First Report on Detection and Complete Genomic Analysis of a Novel CRESS DNA Virus from Sea Turtles.

To date, only a handful of viruses have been identified in sea turtles. Although eukaryotic circular Rep (replication initiation protein)-encoding single-stranded DNA (CRESS DNA) viruses have been reported from a wide variety of terrestrial species, and some of these viruses have been associated with clinical conditions in certain animals, limited information is available on CRESS DNA viruses from marine life. The present study aimed to investigate the presence of CRESS DNA viruses in sea turtles. In the present study, two (samples T3 and T33) of the 34 cloacal samples from 31 sea turtles (found in ocean waters around the Caribbean Islands of St. Kitts and Nevis) tested positive for CRESS DNA viruses by a pan-rep nested PCR assay. The partial Rep sequence of T3 shared 75.78% of a deduced amino acid (aa) identity with that of a CRESS DNA virus (classified under family Circoviridae) from a mollusk. On the other hand, the complete genome (2428 bp) of T33 was determined by an inverse nested PCR assay. The genomic organization of T33 mirrored those of type II CRESS DNA viral genomes of cycloviruses, characterized by the putative "origin of replication" in the 5'-intergenic region, and the putative Capsid (Cap)- and Rep-encoding open reading frame on the virion-sense- and antisense-strand, respectively. The putative Rep (322 aa) of T33 retained the conserved "HUH endonuclease" and the "super 3 family helicase" domains and shared pairwise aa identities of ~57% with unclassified CRESS DNA viruses from benthic sediment and mollusks. Phylogenetically, the T33 Rep formed a distinct branch within an isolated cluster of unclassified CRESS DNA viruses. The putative Cap (370 aa) of T33 shared maximum pairwise aa identity of 30.51% with an unclassified CRESS DNA virus from a capybara. Except for a blood sample from T33 that tested negative for CRESS DNA viruses, other tissue samples were not available from the sea turtles. Therefore, we could not establish whether the T3 and T33 viral strains infected the sea turtles or were of dietary origin. To our knowledge, this is the first report on the detection of CRESS DNA viruses from sea turtles, adding yet another animal species to the rapidly expanding host range of these viruses. Complete genome analysis of T33 identified a novel, unclassified CRESS DNA virus, providing insights into the high genetic diversity between viruses within the phylum Cressdnaviricota. Considering that sea turtles are an at-risk species, extensive studies on virus discovery, surveillance, and pathogenesis in these marine animals are of the utmost importance.

Detection and Complete Genomic Analysis of Porcine circovirus 3 (PCV3) in Diarrheic Pigs from the Dominican Republic: First Report on PCV3 from the Caribbean Region.

The increasing detection of Porcine circovirus 3 (PCV3, family Circoviridae) in clinically ill pigs worldwide has raised concerns on the implications of the virus on porcine health and the pork industry. Although pork production constitutes an important component of the livestock economy and is a major source of animal protein in the Caribbean Islands, there are no reports on PCV3 in pigs from the region so far. In the present study, PCV3 was detected in 21% (21/100) of diarrheic pigs (sampled at three farms) from the Caribbean nation of the Dominican Republic (DR). Although the sample size varied between porcine age groups, the highest PCV3 detection rates (35.3% each, respectively) were observed in piglets and growers. Co-infections with PCV2 and porcine adenovirus were observed in 38.09% and 9.52% of the PCV3 positive samples, respectively. The complete genomes of 11 DR PCV3 strains were analyzed in the present study, revealing a unique deletion (corresponding to nucleotide residue at position 1165 of reference PCV3 sequences) in one of the DR PCV3 sequences. Based on sequence identities and phylogenetic analysis (open reading frame 2 and complete genome sequences), the DR PCV3 strains were assigned to genotype PCV3a, and shared high sequence homologies (>98% identities) between themselves and with those of other PCV3a (Clade-1) strains, corroborating previous observations on the genetic stability of PCV3 worldwide. To our knowledge, this is the first report on the detection and molecular characterization of PCV3 in pigs from the Caribbean region, providing important insights into the expanding global distribution of the virus, even in isolated geographical regions (the Island of Hispaniola). Our findings warrant further investigations on the molecular epidemiology and economic implications of PCV3 in pigs with diarrhea and other clinical conditions across the Caribbean region.

Detection and Complete Genome Analysis of Porcine Circovirus 2 (PCV2) and an Unclassified CRESS DNA Virus from Diarrheic Pigs in the Dominican Republic: First Evidence for Predominance of PCV2d from the Caribbean Region.

