Molecular characterization of extended-spectrum ß-lactamases-producing Enterobacteriaceae species in ground beef and chicken meat.

The objectives of this study were i) to characterize extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-E) using pheno- and genotyping methods, ii) to evaluate the antimicrobial resistance pattern against 10 antibiotics, and iii) to investigate class 1 integron (intI1) in 80 Enterobacteriaceae isolates obtained from chicken meat (n = 40; 47 isolates) and ground beef (n = 40; 33 isolates) samples. Through the study, we found that 55 (68.7 %) of 80 Enterobacteriaceae isolates were capable of ß-lactamase activity, and 38 (47.5 %) of them were multi-drug-resistant (MDR). The ground meat-origin isolates are 1.2 times more likely to produce imipenem resistance compared to chicken-meat-origin isolates (z = 2.1, p < 0.05, OR = 1.42). ESBL-E was found in 18 (22.5 %) of the isolates, 16.3 % of chicken meat and 6.3 % of ground beef origin. The bla genes were detected in 14 isolates [bla-TEM (n = 10; 12.5 %); bla-SHV (n = 4; 5.0 %); bla-CTX-M (n = 0)], where the predominant species were Escherichia (E.) coli and Citrobacter braakii. The nine ESBL-E isolates were MDR. Twenty-eight (35.0 %) of 80 isolates were found to be resistant to at least one third-generation cephalosporin, and eight (28.6 %) of them were also ESBL-E. Eleven of 16 (48.5 %) carbapenem-resistant isolates were ESBL-E. The intI1 gene was found in 13 (16.3 %) isolates, five of which were ESBL-E, and four of which were MDR. Co-existing with bla-TEM and the intI1 isolate was ESBL-E. coli, which was resistant to nine antibiotics. In conclusion, chicken meat and ground beef may pose a potential risk of containing ESBL-E, and bla genes which could be spread to the entire food chain.

Effects of Lactobacillus acidophilus administration to anorexigenic neuropeptides and some biochemical parameters on rats fed with a high-fat diet.

AIM: One of the rapidly rising global public health concern is obesity. Over the past three decades, the prevalence of obesity has doubled/tripled in several nations around the world, most likely as a result of urbanization, sedentary lifestyles, and increased intake of high-calorie processed foods. In this study, it was aimed to investigate the effects of Lactobacillus acidophilus administration on rats exposed to high-fat diet experimentally on anorexigenic peptides in the brain and some biochemical parameters in the serum. METHODS: In the study, 4 different experimental groups were formed. Group 1 was designated as the control group and fed with a standard rat chow (SD). Group 2 was designated as the high-fat diet (HFD) fed group. Group 3 fed with SD and L. acidophilus probiotic administered. Group 4 fed with HFD and L. acidophilus probiotic administered. At the end of the experiment, leptin, serotonin, glucagon-like peptide-1 (GLP-1) levels were measured in the brain tissue and serum. Glucose, total cholesterol (TC), triglyceride (TG), total protein (TP), albumin, uric acid, aspartate transaminase (AST), alanine aminotransferase (ALT) levels were determined in the serum. RESULTS: At the end of the study, it was found that there was an increase in body weight and body mass index in Group 2 compared to Group 1. It was determined that the levels of AST, ALT, TG, TC, glucose, leptin in the serum were significantly high (P < 0.05). The levels of GLP-1 and serotonin in the serum and in the brain were significantly low (P < 0.05). There was a significant decrease in TG and TC in Groups 3 and 4 compared to Group 2 (P < 0.05). The leptin hormone levels in serum and brain were significantly higher in Group 2 than in other groups (P < 0.05). GLP-1 and serotonin levels were found to be significantly low (P < 0.05). The leptin levels in the serum of Groups 3 and 4 decreased significantly compared to Group 2 (P < 0.05). CONCLUSION: It was found that probiotic supplementation in high-fat diet had positive effects on anorexigenic peptides. It was concluded that L. acidophilus probiotic can be recommended as a food supplement in the treatment of obesity.

