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In this study, infectious bronchitis (IB)-suspect problems were observed in 2 broiler and 8 breeder chicken flocks vaccinated with H120 strain, and a total of 220 tracheal swabs were taken from 60 broiler chickens of 2 flocks (30 swabs/flock), and 160 breeder chickens of 8 flocks (20 swabs/flock) to detect infectious bronchitis virus (IBV) by a reverse transcriptase polymerase chain reaction (RT-PCR) and to isolate IBV from chickens showing respiratory sings and nephritis lesions. Detection of 8 Turkish IBVs from the tracheal swab samples directly by RT-PCR was confirmed by both IBV isolation from allantoic fluids of embryonated eggs and by the same RT-PCRs from these samples. Basic Local Alignment Search Tool (BLAST) results revealed that all 8 sequences from our Turkish IBV isolates were related to that of EU780077 (IS/1494/06 IBV strain) with a 99% identity. Phylogenetic and alignment analyses of the sequences of two representative isolates as TR_IS1494_06_B12011 from one broiler flock and TR_1494_06_7201 from one breeder flock also confirmed this close relation to IS/1494/06 IBV genotype. As a result, Turkish IBVs have been isolated for the first time from chickens, which were not protected from IB despite IBV H120 strain vaccination, and genotyped as IS/1494/06 in Turkey suggesting the transfer of this IBV from the countries of Middle East.

This study was conducted to evaluate the capability of the Vitek immunodiagnostic assay system easy Salmonella (VIDAS ESLM) method and a specific real-time PCR system (LightCycler, LCPCR) to complement the International Organization for Standardization Method 6579 (ISO) in detecting Salmonella from a total of 105 naturally contaminated samples comprised of poultry meat and poultry meat products. The detection limit of ISO and LCPCR was 9 cfu/mL for both poultry meat and poultry meat products, whereas that of VIDAS ESLM with both sample types was determined to be 90 cfu/mL. Twelve (33.33%), 11 (30.55%), and 18 (50.00%) out of 36 poultry meat samples were positive for Salmonella by ISO, VIDAS ESLM, and LCPCR, respectively. Salmonella detection rates from poultry meat products were 5.80% for ISO and 8.69% for LCPCR, whereas none of these products tested positive by VIDAS ESLM. In poultry meat samples, VIDAS ESLM and LCPCR detection results were in substantial agreement with ISO, with the relative accuracy, sensitivity, and specificity rates of 97.2, 91.7, and 100%, respectively, for VIDAS ESLM and 83.3, 100, and 75%, respectively, for LCPCR. This is the first report on the evaluation of both VIDAS ESLM and LCPCR to complement ISO for the rapid detection of Salmonella in poultry meat and meat products. We determined that both VIDAS ESLM and LCPCR have the potential to complement the ISO standard culture method in the rapid screening of Salmonella from naturally contaminated poultry meats. For the poultry meat products, VIDAS ESLM and LCPCR can be used for rapid primary screening, and they should be complemented absolutely by ISO. Although LCPCR can preferentially be used for initial screening poultry meat products, the results should definitely be confirmed by ISO. Also, the VIDAS ESLM did not seem to be a suitable method for detecting Salmonella in poultry meat products.

