No abstract

Comlek peyniri is a typical artisanal cheese in Central Anatolia. This type of cheese was made by using the indigenous lactic acid bacteria (LAB) flora of cow or ewes' milk. Majority of the samples were taken from fresh cheese because the aim was to isolate homofermentative LAB. Initially 661 microbial isolates were obtained from 17 cheese samples. Only 107 were found to be homofermentative LAB. These isolates were selected and identified by using both phenotypic and molecular methods. Phenotypic identification included curd formation from skim milk, catalase test, Gram staining and light microscopy, growth at different temperatures and salt concentrations, arginine hydrolysis, gas production from glucose, and carbohydrate fermentation. Molecular identification was based on the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the 16S rRNA gene-ITS (internally transcribed spacer) region. By combining the phenotypic and molecular identification results, isolates belonging to each of the following genera were determined at species or subspecies level : 54 Lactococcus lactis subsp. lactis, 21 Enterococcus faecium, 3 Ec. faecalis, 2 Ec. durans, 10 Ec. sp., 15 Lactobacillus paracasei subsp. paracasei, and 2 Lb. casei strains. Technological characterisation was also performed by culturing each of the strains in UHT skim milk, and by monitoring pH change and lactic acid production at certain time intervals through the 24 h incubation. Results of the technological characterisation indicated that 33 % of the isolates (35 strains) were capable of lowering the pH of UHT milk below 5 . 3 after 6 h incubation at 30 8C. Thirty four of these strains were Lc. lactis subsp. lactis, and only one was an Ec. faecium strain.

a b s t r a c tStreptococcus thermophilus is a commonly used starter bacterium in dairy industry. It reduces the pH of milk rapidly and equilibrates the medium for the growth of Lactobacillus delbrueckii subsp. bulgaricus during yoghurt fermentation. Efforts to increase the diversity of artisanal yoghurt starters are not only important to bring new strains with novel and desirable characteristics, but also for the preservation of natural diversity which diminishes with the overuse and spread of industrial starters to natural resources. In the present study, 14 artisanal yoghurt samples were processed for the isolation of promising strains for yoghurt starter culture production and 66 strains were subsequently characterized. They were all identified as S. thermophilus using species-specific PCR and 16S rRNA gene sequencing. Genotypic diversity at the strain level was investigated by pulsed field gel electrophoresis (PFGE), and 22 homology groups were obtained. Further phenotypic characterization unearthed a significant phenotypic heterogeneity within homology groups, mostly with atypical novel character. Only 7 out of 66 strains showed S. thermophilus type-strain like phenotypic traits. Majority of the isolates were determined to be protease positive and fast milk acidifier to be used as yoghurt starter culture.

E . Y A V U Z , H . G U N E S , S . H A R S A A N D A . F . Y E N I D U N Y A . 2004.Aims: Molecular characterization of extracellular enzyme producing thermophilic bacilli from Balcova geothermal site. Methods and Results: Three types of geothermal samples were collected: mud, re-injection water, and samples from uncontrolled hydrothermal vents. Isolates grown at 55°C in culture media prepared in sterilized re-injection water, were screened for extracellular enzyme activity by using eight different substrates: casein, carboxymethylcellulose, pectin, polygalacturonic acid (PGA), soluble starch, Tween 20 and 80, and xylan. In total, 109 thermoaerophilic isolates were selected. All of the isolates could hydrolyse Tween 20 (100%) but not Tween 80. Soluble starch was hydrolysed by 96%, casein by 55%, xylan and carboxymethylcellulose by 9%, and pectin and PGA by 2% of the isolates. The isolates were grouped into 14 different homology groups by the restriction pattern analysis of 16S-internal transcribed spacer (ITS) rDNA RFLP. Each of the RFLP groups was also studied by 16S rRNA gene partial sequence analysis. Plasmid DNA profiles revealed that 15 of the isolated strains contained small plasmid DNA molecules ranging in size from 12 000 to 35 000 bp. Conclusions: Combined analysis of 16S-ITS rDNA RFLP and 16S rRNA gene partial sequence results indicated the presence of novel or existing species of Anoxybacillus (nine species) and Geobacillus (three species).

