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Avian mycoplasmas are associated with respiratory disease, synovitis, poor quality of day-old chicks and poor performance. The main approach used for the diagnosis of avian mycoplasmas is isolation and identification of the microorganism. Since the bacteria are slow growing fastidious organisms, conventional methods are time consuming, laborious and require experienced personnel. For this reason, we aimed to develop a rapid detection method for Mycoplasma gallisepticum and Mycoplasma synoviae by quantitative real-time polymerase chain reaction (qPCR). For this purpose, the lipoprotein (lp) and variable lipoprotein hemagglutinin (vlhA) genes were used to detect M. gallisepticum and M. synoviae, respectively. The limit of detection (LOD) of the assay was determined to be 100- 101 DNA/µl from artificially contaminated swab samples. The specificity and sensitivity ratios were 100%. Overall, these results indicate that this qPCR method can be accurately used for detection of MG and MS.

The direct effect of fungi on human health makes the fungal diversity in the air an important and necessary subject for examination. Studies that determine fungal diversity generally depends on culture methods. Developed culture-independent methods eliminate many disadvantages of existing culture-dependent methods. In our study, duplicate air samples were collected on 5 different days at a Microbiology Research Laboratory to compare these methods. Samples were collected in 3 different groups: (A) culture method and ITS-targeted Sanger DNA sequencing with 100 L samples, (B) culture method and ITS-targeted Sanger DNA sequencing with 1000 L samples, (C) filter method and next generation sequencing with 1000 L air sample. The Groups A, B and C defines culture, isolation and SANGER DNA-based methods, culture and next generation sequencing methods and directly next generation sequencing methods, respectively. Method-A failed to represent real fungal diversity. Method-B did represent the diversity but did not truly represent relative abundance of the species. Method-C can be completed in 2 days whereas Method-A and B can be completed in 2 weeks and 1 week, respectively. In conclusion, we recommend direct DNA isolation which is followed by ITS-targeted NGS in order to study fungal diversity in indoor air environments.

A one-step reverse transcription and real-time PCR (RT-qPCR) test was developed for rapid screening (40 minutes) of the Spike N501Y and HV69-70del mutations in SARS-CoV-2 positive samples. The test also targets a conserved region of SARS-CoV-2 Orf1ab as an internal control. The samples containing both the N501Y and HV69-70del mutations are concluded as VOC-202012/01 positive. Samples suspected to be positive for B.1.351 or P.1 are the N501Y positive and HV69-70del negative cases. Limit of detection (LOD) of the kit for Orf1ab target is 500 copies/mL, while that of the N501, Y501 and HV69-70del targets are 5000 copies/mL. The developed assay was applied to 165 clinical samples containing SARS-CoV-2 from 32 different lineages. The SARS-CoV-2 lineages were determined via the next-generation sequencing (NGS). The RT-qPCR results were in 100% agreement with the NGS results that 19 samples were N501Y and HV69-70del positive, 10 samples were N501Y positive and HV69-70del negative, 1 sample was N501Y negative and HV69-70del positive, and 135 samples were N501Y and HV69-70del negative. All the VOC-202012/01 positive samples were detected in people who have traveled from England to Turkey. The RT-qPCR test and the Sanger sequencing was further applied to 1000 SARS-CoV-2 positive clinical samples collected in Jan2021 from the 81 different provinces of Turkey. The RT-qPCR results were in 100% agreement with the Sanger sequencing results that 32 samples were N501Y positive and HV69-70del negative, 4 samples were N501Y negative and HV69-70del positive, 964 samples were N501Y and HV69-70del negative. The specificity of the 40 minutes RT-qPCR assay relative to the sequencing-based technologies is 100%. The developed assay is an advantageous tool for timely and representative estimation of the N501Y positive variants’ prevalence because it allows testing a much higher portion of the SARS-CoV-2 positives in much lower time compared to the sequencing-based technologies.

The aim of this study was to investigate the effect of high-influent-concentration suspended solids (SS) on the cultivation, structure and long-term stability of aerobic granular sludge (AGS). Cultivation and long-term stability of AGS were monitored in two pilot-scale sequencing batch reactors fed with raw (R1) and settled (R2) domestic wastewater, representing high and medium SS content, respectively. The real domestic wastewater had high chemical oxygen demand (COD) content (1100 +/- 270 mg COD L-1). Aerobic granular sludge was cultivated in 44 days (R1) and 25 days (R2) under the conditions of high settling velocity (18 m h(-1)) and high organic loading rate (OLR) (2.1-2.4 kg COD m(3) day). The AGS in both reactors had similar structural properties during long-term operation and remained structurally and functionally stable during the last five months of operation. Comparative evaluation of the results indicated that the high influent SS content of the real domestic wastewater had a positive influence on maintaining significantly lower SVI30 and relatively lower effluent SS concentration. Moreover, a higher influent SS content resulted in smaller mature granules during the stable period. Microbial community analyses helped to understand the aerobic granular sludge structure and showed that the sludge retention time and OLR affected the granular sludge population. The high influent SS increased biomass detachment from the granular sludge surface and caused wash-out of some bacteria colonizing the exterior of the granular sludge. (C) 2017 Elsevier Ltd. All rights reserved.