Investigation of the effects of atorvastatin and Lactobacillus acidophilus on some hormones and oxidative stress in experimental hypercholesterolemia.

AIM: The investigation of serum leptin, ghrelin, insulin, seratonin hormones, NO, total oxidant/antioxidant status and brain cannaboid 1 receptor protein and apoptotic cell levels in atorvastatin and Lactobacillus acidophilus administrated experimental hypercholesterolemia was aimed in the project. METHODS: In the study, 5 experimental groups were formed. Group 1 was fed with standard rat chow, and Group 2 was fed with 2% cholesterol added standard rat chow for 8 weeks. Group 3 was fed with 2% cholesterol feed and received atorvastatin (20 mg/kg/day) for the last 4 weeks. Group 4 was given L. acidophilus (2 ×108 cfu/kg/day). Group 5 was given atorvastatin and L. acidophilus probiotic in the last 4 weeks of the experiment period. After the experimental period, blood samples were taken from each rat. Rats were sacrificed and brain tissues were taken for analyzes. In sera samples, leptin, ghrelin, insulin, serotonin hormones and NO levels were measured with ELISA. In brain samples, cannabinoid 1 receptor proteins and apoptosis levels were measured by ELISA. Total oxidant and antioxidant levels were investigated with using Rel Assay Kits. RESULTS: The addition of cholesterol to feeds increased the levels of serum cholesterol, insulin and leptin levels; on the other hand, reduced the levels of serotonin and ghrelin. In hypercholesterolemia, total oxidant and NO levels were increased, and total antioxidant levels were decreased. CONCLUSION: The results showed that administrations of L. acidophilus and atorvastatin might be recommended for treatment of hypercholesterolemia.

Quorum sensing systems and related virulence factors in Pseudomonas aeruginosa isolated from chicken meat and ground beef.

The objective of this study was to evaluate 50 [chicken meat (n = 45) and ground beef (n = 5)] Pseudomonas aeruginosa isolates to determine the expression of the lasI and rhl QS systems, related virulence factors, and the presence of N-3-oxo-dodecanoyl homoserine lactone (AHL: 3-O-C12-HSL). For the isolation and identification of P. aeruginosa, conventional culture and oprL gene-based molecular techniques were used. In relation to QS systems, eight genes consisting of four intact and four internal (lasI/R, rhlI/R) genes were analyzed with PCR assay. The two QS systems genes in P. aeruginosa isolates from ground beef (80.00%) and chicken meat (76.00%) were present at quite high levels. The 3-O-C12-HSL was detected in 14.00% of the isolates. Both biofilm formation and motility were detected in 98.00% of the isolates. Protease activity was determined in 54.00% of the isolates. Pyocyanin production was detected in 48.00% of the isolates. The las system scores strongly and positively correlated with the rhl system (p ˂ .01). PYA moderately and positively correlated with protease (p ˂ .05). In addition, there was statistically significance between lasI and protease activity (p < .10), and rhlI and twitching motility (p < .10). In conclusion, the high number of isolates having QS systems and related virulence factors are critical for public health. Pyocyanin, protease, and biofilm formation can cause spoilage and play essential role in food spoilage and food safety.

Histopathological and biochemical evaluation of paeoniflorin administration in an experimental periodontitis model.

The purpose of this study was to evaluate the effects of administered Paeoniflorin (Pae) on periodontal tissues within an experimental periodontitis model. Forty male Wistar rats were used in this study and experimental periodontitis was created in all rats except in the control group (n = 10, first group). In the periodontitis group, experimental periodontitis was created but no other application was performed (n = 10, second group). In the other groups created experimental periodontitis, systemic Pae (n = 10, third group) or saline (n = 10, fourth group) was applied. A biochemical analysis of the gingival vascular endothelial growth factor (VEGF) levels and a histomorphometric analysis (measurements of the area of alveolar bone, alveolar bone resorption, and attachment loss) were performed. In the Pae group, the area of the alveolar bone was increased, while alveolar bone resorption and attachment loss decreased. Gingival VEGF levels increased in all groups that created experimental periodontitis and the greatest increase seen in the Pae group. Histomorphometric and biochemical analyses in this study suggest that Pae has a curative effect on periodontal tissues. However, additional studies are needed to confirm these results.

