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Background: Biogenic sulfuric acid (BSA) corrosion damages sewerage and wastewater treatment facilities but is not well investigated in sludge digesters. Sulfur/sulfide oxidizing bacteria (SOB) oxidize sulfur compounds to sulfuric acid, inducing BSA corrosion. To obtain more information on BSA corrosion in sludge digesters, microbial communities from six different, BSA-damaged, digesters were analyzed using culture dependent methods and subsequent polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE). BSA production was determined in laboratory scale systems with mixed and pure cultures, and in-situ with concrete specimens from the digester headspace and sludge zones. Results: The SOB Acidithiobacillus thiooxidans, Thiomonas intermedia, and Thiomonas perometabolis were cultivated and compared to PCR-DGGE results, revealing the presence of additional acidophilic and neutrophilic SOB. Sulfate concentrations of 10-87 mmol/L after 6-21 days of incubation (final pH 1.0-2.0) in mixed cultures, and up to 433 mmol/L after 42 days (final pH <1.0) in pure A. thiooxidans cultures showed huge sulfuric acid production potentials. Additionally, elevated sulfate concentrations in the corroded concrete of the digester headspace in contrast to the concrete of the sludge zone indicated biological sulfur/sulfide oxidation. Conclusions: The presence of SOB and confirmation of their sulfuric acid production under laboratory conditions reveal that these organisms might contribute to BSA corrosion within sludge digesters. Elevated sulfate concentrations on the corroded concrete wall in the digester headspace (compared to the sludge zone) further indicate biological sulfur/sulfide oxidation in-situ. For the first time, SOB presence and activity is directly relatable to BSA corrosion in sludge digesters. [ABSTRACT FROM AUTHOR]

Background: A major constraint for the effective control and management of helminth parasites is the lack of rapid, high-throughput, routine diagnostic tests to assess the health status of individual animals and herds and to identify the parasite species responsible for these helminthoses. The capability of a multiplex platform for the simultaneous detection of three pasture associated parasite species was evaluated and compared to existing ELISAs. Methods: The recombinant antigens 14.2 kDa ES protein for Cooperia oncophora, major sperm protein for Dictyocaulus viviparus and Cathepsin L1 for Fasciola hepatica were recombinantly expressed either in Escherichia coli or Pichia pastoris. Antigens were covalently coupled onto magnetic beads. Optimal concentrations for coupling were determined following the examination of serum samples collected from experimentally mono-infected animals, before and after their infection with the target species. Absence of cross-reactivity was further determined with sera from calves mono-infected with Haemonchus contortus, Ostertagia ostertagi and Trichostrongylus colubriformis. Examination of negative serum samples was characterised by low median fluorescence intensity (MFI). Results: Establishment of the optimal serum dilution of 1:200 was achieved for all three bead sets. Receiver Operating Characteristic analyses were performed to obtain cut-off MFI values for each parasite separately. Sensitivity and specificity at the chosen cut-off values were close to, or 100 % for all bead sets. Examination of serum samples collected on different days post infection from different animals showed a high reproducibility of the assays. Serum samples were additionally examined with two already established ELISAs, an in-house ELISA using the recombinant MSP as an antigen and a DRG ELISA using Cathepsin L1 for liver fluke. The results between the assays were compared and kappa tests revealed an overall good agreement. Conclusions: A versatile bead-based assay using fluorescence detection (xMAP® technology) was developed to simultaneously detect antibodies against C. oncophora, D. viviparus and F. hepatica in cattle serum samples. This platform provides rapid, high-throughput results and is highly sensitive and specific in comparison to existing serological as well as coproscopical diagnostic techniques. [ABSTRACT FROM AUTHOR]

