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    Biocontrol of Shiga Toxigenic Escherichia coli O157:H7 in Turkish Raw Meatball by Bacteriophage
    (Wiley, 2016) Gencay, Yilmaz Emre; Ayaz, Naim Deniz; Copuroglu, Gizem; Erol, Irfan
    With an Escherichia coli O157:H7 virulent bacteriophage, M8AEC16, biocontrol efficiency of phages on a highly risky, ready-to-eat, traditional delicacy food called raw meatball under different storage conditions was investigated. Phage, belonging to the Myoviridae family, was isolated from the wastewater of a local slaughterhouse and showed a broad lytic activity toward many E.coli O157:H7 strains with high efficiency of plating and O157 specificity. Our experimental study provided favorable results, with 0.69-2.09 log colony-forming unit (cfu)/g E.coli O157:H7 reductions in the first 5h of the replica trials. Major reductions of viable E.coli O157:H7 counts were observed in the beginning of the storage period, reaching up to 1.85 log cfu/g. Although a significant reduction in E.coli O157:H7 was observed with increased phage concentration, storage conditions had minor effect on efficiency of phage biocontrol. This is the first study in Turkey that investigates applicability of phage biocontrol for a traditional food model. Practical ApplicationsPhage addition in preparation stage of a very complex food model, ready-to-eat Turkish raw meatball, is a promising application in decontamination of Escherichia coli O157:H7. Although investigation of its genomic characteristics along with its stability to different food matrices must be completed for further use of the model phage M8AEC16, findings of this work were encouraging, as phages are valuable in biocontrol of important foodborne in this ready-to-eat Turkish delicacy.
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    Phenotypic and genotypic antibiotic resistance profiles of Escherichia coli O157 from cattle and slaughterhouse wastewater isolates
    (Springer, 2015) Ayaz, Naim Deniz; Gencay, Yilmaz Emre; Erol, Irfan
    The aims of this study were to determine the minimal inhibition concentration of 20 different antibiotics on cattle and slaughterhouse wastewater Escherichia coli O157, including both Shiga toxigenic E. coli O157 (STEC O157) and non-Shiga toxigenic strains (non-STEC O157) by the Epsilometer test, and to determine the antibiotic resistance gene profiles of the isolates by PCR. A total of 102 cattle and slaughterhouse wastewater E. coli O157 isolates including 96 E. coli O157:H7(+) (81 non-sorbitol fermenting [NSF] STEC O157:H7, 12 NSF non-STEC O157:H7, and three sorbitol fermenting [SF] non-STEC O157:H7) and six non-STEC O157:H7(-) isolated from 744 cattle and slaughterhouse wastewater samples collected within a 2-year period were assessed. Of 93 NSF E. coli O157:H7 isolates, 19 were resistant to tetracycline and sulfamethoxazole, 14 to trimethoprim, 13 to cefoxitin, 11 to streptomycin, 10 to ampicillin, eight to chloramphenicol, six to cephalothin, four to cefaclor, four to aztreonam, and four to nalidixic acid. In six of the E. coli O157:H7(-) isolates, tetracycline resistance was detected while five of them were also resistant to ampicillin, sulfamethoxazole, and trimethoprim. In PCR analysis, 26.0 % (25/96) of the NSF E. coli O157:H7(+) and all of the E. coli O157:H7(-) isolates harbored one or more antibiotic resistance genes. While tetA, tetB, tetC, strA, strB, and sulI genes were detected from a number of the isolates, tetD, tetE, tetG, cmlA, floR, sulII, aadA, and ampC genes were not detected in any of the isolates. Results suggest a high antibiotic resistance in E. coli O157:H7(+)/H7(-) cattle and wastewater isolates. The majority of our resistant isolates, antibacterial resistance genes did not correlate with observed phenotypic resistance. Other resistance traits and regulatory factors that mediate antibiotic resistance should be included in further antimicrobial resistance investigations.
