16S 23S Research Articles

Molecular Evidence of Chlamydia-Like Organisms in the Feces of Myotis daubentonii Bats.

Several pathogenic viruses are known to colonize bats, and recent analyses indicate that bats are also reservoir hosts for bacterial genera. Chlamydia-like organisms (CLOs) have been detected in several animal species. CLOs have high 16S rRNA sequence similarity to Chlamydiaceae and exhibit similar intracellular lifestyles and biphasic developmental cycles. Our study describes the frequent occurrence of CLO DNA in bat feces, suggesting an expanding host species spectrum for the Chlamydiales As bats can acquire various infectious agents through their diet, prey insects were also studied. We identified CLO sequences in bats that matched best with sequences in prey insects but also CLO sequences not detected in prey insects. This suggests that a portion of CLO DNA present in bat feces is not prey borne. Furthermore, some sequences from bat droppings not originating from their diet might well represent novel, previously unidentified members of the Chlamydiales order.

Site-Specific Cleavage of Ribosomal RNA in Escherichia coli-Based Cell-Free Protein Synthesis Systems.

Cell-free protein synthesis, which mimics the biological protein production system, allows rapid expression of proteins without the need to maintain a viable cell. Nevertheless, cell-free protein expression relies on active in vivo translation machinery including ribosomes and translation factors. Here, we examined the integrity of the protein synthesis machinery, namely the functionality of ribosomes, during (i) the cell-free extract preparation and (ii) the performance of in vitro protein synthesis by analyzing crucial components involved in translation. Monitoring the 16S rRNA, 23S rRNA, elongation factors and ribosomal protein S1, we show that processing of a cell-free extract results in no substantial alteration of the translation machinery. Moreover, we reveal that the 16S rRNA is specifically cleaved at helix 44 during in vitro translation reactions, resulting in the removal of the anti-Shine-Dalgarno sequence. These defective ribosomes accumulate in the cell-free system. We demonstrate that the specific cleavage of the 16S rRNA is triggered by the decreased concentrations of Mg2+. In addition, we provide evidence that helix 44 of the 30S ribosomal subunit serves as a point-of-entry for ribosome degradation in Escherichia coli. Our results suggest that Mg2+ homeostasis is fundamental to preserving functional ribosomes in cell-free protein synthesis systems, which is of major importance for cell-free protein synthesis at preparative scale, in order to create highly efficient technical in vitro systems.

Identification of Bifidobacterium Animalis Ssp. Lactis from Egyptian Women Breast Milk and Feces of Breast Fed Infant Based On 16S-23S rRNA Gene

Bifidobacterium represent one of the major genera of the intestinal tract of human and animals used as probiotics in dairy and nondairy foods for restore the intestinal microflora which confers a health benefit. The identification of Bifidobacterium by phenotypic features is commonly unreliable, time, money, and effort consuming. We sought to improve the Bifidobacterium identification method based on molecular level to identify probiotic bacteria in complex microbial communities. The application of 16S-23S rRNA oligonucleotide primers is the best and most reliable, rapid, and precise species and sub species identification approach. The ribosomal intergenic spacer region (ISR) located between the highly conserved 16S rRNA and 23S rRNA shows a high degree of variation in length and sequence and potential for intra species discrimination and providing the phylogenetic Relationship of the Genus Bifidobacterium spp. Results showed that one of the two primer sets Bflac2-Bflac5 species specific gives positive results differentiating between B. animalis ssp. Lactis isolated from breast fed infants milk of human and that isolated from feces of breast fed infant and detecting reference strain for B. animalis ssp. Lactis DSM10140. DNA sequences of the two strains were submitted to the Genbank NCBI under accession number (KT758845) named as B. animalis ssp. Lactis Egm1 (Egyptian milk) and accession number (KT758846) named as Egf1 Egyptian feces while the second primer give false positive result. Also, we aim to obtain patent protection under Intellectual property rights (IPRs) for B. animalis ssp. Lactis which was isolated from Egyptian resources to be used for a better and healthier food and dairy products.

