Comparative analysis of Streptococcus suis genomes identifies novel candidate virulence-associated genes in North American isolates.

Streptococcus suis is a significant economic and welfare concern in the swine industry. Pan-genome analysis provides an in-silico approach for the discovery of genes involved in pathogenesis in bacterial pathogens. In this study, we performed pan-genome analysis of 208 S. suis isolates classified into the pathogenic, possibly opportunistic, and commensal pathotypes to identify novel candidate virulence-associated genes (VAGs) of S. suis. Using chi-square tests and LASSO regression models, three accessory pan-genes corresponding to S. suis strain P1/7 markers SSU_RS09525, SSU_RS09155, and SSU_RS03100 (>95% identity) were identified as having a significant association with the pathogenic pathotype. The proposed novel SSU_RS09525 + /SSU_RS09155 + /SSU_RS03100 + genotype identified 96% of the pathogenic pathotype strains, suggesting a novel genotyping scheme for predicting the pathogenicity of S. suis isolates in North America. In addition, mobile genetic elements carrying antimicrobial resistance genes (ARGs) and VAGs were identified but did not appear to play a major role in the spread of ARGs and VAGs.

Genetic diversity and evolution of the emerging picornavirus Senecavirus A.

Senecavirus A (SVA) is an emerging picornavirus that causes vesicular disease (VD) in swine. The virus has been circulating in swine in the United Stated (USA) since at least 1988, however, since 2014 a marked increase in the number of SVA outbreaks has been observed in swine worldwide. The factors that led to the emergence of SVA remain unknown. Evolutionary changes that accumulated in the SVA genome over the years may have contributed to the recent increase in disease incidence. Here we compared full-genome sequences of historical SVA strains (identified before 2010) from the USA and global contemporary SVA strains (identified after 2011). The results from the genetic analysis revealed 6.32 % genetic divergence between historical and contemporary SVA isolates. Selection pressure analysis revealed that the SVA polyprotein is undergoing selection, with four amino acid (aa) residues located in the VP1 (aa 735), 2A (aa 941), 3C (aa 1547) and 3D (aa 1850) coding regions being under positive/diversifying selection. Several aa substitutions were observed in the structural proteins (VP1, VP2 and VP3) of contemporary SVA isolates when compared to historical SVA strains. Some of these aa substitutions led to changes in the surface electrostatic potential of the structural proteins. This work provides important insights into the molecular evolution and epidemiology of SVA.

Pseudocowpox virus infection in an American bison (Bison bison).

BACKGROUND: The present report describes a case of pseudocowpox virus (PCPV) infection in a seven-year-old female bison euthanized due to a history of declining condition and sores on the vulva and udder. CASE PRESENTATION: External examination revealed multifocal, raised, keratinized plaques (0.5-2 cm) covering the skin of the ventral surface of the tail, perineum, caudoventral abdomen, udder, both inguinal recesses, and the medial aspects of both thighs. No significant gross lesions were present in the reminder of the tissues examined. Histopathological examination of the affected skin showed moderate epidermal hyperplasia with rete pegs, marked parakeratotic hyperkeratosis with crusts of degenerate neutrophils and cell debris, and few epithelial cells undergoing ballooning degeneration with occasional eosinophilic intracytoplasmic inclusion bodies (3-5 µm Bollinger body). Negative staining electron microscopy from skin revealed typical Parapoxvirus (PPV) particles, which were also confirmed by real-time PCR (Ct =18.6). Metagenomic analysis of the skin samples revealed only poxviruses. The bison parapox B2L envelope gene clustered with other parapox sequences identified from ruminants. CONCLUSIONS: This is the first report of PCPV virus infection in an American bison. Identification of novel susceptible hosts of parapox viruses sheds light on the viral evolution and highlights the importance of potential economic impact of this disease to the bison industry.

Human-Origin Influenza A(H3N2) Reassortant Viruses in Swine, Southeast Mexico.

The genetic diversity of influenza A viruses circulating in swine in Mexico complicates control efforts in animals and presents a threat to humans, as shown by influenza A(H1N1)pdm09 virus. To describe evolution of swine influenza A viruses in Mexico and evaluate strains for vaccine development, we sequenced the genomes of 59 viruses and performed antigenic cartography on strains from 5 regions. We found that genetic and antigenic diversity were particularly high in southeast Mexico because of repeated introductions of viruses from humans and swine in other regions in Mexico. We identified novel reassortant H3N2 viruses with genome segments derived from 2 different viruses that were independently introduced from humans into swine: pandemic H1N1 viruses and seasonal H3N2 viruses. The Mexico swine viruses are antigenically distinct from US swine lineages. Protection against these viruses is unlikely to be afforded by US virus vaccines and would require development of new vaccines specifically targeting these diverse strains.

