ERIC-PCR genotypic characterization of Haemophilus parasuis isolated from Brazilian swine

Haemophilus parasuis infection, known as Glässer’s disease, is characterized by fibrinous polyserositis, arthritis and meningitis in piglets. Although traditional diagnosis is based on herd history, clinical signs, bacterial isolation and serotyping, the molecular-based methods are alternatives for species-specific tests and epidemiologic study. The aim of this study was to characterize H. parasuis strains isolated from different states of Brazil by serotyping, PCR and ERIC-PCR. Serotyping revealed serovar 4 as the most prevalent (24 %), followed by serovars 14 (14 %), 5 (12 %), 13 (8 %) and 2 (2 %), whereas 40 % of the strains were considered as non-typeable. From 50 strains tested 43 (86%) were positive to Group 1 vtaA gene that have been related to virulent strains of H.parasuis. ERIC-PCR was able to type isolates tested among 23 different patterns, including non-typeable strains. ERIC-PCR patterns were very heterogeneous and presented high similarity between strains of the same animal or farm origin. The results indicated ERIC-PCR as a valuable tool for typing H. parasuis isolates collected in Brazil.This study was sponsored by FAPESP (process: 07/04893-9)

Characterization of antibiotic resistance in Listeria spp. isolated from slaughterhouse environments, pork and human infections

Introduction: Listeria species are susceptible to most antibiotics. However, over the last decade, increasing reports of multidrug-resistant Listeria spp. from various sources have prompted public health concerns. The objective of this study was to characterize the antibiotic susceptibility of Listeria spp. and the genetic mechanisms that confer resistance. Methodology: Forty-six Listeria spp. isolates were studied, and their minimal inhibitory concentrations of antibiotics were determined by microdilution using Sensititre standard susceptibility MIC plates. The isolates were screened for the presence of gyrA, parC, lde, lsa(A), lnu(A), and mprF by PCR, and the amplified genes were sequenced. Results: All isolates were susceptible to penicillin, ampicillin, tetracycline, erythromycin, and carbapenems. Resistance to clindamycin, daptomycin, and oxacillin was found among L. monocytogenes and L. innocua, and all species possessed at least intermediate resistance to fluoroquinolones. GyrA, parC, and mprF were detected in all isolates; however, mutations were found only in gyrA sequences. A high daptomycin MIC, as reported previously, was observed, suggesting an intrinsic resistance of Listeria spp. to daptomycin. Conclusions: These results are consistent with reports of emerging resistance in Listeria spp. and emphasize the need for further genotypic characterization of antibiotic resistance in this genusFAPESP, 2010/19005-4CAPE

Isolation and characterization of Pasteurella multocida from pets

Pasteurella multocida é o agente causador de muitas doenças de importância econômica em medicina veterinária e tem sido descrita como um agente de alto potencial zoónotico. No Brasil, ao contrário do que se observa na literatura internacional, informações sobre a frequência deste agente em animais de companhia como cães, gatos e coelhos são inexistentes. O presente estudo teve como proposta isolar cepas de Pasteurella multocida a partir de cães, gatos e coelhos, avaliar a freqüência deste agente nestas espécies animais, caracterizar os isolados de acordo com o tipo capsular e genes codificadores de fatores de virulência através da reação em cadeia pela polimerase, avaliar o perfil de resistência a antimicrobianos dos isolados obtidos e caracterizar as amostras através da PFGE. Foram examinados 640 animais, sendo 191 gatos, 309 cães e 140 coelhos. Dentre os animais examinados 8,1% foram positivos para o isolamento de P. multocida, sendo 10,4% dos gatos, 0,9% dos cães e 20,7% dos coelhos avaliados. Dentre as 93 cepas selecionadas 22,5% foram sensíveis a todas as drogas testadas, 77,4% das cepas foram resistentes a pelo menos uma droga utilizada e 5,3% foram apresentavam resistência a três ou mais drogas. Através da PFGE foram observados 39 pulsotipos com índice discriminatório igual a 0,97. Todos os genes codificadores de fatores de virulência foram detectados em pelo menos um isolado, sendo que nenhum destes esteve presente em 100% das cepas avaliadas.Pasteurella multocida is the causative agent of many diseases of economic importance in veterinary medicine and has been described as an agent a high zoonotic potential. In Brazil, contrary to what is observed in the international literature, information about the frequency of this agent in animals such as dogs, cats and rabbits are nonexistent. This study proposes isolate strains of Pasteurella multocida from dogs, cats and rabbits, evaluate the frequency of this agent in these animal species, characterize the isolates according to the capsular type, and genes encoding virulence factors through the reaction polymerase chain, evaluate the antimicrobial resistance profiles of isolates and characterize the samples by PFGE. We examined 640 animals, 191 cats, 309 dogs and 140 rabbits. Among the animals examined were positive 8.1% for the isolation of P. multocida, and 10.4% of cats, dogs 0.9% and 20.7% of the rabbits studied. Among the 93 selected strains 22.5% were susceptible to all drugs tested, 77.4% of strains were resistant to at least one drug were used and 5.3% showed resistance to three or more drugs. Through PFGE were observed 39 pulsotipos discriminatory index equal to 0.97. All genes encoding virulence factors were detected in at least one isolated, none of which was present in 100% of the strains evaluated

Genotypic characterization of Clostridium perfringens from swine by pulsed field gel electrophoresis (PFGE)

