Erika M. Tóth

Wohlfahrtiimonas chitiniclastica gen. nov., sp. nov., a new gammaproteobacterium isolated from Wohlfahrtia magnifica (Diptera: Sarcophagidae)

New Gammaproteobacteria were isolated from 3rd stage fly larvae of the parasitic fly Wohlfahrtia magnifica. Phylogenetic analysis of the new isolates showed that these bacteria belong to a distinct lineage close to Ignatzschineria larvae, which was originally isolated from the same species of fly. The low similarity values in 16S rRNA gene sequences (93.8-94.8 %), and differences in fatty acid profiles, RiboPrint patterns, MALDI-TOF mass spectra of cell extracts, and physiological and biochemical characteristics differentiate the isolates from the type strain of Ignatzschineria larvae (DSM 13226T), and indicate that our isolates represent a new genus within the Gammaproteobacteria. The major isoprenoid quinone of the strains is Q8, the major fatty acids are C18 : 1 and C14 : 0, and the predominant polar lipids are phosphatidylglycerol, phosphatidylethanolamine and phosphatidylserine. The G+C content of the DNA of the type strain is 44.3 mol%. The name Wohlfahrtiimonas chitiniclastica gen. nov., sp. nov., is proposed for this novel genus and species. The type strain is S5T (=DSM 18708T=CCM 7401T).

Performance characteristics of qPCR assays targeting human- and ruminant-associated Bacteroidetes for microbial source tracking across sixteen countries on six continents

Numerous quantitative PCR assays for microbial fecal source tracking (MST) have been developed and evaluated in recent years. Widespread application has been hindered by a lack of knowledge regarding the geographical stability and hence applicability of such methods beyond the regional level. This study assessed the performance of five previously reported quantitative PCR assays targeting human-, cattle-, or ruminant-associated Bacteroidetes populations on 280 human and animal fecal samples from 16 countries across six continents. The tested cattle-associated markers were shown to be ruminant-associated. The quantitative distributions of marker concentrations in target and nontarget samples proved to be essential for the assessment of assay performance and were used to establish a new metric for quantitative source-specificity. In general, this study demonstrates that stable target populations required for marker-based MST occur around the globe. Ruminant-associated marker concentrations were strongly correlated with total intestinal Bacteroidetes populations and with each other, indicating that the detected ruminant-associated populations seem to be part of the intestinal core microbiome of ruminants worldwide. Consequently tested ruminant-targeted assays appear to be suitable quantitative MST tools beyond the regional level while the targeted human-associated populations seem to be less prevalent and stable, suggesting potential for improvements in human-targeted methods.

Schineria larvae gen. nov., sp. nov., isolated from the 1st and 2nd larval stages of Wohlfahrtia magnifica (Diptera: Sarcophagidae).

Four bacterial strains were isolated from the fly larvae of an obligate parasitic fly, Wohlfahrtia magnifica (Diptera: Sarcophagidae). These isolates were characterized by a polyphasic approach and represent a new lineage of gamma-Proteobacteria as their closest relative is Xylella fastidiosa (87.1% 16S rDNA similarity). The four strains are identical at the 16S rDNA level, the level of similarity between them, based on DNA-DNA hybridization, is high (97.8-102.5%) and they are similar in their physiological and biochemical characteristics, although they differ in their utilization of different sole carbon sources. All produce chitinase. They are obligately aerobic: no growth is detected under anaerobic conditions, even in the presence of NO3- as terminal electron acceptor. Their predominant respiratory quinone is Q-8. The G+C content of their DNA is 42 mol%. Their cell membrane contains phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylserine and two unknown polar lipids. Their main fatty acids are C18:1, C16:0 and C14:0. To accommodate these bacteria, a new genus, Schineria gen. nov., with the type species Schineria larvae sp. nov., is proposed.

Diversity of Alkaliphilic and Alkalitolerant Bacteria Cultivated from Decomposing Reed Rhizomes in a Hungarian Soda Lake

Bacterial communities associated with decomposing rhizomes of Phragmites australis were investigated in Lake Fertő (Neusiedlersee, Hungary). Alkaliphilic and alkalitolerant strains were isolated on cellulose-containing alkaline medium spread with dilutions of scrapings taken from the surface of the decaying plant material. Fifty-one strains were grouped by numerical analysis based on physiological tests and BIOLOG sole carbon source utilization data. The strains identified by 16S rDNA sequence comparisons included members of low G+C Gram positives (Marinibacillus marinus, Bacillus cereus, and Exiguobacterium aurantiacum), high G+C Gram positives (Nesterenkonia halobia and Dietzia natronolimnea), α-proteobacteria (Pannonibacter phragmitetus), and γ-proteobacteria (Pseudomonas pseudoalcaligenes and Halomonas venusta). Most of the strains were characterized by aerobic chemoorganotrophic respiratory metabolism and utilized several different carbon sources, although no direct cellulolytic activity was observed. Results of the pH and salt tolerance tests revealed optimuma in most cases at pH 11 and at the presence of 2.5–5% NaCl. These bacteria probably occupy niches in the aerobic, alkaline, water-influenced environments on the decomposing reed surfaces.