Histopathological and biochemical evaluation of the effect of Paeoniflorin on the periodontium during and after periodontitis formation in rats.

OBJECTIVES: Appraise the effect of systemic Paeoniflorin (Pae) application on the periodontium during and after induction of experimental periodontitis in the presence of ligature and after its removal. DESIGN: Seventy male Wistar rats were separated into seven equal groups. The first group was reserved as healthy control group (Group 1: no periodontitis, no medication) and experimental periodontitis was induced with ligature in the remaining rats. In experimental periodontitis groups, Pae or saline was administered systemically in three differet periods; during the experimental periodontitis induction (period 1), after experimental periodontitis induction which ligature removed (period 2) or ligature kept in position (period 3). Only, one of the groups acted as the control periodontitis group and received no treatment. Experimental periodontitis groups were as follows; Group 2: medication in period 1, Group 3: periodontitis and no medication, Group 4: medication in period 2, Group 5: saline application in period 2, Group 6: medication in period 3, Group 7: saline application in period 3. Matrix metalloproteinases-9 (MMP-9) levels and interleukin-10 (IL-10) levels were detected biochemically and histomorphometric analyses were performed. These analyses included measurements of the area of alveolar bone, the level of alveolar bone, and attachment loss. RESULTS: Area of alveolar bone and IL-10 levels were higher in the Pae-administered groups; level of alveolar bone, attachment loss, and MMP-9 levels were correspondingly lower (P < 0.05). The beneficial effects at histomorphometrical and biochemical levels of Pae were the strongest in the rats that were administered Pae after the removal of ligature. CONCLUSIONS: Systemically administered Pae had a positive effect on the healing of periodontal tissues. Pae can be used as a new therapeutic agent for periodontal diseases, but microbiology-based studies and more extensive biochemistry-based experimental and clinical studies are needed to address this possibility.

The effects of the conjunctival Brucella vaccine on some biochemical parameters in sheep.

The genus Brucella causes significant economic losses due to infertility, abortion, stillbirth or weak calves, and neonatal mortality in livestock. Brucellosis is still a zoonosis of public health importance worldwide. In the past, vaccination was administered subcutaneously and nowadays, the conjunctival vaccine is administered. There is no definite information about the changes of the biochemical parameters and antibody response after conjunctival vaccination. In this study, the investigation of the changes in the levels of some biochemical parameters due to the conjunctival vaccination for brucellosis was aimed. Thirty sheep were used as an animal material. The vaccine was done single dose against Brucella melitensis and the blood was drawled from Vena jugularis during 4 months. Antibody levels were determined by serum tube agglutination test. Alkaline phosphatase (ALP), alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), glucose, total protein, and albumin levels were measured using commercial autoanalyzer in sera. The antibody titers (p < 0.001) increased significantly at first month compared to the pre-vaccination, but at the second month began to fall. There was no statistically significant changes in glucose, AST, ALT activity after vaccination (p > 0.005). The significant amount of total protein and ALP decreased after vaccination (p < 0.005). LDH levels and total protein levels were significantly increased (p < 0.005). In conclusion, conjunctival vaccine was considered to be used as a safe to protect the sheep from brucellosis and the results of the study may be used to improve the efficiency of brucellosis eradication programs within livestock management.

Evaluation of PCR methods for detection of Brucella strains from culture and tissues.

