Jitka Černohlávková

Effects of fungicides mancozeb and dinocap on carbon and nitrogen mineralization in soils.

In our study, effects of fungicides mancozeb and dinocap on C and N mineralization were measured in arable and grassland soil. The soils were treated with these fungicides at the application and 10 times lower doses and then incubated at 20 degrees C for 2 weeks. Carbon mineralization (basal and substrate-induced respiration) and nitrogen mineralization (potential ammonification and nitrification) were evaluated 1 and 14 days after the treatment. After 14 days, ammonification was decreased to 48% and 83% at dinocap application dose in arable and grassland soil, respectively. Application dose of mancozeb caused significant decrease of nitrification to 11.2% and 5.6% in arable and grassland soil, respectively. Basal respiration and substrate-induced growth were rather stimulated by fungicides, especially at lower application doses. To conclude, potential risk may exist to soil microorganisms and their activities in soils treated routinely by mancozeb or dinocap.

Variability of soil microbial properties: effects of sampling,handling and storage

We investigated the effect of soil spatial variability within the sampling site scale, the effects of sample sieving (1, 2 and 4mm), and storage conditions up to 32 weeks (wet at 4 degrees C, -20 degrees C and air dried) on microbial biomass C, respiration, ammonification and nitrification activities in arable, grassland and forest soil. In general, all results were dependent on soil type. Arable soil showed the highest spatial variability, followed by grassland and forest soil. Sieving did not cause large differences; however, higher biomass C and respiration activity were observed in the 1mm than in the 4mm fraction. Storage at 4 degrees C seemed to be the most appropriate up to 8 weeks showing only minor changes of microbial parameters. Freezing of soils resulted in large increase of respiration. Dried storage indicated disruption of microbial communities even after 2 weeks.

Staphylococcus petrasii sp. nov. including S. petrasii subsp. petrasii subsp. nov. and S. petrasii subsp. croceilyticus subsp. nov., isolated from human clinical specimens and human ear infections

Thirteen coagulase-negative, oxidase-negative, and novobiocin-susceptible staphylococci were isolated from human clinical specimens. The isolates were differentiated from known staphylococcal species on the basis of 16S rRNA, hsp60, rpoB, dnaJ, tuf, and gap gene sequencing, automated ribotyping, (GTG)5-PCR fingerprinting, and MALDI-TOF MS analysis. Phylogenetic analysis based on the 16S rRNA gene sequence indicated phylogenetic relatedness of the analyzed strains to Staphylococcus haemolyticus, Staphylococcus hominis, Staphylococcus devriesei, and Staphylococcus lugdunensis. DNA-DNA hybridization experiments between representative strains CCM 8418(T), CCM 8421(T), and the closest phylogenetic neighbors confirmed that the isolates represent novel Staphylococcus species, for which the name Staphylococcus petrasii sp. nov. is proposed. Genotypic and phenotypic analyses unambiguously split the strains into two closely related subclusters. Based on the results, two novel subspecies S. petrasii subsp. petrasii subsp. nov. and S. petrasii subsp. croceilyticus subsp. nov. are proposed, with type strains CCM 8418(T) (=CCUG 62727(T)) and CCM 8421(T) (=CCUG 62728(T)), respectively.

Vagococcus entomophilus sp. nov., from the digestive tract of a wasp (Vespula vulgaris)

Three unknown Gram-stain-positive, catalase-negative, facultatively anaerobic and coccus-shaped strains of bacteria were isolated from the digestive tracts of wasps (Vespula vulgaris). Analysis of 16S rRNA gene sequences revealed that these strains had identical sequences and showed that Vagococcus salmoninarum, with 96.2% sequence similarity, was the closest phylogenetic neighbour. Further analyses based on hsp60 and pheS gene sequences of representatives of the family Enteroccocaceae and genotypic and phenotypic characterization using (GTG)5-PCR fingerprintings, EcoRI ribotyping, DNA G+C content, whole-cell protein profiling, cellular fatty acid profiles analysis and extensive biotyping confirmed that the investigated strains were representatives of a novel bacterial species within the genus Vagoccocus for which the name Vagoccocus entomophilus sp. nov. is proposed. The type strain is VOSTP2(T) ( = DSM 24756(T) = CCM 7946(T)).