Identification of Bifidobacterium Animalis Ssp. Lactis from Egyptian Women Breast Milk and Feces of Breast Fed Infant Based On 16S-23S rRNA Gene

Bifidobacterium represent one of the major genera of the intestinal tract of human and animals used as probiotics in dairy and nondairy foods for restore the intestinal microflora which confers a health benefit. The identification of Bifidobacterium by phenotypic features is commonly unreliable, time, money, and effort consuming. We sought to improve the Bifidobacterium identification method based on molecular level to identify probiotic bacteria in complex microbial communities. The application of 16S-23S rRNA oligonucleotide primers is the best and most reliable, rapid, and precise species and sub species identification approach. The ribosomal intergenic spacer region (ISR) located between the highly conserved 16S rRNA and 23S rRNA shows a high degree of variation in length and sequence and potential for intra species discrimination and providing the phylogenetic Relationship of the Genus Bifidobacterium spp. Results showed that one of the two primer sets Bflac2-Bflac5 species specific gives positive results differentiating between B. animalis ssp. Lactis isolated from breast fed infants milk of human and that isolated from feces of breast fed infant and detecting reference strain for B. animalis ssp. Lactis DSM10140. DNA sequences of the two strains were submitted to the Genbank NCBI under accession number (KT758845) named as B. animalis ssp. Lactis Egm1 (Egyptian milk) and accession number (KT758846) named as Egf1 Egyptian feces while the second primer give false positive result. Also, we aim to obtain patent protection under Intellectual property rights (IPRs) for B. animalis ssp. Lactis which was isolated from Egyptian resources to be used for a better and healthier food and dairy products.

Comparison of methods for the identification and sub-typing of O157 and non-O157 Escherichia coli serotypes and their integration into a polyphasic taxonomy approach

AbstractPhenotypic, chemotaxonomic and genotypic data from 12 strains ofEscherichia coli werecollected, including carbon source utilisation profiles, ribotypes, sequencing data of the 16S–23S rRNA internal transcribed region (ITS) and Fourier transform-infrared (FT-IR) spectroscopic profiles. The objectives were to compare several identification systems forE. coliand to develop and test a polyphasic taxonomic approach using the four methodologies combined for the sub-typing of O157 and non-O157E. coli. The nucleotide sequences of the 16S–23S rRNA ITS regions were amplified by polymerase chain reaction (PCR), sequenced and compared with reference data available at the GenBank database using the Basic Local Alignment Search Tool (BLAST) . Additional information comprising the utilisation of carbon sources, riboprint profiles and FT-IR spectra was also collected. The capacity of the methods for the identification and typing ofE. colito species and subspecies levels was evaluated. Data were transformed and integrated to present polyphasic hierarchical clusters and relationships. The study reports the use of an integrated scheme comprising phenotypic, chemotaxonomic and genotypic information (carbon source profile, sequencing of the 16S–23S rRNA ITS, ribotyping and FT-IR spectroscopy) for a more precise characterisation and identification ofE. coli. The results showed that identification ofE. colistrains by each individual method was limited mainly by the extension and quality of reference databases. On the contrary, the polyphasic approach, whereby heterogeneous taxonomic data were combined and weighted, improved the identification results, gave more consistency to the final clustering and provided additional information on the taxonomic structure and phenotypic behaviour of strains, as shown by the close clustering of strains with similar stress resistance patterns.

Phylogenetic position reevaluation of Kyrtuthrix and description of a new species K. huatulcensis from Mexico´s Pacific coast

Benthic marine heterocytous cyanoprokaryotes of Mexico´s tropical coast are being recognized as an important and conspicuous component of the supralittoral and intertidal zones usually described as an extreme and low diversity biotope. Although Kyrtuthrix has been reported from different coasts worldwide, its complex morphology has led to differing taxonomic interpretations and positioning. Ten marine supra and intertidal populations of Kyrtuthrix were analyzed using a detailed morphological approach, complemented with ecological and geographical information as well as DNA sequence data of the 16S rRNA gene and associated 16S–23S ITS. Kyrtuthrix huatulcensis is described as a new species, different from K. dalmatica Ercegovic and K. maculans (Gomont) Umezaki based primarily on morphological data. Our material has smaller dimensions in thalli, filaments, trichomes and cells, and possesses differences in qualitative characters as well. Our material is epilithic as in K. maculans but not endolithic as in K. dalmatica. The analysis of molecular data, mainly 16S rRNA gene sequence, show that this genus is within the Rivulariaceae clade, and outside of the Scytonemataceae clade. ITS secondary structure comparison with some of the closer representatives of Rivulariaceae shows considerable differences with these related genera. Based on the evidence gathered in this study, inclusion of Kyrtuthrix within the Rivulariaceae, rather than the Scytonemataceae, is fully supported.

