Saccharopolyspora karakumensis sp. nov., Saccharopolyspora elongata sp. nov., Saccharopolyspora aridisoli sp. nov., Saccharopolyspora terrae sp. nov. and their biotechnological potential revealed by genome analysis

Exploration of unexplored habitats for novel actinobacteria with high bioactivity potential holds great promise in the search for novel entities. During the course of isolation of actinobacteria from desert soils, four actinobacteria, designated as 5K548^T, 7K502^T, 16K309^T and 16K404^T, were isolated from the Karakum Desert and their bioactivity potential as well as taxonomic provenances were revealed by comprehensive genome analyses. Pairwise sequence analyses of the 16S rRNA genes indicated that the four strains are representatives of putatively novel taxa within the prolific actinobacterial genus Saccharopolyspora. The strains have typical chemotaxonomic characteristics of the genus Saccharopolyspora by having meso-diaminopimelic acid as diagnostic diaminoacid, arabinose, galactose and ribose as whole-cell sugars. Consistent with this assignment, all of the isolates contained phosphatidylcholine in their polar lipid profiles and MK-9(H₄) as the predominant menaquinone. The sizes of the genomes of the isolates ranged from 6.0 to 10.2 Mb and the associated G + C contents from 69.6 to 69.7 %. Polyphasic characterizations including determination of overall genome relatedness indices revealed that the strains are representatives of four novel species in the genus Saccharopolyspora. Consequently, isolates 5K548^T, 7K502^T, 16K404^T and 16K309^T are proposed as novel Saccharopolyspora species for which the names of Saccharopolyspora karakumensis sp. nov., Saccharopolyspora elongata sp. nov., Saccharopolyspora aridisoli sp. nov. and Saccharopolyspora terrae sp. nov. are proposed, respectively. Comprehensive genome analysis for biosynthetic gene clusters showed that the strains have high potential for novel secondary metabolites. Moreover, the strains harbour many antimicrobial resistance genes providing more evidence for their potentiality for bioactive metabolites.     

Biofouling of reverse osmosis membranes used in river water purification for drinking purposes: analysis of microbial populations

Biofouling in water treatment processes represents one of the most frequent causes of plant performance decline. Investigation of clogged membranes (reverse osmosis membranes, microfiltration membranes and ultrafiltration membranes) is generally performed on fresh membranes. In the present study, a multidisciplinary autopsy of a reverse osmosis membrane (ROM) was conducted. The membrane, which was used in sulfate-rich river water purification for drinking purposes, had become inoperative after 6 months because of biofouling and was later stored for 18 months in dry conditions before analysis. SSU rRNA gene library construction, clone sequencing, T-RFLP, light microscope, and scanning electron microscope (SEM) observations were used to identify the microorganisms present on the membrane and possibly responsible for biofouling at the time of removal. The microorganisms were mainly represented by bacteria belonging to the phylum Actinobacteria and by a single protozoan species belonging to the Lobosea group. The microbiological analysis was interpreted in the context of the treatment plant operations to hypothesize as to the possible mechanisms used by microorganisms to enter the plant and colonize the ROM surface.     

Deciphering microbial landscapes of fish eggs to mitigate emerging diseases

Animals and plants are increasingly suffering from diseases caused by fungi and oomycetes. These emerging pathogens are now recognized as a global threat to biodiversity and food security. Among oomycetes, Saprolegnia species cause significant declines in fish and amphibian populations. Fish eggs have an immature adaptive immune system and depend on nonspecific innate defences to ward off pathogens. Here, meta-taxonomic analyses revealed that Atlantic salmon eggs are home to diverse fungal, oomycete and bacterial communities. Although virulent Saprolegnia isolates were found in all salmon egg samples, a low incidence of Saprolegniosis was strongly correlated with a high richness and abundance of specific commensal Actinobacteria, with the genus Frondihabitans (Microbacteriaceae) effectively inhibiting attachment of Saprolegniato salmon eggs. These results highlight that fundamental insights into microbial landscapes of fish eggs may provide new sustainable means to mitigate emerging diseases.     

Actinobacterial diversity from marine sediments collected in Mexico

Seventeen different media known to support the growth and isolation of members of the class Actinobacteria were evaluated as selective isolation media for the recovery of this microbial group from marine sediments samples collected in the Gulf of California and the Gulf of Mexico. A general selective isolation procedure was employed for six sediments and nearly 300 actinomycetes were recovered from the selective isolation plates. Full 16S rRNA gene sequencing revealed that the isolates belonged to several actinobacterial taxa, notably to the genera Actinomadura, Dietzia, Gordonia, Micromonospora, Nonomuraea, Rhodococcus, Saccharomonospora, Saccharopolyspora, Salinispora, Streptomyces, “Solwaraspora” and Verrucosispora. Previous works on marine sediments have been restricted to the isolation of members of the genera Micromonospora, Rhodococcus and Streptomyces. This study provides further evidence that Actinobacteria present in marine habitats are not restricted to the Micromonospora-Rhodococcus-Streptomyces grouping. Indeed, this first systematic study shows the extent of actinobacterial diversity that can be found in marine sediments collected in Mexico and probably, worldwide. The 16S rRNA gene sequences of marine isolates A1, AA2, AA6, AB1, AB2, AG1, AI2, AK1, AL2, AO1, AO3, AR1, AW1, B1, BB1, BC1, C5, R1, R2, R3, AV1, AE1, AI1, AN1 and AP1 determined in this study have been deposited under GenBank accession numbers EU714241–EU714258 and FJ462359–FJ462365, respectively.     

