First detection of PCV4 in swine in the United States: codetection with PCV2 and PCV3 and direct detection within tissues.

Since PCV4 was first described in 2019, the virus has been identified in several countries in Southeast Asia and Europe. Most studies have been limited to detecting PCV4 by PCR. Thus, PCV4 has an unclear association with clinical disease. This study utilized 512 porcine clinical lung, feces, spleen, serum, lymphoid tissue, and fetus samples submitted to the ISU-VDL from June-September 2023. PCV4 was detected in 8.6% of samples with an average Ct value of 33. While detection rates among sample types were variable, lymphoid tissue had the highest detection rate (18.7%). Two ORF2 sequences were obtained from lymphoid tissue samples and had 96.36-98.98% nucleotide identity with reference sequences. Direct detection of PCV4 by RNAscope revealed viral replication in B lymphocytes and macrophages in lymph node germinal centers and histiocytic and T lymphocyte infiltration in the lamina propria of the small intestine. PCV4 detection was most commonly observed in nursery to finishing aged pigs displaying respiratory and enteric disease. Coinfection with PCV2, PCV3, and other endemic pathogens was frequently observed, highlighting the complex interplay between different PCVs and their potential roles in disease pathogenesis. This study provides insights into the frequency of detection, tissue distribution, and genetic characteristics of PCV4 in the US.

Infection and Coinfection of Porcine-Selected Viruses (PPV1 to PPV8, PCV2 to PCV4, and PRRSV) in Gilts and Their Associations with Reproductive Performance.

Seven novel porcine parvoviruses (nPPVs) (PPV2 through PPV8) have been described, although their pathogenicity and possible effects on porcine reproductive failure (PRF) are undefined. In this study, these nPPVs were assessed in gilts from Colombia; their coinfections with PPV1, PCV2, PCV3, PCV4, and PRRSV and an association between the nPPVs and the reproductive performance parameters (RPPs) in sows were determined. For this, 234 serum samples were collected from healthy gilts from 40 herds in five Colombian regions, and the viruses were detected via real-time PCR. The results confirmed the circulation of PPV2 through PPV7 in Colombia, with PPV3 (40%), PPV5 (20%), and PPV6 (17%) being the most frequent. Additionally, no PCV4 or PPV8 was detected. PPV2 to PPV7 were detected in concurrence with each other and with the primary PRF viruses, and these coinfections varied from double to sextuple coinfections. Additionally, the association between nPPVs and PRF primary viruses was statistically significant for the presence of PPV6 in PCV3-positive (p < 0.01) and PPV5 in PPRSV-positive (p < 0.05) gilts; conversely, there was a significant presence of PPV3 in both PCV2-negative (p < 0.01) and PRRSV-negative (p < 0.05) gilts. Regarding the RPPs, the crude association between virus detection (positive or negative) and a high or low RPP was only statistically significant for PCV3 and the farrowing rate (FR), indicating that the crude odds of a low FR were 94% lower in herds with PCV3-positive gilts. This finding means that the detection of PCV3 in gilts (PCV3-positive by PCR) is associated with a higher FR in the farm or that these farms (with positive gilts) have lower odds (OR 0.06, p-value 0.0043) of a low FR. Additionally, a low FR tended to be associated with the detection of PPV4 and PPV5 (p-value < 0.20). This study is important for establishing the possible participation of nPPVs in PRF.

The Novel Porcine Parvoviruses: Current State of Knowledge and Their Possible Implications in Clinical Syndromes in Pigs.