High intraspecies heterogeneity within Staphylococcus sciuri and rejection of its classification into S. sciuri subsp. sciuri, S. sciuri subsp. carnaticus and S. sciuri subsp. rodentium.

A polyphasic taxonomic approach was applied to strains of the species Staphylococcus sciuri in order to clarify the taxonomic legitimacy of the delineation of S. sciuri into S. sciurisubsp.sciuri, S. sciurisubsp.carnaticus and S. sciurisubsp.rodentium. A group of 81 S. sciuri isolates obtained from human (n=62) and veterinary (n=17) clinical materials and foods (n=2) and ten reference and type strains obtained from the Czech Collection of Microorganisms were characterized by extensive biotyping using conventional tests and commercial identification kits (ID 32 Staph, STAPHYtest, Biolog Microbial ID System), matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry, automated ribotyping with EcoRI restriction enzyme, 16S-23S rRNA gene intergenic transcribed spacer PCR fingerprinting and repetitive sequence-based PCR fingerprinting with the (GTG)5 primer. Selected strains representing different ribotypes were further characterized using sequencing of the β-subunit of RNA polymerase (rpoB) gene. Individual techniques revealed high heterogeneity within the analysed S. sciuri strains but differentiation of the investigated strains into groups corresponding to the aforementioned S. sciuri subspecies and supported by these techniques was not clearly revealed. Based on obtained results and data retrieved from literature we propose rejecting the separation of S. sciuri species into S. sciurisubsp.sciuri, S. sciurisubsp.carnaticus and S. sciurisubsp.rodentium and we suggest reclassification these subspecies as S. sciuri with the type strain W.E. Kloos SC 116T (=ATCC 29062T=BCRC 12927T=CCM 3473T=CCUG 15598T=CNCTC 5683T=DSM 20345T=JCM 2425T=NCTC 12103T).

Molecular typing and genetic characterization of Mycobacterium avium subsp. hominissuis isolates from humans and swine in Japan.

Mycobacterium avium subsp. hominissuis (MAH) causes disease in both humans and swine; however, the genetic variations in MAH isolates are unclear. The aim of this study was to elucidate the genetic variations in MAH isolates from humans and swine in Japan. We analysed the 16S-23S rDNA internal transcribed spacer (ITS) sequence and variable number of tandem repeats (VNTRs) using the Mycobacterium avium tandem repeat loci, prevalence of ISMav6 and clarithromycin resistance for MAH isolates from patients with pulmonary MAC (pMAC) disease (n=69), and HIV-seropositive and blood culture-positive (HIV-MAC) patients (n=28) and swine (n=23). In the minimum spanning tree based on VNTR analysis, swine MAC isolates belonged to a cluster distinguishable from that of human pMAC isolates. Isolates from HIV-MAC were scattered throughout both clusters. The three major distinct sequevars, Mav-A, Mav-B and Mav-F, were determined according to 16S-23S rDNA ITS sequence analysis in addition to three new sequevars, Mav-Q, Mav-R and Mav-S. Mav-A and Mav-F comprised the majority of human pMAC strains; in contrast, Mav-B predominated in swine isolates. Distribution of ITS sequevars in the minimum spanning tree based on VNTR analysis showed similar clusters of isolates from different origins, i.e. human pMAC, HIV-MAC and swine. These results, together with ISMav6 possession and clarithromycin resistance, revealed the genetic diversity of MAH strains recovered from humans and swine. Molecular epidemiology and genetic characterization in the present study showed the distinctive genetic evolutionary lineage of MAH strains isolated from human pMAC diseases and swine.

Pistricoccus aurantiacus gen. nov., sp. nov., a moderately halophilic bacterium isolated from a shark.

