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      Prevalence and molecular characteristics of Staphylococcus aureus, including methicillin resistant strains, isolated from bulk can milk and raw milk products in pastoral communities of South-West Uganda

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          Abstract

          Background

          Staphylococcus aureus strains are now regarded as zoonotic agents. In pastoral settings where human-animal interaction is intimate, multi-drug resistant microorganisms have become an emerging zoonotic issue of public health concern. The study of S. aureus prevalence, antimicrobial resistance and clonal lineages in humans, animals and food in African settings has great relevance, taking into consideration the high diversity of ethnicities, cultures and food habits that determine the lifestyle of the people. Little is known about milk carriage of methicillin resistant S. aureus strains (MRSA) and their virulence factors in Uganda. Here, we present the prevalence of MRSA in bulk can milk and raw milk products in pastoral communities of south-west Uganda. We also present PFGE profiles, spa-types, as well as frequency of enterotoxins genes.

          Methods

          S. aureus was identified by the coagulase test, susceptibility testing by the Kirby-Bauer disc diffusion and E-test methods and MRSA by detection of the mecA gene and SCC mec types. The presence of Panton – Valentine Leucocidin ( PVL) genes and staphylococcal enterotoxins was determined by PCR, while genotyping was by PFGE and spa typing.

          Results

          S. aureus were isolated from 30/148 (20.3%) milk and 11/91(12%) sour milk samples. mecA gene carriage, hence MRSA, was detected in 23/41 (56.1%) of the isolates, with 21 of the 23 (91.3%) being SCC mec type V; while up to 30/41 (73.2%) of the isolates were resistant to tetracycline. Only five isolates carried the PVL virulence gene, while PFGE typing revealed ten clusters (ranging from two seven isolates each) that comprised 83% of the sample, and only eight isolates with unique pulsotypes. The largest PFGE profile (E) consisted of seven isolates while t7753, t1398, and t2112 were the most common spa-types. Thirty seven of the 41 strains (90.2%) showed at least one of the eight enterotoxin genes tested, with sem 29 (70.7%), sei 25 (61%) and s eg 21 (51.2%) being the most frequently observed genes.

          Conclusion

          This is the first study to demonstrate MRSA and enterotoxin genes in raw milk and its products in Uganda. The fact that over 90% of the isolates carried at least one gene encoding enterotoxins shows a high risk of spread of foodborne diseases through milk in this setting.

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          Most cited references28

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          Novel multiplex PCR assay for characterization and concomitant subtyping of staphylococcal cassette chromosome mec types I to V in methicillin-resistant Staphylococcus aureus.

          Staphylococcal cassette chromosome mec (SCCmec) typing is essential for understanding the molecular epidemiology of methicillin-resistant Staphylococcus aureus (MRSA). SCCmec elements are currently classified into types I to V based on the nature of the mec and ccr gene complexes, and are further classified into subtypes according to their junkyard region DNA segments. Previously described traditional SCCmec PCR typing schemes require multiple primer sets and PCR experiments, while a previously published multiplex PCR assay is limited in its ability to detect recently discovered types and subtypes such as SCCmec type V and subtypes IVa, b, c, and d. We designed new sets of SCCmec type- and subtype-unique and specific primers and developed a novel multiplex PCR assay allowing for concomitant detection of the methicillin resistance (mecA gene) (also serving as an internal control) to facilitate detection and classification of all currently described SCCmec types and subtypes I, II, III, IVa, b, c, d, and V. Our assay demonstrated 100% sensitivity and specificity in accurately characterizing 54 MRSA strains belonging to the various known SCCmec types and subtypes, when compared with previously described typing methods. Further application of our assay in 453 randomly selected local clinical isolates confirmed its feasibility and practicality. This novel assay offers a rapid, simple, and feasible method for SCCmec typing of MRSA, and may serve as a useful tool for clinicians and epidemiologists in their efforts to prevent and control infections caused by this organism.
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            Staphylococcal enterotoxins.