Seven different PCR primers targeting 18 different myxomycetes genera were designed and ribosomal DNA ITS1-5.8SrDNA-ITS2 regions of 52 morphologically characterized species from Turkey were amplified and sequenced. The studied species belong to the genera Arcyria, Badhamia, Ceratiomyxa, Collaria, Comatricha, Cribraria, Diachea, Diderma, Didymium, Enerthenema, Lamproderma, Lycogala, Perichaena, Physarum, Reticularia, Stemonitis, Stemonitopsis, and Trichia. Significant ITS region size and sequence variations were observed among the analyzed species. The ITS based phylogenetic tree did not reflect evolutionary relationships at genus or higher level. The only exception was the genus Didymium, which formed a clearly distinctive cluster in the phylogenetic tree containing 18 different genera. It was concluded that interspecies variation of the ITS1-5.8S-ITS2 region is high enough to differentiate species, but this region did not reflect the evolutionary relationships between the myxomycetes species. Hence, we suggested that sequencing of the ITS region could only be used as a supporting tool for the phenotypic identification, not for revealing the evolutionary relationships between the myxomycetes species.

In this study, the effect of heat and chromium (Cr) heavy metal interactions on wheat seedlings (Triticum aestivum L. cv. Ç-1252 and Gun91) was investigated by measuring total chlorophyll and carotenoid levels, catalase (CAT) and ascorbate peroxidase (APX) antioxidant enzyme activities, and MYB73, ERF1 and TaSRG gene expression. Examination of pigment levels demonstrated a decrease in total chlorophyll in both species of wheat under combined heat and heavy metal stress, while the carotenoid levels showed a slight increase. APX activity increased in both species in response to heavy metal stress, but the increase in APX activity in the Gun91 seedlings was higher than that in the Ç-1252 seedlings. CAT activity increased in Gun91 seedlings but decreased in Ç-1252 seedlings. These results showed that Gun91 seedling had higher resistance to Cr and Cr + heat stresses than the Ç-1252 seedling. The quantitative molecular analyses implied that the higher resistance was related to the overexpression of TaMYB73, TaERF1 and TaSRG transcription factors. The increase in the expression levels of these transcription factors was profound under combined Cr and heat stress. This study suggests that TaMYB73, TaERF1 and TaSRG transcription factors regulate Cr and heat stress responsive genes in wheat.

Aim To detect the unknown components of the oral microbiome and the effects of root canal treatment in a Turkish population and to evaluate the changes in microbial diversity in the root canals before and after treatment. Methodology Single-rooted central and lateral maxillary incisors with one canal were chosen from 20 patients. Baseline samples of intact intracanal microbiota were collected from 20 root canals of permanent teeth with necrotic pulps using sterile paper points. After root canal preparation, the root canals were filled with a calcium hydroxide paste for 7 days. Calcium hydroxide was removed from root canal with 2.5% sodium hypochlorite and 17% EDTA using passive ultrasonic irrigation (PUI). A second bacteriologic samples were taken with sterile paper points prior to root filling. The samples were processes with DNase-I treatment using next-generation sequencing (NGS). Reduction in bacterial numbers during root canal treatment was evaluated using real-time quantitative PCR (qPCR). All statistical analyses were conducted using the MINITAB 17 software (Minitab Ltd. Co., Coventry, UK). A one-sample t-test was used to analyse the data. Statistical significance was accepted at P Results Relative abundances of Mycoplasma sp., Paludibacter sp., Tannerella sp., Prevotella spp. and an uncultured species from the order Bacteroidales decreased with root canal preparation and medication (98.7%, 99.8%, 98.8%, 97.7% and 99.3%, respectively), whilst the relative abundances of Methylobacterium sp., Corynebacterium sp. and Streptococcus infantis increased (93.1%, 94.8% and 99.4%, respectively). Considerable numbers of Streptophyta species were detected before and after treatment. The ratio of Agrobacterium sp. in the treated teeth community and the ratio of order Streptophyta in the infected canals had negative correlations with the success of bacterial elimination. Conclusions The combination of NGS and qPCR techniques resulted in detection of previously unknown components of the oral microbiome and the effects of root canal treatment on their relative abundance in a Turkish population.

