Occurrence, Virulence and Antimicrobial Susceptibility Profiles of Cronobacter spp. from Ready-to-Eat Foods.

Cronobacter spp. can cause foodborne diseases in infants, but Cronobacter infections in healthy adults and vulnerable people have also been reported. These bacteria have ubiquitous nature and can contaminate various foods. Therefore, we assessed the presence of Cronobacter spp. in popularly consumed ready-to-eat (RTE) food products. In the present study, 51 (15%) of the 340 RTE food samples were contaminated with Cronobacter spp The highest contamination rates were found in spices (46.7%), meat-free cig koftes (44.4%), desserts (23.3%), cereals (23.1%), doners (12.2%), and ice cream (11.1%). Phenotypic and molecular methods, including 16S rRNA, gluA, rpoB, cgcA genes, and fusA allele sequencing were tested to identify Cronobacter species. Of the 51 contaminated samples, 54 isolates were identified as C. sakazakii (n = 43), C. malonaticus (n = 7), C. muytjensii (n = 3) and C. turicensis (n = 1) using fusA analysis. These isolates were assigned to 15 different fusA alleles, two of which (191 and 192) were new alleles. Putative virulence factors such as the ompA and zpx gene, biofilms, and siderophores were detected in most of the Cronobacter isolates (> 85%). Cronobacter isolates were resistant to cephalothin (85.2%), cefoxitin (33.3%), cefotaxime (14.8%), ampicillin (11.1%), cefepime (5.6%), aztreonam (5.6%), and piperacillin (1.9%). The multidrug resistance (against three or more classes of antimicrobial agents) was 7.4%. The results indicated presence of Cronobacter spp. in RTE foods, which may be a risk to human health. It is important to adopt rigorous hygiene and sanitization practices to ensure the microbiological safety of these foods consuming without any processing.

Prevalence and antimicrobial resistance of Listeria species and molecular characterization of Listeria monocytogenes isolated from retail ready-to-eat foods.

A wide variety of foods can be contaminated with Listeria species, especially L. monocytogenes. Ready-to-eat (RTE) foods are predominantly associated with human listeriosis caused by L. monocytogenes. Therefore, this study aimed to assess the presence of Listeria species in RTE foods and to characterize L. monocytogenes isolates by means of detection of virulence markers, serotypes and genetic relatedness. Of the 300 RTE food samples, 59 (19.7%) were positive for Listeria species: L. innocua (13.3%), L. monocytogenes (5%), L. welshimerii (2.3%), L. grayi subsp. murrayi (1.3%), L. grayi (1%), L. ivanovii (1%) and L. ivanovi subsp. londoniensis (0.3%). All L. monocytogenes isolates identified were positive for the actA, iap, inlA, inlB, inlC, inlJ, plcA and prfA virulence genes and biofilm. The isolates were serotyped as 1/2c (33.3%), 4b (26.7%), 1/2a (26.7%), 1/2b (6.7%) and 3c (6.7%) by the multiplex-PCR and agglutination methods. PCR-restriction fragment length polymorphism with AluI and MluCI resulted in three and two profiles, respectively. Pulsed-field gel electrophoresis differentiated the L. monocytogenes isolates into 15 ApaI and 12 AscI patterns. Antimicrobial resistance of all Listeria isolates was determined by the disk diffusion method. Most L. monocytogenes isolates were sensitive to antimicrobials used in the treatment of listeriosis. This study shows the presence of potential pathogenic and antimicrobial-resistant L. monocytogenes in RTE foods that may lead to consumer health risks.

Molecular Characterization and Toxin Profiles of Bacillus spp. Isolated from Retail Fish and Ground Beef.

Bacillus species are common in the environment due to their spore-forming ability and nutritional versatility and cause food contamination. Bacilli play a significant role in foodborne illnesses and food spoilage. In this study, 52 Bacillus isolates from retail fish and ground beef were identified and differentiated based on 16S rRNA, gyrB, and rpoB gene sequencing. The presence of genes encoding emetic toxin (ces), hemolytic enterotoxin hemolysin BL (hbl), nonhemolytic enterotoxin (nhe) and cytotoxin K (cytK1) was assessed in all Bacillus isolates. The ability of the Bacillus isolates to produce several extracellular enzymes that contribute to pathogenicity and food spoilage was investigated. The 16S rRNA, rpoB, and gyrB gene sequence similarities of the Bacillus isolates tested were 96.1%, 83.2%, and 77.5%, respectively. The gyrB gene demonstrated a higher degree of sequence variation than the 16S rRNA and rpoB genes. The prevalence of Bacillus isolates producing at least two of the genes of the HBL and NHE complexes was 23.1% and 15.4%, respectively. Of the B. cereus isolates, 10 (41.7%) possessed two or more enterotoxin genes. None of the isolates carried the ces and cytK1 genes. All isolates were positive for the production of enzymes such as protease, lipase, gelatinase, and DNase. However, only 92.3% of the tested isolates were positive for amylase. In conclusion, our results revealed that the presence of genes involved in toxin production and enzyme production in meat-originated B. cereus and other Bacillus isolates may cause spoilage of food and pose a health risk for consumers. PRACTICAL APPLICATION: Bacillus species can be found in various foods due to their ubiquitous nature. Bacillus spp., especially B. cereus, are associated with food poisoning and other infections in humans. Toxins and many extracellular enzymes produced by Bacillus spp. are the causative agents of foodborne outbreaks, food spoilage, and low-quality food with significantly reduced edibility. This study highlights the characterization of Bacillus spp. and presence of potentially pathogenic Bacillus species in meats.

