Bartonella infections in fleas (Siphonaptera : Pulicidae) and lack of Bartonellae in ticks (Acari : Ixodidae) from Hungary

Fleas (95 Pulex irritans, 50 Ctenocephalides felis, 45 Ctenocephalides canis) and ixodid ticks (223 Ixodes ricinus, 231 Dermacentor reticulatus, 204 Haemaphysalis concinna) were collected in Hungary and tested, in assays based on PCR, for Bartonella infection. Low percentages of P. irritans (4.2%) and C. felis (4.0%) were found to be infected. The groEL sequences of the four isolates from P. irritans were different from all the homologous sequences for bartonellae previously stored in GenBank but closest to those of Bartonella sp. SE-Bart-B (sharing 96% identities). The groEL sequences of the two isolates from C. felis were identical with those of the causative agents of cat scratch disease, Bartonella henselae and Bartonella clarridgeiae, respectively. The pap31 sequences of B. henselae amplified from Hungarian fleas were identical with that of Marseille strain. No Bartonella-specific amplification products were detected in C. canis, L ricinus, D. reticulatus and H. concinna pools

Molekuláris faj-diverzitás vizsgáló módszerek fajkompozíció torzításának elemzése, optimális eljárás kidolgozása és gyakorlati alkalmazása = Analysis of method inherent species composition biases of molecular species diversity analyses, method optimatisation and practical application

A baktérium közösségek szerkezetéről molekuláris módszerekkel levont következtetések feltevése, hogy a vizsgálati lépések során a minta faj-abundancia viszonyai nem változnak. A kutatások ennek ellenkezőjét sugallják. Vizsgálataink a multitemplát PCR, ill. a klónkönyvtár készítés közösségszerkezet-torzító hatásait térképezte fel. A preferenciális amplifikációt bacteria specifikus primerekkel elemeztük, az annelációs hőmérséklet, ill. a ciklusszám hatására. Ismert genomú törzsek DNS keverékein a PCR termékek mennyiségi viszonyait TRFLP-vel vizsgáltuk. 1:1 arányú templátkeverékeknél az illeszkedési hibával bíró a tökéletesen illeszkedő templáttal szemben hátrányba került. Az aránytorzítás exponenciális összefüggést mutat az annelációs hőmérséklettel. A PCR ciklusszámnak elhanyagolható hatása van a PCR termékek arányaira. Környezeti minták vizsgálatai alapján javasoljuk a kis annelációs hőmérsékletek használatát a torzulások elkerülésére. A klónozás során fellépő preferenciális ligációt TA vektor segítségével vizsgáltuk. A 16S-23S spacer elemzésnél jellemző inzertméret különbségek esetén a klónkönyvtárak összetétele erősen eltolódott a rövidebb inzertek felé. Ha az inzertek csak szekvenciában különböztek, enyhe torzulást kaptunk. Vagyis a nagy hossz heterogenitású génszakaszokra épülő klónozásos vizsgálatoknál a rövidebb inzertet adó fajok túlreprezentáltak lesznek. A környezeti klónokból az első 12 faj izolálása (Bacteria és Archaea) megtörtént. | There are several biasing factors in molecular microbiological community analyses though the quantitative aspect of multitemplate PCR and ligation at clone library construction is poorly investigated. The biases were investigated using bacteria specific primers, focusing on the effect of primer mismatch, annealing temperature and PCR cycle numbers. The distortion of template-to-product ratio was measured using TRFLP analysis. At 1:1 genomic DNA template mixtures, primer mismatches presented serious bias, with preferential amplification of the template with perfectly matching sequence. The extent of preferential amplification showed exponential relation with increasing annealing temperatures. The number of PCR cycles had little influence on template-to-product ratios. Analysing environmental samples, the use of low annealing temperature was recommended to reduce preferential amplification. Blunt-end cloning ligation step was investigated for the insert length heretogeneity. Strong preferential ligation of the shorter PCR amplicon was shown. Slightly skewed ratios were detected at the same insert length and GC content. These findings indicate that members of a diverse microbial sample may be excluded during clone library construction because of preferential ligation, this way biasing the true community picture.Environmental clones detected by the optimised techniques were targeted to culture. 12 new species (Bacteria and Archaea) were isolated and partially characterised