Complete genome sequence of Jonesia denitrificans type strain (Prevot 55134)

Jonesia denitrificans (Prevot 1961) Rocourt et al. 1987 is the type species of the genus Jonesia, and is of phylogenetic interest because of its isolated location in the actinobacterial suborder Micrococcineae. J. denitrificans is characterized by a typical coryneform morphology and is able to form irregular nonsporulating rods showing branched and club-like forms. Coccoid cells occur in older cultures. J. denitrificans is classified as a pathogenic organism for animals (vertebrates). The type strain whose genome is described here was originally isolated from cooked ox blood. Here we describe the features of this organism, together with the complete genome sequence and annotation. This is the first completed genome sequence of a member of the genus for which a complete genome sequence is described. The 2,749,646 bp long genome with its 2558 protein-coding and 71 RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Expression of the <Emphasis Type="Italic">HEV2.1</Emphasis> gene promoter in transgenic <Emphasis Type="Italic">Hevea brasiliensis</Emphasis>

In this article we describe the identification of endophytic bacteria belonging to three groups isolated from shoot tip cultures of banana cv. Grand Naine in a recent study (Thomas et al. 2008) based on partial 16S rRNA gene sequence homology analysis. The first group included banana stocks that displayed obvious colony growth on MS based tissue culture medium during the first in vitro passage. The second group constituted stocks that were tissue index-negative for cultivable bacteria initially but turned index-positive after a few to several (4–8) in vitro passages while the third group formed one sub-stock that turned index-positive after about 18 passages. The organisms belonged to about 20 different genera comprising of α, β, γ-proteobacteria, Gram-positive firmicutes and actinobacteria. Visibly expressing easily cultured organisms during the first in vitro passage included Enterobacter, Klebsiella, Ochrobactrum, Pantoea, Staphylococcus and Bacillus spp. Organisms of second group that were not detected or non-culturable originally constituted Brevundimonas, Methylobacterium, Alcaligenes, Ralstonia, Pseudomonas, Corynebacterium, Microbacterium, Staphylococcus, Oceanobacillus and Bacillus spp. while the third group that turned cultivable after extended in vitro culturing included mostly non-filamentous actinobacteria (Brachybacterium, Brevibacterium, Kocuria and Tetrasphaera spp.). The identification results suggested that the endophytes of second and third groups were not strictly obligate or fastidious microbes but those surviving in viable but-non-culturable (VBNC) state and displaying gradual activation to cultivable form during continuous tissue culturing. Several of the organisms isolated are known as beneficial ones in agriculture while some organisms have possible implications in human health. The use of tissue cultures for isolating uncommon endophytes is discussed. Supply of live bacterial cultures or genetic material for research purpose is subject to their revival from glycerol stocks (as some of the organisms showed poor tolerance) and the requestor obtaining written permission from the Director General, Indian Council of Agricultural Research, New Delhi-110001.     

Physiological adaptation to low temperatures of strains of Rhizobium leguminosarum bv. viciae associated with Lathyrus spp.(1)

Phylogenetic analyses of ribosomal RNA gene sequences (rDNAs) retrieved from an Australian desert soil sample (Sturt National Park) revealed the presence of a number of clones which branched deeply from the high GC Gram-positive division line of descent. The most abundant group of these clones were related to Rubrobacter. An oligonucleotide probe was designed to have broad specificity to Rubrobacter and relatives. This probe was used to interrogate eight rDNA libraries representing four distinct land forms within the Australian arid zone. Relative abundance of Rubrobacter-relatives in these samples ranged from 2.6 to 10.2%. Clones from these libraries were selected for sequence analysis on the basis of a heteroduplex mobility assay to maximise the diversity represented in the sample. Phylogenetic analyses of these rDNA clones and Rubrobacter-related clones reported in the literature show strong support for three distinct groups. Database-searching revealed 'Rubrobacteria' were relatively abundant in a number of published soil rDNA libraries but absent from others. A PCR assay for group-1 'Rubrobacteria' was used to test for their presence in 21 environmental samples. Only marine and arid-zone soil samples gave positive PCR results. Taken together these results indicate 'Rubrobacteria' are a widespread group of variable abundance and diversity.     