A novel Gram-stain negative, non-motile, moderately halophilic, facultatively anaerobic and spherical bacterium designated strain SS9T was isolated from the gill homogenate of a shark. Cells of SS9T were observed to be 0.8-1.2μm in diameter. The strain was found to grow optimally at 33°C, pH 7.0-8.0 and in the presence of 6.0% (w/v) NaCl. On the basis of 16S rRNA gene phylogeny, strain SS9T can be affiliated with the family Halomonadaceae and is closely related to Chromohalobacter marismortui NBRC 103155T (95.6% sequence similarity), Halomonas ilicicola SP8T (95.6%) and Chromohalobacter salexigens DSM 3043T (95.5%). Multilocus sequence analysis of strain SS9T using the housekeeping genes 16S rRNA, 23S rRNA, gyrB, rpoD and secA revealed the strain's distinct phylogenetic position, separate from other known genera of the family Halomonadaceae. Strain SS9T was found to contain ubiquinone-9 (Q-9) as the predominant ubiquinone and C18:1 ω7c, C16:0 and summed feature 3 (C16:1 ω7c and/or iso-C15:0 2-OH) as the major fatty acids. The major polar lipids of strain SS9T were identified as phosphatidylglycerol and phosphatidylethanolamine. The DNA G+C content of strain SS9T was determined to be 60.4mol%. It is evident from phylogenetic, genotypic, phenotypic and chemotaxonomic results that strain SS9T represents a novel species in a new genus, for which the name Pistricoccus aurantiacus gen. nov., sp. nov. is proposed. The type strain is SS9T (=KCTC 42586T=MCCC 1H00111T).

Detection of the new cosmopolitan genus Thermoleptolyngbya (Cyanobacteria, Leptolyngbyaceae) using the 16S rRNA gene and 16S–23S ITS region

Cyanobacteria are widespread prokaryotes that are able to live in extreme conditions such as thermal springs. Strains attributable to the genus Leptolyngbya are among the most common cyanobacteria sampled from thermal environments. Leptolyngbya is a character-poor taxon that was demonstrated to be polyphyletic based on molecular analyses. The recent joining of 16S rRNA gene phylogenies with 16S–23S ITS secondary structure analysis is a useful approach to detect new cryptic taxa and has led to the separation of new genera from Leptolyngbya and to the description of new species inside this genus and in other related groups.In this study, phylogenetic investigations based on both the 16S rRNA gene and the 16S–23S ITS region were performed alongside 16S rRNA and 16S–23S ITS secondary structure analyses on cyanobacteria of the family Leptolyngbyaceae. These analyses focused on filamentous strains sampled from thermal springs with a morphology ascribable to the genus Leptolyngbya. The phylogenetic reconstructions showed that the Leptolyngbya-like thermal strains grouped into a monophyletic lineage that was distinct from Leptolyngbya. The 16S–23S ITS secondary structure results supported the separation of this cluster. A new genus named Thermoleptolyngbya was erected to encompass these strains, and two species were described inside this new taxon: T. albertanoae and T. oregonensis.

Porphyromonas loveana sp. nov., isolated from the oral cavity of Australian marsupials.

An obligatory anaerobic, Gram-stain-negative coccobacillus with black-pigmented colonies was isolated from the oral cavity of selected Australian marsupial species. Phenotypic and molecular criteria showed that this bacterium was a distinct species within the genus Porphyromonas, and was closely related to Porphyromonas gingivalis and Porphyromonas gulae. This putative novel species and P. gulae could be differentiated from P. gingivalis by catalase activity. Further characterization by multi-locus enzyme electrophoresis of glutamate dehydrogenase and malate dehydrogenase enzyme mobility and matrix-assisted laser desorption ionization time-of-flight MS showed that this putative novel species could be differentiated phenotypically from P. gingivalis and P. gulae. Definitive identification by 16S rRNA gene sequencing showed that this bacterium belonged to a unique monophyletic lineage, phylogenetically distinct from P. gingivalis (94.9 % similarity) and P. gulae (95.5 %). This also was supported by 16S-23S rRNA intergenic spacer region and glutamate dehydrogenase gene sequencing. A new species epithet, Porphyromonas loveana sp. nov., is proposed for this bacterium, with DSM 28520T (=NCTC 13658T=UQD444T=MRK101T), isolated from a musky rat kangaroo, as the type strain.

