Identification and characterization of recent retrovirus in Rhinolophus ferrumequinum bats.

An investigation into retrovirus was conducted in six species of bats (Myotis aurascens, Myotis petax, Myotis macrodactylus, Miniopterus fuliginosus, Rhinolophus ferrumequinum, and Pipistrellus abramus) inhabiting South Korea. Exogenous retroviruses (XRVs) were detected in the tissue samples of R. ferrumequinum individuals by PCR assay. Proviruses were identified in all tissue samples through viral quantification using a digital PCR assay per organ (lung, intestine, heart, brain, wing, kidney, and liver), with viral loads varying greatly between each organ. In phylogenetic analysis based on the whole genome, the Korean bat retroviruses and the R. ferrumequinum retrovirus (RfRV) strain formed a new clade distinct from the Gammaretrovirus clade. The phylogenetic results determined these viruses to be RfRV-like viruses. In the Simplot comparison, Korean RfRV-like viruses exhibited relatively strong fluctuated patterns in the latter part of the envelope gene area compared to other gene areas. Several point mutations within this region (6,878-7,774 bp) of these viruses were observed compared to the RfRV sequence. One Korean RfRV-like virus (named Y4b strain) was successfully recovered in the Raw 264.7 cell line, and virus particles replicated in the cells were confirmed by transmission electron microscopy. RfRVs (or RfRV-like viruses) have been spreading since their first discovery in 2012, and the Korean RfRV-like viruses were assumed to be XRVs that evolved from RfRV.IMPORTANCER. ferrumequinum retrovirus (RfRV)-like viruses were identified in greater horseshoe bats in South Korea. These RfRV-like viruses were considered exogenous retroviruses (XRVs) that emerged from RfRV. Varying amounts of provirus detected in different organs suggest ongoing viral activity, replication, and de novo integration in certain organs. Additionally, the successful recovery of the virus in the Raw 264.7 cell line provides strong evidence supporting their status as XRVs. These viruses have now been identified in South Korea and, more recently, in Kenya since RfRV was discovered in China in 2012, indicating that RfRVs (or RfRV-like viruses) have spread worldwide.

Genetic characterization and pathogenicity in a mouse model of newly isolated bat-originated mammalian orthoreovirus in South Korea.

Mammalian orthoreoviruses (MRVs) infect a wide range of hosts, including humans, livestock, and wildlife. In the present study, we isolated a novel Mammalian orthoreovirus from the intestine of a microbat (Myotis aurascens) and investigated its biological and pathological characteristics. Phylogenetic analysis indicated that the new isolate was serotype 2, sharing the segments with those from different hosts. Our results showed that it can infect a wide range of cell lines from different mammalian species, including human, swine, and non-human primate cell lines. Additionally, media containing trypsin, yeast extract, and tryptose phosphate broth promoted virus propagation in primate cell lines and most human cell lines, but not in A549 and porcine cell lines. Mice infected with this strain via the intranasal route, but not via the oral route, exhibited weight loss and respiratory distress. The virus is distributed in a broad range of organs and causes lung damage. In vitro and in vivo experiments also suggested that the new virus could be a neurotropic infectious strain that can infect a neuroblastoma cell line and replicate in the brains of infected mice. Additionally, it caused a delayed immune response, as indicated by the high expression levels of cytokines and chemokines only at 14 days post-infection (dpi). These data provide an important understanding of the genetics and pathogenicity of mammalian orthoreoviruses in bats at risk of spillover infections.IMPORTANCEMammalian orthoreoviruses (MRVs) have a broad range of hosts and can cause serious respiratory and gastroenteritis diseases in humans and livestock. Some strains infect the central nervous system, causing severe encephalitis. In this study, we identified BatMRV2/SNU1/Korea/2021, a reassortment of MRV serotype 2, isolated from bats with broad tissue tropism, including the neurological system. In addition, it has been shown to cause respiratory syndrome in mouse models. The given data will provide more evidence of the risk of mammalian orthoreovirus transmission from wildlife to various animal species and the sources of spillover infections.

PCR-Based Detection and Genetic Characterization of Parainfluenza Virus 5 Detected in Pigs in Korea from 2016 to 2018.