The genus Brucella causes significant economic losses due to infertility, abortion, stillbirth or weak calves, and neonatal mortality in livestock. Brucellosis is still a zoonosis of public health importance worldwide. The study was aimed to optimize and evaluate PCR assays used for the diagnosis of Brucella infections. For this aim, several primers and PCR protocols were performed and compared with Brucella cultures and biological material inoculated with Brucella. In PCR assays, genus- or species-specific oligonucleotide primers derived from 16S rRNA sequences (F4/R2, Ba148/928, IS711, BruP6-P7) and OMPs (JPF/JPR, 31ter/sd) of Brucella were used. All primers except for BruP6-P7 detected the DNA from reference Brucella strains and field isolates. In spiked blood, milk, and semen samples, F4-R2 primer-oriented PCR assays detected minimal numbers of Brucella. In spiked serum and fetal stomach content, Ba148/928 primer-oriented PCR assays detected minimal numbers of Brucella. Field samples collected from sheep and cattle were examined by bacteriological methods and optimized PCR assays. Overall, sensitivity of PCR assays was found superior to conventional bacteriological isolation. Brucella DNA was detected in 35.1, 1.1, 24.8, 5.0, and 8.0% of aborted fetus, blood, milk, semen, and serum samples by PCR assays, respectively. In conclusion, PCR assay in optimized conditions was found to be valuable in sensitive and specific detection of Brucella infections of animals.

[Detection of the first QnrS gene positivity in aquatic Aeromonas spp. isolates in Turkey]. / Türkiye'de su kaynakli Aeromonas spp. izolatlarinda saptanan ilk QnrS gen pozitifligi.

Aeromonas spp. are oxidase positive, gram-negative, facultative anaerobic bacilli that are widely distributed in aquatic environments. A.hydrophila, A.sobria and A.bestiarum may cause severe infections in both human and cold-blooded animals. Environmental persistance of quinolones that are widely used in both human and veterinary medicine plays an important role in the selection of resistant mutants. Plasmid-mediated resistance is one of the main mechanisms involved in quinolone resistance, and qnr, qepA, aac(6')-Ib-cr, oqxAB genes are identified as resistance determinants. Determination of various types of qnr gene in different bacteria mainly in Enterobacteriaceae, suggests that they are widely distributed in nature. Recently, plasmid-mediated quinolone resistance was defined among Aeromonas species isolated from water. The aim of this study was to investigate the presence of qnr genes among aquatic Aeromonas spp. in Turkey. A total of 45 Aeromonas strains isolated from water and fishes collected from three different geographical regions (Aegean, Mediterranean and Blacksea) in Turkey, were included in the study. The isolates were identified at species level by the use of 16S rDNA-RFLP (Restriction fragment length polymorphism) analysis and multiplex polymerase chain reaction (M-PCR). Among the isolates, 20 were identified as A.sobria, 10 as A.hydrophila, nine as A.salmonicida, four as A.bestiarum and two as A.veronii. The plasmid-mediated quinolone resistance determinants, qnrA, qnrB, qnrC and qnrS genes, were investigated by M-PCR, and sequence analysis was performed for nine qnr-positive isolates. According to the sequence analysis of the genes, qnr genes were characterized in six A.sobria, in two A.bestiarum and in one A.hydrophila isolate (9/45; 20%). When the sequence was compared with GenBank database, this gene was found as qnrS2. All qnrS-positive Aeromonas spp. isolates were ciprofloxacin-susceptible, while five of them were resistant to nalidixic acid. This study is the first research about the plasmid-mediated quinolone resistance and the presence of qnrS2 genes among Aeromonas spp. isolated from fishes and water in Turkey. In conclusion, various resistance genes of aquatic bacteria may constitute a potential risk for the transmission of those genes to other bacteria as well as clinical isolates.

[Nitrate reductase assay for the rapid detection of Staphylococcus aureus methicillin resistance: a breakpoint susceptibility testing method]. / Staphylococcus aureus metisilin direncinin hizli saptanmasinda nitrat redüktaz testi: Bir sinir deger duyarlilik test yöntemi.

Methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of hospital- and community-acquired infections. Therefore rapid and accurate detection of MRSA is essential for infection control and prevention of nosocomial spread. In this study, the efficacy of a nitrate reductase assay (NRA) as a breakpoint susceptibility testing method was evaluated for the rapid detection of methicillin resistance in S.aureus A total of 135 S.aureus clinical isolates from our collection were tested for methicillin susceptibility by NRA breakpoint susceptibility method and by broth microdilution method. For NRA breakpoint susceptibility testing three tubes including growth control tube (without drug), test tube (with 4 mg/L cefoxitin) and lyophilized test tube (with 4 mg/L cefoxitin) were used. 50 µl of 0.5 McFarland bacterial suspension of each isolate was inoculated into the tubes. All tubes were incubated at 35ºC. After five-hour incubation, 500 µl of freshly prepared reagent [2 units of 0.2% sulfanilamide, 2 units of 0.1% N-(1-naphthyl) ethylenediamine dihydrochloride and 1 unit of concentrated hydrochloric acid] was added into each tube and a color change was watched for. The color changed to violet/purple, if there was bacterial growth. If the color changed to violet/purple in all three tubes, the isolate was identified as methicillin-resistant. If the color changed in growth control tube but not in the test and lyophilized tube, the isolate was identified as methicillin-susceptible. Among 135 isolates tested, 97 had mecA gene and were methicillin-resistant by both microdilution and NRA breakpoint susceptibility method. The remaining 38 clinical isolates did not have this gene and were susceptible to methicillin by both methods used. All results were concordant to the PCR which was considered as the gold standard method. Specificity, sensitivity, positive and negative predictive values were 100%. NRA breakpoint susceptibility test in tubes is an inexpensive and reproducible method. This method can easily be used in many laboratories and does not require skilled personnel. In addition, test tubes are prepared by lyophilisation to provide long shelf-life which gives an important advantage.

Phenotypic and molecular characterization of Yersinia ruckeri isolates from rainbow trout (Oncorhynchus mykiss, Walbaum, 1792) in Turkey.

The aim of the study was the phenotypic and molecular characterization of Yersinia (Y) ruckeri strains, the causative agent of Enteric Redmouth Disease (ERM), by antibiotyping, random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) and sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns of whole cell proteins. For this aim, a total of 97 Y ruckeri isolates were analyzed. The isolates were distinguished into ten antibiotypes and six phenotypes according to their resistance properties and whole cell protein profiles, respectively. Also, a glycoprotein band of approximately 25.5 kDa was observed in all Y ruckeri strains tested. In all strains, six different RAPD types were observed. In conclusion, Y ruckeri strains isolated from rainbow trout of fish farms in Turkey showed variation according to their phenotypic and genotypic characteristics, and the use of these three typing techniques in double and triple combinations could be more useful for discriminating the strains.

Molecular typing and cdt genes prevalence of Campylobacter jejuni isolates from various sources.

The genetic diversity of 168 Campylobacter jejuni isolates originating from human (n=30), cattle (n=36), sheep (n=44), dog (n=35), and poultry (n=21) and cdt genes prevalence of the isolates were investigated. To determine the genetic diversity of these strains, random amplified polymorphic DNA-polymerase chain reaction (PCR) using a random primer (M13) was performed. The numbers of genotypes determined in human, cattle, sheep, dog, and poultry isolates were 19, 18, 17, 18, and 6, respectively. To find out the prevalence of cdt genes in C. jejuni isolates simultaneously, a multiplex PCR was performed. The prevalence of the separate cdt genes was found to vary from 69% to 100% for cdtA, 92% to 100% for cdtB, and 39% to 98% for cdtC. These rates without host discriminating were 95%, 98%, and 93% for cdtA, cdtB, and cdtC, respectively. The prevalence of all three cdt genes in strains originating from human, cattle, sheep, dog, and poultry were 87%, 67%, 84%, 89%, and 39%, respectively. These results showed the relatively high genetic heterogeneity and variation of cdt genes among C. jejuni isolates from various sources except for poultry isolates. This study gives baseline data on molecular characterization of C. jejuni strains from different sources.

Development and evaluation of a multiplex PCR assay for simultaneous detection of Flavobacterium psychrophilum, Yersinia ruckeri and Aeromonas salmonicida subsp. salmonicida in culture fisheries.