Unexpected structured intraspecific diversity of thioautotrophic bacterial gill endosymbionts within the Lucinidae (Mollusca: Bivalvia)

In chemoautotrophic associations, sequence comparison of 16S rRNA has been the method of choice to study bacterial diversity in the context of host/symbiont coevolution. However, the relative low rate of evolution of 16S rRNA has been shown to result in a diminished capacity to discriminate between closely related bacterial strains or species. Within chemoautotrophic associations, as described in several studies, the use of other genetic markers may reveal previously unobserved strain diversity among gill endosymbionts. Herein, we conducted a survey of symbionts harbored by six species within the Lucinidae family using five genetic markers (dnaE, gyrB, aprA, cbbL genes and the 16S–23S internal transcribed spacer). Thus, within the already described SoLuc_1 bacterial species shared by six host species we observed an obvious bacterial strain diversity. This diversity is structured in function of the geographic location of the hosts and not in function of ecological parameters or host phylogeny. Interestingly at the local level (same seagrass bed environment), we observed an unexpected specificity in host species/bacterial strain association (i.e., all individuals of the same host species harbor the same symbiotic sequence type). This specificity of association implied that there was a control of symbiont strain acquisition by the host, which was confirmed by a cross-infection experiment of starved adults performed in our study. Based upon our results and other evidence from the literature, we hypothesize that this pattern may be due to a “capture/escape” type antagonist evolution of the two partners.

First report of the melon stem rot disease in protected cultivation caused by Pseudomonas fluorescens

In recent years, melon culture in East China represented by Shanghai district has been severely affected by a stem rot disease. Symptoms were described as the presence of badly water-soaked soft rot associated with spilled brown juice in the later stages. Based on our disease diagnosis study, 30 bacteria were isolated from the stem tissues of affected plants and one bacterial pathogen, named EXXP-01, was highly virulent on melon stem dipped in bacterial suspension. This bacterium has caused significant damage in late autumn melon cultures, and a disease severity analysis presented as decay diameter of leaves and length of the stem rot suggested that the isolated pathogen not only caused extensive damage to the stem of melon, but also posed potential harm to the whole melon plant including seedlings and leaves. The strain EXXP-01 was also found to be most closely related to P. fluorescens based on physiological and morphological characterization, and also the sequencing analysis of 16S rDNA gene and 16S–23S rDNA internal transcribed spacer gene. Moreover, sequence analysis of another housekeeping gene, the RNA polymerase β-subunit gene (rpoB), indicated that P. fluorescens EXXP-01 could be classified as P. fluorescens bv. I. This confirmed that P. fluorescens is the causal agent of melon stem soft rot. Further bactericidal test in vitro suggested that the strain EXXP-01 has strong resistance to various germicides and should be worthy of attention.

Proposal of Helicobacter canicola sp. nov., previously identified as Helicobacter cinaedi, isolated from canines

During the course of our taxonomic investigation of Helicobacter cinaedi, it was realized that the strains isolated from dogs, which have been identified as H. cinaedi, showed different biochemical traits than did the isolates obtained from humans. None of the three dog isolates could reduce nitrate to nitrite, whereas all of the human H. cinaedi isolates could do so. The dog isolates showed a strong positive alkaline phosphatase reaction and could grow at 42°C, however the human isolates showed negative to very weak responses to those tests. The GyrA protein based phylogenetic analysis showed that the three isolates from dogs formed a slightly distinct cluster from the human isolate cluster. Phylogenetic analysis of the 16S rRNA, 23S rRNA, gyrB, and hsp60 gene sequences further confirmed that the dog isolates differed from the human H. cinaedi isolate cluster. The whole-genome in silico DNA similarities of each isolate based on their full genome sequences revealed that the isolates from dogs shared more than 94.9% ANIb (average nucleotide identity based on BLAST), while 94.0% ANIb were found between the isolates from dogs and the humans, including the H. cinaedi type strain ATCC BAA-847T (=CCUG 18818T). From these data, we propose a new species, ‘H. canicola’ sp. nov., for the isolates from dogs. The type strain is PAGU 1410T (CCUG 33887T=LMG 29580T).

Genetic diversity of Bartonella genotypes found in the striped field mouse (Apodemus agrarius) in Central Europe.