Seasonal and Spatial Changes of Planktonic Bacterial Communities Inhabiting the Natural Thermal Lake Hévíz, Hungary

Lake Hévíz is a unique thermal spa located in Hungary. Owing to the thermal springs nourishing the lake, it has a relatively rapid water turnover. In spring 2011 a comprehensive embankment reconstruction was performed to preserve the water supply of the surrounding wetland habitats. The physical and chemical parameters as well as the planktonic microbial communities were studied with special respect to the effect of the disturbance of the water of Lake Hévíz. According to the abiotic components, both temporal and spatial differences were revealed with the exception of autumn samples. The reconstruction resulted in a short term but dramatic alteration of the total planktonic bacterial and cyanobacterial community structures as revealed by denaturing gradient gel electrophoresis. In addition, greater seasonal than spatial differences of bacterial communities were also observed. Planktonic bacterial community composition of Lake Hévíz included mainly typical freshwater species within phylum Actinobacteria, Chloroflexi, Cyanobacteria and class Alpha-, Beta- and Gamma-proteobacteria. Most of them were aerobic or facultative anaerobic heterotrophic but chemolitotrophic (e.g. Thiobacillus) or photolithotrophic (e.g. Cyanobacteria and Chloroflexi) autotrophic microbes were also identified

Magyarországi szikes vizek bakteriális biodiverzitásának megismerése, extremofil szervezetek polifázikus taxonómiai jellemzése = Bacterial biodiversity of Hungarian soda lakes and polyphasic taxonomic characterisation of extremophile microorganisms

Magyarországi szikes vizek mikrobaközösségeit szezonális mintavételezéseket követően tenyésztésen alapuló és tenyésztéstől független molekuláris biológiai diverzitás elemző módszerek együttes alkalmazásával vizsgáltuk 2002 és 2006 között. E vizsgálatok keretében betekintést nyertünk a Velencei-tó, a kiskunsági Kelemen-szék, Zab-szék és Böddi-szék, valamint a tiszántúli Nagy-vadas-tó és Fehér-szik vizének és üledékének aerob mikrobiális folyamataiban résztvevő baktériumközösségek faji összetételébe, a szikes (extrém) élőhelyekhez adaptálódott autochton alkalofil és halofil mikrobaközösségek szerkezetébe. Megismertük a törzsek anyagcsere potenciálját és ökológiai toleranciáját. 16S rRNS és dsrAB funkcionális gén egyidejű vizsgálatán alapuló kombinált eljárással feltártuk a szikes vizek nádasainak rizoszférájában élő anaerob szulfátredukáló baktériumközösségek eddig még ismeretlen faji struktúráját. Új mikrobiológiai módszerek (BIOLOG és DGGE) hazai bevezetésével és alkalmazásával összehasonlítottuk a különböző szikes vizek mikrobaközösségeinek szerkezetében és aktivitásában megnyilvánuló változásokat, következtetéseket vontunk le a mikrobaközösségek anyagforgalmi dinamikájára vonatkozóan. A tavi környezetek kémiai sajátosságainak figyelembevételével kifejlesztett szelektív táptalajokon több a tudományra nézve új extremofil baktérium taxont sikerült tenyésztésbe vonni és leírni. Megkezdődött a törzsek gyakorlati célú alkalmazása: erőművi pernye komposztálásos ártalmatlanítására. | Microbial communities of Hungarian soda lakes were studied by cultivation-based and cultivation-independent methods following seasonal samplings between 2002 and 2006. The species composition of aerobic bacterial communities and the structure of autochthonous alkaliphilic and halophilic microbial communities adapted to sodic (extreme) water and sediment habitats were investigated in Lake Velencei, in Kelemen-szék, Zab-szék and Böddi-szék (Kiskunság National Park), in Nagy-vadas and Fehér-szik (Tiszántúl region). The metabolic potential and ecological tolerance of the strains were also studied. A combined 16S rRNA and dsrAB gene based approach was applied to reveal the so far unknown species diversity of sulphate reducing bacteria inhabiting the reed rhizosphere of soda lakes. Novel microbiological methods (BIOLOG and DGGE) were introduced and used first in Hungary to compare the changes of the structure and activity of microbial communities originating from different soda lakes and to ascertain the possible contributions of microbial communities to the local carbon cycling. New selective media were developed considering the physico-chemical characteristics of the lakes from which several novel bacterial taxa were managed to cultivate and describe. Experiments have been started with some of these alkali-halophilic strains for their biotechnological application in compost production

