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Background This study aimed to investigate the genetic diversity of OXA-51-like, OXA-23-like, OXA-24, and OXA-58-like genes and the role of β-lactamases in carbapenem resistance among multidrug resistant Acinetobacter baumannii strains recovered from patients in intensive care units (ICUs). Methods Non-duplicate clinical isolates of A. baumannii from ICUs that were identified as imipenem and meropenem resistant were collected. Antimicrobial susceptibilities were determined by PhoenixTM system (Becton Dickinson, USA). Minimum inhibitory concentrations (MICs) for imipenem and meropenem were determined by using gradient strip method (E-test) and interpreted according to CLSI. Presence of carbapenemase activity was determined by the modified Hodge test (MHT) and detection of metallo-β-lactamase (MBL) was performed by the double-disk synergy test (DDST) and MBL E-test. Detection of the four groups of OXA carbapenemase genes (OXA-23, OXA-24, OXA-51, and OXA-58) was carried out using a multiplex PCR assay. Sequencing of the products in both directions was performed by ABI 3130XL genetic analyzer (Life Technologies Corporation, CA, USA). The resulting DNA sequence was analyzed by the BLAST program, available at the NCBI website. Results Sixty-one non-duplicate, multidrug resistant clinical A. baumannii isolates were studied. MHTs were positive for all 61 A. baumannii strains, but none of them showed MBL activity. As determined through multiplex PCR, all of the 61 isolates had blaOXA-51 genes including blaOXA-64, blaOXA-66, and blaOXA-91, 50 isolates had blaOXA-23, and 11 isolates had blaOXA-58 genes. Alleles encoding OXA-24-like enzymes were not detected in any isolates. Conclusions This study indicated that the major cause of carbapenem resistance in our region was OXA-type car-bapenemase encoded by blaOXA-51, blaOXA-23, and blaOXA-58 genes and as we know, this is the first report from Turkey identifying blaOXA-51-like sequences.

WOS:000564336200005 PubMed: 32800227 Heavy metal acclimated bacteria are profoundly the preferred choice for bioremediation studies. Bacteria get acclimated to toxic concentrations of heavy metals by induction of specific enzymes and genetic selection favoring new metabolic abilities leading to activation of one or several of resistance mechanisms creating bacterial populations with differences in resistance profile and/or level. Therefore, to use in bioremediation processes, it is important to discriminate acclimated bacterial populations and choose a more resistant strain. In this study, we discriminated heavy metal acclimated bacteria by using Attenuated Total Reflectance Fourier Transform Infrared (ATR-FTIR) spectroscopy and multivariate analysis methods namely Hierarchical Cluster Analysis (HCA), Principal Component Analysis (PCA) and Soft Independent Modeling of Class Analogy (SIMCA). Two acclimation methods, acute and gradual, were used which cause differences in molecular changes resulting in bacterial populations with different molecular and resistance profiles. Brevundimonas sp., Gordonia sp., and Microbacterium oxydans were exposed to the toxic concentrations of Cd (30 mu g/ml) or Pb (90 mu g/ml) by using broth medium as a growth media. Our results revealed that PCA and HCA clearly discriminated the acute-acclimated, gradual acclimated, and control bacteria from each other in protein, carbohydrate, and whole spectral regions. Furthermore, we classified acclimated (acute and gradual) and control bacteria more accurately by using SIMCA with 99.9% confidence. This study demonstrated that heavy metal acclimated and control group bacteria can be discriminated by using chemometric analysis of FTIR spectra in a powerful, cost-effective, and handy way. In addition to the determination of the most appropriate acclimation procedure, this approach can be used in the detection of the most resistant bacterial strains to be used in bioremediation studies. Environment, Atmosphere, Earth and Marine Sciences Research Group (CAYDAG) of the Scientific and Technological Research Council of Turkey (TUBITAK)Turkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [113Y515] This work was supported by the Environment, Atmosphere, Earth and Marine Sciences Research Group (CAYDAG) of the Scientific and Technological Research Council of Turkey (TUBITAK) (Project No. 113Y515).

The use of biological means for ground improvement have become popular, which generally works through the process called microbially-induced calcium carbonate precipitation (MICP). Many studies ind...