Serotype and Genotype (Multilocus Sequence Type) of Streptococcus suis Isolates from the United States Serve as Predictors of Pathotype.

Streptococcus suis is a significant cause of mortality in piglets and growing pigs worldwide. The species contains pathogenic and commensal strains, with pathogenic strains causing meningitis, arthritis, endocarditis, polyserositis, and septicemia. Serotyping and multilocus sequence typing (MLST) are primary methods to differentiate strains, but the information is limited for strains found in the United States. The objective of this study was to characterize the diversity of 208 S. suis isolates collected between 2014 and 2017 across North America (mainly the United States) by serotyping and MLST and to investigate associations between subtype and pathotype classifications (pathogenic, possibly opportunistic, and commensal), based on clinical information and site of isolation. Twenty serotypes were identified, and the predominant serotypes were 1/2 and 7. Fifty-eight sequence types (STs) were identified, and the predominant ST was ST28. Associations among serotypes, STs, and pathotypes were investigated using odds ratio and clustering analyses. Evaluation of serotype and ST with pathotype identified a majority of isolates of serotypes 1, 1/2, 2, 7, 14, and 23 and ST1, ST13, ST25, ST28, ST29, ST94, ST108, ST117, ST225, ST373, ST961, and ST977 as associated with the pathogenic pathotype. Serotypes 21 and 31, ST750, and ST821 were associated with the commensal pathotype, which is composed of isolates from farms with no known history of S. suis-associated disease. Our study demonstrates the use of serotyping and MLST to differentiate pathogenic from commensal isolates and establish links between pathotype and subtype, thus increasing the knowledge about S. suis strains circulating in the United States.

Influenza C Virus in Cattle with Respiratory Disease, United States, 2016-2018.

We identified influenza C virus (ICV) in samples from US cattle with bovine respiratory disease through real-time PCR testing and sequencing. Bovine ICV isolates had high nucleotide identities (≈98%) with each other and were closely related to human ICV strains (≈95%). Further research is needed to determine bovine ICV's zoonotic potential.

Complete genome characterization of a rotavirus B (RVB) strain identified in Alpine goat kids with enteritis reveals inter-species transmission with RVB bovine strains.

Rotavirus B (RVB) has been associated with enteric disease in many animal species. An RVB strain was identified in pooled intestinal samples from Alpine caprine kids (between 2 and 3 days of age) experiencing high (>90 %) morbidity, and the complete caprine RVB genome was characterized. Histology revealed villus atrophy, the samples tested positive for RVB by real-time RT-PCR and metagenomic next-generation sequencing identified only RVB and orf virus. In the VP4 gene segment, the caprine RVB strain had a higher percentage nucleotide identity to the Indian bovine RVB strains than to the Japanese bovine RVB strains, but the VP7, VP6, VP2, NSP1, NSP2 and NSP5 gene segments of the American caprine RVB strain were genetically related to the Japanese bovine RVB strains. The results indicate a lack of RVB sequences to understand reassortment or the evolutionary relationship of RVB strains from cattle and goats.

Complete Genome Sequencing of Influenza A Viruses within Swine Farrow-to-Wean Farms Reveals the Emergence, Persistence, and Subsidence of Diverse Viral Genotypes.