Background Evaluation of xenobiotics biodegradation potential, shown here for benzotriazoles (corrosion inhibitors) and sulfamethoxazole (sulfonamide antibiotic) by microbial communities and/or pure cultures normally requires time intensive and money consuming LC/GC methods that are, in case of laboratory setups, not always needed. Results The usage of high concentrations to apply a high selective pressure on the microbial communities/pure cultures in laboratory setups, a simple UV-absorbance measurement (UVAM) was developed and validated for screening a large number of setups, requiring almost no preparation and significantly less time and money compared to LC/GC methods. This rapid and easy to use method was evaluated by comparing its measured values to LC-UV and GC-MS/MS results. Furthermore, its application for monitoring and screening unknown activated sludge communities (ASC) and mixed pure cultures has been tested and approved to detect biodegradation of benzotriazole (BTri), 4- and 5-tolyltriazole (4-TTri, 5-TTri) as well as SMX. Conclusions In laboratory setups, xenobiotics concentrations above 1.0 mg L-1 without any enrichment or preparation could be detected after optimization of the method. As UV-AM does not require much preparatory work and can be conducted in 96 or even 384 well plate formats, the number of possible parallel setups and screening efficiency was significantly increased while analytic and laboratory costs were reduced to a minimum. [ABSTRACT FROM AUTHOR]

Background VKORC1 has been identified some years ago as the gene encoding vitamin K epoxide reductase (VKOR) - the target protein for coumarin derivates like warfarin or phenprocoumon. Resistance against warfarin and other coumarin-type anticoagulants has been frequently reported over the last 50 years in rodents due to problems in pest control as well as in thrombophilic patients showing variable response to anticoagulant treatment. Many different mutations have already been detected in the VKORC1 gene leading to warfarin resistance in rats, mice and in humans. Since the conventional in vitro dithiothreitol (DTT)- driven VKOR enzymatic assay often did not reflect the in vivo status concerning warfarin resistance, we recently developed a cell culture-based method for coexpression of VKORC1 with coagulation factor IX and subsequent measurement of secreted FIX in order to test warfarin inhibition in wild-type and mutated VKORC1. Results In the present study, we coexpressed wild-type factor IX with 12 different VKORC1 variants which were previously detected in warfarin resistant rats and mice. The results show that amino acid substitutions in VKORC1 maintain VKOR activity and are associated with warfarin resistance. When we projected in silico the amino acid substitutions onto the published three-dimensional model of the bacterial VKOR enzyme, the predicted effects matched well the catalytic mechanism proposed for the bacterial enzyme. Conclusions The established cell-based system for coexpression of VKORC1 and factor IX uses FIX activity as an indicator of carboxylation efficiency. This system reflects the warfarin resistance status of VKORC1 mutations from anticoagulant resistant rodents more closely than the traditional DTT-driven enzyme assay. All mutations studied were also predicted to be involved in the reaction mechanism. [ABSTRACT FROM AUTHOR]

Background Sulfamethoxazole (SMX, sulfonamide antibiotic) biodegradation by activated sludge communities (ASC) is still only partly understood. The present work is focusing on nine different bacteria species capable of SMX biodegradation that were isolated from SMXacclimated ASC. Results Initially 110 pure cultures, isolated from activated sludge, were screened by UV-absorbance measurements (UV-AM) for their SMX biodegradation potential. Identification via almost complete 16S rRNA gene sequencing revealed five Pseudomonas spp., one Brevundimonas sp., one Variovorax sp. and two Microbacterium spp.. Thus seven species belonged to the phylum Proteobacteria and two to Actinobacteria. These cultures were subsequently incubated in media containing 10 mg L-1 SMX and different concentrations of carbon (sodium-acetate) and nitrogen (ammonium-nitrate). Different biodegradation patterns were revealed with respect to media composition and bacterial species. Biodegradation, validated by LC-UV measurements to verify UV-AM, occurred very fast with 2.5 mg L-1 d-1 SMX being biodegraded in all pure cultures in, for UV-AM modified, R2A-UV medium under aerobic conditions and room temperature. However, reduced and different biodegradation rates were observed for setups with SMX provided as co-substrate together with a carbon/nitrogen source at a ratio of DOC:N - 33:1 with rates ranging from 1.25 to 2.5 mg L-1 d-1. Conclusions Media containing only SMX as carbon and nitrogen source proved the organisms' ability to use SMX as sole nutrient source where biodegradation rates decreased to 1.0 - 1.7 mg L-1 d-1. The different taxonomically identified species showed specific biodegradation rates and behaviours at various nutrient conditions. Readily degradable energy sources seem to be crucial for efficient SMX biodegradation. [ABSTRACT FROM AUTHOR]

