Translational control of the activation of transcription factor NF-κB and production of type I interferon by phosphorylation of the translation factor eIF4E.

Type I interferon is an integral component of the antiviral response, and its production is tightly controlled at the levels of transcription and translation. The eukaryotic translation-initiation factor eIF4E is a rate-limiting factor whose activity is regulated by phosphorylation of Ser209. Here we found that mice and fibroblasts in which eIF4E cannot be phosphorylated were less susceptible to virus infection. More production of type I interferon, resulting from less translation of Nfkbia mRNA (which encodes the inhibitor IκBα), largely explained this phenotype. The lower abundance of IκBα resulted in enhanced activity of the transcription factor NF-κB, which promoted the production of interferon-ß (IFN-ß). Thus, regulated phosphorylation of eIF4E has a key role in antiviral host defense by selectively controlling the translation of an mRNA that encodes a critical suppressor of the innate antiviral response.

Biological and molecular characterization of a resistance-breaking isolate of citrus tristeza virus from Uruguay and its effects on Poncirus trifoliata growth performance.

Resistance-breaking (RB) isolates of citrus tristeza virus (CTV) can replicate and move systemically in Poncirus trifoliata, a rootstock widely used for management of decline caused by CTV and other purposes. In Uruguay, severe CTV isolates are prevalent, and an RB isolate (designated as RB-UY1) was identified. In order to predict the implications of this genotype circulating in citrus crops grafted on trifoliate rootstocks, the aim of this work was to determine the biological and molecular characteristics of this isolate, the efficiency of its transmission by Toxoptera citricida, and its effects on plant growth performance of P. trifoliata. Our results show that RB-UY1 can be classified as a mild isolate, that it is phylogenetically associated with the RB1 group, and that it is efficiently transmitted by T. citrida. They also suggest that the RB-UY1 isolate should not affect the performance of citrus crops grafted on trifoliate rootstocks, although some growth parameters of P. trifoliata seedlings were affected four years after inoculation.

High abundance of high-risk Human Papillomavirus genotypes in wastewater in Uruguay.

The aim of this study was to determine the frequency of Human Papillomavirus (HPV) genotypes in wastewater of Salto city, Uruguay, in order to obtain a general overview of the circulating genotypes in their population. HPV was detected in 34% (32/93) of the wastewater samples collected and analyzed during 2020/21 in Salto city, Uruguay. Thirty-three genotypes were observed, of which 16 presented read abundance higher than 1%, including both high-risk (HR) and low-risk (LR) genotypes. HR genotypes 31, 16, 58, 52, 33 and 59 were detected representing 40% (163,220 reads) of the total read abundance, with genotypes 31 (64,365), 16 (39,337) and 58 (36,332) being the most abundant. LR genotypes 72, 6, 11 and 40 were also detected in a high frequency, accounting for 37% (148,359) of the HPV reads. This study highlights the high frequency of HR genotypes of HPV, circulating in the population of Salto city which is a burden in public health mainly due to the devastating impact of cervical cancer in women.

[Neurotropic bovine astrovirus-associated encephalitis: An underdiagnosed disease in South America?] / Encefalitis asociada a astrovirus bovino neurotrópico, ¿una enfermedad subdiagnosticada en Sudamérica?

We describe a case of neurotropic bovine astrovirus-associated encephalitis in a Jersey dairy cow from the department of San José, Uruguay. This represents the second case of this condition reported in the Southern Hemisphere. The cow was the only one affected in a herd of 70 cows, showing neurological signs with a 2-day clinical course, before dying spontaneously. Histopathological examination revealed lymphocytic, histiocytic, and plasmacytic meningoencephalitis with neuronal necrosis, without detectable inclusion bodies. Other infectious agents, including Rabies virus(Lyssavirus), Bovine alphaherpesvirus-1 and Bovine alphaherpesvirus-5(Varicellovirus), Bovine viral diarrhea virus(Pestivirus), West Nile virus(Flavivirus), Listeria monocytogenes, Histophilus somni and other bacteria, were not detected in the brain. We propose that given the recent discovery of neurotropic astroviruses in various mammalian species, including humans, cases of astrovirus encephalitis may have gone undetected in South America. We briefly discuss the differential pathologic diagnosis of infectious bovine encephalitis.

Real-Time Genomic Surveillance for SARS-CoV-2 Variants of Concern, Uruguay.