Influenza A viruses (IAVs) are endemic in swine and represent a public health risk. However, there is limited information on the genetic diversity of swine IAVs within farrow-to-wean farms, which is where most pigs are born. In this longitudinal study, we sampled 5 farrow-to-wean farms for a year and collected 4,190 individual nasal swabs from three distinct pig subpopulations. Of these, 207 (4.9%) samples tested PCR positive for IAV, and 124 IAVs were isolated. We sequenced the complete genomes of 123 IAV isolates and found 31 H1N1, 26 H1N2, 63 H3N2, and 3 mixed IAVs. Based on the IAV hemagglutinin, seven different influenza A viral groups (VGs) were identified. Most of the remaining IAV gene segments allowed us to differentiate the same VGs, although an additional viral group was identified for gene segment 3 (PA). Moreover, the codetection of more than one IAV VG was documented at different levels (farm, subpopulation, and individual pigs), highlighting the environment for potential IAV reassortment. Additionally, 3 out of 5 farms contained IAV isolates (n = 5) with gene segments from more than one VG, and 79% of all the IAVs sequenced contained a signature mutation (S31N) in the matrix gene that has been associated with resistance to the antiviral amantadine. Within farms, some IAVs were detected only once, while others were detected for 283 days. Our results illustrate the maintenance and subsidence of different IAVs within swine farrow-to-wean farms over time, demonstrating that pig subpopulation dynamics are important to better understand the diversity and epidemiology of swine IAVs.IMPORTANCE On a global scale, swine are one of the main reservoir species for influenza A viruses (IAVs) and play a key role in the transmission of IAVs between species. Additionally, the 2009 IAV pandemics highlighted the role of pigs in the emergence of IAVs with pandemic potential. However, limited information is available regarding the diversity and distribution of swine IAVs on farrow-to-wean farms, where novel IAVs can emerge. In this study, we studied 5 swine farrow-to-wean farms for a year and characterized the genetic diversity of IAVs among three different pig subpopulations commonly housed on this type of farm. Using next-generation-sequencing technologies, we demonstrated the complex distribution and diversity of IAVs among the pig subpopulations studied. Our results demonstrated the dynamic evolution of IAVs within farrow-to-wean farms, which is crucial to improve health interventions to reduce the risk of transmission between pigs and from pigs to people.

Deletion of a 197-Amino-Acid Region in the N-Terminal Domain of Spike Protein Attenuates Porcine Epidemic Diarrhea Virus in Piglets.

We previously isolated a porcine epidemic diarrhea virus (PEDV) strain, PC177, by blind serial passaging of the intestinal contents of a diarrheic piglet in Vero cell culture. Compared with the highly virulent U.S. PEDV strain PC21A, the tissue culture-adapted PC177 (TC-PC177) contains a 197-amino-acid (aa) deletion in the N-terminal domain of the spike (S) protein. We orally inoculated neonatal, conventional suckling piglets with TC-PC177 or PC21A to compare their pathogenicities. Within 7 days postinoculation, TC-PC177 caused mild diarrhea and lower fecal viral RNA shedding, with no mortality, whereas PC21A caused severe clinical signs and 55% mortality. To investigate whether infection with TC-PC177 can induce cross-protection against challenge with a highly virulent PEDV strain, all the surviving piglets were challenged with PC21A at 3 weeks postinoculation. Compared with 100% protection in piglets initially inoculated with PC21A, 88% and 100% TC-PC177- and mock-inoculated piglets had diarrhea following challenge, respectively, indicating incomplete cross-protection. To investigate whether this 197-aa deletion was the determinant for the attenuation of TC-PC177, we generated a mutant (icPC22A-S1Δ197) bearing the 197-aa deletion from an infectious cDNA clone of the highly virulent PEDV PC22A strain (infectious clone PC22A, icPC22A). In neonatal gnotobiotic pigs, the icPC22A-S1Δ197 virus caused mild to moderate diarrhea, lower titers of viral shedding, and no mortality, whereas the icPC22A virus caused severe diarrhea and 100% mortality. Our data indicate that deletion of this 197-aa fragment in the spike protein can attenuate a highly virulent PEDV, but the virus may lose important epitopes for inducing robust protective immunity.IMPORTANCE The emerging, highly virulent PEDV strains have caused substantial economic losses worldwide. However, the virulence determinants are not established. In this study, we found that a 197-aa deletion in the N-terminal region of the S protein did not alter virus (TC-PC177) tissue tropism but reduced the virulence of the highly virulent PEDV strain PC22A in neonatal piglets. We also demonstrated that the primary infection with TC-PC177 failed to induce complete cross-protection against challenge by the highly virulent PEDV PC21A, suggesting that the 197-aa region may contain important epitopes for inducing protective immunity. Our results provide an insight into the role of this large deletion in virus propagation and pathogenicity. In addition, the reverse genetics platform of the PC22A strain was further optimized for the rescue of recombinant PEDV viruses in vitro This breakthrough allows us to investigate other virulence determinants of PEDV strains and will provide knowledge leading to better control PEDV infections.

A porcine enterovirus G associated with enteric disease contains a novel papain-like cysteine protease.