Aquitalea pelogenes sp. nov., isolated from mineral peloid

Strain P1297T was isolated in the frame of a project aimed on the psychrotolerant microbiota occurring in water sources. The strain initially identified as a tentative species of the genus Aeromonas was rod-shaped, Gram-stain-negative, facultatively anaerobic and oxidase-positive. Subsequently, 16S rRNA gene sequence analysis placed strain P1297T within the class Betaproteobacteria and showed Aquitalea magnusonii TRO-001DR8T as the closest phylogenetic relative with 99.28 % 16S rRNA gene sequence similarity. Digital DDH and average nucleotide identity (ANI) were determined to evaluate the genomic relationship between strain P1297T and Aquitalea magnusonii CCM 7607T. Digital DDH estimation (31.3 ± 2.46 %) as well as ANI (85.6001 %; reciprocal value 85.3277 %) proved the dissimilarity of strain P1297T. Further investigation using phenotyping, automated ribotyping, whole-cell protein profiling and PCR-fingerprinting methods showed a distinct taxonomic position of strain P1297T among hitherto described species of the genus Aquitalea. DNA-DNA hybridization experiments revealed low binding values between strain P1297T and Aquitalea magnusonii CCM 7607T (57 ± 3 %) and Aquitalea denitrificans CCM 7935T (41 ± 5 %). The DNA G+C content of strain P1297T was 60.3 mol%. The predominant fatty acids were C16 : 1ω7c/ iso-C15 : 0 2-OH (47.0 %), C16 : 0 (24.5 %) and C18 : 1ω7c (10.6 %), and the quinone system contained predominantly ubiquinone Q-8. The polar lipids detected were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, two unidentified phospholipids and one unidentified aminophospholipid. Obtained results of genotypic and chemotaxonomic methods clearly proved that strain P1297T represents a novel species of the genus Aquitalea, for which the name Aquitalea pelogenes sp. nov. is proposed. The type strain is P1297T ( = CCM 7557T = LMG 28989T = CCUG 67440T).

Classification of strain CCM 4446T as Rhodococcus degradans sp. nov.

Strain CCM 4446T, with notable biodegradation capabilities, was investigated in this study in order to elucidate its taxonomic position. Chemotaxonomic analyses of quinones, polar lipids, mycolic acids, polyamines and the diamino acid of the cell-wall peptidoglycan corresponded with characteristics of the genus Rhodococcus. Phylogenetic analysis, based on the 16S rRNA gene sequence, assigned strain CCM 4446T to the genus Rhodococcus and placed it in the Rhodococcus erythropolis 16S rRNA gene clade. Further analysis of catA and gyrB gene sequences, automated ribotyping with EcoRI restriction endonuclease, whole-cell protein profiling, DNA-DNA hybridization and extensive biotyping enabled differentiation of strain CCM 4446T from all phylogenetically closely related species, i.e., Rhodococcus baikonurensis, Rhodococcus qingshengii, Rhodococcus erythropolis and Rhodococcus globerulus. The results obtained show that the strain investigated represents a novel species within the genus Rhodococcus, for which the name Rhodococcus degradans sp. nov., is proposed. The type strain is CCM 4446T ( = LMG 28633T).

Description of Pseudomonas gregormendelii sp. nov., a Novel Psychrotrophic Bacterium from James Ross Island, Antarctica.