Background: Group A streptococci (GAS) are the most common bacterial cause of acute pharyngitis and account for 15-30 % of cases of acute pharyngitis in children and 5-10 % of cases in adults. In this study, a real-time quantitative PCR (qPCR) based GAS detection assay in pharyngeal swab specimens was developed. Methods: The qPCR assay was compared with the gold standard bacterial culture and a rapid antigen detection test (RADT) to evaluate its clinical performance in 687 patients. The analytical sensitivity of the assay was 240 cfu/swab. Forty-five different potential cross-reacting organisms did not react with the test. Four different laboratories for the reproducibility studies were in 100 % (60/60) agreement for the contrived GAS positive and negative swab samples. Results: The relative sensitivities of the RADT and the qPCR test were 55.9 and 100 %; and the relative specificities were 100 and 96.3 %, respectively. Duration of the total assay for 24 samples including pre-analytical processing and analysis changed between 42 and 55 min depending on the type of qPCR instrument used. A simple DNA extraction method and a low qPCR volume made the developed assay an economical alternative for the GAS detection. Conclusion: We showed that the developed qPCR test is rapid, cheap, sensitive and specific and therefore can be used to replace both antigen detection and culture for diagnosis of acute GAS pharyngitis.

Physicochemical and microbiological characterization of anoxic sediments taken from seven highly polluted sites of the Marmara Sea was carried out. The 16S rRNA based microbial community structure analyses were performed using domain-specific PCR followed by denaturant gradient gel electrophoresis (DGGE) and sequencing of characteristic bands. The results showed that the microbial communities in these sediments were diverse and evenly distributed. Relating the prokaryotic and geochemical variables through statistical tools revealed that the microbial diversity in the sediments significantly related to depth, and S, Mn and Fe content of the sediments. Fermentative bacteria, denitrifying bacteria and hydrogenotrophic methanogens were dominant whereas sulfate reducing bacteria were absent in the DGGE patterns. This unusual microbial community structure implied that the newly discovered anaerobic methane oxidation coupled to denitrification process may occur in these subseafloor environments. (C) 2008 Elsevier Ltd. All rights reserved.

In this study, two full-scale upflow anaerobic sludge blanket (UASB) reactors, namely TUASB and CUASB, at the wastewater treatment plants of the Tekirdaǧ Alcohol (Raki) and Canakkale Alcohol (Cognac) distilleries were investigated in terms of performance, acetoclastic methanogenic capacity and microbial composition. The results were compared with a previously studied other UASB reactor (IUASB) at the wastewater treatment plant of the Istanbul Alcohol (Raki) Distillery from which the two reactors (TUASB and CUASB) were seeded. The IUASB reactor performed well achieving COD removal efficiencies of no lower than 85% at organic logding rates (OLRs) in the range of 6–11 kg COD m−3 day−1 between 1996 and 2001. During the last one year of operation, between 2000 and 2001, performance of the CUASB reactor in terms of COD removal efficiency was 70–80% at OLRs in a range of 1–4.5 kg COD m−3 day−1 whereas it was 60–80% at OLRs in a range of 2.5–8.5 kg COD m−3 day−1 in the TUASB reactor. At the end of year 2000, specific methanogenic activity (SMA) tests were carried out to determine potential loading capacity and optimum operating conditions of the IUASB, CUASB and TUASB reactors. The potential methane production (PMP) rates of the CUASB, IUASB and TUASB reactors were measured as 230 cm3 CH4 gVSS−1 day−1, 350 cm3 CH4 gVSS−1 day−1 and 376 cm3 CH4 gVSS−1 day−1 respectively. When the PMP rates were compared with actual methane production (AMP) rates obtained from the three UASB reactors, AMP/PMP ratios were evaluated to be 0.18, 0.12 and 0.13 for CUASB, TUASB and IUASB reactors respectively. This showed that the CUASB, TUASB and IUASB reactors were using only 18%, 12% and 13% of their potential acetoclastic methanogenic capacity respectively. These results can be interpreted that the three UASB reactors were underloaded compared with their potential acetoclastic methanogenic capacities. It was, therefore, recommended that the three UASB reactors should be loaded at higher organic loading rates or sludge withdrawn in order to maintain an AMP/PMP ratio of 0.6–0.7, which can ensure desired reactor performance with safer operation. Results of epifluoresence microscopic examinations showed that the percentage of total autofluorescent methanogens was approximately 30% of the total population in sludges from the TUASB and IUASB reactors whereas it was 20% in sludge from the CUASB reactor. The two UASB reactors treating raki distillery wastewaters contained sludges having a higher percentage of autofluorescent methanogenic population and higher acetoclastic methanogenic activity. Copyright © 2004 Society of Chemical Industry

Vibrio anguillarum expresses several virulence factors and causes hemorrhagic septicemia accompanied by serious losses in marine fish. Transferrin is a glycoprotein, also known as a multitasking protein, which is mainly synthesized by the liver. It has a fundamental role in the immune system. In the present study, the transferrin gene expression of sea bass (Dicentrarchus labrax) was investigated during an experimental infection with V. anguillarum. Fish samples were examined by hematological and serological methods as well as real-time polymerase chain reaction. The infection was performed via water. The sampled fish displayed vibriosis infection symptoms, both internally and externally. The transferrin saturation in the diseased fish serum decreased dramatically and the transferrin gene expression increased during the first 2 days; however, it decreased in the subsequent days.