We developed a genomic surveillance program for real-time monitoring of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) in Uruguay. We report on a PCR method for SARS-CoV-2 VOCs, the surveillance workflow, and multiple independent introductions and community transmission of the SARS-CoV-2 P.1 VOC in Uruguay.

Evidence of increasing diversification of emerging Severe Acute Respiratory Syndrome Coronavirus 2 strains.

On 30th January 2020, an outbreak of atypical pneumonia caused by a novel betacoronavirus, named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was declared a public health emergency of international concern by the World Health Organization. For this reason, a detailed evolutionary analysis of SARS-CoV-2 strains currently circulating in different geographic regions of the world was performed. A compositional analysis as well as a Bayesian coalescent analysis of complete genome sequences of SARS-CoV-2 strains recently isolated in Europe, North America, South America, and Asia was performed. The results of these studies revealed a diversification of SARS-CoV-2 strains in three different genetic clades. Co-circulation of different clades in different countries, as well as different genetic lineages within different clades were observed. The time of the most recent common ancestor was established to be around 1st November 2019. A mean rate of evolution of 6.57 × 10-4 substitutions per site per year was found. A significant migration rate per genetic lineage per year from Europe to South America was also observed. The results of these studies revealed an increasing diversification of SARS-CoV-2 strains. High evolutionary rates and fast population growth characterizes the population dynamics of SARS-CoV-2 strains.

Hand-foot-and-mouth disease in uruguay: Coxsackievirus A6 identified as causative of an outbreak in a rural childcare center.

To know the epidemiological context of hand-foot-and-mouth disease (HFMD) in a region of Uruguay and to identify the Enterovirus responsible for an outbreak in a rural childcare center in 2018. Swab samples from skin lesions and/or stools samples were collected from children suffering HFMD during an outbreak in a rural childcare center. Samples were subject to viral RNA extraction and reverse-transcription polymerase chain reaction towards VP1 coding segment, to identify the Enterovirus type by sequencing and phylogenetic analysis. Total of 149 cases of HFMD affecting 98 boys and 51 girls were reported in Salto Province-Uruguay in 2018. Total 60% of the cases were originated from outbreaks, which occurred in ten educative and childcare institutions from both urban and rural areas. Coxsackievirus-6 (CV-A6) was identified as responsible for one of the rural outbreaks. Uruguayan strains were more related to strains reported in Russia, Turkey, and Germany (2014-2017) than to strains reported in Brazil and Argentina from 2015 to 2016. This is the first report of CV-A6-associated HFMD in Uruguay, evidencing a wide geographic range of the virus in the Latin American region. Our report also warns about CV-A6-associated HFMD during winter, contrarily to most reports that register HFMD during summer and fall seasons.

An extensive field study reveals the circulation of new genetic variants of subtype 1a of bovine viral diarrhea virus in Uruguay.

Bovine viral diarrhea virus (BVDV) is a major pathogen worldwide, causing significant economic losses to the livestock sector. In Uruguay, BVDV seroprevalence at the farm level is >80%. In this work, 2546 serum, blood or tissue samples collected from animals suspected of being affected by BVD between 2015 and 2017 were analyzed by reverse transcription PCR and sequencing. Analysis of the BVDV genomic regions 5'UTR/Npro, Npro and E2 revealed that BVDV-1a, 1i and 2b circulate in the country, with BVDV-1a being the most prevalent subtype. Population dynamics studies revealed that BVDV-1a has been circulating in our herds since ~1990. This subtype began to spread and evolve, accumulating point mutations at a rate of 3.48 × 10-3 substitutions/site/year, acquiring specific genetic characteristics that gave rise to two local genetic lineages of BVDV-1a. These lineages are divergent from those circulating worldwide, as well as the vaccine strain currently used in Uruguay. The most notable differences between field and vaccine strains were found in the E2 glycoprotein, suggesting that the amino acid substitutions could result in failure of cross-protection/neutralization after vaccination. This is the first study that compares Uruguayan BVDV field and vaccine strains with other BVDV strains from throughout the world. The results obtained in this study will be very useful for developing a suitable immunization program for BVDV in Uruguay by identifying local field strains as candidates for vaccine development.

Borrelia burgdorferi sensu lato infecting Ixodes auritulus ticks in Uruguay.