Bacteria Isolated from the Different Developmental Stages and Larval Organs of the Obligate Parasitic Fly, Wohlfahrtia magnifica (Diptera: Sarcophagidae)

Wohlfahrtia magnifica (Diptera: Sarcophagidae) is the major myiasis-causing fly species in the whole of Eurasia for most important domestic animals. The aim of the present work was to obtain data on the culturable bacteria isolated under aerobic conditions from this fly: bacteria were isolated from all developmental stages (larvae, pupa, and imago) of Wohlfahrtia magnifica, and the third-stage larval organs were also sampled. To determine the possible antagonistic effects between the dominant bacterial groups, an antibiosis assay was carried out. Plating and isolation of bacteria was performed by classical microbiological methods. Characterization of the isolated strains was carried out via a polyphasic approach; classical phenotypic tests, chemotaxonomical examinations, and 16S rDNA sequence analyses were also applied. In the case of maggot macerate samples, members of the family Enterobacteriaceae were characteristic. Members of a new genus (Schineria) belonging to the γ subdivision of proteobacteria were also isolated. According to our data, the shifts in the Schineria and Proteus populations within the larvae are strongly influenced by their interactions with each other and among the members of the family Enterobacteriaceae. The pupa and imago samples contained several other Gram-negative bacteria (Stenotrophomonas, Brevundimonas, etc.). Among Gram-positive bacteria, in all maggot macerate samples, members of the genus Bacillus and the Arthrobacter–Micrococcus group of actinobacteria were dominant (neither of them was a producer or sensitive to the compounds of other microorganisms), and bacteria related to the genus Corynebacterium were also found. From the larvae Aureobacterium liquefaciens and Enterococcus faecalis were isolated, and from the pupae Dietzia maris and Enterococcus faecalis. In the samples of third-stage larval organs, the dominant groups were the same as in the third-stage larval macerate sample; however, several additional genera/species were observed (Rhodococcus fascians, Streptomyces sp., Rathayibacter sp., Bacillus thuringiensis/cereus).

Bacillus aurantiacus sp. nov., an alkaliphilic and moderately halophilic bacterium isolated from Hungarian soda lakes

Three alkaliphilic and moderately halophilic strains designated K1-5T, K1-10 and B1-1, characterized by optimal growth at pH 9.0-10.0 and at 3-7 % (w/v) NaCl, were isolated from extremely shallow, alkaline soda lakes located in Hungary. Cells of the strains are Gram-positive, straight rods and form a central to subterminal, ellipsoidal endospore. The isolates are strictly aerobic, catalase-positive, oxidase-negative and contain a peptidoglycan of type A1 gamma based on meso-diaminopimelic acid. In strain K1-5T, menaquinone-7 (MK-7) is the predominant isoprenoid quinone and anteiso-C15 : 0 is the major cellular fatty acid. The DNA G+C content of strain K1-5T is 42.9 mol%. 16S rRNA gene-based phylogenetic analysis revealed that the strains exhibit levels of sequence similarity of less than 95.8 % to known Bacillus species. According to the polyphasic characterization, the strains represent a novel species, for which the name Bacillus aurantiacus sp. nov. is proposed. The type strain is K1-5T (=DSM 18675T =CCM 7447T =NCAIM B002265T).

Nocardioides daphniae sp. nov., isolated from Daphnia cucullata (Crustacea: Cladocera)

A Gram-positive, rod-shaped or coccoid, yellow-pigmented bacterial strain, D287(T), was isolated from the water flea Daphnia cucullata (Crustacea: Cladocera) collected from Lake Balaton in Hungary. Phylogenetic analysis on the basis of 16S rRNA gene sequence comparisons revealed that the strain represented a distinct lineage within the cluster of the genera Nocardioides and Marmoricola. The following characteristics were consistent with the affiliation of strain D287(T) to the genus Nocardioides: peptidoglycan based on LL-2,6-diaminopimelic acid, MK-8(H(4)) as the major menaquinone, iso-C(16:0) as the predominant cellular fatty acid, the presence of phosphatidylglycerol and diphosphatidylglycerol and a DNA G+C content of 69.9 mol%. Owing to characteristic differences in physiological traits and levels of 16S rRNA gene sequence similarity to its phylogenetically closest neighbours that were below 97%, strain D287(T) is considered to represent a novel species of the genus Nocardioides, for which the name Nocardioides daphniae sp. nov. is proposed. The type strain is D287(T) (=DSM 18664(T)=CCM 7403(T)).