Poly[(maleic anhydride)‐co‐(vinyl acetate)] (MAVA) copolymer was synthesized by free radical polymerization reaction, in methyl ethyl ketone at 80 °C, using benzoyl peroxide as the initiator. The copolymer was then modified with a biomolecule, noradrenaline (NA). The modification reaction was performed at 70 °C in dimethylformamide containing triethylamine as the catalyst. The modified polymer was named MAVA/NA. Structural characterization of the copolymer and the modified product was carried out by Fourier transform infrared (FTIR) and 1H NMR and 13C NMR spectroscopy. The FTIR, 1H NMR and 13C NMR spectra confirmed that NA was successfully covalently bound to the MAVA copolymer backbone. Surface morphology was visualized by atomic force microscopy. The cumulative release of NA from MAVA/NA was determined in phosphate buffered saline solution for 7 days at 37 °C and compared with MAVA. Cytotoxicity of the MAVA/NA was evaluated by using a mouse fibroblast cell line (L929). Results obtained indicated that MAVA/NA had almost no toxicity and no negative effect on cell viability at 250 µg mL−1 concentration. © 2012 Society of Chemical Industry

No abstract

SummaryThe 16S-ITS (internal transcribed spacer) region of the rrn operon was amplified by polymerase chain reaction (PCR). The amplification products were analysed by restriction fragment length polymorphism (RFLP) using a set of restriction enzymes, AluI, HaeIII, and TaqI. Restriction pattern analyses revealed that TaqI restriction enzyme could clearly differentiate the nine reference strains of Lactobacillus used in the study.

The capacity of 16S internal transcribed spacer (16S-ITS) rRNA gene RFLP to differentiate 16 type strains and nine clinical isolates of staphylococci was evaluated. The 16S rRNA gene was amplified together with the ITS region and the amplification products were digested with TaqI restriction enzyme. Analysis of the 16S-ITS rRNA gene RFLP profiles differentiated each of the 16 type strains into distinct RFLP haplotypes. INTRODUCTIONThe habitat of staphylococci is skin, skin glands and mucous membranes of humans and many animals. The genus Staphylococcus includes both pathogenic and saprophytic strains that have been isolated from animal products such as meat and milk, and from environmental sources such as soil, sea water, fresh water, dust and air samples (Kloos et al., 1991).Coagulase-positive species such as Staphylococcus aureus are the cause of many types of infection (Forbes et al., 1998). During the last two decades, coagulase-negative staphylococci (CNS) have also emerged as pathogens causing medical-device-related infections (von Eiff et al., 2002). Because of the pathogenic potential of these bacteria, effective methods are required for their in vitro identification. Beside fatty acid analyses, many diagnostic tests rely on a miniaturized phenotypic characterization and a set of biochemical reactions. These methods enable the identification of S. aureus isolates, but often fail in the identification of CNS (Stoakes et al., 1994;Renneberg et al., 1995;Martineau et al., 1996). Analysis of specific regions of genomic DNA, on the other hand, has produced much more discriminative data. For example, several genomic targets have been effectively used for the identification of Staphylococcus species, including the 16S rRNA gene (Bialkowska-Hobrzanska et al., 1990;De Buyser et al., 1992), the tRNA gene intergenic spacer (Maes et al., 1997), the internal transcribed spacer (Couto et al., 2001), the heat-shock protein 60 (HSP60) gene (Goh et al., 1996), the chaperonin 60 gene (Goh et al., 1997), the femA gene (Vannuffel et al., 1999), the sodA gene (Poyart et al., 2001), the gap gene (Yugueros et al., 2000) and the nuc gene (Brakstad et al., 1992). Recently, an enterobacterial repetitive intergenic consensus PCR and BOX-PCR were also used in the identification of Staphylococcus epidermidis strains (Wieser & Busse, 2000). . The nine clinical isolates were S. capitis subsp. capitis (n ¼ 2), S. caprae (n ¼ 2), S. cohnii subsp. cohnii (n ¼ 1) and S. epidermidis (n ¼ 4). The clinical isolates were identified by basic phenotypic tests and the Staph ID 32 system (bioMérieux). All strains were cultured in tryptic soy broth (TSB; Merck) at 37 8C and stored in 25 % (v/v) glycerol at À80 8C.16S-ITS rRNA gene PCR. Genomic DNA was isolated as described by Arciola et al. (2001). Briefly, 100 ìl of overnight culture prepared in 5 ml TSB at 37 8C with shaking was pelleted by centrifugation. Cell pellets were resuspended in 45 ìl deionized water and 5 ìl lysostaphin solution (100 ìg ml À1 in dH 2 O; Sigma) and incubated for 10 min...