In the southern cone of South America different haplotypes of Borrelia burgdorferi sensu lato (Bbsl) have been detected in Ixodes spp. from Argentina, southern Brazil, Chile, and Uruguay. So far, Lyme borreliosis has not been diagnosed in Uruguay and the medical relevance of the genus Ixodes in South America is uncertain. However, the growing number of new genospecies of Bbsl in the southern cone region and the scarce information about its pathogenicity, reservoirs and vectors, highlights the importance of further studies about spirochetes present in Uruguay and the region. The aim of this study was to determine the presence of Bbsl in Ixodes auritulus ticks collected from birds and vegetation in two localities of southeastern Uruguay. In total 306 I. auritulus were collected from 392 passerine birds sampled and 1110 ticks were collected by flagging in vegetation. Nymphs and females were analyzed for Borrelia spp. by PCR targeting the flagellin (fla) gene and the rrfA-rrlB intergenic spacer region (IGS). The phylogenetic analysis of Borrelia spp. positive samples from passerine birds and vegetation revealed the presence of four fla haplotypes that form a clade within the Bbsl complex. They were closely related to isolates of Borrelia sp. detected in I. auritulus from Argentina and Canada.

Bovine coronavirus in Uruguay: genetic diversity, risk factors and transboundary introductions from neighboring countries.

Bovine coronavirus (BCoV) is a recognized cause of severe neonatal calf diarrhea, with a negative impact on animal welfare, leading to economic losses to the livestock industry. Cattle production is one of the most important economic sectors in Uruguay. The aim of this study was to determine the frequency of BCoV infections and their genetic diversity in Uruguayan calves and to describe the evolutionary history of the virus in South America. The overall detection rate of BCoV in Uruguay was 7.8% (64/824): 7.7% (60/782) in dairy cattle and 9.5% (4/42) in beef cattle. The detection rate of BCoV in samples from deceased and live calves was 10.0% (6/60) and 7.6% (58/763), respectively. Interestingly, there was a lower frequency of BCoV detection in calves born to vaccinated dams (3.3%, 8/240) than in calves born to unvaccinated dams (12.2%, 32/263) (OR: 4.02, 95%CI: 1.81-8.90; p = 0.00026). The frequency of BCoV detection was higher in colder months (11.8%, 44/373) than in warmer months (1.5%, 3/206) (OR: 9.05, 95%CI: 2.77-29.53, p = 0.000013). Uruguayan strains grouped together in two different lineages: one with Argentinean strains and the other with Brazilian strains. Both BCoV lineages were estimated to have entered Uruguay in 2013: one of them from Brazil (95%HPD interval: 2011-2014) and the other from Argentina (95%HPD interval: 2010-2014). The lineages differed by four amino acid changes, and both were divergent from the Mebus reference strain. Surveillance should be maintained to detect possible emerging strains that can clearly diverge at the antigenic level from vaccine strains.

Fetal Pathology in an Aborted Holstein Fetus Infected With Bovine Parainfluenza Virus-3 Genotype A.

Bovine parainfluenza virus-3 (BPIV-3) is a recognized respiratory pathogen of cattle, and it has also been identified in aborted fetuses. However, little is known of this agent as a reproductive pathogen and detailed descriptions of fetal pathology on natural cases are lacking in the scientific literature. This article describes and illustrates lesions in a fetus spontaneously aborted by a first-calving Holstein heifer, naturally infected with BPIV-3 genotype A, broadening the current knowledge on fetal pathology by this virus. Fetal autopsy revealed diffusely reddened, rubbery and unexpanded lungs. Histologically, there was necrotizing bronchiolitis/alveolitis with intraluminal fibrin exudate and syncytial cells in the bronchiolar/alveolar spaces, and non-suppurative peribronchiolitis and perivascular interstitial pneumonia. In the small intestine there was multifocal necrotizing cryptitis and occasional necrotic syncytial enterocytes. Intralesional and extralesional BPIV-3 antigen was detected by immunohistochemistry in the lung and small intestine, and BPIV-3a was identified in fetal tissues by RT-PCR and sequencing.

Description of a human Bocavirus recombinant strain in the Americas.