Identification of unknown pathogens in pigs displaying enteric illness is difficult due to the large diversity of bacterial and viral species found within faecal samples. Current methods often require bacterial or viral isolation, or testing only a limited number of known species using quantitative PCR analysis. Herein, faeces from two 25-day-old piglets with diarrhoea from Texas, USA, were analysed by metagenomic next-generation sequencing to rapidly identify possible pathogens. Our analysis included a bioinformatics pipeline of rapid short-read classification and de novo genome assembly which resulted in the identification of a porcine enterovirus G (EV-G), a complete genome with substantial nucleotide differences (>30 %) among current sequences, and a novel non-structural protein similar in sequence to the Torovirus papain-like cysteine protease (PLpro). This discovery led to the identification and circulation of an EV-G with a novel PLpro in the USA that has not been previously reported.

Genome characterization of Turkey Rotavirus G strains from the United States identifies potential recombination events with human Rotavirus B strains.

Rotavirus G (RVG) strains have been detected in a variety of avian species, but RVG genomes have been published from only a single pigeon and two chicken strains. Two turkey RVG strains were identified and characterized, one in a hatchery with no reported health issues and the other in a hatchery with high embryo/poult mortality. The two turkey RVG strains shared only an 85.3 % nucleotide sequence identity in the VP7 gene while the other genes possessed high nucleotide identity among them (96.3-99.9 %). Low nucleotide percentage identities (31.6-87.3 %) occurred among the pigeon and chicken RVG strains. Interestingly, potential recombination events were detected between our RVG strains and a human RVB strain, in the VP6 and NSP3 segments. The epidemiology of RVG in avian flocks and the pathogenicity of the two different RVG strains should be further investigated to understand the ecology and impact of RVG in commercial poultry flocks.

Comparative In Vitro and In Vivo Studies of Porcine Rotavirus G9P[13] and Human Rotavirus Wa G1P[8].

UNLABELLED: The changing epidemiology of group A rotavirus (RV) strains in humans and swine, including emerging G9 strains, poses new challenges to current vaccines. In this study, we comparatively assessed the pathogenesis of porcine RV (PRV) G9P[13] and evaluated the short-term cross-protection between this strain and human RV (HRV) Wa G1P[8] in gnotobiotic pigs. Complete genome sequencing demonstrated that PRV G9P[13] possessed a human-like G9 VP7 genotype but shared higher overall nucleotide identity with historic PRV strains. PRV G9P[13] induced longer rectal virus shedding and RV RNAemia in pigs than HRV Wa G1P[8] and generated complete short-term cross-protection in pigs challenged with HRV or PRV, whereas HRV Wa G1P[8] induced only partial protection against PRV challenge. Moreover, PRV G9P[13] replicated more extensively in porcine monocyte-derived dendritic cells (MoDCs) than did HRV Wa G1P[8]. Cross-protection was likely not dependent on serum virus-neutralizing (VN) antibodies, as the heterologous VN antibody titers in the sera of G9P[13]-inoculated pigs were low. Thus, our results suggest that heterologous protection by the current monovalent G1P[8] HRV vaccine against emerging G9 strains should be evaluated in clinical and experimental studies to prevent further dissemination of G9 strains. Differences in the pathogenesis of these two strains may be partially attributable to their variable abilities to replicate and persist in porcine immune cells, including dendritic cells (DCs). Additional studies are needed to evaluate the emerging G9 strains as potential vaccine candidates and to test the susceptibility of various immune cells to infection by G9 and other common HRV/PRV genotypes. IMPORTANCE: The changing epidemiology of porcine and human group A rotaviruses (RVs), including emerging G9 strains, may compromise the efficacy of current vaccines. An understanding of the pathogenesis and genetic, immunological, and biological features of the new emerging RV strains will contribute to the development of new surveillance and prevention tools. Additionally, studies of cross-protection between the newly identified emerging G9 porcine RV strains and a human G1 RV vaccine strain in a susceptible host (swine) will allow evaluation of G9 strains as potential novel vaccine candidates to be included in porcine or human vaccines.

Longitudinal study of Staphylococcus aureus colonization and infection in a cohort of swine veterinarians in the United States.