This study aims to determine the antibiotic resistance phenotypes of Salmonella isolates of broiler meat and chicken samples against a total of 23 antimicrobial agents. For this, 64 meat and 79 chicken Salmonella isolates, which were obtained from retail meat and farm samples analyzed in our laboratory from February 2006 to January 2009, were used. All Salmonella isolates were resistant to at least 4 antimicrobials tested, with the common highest resistance rates to erythromycin (100.0%), bacitracin (99.3%), and rifampicin (98.6%), regardless of the sample type. Additionally, meat isolates were 100.0% resistant to bacitracin, clindamycin, erythromycin, rifampicin, and chicken isolates were 100.0% resistant to erythromycin, spiramycin, penicillin G, doxycycline hydrochloride, and tetracycline. Four to 7 Multi Drug Resistant (MDR) Salmonella conformed 82.83% of all meat isolates, and 10, 12, 15, and 17 MDR Salmonella conformed 55.68% of all chicken isolates analyzed. The B/DA/E/RD and N/B/CT/E/SP/P/RD/DO/OT/TE resistances were the most common patterns as 35.94% and 13.92% within meat and chicken isolates, respectively. The considerably high resistance rates and MDR types to commonly used antimicrobial agents among the Salmonella isolates tested pose a significant risk factor for the treatment of foodborne Salmonella infections in humans, Bu çalışmada tavuk eti ve tavuk orijinli Salmonella izolatlarında 23 antimikrobiyal ajana karşı dirençlilik fenotiplerinin belirlenmesi amacı ile perakende satış yerleri ve çiftlik örneklerinden elde edilen 64 adet et ve 79 adet tavuk Salmonella izolatı, Şubat 2006 ile Ocak 2009 arasında laboratuvarımızda analiz edildi. Tüm Salmonella izolatlarının, örnek tipi göz önünde bulundurulmaksızın, test edilen antimikrobiyalleden en yüksek direnç oranı ile sırası ile eritromisin’e (%100.0), basitrasin’e (%99.3) ve rifampisin’e (%98.6) olmak üzere en az 4 antimikrobiyale dirençli olduğu belirlendi. Ayrıca, et izolatlarının basitrasin, klindamisin, eritromisin, rifampisin’e; tavuk izolatlarının ise eritromisin, spiramisin, penisilin G, doksisiklin hidroklorür ve tetrasiklin’e %100.0 dirençli olduğu bulundu. Tüm et izolatlarının %82.83’ü 4 ile 7 Çoklu İlaç Direnci (ÇİD) gösterirken, tüm tavuk izolatlarının %55.68’i 10, 12, 15 ve 17 antimikrobiyale ÇİD gösterdiği belirlendi. Et izolatlarında %35.94 oranında ve tavuk izolatlarında ise %13.92 oranında olmak üzere en sık rastlanan ÇİD paternleri sırasıyla B/DA/E/RD ve N/B/CT/E/SP/P/RD/DO/OT/TE olarak bulundu. Salmonella izolatlarında karşılaşılan bu yüksek dirençlilik oranlarının ve ÇİD paternlerinin insanlardaki Salmonella infeksiyonlarının tedavisinde risk oluşturabileceği belirtildi

This study aims to determine the serogroup profiles of randomly collected 46 chicken meat and 15 turkey meat samples, following the U. S. Food and Drug Administration's (FDA) Bacteriological Analytical Manual Chapter 5: Salmonella and International Organization for Standardization (ISO) Method 6579 culture methods. The total number of poultry meat samples with more than one serogroup isolated by the FDA and ISO culture methods were 10 (37.0%) and 21 (77.8%) of 27, respectively. Presence of multiple serogroups per sample was more frequently observed in chicken meat samples than in turkey meat samples. The profile of Salmonella serogroup isolates of chicken meat samples in descending order were serogroups D and E4 (15.8%), B and C2 (8.8%), C1 (5.3%), G (3.5%), and E1 and F (1.7%). The serogroup distribution of turkey meat sample isolates were serogroups B (27.2%), E4 (18.2%), and C2 (9.1%). On the basis of our findings that a selective plate in Salmonella culture method can harbor more than one serogroup, and that the FDA and ISO methods could detect different serogroups from chicken and turkey meats, we suggest screening multiple suspect colonies from each plate, if possible, and considering the collective and comparative use of the FDA and ISO culture methods and/or including several selective and differential media to ensure the detection of Salmonella and the possible detection of multiple serogroups from samples.

In this work, we describe a rapid detection procedure for Mycoplasma gallisepticum from chicken tracheal swabs by real-time polymerase chain reaction (PCR) by LightCycler system, where we were able to monitor the amplification of the newly synthesized M gallisepticum-specific PCR product as a proportionally increasing fluorescent signal by using the double-stranded DNA binding dye SYBR Green I and have identified M. gallisepticum-specific PCR products by DNA melting curve analysis by plotting the first negative derivative (-d[F1]/dT) of fluorescence over temperature. Detection limits of the PCR were found to be 3 and 3000 colony-forming units ml(-1) with pure culture of M. gallisepticum and artificially spiked samples, respectively. Out of 96 tracheal swabs, 68 were taken from live chickens and 28 were taken by scraping the mucosal surface of the trachea (SMST) of necropsied chickens. All of the 18 PCR-positive results were from the swabs taken by the SMST method, whereas all of the samples taken from live chickens were negative. Thus, the PCR with the SMST method had a sensitivity and a specificity of 64.2% (18 of 28 chickens) and 100%, respectively. The total time required for template preparation from tracheal swab samples and real-time PCR was approximately 65 min. These results indicate that real-time PCR with the LightCycler technology is a rapid and sensitive test to identify M. gallisepticum-infected flocks if a proper sampling is applied.