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    Prevalence and molecular characterization of sorbitol fermenting and non-fermenting Escherichia coli O157:H7+/H7– isolated from cattle at slaughterhouse and slaughterhouse wastewater
    (Elsevier, 2014) Ayaz, Naim Deniz; Gencay, Yilmaz Emre; Erol, Irfan
    The prevalence and seasonal distribution of E. colt 0157:H7(+)/H7(-) in an array of aged cattle at slaughter and its dissemination with slaughterhouse wastewater over a two year period in Turkey were investigated. For this purpose, a total of 720 samples (240 rectoanal mucosal swap [RAMS], 240 carcass sponge and 240 bile samples) of 240 cattle categorized according to age, gender, breed and sampling site were collected along with additional 24 wastewater samples and were subjected to immunomagnetic separation based cultivation technique to efficiently isolate E. coli 0157 from the background flora. Identification (rfbE(0157),fliC(h7)), detection of major virulence factors (sbc7, sU2, eaeA, hly, lpfA1-3 and espA), intimin variants (eae-alpha 1, eae-alpha 2, eae-beta 3, eae-beta 1, eae-beta 2, eae-gamma l and eae-gamma 2/0) and shiga toxin variants (stx70 stx(1d), stx(2c), stx(2d), stx(2e), stx(2f)and stx(2g)) of all the isolates were assessed by PCR. From 10(42%) of RAMS and 11(4.6%) of carcass sponge samples and 5 (20.8%) of slaughterhouse wastewater samples, a total of 102 colonies (99 sorbitol negative and 3 sorbitol positive) were isolated. Overall, 17 (7.1%) and 15 (6.3%) of 240 sampled cattle were shown to harbor E. coli 0157 and E. colt 0157:H7, respectively either in their RAMS or carcass sponge samples analyzed. Statistically significant differences between categories; season, age, gender and breed of cattle were not observed (p > 0.05). None of the isolated E. coli 0157:H7+/H7- strains harbored any of the investigated intimin types other than eaeyi or shiga toxin variants stx(1d), StX(2e), six(2f) or StX(2g) while all were lpfA1-3+ except 5 E. coli 0157:H7- strains. Intimin variant eaey, and shiga toxin 1 variant stx/c were detected from all of the eaeA+ (97/102, 95.1%) and stxt (32/102,313%) strains, respectively while from sbd" (80/102, 78.4%) isolates, both stx(2c) (68/80, 85.0%) and 5tx(2d) (12/80, 15.0%) variants were determined. In the last decade, prevalence of E. coli 0157:H7 has an increasing trend in cattle. Slaughterhouses are the significant sources of environmental contamination with E. coli 0157:H7. Isolation and molecular characterization of sorbitol fermenting E. coli 0157:H7 are a novel finding and may lead to a revision of reference isolation procedure of E. coli 0157:H7 in future. (C) 2014 Elsevier B.V. All rights reserved.
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    The relationship between seropositivity and tissue cysts in sheep naturally infected with Toxoplasma gondii
    (Scientific Technical Research Council Turkey-Tubitak, 2014) Yildiz, Kader; Kul, Oguz; Gokpinar, Sami; Atmaca, Hasan Tarik; Gencay, Yilmaz Emre; Gazyagci, Aycan Nuriye; Gurcan, Ismayil Safa
    Skeletal muscles (tongue, masseter, leg, intercostal, and diaphragmatic muscles) and brain samples of 100 sheep at slaughter were analysed for the presence of T. gondii tissue cysts along with serum IgG titres. Two methods of isolation by percoll gradient centrifugation and tissue microarray (TMA) technique with immunoperoxidase staining were used. Seropositivity was detected in 88% (88/100) of sheep sera analysed by indirect fluorescent antibody test. Tissue cysts were observed in 46 (52.3%, 46/88) and 15 (17%, 15/88) of the seropositive sheep with the isolation technique and TMA and immunoperoxidase staining, respectively. The diameters of the tissue cysts were 25-58 x 25-62 (mean 34 x 36) mu m. The relationship between the presence of tissue cysts and seropositivity in sheep was statistically significant at 1/16 (P < 0.01) and at 1/64 and 1/128 (P < 0.001) serum dilutions.
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    Sheep as an important source of E-coli O157/O157:H7 in Turkey
    (Elsevier Science Bv, 2014) Gencay, Yilmaz Emre
    Escherichia coli O157:H7 is a globally important foodborne pathogen and has been mainly associated with cattle as the reservoir. However, accumulating data shows the importance of sheep as an E. coli O157:H7 vehicle. The presence of E. coli O157/O157:H7 in recto-anal mucosal swap and carcass sponge samples of 100 sheep brought to the slaughterhouse in Kirikkale were analyzed over a year. Molecular characteristics (stx(1), stx(2), eaeA, hly, lpfA1-3, espA, eae-alpha 1, eae-alpha 2, eae-beta, eae-beta 1, eae-beta 2, eae-gamma 1, eae-gamma 2/theta, stx1(c),stx1(d), Stx2(c), stx2(d), stx2(e), stx2(f), stx2(g), bla(ampC), tet(A), tet(B), tet(C), tet(D), tet(E), tet(G), sul1, sul2, floR, cmlA, strA, strB and aadA) of 79 isolates were determined and minimum inhibitory concentrations of 20 different antibiotics were investigated. E. colt O157/O157:H7 was found in 18% of sheep included in the study and was more prevalent in yearlings than lambs and mature sheep, and male than female sheep, though none of the categories (season, sex or age range) had significant effect on prevalence. Furthermore, Shiga-toxigenic E. colt (STEC) O157:H7 was determined in 2% and 8% of sheep feces and carcasses, respectively. Additionally, lpfA1-3 and eae-gamma 1 were detected in all isolates. None of the isolates showed resistance against investigated antibiotics, even though 4 sorbitol fermenting E. colt O157 isolates were positive for tet(A), sul1 and aadA. This is the first study in Turkey that reveals the potential public health risk due to the contamination of sheep carcasses with potentially highly pathogenic STEC O157:H7 strains. (C) 2014 Elsevier B.V. All rights reserved.