Investigation of some changes and clonal relationship in enterococci isolates due to relocation of a hospital.

Objectives: To investigate the isolation rates, antimicrobial resistance rates, minimum inhibitory concentration values of antimicrobial agents, and clonal relationships of Enterococcus faecalis and Enterococcus faeciumdue to the relocation of a hospital to a newly constructed building. METHODS: The comparative, prospective study was conducted at adult general intensive care units of the Mus State Hospital, Mus, Turkey, in two phases; before the relocation from January 25 to December 1, 2014, and after the relocation from February 10 to May 24, 2015. Rectal swab samples were collected 72 hours post-hospitalisation. Identification of Enterococcus faecalis and Enterococcus faeciumisolates was determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and antimicrobial resistance with minimum inhibitory concentration values was detected with Vitek 2 system. The clonal relatedness among the strains was investigated by pulsed-field gel electrophoresis. Data was analysed using SPSS 23. RESULTS: Of the 69 patients, 37(53.62%) were related to pre-relocation phase; 20(54.1%) females and 17(45.9%) males with mean age 62.81±21.71 years. There were 32(46.37%) patients in the post-relocation phase; 13(40.6%) females and 19(59.4%) males with mean age 62.69±21.35 years (p>0.05). Of the 84 enterococci strains isolated, 51(60.7%) were Enterococcus faecium; 28(55%) before relocation and 23(45%) after relocation (p=0.77). The remaining 33(39.3%) isolates were Enterococcus faecalis; 16(48.5%) before relocation and 17(51.5%) after relocation (p=0.73). Multiple strains were located in 7(18.9%) patients before relocation and in 7(21.9%) after relocation. In 1(3.1%) patient after relocation, 2(8.7%) Enterococcus faecium isolates with different resistance and pulsed-field gel electrophoresis patterns were detected. There were no significant differences between the isolation and antibiotic resistance rates before and after relocation (p>0.05), and a clonal relation between the isolates was not detected (p>0.05). Decreased minimum inhibitory concentration values were noted for some antibiotics. CONCLUSIONS: Clonal relationship between the isolates and change in the rates of isolation and antimicrobial resistance of Enterococcus faecalis and Enterococcus faecium was not detected due to relocation. Minimum inhibitory concentration values could be used to reveal relocation-related changes in isolates obtained from patients hospitalised in intensive care units.

Rapid method for detection of Vibrio cholerae from drinking water with nanomaterials enhancing electrochemical biosensor.

Cholera is a potentially fatal diarrheal disease caused by Vibrio cholerae and is spread to humans from contaminated food and water. In order to prevent spread of epidemic chlorea, the development of novel sensitive, selective, user-friendly, cost-effective and rapid detection systems to detect of V. cholerae are necessary. Therefore, in this study, it was aimed to develop a specific, electrochemical immunoassay with high selectivitiy and sensitivity for detection of V. cholerae from drinking water using in house synthesized Gold Nanoparticles (AuNPs). The synthesized AuNPs were characterized by UV/Vis spectroscopy, Dynamic Light Scattering (DLS) and Atomic Force Microscopy (AFM) and electrochemical techniques were applied to confirm the succesful fabrication of the immunosensor. Also, this study focuses on the development of an antibody sensor for V. cholerae detection using a standard immunoassay without using nanoparticle. To accomplish that, in house spherical synthesized AuNPs at various sizes were synthesized, conjugated with secondary antibody-horseradish peroxidase enzyme (HRP) complex and their possible effect on the lowest detection limit of V. cholerae was investigated in comparison to commercially available AuNPs. The AuNPs-immunosensor on the results enabled the quantification of V. cholerae in a wide concentration range with a high sensitivity limit of detection (1 Colony-Forming Units/Milliliter) and specificity. Although the effect of 33 and 54 nm AuNPs on the process is close to each other, it has been observed that there is a 34% loss of efficiency when the size of the nanoparticle increases. With this study, a novel V. cholerae specific immunosensor was developed and the effects of in house synthesized AuNPs with various diameter on this developed biosensor were investigated in detail.

Molecular Typing of Different Salmonella Serotypes by Multiple-Locus Analysis of Tandem Repeats.