In this study, changes in microbial diversity and polyhydroxyalkanoate storage ability of activated sludge under aerobic dynamic feeding conditions were investigated. Two sequencing batch reactors were operated with and without nitrogen limitation, by applying a moderate sludge retention time. Polymer storage abilities of the biomasses were improved significantly under dynamic conditions, in terms of specific polymer storage rate, polymer storage yield and polymer content of activated sludge. Moreover, aerobic dynamic feeding conditions together with nutrient limitation further improved the storage ability of the mixed population. Polymer storage yields of the biomass enriched under nitrogen-sufficient and nitrogen-deficient conditions were 0.43 and 0.61 Cmmol PHA/Cmmol substrate, respectively. This study also contributes to the knowledge of activated sludge microbiology, providing detailed information about temporal changes in community structure under dynamic conditions. Microbial community structure was determined by 16S rDNA clone library construction. Also changes in communities under different operating conditions were monitored by DGGE analysis based on bacterial 16S rDNA. The beta subclass of Proteobacteria was the most abundant phylum in both reactors during the operation periods. Changes in the community structure occurred in terms of relative abundance of the operational taxonomic units (OTUs) rather than the OTU types present in the system.

The aim of this study was to determine the effect of toluene on an anaerobic sludge taken from a full-scale upflow anaerobic sludge blanket (UASB) reactor in terms of potential activity and composition of acetoclastic methanogens. Specific methanogenic activity (SMA) test results showed that 5%, 9.5%, 14%, 24%, 29%, 38% and 62% inhibition occurred in the potential methane production (PMP) rate of the sludge at toluene concentrations of 0.1 mM, 0.2 mM, 0.3 mM, 0.4 mM, 0.5 mM, 0.6 mM and 1 mM, respectively. Fluorescence in situ hybridization (FISH) results showed that relative abundance of archaeal cells was approx. 19% throughout the SMA tests. The anaerobic sludge was dominated by acetoclastic genus Methanosaeta which were slightly affected by increasing toluene concentrations do not have any effect on relative abundance of Methanosaeta spp., which was between 73% +/- 1.6 and 68% +/- 2.1 of the archaeal population.

Spatial (10 different locations) and temporal (2 years) changes in characteristics of the Marmara Sea Sediments were monitored to determine interactions between the chemical and microbial diversity. The sediments were rich in terms of hydrocarbon, nitrate, Ni and microbial cell content. Denitrifying, sulfate reducing, fermentative and methanogenic organisms were co-abundant in 15 cm below the sea floor. The local variations in the sediments' characteristics were more distinctive than the temporal ones. The sulfate and nitrate contents were the main drivers of the changes in the microbial community compositions. N and P were limited for microbial growth in the sediments, and their levels determined the total cell abundance and activity. Seasonal shifts in temperatures of the shallow sediments were also reflected in the active cell abundances. It was concluded that the Marmara Sea is a promising ecosystem for the further investigation of the ecologically important microbial processes. (C) 2011 Elsevier Ltd. All rights reserved.

A full-scale upflow anaerobic sludge blanket (UASB) reactor was investigated in terms of archaeal composition, acetoclastic methanogenic capacity and performance over a 2-year period. Performance of the reactor in terms of COD removal efficiency varied between 60% and 80% at organic loading rates (OLRs) in the range of 2.5-12 kg COD m-3 d-1. The reactor had been operated under a F/M (food to microorganisms) ratio of 0.02-0.03 gCOD gTVS-1 d-1, which is much lower than the typical values reported for similar reactors. According to specific methanogenic activity (SMA) tests the anaerobic sludge was operating at only 12-34% of its potential acetoclastic methanogenic capacity. These results demonstrated that the UASB reactor was under loaded compared to its maximum loading capacity. All other operational parameters had been maintained within their desired ranges. The SMA test and Fluorescence in situ hybridization (FISH) results revealed that a decrease in the acetoclastic methanogenic activity of the UASB sludge from 344 mL CH4 gTVS-1 d-1 to 109 mL CH4 gTVS-1 d-1 coincided with a decrease in the relative abundance of acetoclastic Methanosaeta from 90%+/-1.2 to 79%+/-1.4 of the archaeal population, and an increase in the relative abundance of hydrogenotrophic Methanobacteriales from non-detectable levels to 24%+/-0.7% of the archaeal population during the 2-year operation of the reactor. The relative abundance of archaeal cells within the UASB sludge was in the range of 15-17%.