Spleen and lung virome analysis of South American fur seals (Arctocephalus australis) collected on the southern Brazilian coast.

South American fur seals (Arctocephalus australis) are believed to reach the coast of Rio Grande do Sul (RS) through sea currents. They live in colonies and are frequently found resting on the beach. However, it is also common to find dead pinnipeds on beaches, sharing the environment with humans, domestic animals and other wild species on the coast and facilitating the transmission of pathogens. In the present study, a metagenomic approach was applied to evaluate the viral diversity in organs of fur seals found deceased along the coast of the state of RS, southern Brazil. The lungs and spleens of 29 animals were collected, macerated individually, pooled separately (one pool for lungs and another for spleens) and sequenced using the Illumina MiSeq platform. Sequences more closely related to members of the Anelloviridae and Circoviridae families were detected. Nine putative new species of anellovirus and one putative new genus, named Nitorquevirus, were described. Additionally, the circovirus sequences found in the lungs of A. australis have a common ancestor with PCV3, a proposed swine pathogen. Our study expanded the knowledge about viral communities in pinnipeds and could be useful for monitoring new viruses and potential viral sharing among wildlife, domestic animals, and humans.

Changes in the ceca microbiota of broilers vaccinated for coccidiosis or supplemented with salinomycin.

The objective of this study was to characterize differences in the cecal microbiota of chickens vaccinated for coccidiosis or receiving salinomycin in the diet. In this study, 140 male 1-day-old broiler chickens were divided in 2 groups: vaccine group (live vaccine) vaccinated at the first day and salinomycin group (125 ppm/kg since the first day until 35 d of age). Each treatment was composed for 7 replicates of 10 birds per pen. At 28 d, the cecal content of one bird per replicate was collected for microbiota analysis. The genetic sequencing was conducted by the Miseq Illumina platform. Vaccine group showed lower body weight, weight gain, and poorer feed conversion in the total period (P < 0.05). Bacterial 16S rRNA genes were classified as 3 major phyla (Bacteroidetes, Firmicutes, and Proteobacteria), accounting for more than 98% of the total bacterial community. The microbiota complexity in the cecal was estimated based on the α-diversity indices. The vaccine did not reduce species richness and diversity (P > 0.05). The richness distribution in the salinomycin group was larger and more uniform than the vaccinated birds. Salinomycin group was related to the enrichment of Bacteroidetes, whereas Firmicutes and Proteobacteria phyla were in greater proportions in the vaccine group. The last phylum includes a wide variety of pathogenic bacteria. The vaccine did not decrease the species richness but decreased the percentage of Bacteroidetes, a phylum composed by genera that produce short-chain fatty acids improving intestinal health. Vaccine group also had higher Proteobacteria phylum, which may help explain its poorer performance.

Novel Gyrovirus genomes recovered from free-living pigeons in Southern Brazil.

Wild birds carry a number of infectious agents, some of which may have pathogenic potential for the host and others species, including humans. Domestic pigeons (Columba livia) are important targets of study since these increasingly cohabit urban spaces, being possible spillover sources of pathogens to humans. In the present study, two genomes (PiGyV_Tq/RS/Br and PiGyV_RG/RS/Br), representative of Gyrovirus genus, family Anelloviridae, were detected in sera of free-living pigeons collected in Southern Brazil. The genomes exhibit less than 50% identity to previously described members of Gyrovirus genus, suggesting that they constitute a new viral species circulating in pigeons, to which the name "pigeon gyrovirus (PiGyV)" is proposed. The current study characterizes these two PiGyV genomes which, to date, are the first gyrovirus species identified in domestic pigeons.

Virome of crab-eating (Cerdocyon thous) and pampas foxes (Lycalopex gymnocercus) from southern Brazil and Uruguay.

Crab-eating (Cerdocyon thous) and Pampas foxes (Lycalopex gymnocercus) are wild canids distributed in South America. Domestic dogs (Canis lupus familiaris) and wild canids may share viral pathogens, including rabies virus (RABV), canine distemper virus (CDV), and canine parvovirus 2 (CPV-2). To characterize the virome of these wild canid species, the present work evaluated the spleen and mesenteric lymph node virome of 17 crab-eating and five Pampas foxes using high-throughput sequencing (HTS). Organ samples were pooled and sequenced using an Illumina MiSeq platform. Additional PCR analyses were performed to identify the frequencies and host origin for each virus detected by HTS. Sequences more closely related to the Paramyxoviridae, Parvoviridae and Anelloviridae families were detected, as well as circular Rep-encoding single-stranded (CRESS) DNA viruses. CDV was found only in crab-eating foxes, whereas CPV-2 was found in both canid species; both viruses were closely related to sequences reported in domestic dogs from southern Brazil. Moreover, the present work reported the detection of canine bocavirus (CBoV) strains that were genetically divergent from CBoV-1 and 2 lineages. Finally, we also characterized CRESS DNA viruses and anelloviruses with marked diversity. The results of this study contribute to the body of knowledge regarding wild canid viruses that can potentially be shared with domestic canids or other species.