We report here high rates (47.5%, 48/101) of detection of porcine circovirus 2 (PCV2) in diarrheic pigs from three pig farms in the Dominican Republic. Seventeen of the PCV2 positive samples, representing the three pig farms, different age groups and sampling periods (2020-2021), were amplified for the complete PCV2 genome. Based on analysis of open reading frame 2 and complete genome sequences, the 17 PCV2 strains were assigned to the PCV2d genotype. Significant differences were observed in PCV2 detection rates between the vaccinated (20% (10/50)) and unvaccinated (62.5% (10/16) and 80% (28/35)) farms, corroborating previous observations that PCV2a-based vaccines confer protection against heterologous PCV2 genotypes. The present study is the first to report detection and molecular characterization of PCV2 from the Dominican Republic, warranting large-scale molecular epidemiological studies on PCV2 in pig farms and backyard systems across the country. For the first time, PCV2d was identified as the predominant PCV2 genotype in a study from the Caribbean region, suggesting that a genotype shift from PCV2b to PCV2d might be happening in the Caribbean region, which mirrored the current PCV2 genotype scenario in many other parts of the world. Besides PCV2, we also identified a pigeon circovirus-like virus, and a circular Replication-associated protein (Rep)-encoding single-stranded (CRESS) DNA virus, which was characterized for the complete genome. The CRESS DNA virus shared a similar genomic organization and was related to unclassified CRESSV2 DNA viruses (belonging to the Order Cirlivirales) from porcine feces in Hungary, indicating that related unclassified CRESS DNA viruses are circulating among pigs in different geographical regions, warranting further studies on the epidemiology and biology of these novel viruses.

Novel Cyclovirus Species in Dogs with Hemorrhagic Gastroenteritis.

Nested PCRs with circovirus/cyclovirus pan-rep (replicase gene) primers detected eukaryotic circular Rep-encoding single-stranded DNA (CRESS DNA) viruses in three (samples CN9E, CN16E and CN34) of 18 canine parvovirus-2-positive fecal samples from household dogs with hemorrhagic gastroenteritis on the Caribbean island of Nevis. The complete genomes of CRESS DNA virus CN9E, CN16E and CN34 were determined by inverse nested PCRs. Based on (i) genome organization, (ii) location of the putative origin of replication, (iii) pairwise genome-wide sequence identities, (iv) the presence of conserved motifs in the putative replication-associated protein (Rep) and the arginine-rich region in the amino terminus of the putative capsid protein (Cp) and (v) a phylogenetic analysis, CN9E, CN16E and CN34 were classified as cycloviruses. Canine-associated cycloviruses CN16E and CN34 were closely related to each other and shared low genome-wide nucleotide (59.642-59.704%), deduced Rep (35.018-35.379%) and Cp (26.601%) amino acid sequence identities with CN9E. All the three canine-associated cycloviruses shared < 80% genome-wide pairwise nucleotide sequence identities with cycloviruses from other animals/environmental samples, constituting two novel species (CN9E and CN16E/34) within the genus Cyclovirus. Considering the feeding habits of dogs, we could not determine whether the cycloviruses were of dietary origin or infected the host. Interestingly, the CN9E putative Rep-encoding open reading frame was found to use the invertebrate mitochondrial genetic code with an alternative initiation codon (ATA) for translation, corroborating the hypothesis that cycloviruses are actually arthropod-infecting viruses. To our knowledge, this is the first report on the detection and complete genome analysis of cycloviruses from domestic dogs.

Phylogenetic analysis of G1P[6] group A rotavirus strains detected in Northeast Brazilian children fully vaccinated with Rotarix™.

In 2009 the World Health Organization recommended the use of group A rotavirus (RVA) vaccines in all national immunization programs (NIPs) in order to control severe RVA gastroenteritis disease. In Brazil, Rotarix™ was introduced in the NIP in March 2006, and a significant reduction in mortality rates among children ≤ 5 years old was observed, especially in the Northern and Northeastern Brazil. In the current study the 11 gene segments of six Brazilian G1P[6] RVA strains, isolated in 2009 and 2010 from vaccinated children, were analyzed in order to investigate if the genetic composition of these strains might help to elucidate why they were able to cause acute gastroenteritis in vaccinated children. All six Brazilian RVA strains revealed a complete Wa-like genotype constellation: G1-P[6]-I1-R1-C1-M1-A1-N1-T1-E1-H1. Phylogenetic analysis showed that all six strains were nearly identical and showed a close genetic relationship with contemporary typical human Wa-like RVA strains. These results suggests that the fact that these strains were able to cause acute gastroenteritis in vaccinated children is likely not due to the genetic background of the strains, but rather to other factors such as host relating factors, co-infecting pathogens or vaccine efficacy. P[6] RVA strains are detected rather occasionally in humans in most regions of the world, except for South Asia and Sub-Saharan Africa. However, recently two studies conducted in Brazil showed the circulation of G12P[6] and G2P[6]. This is the first report on the detection and complete genome analyses of G1P[6] RVA strains in Brazil. Surveillance studies will be crucial to further investigate the prevalence of this genotype in the Brazilian population, and the efficacy of current licensed vaccines, which do not contain the P[6] genotype.