Diversity and taxonomic revision of methanogens and other archaea in the intestinal tract of terrestrial arthropods.

Methane emission by terrestrial invertebrates is restricted to millipedes, termites, cockroaches, and scarab beetles. The arthropod-associated archaea known to date belong to the orders Methanobacteriales, Methanomassiliicoccales, Methanomicrobiales, and Methanosarcinales, and in a few cases also to non-methanogenic Nitrososphaerales and Bathyarchaeales. However, all major host groups are severely undersampled, and the taxonomy of existing lineages is not well developed. Full-length 16S rRNA gene sequences and genomes of arthropod-associated archaea are scarce, reference databases lack resolution, and the names of many taxa are either not validly published or under-classified and require revision. Here, we investigated the diversity of archaea in a wide range of methane-emitting arthropods, combining phylogenomic analysis of isolates and metagenome-assembled genomes (MAGs) with amplicon sequencing of full-length 16S rRNA genes. Our results allowed us to describe numerous new species in hitherto undescribed taxa among the orders Methanobacteriales (Methanacia, Methanarmilla, Methanobaculum, Methanobinarius, Methanocatella, Methanoflexus, Methanorudis, and Methanovirga, all gen. nova), Methanomicrobiales (Methanofilum and Methanorbis, both gen. nova), Methanosarcinales (Methanofrustulum and Methanolapillus, both gen. nova), Methanomassiliicoccales (Methanomethylophilaceae fam. nov., Methanarcanum, Methanogranum, Methanomethylophilus, Methanomicula, Methanoplasma, Methanoprimaticola, all gen. nova), and the new family Bathycorpusculaceae (Bathycorpusculum gen. nov.). Reclassification of amplicon libraries from this and previous studies using this new taxonomic framework revealed that arthropods harbor only CO2 and methyl-reducing hydrogenotrophic methanogens. Numerous genus-level lineages appear to be present exclusively in arthropods, suggesting long evolutionary trajectories with their termite, cockroach, and millipede hosts, and a radiation into various microhabitats and ecological niches provided by their digestive tracts (e.g., hindgut compartments, gut wall, or anaerobic protists). The distribution patterns among the different host groups are often complex, indicating a mixed mode of transmission and a parallel evolution of invertebrate and vertebrate-associated lineages.

Evaluation of prokaryotic diversity of five hot springs in Eritrea.

BACKGROUND: Total community rDNA was used to determine the diversity of bacteria and archaea from water, wet sediment and microbial mats samples of hot springs in the Eastern lowlands of Eritrea. The temperatures of the springs range from 49.5 °C to 100 °C while pH levels varied from 6.97 to 7.54. Akwar and Maiwooi have high carbonate levels. The springs near the seashore, Garbanabra and Gelti, are more saline with higher levels of sodium and chlorides. Elegedi, situated in the Alid volcanic area, has the highest temperature, iron and sulfate concentrations. RESULTS: The five hot springs shared 901 of 4371 OTUs recovered while the three sample types (water, wet sediment and microbial mats) also shared 1429 OTUs. The Chao1 OTU estimate in water sample was significantly higher than the wet sediment and microbial mat samples. As indicated by NMDS, the community samples at genus level showed location specific clustering. Certain genera correlated with temperature, sodium, carbonate, iron, sulfate and ammonium levels in water. The abundant phyla included Proteobacteria (6.2-82.3%), Firmicutes (1.6-63.5%), Deinococcus-Thermus (0.0-19.2%), Planctomycetes (0.0-11.8%), Aquificae (0.0-9.9%), Chlorobi (0.0-22.3%) and Bacteroidetes (2.7-8.4%). CONCLUSION: There were significant differences in microbial community structure within the five locations and sample types at OTU level. The occurence of Aquificae, Deinococcus-Thermus, some Cyanobacteria and Crenarchaeota were highly dependent on temperature. The Halobacterium, unclassified Thaumarchaeota, Actinobacteria and Cyanobacteria showed significant correlation with salinity occurring abundantly in Garbanabra and Gelti. Firmicutes and unclassified Rhodocylaceae were higher in the microbial mat samples, while Archaea were prominent in the wet sediment samples.