Gozen, Ayse Gul/0000-0001-5479-051X WOS:000467052400013 PubMed: 30578608 In the environment, bacteria can be exposed to the concentration gradient of toxic heavy metals (gradual) or sudden high concentration of them (acute). In both situations, bacteria get acclimated to toxic heavy metal concentrations. Acclimation causes metabolic and molecular changes in bacteria. In this study, we aimed to understand whether there are differences between molecular profiles of the bacteria (Brevundimonas, Gordonia and Microbacterium) which are under acute or gradual exposure to cadmium or lead by using ATR-FTIR spectroscopy. Our results revealed the differences between the acclimation groups in membrane dynamics including changes in the structure and composition of the membrane lipids and proteins. Furthermore, protein concentrations decreased in acclimated bacterial groups. Also, a remarkable increase in exopolymer production occurred in acclimated groups. Interestingly, bacteria under acute cadmium exposure produced the significantly higher amount of exopolymer than they did under gradual exposure. On the contrary, under lead exposure gradually acclimate strains produced significantly higher amounts of exopolymer than those of acutely acclimated ones. This information can be used in bioremediation studies to obtain bacterial strains producing a higher amount of exopolymer.

Gozen, Ayse Gul/0000-0001-5479-051X; Tuncer, Sinem/0000-0002-8947-8646; Mingu-Akmete, Sara/0000-0002-7332-0764 WOS:000486357300014 PubMed: 31394353 Heavy metal acclimation of bacteria is of particular interest in many aspects. It could add to our understanding of adaptation strategies applied by bacteria, as well as help us in devising ways to use such adaptive bacteria for bioremediation. In this study, we have explored the changes in the DNA of an aquatic Gordonia sp. acclimated to silver, cadmium, and lead. We have measured the changes in the DNA extracted from the acclimated bacteria by using ATR-FTIR coupled with unsupervised and supervised pattern recognition algorithms. Although whole-cell FTIR studies do reveal nucleic acid changes, the special care should be taken when considering marker nucleic acid bands in such spectra, as various other cell or tissue constituents also yield IR bands in the same region. An FTIR study on isolated DNA can be used to avoid this problem. The IR spectral profiles of the DNA molecules revealed significant changes in the backbone and sugar conformations of upon acclimation. We then further analyzed the DNA's global cytosine-methylation patterns of the heavy metal-acclimated bacteria. We aimed to find out whether epigenetic mechanisms operate in bacteria for survival and growth in inhibitory heavy metal concentrations or not. We found hypermethylation in Cd acclimation but hypomethylation for both Pb and Ag in Gordonia sp. Our results imply that changes in the conformational and methylation states of DNA seem to let bacteria to thrive in otherwise inhibitory conditions and mark the involvement of epigenetic modulation in acclimation processes. Middle East Technical University Scientific Research Project Coordination Unit [BAP-01-08-2017-003] This study was supported by the Middle East Technical University Scientific Research Project Coordination Unit (Project No: BAP-01-08-2017-003).

Recent studies carried on germ –free (GF) animal models suggest that the gut microbiota (GM) may play a role in the regulation of anxiety, mood, and cognitive abilities such as memory and learning processes. Consistently, any treatment disturbing the gut microbiota, including the overuse of antibiotics, may influence the brain functions and impact behavior. In the present study, to address this issue, two wide-spectrum antibiotics (ampicillin and cefoperazone, 1 g/l) were repeatedly applied throughout a 6-week period to initially 21-day-old male BALB/c mice. Antibiotics were administered separately or in a mixed fashion. On the completion of the antibiotic treatment, all mice were subjected to the behavioral tests. The serum levels of corticosterone and brain-derived neurotropic factor (BDNF) were assessed. Gut microbiota profiles were obtained by using denaturing gradient gel electrophoresis system, DGGE, from fecal samples. Ampicillin had a greater impact on both, gut microbiota composition and mice behavior compared to cefoperazone. All antibiotic-treated groups manifested a decrease in the locomotor activity and reduced recognition memory. However, the ampicillin-treated groups showed a higher anxiety level as assessed by the open field and the elevated plus maze tests and an increased immobility (behavioral despair) in the forced swim test. Obtained results evidently show that in mice, a repeated antibiotic treatment applied during adolescence, parallel to the changes in GM, affects locomotor activity, affective behavior and cognitive skills in young adults with ampicillin specifically enhancing anxiety- and depressive-like responses. Lower levels of serum BDNF were not associated with cognitive impairment but with changes in affective-like behaviors. Repeated administration of neither ampicillin nor cefoperazone affected basal serum corticosterone levels. This is one of the few studies demonstrating changes in a behavioral phenotype of young-adult subjects who were previously exposed to a repeated antibiotic treatment. © 2018 BAP-01-08-2016005-16 The study was supported by the METU Scientific Research Found , BAP-01-08-2016005-16 , to AG Gozen.

