Phenotypic characterization of swine peripheral blood monocyte-derived macrophages and ex vivo infection with Salmonella enterica serovar Typhimurium.

Macrophages are critical mediators of the inflammatory process, playing a relevant role in the pathogenesis of Salmonella Typhimurium. The protocols for isolation, culture, and differentiation of monocytes into macrophages and their interaction with Salmonella are well established in humans and murine models, but little information is available in swine. The aims of this study were to establish an efficient protocol for macrophage culture and to evaluate the interaction of the invA mutant strain and the wild type (WT) Salmonella Typhimurium with porcine macrophages. Peripheral blood monocyte-derived macrophages from pigs were obtained, separated by density-gradient centrifugation, and cultured in Teflon vials for 10 days. After the differentiation period, cultures consisted of 92.4% CD14+ cells. In addition, these cells showed phagocytic ability, demonstrated by the presence of the same amount of WT and invA mutant Salmonella Typhimurium 1 h after interaction with macrophages. The early cytotoxic effect was Salmonella pathogenicity island (SPI)-[1]dependent, in which log-phase WT strains were more efficient (p < 0.01) than the invA mutant strain at inducing the death of macrophages.

Genetic characterization of porcine circovirus type 2 in captive wild boars in southern Brazil.

Porcine circovirus type 2 (PCV2) has been identified in pig population in Brazil since 2000, but scarce studies involving wild boars with PCV2 infection are reported in the country. This study aimed to perform the genetic characterization of PCV2 detected in clinically healthy captive wild boars from farms located in Southern Brazil. Bronchial and mesenteric lymph nodes from 129 clinically healthy captive wild boars were tested by nested PCR for PCV2 detection. Six out of 38 positive samples (29.5%) were submitted to a quantitative real time PCR (qPCR) and genetic sequencing. Viral load up to 1.19 × 109 viral DNA copies/uL was detected in lymph nodes samples by qPCR. According to the ORF2 gene sequence analysis, all PCV2 samples were classified into PCV2b genotype. Comparisons based on a 702 nt region of the ORF2 of all six isolates revealed a high degree of similarity between these isolates. The ORF2 sequences characterized here share 97.1-100% of nucleotide identity and 95.7-100% of amino acid identity with other PCV2b isolated in Brazil from wild boars and feral pigs. This study reports the first detection and genetic characterization of PCV2b in captive wild boars in Brazil and provides important information on PCV2 infection in this domesticated species.

Primeira evidência da infecção pelo vírus da diarreia viral bovina em javalis / First evidence of bovine viral diarrhea virus infection in wild boars

Background: The farming of wild boars has growing due to the interest of the human consumption of this exotic meat. Such a development may pose an increased risk of disease transmission between boars and domestic animals. The wild boar population has increased in South America in the last years due the absence of predator causing economic losses due to direct damage to crops and risk of disease transmission. The genus Pestivirus within the family Flaviviridae are composed by four recognized species by the International Committee on the Taxonomy of Viruses (ICTV): classical swine fever virus (CSFV), border disease virus (BDV), bovine viral diarrhea virus type 1 (BVDV-1) and 2 (BVDV-2). Other putative species denoted as atypical pesitiviruses have been reported as ‘HoBi’-like virus, giraffe pestivirus, Bungowannah pestivirus, Pronghorn antelope virus, atypical porcine pestivirus (APPV), Norwegian rat pestivirus (NrPV) and Rhinolophus affinis bat pestivirus (RaPestV-1). CSFV is commonly detected in wild boars, but despite positive serology, bovine viral diarrhea virus (BVDV) was never detected in this animal species. Thereby, the present communication describes the first detection of BVDV in the lungs of captive boars using RT-PCR and DNA sequencing.Materials, Methods & Results: Forty lung samples from farmed wild boars were collected after slaughter in a commercial abattoir. The organs were crushed separately, centrifuged, and the supernatant was stored for further analysis. The total RNA was isolated using a phenol-based protocol and RT-PCR protocol that amplified 118 bp of 5’ untranslated region (5’UTR) was carried out. One out 40 samples resulted positive. The positive sample had partial fragments of 5’UTR and N terminal autoprotease (Npro) sequenced and analyzed. The strain LV Java/2012 presented 99% of identity in 5’UTR and 98% in Npro region with a BVDV-2 previously reported in bovines in Southern Brazil.[...]