Human bocaviruses (HBoV) are mainly associated with respiratory and gastroenteric infections. These viruses belong to the family Parvoviridae, genus Bocaparvovirus and are classified in four subtypes (HBoV1-4). Recombination and point mutation have been described as basis of parvovirus evolution. In this study three viral sequences were obtained from positives HBoV sewage samples collected in two Uruguayan cities and were characterised by different methods as recombinant strains. This recombination event was localised in the 5' end of VP1 gene and the parental strains belonged to subtypes 3 and 4. These three Uruguayan strains are identical at the nucleotide sequences in the analysed genome region of the virus. As far as we known, this study represents the first detection of HBoV recombinants strains in the Americas.

Phylodynamics of Merkel-cell polyomavirus and human polyomavirus 6: A long-term history with humans.

New human polyomaviruses have been described in the last years, including the Merkel-cell polyomavirus (MCPyV; Human polyomavirus 5) and the Human polyomavirus 6 (HPyV6). Although their infection is usually asymptomatic, in immunocompromised host can cause life-threatening pathologies, such as the Merkel cell carcinoma, an aggressive skin neoplasia associated to the MCPyV. Despite being prevalent viruses in population, epidemiological data from South America are scarce, as well as the characterization of the viral types circulating and their origin. The aims of this work were to describe MCPyV and HPyV6 from environmental samples with different geographical origin and to analyze their phylogenetic and evolutionary histories, particularly for MCPyV. Partial and complete genome sequences were obtained from sewage samples from Argentina, Uruguay and Spain. A total number of 87 sequences were obtained for MCPyV and 33 for HPyV6. Phylogenetic analysis showed that MCPyV sequences distributed according to their geographic origin in Europe/North America, Africa, Asia, South America and Oceania groups, suggesting that viral diversification might have followed human migrations across the globe. In particular, viruses from Argentina associated with Europe/North America and South America genotypes, whereas those from Uruguay and Spain also grouped with Africa genotype, reflecting the origin of the current population in each country, which could arrive not only during ancient human migration but also during recent migratory events. In addition, the South American group presented a high level of clusterization, showing internal clusters that could be related to specific locations, such as French Guiana and Brazil or the Southern region into South America, such as Argentina and Uruguay, suggesting a long term evolutionary process in the region. Additionally, in this work, we carried out the first analysis about the evolutionary history of MCPyV trough the integration of phylogenetic, epidemiological and historical data. Since a strong association is observed between the phylogenetic relationships and the origin of the sampled population, this analysis was based on the hypothesis of co-divergence between the virus and human populations. This analysis resulted in a substitution rate of 5.1 × 10-8 s/s/y (∼5.1% of divergence per million years) for the complete genome of MCPyV, which is in the range of those estimated for other double-stranded DNA viruses. Regarding HPyV6, a South American group with clusterization was observed (sequences from Uruguay). Meanwhile, sequences from Argentina grouped with European ones (France and Spain) and remained separated from those isolated in China, USA or Australia. The analysis of viruses from the environment allowed us to deep characterize prevalent infections in different geographic regions, reveling that viruses circulating in each population reflected its origin and that there are specific lineages associated with South America.

Epidemiology and genetic diversity of classic human astrovirus among hospitalized children with acute gastroenteritis in Uruguay.

Classic Human Astrovirus (Classic HAstV) are one of the most important causes of pediatric acute gastroenteritis (AGE), after rotaviruses and arguably caliciviruses. The aim of this study was to determine the molecular epidemiology of Classic HAstV from 175 clinical samples, being 153 stools and 22 vomits, collected from pediatric hospitalized patients with AGE in Salto city, Uruguay, from January 2011 to December 2012. Classic HAstV were detected and genotyped by using a qualitative Retro Transcription-Polimerase Chain Reaction (RT-PCR) directed to the Open Reading Frame-2 (ORF2) region C. Amplicons were sequenced and phylogenetic analyses were carried out in order to determine genotypes and lineages. Classic HAstV were detected in 18 out of 175 analyzed samples (10.3%) and 14 of them (78.0%) were successfully sequenced being 6 (42.8%) classified as HAstV-1 (1a lineage), 4 (28.6%) as HAstV-2 (2c lineage), and 4 (28.6%) as HAstV-3 (3c lineage). A higher detection of Classic HAstV infections was observed in autumn for both years of surveillance, and the majority of the positive cases were observed in 2011. The group of children between 2 and 5 years old presented the higher percentage of infections. To our knowledge, the present study represents the first report of astrovirus from acute gastroenteritis cases in Uruguay, evidencing its role as a relevant etiologic agent in severe cases of this disease.