454 Pyrosequencing-based assessment of bacterial diversity and community structure in termite guts, mounds and surrounding soils.

Termites constitute part of diverse and economically important termite fauna in Africa, but information on gut microbiota and their associated soil microbiome is still inadequate. In this study, we assessed and compared the bacterial diversity and community structure between termites' gut, their mounds and surrounding soil using the 454 pyrosequencing-based analysis of 16S rRNA gene sequences. A wood-feeder termite (Microcerotermes sp.), three fungus-cultivating termites (Macrotermes michaelseni, Odontotermes sp. and Microtermes sp.), their associated mounds and corresponding savannah soil samples were analyzed. The pH of the gut homogenates and soil physico-chemical properties were determined. The results indicated significant difference in bacterial community composition and structure between the gut and corresponding soil samples. Soil samples (Chao1 index ranged from 1359 to 2619) had higher species richness than gut samples (Chao1 index ranged from 461 to 1527). The bacterial composition and community structure in the gut of Macrotermes michaelseni and Odontotermes sp. were almost identical but different from that of Microtermes and Microcerotermes species, which had unique community structures. The most predominant bacterial phyla in the gut were Bacteroidetes (40-58 %), Spirochaetes (10-70 %), Firmicutes (17-27 %) and Fibrobacteres (13 %) while in the soil samples were Acidobacteria (28-45 %), Actinobacteria (20-40 %) and Proteobacteria (18-24 %). Some termite gut-specific bacterial lineages belonging to the genera Dysgonomonas, Parabacteroides, Paludibacter, Tannerella, Alistipes, BCf9-17 termite group and Termite Treponema cluster were observed. The results not only demonstrated a high level of bacterial diversity in the gut and surrounding soil environments, but also presence of distinct bacterial communities that are yet to be cultivated. Therefore, combined efforts using both culture and culture-independent methods are suggested to comprehensively characterize the bacterial species and their specific roles in these environments.

Phenotypic and genetic characterization of Vibrio cholerae O1 isolated from various regions of Kenya between 2007 and 2010.

INTRODUCTION: Cholera, a disease caused by Vibrio cholerae O1 and O139 remains an important public health problem globally. In the last decade, Kenya has experienced a steady increase of cholera cases. In 2009 alone, 11,769 cases were reported to the Ministry of Public Health and Sanitation. This study sought to describe the phenotypic characteristics of the isolated V. cholerae isolates. METHODS: This was a laboratory based cross-sectional study that involved isolates from different cholera outbreaks. Seventy six Vibrio cholerae O1 strains from different geographical areas were used to represent 2007 to 2010 cholera epidemics in Kenya, and were characterized by serotyping, biotyping, polymerase chain r(PCR), pulsed-field gel electrophoresis (PFGE) and ribotyping along with antimicrobial susceptibility testing. RESULTS: Seventy six Vibrio cholerae O1 strains from different geographical areas were used to represent 2007 to 2010 cholera epidemics in Kenya. Serotype Inaba was dominant (88.2%) compared to Ogawa. The isolates showed varying levels of antibiotic resistance ranging from 100% susceptible to tetracycline, doxycycline, ofloxacin, azithromycin, norfloxacin and ceftriaxone to 100% resistant to furazolidone, trimethoprim-sulfamethoxazole, polymyxin-B and streptomycin. The isolates were positive for ctxA, tcpA (El Tor), rtxC genes and were biotype El Tor variant harboring classical ctxB gene. All the isolates were classified as cholera toxin (CT) genotype 1 as they had mutation in the ctxB at positions 39 and 68. All the isolates had genetically similar NotI PFGE and BglI ribotype patterns. The absence of any observed variation is consistent with a clonal origin for all of the isolates. CONCLUSION: Kenya experienced cholera numerous outbreak from 2007-2010. The clinical Vibrio cholerae O1 isolates from the recent cholera epidemic were serotypes Inaba and Ogawa, Inaba being the predominant serotype. The Vibrio cholerae O1 strains were biotype El Tor variants that produce cholera toxin B (ctx B) of the classical type and were positive for ctxA, tcpA El Tor and rtxC genes.