First evidence of bovine viral diarrhea virus infection in wild boars / Primeira evidência da infecção pelo vírus da diarreia viral bovina em javalis

Background: The farming of wild boars has growing due to the interest of the human consumption of this exotic meat. Such a development may pose an increased risk of disease transmission between boars and domestic animals. The wild boar population has increased in South America in the last years due the absence of predator causing economic losses due to direct damage to crops and risk of disease transmission. The genus Pestivirus within the family Flaviviridae are composed by four recognized species by the International Committee on the Taxonomy of Viruses (ICTV): classical swine fever virus (CSFV), border disease virus (BDV), bovine viral diarrhea virus type 1 (BVDV-1) and 2 (BVDV-2). Other putative species denoted as atypical pesitiviruses have been reported as HoBi-like virus, giraffe pestivirus, Bungowannah pestivirus, Pronghorn antelope virus, atypical porcine pestivirus (APPV), Norwegian rat pestivirus (NrPV) and Rhinolophus affinis bat pestivirus (RaPestV-1). CSFV is commonly detected in wild boars, but despite positive serology, bovine viral diarrhea virus (BVDV) was never detected in this animal species. Thereby, the present communication describes the first detection of BVDV in the lungs of captive boars using RT-PCR and DNA sequencing.Materials, Methods & Results: Forty lung samples from farmed wild boars were collected after slaughter in a commercial abattoir. [...](AU)

Primeira evidência da infecção pelo vírus da diarreia viral bovina em javalis / First evidence of bovine viral diarrhea virus infection in wild boars

Background: The farming of wild boars has growing due to the interest of the human consumption of this exotic meat. Such a development may pose an increased risk of disease transmission between boars and domestic animals. The wild boar population has increased in South America in the last years due the absence of predator causing economic losses due to direct damage to crops and risk of disease transmission. The genus Pestivirus within the family Flaviviridae are composed by four recognized species by the International Committee on the Taxonomy of Viruses (ICTV): classical swine fever virus (CSFV), border disease virus (BDV), bovine viral diarrhea virus type 1 (BVDV-1) and 2 (BVDV-2). Other putative species denoted as atypical pesitiviruses have been reported as ‘HoBi-like virus, giraffe pestivirus, Bungowannah pestivirus, Pronghorn antelope virus, atypical porcine pestivirus (APPV), Norwegian rat pestivirus (NrPV) and Rhinolophus affinis bat pestivirus (RaPestV-1). CSFV is commonly detected in wild boars, but despite positive serology, bovine viral diarrhea virus (BVDV) was never detected in this animal species. Thereby, the present communication describes the first detection of BVDV in the lungs of captive boars using RT-PCR and DNA sequencing.Materials, Methods & Results: Forty lung samples from farmed wild boars were collected after slaughter in a commercial abattoir. The organs were crushed separately, centrifuged, and the supernatant was stored for further analysis. The total RNA was isolated using a phenol-based protocol and RT-PCR protocol that amplified 118 bp of 5 untranslated region (5UTR) was carried out. One out 40 samples resulted positive. The positive sample had partial fragments of 5UTR and N terminal autoprotease (Npro) sequenced and analyzed. The strain LV Java/2012 presented 99% of identity in 5UTR and 98% in Npro region with a BVDV-2 previously reported in bovines in Southern Brazil.[...](AU)

Associação entre fatores de virulência e resistência antimicrobiana de Escherichia coli enterotoxigênicas isoladas de leitões com diarreia no Brasil / Association between virulence factors and antimicrobial resistance from enterotoxigenic Escherichia coli isolated from pigs with diarrhea in Brazil