Liver virome of healthy pigs reveals diverse small ssDNA viral genomes.

Brazil is a major exporter of pork meat worldwide. Swine liver is a common ingredient in food consumed by humans, thus emphasizing the importance of evaluating the presence of associated pathogens in swine liver. To obtain knowledge, this study aimed to provide insights into the viral communities of livers collected from slaughtered pigs from southern Brazil. The 46 livers were processed and submitted for high-throughput sequencing (HTS). The sequences were most closely related to Anelloviridae, Circoviridae and Parvoviridae families. The present work also describes the first Brazilian PCV1 and the first PPV6 and PPV7 from South America. Virus frequencies revelead 63% of samples positive for TTSuV1, 71% for TTSuVk2, 10.8% for PCV, 13% for PPV and 6% for PBov. This report addresses the diversity of the liver virome of healthy pigs and expands the number of viruses detected, further characterizing their genomes to assist future studies.

Highly divergent cattle hepacivirus N in Southern Brazil.

The genus Hepacivirus includes 14 species (Hepacivirus A-N). In this study, we determined a partial genome sequence of a highly divergent bovine hepacivirus (hepacivirus N, HNV) isolate from cattle in Southern Brazil. Previously described HNV isolates have shared 80-99.7% nucleotide sequence identity in the NS3 coding region. However, the sequence determined in this study had 72.6% to 73.8% nucleotide sequence identity to known HNV NS3 sequences. This high divergence could be seen in a phylogenetic tree, suggesting that it represents a new genotype of HNV. These data expand our knowledge concerning the genetic variability and evolution of hepaciviruses.

Characterization of the viral genomes present in commercial batches of horse serum obtained by high-throughput sequencing.

Horses are often used as blood donors for commercial horse serum (HS) production and to manufacture biologicals. HS is an alternative for fetal bovine serum (FBS) used as a supplement for cell culture and vaccine production. Furthermore, HS is also frequently obtained in order to produce antisera toxins and pathogens. The advent of high-throughput sequencing (HTS) has promoted changes in virus detection, since previous knowledge of targets is not required. Thus, the present study aimed to describe the virome of five different batches of commercial HS from New Zealand (three batches) and Brazil and the United States (one batch each) using HTS. Each HS pool were processed and sequenced using an Illumina MiSeq platform. Sequences-related to viruses belonging to the Flaviviridae, Herpesviridae, and Parvoviridae families were detected. Particularly, equine hepacivirus (EqHV), equine pegivirus (EPgV), and Theiler's disease-associated virus (TDAV) were more frequent found in the batches analyzed. The presence of viral genomes in cell culture sera illustrates that these commercial sera can contain a mixture of different viruses and, therefore, can be regarded as potentially infectious for susceptible hosts. Moreover, the innocuity of commercial HS is important for the efficiency and security of diagnostics and the production of biological products.

Correction to: Columbid circoviruses detected in free ranging pigeons from Southern Brazil: insights on PiCV evolution.

Unfortunately, the word "evolution" was found missing in title of the original article which is corrected here by this erratum. The original article has been corrected.

Columbid circoviruses detected in free ranging pigeons from Southern Brazil: insights on PiCV evolution.

Pigeon circovirus (PiCV) is taxonomically classified as a member of the Circovirus genus, family Circoviridae. The virus contains a single stranded DNA genome of approximately 2 kb, with minor length variations among different isolates. The occurrence of PiCV infections in pigeons (Columba livia) has been documented worldwide over the past 20 years; however, in Brazil there were still no reports on PiCV detection. This study identifies seven PiCV genomes recovered from domestic pigeons of South Brazil through high-throughput sequencing and shows a high frequency of PiCV infection, through quantitative real-time PCR. Phylogenetic classification was performed by maximum likelihood analysis of the full genomes, ORF V1 (Rep) and ORF C1 (Cap). The results show that either full genome or Cap based analysis allowed PiCV classification into five major clades (groups A to E), where Brazilian sequences were classified as A, C or D. Recombination analyses were carried out with Simplot and RDP4 and the results show that both Rep and Cap ORFs contain several recombination hotspots, pointing to an important role for such events in PiCV evolution.

