs) of Blastocystis in wild boars in South Korea and confirmed its zoonotic potential. From March 2016 to November 2018, we collected 433 fecal samples throughout the country from trapped or road-killed wild boars. The 18S rRNA gene was used for molecular identification and subtyping and the proportion of PCR-positive samples was 10.4%. We then assessed positive samples for associations with sex, region, and seasonal infection; however, no statistical significance was observed for any variable other than season. Phylogenetic analyses revealed that all sequences belonged to subtype 5 and had 99.5-99.9% identity with sequences obtained from Japanese cattle (Bos taurus) and 97.1% identity with sequences obtained from Chinese. Subtype 5 has been implicated in zoonoses, indicating that Korean wild boars could transmit Blastocystis to humans and other livestock. Our results, in accordance with the One Health concept, strongly support continued interest and efforts by public health and disease control organizations toward transmission prevention.