Gozen, Ayse Gul/0000-0001-5479-051X WOS:000413607000038 PubMed: 28823969 Rapid, cost-effective, sensitive and accurate methodologies to classify bacteria are still in the process of development. The major drawbacks of standard microbiological, molecular and immunological techniques call for the possible usage of infrared (IR) spectroscopy based supervised chemometric techniques. Previous applications of IR based chemometric methods have demonstrated outstanding findings in the classification of bacteria. Therefore, we have exploited an IR spectroscopy based chemometrics using supervised method namely Soft Independent Modeling of Class Analogy (SIMCA) technique for the first time to classify heavy metal-exposed bacteria to be used in the selection of suitable bacteria to evaluate their potential for environmental cleanup applications. Herein, we present the powerful differentiation and classification of laboratory strains (Escherichia coli and Staphylococcus aureus) and environmental isolates (Gordonia sp. and Microbacterium oxydans) of bacteria exposed to growth inhibitory concentrations of silver (Ag), cadmium (Cd) and lead (Pb). Our results demonstrated that SIMCA was able to differentiate all heavy metal-exposed and control groups from each other with 95% confidence level. Correct identification of randomly chosen test samples in their corresponding groups and high model distances between the classes were also achieved. We report, for the first time, the success of IR spectroscopy coupled with supervised chemometric technique SIMCA in classification of different bacteria under a given treatment. (C) 2017 Elsevier B.V. All rights reserved. Scientific and Technological Research Council of TurkeyTurkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [113Y515]; Scientific Research Projects Fund (BAP) of the Middle East Technical UniversityMiddle East Technical University [BAP-01-08-2016-005] The Scientific and Technological Research Council of Turkey (Grant 113Y515) and The Scientific Research Projects Fund (BAP) of the Middle East Technical University (Grant BAP-01-08-2016-005) supported this work. The authors would like to thank Dr. Nihal Simsek Ozek for her valuable comments on the manuscript

Lead and cadmium are frequently encountered heavy metals in industrially polluted areas. Many heavy metal resistant bacterial strains have a high biosorption capacity and thus are good candidates for the removal of toxic metals from the environment. However, as of yet there is no accurate method for discrimination of highly adaptive bacterial strains among the populations present in a given habitat. In this study, we aimed to find distinguishing molecular features of lead and cadmium resistant bacteria using Attenuated Total Reflectance Fourier Transformed Infrared (ATR-FT-IR) spectroscopy and chemometric approaches. Our results demonstrated that both control and metal exposed E. coli and S. aureus strains could be successfully discriminated from each other using hierarchical cluster and principal component analysis methods. Moreover, we found that lead exposed bacterial strains could be successfully discriminated from cadmium exposed ones with a high heterogeneity value. These clear discriminations can be described by the ability of a bacterium to change its metabolism in terms of the content and structure of cellular macromolecules under heavy metal stress. In our case, cadmium and lead-induced genetic response systems in bacteria caused remarkable alterations in overall cellular metabolism. Bacteria deal with a heavy metal stress by altering nucleic acid methylations and lipid and protein synthesis. Heavy metal burden led to the development of relevant metabolic changes in proteins, lipids, and nucleic acids of the resistant bacteria described in this study. Our approach showed that infrared spectra obtained via ATR-FT-IR spectroscopy coupled with chemometric analysis can be utilized for rapid, low-cost, informative, reliable, and operator-independent discrimination of resistant bacterial populations.

Gozen, Ayse Gul/0000-0001-5479-051X; Tuncer, Sinem/0000-0002-8947-8646; WOS:000432008800009 PubMed: 29243883 In this study, the molecular profile changes leading to the adaptation of bacteria to survive and grow at inhibitory silver concentration were explored. The profile obtained through infrared (IR)-based measurements indicated extensive changes in all biomolecular components, which were supported by chemometric techniques. The changes in biomolecular profile were prominent, including nucleic acids. The changes in nucleic acid region (1350-950 cm(-1)) were encountered as a clue for conformational change in DNA. Further analysis of DNA by IR spectroscopy revealed changes in the backbone and sugar conformations. Moreover, Enzyme-Linked Immunosorbent Assay-based measurements of DNA methylation levels were performed to see if epigenetic mechanisms are in operation during bacterial adaptation to this environmental challenge. The results indicated a notable demethylation in Escherichia coli and methylation in Staphylococcus aureus likely to be associated with their elaborate adaptation process to sustain survival and growth. TubitakTurkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [113Y515] Tubitak, Grant/Award number: 113Y515