Parvoviruses (PVs) affect various animal species causing different diseases. To date, eight different porcine parvoviruses (PPV1 through PPV8) are recognized in the swine population, all of which are distributed among subfamilies and genera of the Parvoviridae family. PPV1 is the oldest and is recognized as the primary agent of SMEDI, while the rest of the PPVs (PPV2 through PPV8) are called novel PPVs (nPPVs). The pathogenesis of nPPVs is still undefined, and whether these viruses are putative disease agents is unknown. Structurally, the PPVs are very similar; the differences occur mainly at the level of their genomes (ssDNA), where there is variation in the number and location of the coding genes. Additionally, it is considered that the genome of PVs has mutation rates similar to those of ssRNA viruses, that is, in the order of 10-5-10-4 nucleotide/substitution/year. These mutations manifest mainly in the VP protein, constituting the viral capsid, affecting virulence, tropism, and viral antigenicity. For nPPVs, mutation rates have already been established that are similar to those already described; however, within this group of viruses, the highest mutation rate has been reported for PPV7. In addition to the mutations, recombinations are also reported, mainly in PPV2, PPV3, and PPV7; these have been found between strains of domestic pigs and wild boars and in a more significant proportion in VP sequences. Regarding affinity for cell types, nPPVs have been detected with variable prevalence in different types of organs and tissues; this has led to the suggestion that they have a broad tropism, although proportionally more have been found in lung and lymphoid tissue such as spleen, tonsils, and lymph nodes. Regarding their epidemiology, nPPVs are present on all continents (except PPV8, only in Asia), and within pig farms, the highest prevalences detecting viral genomes have been seen in the fattener and finishing groups. The relationship between nPPVs and clinical manifestations has been complicated to establish. However, there is already some evidence that establishes associations. One of them is PPV2 with porcine respiratory disease complex (PRDC), where causality tests (PCR, ISH, and histopathology) lead to proposing the PPV2 virus as a possible agent involved in this syndrome. With the other nPPVs, there is still no clear association with any pathology. These have been detected in different systems (respiratory, reproductive, gastrointestinal, urinary, and nervous), and there is still insufficient evidence to classify them as disease-causing agents. In this regard, nPPVs (except PPV8) have been found to cause porcine reproductive failure (PRF), with the most prevalent being PPV4, PPV6, and PPV7. In the case of PRDC, nPPVs have also been detected, with PPV2 having the highest viral loads in the lungs of affected pigs. Regarding coinfections, nPPVs have been detected in concurrence in healthy and sick pigs, with primary PRDC and PRF viruses such as PCV2, PCV3, and PRRSV. The effect of these coinfections is not apparent; it is unknown whether they favor the replication of the primary agents, the severity of the clinical manifestations, or have no effect. The most significant limitation in the study of nPPVs is that their isolation has been impossible; therefore, there are no studies on their pathogenesis both in vitro and in vivo. For all of the above, it is necessary to propose basic and applied research on nPPVs to establish if they are putative disease agents, establish their effect on coinfections, and measure their impact on swine production.

Canine circovirus: An emerging or an endemic undiagnosed enteritis virus?

Canine Circovirus (CanineCV) belongs to the family Circoviridae. It is an emerging virus described for the first time in 2011; since then, it has been detected in different countries and can be defined as worldwide distribution virus. CanineCV infects domestic and wild canids and is mainly related to hemorrhagic enteritis in canines. However, it has been identified in fecal samples from apparently healthy animals, where in most cases it is found in coinfection with other viral agents such as the canine parvovirus type-2 (CPV). The estimated prevalence/frequency of CanineCV has been variable in the populations and countries where it has been evaluated, reaching from 1 to 30%, and there are still many concepts to define the epidemiological characteristics of the virus. The molecular characterization and phylo-evolutive analyses that allow to postulate the wild origin and intercontinental distribution of the virus. This review focuses on the importance on continuing research and establish surveillance systems for this emerging virus.

The Prevalence and Genetic Diversity of PCV3 and PCV2 in Colombia and PCV4 Survey during 2015-2016 and 2018-2019.

Four genotypes of circovirus have been recognized in swine, with PCV2 and PCV3 being the most associated with clinical manifestations, while PCV4 does not have a defined disease. In addition, PCV2 is associated with different syndromes grouped as diseases associated with porcine circovirus (PCVAD), while PCV3 causes systemic and reproductive diseases. In the present study, we retrospectively detected PCV2, PCV3, and PCV4 in Colombia during two periods: A (2015-2016) and B (2018-2019). During period A, we evaluated stool pools from the 32 Colombian provinces, finding a higher prevalence of PCV3 compared to PCV2 as well as PCV2/PCV3 co-infection. Furthermore, we determined that PCV3 had been circulating since 2015 in Colombia. Regarding period B, we evaluated sera pools and tissues from abortions and stillborn piglets from the five provinces with the highest pig production. The highest prevalence found was for PCV3 in tissues followed by sera pools, while PCV2 was lower and only in sera pools. In addition, PCV2/PCV3 co-infection in sera pools was also found for this period. The complete genome sequences of PCV3 and PCV3-ORF2 placed the Colombian isolates within clade 1 as the majority in the world. For PCV2, the predominant genotype currently in Colombia is PCV2d. Likewise, in some PCV3-ORF2 sequences, a mutation (A24V) was found at the level of the Cap protein, which could be involved in PCV3 immunogenic recognition. Regarding PCV4, retrospective surveillance showed that there is no evidence of the presence of this virus in Colombia.