Salmonella enterica subspecies enterica causes salmonellosis in humans and animals and is an important cause of food infections worldwide. In recent years, the multiple-locus variable-number of tandem repeat (VNTR) analysis (MLVA), a fast molecular typing method with strong epidemiological discrimination, has facilitated the effective control of diverse infections. This study aimed at the typing of 28 human origined Salmonella enteritidis, Salmonella infantis, and Salmonella typhimurium strains by using a single MLVA protocol. Previously these strains have been identified by pulsed field gel electrophoresis (PFGE) method and it has been shown that each strain produced a distinct PFGE banding profile. One MLVA protocol was tested on 3 serotypes simultaneously and it produced three banding patterns specific to each of the three common Salmonella serotypes. MLVA also constitute a relatively more cost-effective and faster method than PFGE.

Esterification of Fructose-oleic Acid by tert-Butanol/Dimethyl Sulfoxide and by 2-Methyl-2-butanol/Dimethyl Sulfoxide.

In this study two different strategy were followed to obtain a D-fructose-oleic acid ester. One of the strategies has been well established enzymatic synthesis of an ester bond. The other strategy excluded the biocatalyst and only used a mixture of two organic solvents as the reaction media, 2-methyl-2-butanol / dimethyl sulfoxide or tert-butanol / dimethyl sulfoxide for the production of D-fructose-oleic acid ester. Ester products obtained were characterised by using FT-IR, NMR, by MS. Product yield was also assessed by HPLC. Results of structural analyses and yield measurement indicated that two approaches produced almost identical ester products.

Graphene Quantum Dots as Nanozymes for Electrochemical Sensing of Yersinia enterocolitica in Milk and Human Serum.

The genus Yersinia contains three well-recognized human pathogens, including Y. enterocolitica, Y. pestis, and Y. pseudotuberculosis. Various domesticated and wild animals carry Yersinia in their intestines. Spread to individuals arises from eating food or water contaminated by infected human or animal faeces. Interaction with infected pets and domestic stock may also lead to infection. Yersinia is able to multiply at temperatures found in normal refrigerators; hence, a large number of the bacteria may be present if meat is kept without freezing. Yersinia is also rarely transmitted by blood transfusion, because it is able to multiply in stored blood products. Infection with Yersinia can cause yersiniosis, a serious bacterial infection associated with fever, abdominal pain and cramps, diarrhea, joint pain, and symptoms similar to appendicitis in older children and adults. This paper describes a novel immunosensor approach using graphene quantum dots (GQDs) as enzyme mimics in an electrochemical sensor set up to provide an efficient diagnostic method for Y. enterecolitica. The optimum assay conditions were initially determined and the developed immunosensor was subsequently used for the detection of the bacterium in milk and human serum. The GQD-immunosensor enabled the quantification of Y. enterocolitica in a wide concentration range with a high sensitivity (LODmilk = 5 cfu mL-1 and LODserum = 30 cfu mL-1) and specificity. The developed method can be used for any pathogenic bacteria detection for clinical and food samples without pre-sample treatment. Offering a very rapid, specific and sensitive detection with a label-free system, the GQD-based immunosensor can be coupled with many electrochemical biosensors.

From days to hours: Can MALDI-TOF MS system replace both conventional and molecular typing methods with new cut off level for Vancomycin Resistant Enterococcus faecium.

Vancomycin-Resistant E. faecium (VRE) strains from clinical specimens were identified by conventional methods before. Following the phenotype-based identification, all strains were also identified using both BD Phoenix and VITEK MS bioMérieux System. Strains were typed with the Bruker MALDI-TOF MS system, pulsed field gel electrophoresis (PFGE) and 16S rRNA gene sequencing analysis and then the sensitivity compared for each. A cut off value of 850 assigned with Bruker MALDI-TOF MS system was found to give equal sensitivity to that of PFGE. Results obtained were compared with those of molecular typing. The main advantage of MALDI-TOF MS technology over the others was the much shorter analysis time which lasted only a few hours rather than days or a whole week. Also, the Bruker MALDI-TOF MS system was used for typing and compared with the gold standard method and this study is first to report the determined cut off level for typing of VRE strains.

Nanoparticle Enhanced Antibody and DNA Biosensors for Sensitive Detection of Salmonella.