This study applied a molecular-based method to detect parainfluenza virus 5 (PIV5) collected from 2016 to 2018 in nine provinces of Republic of Korea. We demonstrated that PIV5 was detectable in both serum and pooled organs at an average positive rate of 1.78% (99/5566). Among these, the complete genome sequence of 15,246 nucleotides was obtained for 12 field strains. Three out of the 12 strains had the lowest genetic identity (96.20-96.68%) among the 21 porcine PIV5 genomes collected in Germany, China, India, and Republic of Korea from 1998 to 2017. By analyzing a large collection of complete genome sequences of the structural protein-coding F and HN genes, this study proposed a classification of PIV5 into two lineages, 1 and 2, and identified that group 2.2.2 within sub-lineage 2.2 was substantially divergent. The evolution of two structural protein-coding genes was largely under purifying selection. A few codons (6/9 for the F gene, 7/8 for the HN gene) had elevated dN/dS values, which were loaded on internal branches and were predicted to be related to beneficial trait(s) of the virus.

Molecular-based detection, genetic characterization and phylogenetic analysis of porcine circovirus 4 from Korean domestic swine farms.

Porcine circovirus 4 (PCV4), a novel and unclassified member of the genus Circovirus, was first reported in China in 2019. Aiming to provide more evidence about the active circulation of PCV4, this study screened 335 pooled internal organs and detected the virus (i) at a rate of 3.28%, (ii) from both clinically healthy and clinically sick pigs of various age groups, and (iii) in six out of nine provinces of Korea. The complete genomic sequence of the Korean PCV4 strain (E115) was 1,770 nucleotides in length and had 98.5%-98.9% identity to three PCV4 strains currently available at GenBank. Utilizing a set of bioinformatic programs, it was revealed that the Korean PCV4 strain contained several genomic features of (i) a palindrome stem-loop structure with a conserved nonanucleotide, (ii) packed overlapping ORFs oriented in different directions and (iii) two intergenic regions in between genes encoding the putative replication-associated protein (Rep) and capsid (Cap) proteins. This study also predicted the presence of essential elements for the replication of circoviruses in all PCV4 strains, for example the origin of DNA replication, endonuclease and helicase domains of Rep, and the nuclear localization signal on the putative Cap protein. Finally, based on the phylogeny inferred from sequences of the putative Rep protein, this study further clarified the genetic relationships between PCV4 and other CRESS DNA viruses in general and circoviruses in particular.

Serological and Molecular Characterization of Avian Metapneumovirus in Chickens in Northern Vietnam.

Avian Metapneumovirus (aMPV) is a causative agent of respiratory disease complex in turkeys and chickens that has recently been detected in Vietnam. Due to its novelty, this study was conducted to elucidate the distribution of aMPV in several provinces in northern Vietnam. By the application of Enzyme-Linked Immunosorbent Assay (ELISA) and nested Reverse Transcription-Polymerase Chain Reaction (RT-PCR), this study demonstrated the circulation of aMPV in 12 out of 14 cities/provinces with positive rates of 37.6% and 17.2%, respectively. All nested RT-PCR positive samples were aMPV subgroup B. By pairing the detection results with age groups, it was observed that aMPV infections occurred in chickens of all ages. Additionally, by genetic characterization, aMPV strains were demonstrated to not be attenuated vaccine viruses and to belong to at least two genetic clades. Overall, the obtained results provided insights into the prevalence of aMPV and indicated a greater complexity of respiratory diseases in chickens in Vietnam.

Genomic Characterization of a Novel Alphacoronavirus Isolated from Bats, Korea, 2020.

Coronavirus, an important zoonotic disease, raises concerns of future pandemics. The bat is considered a source of noticeable viruses resulting in human and livestock infections, especially the coronavirus. Therefore, surveillance and genetic analysis of coronaviruses in bats are essential in order to prevent the risk of future diseases. In this study, the genome of HCQD-2020, a novel alphacoronavirus detected in a bat (Eptesicus serotinus), was assembled and described using next-generation sequencing and bioinformatics analysis. The comparison of the whole-genome sequence and the conserved amino acid sequence of replicated proteins revealed that the new strain was distantly related with other known species in the Alphacoronavirus genus. Phylogenetic construction indicated that this strain formed a separated branch with other species, suggesting a new species of Alphacoronavirus. Additionally, in silico prediction also revealed the risk of cross-species infection of this strain, especially in the order Artiodactyla. In summary, this study provided the genetic characteristics of a possible new species belonging to Alphacoronavirus.