Genome-wide analysis of codon usage bias in Bovine Coronavirus.

BACKGROUND: Bovine coronavirus (BCoV) belong to the genus Betacoronavirus of the family Coronaviridae. BCoV are widespread around the world and cause enteric or respiratory infections among cattle, leading to important economic losses to the beef and dairy industry worldwide. To study the relation of codon usage among viruses and their hosts is essential to understand host-pathogen interaction, evasion from host's immune system and evolution. METHODS: We performed a comprehensive analysis of codon usage and composition of BCoV. RESULTS: The global codon usage among BCoV strains is similar. Significant differences of codon preferences in BCoV genes in relation to codon usage of Bos taurus host genes were found. Most of the highly frequent codons are U-ending. G + C compositional constraint and dinucleotide composition also plays a role in the overall pattern of BCoV codon usage. CONCLUSIONS: The results of these studies revealed that mutational bias is a leading force shaping codon usage in this virus. Additionally, relative dinucleotide frequencies, geographical distribution, and evolutionary processes also influenced the codon usage pattern.

Molecular epidemiology of group A rotavirus among children admitted to hospital in Salto, Uruguay, 2011-2012: first detection of the emerging genotype G12.

Group A rotavirus (RVA) is the most important etiologic agent of infant acute gastroenteritis (AGE) worldwide. Detection and molecular characterization of RVA in Salto department, Northwestern region of Uruguay, was conducted on 175 clinical samples, being 153 stool and 22 vomit samples, collected from hospitalized children with AGE, between 0-15 years old, from two hospitals of Salto city during 2011 and 2012. RVA was detected and genotyped by seminested multiplex RT-PCR in order to determine G- and P-genotypes. Positive samples were sequenced and phylogenetic analyses were carried out in order to determine lineages and sub-lineages. RVA were detected in 64 (37%) of the samples and the G and P genotypes observed were: 6% G1P[8], 23% G2P[4]/G2P[X]/GXP[4], 23% G3P[8]/G3P[X], 14% G12P[8]/G12P[X], 16% GXP[8], 1,5% G12P[9], 3% G2P[4]/[8], and 16% non-typeable. VP7 and VP4 genotypes related to DS-1 like gene constellation were prevalent during 2011 and those VP7 and VP4 genotypes related to Wa-like constellation were prevalent during 2012 (mainly represented by G3P[8]). Interestingly, RVA was detected in vomit samples in a high prevalence (41%). RVA was observed mainly in the age group between 1 and 5 years old (75% of the cases), and seasonality with a high detection rate in winter season was observed for the two consecutive years of surveillance. To our knowledge, this study represents the first detection and molecular characterization of RVA in Salto department, Northwestern region of Uruguay; and the first identification of the emerging genotype G12 in the country.

Translational control of the innate immune response through IRF-7.

Transcriptional activation of cytokines, such as type-I interferons (interferon (IFN)-alpha and IFN-beta), constitutes the first line of antiviral defence. Here we show that translational control is critical for induction of type-I IFN production. In mouse embryonic fibroblasts lacking the translational repressors 4E-BP1 and 4E-BP2, the threshold for eliciting type-I IFN production is lowered. Consequently, replication of encephalomyocarditis virus, vesicular stomatitis virus, influenza virus and Sindbis virus is markedly suppressed. Furthermore, mice with both 4E- and 4E-BP2 genes (also known as Eif4ebp1 and Eif4ebp2, respectively) knocked out are resistant to vesicular stomatitis virus infection, and this correlates with an enhanced type-I IFN production in plasmacytoid dendritic cells and the expression of IFN-regulated genes in the lungs. The enhanced type-I IFN response in 4E-BP1-/- 4E-BP2-/- double knockout mouse embryonic fibroblasts is caused by upregulation of interferon regulatory factor 7 (Irf7) messenger RNA translation. These findings highlight the role of 4E-BPs as negative regulators of type-I IFN production, via translational repression of Irf7 mRNA.

Detection and Genetic Characterization of Bovine Torovirus in Uruguay.

