Short communication: Recovery of viable Mycobacterium avium subspecies paratuberculosis from retail pasteurized whole milk in Brazil.

Mycobacterium avium ssp. paratuberculosis (MAP) is the etiological agent of paratuberculosis, a chronic granulomatous enteritis that affects all ruminants worldwide. Some researchers have indicated a possible role of MAP in Crohn's disease. Despite extensive research and large and important advances in the past few decades, the etiology of Crohn's disease remains indefinite. The most probable transmission route of MAP from animals to humans is milk and dairy products. Mycobacterium avium ssp. paratuberculosis has already been detected in milk samples worldwide, and some studies have reported that MAP is resistant to pasteurization. In Brazil, MAP has been reported in raw milk samples; however, Brazilian retail pasteurized milk has not yet been tested for viable MAP. The aim of this study was to investigate MAP in pasteurized milk in the region of Viçosa (Minas Gerais, Brazil). Thirty-seven samples were collected and processed for culture of MAP. One colony similar to MAP was observed and confirmed by IS900-nested PCR and sequencing. Analysis revealed 97 to 99% identity with the MAP K-10 strain. This study is the first report of the presence of MAP in retail pasteurized whole milk in Brazil.

Effect of the inhibitors phenylalanine arginyl ß-naphthylamide (PAßN) and 1-(1-naphthylmethyl)-piperazine (NMP) on expression of genes in multidrug efflux systems of Escherichia coli isolates from bovine mastitis.

The multidrug efflux system in bacteria can reduce antibiotic concentration inside the cell, leading to failure in the treatment of bacterial diseases. This study evaluated the influence of two efflux pump inhibitors (EPIs), phenylalanine arginyl ß-naphthylamide (PAßN) and 1-(1-naphthylmethyl)-piperazine (NMP), on the gene expression of three multidrug efflux systems, AcrAB, AcrEF and EmrAB in Escherichia coli bovine mastitis isolates resistant to ampicillin and sulfamethoxazole/trimethoprim simultaneously. Each isolate had at least three multidrug efflux system genes. The acrA and acrB had the lowest expression levels in all treatments, while the emrA or emrB showed the highest expression levels in the presence of ampicillin, sulfamethoxazole/trimethoprim, PAßN and NMP. EPIs also contributed to the decrease in arcF expression when used in combination with ampicillin treatment. Since PAßN showed stronger effects than NMP, it may serve as an alternative to assist in the antimicrobial therapy of mastitis.

Sistemas de efluxo multidrogas em Escherichia coli e uso de inibidores como possíveis adjuvantes na terapia da mastite bovina / Multidrug efflux systems in Escherichia coli and the use of inhibitors as possible adjuvant therapy in bovine mastitis

Vinte e sete isolados de Escherichia coli provenientes de leite de bovinos com mastite clínica foram submetidos a teste de sensibilidade aos principais antimicrobianos usados no tratamento desta doença. Avaliou-se também a efetividade in vitro de dois inibidores de sistemas de efluxo multidrogas, fenilalanina arginyl ß naftilamida (PAβN) e 1-(1-Naphthylmethyl)-piperazine (NMP), utilizando-se a concentração inibitória mínima (CIM) como referência. A CIM e o sistema de efluxo foram detectados com base nas curvas de crescimento, utilizando-se a densidade óptica (D.O550), em diferentes concentrações da droga e na presença e ausência dos inibidores. Apenas quatro isolados apresentaram resistência à ampicilina e ao sulfametoxazol∕trimetoprim, simultaneamente, enquanto todos os 27 foram sensíveis aos demais antimicrobianos. Observaram-se valores para ampicilina variando de 6µg/mL a 250µg/mL e para sulfametoxazol∕trimetoprim de 12µg/mL a 1500µg/mL. Houve redução das CIMs desses antimicrobianos em todos os isolados na presença dos inibidores, exceto para sulfametoxazol∕trimetoprim na presença de NMP. Conclui-se que esses isolados possuem um estreito perfil de resistência e que PAßN apresentou melhor efeito inibitório em relação à ampicilina e ao sulfametoxazol∕trimetoprim, apresentando-se como um candidato a adjuvante no tratamento da mastite...

Twenty-seven isolates of Escherichia coli from cattle with clinical mastitis were subjected to sensitivity tests regarding main antimicrobials used in the treatment of this disease. We also evaluated in vitro effectiveness of two inhibitors of multidrug efflux systems, fenilalanina arginyl ß naftilamida (PAβN) and 1-(1-Naphthylmethyl)-piperazine (NMP), using the minimum inhibitory concentration (MIC) as a reference. MIC and multidrug efflux systems were detected in the growth curves, using optical density (D.O550) at different drug concentrations and the presence and absence of inhibitors. Only four isolates of E. coli were resistant to ampicillin and trimethoprim/sulfamethoxazole, simultaneously. However, all isolates were sensitive to the other antimicrobials. Were observed values ranging from 6mg to 250mg ampicillin/mL, and 12mg to 1500mg/mL trimethoprim/sulfamethoxazole. There was a reduction of the MIC of antimicrobials for all isolates in the presence of inhibitors, except for trimethoprim/sulfamethoxazole in the presence of NMP. In conclusion, these isolates have a narrow resistance profile and PAßN showed better inhibitory effect against ampicillin and trimethoprim/sulfamethoxazole, and is a candidate for the adjuvant treatment of mastitis...