Application of nano-graphene oxide as nontoxic disinfectant against alpha and betacoronaviruses.

New viruses are continuously emerging and recently there have been many great concerns on severe acute respiratory syndrome coronavirus (SARS-CoV-2). Nanographene oxide (nanoGO) has received much attention and is widely investigated to be utilised in therapy for infectious diseases by viruses. Thus, antiviral activity of nanoGO was evaluated using the porcine epidemic diarrhoea virus (PEDV), bovine coronavirus (BCoV), and SARS-CoV-2, which are all Alpha- and Beta-coronavirus. In a virus inhibition assay, the three viruses were inhibited by nanoGO in a dose-dependent manner, including attempts in the presence of high serum solution which partially mimicked biological fluid.

Torque teno virus from Korean domestic swine farms, 2017-2018.

BACKGROUND: Torque teno viruses (TTVs) have been detected worldwide, from a wide range of animals. Up to date, few studies focused on the prevalence of TTVs in general and swine torque teno viruses (TTSuVs) in particular in Korean swine farms. OBJECTIVE: This study aimed to investigate the appearance of TTSuVs and TTVs in sick pigs during the 2017-2018 period. MATERIALS AND METHODS: Molecular-based method using TTSuV1-, TTSuV2- and TTV3-specific primers was used to screen for the viruses from either sera or pooled internal organs of sick pigs. For genetic characterization, genomic sequences of TTVs were sequenced by a primer walking method. Several bioinformatic tools have been utilized to investigate the genomic organization and genetic relationship of TTVs. RESULTS: Two years of prevalence survey reveal that the prevalence of TTSuV2 is about twice that of TTSuV1. Furthermore, we identified TTV of genogroup 3 in swine pooled organ samples. The genome of two strains, M265_Korea_2017 and N119_Korea_2018, are 3,817 bp in size; M265_2017 has three open reading frames (ORFs); and N119_2018 strain has four ORFs. The complete genome nucleotide sequencing of the two strains shows 98.4% homology, and the phylogenetic analysis of Open reading frame (ORF)1 indicates that the strains are located close to TUPB strain subgroup C of genogroup 3. CONCLUSION: Our study provided the information of TTSuVs prevalence in swine farms in Korea and highlighted the presence of TTV genogroup 3 strains in pigs.

Full-length ORF2 sequence-based genetic and phylogenetic characterization of Korean feline caliciviruses.

Feline calicivirus (FCV) is a highly infectious pathogen in cats and widely distributed worldwide with high genetic variation. Full-length open reading frame 2 of 5 from recently isolated Korean FCV isolates were sequenced and compared with those of global isolates. The results of phylogenetic analysis supported dividing global FCV isolates into two genogroups (type I and II) and demonstrated the presence of genogroup II in Korea, indicating their geographic spread in East Asia. High sequence variations in region E of the FCV isolates emphasizes that a novel vaccine needs to be developed to induce protective immunity against various FCV strains.

Application of chitosan as a natural disinfectant against porcine epidemic diarrhoea virus.

Porcine epidemic diarrhoea virus (PEDV) is one of the major pathogens causing acute enteritis, which is characterised by vomiting and watery diarrhoea and commonly leads to high rates of mortality and morbidity in suckling piglets. Chitosan has been regarded as a promising natural disinfectant. In this study, the disinfectant effect and mammalian-cell toxicity of chitosan were evaluated against PEDV using Vero cells. A 0.01% solution of chitosan was determined to be an effective disinfectant. In addition, no evidence of toxicity was observed during the cell toxicity test; on the contrary, chitosan promoted cell proliferation. In conclusion, chitosan is a promising candidate for an effective and safe disinfectant against PEDV as well as other coronaviruses.

Characteristics of Aerococcus viridans isolated from porcine fetuses in Korean farms.

Swine abortion caused by viruses as well as bacteria has caused many economic losses in domestic farms over the years; however, bacterial abortion has not yet been studied in Korea. Several bacterial species were isolated from aborted fetuses (n = 103) for which the cause of death was not viral abortion. Among them, we focused on Aerococcus viridans, which had the highest positive rate within three provinces (Gangwon, Jeonnam and Gyeongnam). A total of 16 isolates were identified as A. viridans by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), and 13 were characterized by both antibiotic resistance and 16S rRNA gene analysis. Based on antibiotic susceptibility testing result, eight antimicrobials could not effectively eliminate the present isolation (more than 40% of isolates can resist these antibiotics), while all except two strains were susceptible to trimethoprim/sulfamethoxazole. Molecular analysis indicated genetic variation among these strains. This study is the first report detecting A. viridans from aborted fetuses in Korean domestic farms.