Genotypic and phenotypic virulence characteristics and antimicrobial resistance of Yersinia spp. isolated from meat and milk products.

A total of 300 food samples including 180 milk and 120 meat products have been examined for the presence of Yersinia spp. using the ISO 10273 and the cold enrichment method. The overall prevalence of Yersinia spp. was 84 (28%). Yersinia enterocolitica was isolated from 18 (6%) of the 300 samples. The other Yersinia species were detected in the samples Yersinia rohdei 15 (5%), Yersinia intermedia 14 (4.7%), Yersinia pseudotuberculosis 12 (4%), Yersinia ruckeri 12 (4%), Yersinia mollaretii 5 (1.7%), Yersinia bercovieri 4 (1.3%), and atypical Yersinia spp. 4 (1.3%). The conventionally identified Y. enterocolitica strains were also confirmed by the 16S rRNA gene sequencing. All Y. enterocolitica strains biotyped as 1A had negative results in the phenotypic virulence tests. The 84 Yersinia strains were also examined genotypically for the presence of virulence genes. None of the Y. enterocolitica and other Yersinia strains contained the ail, ystA, yadA, and virF except only 1 Y. intermedia and 2 Y. enterocolitica strains that were found to be positive for ystB. Antimicrobial resistance of 84 Yersinia to 16 antimicrobial agents was determined by the disk diffusion method. All strains were sensitive to tobramycine and imipenem while resistant to clindamycin. Although 84.5% of the strains were resistant to at least 3 or more antimicrobial agents, 64.3% of them were resistant to 4 or more antimicrobial agents.

Toxigenic genes, spoilage potential, and antimicrobial resistance of Bacillus cereus group strains from ice cream.

Bacillus spp. can be recovered from almost every environment. It is also found readily in foods, where it may cause food spoilage and/or food poisoning due to its toxigenic and pathogenic nature, and extracellular enzymes. In this study, 29 Bacillus cereus group strains from ice cream were examined for the presence of following virulence genes hblC, nheA, cytK and ces genes, and tested for a range of the extracellular enzymes, and antimicrobial susceptibility. The strains were found to produce extracellular enzymes: proteolytic and lipolytic activity, gelatin hydrolysis and lecithinase production (100%), DNase production (93.1%) and amylase activity (93.1%). Of 29 strains examined, 24 (82.8%) showed hemolytic activity on blood agar. Beta-lactamase enzyme was only produced by 20.7% of B. cereus group. Among 29 B. cereus group from ice cream, nheA was the most common virulence gene detected in 44.8% of the strains, followed by hblC gene with 17.2%. Four (13.8%) of the 29 strains were positive for both hblC gene and nheA gene. Contrarily, cytK and ces genes were not detected in any of the strains. Antimicrobial susceptibility of ice cream isolates was tested to 14 different antimicrobial agents using the disc diffusion method. We detected resistance to penicillin and ampicillin with the same rate of 89.7%. Thirty-one percent of the strains were multiresistant to three or more antibiotics. This study emphasizes that the presence of natural isolates of Bacillus spp. harboring one or more enterotoxin genes, producing extracellular enzymes which may cause spoilage and acquiring antibiotic resistance might hold crucial importance in the food safety and quality.

Occurrence and antimicrobial resistance profiles of Salmonella species in retail meat products.

In the present study, a total of 225 retail meat products (poultry meat, ground beef, and beef samples) were tested for the prevalence of Salmonella. Of these, 50 (22.2%) were positive for Salmonella. Overall, the pathogen was detected in 22 (29.3%) samples of poultry meat (n = 75), 16 (21.3%) samples of ground beef (n = 75), and 12 (16%) samples of beef (n = 75). The most common isolate was Salmonella enterica serovar Typhimurium (9.8%), followed by S. bongori species (8.9%) and S. enterica subsp. diarizonae (3.5%). The Salmonella strains isolated were also examined for antimicrobial resistance patterns and production of ß-lactamase enzyme. The resistance levels of the isolates against 14 different antimicrobial agents were tested by the disk diffusion method. None of the strains exhibited resistance to cefotaxime, ciprofloxacin, norfloxacin, or levofloxacin. However, the highest resistance rates in the meat isolates were 64% each for ampicillin and cephazolin and 56% for amoxicillin-clavulanic acid. A total of 62% of the 50 Salmonella strains were multiresistant to three or more antimicrobial agents. The exhibited multiple resistance to four or more antimicrobial drugs was 32%. Furthermore, none of the isolates had ß-lactamase enzyme activity.