We investigated the diversity of Bartonella in Apodemus agrarius, an important rodent of peri-domestic habitats, which has spread into Europe in the past 1000 years. Spleen samples of 344 A. agrarius from Eastern Slovakia were screened for the presence of Bartonella spp. using 16S-23S rRNA internal transcribed spacer region and bacteria were detected in 9% of rodents. Based on sequencing of three housekeeping genes (gltA, rpoB and groEL) Bartonella genotypes were ascribed to the species typical for mice and voles: B. grahamii, B. taylorii and B. birtlesii. However, the study also confirmed presence of genotypes belonging to the B. clarridgeiae/B. rochalimae clade, and the B. elizabethae/B. tribocorum clade, which are not commonly found in woodland rodents. In addition, a potential recombination event between these two genotypes was noted, which highlights an important role of A. agrarius in shaping Bartonella diversity and evolution.

Description and prevalence of Mycoplasma ciconiae sp. nov. isolated from white stork nestlings (Ciconia ciconia).

The mycoplasma strain ST 57T was isolated from the trachea of a clinically healthy, free-ranging white stork nestling in Nielitz, Mecklenburg-Western Pomerania, Germany. Strain ST 57T grew in fried-egg-shaped colonies on mycoplasma (SP4) agar plates and was dependent on sterol for growth. The organism fermented glucose and did not hydrolyse arginine or urea. The optimal growth temperature was 37 °C, with a temperature range from 23 to 44 °C. Strain ST 57Tcould not be identified as a representative of any of the currently described mycoplasma species by alignment of the 16S rRNA gene sequence or 16S-23S intergenic transcribed spacer region, or by immunobinding assays. Thus, this organism appears to be a representative of a novel species, for which the name Mycoplasma ciconiae sp. nov. is proposed. The type strain is ST 57T (=ATCC BAA-2401T=DSM 25251T). Four further strains of this species are included in this description (ST 24=DSM 29908, ST 56 Clone 1=DSM 29054, ST 99=DSM 29909, ST 102=DSM 29010). The prevalence of this mycoplasma species in clinically healthy, white stork nestlings in northern Germany was determined. Our species-specific PCR detected 57.8 % (48/83) of the samples positive for M. ciconiae sp. nov. As this species appears to be widespread in the healthy free-ranging white stork population, we conclude that this species is either apathogenic or an opportunistic pathogen in white storks.

Distribution, diversity and population composition of soybean-nodulating bradyrhizobia from different agro-climatic regions in Ethiopia

The genetic diversity and population composition of bradyrhizobial isolates collected from different parts of north-western and southern Ethiopia were studied. A total of 103 bacterial symbionts were trapped from the soils collected from Ethiopia. Genetic diversity and population composition of the bradyrhizobial isolates were assessed using restriction fragment length polymorphism (RFLP) of 16S–23S rRNA region. The results showed the presence of 30 distinct restriction pattern types in the population. The difference in bradyrhizobial communities between pairs of soil samples were estimated by alpha (H′α), beta (H′β) and gamma (H′γ) diversity. Southern Ethiopia showed the most diverse bradyrhizobial populations based on the many RFLP clusters occupied. The ratio of beta to gamma (H′β/ H′γ) diversity between the bradyrhizobial community compositions was greater in north-western than in the southern region of Ethiopia. The RFLP-based population composition suggested the frequent presence of individuals with admixture of ITS (16S–23S rDNA) region and showed the interlineage transfer of ITS genomic region. Phylogenetic analysis of 16S–23S rRNA sequences revealed the presence of a heterogenous group of Bradyrhizobium in Ethiopian soils. These results have provided new insight into the ecology of Bradyrhizobium nodulating soybean under different environmental conditions in Ethiopia.

16S-gyrB-rpoB multilocus sequence analysis for species identification in the genus Microbispora.