BACKGROUND: People working with pigs are at elevated risk of harboring methicillin resistant S. aureus (MRSA) in their nose, which is attributable to occupational exposure to animals harboring livestock adapted S. aureus. To obtain insight into the biological nature of occupationally related nasal culture positivity, we conducted a longitudinal study of 66 swine veterinarians in the USA. METHODS: The study cohort resided in 15 US states and worked predominantly with swine. Monthly for 18 months, participants self-collected nasal swabs and completed a survey to report recent exposure to pigs and other animals; the occurrence of work related injuries; and any relevant health events such as skin and soft tissue infections or confirmed staphylococcal infections. Nasal swabs were cultured using selective methods to determine the presence of MRSA and methicillin susceptible S. aureus (MSSA), and isolates were characterized by spa typing and MLST. RESULTS: Prevalences of S. aureus (64%, monthly range from 58 to 82%) and MRSA (9.5%; monthly range from 6 to15%) were higher than reported for the US population (30% and 1.5% respectively). Predominant spa types were t034 (ST398, 37%), t002 (ST5, 17%) and t337 (ST9/ST398 13%), a distribution similar to that found in a concurrent study in pigs in the USA. Veterinarians were classified into three groups: Persistent carriers (PC, 52%), Intermittent carriers (IC, 47%) and Non-carriers (NC, 1%). Persistent carriage of a single spa type was observed in 14 (21%) of participants, and paired (first and last) isolates from PC subjects had minor genetic differences. Swabs from PC veterinarians carried higher numbers of S. aureus. Among IC veterinarians, culture positivity was significantly associated with recent contact with pigs. CONCLUSIONS: Exposure to pigs did not lead to prolonged colonization in most subjects, and the higher numbers of S. aureus in PC subjects suggests that unknown host factors may determine the likelihood of prolonged colonization by S. aureus of livestock origin. Exposure to S. aureus and persistent colonization of swine veterinarians was common but rarely associated with S. aureus disease.

Longitudinal study of Senecavirus a shedding in sows and piglets on a single United States farm during an outbreak of vesicular disease.

BACKGROUND: The study highlights the shedding pattern of Senecavirus A (SVA) during an outbreak of vesicular disease in a sow farm from the South-central Minnesota, USA. In this study, 34 individual, mixed parity sows with clinical signs of vesicular lesions and 30 individual piglets from 15 individual litters from sows with vesicular lesions were conveniently selected for individual, longitudinal sampling. Serum, tonsil, rectal, and vesicular swabs were collected on day1 post outbreak, and then again at 1, 2, 3, 4, 6, and 9 weeks post outbreak. Samples were tested at the University of Minnesota Veterinary Diagnostic Laboratory for SVA via Real Time Polymerase Chain Reaction (RT-PCR) RESULTS: In sows, vesicular lesions had the highest concentration of SVA, but had the shortest duration of detection lasting only 2 weeks. Viremia was detected for 1 week post outbreak, and quickly declined thereafter. SVA was detected at approximately the same frequency for both tonsil and rectal swabs with the highest percentage of SVA positive samples detected in the first 6 weeks post outbreak. In suckling piglets, viremia quickly declined 1 week post outbreak and was prevalent in low levels during the first week after weaning (4 weeks post outbreak) and was also detected in piglets that were co-mingled from a SVA negative sow farm. Similar to sows, SVA detection on rectal and tonsil swabs in piglets lasted approximately 6 weeks post outbreak. CONCLUSION: The study illustrates the variation of SVA shedding patterns in different sample types over a 9 week period in sows and piglets, and suggests the potential for viral spread between piglets at weaning.

Porcine Epidemic Diarrhea Virus and Discovery of a Recombinant Swine Enteric Coronavirus, Italy.

Porcine epidemic diarrhea virus (PEDV) has been detected sporadically in Italy since the 1990s. We report the phylogenetic relationship of swine enteric coronaviruses collected in Italy during 2007-2014 and identify a drastic shift in PEDV strain variability and a new swine enteric coronavirus generated by recombination of transmissible gastroenteritis virus and PEDV.

Detection of a novel circovirus PCV3 in pigs with cardiac and multi-systemic inflammation.

BACKGROUND: Porcine circovirus 2 causes different clinical syndromes resulting in a significant economic loss in the pork industry. Three pigs with unexplained cardiac and multi-organ inflammation that tested negative for PCV2 and other known porcine pathogens were further analyzed. METHODS: Histology was used to identify microscopic lesions in multiple tissues. Metagenomics was used to detect viral sequences in tissue homogenates. In situ hybridization was used to detect viral RNA expression in cardiac tissue. RESULTS: In all three cases we characterized the genome of a new circovirus we called PCV3 with a replicase and capsid proteins showing 55 and 35 % identities to the genetically-closest proteins from a bat-feces associated circovirus and were even more distant to those of porcine circovirus 1 and 2. Common microscopic lesions included non-suppurative myocarditis and/or cardiac arteriolitis. Viral mRNA was detected intralesionally in cardiac cells. Deep sequencing in tissues also revealed the presence of porcine astrovirus 4 in all three animals as well as rotavirus A, porcine cytomegalovirus and porcine hemagglutinating encephalomyelitis virus in individual cases. CONCLUSION: The pathogenicity and molecular epidemiology of this new circovirus, alone or in the context of co-infections, warrants further investigations.