Full-Genome Sequence of Porcine Circovirus type 3 recovered from serum of sows with stillbirths in Brazil.

Two full-genome sequences of porcine circovirus type 3 (PCV3) are reported. The genomes were recovered from pooled serum samples from sows who had just delivered litters with variable numbers of stillbirths. The two circular genomes (PCV3-BR/RS/6 and PCV3-BR/RS/8) are 2,000 nucleotides long and contain two open reading frames (ORFs) oriented in opposite directions that encode the putative capsid (Cap) and replicase (Rep) proteins. The intergenic region contains a stem-loop motif, as reported for other circoviruses. Rolling circle replication motifs and putative helicase domains were identified in the Rep coding region. The degree of overall nucleotide similarity between the genomes reported here and those available at GenBank was higher than 97%. No PCV3 sequence was detected in pooled serum samples from sows which had no stillbirths on the same farms. However, further studies are necessary to confirm the association between PCV3 and the occurrence of stillbirths.

Swine Influenza Virus and Association with the Porcine Respiratory Disease Complex in Pig Farms in Southern Brazil.

Despite the putative endemic status of swine influenza A virus (swIAV) infections, data on the occurrence of swine influenza outbreaks are scarce in Brazil. The aim of this study was to detect and subtype swIAVs from six outbreaks of porcine respiratory disease complex (PRDC) in southern Brazil. Nasal swabs were collected from 66 piglets with signs of respiratory disease in six herds. Lung tissue samples were collected from six necropsied animals. Virus detection was performed by PCR screening and confirmed by virus isolation and hemagglutination (HA). Influenza A subtyping was performed by a real-time reverse transcriptase PCR (rRT-PCR) to detect the A(H1N1)pdm09; other swIAV subtypes were determined by multiplex RT-PCR. In lung tissues, the major bacterial and viral pathogens associated with PRDC (Pasteurella multocida, Mycoplasma hyopneumoniae, Actinobacillus pleuropneumoniae, Haemophilus parasuis and PCV2) were investigated. In some affected pigs, clinico-pathological evaluations were conducted. Influenza A was detected by screening PCR in 46 of 66 swab samples and from five of six lungs. Virus was recovered from pigs of all six herds. Subtype A(H1N1)pdm09 was detected in four of six herds and H1N2 in the other two herds. In lung tissues, further agents involved in PRDC were detected in all cases; Pasteurella multocida was identified in five of six samples and Mycoplasma hyopneumoniae in three of six. Actinobacillus pleuropneumoniae (1/6), Haemophilus parasuis (1/6) and PCV2 (1/6) were also detected. These findings indicate that subtypes A(H1N1)pdm09 and H1N2 were present in pigs in southern Brazil and were associated with PRDC outbreaks.

Neutralizing antibodies to bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) and its subtypes.

This study was carried out to determine whether the sensitivity of serum neutralization (SN) tests would be affected by the use of distinct subtypes of bovine herpesvirus 1 (BoHV-1) and 5 (BoHV-5) as test challenge viruses. Bovine sera collected from a randomized sample (n=287) were tested in a 24h incubation SN against three type 1 viruses (BoHV-1.1 strains "Los Angeles" (LA) and "EVI 123"; BoHV-1.2a strain "SV 265") and three type 5 viruses (BoHV-5a strain "EVI 88"; BoHV-5b strain "A 663" and BoHV-5c "ISO 97"). SN sensitivity varied greatly depending on the test challenge virus used in the test, particularly when results against each virus were considered individually, where it ranged from 77% (detecting 80 out of 104 antibody-positive sera) with ISO 97 to 91% (95/104) with BoHV-1.1 strain LA. All tests to single viruses revealed a significantly low sensitivity (McNemar's; p<0.05). Maximum sensitivity (104/104) was achieved when positive results to a particular combination of four of the challenge viruses (LA+EVI 123+SV 265+A 663) or some combinations of five viruses (or all six viruses) were added cumulatively. These results provide evidence for no association between any particular virus type/subtype and higher SN sensitivity. In addition, it was clearly shown that when SN is performed with single test challenge viruses, sensitivity can vary so significantly that might compromise control or eradication efforts. Performing SN against a number of different viruses demonstrated to improve significantly the test's sensitivity.