Bacteria-related pathogenic diseases are one of the major health problems throughout the world. Salmonella is a genus of rod-shaped Gram-negative enterobacteria of which more than 2600 serotypes have been identified. Infection with Salmonella can cause salmonellosis, a serious bacterial toxi-infection syndrome associated with gastroenteritis, and paralyphoid and typhoid fevers. Its rapid and sensitive detection is a key to the prevention of problems related to health. This paper describes the development of antibody and DNA sensors for Salmonella detection using a microfluidic-based electrochemical system. Commercial Salmonella typhimurium and Salmonella typhimurium from human stool samples were investigated using standard and nanomaterial-amplified antibody sensors. S. typhimurium could be detected down to 1 cfu mL-1. The specificity of immunoassay was tested by studying with non-specific bacteria including E. coli and S. aureus that revealed only 2.01% and 2.66% binding when compared to the target bacterium. On the other hand, the quantification of Salmonella DNA was investigated in a concentration range of 0.002⁻200 µM using the developed DNA biosensor that demonstrated very high specificity and sensitivity with a detection limit of 0.94 nM. Our custom-designed microfluidic sensor offers rapid, highly sensitive, and specific diagnostic assay approaches for pathogen detection.

[Comparison of MALDI-TOF and 16S rRNA methods in identification of viridans group streptococci]. / Viridans grup streptokok tanimlamasinda MALDI-TOF ve 16S rRNA yöntemlerinin karsilastirilmasi.

Accurate identification of viridans group streptococci (VGS) frequently encountered as a causative agent of infective endocarditis is always a challenge for the clinical microbiology laboratory. Clinical microbiology laboratories generally use semi automatic/full automatic systems, molecular methods and also conventional methods for the identification of these bacteria. There are recent published studies that have used MALDI-TOF (Matrix Assisted Laser Ionization Mass Spectrometry-Time of Flight) systems in the identification of VGS. The aim of the study was to compare the performance of the conventional methods, semi automatic and MALDI-TOF MS system used in identification of VGS in oral microbiota of persons under the risk of infective endocarditis, with the gold standard method 16S rRNA sequence analysis and to create a diagnosis algorithm for the identification of VGS in clinical microbiology laboratories according to the obtained data.The study was conducted with 51 VGS strains isolated from oral microbiota of the patients with rheumatologic cardiac, valve and/or prosthetic valve diseases, under the risk of development of infective endocarditis, who have admitted to Ankara Numune Training and Research Hospital, Department of Cardiology, between February-June 2015. Standard microbiology procedures, optochin susceptibility and bile solubility tests were done for the isolation of bacteria. Bacteria were also identified with APISTREP (bioMérieux, France) and MALDI-TOF MS Bruker Microflex (Bruker Biotyper; Bruker Daltonics, Bremen, Germany) methods. BSF-8 (5´-AGAGTTTGATCCTGGCTCAG-3´) and BSR-534(5´-ATTACCGCGGCTGCTGGC-3´) primers were used in the 16S rRNA sequence analysis of bacteria. ABI PRISM 3100 Avan t Genetic Analyzer (Applied Biossytems, Foster City, CA, USA) were used for the sequence analysis. Electropherograms were analyzed in SeqScape Software (Applied Biosystems, Foster City, CA, USA) and compared with the reference sequences in GenBank with BLASTN (NCBI). According to the result of optochin and bile solubility tests, with API STREP system, 16 (31,37%) of the isolates were identified as Mitis group, 15 (29.41%) as Anginosus group, 9 (17.5%) as Salivarius group, 7 (13,73%) as Sanguinis group and 4 (7.84%) as Bovis group among optochin and bile resistant alpha hemolytic streptococci. Moreover, of the same isolates 20 (39.22%) were identified as Mitis group, 14 (27.45%) as Anginosus group, 13 (25.49%) as Salivarius group and 4 (7.84%) as Sanguinis group with MALDI-TOF system. In the identification with 16S rRNA, 25 (49.02%) of the isolates were identified as Mitis group, 13 (25.49%) as Anginosus group, 12 (23.53%) as Salivarius group and 1 (1.96%) as Sanguinis group. According to the results, it was determined that 33 (64.70%) of the isolates identified in MALDI-TOF MS system and 31 (60.78%) of the isolates identified in API STREP system were compatible with 16S rRNA sequence analysis method. For Mitis group, API STREP test sensitivity was 48.00% and specificity was 84.62% and MALDI-TOF system sensitivity was 80.00% and specificity was 100%. As VGS identification is a complicated process, we believe a single method will be insufficient for the identification of these isolates in clinical microbiology laboratories. We suggest that MALDI-TOF system can be used for VGS diagnosis, however, optochin test and/or molecular methods should also be included in the diagnosis algorithm when necessary.