The genus Microbispora has been considered difficult to define taxonomically. While 16S rRNA gene analysis is required to determine phylogenetic relationships among species in this genus, most 16S rRNA gene-based phylogenetic tree topologies are not reliable. The genus Microbispora currently contains eight species along with six reclassified species (Microbispora chromogenes, Microbispora diastatica, Microbispora parva, Microbispora indica, Microbispora karnatakensis, Microbispora rosea) and Microbispora rosea subsp. aerata, a taxon composed of three further reclassified species (Microbispora aerata, Microbispora thermodiastatica, and Microbispora thermorosea). 16S rRNA, 23S rRNA, gyrB, and rpoB gene sequences were obtained for the type strains of Microbispora species, and eleven endophytic isolates from a Brazilian medicinal plant, Vochysia divergens. Using the concatenated sequence, most Microbispora type strains could be distinguished with high probability support. Based on these analyses, we propose that five of the species reclassified within the subspecies of M. rosea (M. chromogenes, M. karnatakensis, M. parva, M. aerata and M. thermorosea) are distinct from M. rosea and so should be retained as distinct species. The concatenated 16S-gyrB-rpoB gene phylogenic tree had significant probability support and topology. We propose the use of concatenated 16S-gyrB-rpoB gene sequences to determine phylogenetic relationships within the genus Microbispora. We also suggest that strains sharing >98.1% 16S-gyrB-rpoB gene sequences similarity be defined as a single species, based on results from this analysis. Seven of the strains isolated from V. divergens were not related to any previously described Microbispora species.

Reclassification of Bacillus isronensisShivaji et al. 2009 as Solibacillus isronensis comb. nov. and emended description of genus Solibacillus Krishnamurthi et al. 2009.

An investigation into the taxonomic position of Bacillus isronensis MTCC 7902T revealed that the strain shares a common phylogenetic lineage with Solibacillus silvestris MTCC 10789T. It displays considerable overlap in phenotypic properties with the genus Solibacillus, including endospore shape and position, oxidase and catalase activities, presence of iso-C15 : 0, C16 : 1ω7c alcohol and iso-C17 : 1ω7c as major cellular fatty acids, A4α-type cell-wall peptidoglycan, polar lipids and menaquinone pattern. These features reinforce the findings of molecular phylogenetic analyses based on 16S rRNA gene, gyrB gene and 16S-23S internal transcribed spacer (ITS) region sequences and, in line with the recommendations of Kämpfer et al. [Int J Syst Evol Microbiol 56 (2006), 781-786], provide justification for the transfer of Bacillus isronensis from the genus Bacillus to Solibacillus as Solibacillus isronensis comb. nov. The type strain is B3W22T ( = MTCC 7902T = DSM 21046T = JCM 13838T). An emended description of the genus Solibacillus is also provided.

Q-PCR Based Culture-Independent Enumeration and Detection of Enterobacter: An Emerging Environmental Human Pathogen in Riverine Systems and Potable Water.

The availability of safe and pristine water is a global challenge when large numbers of natural and anthropogenic water resources are being depleted with faster rate. The remaining water resources are severely contaminated with various kinds of contaminants including microorganisms. Enterobacter is one of the fecal coliform bacteria of family Enterobacteriaceae. Enterobacter was earlier used as an indicator bacterium along with other fecal Coliforms namely Escherichia coli, Citrobacter, and Klebsiella, but it is now known to cause various diseases in human beings. In this study, we have collected 55 samples from potable water and riverine system and proved their presence using their conserved sequences of 16S rRNA and 23S rRNA genes with the help of SYBR green real-time PCR, which showed very high specificity for the detection of Enterobacter. The Enterobacter counts in potable water were found to 1290 ± 32.89 to 1460 ± 39.42 cfu/100 ml. The Enterobacter levels in surface water were 1.76 × 104 ± 492, 1.33 × 104 ± 334, 1.15 × 104 ± 308, 2.56 × 104 ± 802, 2.89 × 104 ± 962, 8.16 × 104 ± 3443 cfu/100 ml; the levels of Enterobacter contamination associated with hydrophytes were 4.80 × 104 ± 1804, 3.48 × 104 ± 856, 8.50 × 104 ± 2074, 8.09 × 104 ± 1724, 6.30 × 104 ± 1738, 3.68 × 104 ± 949 cfu/10 g and the Enterobacter counts in sediments of the river, were 2.36 × 104 ± 703, 1.98 × 104 ± 530, 9.92 × 104 ± 3839, 6.80 × 104 ± 2230, 8.76 × 104 ± 3066 and 2.34 × 104 ± 732 cfu/10 g at the sampling Site #1, Site #2, Site #3, Site #4, Site #5, and Site #6, respectively. The assay could be used for the regular monitoring of potable water and other water reservoirs to check waterborne outbreaks.