Molecular characterization of Salmonella spp. and Listeria monocytogenes strains from biofilms in cattle and poultry slaughterhouses located in the federal District and State of Goiás, Brazil.

Listeria monocytogenes and Salmonella spp. are considered important foodborne pathogens that are commonly associated with foods of animal origin. The aim of this study was to perform molecular characterization of L. monocytogenes and Salmonella spp. isolated from biofilms of cattle and poultry slaughterhouses located in the Federal District and State of Goiás, Brazil. Fourteen L. monocytogenes isolates and one Salmonella sp. were detected in poultry slaughterhouses. No isolates were detected in cattle slaughterhouses. All L. monocytogenes isolates belonged to lineage II, and 11 different pulsotypes were detected. Pulsed-field gel electrophoresis analysis revealed the dissemination of two strains within one plant, in addition to the regional dissemination of one of them. The Salmonella isolate was identified via whole genome sequencing as Salmonella enterica serovar Minnesota ST548. In the sequence analysis, no premature stop codons were detected in the inlA gene of Listeria. All isolates demonstrated the ability to adhere to Caco-2 cells, while 50% were capable of invading them. Antimicrobial resistance was detected in 57.1% of the L. monocytogenes isolates, and resistance to sulfonamide was the most common feature. The tetC, ermB, and tetM genes were detected, and four isolates were classified as multidrug-resistant. Salmonella sp. was resistant to nine antimicrobials and was classified as multidrug-resistant. Resistance genes qnrB19, blaCMY-2, aac(6')-Iaa, sul2, and tetA, and a mutation in the parC gene were detected. The majority (78.5%) of the L. monocytogenes isolates were capable of forming biofilms after incubation at 37°C for 24 h, and 64.3% were capable of forming biofilms after incubation at 12°C for 168 h. There was no statistical difference in the biofilm-forming capacity under the different evaluated conditions. Salmonella sp. was capable of forming biofilms at both tested temperatures. Biofilm characterization was confirmed by collecting the samples consistently, at the same sampling points, and by assessing biofilm formation in vitro. These results highlight the potential risk of cross-contamination in poultry slaughterhouses and the importance of surveillance and pathogen control maintenance programs within the meat production industry.

Assessment of Internalin A Gene Sequences and Cell Adhesion and Invasion Capacity of Listeria monocytogenes Strains Isolated from Foods of Animal and Related Origins.

Listeria monocytogenes is a foodborne pathogen of global relevance that causes outbreaks and sporadic cases of listeriosis, acquired through the consumption of contaminated products, including milk or meat products and ready-to-eat meat products subjected to intensive handling. The objective of the present study was to classify L. monocytogenes isolated from various food-related sources in the Federal District of Brazil and surrounding areas to sequence internalin A (inlA) genes from these isolates and assess their adhesion and invasion capacity using Caco-2 cells. In addition, 15 were classified as group I, 3 as group II, and 7 classified as group IV. Premature stop codons (PMSCs) at the nucleotide position 976 (GAA→TAA) of the inlA gene were identified in 5 of the 25 isolates. Adhesion and invasion tests in Caco-2 cells showed that all the isolates were capable of adhesion and cellular invasion, with isolates containing PMSCs exhibiting on average higher invasion capacity than those without PMSCs (p = 0.041) and a median of adhesion very distinctive from those without stop codons. These results are the first report of PMSCs in the inlA gene of L. monocytogenes from the Federal District of Brazil and Brazil.

Isolamento de Listeria monocytogenes em salsichas comercializadas no Distrito Federal / Isolation of Listeria monocytogenesin sausages commercialized in the Distrito Federal

A bactéria Listeria monocytogenes é o patógeno causador da listeriose, importante doença transmitida por alimentos que acomete humanos e animais podendo causar morte. Foram coletadas 100 amostras de salsicha de mercados do Distrito Federal. A metodologia usada para o isolamento foi a descrita na Instrução Normativa Nº 40 do Ministério da Agricultura, Pecuária e Abastecimento. As amostras que apresentaram motilidade característica de Listeria sp p. foram submetidas a técnica de PCR para confirmação da espécie L. monocytogenes. Os resultados positivos para Listeria sp p. foram vistos em 14 amostras e em 5 confirmou-se a presença de L. monocytogenes. Através dos resultados desse experimento conclui-se que salsichas podem estar contaminadas com bactérias do gênero Listeria sp p. e L. monocytogenes, oferecendo risco de listeriose aos consumidores.