Genotyping of PCV3 based on reassembled viral gene sequences.

Porcine circovirus type 3 (PCV3) has been reported in many countries such as USA, China, Korea and many European countries during 2015-2018. The six PCV3 strains named IH, SJ, N5, N10, N13 and N62 were detected out of 220 samples by PCR methods while the prevalence our study was conducted in 2017 to 2018. The six detected strains were hard to genotype with reference viruses due to their diverse phylogenetic relationship. PCV3 capsid, ORF3 and replicase protein coding genes were reassembled at the nucleotide sequence level, then 16 new reassembled PCV3 sequences were generated. Based on the maximum likelihood mapping analysis of 303 PCV3 sequences a model with a combination of replicase, ORF3 and capsid protein coding genes was selected as the most appropriate target for genotyping, which provided the best support for the clade classification into three genotypes and several subtypes (genotype 1, genotype 2; subtype: a and b, genotype 3; subtype a, b, c, d, e, f, g, h). This study, the IH_Korea_2017 and N62_Korea_2018 strains belong to genogroup 3 (subtype a) the SJ_Korea_2017 strain genogroup 3 (subtype g) and the N5, N10, N13 Korea_ 2018 strains genogroup 3 (subtype f), respectively. In conclusion, this study may provide insights to classification of PCV3 genotypes around the world.

Molecular Characterization of Porcine Epidemic Diarrhea Virus and Its New Genetic Classification Based on the Nucleocapsid Gene.

Porcine epidemic diarrhea virus (PEDV) causes continuous, significant damage to the swine industry worldwide. By RT-PCR-based methods, this study demonstrated the ongoing presence of PEDV in pigs of all ages in Korea at the average detection rate of 9.92%. By the application of Bayesian phylogenetic analysis, it was found that the nucleocapsid (N) gene of PEDV could evolve at similar rates to the spike (S) gene at the order of 10-4 substitutions/site/year. Based on branching patterns of PEDV strains, three main N gene-base genogroups (N1, N2, and N3) and two sub-genogroups (N3a, N3b) were proposed in this study. By analyzing the antigenic index, possible antigenic differences also emerged in both the spike and nucleocapsid proteins between the three genogroups. The antigenic indexes of genogroup N3 strains were significantly lower compared with those of genogroups N1 and N2 strains in the B-cell epitope of the nucleocapsid protein. Similarly, significantly lower antigenic indexes in some parts of the B-cell epitope sequences of the spike protein (COE, S1D, and 2C10) were also identified. PEDV mutants derived from genetic mutations of the S and N genes may cause severe damage to swine farms by evading established host immunities.

New genotype classification and molecular characterization of canine and feline parvoviruses.

BACKGROUND: Canine parvovirus (CPV) and feline panleukopenia (FPV) cause severe intestinal disease and leukopenia. OBJECTIVES: In Korea, there have been a few studies on Korean FPV and CPV-2 strains. We attempted to investigate several genetic properties of FPV and CPV-2. METHODS: Several FPV and CPV sequences from around world were analyzed by Bayesian phylo-geographical analysis. RESULTS: The parvoviruses strains were newly classified into FPV, CPV 2-I, CPV 2-II, and CPV 2-III genotypes. In the strains isolated in this study, Gigucheon, Rara and Jun belong to the FPV, while Rachi strain belong to CPV 2-III. With respect to CPV type 2, the new genotypes are inconsistent with the previous genotype classifications (CPV-2a, -2b, and -2c). The root of CPV-I strains were inferred to be originated from a USA strain, while the CPV-II and III were derived from Italy strains that originated in the USA. Based on VP2 protein analysis, CPV 2-I included CPV-2a-like isolates only, as differentiated by the change in residue S297A/N. Almost CPV-2a isolates were classified into CPV 2-III, and a large portion of CPV-2c isolates was classified into CPV 2-II. Two residue substitutions F267Y and Y324I of the VP2 protein were characterized in the isolates of CPV 2-III only. CONCLUSIONS: We provided an updated insight on FPV and CPV-2 genotypes by molecular-based and our findings demonstrate the genetic characterization according to the new genotypes.