Evidence of bovine leukemia virus circulating in sheep and buffaloes in Colombia: insights into multispecies infection.

Bovine leukemia virus (BLV) is the causative agent of leukemia/lymphoma in cattle. However, previous evidence has shown its presence in other species of livestock as well as in humans, suggesting that other species can be accidental hosts of the virus. In viral infections, receptors that are common to different animal species are proposed to be involved in cross-species infections. For BLV, AP3D1 has been proposed to be its receptor, and this protein is conserved in most mammalian species. In Colombia, BLV has been reported in cattle with high prevalence rates, but there has been no evidence of BLV infections in other animal species. In this study, we tested for the virus in sheep (n = 44) and buffaloes (n = 61) from different regions of Colombia by nested PCR, using peripheral blood samples collected from the animals. BLV was found in 25.7% of the animals tested (12 buffaloes and 15 sheep), and the results were confirmed by Sanger sequencing. In addition, to gain more information about the capacity of the virus to infect these species, the predicted interactions of AP3D1 of sheep and buffaloes with the BLV-gp51 protein were analyzed in silico. Conserved amino acids in the binding domains of the proteins were identified. The detection of BLV in sheep and buffaloes suggests circulation of the virus in multiple species, which could be involved in dissemination of the virus in mixed livestock production settings. Due to the presence of the virus in multiple species and the high prevalence rates observed, integrated prevention and control strategies in the livestock industry should be considered to decrease the spread of BLV.

The first report of porcine parvovirus 7 (PPV7) in Colombia demonstrates the presence of variants associated with modifications at the level of the VP2-capsid protein.

There are a wide variety of porcine parvoviruses (PPVs) referred to as PPV1 to PPV7. The latter was discovered in 2016 and later reported in some countries in America, Asia, and Europe. PPV7 as a pathogenic agent or coinfection with other pathogens causing disease has not yet been determined. In the present study, we report the identification of PPV7 for the first time in Colombia, where it was found retrospectively since 2015 in 40% of the provinces that make up the country (13/32), and the virus was ratified for 2018 in 4/5 provinces evaluated. Additionally, partial sequencing (nucleotides 380 to 4000) was performed of four Colombian strains completely covering the VP2 and NS1 viral genes. A sequence identity greater than 99% was found when comparing them with reference strains from the USA and China. In three of the four Colombian strains, an insertion of 15 nucleotides (five amino acids) was found in the PPV7-VP2 capsid protein (540-5554 nt; 180-184 aa). Based on this insertion, the VP2 phylogenetic analysis exhibited two well-differentiated evolutionarily related groups. To evaluate the impact of this insertion on the structure of the PPV7-VP2 capsid protein, the secondary structure of two different Colombian strains was predicted, and it was determined that the insertion is located in the coil region and not involved in significant changes in the structure of the protein. The 3D structure of the PPV7-VP2 capsid protein was determined by threading and homology modeling, and it was shown that the insertion did not imply a change in the shape of the protein. Additionally, it was determined that the insertion is not involved in suppressing a potential B cell epitope, although the increase in length of the epitope could affect the interaction with molecules that allow a specific immune response.

Field infection of a gilt and its litter demonstrates vertical transmission and effect on reproductive failure caused by porcine circovirus type 3 (PCV3).