The identification of foodborne microorganisms and their endospores in food products are important for food safety. The present work compares Bacillus ( Bacillus licheniformis , Bacillus circulans and Bacillus subtilis ) and Micrococcus ( Micrococcus luteus ) species with Fourier transform infrared (FTIR) spectroscopy. Our results show that there are several characteristic peaks belonging to both the Micrococcus and Bacillus species which can be used for the identification of these foodborne bacteria and their endospores. For Micrococcus species, a new band was observed at 1338 cm −1 which may be due to acetate oxidation via the carboxylic acid cycle. The bands at 1313 cm −1 and 1256 cm −1 can be explained by an exopolymer formation and the other bands at 1074 cm −1 and 550 cm −1 , may be due to the glycogen-like storage material in Micrococcus spp. There are also characteristic peaks at 993 cm −1 and 801 cm −1 for these bacterial species. Different Bacillus species also showed characteristic peaks at 1000–500 cm −1 region. Dipicolinic acid (DPA) bands at ∼728 cm −1 and ∼703 cm −1 seen only in B. circulans were the marker of an endospore formation.

High concentrations of heavy metals can be toxic for bacteria. However, after prolonged exposure, bacteria can become acclimated and begin to be able to grow in the presence of heavy metals. Acclimation can involve alterations of metabolism and molecular structures. Our aim was to examine these alterations in cobalt-acclimated bacteria via attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy on viable samples. Bacillus sp. and Pseudomonas sp. isolated from a temperate shallow lake and a well-established strain of E. coli were investigated. Our results revealed consistent, wide-spread changes in cell membrane and cell wall dynamics of Bacillus sp. and E. coli, including a decrease in peptidoglycan content of Bacillus sp. and increased lipid ordering of the membrane in both bacteria. Furthermore, a decrease in RNA and protein concentrations of Bacillus sp. was measured. All three bacteria studied showed a decrease in conformational freedom of proteins following cobalt acclimation. Interestingly, both Bacillus sp. and E. coli showed slight but significant alterations in their DNA conformations which might imply a methylation-mediated memory formation leading to epigenetic modulation for cobalt adaptation.

Gozen, Ayse Gul/0000-0001-5479-051X WOS: 000312972200020 PubMed: 23039919 Freshwater fish, Alburnus alburnus (bleak), were captured from Lake Mogan, situated in Ankara, during spring. The surface mucus of the fish was collected and associated bacteria were cultured and isolated. By sequencing PCR-amplified 16S RNA encoding genes, the isolates were identified as members of 12 different genera: Acinetobacter, Aeromonas, Bacillus, Brevundimonas, Gordonia, Kocuria, Microbacterium, Mycobacterium, Pseudomonas, Rhodococcus, and Staphylococcus, in addition to one strain that was unidentified. The mucus-dwelling bacterial isolates were tested for resistance against ampicillin, kanamycin, streptomycin and chloramphenicol. About 95% of the isolates were found to be resistant to ampicillin, 93% to chloramphenicol, and 88% to kanamycin and streptomycin. A Microbacterium oxydans and the unidentified environmental isolate were resistant to all four antibiotics tested at very high levels (>1600 mu g/ml ampicillin and streptomycin; >1120 mu g/ml kanamycin; >960 mu g/ml chloramphenicol). Only a Kocuria sp. was sensitive to all four antibiotics at the lowest concentrations tested (3.10 mu g/ml ampicillin and streptomycin; 2.15 mu g/ml kanamycin; 1.85 mu g/ml chloramphenicol). The rest of the isolates showed different resistance levels. Plasmid isolations were carried out to determine if the multiple antibiotic resistance could be attributed to the presence of plasmids. However, no plasmid was detected in any of the isolates. The resistance appeared to be mediated by chromosome-associated functions. This study indicated that multiple antibiotic resistance at moderate to high levels is common among the current phenotypes of the fish mucus-dwelling bacterial populations in this temperate, shallow lake which has not been subjected to any aquaculturing so far but under anthropogenic effect being in a recreational area. (C) 2012 Elsevier Ltd. All rights reserved. Middle East Technical UniversityMiddle East Technical University [BAP-08-11-DPT-2002K120510] This work was supported by the Faculty Development Program (OYP) of the Middle East Technical University (grant BAP-08-11-DPT-2002K120510). The authors thank Ms. Gul Inan M.Sc. for her help with statistical analyses and Mr. Robert W. Murdoch Ph.D. for his help with language editing.