During the microbiological research performed within the scope of activities of Czech expeditions based at the Johann Gregor Mendel Station at James Ross Island, Antarctica, two psychrotrophic gram-stain negative non-fluorescent strains CCM 8506T and CCM 8507 from soil were extensively characterized using genotypic and phenotypic methods. Initial characterization using ribotyping with HindIII restriction endonuclease and phenotyping implies that both isolates belong to a single Pseudomonas species. Sequencing of rrs, rpoB, rpoD and glnA genes of strain CCM 8506T confirmed affiliation of investigated strains within the genus Pseudomonas. Further investigation using automated ribotyping with EcoRI (RiboPrinter® Microbial Characterisation System), whole-cell protein profiling using the Agilent 2100 Bioanalyzer system, extensive biochemical testing and DNA–DNA hybridization experiments confirmed that both investigated strains are members of a single taxon which is clearly separated from all hitherto described Pseudomonas spp. Based on all findings, we describe a novel species Pseudomonas gregormendelii sp. nov. with the type strain CCM 8506T (=LMG 28632T).

Enterococcus alcedinis sp. nov., isolated from common kingfisher (Alcedo atthis)

Two Gram-positive, catalase-negative bacterial strains were isolated from the cloaca of common kingfishers (Alcedo atthis). Repetitive sequence-based PCR fingerprinting using the (GTG)5 primer grouped these isolates into a single cluster separated from all known enterococcal species. The two strains revealed identical 16S rRNA gene sequences placing them within the genus Enterococcus with Enterococcus aquimarinus LMG 16607(T) as the closest relative (97.14 % similarity). Further taxonomic investigation using sequencing of the genes for the superoxide dismutase (sodA), phenylalanyl-tRNA synthase alpha subunit (pheS) and the RNA polymerase alpha subunit (rpoA) as well as application of whole-cell protein fingerprinting, automated ribotyping and extensive phenotyping confirmed that both strains belong to the same species. Based on data from this polyphasic study, these strains represent a novel species of the genus Enterococcus, for which the name Enterococcus alcedinis sp. nov. is proposed. The type strain is L34(T) (= CCM 8433(T) = LMG 27164(T)).

Staphylococcus petrasii subsp. pragensis subsp. nov., occurring in human clinical material

Seven coagulase-negative, oxidase-negative and novobiocin-susceptible staphylococci assigned tentatively as Staphylococcus petrasii were investigated in this study in order to elucidate their taxonomic position. All strains were initially shown to form a genetically homogeneous group separated from remaining species of the genus Staphylococcus by using a repetitive sequence-based PCR fingerprinting with the (GTG)5 primer. Phylogenetic analysis based on 16S rRNA gene, hsp60, rpoB, dnaJ, gap and tuf sequences showed that the group is closely related to Staphylococcus petrasii but separated from the three hitherto known subspecies, S. petrasii subsp. petrasii, S. petrasii subsp. croceilyticus and S. petrasii subsp. jettensis. Further investigation using automated ribotyping, MALDI-TOF mass spectrometry, fatty acid methyl ester analysis, DNA-DNA hybridization and extensive biotyping confirmed that the analysed group represents a novel subspecies within S. petrasii, for which the name Staphylococcus petrasii subsp. pragensis subsp. nov. is proposed. The type strain is NRL/St 12/356(T) ( = CCM 8529(T) = LMG 28327(T)).

Isolation of human pathogen Escherichia albertii from faeces of seals (Leptonychotes weddelli) in James Ross Island, Antarctica

A set of nine gram-negative fermenting rods biochemically identified as Escherichia coli was isolated from faeces of seals. These bacteria were characterized by phenotypic classification, 16S rDNA sequence analyses, automated ribotyping, study of whole-cell protein profiles by SDS-PAGE and finally by bacteriocin production. The results of our polyphasic taxonomic study supported the recognition of P4652, P4653 and P4740 isolates as true members of Escherichia albertii species – probably a major enteric human pathogen. To our best knowledge, this is the first evidence showing that E. albertii produces bacteriocin, and colicin D. Obtained data unambiguously showed inconvenience of commercial identification systems to distinguish both Escherichia species due to missing data of E. albertii in the commercial databases. The results of Escherichia isolates taxonomy suggest seals as a novel source of human and animal pathogen, E. albertii in the Antarctic region.