Clostridium perfringens é um importante patógeno envolvido em doenças entéricas dos animais domésticos e quadros de toxinfecção alimentar em humanos. Embora as infecções causadas por C. perfringens biotipo C e A em suínos sejam amplamente estudadas, existem poucos relatos que descrevem as reais correlações genéticas existentes na cadeia epidemiologia das clostridioses para esta espécie animal, assim como a transmissão do agente através da fêmea lactante, e a eliminação e perpetuação do agente no momento do abate. O presente estudo teve como objetivo o isolamento e a caracterização genotípica através da eletroforese em gel de campo pulsado (PFGE) de cepas de C. perfringens isoladas a partir de fezes e de carcaças de suínos no momento do abate, fezes de fêmeas suínas e seus leitões e de amostras de farinha de carne e ossos. Foi ainda realizada a comparação dessas cepas com cepas isoladas a partir de leitões com enterite. A freqüência de isolamento do agente em carcaças, em fezes de leitões terminados e a partir de farinha de carne e osso foram, 44,2%, 52,5%, e 32,2% respectivamente. De acordo com a reação em cadeia pela polimerase (PCR) foram detectadas somente as toxinas alfa e beta 2, sendo esta ultima detectada somente nos casos de enterite. Por meio da PFGE as amostras foram caracterizadas em 97 perfis genéticos com um alto índice discriminatório. As amostras isoladas de carcaça apresentaram alta similaridade em relação às de origem fecal, os isolados de farinha de carne apresentaram perfis similares aos obtidos em isolados de fezes de fêmeas, leitões sadios e leitões com enterite. E os isolados de leitões com e sem enterite apresentaram baixa similaridade em relação aos isolados de fêmeas.Clostridium perfringens is an important enteric disease pathogen of domestic animals and foodborne diseases of human beings. Although infections caused by C. Perfringens type C and A in swine are well studied, just few reports describe genetic relationship among strains in the epidemiological chain of Swine Clostridioses, as well as the transmission of the microorganism by the nursing female its elimination and maintenance at slaughterhouses. The aim of the present study was the isolation and genotypic characterization by Pulsed-Field gel eletrophoresis (PFGE), of C. Perfringens strains from feces and carcasses from at slaughterhouse pigs, feces from sows and their piglets, and samples of meat and bone meal. The microorganism isolation frequencies in carcasses, finishing pig feces, and meat and bone meal were 44.2%, 52.5%, 32.2%, respectively. According to Polymerase Chain Reaction (PCR) assay, only the alfa and beta 2 toxins were detected; whereas beta 2 toxin was detected barely in cases of enteritis. By means of the PFGE assay techinique the samples were characterized in 97 genetic profiles with a high discriminatory level. Strains from carcasses samples presented high similarity to strains from fecal origin. strains from meat and bone meal samples showed similar profiles to strains from sow feces samples; of sows, assimptomatic piglets and piglets with enteritis. And strains isolated from piglets (assimptomatic and with enteritis) presented low similarity to strains from sow feces samples

Genotypic characterization of Clostridium perfringens from swine by pulsed field gel electrophoresis (PFGE)

Clostridium perfringens é um importante patógeno envolvido em doenças entéricas dos animais domésticos e quadros de toxinfecção alimentar em humanos. Embora as infecções causadas por C. perfringens biotipo C e A em suínos sejam amplamente estudadas, existem poucos relatos que descrevem as reais correlações genéticas existentes na cadeia epidemiologia das clostridioses para esta espécie animal, assim como a transmissão do agente através da fêmea lactante, e a eliminação e perpetuação do agente no momento do abate. O presente estudo teve como objetivo o isolamento e a caracterização genotípica através da eletroforese em gel de campo pulsado (PFGE) de cepas de C. perfringens isoladas a partir de fezes e de carcaças de suínos no momento do abate, fezes de fêmeas suínas e seus leitões e de amostras de farinha de carne e ossos. Foi ainda realizada a comparação dessas cepas com cepas isoladas a partir de leitões com enterite. A freqüência de isolamento do agente em carcaças, em fezes de leitões terminados e a partir de farinha de carne e osso foram, 44,2%, 52,5%, e 32,2% respectivamente. De acordo com a reação em cadeia pela polimerase (PCR) foram detectadas somente as toxinas alfa e beta 2, sendo esta ultima detectada somente nos casos de enterite. Por meio da PFGE as amostras foram caracterizadas em 97 perfis genéticos com um alto índice discriminatório. As amostras isoladas de carcaça apresentaram alta similaridade em relação às de origem fecal, os isolados de farinha de carne apresentaram perfis similares aos obtidos em isolados de fezes de fêmeas, leitões sadios e leitões com enterite. E os isolados de leitões com e sem enterite apresentaram baixa similaridade em relação aos isolados de fêmeas.Clostridium perfringens is an important enteric disease pathogen of domestic animals and foodborne diseases of human beings. Although infections caused by C. Perfringens type C and A in swine are well studied, just few reports describe genetic relationship among strains in the epidemiological chain of Swine Clostridioses, as well as the transmission of the microorganism by the nursing female its elimination and maintenance at slaughterhouses. The aim of the present study was the isolation and genotypic characterization by Pulsed-Field gel eletrophoresis (PFGE), of C. Perfringens strains from feces and carcasses from at slaughterhouse pigs, feces from sows and their piglets, and samples of meat and bone meal. The microorganism isolation frequencies in carcasses, finishing pig feces, and meat and bone meal were 44.2%, 52.5%, 32.2%, respectively. According to Polymerase Chain Reaction (PCR) assay, only the alfa and beta 2 toxins were detected; whereas beta 2 toxin was detected barely in cases of enteritis. By means of the PFGE assay techinique the samples were characterized in 97 genetic profiles with a high discriminatory level. Strains from carcasses samples presented high similarity to strains from fecal origin. strains from meat and bone meal samples showed similar profiles to strains from sow feces samples; of sows, assimptomatic piglets and piglets with enteritis. And strains isolated from piglets (assimptomatic and with enteritis) presented low similarity to strains from sow feces samples