Bovine torovirus (BToV) is an enteric pathogen that may cause diarrhea in calves and adult cattle, which could result in economic losses due to weight loss and decreased milk production. This study aimed to report the presence, the genetic characterization and the evolution of BToV in calves in Uruguay. BToV was detected in 7.9% (22/278) of fecal samples, being identified in dairy (9.2%, 22/239) but not beef (0.0%, 0/39) calves. BToV was detected in both diarrheic (14%, 6/43) and non-diarrheic (13.2%, 5/38) dairy calves. In addition, BToV was detected in the intestinal contents of 14.9% (7/47) of naturally deceased dairy calves. A complete genome (28,446 nucleotides) was obtained, which was the second outside Asia and the first in Latin America. In addition, partial S gene sequences were obtained to perform evolutionary analyses. Nucleotide and amino acid substitutions within and between outbreaks/farms were observed, alerting the continuous evolution of the virus. Through Bayesian analysis using BEAST, a recent origin (mid-60s) of BToV, possibly in Asia, was estimated, with two introductions into Uruguay from Asia and Europe in 2004 and 2013, respectively. The estimated evolutionary rate was 1.80 × 10-3 substitutions/site/year. Our findings emphasize the importance of continued surveillance and genetic characterization for the effective management and understanding of BToV's global epidemiology and evolution.

Development of an IgY-Based Treatment to Control Bovine Coronavirus Diarrhea in Dairy Calves.

Bovine Coronavirus (BCoV) is a major pathogen associated with neonatal calf diarrhea. Standard practice dictates that to prevent BCoV diarrhea, dams should be immunized in the last stage of pregnancy to increase BCoV-specific antibody (Ab) titers in serum and colostrum. For the prevention to be effective, calves need to suck maternal colostrum within the first six to twelve hours of life before gut closure to ensure a good level of passive immunity. The high rate of maternal Ab transfer failure resulting from this process posed the need to develop alternative local passive immunity strategies to strengthen the prevention and treatment of BCoV diarrhea. Immunoglobulin Y technology represents a promising tool to address this gap. In this study, 200 laying hens were immunized with BCoV to obtain spray-dried egg powder enriched in specific IgY Abs to BCoV on a large production scale. To ensure batch-to-batch product consistency, a potency assay was statistically validated. With a sample size of 241, the BCoV-specific IgY ELISA showed a sensitivity and specificity of 97.7% and 98.2%, respectively. ELISA IgY Abs to BCoV correlated with virus-neutralizing Ab titers (Pearson correlation, R2 = 0.92, p < 0.001). Most importantly, a pilot efficacy study in newborn calves showed a significant delay and shorter duration of BCoV-associated diarrhea and shedding in IgY-treated colostrum-deprived calves. Calves were treated with milk supplemented with egg powder (final IgY Ab titer to BCoV ELISA = 512; VN = 32) for 14 days as a passive treatment before a challenge with BCoV and were compared to calves fed milk with no supplementation. This is the first study with proof of efficacy of a product based on egg powder manufactured at a scale that successfully prevents BCoV-associated neonatal calf diarrhea.

Epidemiology Update of Hepatitis E Virus (HEV) in Uruguay: Subtyping, Environmental Surveillance and Zoonotic Transmission.

Hepatitis E Virus (HEV) infection is an emergent zoonotic disease of increasing concern in developed regions. HEV genotype 3 (HEV-3) is mainly transmitted through consumption of contaminated food in high-income countries and is classified into at least 13 subtypes (3a-3n), based on p-distance values from complete genomes. In Latin America, HEV epidemiology studies are very scant. Our group has previously detected HEV3 in clinical cases, swine, wild boars, captive white-collared peccaries, and spotted deer from Uruguay. Herein, we aimed to provide novel insights and an updated overview of the molecular epidemiology of zoonotic HEV in Uruguay, including data from wastewater-based surveillance studies. A thorough analysis of HEV whole genomes and partial ORF2 sequences from Uruguayan human and domestic pig strains showed that they formed a separate monophyletic cluster with high nucleotide identity and exhibited p-distance values over the established cut-off (0.093) compared with reference subtypes' sequences. Furthermore, we found an overall prevalence of 10.87% (10/92) in wastewater, where two samples revealed a close relationship with humans, and animal reservoirs/hosts isolates from Uruguay. In conclusion, a single, new HEV-3 subtype currently circulates in different epidemiological settings in Uruguay, and we propose its designation as 3o along with its reference sequence.