Background: Canine vector-borne infections have gained importance in Germany due to growing tourist traffic and an increased import of dogs from abroad. Endemic regions for pathogens such as Leishmania infantum, Hepatozoon canis, Ehrlichia canis, Anaplasma platys and Dirofilaria spp. are the Mediterranean area and southeastern Europe. Babesia species and Anaplasma phagocytophilum are present all over Europe. The objective of this retrospective study was to evaluate the prevalence of vector-borne infections in dogs imported from defined endemic countries in the Mediterranean area and southeastern Europe. Methods: Medical records and laboratory test results of 345 dogs that were imported to Germany from 17 endemic countries and that were presented to the Small Animal Clinic at Freie Universität Berlin between 2007 and 2015 were retrospectively reviewed. A total of 1368 test results from external laboratories were descriptively analysed including 576 and 792 test results of direct and indirect detection methods, respectively. Results: Overall, 35% (122/345 dogs) were positive for at least one pathogen. Concurrent infections with two to four pathogens were detected in 8% of the dogs (27/345). The positive results were: L. infantum 21% (66/314 dogs; methods: PCR 20/79, IFAT or ELISA 63/308 dogs), E. canis 16% (45/278 dogs; methods: PCR 8/68, IFAT 43/257 dogs), H. canis 11% (3/28 dogs; method: PCR), Babesia spp. 10% (25/251 dogs; methods: Babesia spp. PCR 3/98, B. canis/vogeli IFAT or ELISA 22/214 and B. gibsoni IFAT 0/13 dogs), Dirofilaria spp. 7% (13/178 dogs; methods: D. immitis Ag-ELISA 8/156, Knott's test 7/95, microfilariae PCR 5/23 dogs) and A. platys 5% (1/21 dogs; method: PCR). None of 8 tested dogs were positive in a combined Babesia spp./Hepatozoon spp. PCR test. Conclusions: Dogs, which are imported from countries which are endemic for vector-borne infections should be thoroughly tested using direct and indirect detection methods. Potential owners of imported dogs should be informed about the diseases, risks and incubation periods. [ABSTRACT FROM AUTHOR]

Background: Fasciola hepatica and Ostertagia ostertagi infections are widespread in cattle population of Europe, however data on their prevalence in Poland are only fragmentary. Therefore, the cross-sectional study was carried out to determine the herd-level seroprevalence of F. hepatica and O. ostertagi infection in dairy cattle population in the central and north-eastern provinces Poland, and to identify basic local risk factors for these infections. In total, 598 herds were enrolled, 394 (65.9%) in the north-eastern province and 204 (34.1%) in the central province. In each herd the questionnaire survey was conducted and bulk-tank milk (BTM) sample was collected and screened using two indirect immunoenzymatic tests. Optical density ratio (ODR) was regarded as the quantitative proxy of exposure to either of the two parasites. Results: Both Fasciola and Ostertagia ELISA ODR in the north-eastern province was significantly higher than ODR in the central province. At the cut-off value of ODR = 0.27 the true herd-level seroprevalence of F. hepatica was 79.6% (95% CI: 74.0%, 84.3%) in the north-eastern province and 13.0% (95% CI: 5.3%, 21.7%) in the central province. At the cut-off of ODR = 0.50151 of 188 herds (80.3%, 95% CI: 74.1%, 85.4%) were seropositive for O. ostertagi in the north-eastern province and only 70 of 136 herds (51.5%, 95% CI: 43.1%, 59.7%) were seropositive in the central province. Location of a herd in the north-eastern province, longer grazing period practiced in a herd and > 50%-share of grazing grass in roughage were all positively related to the increase in exposure to both parasites. Moreover, the use of hay or haylage as main roughage proved to be positively related to the increase in exposure to F. hepatica. Conclusions: F. hepatica and O. ostertagi are widespread in cattle population in Poland, however their occurrence at a herd-level varies between different regions of Poland. This diversity can only partly be explained by different herd management, and appears linked to environmental and climate conditions typical for these regions. [ABSTRACT FROM AUTHOR]