BACKGROUND: PCV3 is a member of the Circovirus family, associated with disease and mortality in pigs. It is not clear whether PCV3 putatively causes clinical symptoms and disease. In the present case, we reported a gilt infected with PCV3 associated with reproductive failures, vertical transmission, tissue lesions, viral replication by in situ hybridization, and the hypothesis that some strains of PCV3 clade one are associated with reproductive failures at the field level. CASE PRESENTATION: In May 2019, a pig farm in Colombia reported increased reproductive failures, and the presence of PCV3 in gilts and sows was established in a single form or coinfections, mainly with PCV2 and PPV7. Ten sows with a single infection with PCV3 were found, and one gilt with a pre-farrowing serum viral load above 103 was studied. This gilt was followed up during the pre-farrowing, farrowing period and on her litter for 6 weeks. During dystocic farrowing, a mummy and ten piglets were released, including two weak-born piglets. The highest viral loads for PCV3 were found in the mummy and the placenta. In the weak-born piglets, there were viral loads both in serum and in tissues, mainly in the mesenteric ganglia and lung. Replication of PCV3 in these tissues was demonstrated by in situ hybridizations. PCV3 was also found in the precolostrum sera of piglets and colostrum, showing vertical transmission. The viral load in piglets decreased gradually until week six of life. The viral genome's complete sequencing was made from the mummy, and its analysis classified it as PCV3 clade one. CONCLUSIONS: This report confirms that PCV3 can cause disease at the field level, and putatively, in this case, we find the generation of reproductive failures. The ability of PCV3 to cause disease as a putative pathogen may be associated with the viral load present in the pig and the strain that is affecting the farm. For this case, we found that viral loads above 103 (4.93 log genomic copies / mL) in the gilt were associated with clinical manifestation and that some PCV3 strains belonging to clade one are more associated with the reproductive presentation.

First detection and full genomic analysis of Canine Circovirus in CPV-2 infected dogs in Colombia, South America.

Canine Circovirus (CanineCV) is an emerging virus which since its first report in USA in 2012, it has been described worldwide. It was the second mammalian circovirus species identified in dogs and its role in canine enteritis is still being uncertain as much as its association in disease with the Canine Parvovirus-2 (CPV-2). Here, we aim to confirm for the first time the presence of CanineCV in Colombia and to develop phylogenetic evolutive analyses of CanineCV in CPV-2 positive animals. DNA from samples were extracted and PCR, full genome sequencing and phylogenetic analysis was performed to detect and characterize CanineCV. From a total of 30 CPV-2 positive samples, 16.6% (n = 5) were positives for CanineCV. Sequencing analysis of Colombian CanineCV wild-type strains displayed high identity to each other (99.5-99.7% nt; 99.7% aa). The full genome phylogenetic analysis confirmed that worldwide reported CanineCV strains were separated into four distinct genotypes in addition to a European origin of the South American CanineCV strains. This study demonstrated the importance of continue surveillance of emerging viruses in canine populations and confirm for the first time the circulation and origin of CanineCV in Colombia.

First detection of porcine circovirus type 3 in Colombia and the complete genome sequence demonstrates the circulation of PCV3a1 and PCV3a2.

Porcine circovirus 3 (PCV3) was recently discovered and is a new species of the genus circovirus. Clinically, it is associated with absence of symptoms or with different clinical syndromes. It has been reported in different countries of America, Europe and Asia. Last year, in Colombia, some farms have reported symptoms similar to those caused by PCV2. Samples were taken from two farms located in the centre of the country, and the presence of PCV3 was determined by PCR in two samples, one from a pool of sera and another from mesenteric lymph node. The strains were fully sequenced (GenBank accession numbers MH327784 and MH327785) and classified into subgroups a1 and a2. According to this classification and its analysis, strain a2 is located within the group called "Linker" that may be evolving towards group "b". In addition to the above, the two Colombian strains were compared with 104 strains reported in the GenBank database. The phylogenetic tree obtained grouped according to the classification of subgroups a1, a2, b1 and b2. It was found that subgroups a1 and a2 were well grouped when comparing whole genomes, but the same was not observed with the strains of group "b". In the latter, no subgroups were evidenced when comparing complete genomes. It is suggested that a new classification of PCV3 subgroups should be proposed, based on whole genome sequences. This is the first report of PCV3 in Colombia and its complete genome sequence.