A retrospective search for bovine respiratory syncytial virus (BRSV) antigens in histological specimens by immunofluorescence and immunohistochemistry

Bovine respiratory syncytial virus (BRSV) has been only sporadically identified as a causative agent of respiratory disease in Brazil. This contrasts with frequent reports of clinical and histopathological findings suggestive of BRSV-associated disease. In order to examine a possible involvement of BRSV in cases of calf pneumonia, a retrospective search was performed for BRSV antigens in histological specimens submitted to veterinary diagnostic services from the states of Rio Grande do Sul and Minas Gerais. Ten out of 41 cases examined (24.4 percent) were positive for BRSV antigens by immunohistochemistry (IPX). Eight of these cases (19.5 percent) were also positive by indirect immunofluorescence (IFA), and 31 cases (75.6 percent) were negative in both assays. In the lungs, BRSV antigens were predominantly observed in epithelial cells of bronchioles and less frequently found in alveoli. In one case, antigens were detected only in the epithelium of the alveolar septae. The presence of antigen-positive cells was largely restricted to epithelial cells of these airways. In two cases, positive staining was also observed in cells and cellular debris in the exudate within the pulmonary airways. The clinical cases positive for BRSV antigens were observed mainly in young animals (2 to 12 month-old) from dairy herds. The main microscopic changes included bronchointerstitial pneumonia characterized by thickening of alveolar septae adjacent to airways by mononuclear cell infiltrates, and the presence of alveolar syncytial giant cells. In summary, the results demonstrate the suitability of the immunodetection of viral antigens in routinely fixed tissue specimens as a diagnostic tool for BRSV infection. Moreover, the findings provide further evidence of the importance of BRSV as a respiratory pathogen of young cattle in southeastern and southern Brazil.

O Vírus Respiratório Sincicial Bovino (BRSV) tem sido raramente identificado como agente etiológico de doença respiratória em bovinos no Brasil. Isso contrasta com freqüentes relatos clínicos e histopatológicos sugestivos de enfermidade associada ao BRSV. Com o objetivo de investigar o possível envolvimento do BRSV em casos de pneumonia em bovinos jovens, realizou-se uma pesquisa retrospectiva para antígenos do BRSV em cortes histológicos de materiais submetidos a quatro serviços de diagnóstico nos Estados do Rio Grande do Sul e Minas Gerais. Dez entre 41 casos examinados (24,4 percent) foram positivos para antígenos do BRSV por imunohistoquímica (IPX). Oito desses casos (19,5 percent) também foram positivos por imunofluorescência (IFA) e 31 casos (75,6 percent) foram negativos em ambos os testes. Nos pulmões, antígenos virais foram detectados predominantemente nas células epiteliais dos bronquíolos e menos freqüentemente nos alveólos. Em um caso, antígenos virais foram detectados exclusivamente no epitélio alveolar. A presença de células positivas restringiu-se ao epitélio dessas vias aéreas. Em dois casos, antígenos virais foram detectados em células descamativas e em restos celulares no exsudato das vias aéreas. Os casos positivos para antígenos do BRSV eram oriundos principalmente de animais jovens (2 a 12 meses de idade) de rebanhos leiteiros. As principais alterações histológicas observadas foram pneumonia bronco-intersticial caracterizada por espessamento dos septos alveolares adjacentes às vias aéreas, infiltrados de células mononucleares e presença de células gigantes multinucleadas nos alveólos. Os resultados obtidos demonstram a utilidade dos métodos de detecção de antígenos em cortes histológicos para o diagnóstico da infecção pelo BRSV. Além disso, esses resultados confirmam evidências anteriores sobre a importância do BRSV como patógeno respiratório no sudeste e sul do Brasil.