Bacterial cold water disease, enteric red mouth disease and frunculosis are the common bacterial diseases of fish worldwide. The etiologic agents of these diseases are Flavobacterium (F.) psychrophilum, Yersinia (Y.) ruckeri and Aeromonas (A.) salmonicida subsp. salmonicida, respectively. In this study, a multiplex polymerase chain reaction (m-PCR) method with YER8/10-Fer3/4-FP1/3 primer pairs which can identify these fish pathogens simultaneously was developed and optimized. In optimized conditions, neither false specific nor nonspecific amplification occurred. The detection limits of the m-PCR method using DNA extracts from dilutions of pure cultures of bacteria were 35 pg for Y. ruckeri and F. psychrophilum and 70 pg for A. salmonicida subsp. salmonicida. It was determined that 15 CFU Y. ruckeri and F. psychrophilum and 30 CFU A. salmonicida subsp. salmonicida could be detected by m-PCR developed using genomic DNA extracted from dilutions of the suspensions. The detection limits in the presence of tissue debris were 125 CFU for Y. ruckeri and F. psychrophilum and 250 CFU for A. salmonicida subsp. salmonicida. In conclusion, we submit that the m-PCR method developed and optimized in this study can be used for accurate and rapid identification of these bacteria.

Molecular typing of Staphylococcus aureus strains from ovine mastitis by pulsed-field gel electrophoresis and polymerase chain reaction based on coagulase and protein A gene polymorphisms.

Staphylococcus aureus is one of the most important etiologic agents of ovine mastitis. To develop effective control measures for mastitis, it is important to type S. aureus strains that have considerable genetic heterogeneity. In the current study, 47 S. aureus strains isolated from ovine mastitis were typed by polymerase chain reaction (PCR) based on coagulase (coa) and protein A (spa) polymorphisms and by pulsed-field gel electrophoresis (PFGE). Eight different coa types and 4 spa types were identified by PCR. While the most prevalent coa type was CG2 (42.56%), the spa types S4 and S1 were the most commonly observed (44.68% and 38.29%, respectively). Nineteen different pulsotypes were identified, and 12 of these were represented by a single isolate. Pulsotypes J and K were predominant and each represented 9 isolates (19.14%). All isolates belonging to J and K pulsotypes were CG2. Although all 9 isolates belonging to the J pulsotype were S4, all isolates in the K pulsotype were S1. While PFGE was found to be the best discriminatory technique for distinguishing strains, coa and spa types were found to be in correlation with PFGE types and can be used for quick, preliminary epidemiologic studies for detecting strains that may cause mastitis.

Mouse intestine colonization ability of Campylobacter coli strains.

Campylobacter coli is an etiological agent of gastrointestinal and extraintestinal infections in man and animals, and can be found as a commensal in gastrointestinal tract of animals. In this study, we aimed to determine differences among C coli strains in colonization of the intestinal tract of mice. Seven C coli strains isolated from diarrheic patients, asymptomatic hosts and chicken carcasses were used for this study. Each strain was inoculated with 0.1 ml of a bacterial suspension (3 x 10(8) CFU/ml) to 5 weanling mice, intragastrically. For the isolation of C coli, faecal pellets collected before inoculation and after inoculation at particular intervals were cultured on Campylobacter Selective Agar. Seven C. coli strains were divided into 3 colonization groups, based on faecal shedding. Group I showed immediate colonization, with prolonged excretion of organism in all mice. Group II showed delayed and short time colonization of C. coli. Group III could not colonize mice. Division of isolates into colonization groups was as follows: Group I included 3 strains from gastrointestinal disease; Group II included 2 strains from asymptomatic hosts and Group III included 2 strains from chicken carcasses. The study showed that there were marked differences among C coli strains with respect to their colonization potential and it may depend upon the origin of the strain. For understanding the complete pathogenesis of Campylobacter spp., a greater number of strains from different sources and geographical locations require to be tested in further investigations in the light of our findings.

Slime production and antibiotic resistance of Enterococcus faecalis isolated from arthritis in chickens.