Longitudinal comparison of the humoral immune response and viral load of Porcine Circovirus Type 2 in pigs with different vaccination schemes under field conditions.

Background: Porcine Circovirus type 2 (PCV2) infections are distributed worldwide and cause Porcine Circovirus Associated Disease (PCVAD). To minimize the impact of PCV2 infection on swine health and production, different vaccination schemes have been used since 2006. However, the association between vaccination schemes, virus load and disease under field conditions are not completely understood. Therefore, the objective of this study was to compare the effect of two different PCV2 vaccination schemes on the humoral response and PCV2 load in pigs after weaning under field conditions. Methods: Two commercial pig farms (Farm A and B), endemically infected with PCV2, which were using two different PCV2 subunit vaccinations schemes for sow, gilts and piglets, were selected. We designed a longitudinal study and measured IgG levels by ELISA and virus load by quantitative PCR in pigs after weaning. Forty 3-week old piglets were randomly selected at weaning and followed for 20 weeks. IgG levels and virus loads were compared within and between farms and considered statistically different if the non-parametric Kruskal Wallis test p value was lower than 0.05. Results: We found that low virus loads were maintained in pigs from both farms regardless of the vaccination scheme used (p>0.05). However, there was significant difference in the mean IgG levels observed over time (p<0.05), suggesting that different humoral immune response are not necessarily associated with different virus loads observed over time. Conclusions: These results are important because they can help to prevent PCV2 infections using different vaccination schemes to minimize the effect of PCVAD on swine health and production.

Molecular characterization of rotaviruses isolated from calves with bovine neonatal diarrhea (BND) in Colombia / Caracterizacion molecular de rotavirus aislados a partir de terneros con diarrea neonatal bovina (dnb) en colombia

Bovine rotaviruses are one of the main agents involved in the presentation of Bovine Neonatal Diarrhea (BND), a disease that affects calves in the first month of life. Objective: The present study aimed to determine the types of bovine rotaviruses that affect dairy herds in the Sabana region of Bogotá, Colombia. Materials and methods: 132 fecal samples were obtained from calves of less than five weeks of age and subjected to an ELISA test. MA104 cell cultures were infected with positive samples in order to isolate rotaviruses. The presence of the viral genome was confirmed by amplification and sequencing of a region of the viral VP7 protein-encoding gene. Results: Of the 132 samples, 26 (19, 7%) were ELISA-positive and nine samples were used for viral isolation. PCR amplification was achieved in all infected cultures. Sequencing showed homology of five samples to the G6 genotype. In addition, the presence of the G10 genotype was first determined for the country. Discussion: A greater presence of the G6 genotype from lineage V was found in the Sabana region of Bogota, showing a high prevalence in cattle and association with the presence of BND. The presence of the G10 genotype is a new report for the country and constitutes a new element of investigation in these viruses.

Los rotavirus bovinos son unos de los principales agentes involucrados en la presentación del síndrome de Diarrea Neonatal Bovina (DNB), una enfermedad que afecta terneros en el primer mes de vida. Objetivo: El presente estudio buscó determinar los tipos de rotavirus que afectan los hatos ganaderos de leche en la región de la Sabana de Bogotá, Colombia. Materiales y métodos: Se evaluaron a través de una prueba de ELISA, 132 muestras de materia fecal provenientes de terneros de menos de cinco semanas de edad. Con las muestras positivas, se infectaron células MA 104 con el fin hacer aislamiento. La presencia del genoma viral se verificó por amplificación de una región del gen que codifica para la proteína viral VP-7 y luego se secuenció. Resultados: De las 132 muestras evaluadas, 26 (19,7%) fueron positivas por ELISA. De estas, 9 muestras se emplearon para aislamiento viral. La amplificación de genoma viral por PCR se obtuvo en todos los cultivos infectados. La secuenciación evidenció una homología de 5 muestras con el genotipo G6 y la presencia del genotipo G10, que se encontró por primera vez en el país. Discusión: En la Sabana de Bogotá se encontró una mayor presencia del genotipo G6, linaje V, que tiene alta prevalencia en bovinos y está asociado mayoritariamente con la presencia de DNB. La presencia del genotipo G10 constituye un elemento nuevo de investigación en estos virus.