            Staphylococcus aureus is a major human pathogen that produces a wide array of toxins, thus causing various types of disease symptoms. Staphylococcal enterotoxins (SEs), a family of nine major serological types of heat stable enterotoxins, are a leading cause of gastroenteritis resulting from consumption of contaminated food. In addition, SEs are powerful superantigens that stimulate non-specific T-cell proliferation. SEs share close phylogenetic relationships, with similar structures and activities. Here we review the structure and function of each known enterotoxin.
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              Implication of milk and milk products in food-borne diseases in France and in different industrialised countries.

              A study was carried out to estimate the proportion of diseases due to milk and milk products among food-borne diseases recorded in France and in other countries since 1980. Particular attention was given to whether the milk involved was heat-treated or not. Four etiologic agents were considered: Salmonella spp., Staphylococcus aureus, Listeria monocytogenes, and pathogenic Escherichia coli. An overview of food-borne disease annual reports from seven countries indicated that milk and milk products were implicated in 1-5% of the total bacterial outbreaks; however, details about the type of product and milk involved were usually not provided. When considering 60 outbreaks and four single cases described in the literature and implicating milk and milk products, confirmed or suspected food vehicles were distributed as follows: milk, 39.1%, cheese, 53.1%, other milk products, 7.8%. Overall, 32.8% of the food vehicles were made from pasteurised milk; 37.5% from raw milk; 10.9% from milk stated as "unpasteurised"; and 18.8% from unspecified milk. Salmonella spp. were responsible for 29 outbreaks, L. monocytogenes for 10 outbreaks and four well-documented single cases, pathogenic E. coli for 11 outbreaks, and S. aureus for 10 outbreaks. Analysis of unpublished data about food-borne disease outbreaks, listeriosis excluded, collected by the coordinator of the French surveillance system from 1992 to 1997, revealed 69 documented outbreaks for which milk and milk products were confirmed as the vehicle by the isolation of the etiologic agent. The food vehicles were distributed as follows: milk, 10%; cheese, 87%; others, 3%. UHT milk accounted for 1.5%, raw milk and raw milk products for 48%, and milk and milk products from unspecified milk for 50.5% of the 69 outbreaks. S. aureus was by far the most frequent pathogen associated with these outbreaks (85.5% of the outbreaks), followed by Salmonella (10.1%). This study demonstrates the limitations of the surveillance systems and the difficulties in estimating the contribution of milk and milk products to food-borne diseases. In particular, it was not possible to find out in many outbreaks what heat treatment, if any, the milk had undergone.
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                Author and article information

                Contributors
                basiimwe@chs.mak.ac.ug
                baldan.rossella@hsr.it
                trovato.alberto@hsr.it
                cirillo.daniela@hsr.it
                Journal
                BMC Infect Dis
                BMC Infect. Dis
                BMC Infectious Diseases
                BioMed Central (London )
                1471-2334
                13 June 2017
                13 June 2017
                2017
                : 17
                : 422
                Affiliations
                [1 ]GRID grid.414603.4, Emerging Bacterial Pathogens Unit, , IRCCS, ; Via Olgettina 58, Milan, Italy
                [2 ]GRID grid.15496.3f, , Universita Vita-Salute San Raffaele, ; Via Olgettina 58, Milan, Italy
                [3 ]ISNI 0000 0004 0620 0548, GRID grid.11194.3c, Department of Medical Microbiology, , Makerere University College of Health Sciences, ; P O Box 7072, Kampala, Uganda
                Article
                2524
                10.1186/s12879-017-2524-4
                5470224
                28610560
                8e2dd839-4c37-4f48-9067-8b0cf33b970c
                © The Author(s). 2017

                Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 14 February 2017
                : 5 June 2017
                Categories
                Research Article
                Custom metadata
                © The Author(s) 2017

                Infectious disease & Microbiology
                Infectious disease & Microbiology

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