Slime production and antibiotic susceptibility in staphylococci isolated from clinical samples.

A total of 187 isolates from several clinical specimens were identified to species level as 129 Staphylococcus aureus strains and 58 coagulase-negative staphylococci (CNS) strains by the API Staph System (Biomerieux). Slime production was detected both by the conventional Christensen's method as well as by the Congo red agar method. Seventy-two strains of staphylococci isolates (38.5%) were found to be slime producers by Christensen's test tube method whereas 58 strains (31%) were slime positive with Congo red agar method. There was no statistically significant difference between the two methods for the detection of slime production (P > 0.05). Susceptibility of isolates against antimicrobial agents was tested by the disk diffusion method. Staphylococcal species had resistance to one or more antibiotics. Among the various antimicrobial agents, oxacillin (71.1%) and erythromycin (47.1%) showed higher resistance than most of the agents used against all isolates. Oxacillin resistant S. aureus (ORSA) and oxacillin resistant coagulase-negative staphylococci (ORCNS), 97 (75.2%) and 36 (62.1%) respectively were frequently observed in strains isolated from clinical materials. Among the ORSA strains, two strains were resistant to vancomycin. Moreover, 96 (74.4%) of 129 S. aureus strains were positive for beta-lactamase enzyme. However, 78 (81.25%) of 96 beta-lactamase positive S. aureus strains were beta-lactamase positive ORSA isolates, but none of them had vancomycin resistance.

Slime production and antibiotic susceptibility in staphylococci isolated from clinical samples

A total of 187 isolates from several clinical specimens were identified to species level as 129 Staphylococcus aureus strains and 58 coagulase-negative staphylococci (CNS) strains by the API Staph System (Biomerieux). Slime production was detected both by the conventional Christensen's method as well as by the Congo red agar method. Seventy-two strains of staphylococci isolates (38.5 percent) were found to be slime producers by Christensen's test tube method whereas 58 strains (31 percent) were slime positive with Congo red agar method. There was no statistically significant difference between the two methods for the detection of slime production (P > 0.05). Susceptibility of isolates against antimicrobial agents was tested by the disk diffusion method. Staphylococcal species had resistance to one or more antibiotics. Among the various antimicrobial agents, oxacillin (71.1 percent) and erythromycin (47.1 percent) showed higher resistance than most of the agents used against all isolates. Oxacillin resistant S. aureus (ORSA) and oxacillin resistant coagulase-negative staphylococci (ORCNS), 97 (75.2 percent) and 36 (62.1 percent) respectively were frequently observed in strains isolated from clinical materials. Among the ORSA strains, two strains were resistant to vancomycin. Moreover, 96 (74.4 percent) of 129 S. aureus strains were positive for blactamase enzyme. However, 78 (81.25 percent) of 96 b-lactamase positive S. aureus strains were b-lactamase positive ORSA isolates, but none of them had vancomycin resistance.

Antimicrobial activity studies on some piperidine and pyrrolidine substituted halogenobenzene derivatives.

The in vitro antibacterial and antifungal activities of the compounds synthesised from some 1,2,3,5-tetrahalogeno benzenes in presence of sodium piperidide and sodium pyrrolidide (2,6-dipiperidino-1,4-dihalogenobenzenes; 2,6-dipyrrolidino-1,4-dibromobenzene; 2,4,6-tripyrrolidino chlorobenzene; and 1,3-dipyrrolidino benzene) were investigated. The in vitro antimicrobial activities were screened against the standard strains: Staphylococcus aureus ATCC 25923 and Bacillus subtilis ATCC 6633 as Gram positive, Yersinia enterocolitica ATCC 1501, Escherichia coli ATCC 11230 and Klebsiella pneumoniae as Gram negative, and Candida albicans as yeast-like fungus. Compounds (3, 5, 6, 7) inhibited the growth of all the test strains at MIC values of 32-512 microg/ml. None of the four compounds (1, 2,4,8) studied showed antimicrobial activity against any of the test strains within the MIC range 0.25-512 micro/ml.

Antimicrobial activities of N-(2-hydroxy-1-naphthalidene)-amino acid(glycine, alanine, phenylalanine, histidine, tryptophane) Schiff bases and their manganese(III) complexes.

The in vitro antibacterial and antifungal activities of five different amino acid Schiff bases derived from the reaction of 2-hydroxy-1-naphthaldehyde with glycine, L-alanine L-phenylalanine, L-histidine, L-tryptophane and the manganese(III) complexes of these bases were investigated. Structures of the Schiff bases were proven by 1H-NMR. In vitro activities against some Gram-positive (Staphylococcus aureus and Bacillus polymyxa) and Gram-negative (Escherichia coli) bacteria and the fungus Candida albicans were determined. The antimicrobial activities tended to decrease with the increasing size of the amino acid residues.