Slime factor production and antibiotic resistance of 67 Enterococcus faecalis strains isolated from chicken arthritis were investigated in this study. Slime factor productions of enterococci were found as 59.7%. The antibiotic resistances were investigated by testing gentamycin, penicillin, streptomycin, vancomycin, danofloxacin, and enrofloxacin. The resistance rates were found as 62.68%, 76.11%, 67.16%, 13.43%, 47.76%, 43.28%, respectively. For slime factor positive enterococci, the antibiotic resistance rates were found as follows respectively; 82.50%, 87.50%, 92.50%, 17.50%, 72.50%, and 60.00%. In conclusion; the slime factor might play a role as a colonization factor for chicken arthritis and slime factor positive enterococci were found to be more resistant to these antibiotics. The resistance rates between slime factor positive and negative enterococci against the tested antibiotics except for vancomycin were found statistically significant (p<0.05).

Detection of methicillin resistance and slime factor production of Staphylococcus aureus in bovine mastitis / Detecção de resistência a meticilina e produção do fator slime por Staphylococcus aureus em mastite bovina

This study aimed to detect methicillin resistant and slime producing Staphylococcus aureus in cases of bovine mastitis. A triplex PCR was optimized targetting 16S rRNA, nuc and mecA genes for detection of Staphylococcus species, S. aureus and methicillin resistance, respectively. Furthermore, for detection of slime producing strains, a PCR assay targetting icaA and icaD genes was performed. In this study, 59 strains were detected as S. aureus by both conventional tests and PCR, and 13 of them were found to be methicillinresistant and 4 (30.7%) were positive for mecA gene. Although 22 of 59 (37.2%) S. aureus isolates were slimeproducing in Congo Red Agar, in PCR analysis only 15 were positive for both icaA and icaD genes. Sixteen and 38 out of 59 strains were positive for icaA and icaD gene, respectively. Only 2 of 59 strains were positive for both methicillin resistance and slime producing, phenotypically, suggesting lack of correlation between methicillin resistance and slime production in these isolates. In conclusion, the optimized triplex PCR in this study was useful for rapid and reliable detection of methicillin resistant S. aureus. Furthermore, only PCR targetting icaA and icaD may not sufficient to detect slime production and further studies targetting other ica genes should be conducted for accurate evaluation of slime production characters of S. aureus strains.

Este estudo objetivou a detecção de Staphylococcus aureus resistente a meticilina e produtor do fator slime em casos de mastite bovina. Um PCR triplex foi otimizado, com alvo no genes 16SrRNA, nuc e mecA para detecção de Staphylococcus spp, S. aureus e resistencia a meticilina, respectivamente. Para detecção das cepas produtoras do fator slime, empregou-se um PCR com alvo nos genes icaA e icaD. No estudo, 59 cepas foram identificadas como S. aureus por testes convencionais e PCR, sendo 13 resistentes a meticilina e quatro positivas para o gene mecA. Embora 22 das 59 cepas tenham sido produtoras do fator slime em Agar Vermelho Congo, no teste PCR somente 15 foram positivas para os genes icaA e icaD. Dezesseis e 38 das 59 cepas foram positivas para os genes icaA e icaD, respectivamente. Somente duas das 59 cepas foram positivas simultaneamente para resistência a meticilina e produção do fator slime, sugerindo falta de correlação entre estas características. Em conclusão, o PCR triplex otimizado neste trabalho mostrou-se ser um método rápido e confiável para detecção de S.aureus meticilina resistente. Por outro lado, somente PCR para os genes icaA e icaD pode não ser suficiente para detectar produção de fator slime e outros estudos com alvo em outros genes ica são necessários para um avaliação correta da produção do fator slime por S. aureus.

Detection of methicillin resistance and slime factor production of Staphylococcus aureus in bovine mastitis.