Detection of Actinobacteria cultivated from environmental samples reveals bias in universal primers

AIMS: The aims of this study were to develop media to cultivate actinomycetes, screen the resulting isolates with Actinobacteria-specific primers, and examine the efficacy of detection of the actinobacterial isolates with universal primers. METHODS AND RESULTS: Soil-extract medium was developed for a terrestrial bluff environment. Recovered isolates were subjected to polymerase chain reaction (PCR) with taxon-specific primers to identify Actinobacteria. Universal bacterial primers 24f and 1492r (modified and original versions) were used to amplify the 16S rRNA gene from the putative Actinobacteria. While both reverse primers failed to provide amplification products from 20% to 50% of the isolates, the 1492r primer detected Actinobacteria more effectively than 1492r-mod. The region of the gene containing the annealing site for the 1492r primers from 15 isolates that failed to amplify showed no differences in nucleotide sequence to the original 1492r primer. CONCLUSIONS: Universal 16S rRNA gene primers are not capable of amplifying this gene from all bacteria within an environmental sample. Some Actinobacteria may share 100% sequence similarity to universal primers but remain undetected. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings are important for studies of particular taxa in environmental samples where reactions utilizing universal primers may not reveal the extent of their presence and diversity.     

Natronosporangium hydrolyticum gen. nov., sp. nov., a haloalkaliphilic polyhydrolytic actinobacterium from a soda solonchak soil in Central Asia

During a cultural diversity survey on hydrolytic bacteria in saline alkaline soils, a hydrolytic actinobacterium strain ACPA39^T was enriched and isolated in pure culture from a soda solonchak soil in southwestern Siberia. It forms a substrate mycelium with rod-shaped sporangia containing 1–3 exospores. The isolate is obligately alkaliphilic, growing at pH 7.5–10.3 (optimum at 8.5–9.0) and moderately halophilic, tolerating up to 3 M total Na⁺ in the form of sodium carbonates. It is an obligately aerobic, organoheteroterophic, saccharolytic bacterium, utilizing various sugars and alpha/beta-glucans as growth substrates. According to the 16S rRNA gene-based phylogenetic analysis, strain ACPA39^T forms a distinct branch within the family Micromonosporaceae, with the sequence identities below 94.5% with type strains of other genera. This is confirmed by phylogenomic analysis based on the 120 conserved single copy protein-based markers and genomic indexes (ANI, AAI). The cell-wall of ACPA39^T contained meso-DAP, glycine, glutamic acid and alanine in a equimolar ratio, characteristic of the peptidoglycan type A1γ'. The whole-cell sugars include galactose and xylose. The major menaquinone is MK-10(H₄). The identified polar lipids consist of phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylinositol. The polar lipid fatty acids were dominated by anteiso-C_17:0, iso-C_16:0, iso-C_17:0, 10 Me-C_18:0 and C_18:1ω9. Based on the distinct phylogeny, the chemotaxonomy features and unique phenotypic properties, strain ACPA39^T (DSM 106523^T = VKM 2772^T) is classified into a new genus and species in the family Micromonosporaceae for which the name Natronosporangium hydrolitycum gen. nov., sp. nov. is proposed.     

Phylogenetic diversity of sediment bacteria from the southern Cretan margin,Eastern Mediterranean Sea

This study is the first culture-independent report on the regional variability of bacterial diversity in oxic sediments from the unexplored southern Cretan margin (SCM). Three main deep basins (water column depths: 2670–3603 m), located at the mouth of two submarine canyons (Samaria Gorge and Paximades Channel) and an adjacent slope system, as well as two shallow upper-slope stations (water column depths: 215 and 520 m), were sampled. A total of 454 clones were sequenced and the bacterial richness, estimated through five clone libraries using rarefaction analysis, ranged from 71 to 296 unique phylotypes. The average sequence identity of the retrieved Cretan margin sequences compared to the >1,000,000 known rRNA sequences was only 93.5%. A diverse range of prokaryotes was found in the sediments, which were represented by 15 different taxonomic groups at the phylum level. The phylogenetic analysis revealed that these new sequences grouped with the phyla Acidobacteria, Planctomycetes, Actinobacteria, Gamma-, Alpha- and Delta-proteobacteria. Only a few bacterial clones were affiliated with Chloroflexi, Bacteroidetes, Firmicutes, Gemmatimonadetes, Verrucomicrobia, Nitrospirae, Beta-proteobacteria, Lentisphaerae and Dictyoglomi. A large fraction of the retrieved sequences (12%) did not fall into any taxonomic division previously characterized by molecular criteria, whereas four novel division-level lineages, termed candidate division SCMs, were identified. Bacterial community composition demonstrated significant differences in comparison to previous phylogenetic studies. This divergence was mainly triggered by the dominance of Acidobacteria and Actinobacteria and reflected a bacterial community different from that currently known for oxic and pristine marine sediments.