Molecular Typing and Antimicrobial Susceptibility of Methicillin-Resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) in raw milk can be transmitted from animals to humans, and in Tanzania raw milk is sold in local markets and consumed as purchased. This study was performed to determine the molecular characteristics and antimicrobial susceptibility pattern of MRSA strains isolated from raw bovine milk sold at local markets in Tanzania. A total of 117 raw milk samples were cultured on Baird-Parker medium to isolate S. aureus and PCR was used for amplification of gltB gene for S. aureus identification and the presence of mecA gene for methicillin-resistant strains. Coagulase-negative (CN) S. aureus were reconfirmed using tube coagulase, DNase, and API Staph tests. MRSA isolates were spa typed whereas antimicrobial susceptibility testing was performed by the disc diffusion method. Forty-six coagulase positives (CP) and two CN S. aureus were identified. Most strains were resistant to penicillin (72%), and 3 isolates: 2 CN S. aureus and 1 coagulase-negative Staphylococci (CNS), were phenotypically resistant to vancomycin, oxacillin, and cefoxitin and were confirmed to carry mecA. Resistance to clindamycin, trimethoprim-sulfamethoxazole, and tetracycline was 23.9%, 30.4%, and 41.3%, respectively. Twelve isolates exhibited multidrug resistance; however, only one mecA positive strain among the three was typeable and belonged to spa type t2603. This study reports for the first time the presence of CN variant of MRSA, which was assigned the spa type t2603, and the presence of multidrug resistant S. aureus isolates from bovine milk in Morogoro, Tanzania

Adaptation of recombinase-based in vivo expression technology (RIVET) to avian pathogenic Escherichia coli (APEC): Construction of a functional reporter strain and test-screen of the APEC RIVET library in chickens

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Identification of avian pathogenic Escherichia coli genes that are induced in vivo during infection in chicken

Avian pathogenic E. coli (APEC) is associated with extra-intestinal infections in poultry causing a variety of diseases collectively known as colibacillosis. Host and bacterial factors influencing and/or responsible for carriage and systemic translocation of APEC inside the host are poorly understood. Identification of such factors could help in the understanding of its pathogenesis and in the subsequent development of control strategies. Recombination based in vivo expression technology (RIVET) was used to identify APEC genes specifically expressed during infection in chickens. A total of 21 clones with in vivo-induced promoters were isolated from chicken's liver and spleen, indicative for systemic infection. DNA sequencing of the cloned fragments revealed that twelve of the genes were E. coli conserved genes (metH, lysA, pntA, purL, serS, ybjE, ycdK (rutC), wcaJ, gspL, sdsR ylbE and yjiY), six of the genes were phage-related/associated, and three genes were pathogen-specific (tkt1, irp2 and eitD). These genes are involved in various cellular functions such as metabolism, cell envelope and integrity, transport systems and virulence. Others were phage-related or have yet unknown functions.status: publishe

High prevalence iron receptor genes of avian pathogenic Escherichia coli

Avian pathogenic Escherichia coli are known to cause significant losses in the poultry industry worldwide. Although prophylactic measures based on vaccination are advisable, until now no full heterologous protection against colibacillosis has been achieved. Since iron is an essential nutrient to these bacteria, the aim of this study was to investigate the prevalence of 12 outer-membrane iron receptor genes in 239 pathogenic strains isolated from clinical cases of colibacillosis in chickens. Five multiplex polymerase chain reactions were developed as a tool for efficient screening. Among the 239 avian E. coli isolates, 100% were positive for fhuE and fepA, 96.2% for fiu, 92.9% for cir, 92.5% for iroN, 87.4% for iutA, 63.2% for fecA, 53.1% for fyuA, 46.9% for fhuA, 45.6% for ireA, 41.8% for chuA and 4.6% for iha.status: publishe

Identification of avian pathogenic Escherichia coli genes that are induced in vivo during infection in chickens

Avian pathogenic Escherichia coli (APEC) is associated with extraintestinal infections in poultry causing a variety of diseases collectively known as colibacillosis. The host and bacterial factors influencing and/or responsible for carriage and systemic translocation of APEC inside the host are poorly understood. Identification of such factors could help in the understanding of its pathogenesis and in the subsequent development of control strategies. Recombination-based in vivo expression technology (RIVET) was used to identify APEC genes specifically expressed during infection in chickens. A total of 21 clones with in vivo-induced promoters were isolated from chicken livers and spleens, indicative of systemic infection. DNA sequencing of the cloned fragments revealed that 12 of the genes were conserved E. coli genes (metH, lysA, pntA, purL, serS, ybjE, ycdK [ rutC], wcaJ, gspL, sdsR, ylbE, and yjiY), 6 of the genes were phage related/associated, and 3 genes were pathogen specific (tkt1, irp2, and eitD). These genes are involved in various cellular functions, such as metabolism, cell envelope and integrity, transport systems, and virulence. Others were phage related or have yet-unknown functions