This study aimed to detect methicillin resistant and slime producing Staphylococcus aureus in cases of bovine mastitis. A triplex PCR was optimized targetting 16S rRNA, nuc and mecA genes for detection of Staphylococcus species, S. aureus and methicillin resistance, respectively. Furthermore, for detection of slime producing strains, a PCR assay targetting icaA and icaD genes was performed. In this study, 59 strains were detected as S. aureus by both conventional tests and PCR, and 13 of them were found to be methicillin resistant and 4 (30.7%) were positive for mecA gene. Although 22 of 59 (37.2%) S. aureus isolates were slime-producing in Congo Red Agar, in PCR analysis only 15 were positive for both icaA and icaD genes. Sixteen and 38 out of 59 strains were positive for icaA and icaD gene, respectively. Only 2 of 59 strains were positive for both methicillin resistance and slime producing, phenotypically, suggesting lack of correlation between methicillin resistance and slime production in these isolates. In conclusion, the optimized triplex PCR in this study was useful for rapid and reliable detection of methicillin resistant S. aureus. Furthermore, only PCR targetting icaA and icaD may not sufficient to detect slime production and further studies targetting other ica genes should be conducted for accurate evaluation of slime production characters of S. aureus strains.

[Investigation of biofilm formation and relationship with genotype and antibiotic susceptibility of Pseudomonas aeruginosa strains isolated from patients with cystic fibrosis]. / Kistik fibrozlu hastalardan izole edilen Pseudomonas aeruginosa izolatlarinin biyofilm olusturma yeteneklerinin arastirilmasi ve bu özelligin genotip ve antibiyotik duyarliligi ile iliskisinin belirlenmesi.

Pseudomonas aeruginosa is a frequent cause of respiratory infections in cystic fibrosis (CF) patients. P. aeruginosa strains isolated from these patients have often a mucoid phenotype at advanced disease. This mucoid structure contains a dense amount of alginate type polysaccharide which facilitates bacterial attachment to lung epithelia and provides protection from the immune system due to biofilm formation. The aims of this study were to investigate the biofilm formation and the relation of this property with genotype and antibiotic susceptibilities of P. aeruginosa strains isolated from CF patients. The biofilm formation was determined by using the Congo Red agar and Christensen methods. RAPD-PCR (Random amplification of polymorphic DNA polymerase chain reaction) and disc diffusion methods were used for genotyping and antibiotic susceptibility testing, respectively. Biofilm production was found positive in 33.3% (20/60) of P. aeruginosa tested. While 9 of these 20 isolates were of mucoid colony morphotype, among the 40 biofilm negative isolates mucoid colony was detected in 16 of them. RAPD genotyping based on 70% similarity yielded 19 (A-S) clusters and subtypes related to five of these clusters (K1, K2, N1, N2, Q1, Q2, R1, R2, S1, S2) making up a total of 24 genotypes. Nine of these genotypes composed of biofilm positive isolates and 15 were biofilm negative ones. Most of the biofilm positive strains belonged to K1 (n = 5) and K2 (n = 6) genotypes while biofilm negative isolates were in the L (n = 8) and O (n = 7) genotypes. The comparison of antibiotic susceptibilities in both groups revealed no statistically significant difference (p > 0.0%). However, highest rate of resistance was detected for tobramycin and lowest rate for piperacillin/tazobactam. The data obtained from this study indicated that biofilm negative and positive P. aeruginosa isolates clustered in different groups. These results should be supported with larger scale multi-center studies which may provide information about P. aeruginosa dynamics in CF lungs.

Effects of trichophytosis on serum zinc levels in calves.

The purpose of this study was to determine the zinc levels in calves with trichophytosis and to research the importance of zinc for fungi. The sera of 20 calves with trichophytosis and 10 healthy calves were used in this study. Zinc levels of the sera were measured by the atomic absorption spectrophotometer method. Serum zinc levels of diseased and healthy animals were found to be 42.0+/-16.6 microg/dL and 75.8+/-5.9 microg/dL, respectively. Serum zinc levels of diseased calves were lower than healthy ones and this difference were found to be important statistically (p<0.001), whereas there is no statistical difference on the levels of lymphocyte, monocyte, granulocyte, erythrocyte, hemoglobin, hematocrit, and mean corpuscular volume between groups. These parameters were not influenced by low zinc levels.