Detection of Salmonella by bacteriologic methods is known to be time consuming. Therefore, we have developed a real-time probe-specific polymerase chain reaction (PCR) to rapidly detect Salmonella invA gene-based PCR products from chicken feces and carcasses by a fluorescence resonance energy transfer assay. The sensitivity and the specificity of this system were determined as 3 colony-forming units ml(-1) and 100%, respectively. Overnight tetrathionate broth enrichment cultures of chicken feces and carcass samples were used in template preparation for PCR. Also, a standard bacteriology was performed (National Poultry Improvement Plan-U.S. Department of Agriculture, Bacteriological Analytical Manual-Food and Drug Administration Center for Food Safety and Applied Nutrition) for confirmation. Seventy-two cloacal swab, 147 intestine, and 50 carcass (neck) samples were examined. Thirteen (8.8%) and 25 (17%) of the intestinal samples were found to harbor Salmonella by bacteriology and PCR, respectively. Forty-five of 50 (90%) carcass samples were Salmonella positive by both methods. Salmonella was not detected from cloacal swab samples. Results indicate that this assay has the potential for use in routine monitoring and detection of Salmonella in infected flocks and carcasses.

This study aimed to determine the prevalence of Escherichia coli O157 in retail red meat and meat products with the Vitek Immunodiagnostic Assay System, including H7 Escherichia coli phage technology (VIDAS ECPT), and a real-time polymerase chain reaction system (LightCycler PCR; LCPCR). A total of 106 red meat and meat product samples were analyzed with VIDAS ECPT and LCPCR. VIDAS ECPT presumptive positive samples were subjected to VIDAS Immuno Concentration E. coli O157, followed by culture and serology. Among the 27 out of 72 (37.50%) red meat samples and 3 out of 34 (8.82%) red meat products that tested positive by VIDAS ECPT, 5.55% and 0.00% were confirmed positive for E. coli O157 (but not H7), respectively. Red meat and red meat product samples were 73.61% and 20.58% positive by LCPCR. The 5.55% prevalence of E. coli O157 in red meats poses a significant risk for consumers and indicates insufficient hygiene management both at the farm and during the slaughtering and meat handling processes in Turkey. This is the first report on the detection of E. coli O157 by VIDAS ECPT and LCPCR in naturally contaminated red meat and meat products. Bu çalışmanın amacı, Escherichia coli O157 (E. coli O157)’yi de içeren Escherichia coli Vitek Ultra Performans Immunodiagnostik Assay Sistemi Faj Teknolojisi (VIDAS ECPT) ve bir gerçek zamanlı Polimeraz Zincir Reaksiyonu sistemi (LightCycler PCR- LCPCR) ile kırmızı etlerde ve et ürünlerinde E. coli O157’nin prevalansının belirlenmesidir. Çalışmada, 106 kırmızı et ve et ürünü örneği VIDAS ECPT ve LCPCR ile analiz edildi. VIDAS ECPT şüpheli pozitif örnekler, VIDAS Immuno-konsantrasyon E. coli O157 (ICE) testine tabi tutuldu, ardından kültür ve seroloji uygulandı. VIDAS ECPT ile 72 kırmızı et örneğinin 27’si (% 37,50) ve 34 et ürünü örneğinin 3’ü (% 8,82) pozitif iken, bu örneklerin sırasıyla % 5,55 ve % 0,00 oranında E. coli O157 (H7 hariç) pozitif olduğu doğrulandı. Kırmızı et örneklerinin % 73,61’inin, kırmızı et ürünü örneklerinin ise % 20,58’inin LCPCR ile E. coli O157 pozitif olduğu belirlendi. Sonuç olarak, kırmızı etlerde % 5,55 oranında E. coli O157 bulunması tüketici yönünden risk oluşturmaktadır. Bu durum ülkemizde çift lik, kesimhane ve et işleme yerlerinde hijyen koşullarının yetersizliğinin bir göstergesidir. Bu çalışma, doğal kontamine kırmızı et ve et ürünlerinde E. coli O157’nin varlığının VIDAS ECPT ve LCPCR ile belirlendiği rapor edilen ilk çalışmadır

This study aims to determine the presence of Listeria monocytogenes, an important infectious agent in humans, and other Listeria spp. in retail dairy products using the Vitek Immunodiagnostic Assay System LDUO (VIDAS LDUO) rapid detection method and ISO method 11290-1 (ISO 11290-1). One hundred dairy products (10 pasteurised milk, 10 kashar cheese, 20 white cheese, 18 mihalic cheese, 12 curd cheese, 20 ice cream, and 10 butter samples), collected randomly from food stores in Bursa, Turkey were analysed using the VIDAS LDUO and ISO 11290-1 methods. The results showed that one dairy sample (1%), a mihalic cheese, was positive for both Listeria spp. and L. monocytogenes. This finding highlights the risk of Listeria contamination of cheese manufactured from raw milk.