A comprehensive microbiological safety approach for agarose encapsulated porcine islets intended for clinical trials.

BACKGROUND: The use of porcine islets to replace insulin-producing islet ß-cells, destroyed during the diabetogenic disease process, presents distinct challenges if this option is to become a therapeutic reality for the treatment of type 1 diabetes. These challenges include a thorough evaluation of the microbiological safety of the islets. In this study, we describe a robust porcine islet-screening program that provides a high level of confidence in the microbiological safety of porcine islets suitable for clinical trials. METHODS: A four-checkpoint program systematically screens the donor herd (Large White - Yorkshire × Landrace F1 hybrid animals), individual sentinel and pancreas donor animals and, critically, the islet macrobeads themselves. Molecular assays screen for more than 30 known viruses, while electron microscopy and in vitro studies are employed to screen for potential new or divergent (emergent) viruses. RESULTS: Of 1207 monthly samples taken from random animals over a 2-year period, only a single positive result for Transmissible gastroenteritis virus was observed, demonstrating the high level of biosecurity maintained in the source herd. Given the lack of clinical signs, positive antibody titers for Porcine reproductive and respiratory syndrome virus, Porcine parvovirus, and Influenza A confirm the efficacy of the herd vaccination program. Porcine respiratory coronavirus was found to be present in the herd, as expected for domestic swine. Tissue homogenate samples from six sentinel and 11 donor animals, over the same 2-year period, were negative for the presence of viruses when co-cultured with six different cell lines from four species. The absence of adventitious viruses in separate islet macrobead preparations produced from 12 individual pancreas donor animals was confirmed using validated molecular (n = 32 viruses), in vitro culture (cells from four species), and transmission electron microscopy assays (200 cell profiles per donor animal) over the same 2-year period. There has been no evidence of viral transmission following the implantation of these same encapsulated and functional porcine islets into non-immunosuppressed diabetic cynomolgus macaques for up to 4 years. Isolated peripheral blood mononuclear cells from all time points were negative for PCV (Type 2), PLHV, PRRSV, PCMV, and PERV-A, PERV-B, and PERV-C by PCR analysis in all six recipient animals. CONCLUSION: The four-checkpoint program is a robust and reliable method for characterization of the microbiological safety of encapsulated porcine islets intended for clinical trials.

Rapid detection, complete genome sequencing, and phylogenetic analysis of porcine deltacoronavirus.

In February 2014, porcine deltacoronavirus (PDCoV) was identified in the United States. We developed a PDCoV real-time reverse transcription PCR that identified PDCoV in 30% of samples tested. Four additional PDCoV genomes from the United States were sequenced; these had ≈99%-100% nt similarity to the other US PDCoV strains.

Widespread rotavirus H in commercially raised pigs, United States.

We investigated the presence in US pigs of rotavirus H (RVH), identified in pigs in Japan and Brazil. From 204 samples collected during 2006-2009, we identified RVH in 15% of fecal samples from 10 US states, suggesting that RVH has circulated in the United States since 2002, but probably longer.

Distinct characteristics and complex evolution of PEDV strains, North America, May 2013-February 2014.

Porcine epidemic diarrhea virus (PEDV), which emerged in the United States in 2013, has spread throughout North America. Limited availability of PEDV complete genomes worldwide has impeded our understanding of PEDV introduction into the United States. To determine the relationship between the North American strains and global emerging and historic PEDV strains, we sequenced and analyzed complete genomes of 74 strains from North America; the strains clustered into 2 distinct clades. Compared with the initially reported virulent US PEDV strains, 7 (9.7%) strains from 4 states contained insertions and deletions in the spike gene (S INDELs). These S INDEL strains share 99.8%-100% nt identity with each other and 96.2%-96.7% nt identity with the initial US strains. Furthermore, the S INDEL strains form a distinct cluster within North American clade II, sharing 98.6%-100% nt identity overall. In the United States, the S INDEL and original PEDV strains are co-circulating and could have been introduced simultaneously.