Disinfection by hydrogen peroxide nebulization increases susceptibility to avian pathogenic Escherichia coli

BACKGROUND: Avian pathogenic Escherichia coli (APEC) are the major cause of economic losses in the poultry industry worldwide. Traditionally, antibiotics are used to treat and prevent colibacillosis in broilers. Due to resistance development other ways of preventing/treating the disease have to be found. Therefore during this study the nebulization of low concentrations of hydrogen peroxide (H2O2) was tested in the presence of chickens to lower pathogenicity of APEC. RESULTS: Significantly higher total lesion scores and higher E. coli concentrations were found in the spleen of chickens exposed to 2% H2O2 compared to those exposed to 1% H2O2 and control chickens which had been exposed to nebulization with distilled water. Higher total lesions scores and E. coli concentrations in the spleen were found in chickens exposed to 1% H2O2 in comparison to control chickens (not significant). CONCLUSION: H2O2 is rendering animals more prone to APEC infection contraindicating H2O2 nebulization in the presence of chickens.status: publishe

Bioluminescent avian pathogenic Escherichia coli for monitoring colibacillosis in experimentally infected chickens

Avian pathogenic Escherichia coli (APEC) are responsible for significant economic losses in the poultry industry. In this study, a model for investigating the pathogenesis of APEC infections was established. APEC strain CH2 (O78) was marked with the luciferase operon (luxCDABE) using a Tn7 transposon and tissues of experimentally infected chickens were analysed for a correlation between the bioluminescent signal and the number of bacteria. Transposition of the lux operon into the chromosome of the APEC isolate did not affect sensitivity to lytic bacteriophages and there was no effect on virulence in an intratracheal infection model in 1-day-old chicks, although results with a subcutaneous infection model were inconclusive. A correlation between the number of bacteria and the luminescent signal was found in liquid medium, as well as in homogenised heart, liver, spleen and lung of 4-week-old experimentally infected chickens. This study showed that lux could be used for identification of the infecting strain after experimental infection with APEC in poultry.publisher: Elsevier articletitle: Bioluminescent avian pathogenic Escherichia coli for monitoring colibacillosis in experimentally infected chickens journaltitle: The Veterinary Journal articlelink: http://dx.doi.org/10.1016/j.tvjl.2016.07.011 content_type: article copyright: © 2016 Elsevier Ltd. All rights reserved.status: publishe

A cocktail of in vitro efficient phages is not a guarantee for in vivo therapeutic results against avian colibacillosis

Avian pathogenic Escherichia coli (APEC) causes colibacillosis in poultry, leading to important economic losses worldwide. To cure APEC-infected chickens, a cocktail of four different APEC-specific bacteriophages (phages) was composed and tested. Specific phages were selected from a collection of phages isolated in Belgium. The selection was based on their obligate lytic infection cycle, a broad host range, low cross-resistance and low frequency of development of resistant APEC mutants. Genome analysis of the phages indicated they were close relatives of T4 and N4, considered to be safe in vivo. Chickens were intratracheally infected with APEC strain CH2 (serogroup O78), causing a mortality of about 50% during the seven days following the infection. The phage cocktail was administered 2h after the infection, via three different ways: intratracheally, intra-esophageally or via the drinking water. Treated groups did not show a significant decrease in mortality, lesion scores or weight loss compared to untreated groups, although the APEC-specific phages could be re-isolated from the lung and heart of chickens that were euthanized. Moreover, the re-isolated bacteria from infected chickens had remained sensitive to the phage cocktail. Our results indicate that the efficiency of the phage cocktail used in treating CH2-infected chickens in vivo is negligible, even though in vitro, the phages in the cocktail were able to efficiently lyse the APEC strain CH2. Our results emphasize that the 'traditional' pathway of isolation, followed by phenotypical and genotypical characterization of phages composing the cocktail, does not lead to success in phage therapy in all cases.publisher: Elsevier articletitle: A cocktail of in vitro efficient phages is not a guarantee for in vivo therapeutic results against avian colibacillosis journaltitle: Veterinary Microbiology articlelink: http://dx.doi.org/10.1016/j.vetmic.2013.10.021 content_type: article copyright: Copyright © 2013 Elsevier B.V. All rights reserved.status: publishe