This study aimed to compare a real-time PCR (rPCR) test with improved detection limit to serology and culture for the detection of Mycoplasma gallisepticum (MG) infection in chicken breeder flocks. Six hundred and forty-six blood and tracheal swab samples belonging to 31 grandparent chicken breeder flocks were tested by rPCR. The detection limit of rPCR was 0.9 pg mu l(-1), with pure MG S6 strain DNA and 100 colony forming units (CFUs) ml(-1), where both pure culture and tracheal swabs were artificially spiked with the same strain. The seropositive flock rate based on both MG RPA and HI were calculated as 48.4% (15/ 31) and 32.3% (10/31), respectively, while culture- and rPCR-positive flock rates were 16.1% (5/31) and 29.0% (9/31), respectively. On flock-based analysis, culture method detected 5 out of 10 MG seropositive flocks (sensitivity 50%, specificity 100%), whereas rPCR detected 8 out of 10 flocks (sensitivity 80%, specificity 95%). Agreements between serology and culture, and serology and rPCR were 83.9% and 90.3%, respectively. On individual sample-based analysis, culture method detected 26 out of 78 MG seropositive chicken (sensitivity 33%, specificity 100%), whereas rPCR detected 51 out of 78 MG seropositive chickens (sensitivity 65%, specificity 96%). There was 91.9% and 91.4% agreement between serology and culture, and serology and rPCR, respectively. Results of this study indicate that the rPCR with improved in vitro detection limit could detect MG in seropositive chicken flocks. Therefore, we advise the use of rPCR and/or culture for confirmation of serology results obtained from screening MG infection in chicken flocks.

This study presents Salmonella Enteritidis incidence in chicken layer flocks in Turkey determined by real-time PCR (rPCR) and by International Organization for Standardization (ISO) method 6579:2002/Amd 1:2007. A total of 259 samples, composed of 1,036 individual samples each pooled into 4, including 175 cloacal swab, 14 intestine, 35 gizzard swab, and 35 cecal swab samples, belonging to 6 major companies, were collected from 50 layer flocks and tested by rPCR and ISO culture methods. Overall incidence of Salmonella in layer flocks by rPCR and culture was 61.0 and 55.6%, respectively, where 70.1% of these Salmonella isolates were determined as Salmonella Enteritidis. Incidences of Salmonella Enteritidis in culture-positive samples were 65.3% in cloacal swabs, 50.0% in intestines, 73.9% in gizzard swabs, and 87.5% in cecal swabs. The rPCR results were in 100% agreement (100% sensitivity and specificity) with culture results when cecal swabs were selected as the sample type. The relative accuracy of rPCR was 92.4, 91.4, and 84% for intestine, gizzard, and cloacal swab samples, respectively. As a result, by using rPCR and ISO culture, we determined that the Salmonella Enteritidis incidence in layer flocks in Turkey was high and that the use of cecal swab and intestine samples in Salmonella detection would yield reliable results. To reduce this high Salmonella Enteritidis incidence in layer flocks, Salmonella Enteritidis-specific vaccination should be implemented properly in conjunction with a well-designed biosecurity plan, including verifiable corrective actions.

Aims: In this study, a capillary polymerase chain reaction (cPCR) was applied for Salmonella detection from poultry meat. Methods and Results: Salmonella detection limits of the optimized cPCR were determined with DNA templates from the samples of tetrathionate broth (TTB), Rappaport Vassiliadis broth (RVB) and selenite cystine broth (SCB) artificially contaminated with 10-fold dilutions of 6 × 108 CFU ml−1 of pure Salmonella enterica ssp. enterica serovar Enteritidis 64K stock culture. Detection limits of cPCR from TTB, RVB and SCB were found as 6, 6 × 101 and 6 × 104 CFU ml−1, respectively. In addition, detection limits of bacteriology were also determined as 6 CFU ml−1 with TTB and SCB, and 6 × 101 CFU ml−1 with RVB. A total of 200 samples, consisting of 100 chicken and 100 turkey meat samples, were tested with optimized cPCR and bacteriology. Eight and six per cent of the chicken meat samples were found to harbour Salmonella by cPCR and standard bacteriology, respectively. Of six Salmonella isolates, four belonged to serogroup D, two to serogroup B. Conclusions: The TTB cultures of both artificially and naturally contaminated samples were found to be superior to those of RVB and SCB cultures in their cPCR results. This cPCR, utilizing template from 18-h TTB primary enrichment broth culture, takes approximately 40 min in the successful detection of Salmonella from poultry meat. Significance and Impact of the Study: This study shows that cPCR from TTB enrichment culture of poultry meat would enable rapid detection of Salmonella in laboratories with low sample throughput and limited budget.