A Novel Synonymous Mutation of SARS-CoV-2: Is This Possible to Affect Their Antigenicity and Immunogenicity?

The S glycoprotein of coronaviruses is important for viral entry and pathogenesis with most variable sequences. Therefore, we analyzed the S gene sequences of SARS-CoV-2 to better understand the antigenicity and immunogenicity of this virus in this study. In phylogenetic analysis, two subtypes (SARS-CoV-2a and -b) were confirmed within SARS-CoV-2 strains. These two subtypes were divided by a novel synonymous mutation of D614G. This may play a crucial role in the evolution of SARS-CoV-2 to evade the host immune system. The region containing this mutation point was confirmed as a B-cell epitope located in the S1 domain, and SARS-CoV-2b strains exhibited severe reduced antigenic indexes compared to SARS-CoV-2a in this area. This may allow these two subtypes to have different antigenicity. If the two subtypes have different serological characteristics, a vaccine for both subtypes will be more effective to prevent COVID-19. Thus, further study is urgently required to confirm the antigenicity of these two subtypes.

PCR-based detection and genetic characterization of porcine parvoviruses in South Korea in 2018.

BACKGROUND: with the advantage of sequencing technology, many novel porcine parvoviruses (PPV) rather than PPV1 has been reported. This study ultilized specific PCR- based method and gene- based analysis to study the presence and genetic diversity of porcine parvoviruses in South Korea in 2018. RESULTS: The present study was conducted in 2018 and found PPV1 and PPV7 in nine out of 151 field samples (organs and semen) by the PCR method. Among these, the complete genome sequences of five strains (N2, N91, N108, N133, and N141) were recovered. Phylogenic analysis revealed that the strains N2, N91, and N108 belong to the PPV1 genotype, while N133 and N141 belong to PPV7 genotype. The PPV7 strains collected in this study had deletion mutations in the VP2 gene but differed from that of PPV7 strains collected in 2017. Among the PPV1 strains, the amino acid variations in the B cell epitopes of the VP2 protein were observed between three Korean PPV1 field strains (N2, N91, and N108) and the reference PPV1 strains. Those substitutions resulted in six out of 12 predicted epitopes having significant differences in antigenic index compared to the other PPV1 strains. CONCLUSIONS: This study confirmed the presence of different genotypes of porcine parvoviruses in South Korea. The PPVs circulating in South Korea were phylogenetically classified as PPV1 and PPV7 genotypes. Three Korean PPV1 strains collected in 2018 were predicted to have antigenic alteration in VP2 compared to several reference strains of PPV1.

Molecular-based investigation and genetic characterization of porcine stool-associated RNA virus (posavirus) lineages 1 to 3 in pigs in South Korea from 2017 to 2019.

Recent results on the detection and genetic characterization of stool-associated RNA viruses from different species have increased the knowledge about the extreme genetic diversity of picornaviruses. This study aimed to investigate the presence of unclassified porcine stool-associated RNA viruses (posaviruses) in South Korea and to elucidate the molecular evolution of the viruses. By RT-PCR, posaviruses 1 and 3 were exclusively found in fecal samples and consistently detected in three consecutive years in six of eight provinces, with 148/697 (21.2%) and 33/84 (39.3%) positive samples and farms, respectively. Every age group but the older age groups (finisher, sow) had significantly higher positive rates of posavirus 1 than posavirus 3. An analysis of the RNA-dependent RNA polymerase sequences by likelihood mapping and maximum-likelihood-based phylogenetic analysis revealed that stool-associated RNA viruses formed four supergroups that were well separated from all recognized families of the order Picornavirales. Five genomes of Korean posaviruses generated in this study were phylogenetically grouped with posavirus 1 and posavirus 3 and were predicted to have the typical genome organization of picornaviruses.

Regulation of porcine endogenous retrovirus by dual LTR1+2 (Long Terminal Region) miRNA in primary porcine kidney cells.