Background: Enterotoxigenic Escherichia coli (ETEC) is the etiologic agent of post weaning colibacillosis, one of the most important diseases in pig farming. The pathogenesis of the disease is associated with two virulence factors (VF), fimbriae and enterotoxins. In veterinary medicine, the use of antibiotics can lead to the selection of resistant bacteria. The association of VF and antibiotic resistance is an important mechanism for bacterial survival under adverse conditions. This study aimed to determine the VF and antimicrobial susceptibility of ETEC isolates from piglets with diarrhea and analyze the association between these factors.Materials, Methods & Results: A total of 185 rectal swabs were collected from weaned piglets in Brazilian farms of the states of Rio Grande do Sul, Santa Catarina, Paraná, Minas Gerais and Goiás. The isolation of ETEC was carried out on blood and MacConkey Agar and characterization by biochemical tests and detection by PCR of fimbrial genes F4, F45, F6, F18 and F41, and toxins genes LT, STa, STb and STx2e. Antimicrobial susceptibility profiles were determined by Agar diffusion test for amoxicillin, ampicillin, ceftiofur, ciprofloxacin, colistin, doxycycline, enrofloxacin, florfenicol, gentamicin, neomycin, norfloxacin, oxytetracycline, streptomycin, tetracycline, lincomycin + spectinomycin and sulfamethoxazole + trimethoprim. The association between VF and antimicrobials resistance results was determined by Chi-square and Fisher test (P ≤ 0.05). A total of 376 isolates were analyzed. The frequencies of fimbriae and toxins amplified were: F4 (31.6%), F18 (18.9%), F5 (4.2%) and toxins STa (43.1%), STb (24.7%), LT (21.8%) and STx2e (5.3%). Antibiotic resistance was higher to tetracycline (96.3%), florfenicol (95.2%), oxytetracycline (93.62%) and doxycycline (90.7%). Lowest levels of resistance were to ceftiofur (2.1%), colistin (9.8%), lincomycin + spectinomicin (15.4%) and neomycin (23.1%).[...](AU)

Associação entre fatores de virulência e resistência antimicrobiana de Escherichia coli enterotoxigênicas isoladas de leitões com diarreia no Brasil / Association between virulence factors and antimicrobial resistance from enterotoxigenic Escherichia coli isolated from pigs with diarrhea in Brazil

Background: Enterotoxigenic Escherichia coli (ETEC) is the etiologic agent of post weaning colibacillosis, one of the most important diseases in pig farming. The pathogenesis of the disease is associated with two virulence factors (VF), fimbriae and enterotoxins. In veterinary medicine, the use of antibiotics can lead to the selection of resistant bacteria. The association of VF and antibiotic resistance is an important mechanism for bacterial survival under adverse conditions. This study aimed to determine the VF and antimicrobial susceptibility of ETEC isolates from piglets with diarrhea and analyze the association between these factors.Materials, Methods & Results: A total of 185 rectal swabs were collected from weaned piglets in Brazilian farms of the states of Rio Grande do Sul, Santa Catarina, Paraná, Minas Gerais and Goiás. The isolation of ETEC was carried out on blood and MacConkey Agar and characterization by biochemical tests and detection by PCR of fimbrial genes F4, F45, F6, F18 and F41, and toxins genes LT, STa, STb and STx2e. Antimicrobial susceptibility profiles were determined by Agar diffusion test for amoxicillin, ampicillin, ceftiofur, ciprofloxacin, colistin, doxycycline, enrofloxacin, florfenicol, gentamicin, neomycin, norfloxacin, oxytetracycline, streptomycin, tetracycline, lincomycin + spectinomycin and sulfamethoxazole + trimethoprim. The association between VF and antimicrobials resistance results was determined by Chi-square and Fisher test (P ≤ 0.05). A total of 376 isolates were analyzed. The frequencies of fimbriae and toxins amplified were: F4 (31.6%), F18 (18.9%), F5 (4.2%) and toxins STa (43.1%), STb (24.7%), LT (21.8%) and STx2e (5.3%). Antibiotic resistance was higher to tetracycline (96.3%), florfenicol (95.2%), oxytetracycline (93.62%) and doxycycline (90.7%). Lowest levels of resistance were to ceftiofur (2.1%), colistin (9.8%), lincomycin + spectinomicin (15.4%) and neomycin (23.1%).[...]