Microbiological investigation of an industrial ultra pure supply water plant using cultivation-based and cultivation-independent methods

Ultra pure waters (UPW), characterized by extremely low salt and nutrient concentrations, can suffer from microbial contamination which causes biofouling and biocorrosion, possibly leading to reduced lifetime and increased operational costs. Samples were taken from an ultra pure supply water producing plant of a power plant. Scanning electron microscopic examination was carried out on the biofilms formed in the system. Biofilm, ion exchange resin, and water samples were characterized by culture-based methods and molecular fingerprinting (terminal restriction fragment length polymorphism [T-RFLP] analysis and molecular cloning). Identification of bacteria was based on 16S rDNA sequence comparison. A complex microbial community structure was revealed. Nearly 46% of the clones were related to as yet uncultured bacteria. The community profiles of the water samples were the most diverse and most of bacteria were recruited from bacterial communities of tube surface and ion exchange resin biofilms. Microbiota of different layers of the mixed bed ion exchange resin showed the highest similarity. Most of the identified taxa (dominated by β-Proteobacteria) could take part in microbially influenced corrosion.

Thermobifida cellulolytica sp. nov., a novel lignocellulose-decomposing actinomycete

Four actinomycete strains, isolated from the overheated region of manure compost, were assigned to the genus Thermobifida on the basis of morphological, physiological and biochemical characteristics. All strains produced single, ovoid, heat-sensitive spores on dichotomically branched aerial hyphae. On the basis of chemotaxonomic traits, these isolates showed strong affinity towards members of the genus Thermobifida. Cell-wall analysis revealed the presence of meso-diaminopimelic acid, but no other characteristic amino acids or sugars in the murein (cell wall type III). According to polar lipid analysis, all strains showed PL II-type phospholipid composition; phosphatidylethanolamine and glycolipid were detected together with some unidentified phospholipids. The isoprenoid quinone composition of the new isolates differed slightly from that of the other two Thermobifida species described thus far. The partial 16S rDNA sequence similarity of the four strains reached 99.8-100%, whereas a nearly complete 16S rDNA sequence of TB100T, the representative strain of this collection, showed only 97.4 and 97.8% similarity to the corresponding rDNA sequences of the type strains of Thermobifida fusca and Thermobifida alba, respectively. These four isolates constituted a homogeneous group with levels of DNA-DNA homology ranging from 94.6 to 99.1%. The DNA-DNA relative homology values of strain TB100T to Thermobifida fusca ATCC 27730T and Thermobifida alba DSM 43795T were 48.1 and 57%, respectively. On the basis of phenotypic, chemotaxonomic and genotypic data, the strains are assigned to a new species within the genus Thermobifida under the name Thermobifida cellulolytica sp. nov. The type strain is TB100T (= DSM 44535T = NCAIM B01997T).

Chryseobacterium hungaricum sp. nov., isolated from hydrocarbon-contaminated soil

The taxonomic position of a strain isolated from kerosene-contaminated soil in Hungary and formerly misidentified as Brevundimonas vesicularis was examined using a polyphasic approach. The isolate, designated CHB-20p(T), could be clearly assigned to the genus Chryseobacterium (family Flavobacteriaceae) on the basis of 16S rRNA gene sequence similarity. Strain CHB-20p(T), a moderate oil degrader, was a Gram-negative, aerobic, mesophilic microbe with a temperature optimum of 28-30 degrees C. Predominant fatty acids were iso-C(15 : 0), summed feature 3 (comprising C(16 : 1)omega7c and/or iso-C(15 : 0) 2-OH) and iso-C(17 : 0) 3-OH. Menaquinone-6 (MK-6) was the predominant respiratory quinone; MK-5 was present as a minor component. The almost complete 16S rRNA gene sequence of strain CHB-20p(T) shared 94-97 % similarity with sequences of the type strains of species of the genus Chryseobacterium. DNA-DNA relatedness between strain CHB-20p(T) and its closest relative, Chryseobacterium caeni, was lower than 46 %. Moreover, several diagnostic phenotypic properties distinguished strain CHB-20p(T) from C. caeni. On the basis of biochemical, chemotaxonomic and genotypic data, isolate CHB-20p(T) represents a novel species within the genus Chryseobacterium, Chryseobacterium hungaricum sp. nov.; the type strain is CHB-20p(T) (=NCAIM B2269(T)=DSM 19684(T)).