Porcine endogenous retroviruses (PERVs) integrate into germline DNA as proviral genome that enables vertical transmission from parents to their offspring. The provirus usually survives as part of the host genome rather than as an infectious agent, but may become pathogenic if it crosses species barriers. Therefore, replication-competent PERV should be controlled through selective breeding or knockout technologies. Two microRNAs (miRNAs), dual LTR1 and LTR2, were selected to inhibit the expression of PERV in primary porcine kidney cells. The inhibition efficiency of the miRNAs was compared based on their inhibition of different PERV regions, specifically long terminal repeats (LTRs), gag, pol, and env. Gene expression was quantified using real-time polymerase chain reaction and the C-type reverse transcriptase (RT) activity was determined. The messenger RNA (mRNA) expression of the PERV LTR and env regions was determined in HeLa cells co-cultured with primary porcine kidney cells. The mRNA expression of the LTR, gag, pol, and env regions of PERV was dramatically inhibited by dual miRNA from 24 to 144 h after transfection, with the highest inhibition observed for the LTR and pol regions at 120 h. Additionally, the RT activity of PERV in the co-culture experiment of porcine and human cells was reduced by 84.4% at the sixth passage. The dual LTR 1+2 miRNA efficiently silences PERV in primary porcine kidney cells.

Molecular characterization of a Korean porcine epidemic diarrhea virus strain NB1.

In Korea, for the past 30 years (1987-present), porcine epidemic diarrhea (PED) has been established as an endemic situation in which multiple genogroups of classical G1 and G2b, and the recently introduced pandemic G2a, coexisted. Because of the dynamic nature of the virus, continuous field monitoring for PEDV strains is required. This study is the first to reveal prevalence of PEDV in 9 sampling provinces, with an overall detection rate of 6.70%. Porcine endemic diarrhea virus (PEDV) was present in pigs of all ages, especially in the non-PED vaccinated groups. The highest detection rate was in the finisher group (2.34%), followed by that in the newborn group (1.56%). Secondly, using Sanger sequencing, this study recovered a complete genome (28 005 nucleotides long) of NB1 strain from a farm severely affected by PED. Analyses of nucleotide and deduced amino acid sequences showed that NB1 differed from 18 other Korean PEDV mostly in 4 protein coding genes: ORF1a, ORF1b, S, and N. Two amino acid substitutions (V635E and Y681Q) in the COE and S1D neutralizing epitopes of NB1 resulted in antigenic index alteration of the adjacent sites, one of which contributed to a mutation that escaped neutralizing antibodies.

En Corée, pour les 30 dernières années (1987 à ce jour), la diarrhée épidémique porcine (DEP) s'est établie comme une situation endémique dans laquelle de multiples génogroupes des classiques G1 et G2b, ainsi que le G2a pandémique récemment introduit, ont coexisté. Étant donné la nature dynamique du virus, un suivi continu sur le terrain des souches de DEP est requis. La présente étude est la première à révéler la prévalence de DEP dans neuf provinces échantillonnées, avec un taux de détection global de 6,70 %. Le virus de la DEP (VDEP) était présent chez les porcs de tout âge, spécialement dans les groupes d'animaux non-vaccinés contre la DEP. Les animaux dans le groupe en finition avaient taux de détection le plus élevé (2,34 %), suivi par ceux du groupe des nouveau-nés (1,56 %). Deuxièmement, en utilisant le séquençage de Sanger, nous avons récupéré un génome complet (28 005 nucléotides de long) de la souche NB1 sur une ferme sévèrement affectée par la DEP. L'analyse des nucléotides et des séquences d'acides aminés déduites a montré que NB1 différaient de 18 autres VDEP coréens principalement dans quatre gènes codant pour protéines: ORF1a, ORF1b, S, et N. Deux substitutions d'acides aminés (V635E et Y681Q) dans les épitopes neutralisants COE et S1D de NB1 ont résulté en une altération de l'index antigénique des sites adjacents, dont l'un contribuait à une mutation qui échappait aux anticorps neutralisants.(Traduit par Docteur Serge Messier).

Perspectives on the Evolution of Porcine Parvovirus.

Porcine parvovirus (PPV) is one of the main causes of porcine reproductive failure. It is important for swine industries to understand the recent trends in PPV evolution. Previous data show that PPV has two genetic lineages originating in Germany. In this study, two more genetic lineages were defined, one of which was distinctly Asian. Additionally, amino acid substitutions in European strains and Asian strains showed distinct differences in several regions of the VP2 gene. The VP1 gene of the recent PPV isolate (T142_South Korea) was identical to that of Kresse strain isolated in the USA in 1985, indicating that modern PPV strains now resemble the original strains (Kresse and NADL-2). In this study, we compared strains isolated in the 20th century to recent isolates and confirmed the trend that modern strains are becoming more similar to previous strains.