Quantitative differentiation of live cells in biosolids samples, without the use of culturing-based approaches, is highly critical from a public health risk perspective, as recent studies have shown significant regrowth and reactivation of indicator organisms. Persistence of DNA in the environment after cell death in the range of days to weeks limits the application of DNA-based approaches as a measure of live cell density. Using selective nucleic acid intercalating dyes like ethidium monoazide (EMA) and propidium monoazide (PMA) is one of the alternative approaches to detecting and quantifying viable cells by quantitative PCR. These compounds have the ability to penetrate only into dead cells with compromised membrane integrity and intercalate with DNA via their photoinducible azide groups and in turn inhibit DNA amplification during PCRs. PMA has been successfully used in different studies and microorganisms, but it has not been evaluated sufficiently for complex environmental samples such as biosolids. In this study, experiments were performed with Escherichia coli ATCC 25922 as the model organism and the uidA gene as the target sequence using real-time PCR via the absolute quantification method. Experiments with the known quantities of live and dead cell mixtures showed that PMA treatment inhibits PCR amplification from dead cells with over 99% efficiency. The results also indicated that PMA-modified quantitative PCR could be successfully applied to biosolids when the total suspended solids (TSS) concentration is at or below 2,000 mg·liter −1 .

A set of new aziridinyl phosphonates (4a-g) were synthesized by using the Gabriel-Cromwell reaction and its modified version developed in this study and their structures confirmed by HRMS, IR. and NMR spectra. All the compounds were screened for their antibacterial activity. They all showed comparable moderate to good growth inhibitory activity in reference to ampicillin and streptomycin. (C) 2011 Elsevier Masson SAS. All rights reserved.

Vertical migration of Daphnia represents the best-studied predator-avoidance behaviour known; yet the mechanisms underlying the choice to migrate require further investigation to understand the role of environmental context. To investigate the optimal habitat choice of Daphnia under fish predation pressure, first, we selected the individuals exhibiting strong migration behaviour. The animals collected from the hypolimnion during the daytime were significantly larger, being more conspicuous, and in turn performed stronger diel vertical migration (DVM) when exposed to fish cue. We called them strong migrants. Second, we provided the strong migrant D. pulex with food at high and intermediate (1 and 0.4 mg C l(-1), respectively) levels, which were well above the incipient limiting level and of high quality. They traded the benefits of staying in the warm water layer and moved down to the cold water in response to fish cue indicating fish predation. The availability of food allowed the animals to stay in the cold hypolimnion. However, at the low food level (0.1 mg C l(-1)), which is an additional constraint on fitness, Daphnia moved away from the cold hypolimnion. Poor food condition resulted in strong migrant Daphnia to cease migration and remain in the upper warmer water layer. Although temperature is known to be a more important cost factor of DVM than food, our results clearly show that this is only true as long as food is available. It becomes clear that food availability is controlling the direction of vertical positioning when daphnids experience a dilemma between optimising temperature and food condition while being exposed to fish cue. Then they overlook the predation risk. Thus, the optimal habitat choice of Daphnia appears to be a function of several variables including temperature, food levels and fish predation.

Summary 1. Bacterial populations associated with fish have previously been documented to be crucial for the production of chemical signals governing the interactions between predator fish and zooplankton prey. 2. In this study, we investigated the roles of fish and mucus-dwelling bacteria in kairomone production by conducting two sets of experiments related to elimination of bacteria with antibiotics and using fish mucus in bioassays of Daphnia pulex’s diel vertical migration. 3. Daphnia’s migratory response to the antibiotic-treated fish was about half the strength of the response to the fish cue treatment. Furthermore, when the same antibiotic-treated fish were removed from the antibiotic-containing water and transferred into control water for 24 and 48 h, the extent of D. pulex’s migration depended on the length of the incubation period, apparently corresponding to the regeneration of bacterial colonies associated with mucus. The migration pattern observed in the 24 h treatment was similar to that of antibiotic-treated fish. On the other hand, a pronounced migration occurred in the 48 h following antibiotic treatment; here, we found a higher density of fish surface dwelling bacteria than at the start of the experiment. 4. In the experiment involving fish mucus, the mucus-enriched control water induced a weak response similar to antibiotic-treated fish. 5. On the basis of the results from the two experiments, we suggest that both fish and fish mucus-dwelling bacteria interact in the release of kairomone in ecologically relevant quantities.