From years 2000 to 2003, Salmonella was investigated from a total of 1785 samples comprised of chicken intestinal samples, cloacal swabs, drag swabs, litter samples and chick dust samples collected from 191 poultry breeding flocks belonging to 15 different chicken breeding stock companies in the Marmara region, Turkey by a SYBR green-based real-time polymerase chain reaction (SGBRT-PCR), by a probe-specific real-time polymerase chain reaction (PSRT-PCR) and by standardized bacteriology as described in the manual of National Poultry Improvement Plan and Auxillary Provisions, United States Department of Agriculture. Between January 2000 and July 2001, Salmonella was detected at the rates of 5.87% and 4.10% out of a total of 1242 samples by SGBRT-PCR and bacteriology, respectively. From July 2001 until December 2003, Salmonella was found at rates of 11.42% and 5.52% from a total of 543 samples by PSRT-PCR and bacteriology, respectively. The dominant Salmonella serovar was determined as Salmonella enterica subsp. enterica Serovar Enteritidis ( S. Enteritidis), while serogroup C1 and C2 in 2001 and serogroup E1 in 2002 were isolated as additional serovars. As a conclusion, S. Enteritidis seems to be the major problem in poultry breeding flocks in Turkey, and both of the real-time polymerase chain reaction methods were found more sensitive than standard bacteriology for the detection of Salmonella from poultry samples.

This report describes a rapid detection procedure for salmonellae from chicken feces by the combination of tetrathionate primary enrichment (preenrichment [PE])-bacterial lysis-capillary PCR and capillary gel electrophoresis. Pure Salmonella enterica serovar Enteritidis 64K was reisolated and detected by capillary PCR after buffered peptone water and nutrient broth, tetrathionate broth base Hajna (TTBH), and tetrathionate broth (TTB) preenrichments. When the same culture was mixed with intestinal homogenate, bacteriological reisolation and capillary PCR detection was achieved only by TTBH and TTB preenrichments. Capillary gel electrophoresis revealed that a Salmonella genus-specific 281-bp PCR product was detected when Salmonella strains but not non- Salmonella strains were tested. The detection limit of capillary PCR with whole-cell DNA extracted from pure Salmonella enterica serovars Enteritidis 64K, Typhimurium LT2-CIP60-62, and Gallinarum 64K was 3, 3, and 9 CFU ml −1 , respectively. The detection limit of capillary PCR from whole-cell DNA extracted from intestinal homogenate artificially contaminated with the same three strains was 3, 3, and 7 CFU ml −1 , respectively. We compared the results of the capillary PCR and bacteriological examination from the natural samples. Thirty-five of 53 naturally contaminated samples produced a specific PCR product. In 9 of the 35 PCR-positive samples, Salmonella could not be detected bacteriologically either by PE or a primary and delayed secondary enrichment (DSE) combination. In the 18 PCR-negative samples, 4 samples were found to harbor Salmonella by both PE and DSE and 14 samples were positive after DSE. Fifty-three additional intestinal homogenate samples, which were negative by their PE and DSE in bacteriological examination, were found to be also negative by their PCRs. The total time required to detect Salmonella with the capillary PCR method we used was approximately 20 h. If samples are from clinically diseased birds, the total time for PCR and detection is reduced to 2 h since the 18-h PE is not required. These results indicate that TTB enrichment, bacterial lysis, and genus-specific capillary PCR combined with capillary gel electrophoresis constitute a sensitive and selective procedure which has the potential to rapidly identify Salmonella -infected flocks.

The possible use of a genus-specific polymerase chain reaction (PCR) for the identification of Salmonella colonies was tested on 14 reference strains and 38 clinical Salmonella strains isolated from chickens by the standard method of the National Poultry Improvement Plan and Auxiliary Provisions (NPIP), USDA, U.S.A. All the PCRs using primer pairs InvA1 and InvA2 (5) gave 281 bp amplification product specific for the Salmonella genus and produced results identical (100%) to those obtained with biochemical and serological methods. Thus, the genus-specific capillary PCR for the identification of a colony of Salmonellae from a selective agar plate was performed within approximately 2 h and the total time required for definitive diagnosis of infection was 2 days using primary enrichment (PE) and 7 days using delayed secondary enrichment (DSE).

Sulphate-reducing bacteria (SRB) were found in all of 200 bovine faeces examined. The number of SRB in bovine faeces ranged from 5 times 102 to 6 times 108 bacteria g-1. Of 50 isolates identified, all were assigned to the genus Desulfovibrio.