Unique picoeukaryotic algal community under multiple environmental stress conditions in a shallow, alkaline pan

Winter phytoplankton communities in the shallow alkaline pans of Hungary are frequently dominated by picoeukaryotes, sometimes in particularly high abundance. In winter 2012, the ice-covered alkaline Zab-sze´k pan was found to be extraordinarily rich in picoeukaryotic green algae (42–82 9 106 cells ml-1) despite the simultaneous presence of multiple stressors (low temperature and light intensity with high pH and salinity). The maximum photosynthetic rate of the picoeukaryote community was 1.4 lg C lg chlorophyll a-1 h-1 at 125 lmol m-2 s-1. The assimilation rates compared with the available light intensity measured on the field show that the community was considerably light-limited. Estimated areal primary production was 180 mg C m-2 d-1. On the basis of the 18S rRNA gene analysis (cloning and DGGE), the community was phylogenetically heterogeneous with several previously undescribed chlorophyte lineages, which indicates the ability of picoeukaryotic communities to maintain high genetic diversity under extreme conditions

Microbiological investigations on the water of a thermal bath at Budapest

Thermal baths are unique aquatic environments combining a wide variety of natural and anthropogenic ecological factors, which also appear in their microbiological state. There is limited information on the microbiology of thermal baths in their complexity, tracking community shifts from the thermal wells to the pools. In the present study, the natural microbial community of well and pool waters in Gellért bath was studied in detail by cultivation-based techniques. To isolate bacteria, 10% R2A and minimal synthetic media (with “bath water”) with agar–agar and gellan gum were used after prolonged incubation time; moreover, polyurethane blocks covered with media were also applied. Strains were identified by sequencing their 16S rRNA gene after grouping them by amplified rDNA restriction analysis. From each sample, the dominance of Alphaproteobacteria was characteristic though their diversity differed among samples. Members of Actinobacteria, Firmicutes, Beta- and Gamma-proteobacteria, Deinococcus–Thermus, and Bacteroidetes were also identified. Representatives of Deinococcus–Thermus phylum appeared only in the pool water. The largest groups in the pool water belonged to the Tistrella and Chelatococcus genera. The most dominant member in the well water was a new taxon, its similarity to Hartmannibacter diazotrophicus as closest relative was 93.93%

Remarkable impact of PAHs and TPHs on the richness and diversity of bacterial species in surface soils exposed to long-term hydrocarbon pollution

Nowadays, because of substantial use of petroleum-derived fuels the number and extension of hydrocarbon polluted terrestrial ecosystems is in growth worldwide. In remediation of aforementioned sites bioremediation still tends to be an innovative, environmentally attractive technology. Although huge amount of information is available concerning the hydrocarbon degradation potential of cultivable hydrocarbonoclastic bacteria little is known about the in situ long-term effects of petroleum derived compounds on the structure of soil microbiota. Therefore, in this study our aim was to determine the longterm impact of total petroleum hydrocarbons (TPHs), volatile petroleum hydrocarbons (VPHs), total alkyl benzenes (TABs) as well as of polycyclic aromatic hydrocarbons (PAHs) on the structure of bacterial communities of four different contaminated soil samples. Our results indicated that a very high amount of TPH affected positively the diversity of hydrocarbonoclastic bacteria. This finding was supported by the occurrence of representatives of the a-, b-, c-Proteobacteria, Actinobacteria, Flavobacteriia and Bacilli classes. High concentration of VPHs and TABs contributed to the predominance of actinobacterial isolates. In PAH impacted samples the concentration of PAHs negatively correlated with the diversity of bacterial species. Heavily PAH polluted soil samples were mainly inhabited by the representatives of the b-, c- Proteobacteria (overwhelming dominance of Pseudomonas sp.) and Actinobacteria