16S-rRNA-based analysis of bacterial diversity in the gut of fungus-cultivating termites (Microtermes and Odontotermes species).

The interaction between termites and their gut symbionts has continued to attract the curiosity of researchers over time. The aim of this study was to characterize and compare the bacterial diversity and community structure in the guts of three termites (Odontotermes somaliensis, Odontotermes sp. and Microtermes sp.) using 16S rRNA gene sequencing of clone libraries. Clone libraries were screened by restriction fragment length polymorphism and representative clones from O. somaliensis (100 out of 330 clones), Odontotermes sp. (100 out of 359 clones) and Microtermes sp. (96 out 336 clones) were sequenced. Phylogenetic analysis indicated seven bacterial phyla were represented: Bacteroidetes, Spirochaetes, Firmicutes, Proteobacteria, Synergistetes, Planctomycetes and Actinobacteria. Sequences representing the phylum Bacteroidetes (>60 %) were the most abundant group in Odontotermes while those of Spirochaetes (29 %) and Firmicutes (23 %) were the abundant groups in Microtermes. The gut bacterial community structure within the two Odontotermes species investigated here was almost identical at the phylum level, but the Microtermes sp. had a unique bacterial community structure. Bacterial diversity was higher in Odontotermes than in Microtermes. The affiliation and clustering of the sequences, often with those from other termites' guts, indicate a majority of the gut bacteria are autochthonous having mutualistic relationships with their hosts. The findings underscore the presence of termite-specific bacterial lineages, the majority of which are still uncultured.

Diversity of Termitomyces associated with fungus-farming termites assessed by cultural and culture-independent methods.

BACKGROUND: Fungus-cultivating termites make use of an obligate mutualism with fungi from the genus Termitomyces, which are acquired through either vertical transmission via reproductive alates or horizontally transmitted during the formation of new mounds. Termitomyces taxonomy, and thus estimating diversity and host specificity of these fungi, is challenging because fruiting bodies are rarely found. Molecular techniques can be applied but need not necessarily yield the same outcome than morphological identification. METHODOLOGY: Culture-dependent and culture-independent methods were used to comprehensively assess host specificity and gut fungal diversity. Termites were identified using mitochondrial cytochrome oxidase II (COII) genes. Twenty-three Termitomyces cultures were isolated from fungal combs. Internal transcribed spacer (ITS) clone libraries were constructed from termite guts. Presence of Termitomyces was confirmed using specific and universal primers. Termitomyces species boundaries were estimated by cross-comparison of macromorphological and sequence features, and ITS clustering parameters accordingly optimized. The overall trends in coverage of Termitomyces diversity and host associations were estimated using Genbank data. RESULTS AND CONCLUSION: Results indicate a monoculture of Termitomyces in the guts as well as the isolation sources (fungal combs). However, cases of more than one Termitomyces strains per mound were observed since mounds can contain different termite colonies. The newly found cultures, as well as the clustering analysis of GenBank data indicate that there are on average between one and two host genera per Termitomyces species. Saturation does not appear to have been reached, neither for the total number of known Termitomyces species nor for the number of Termitomyces species per host taxon, nor for the number of known hosts per Termitomyces species. Considering the rarity of Termitomyces fruiting bodies, it is suggested to base the future taxonomy of the group mainly on well-characterized and publicly accessible cultures.

Effects of carbon amendment on in situ atrazine degradation and total microbial biomass.

This study elucidates the effects of carbon amendment on metabolic degradation of atrazine (6-chloro-N(2)-ethyl-N(4)-isopropyl-1,3,5-triazine-2,4-diamine) and total microbial biomass in soil. Degradation of (14)C-ring-labelled atrazine was monitored in laboratory incubations of soils supplemented with 0, 10, 100 and 1000 µg g(-1) sucrose concentrations. An experiment to determine the effect of carbon amendment on total microbial biomass and soil respiration was carried out with different concentrations of sucrose and non-labelled atrazine. The soils were incubated at a constant temperature and constant soil moisture at water potential of -15 kPa and a soil density of 1.3 g cm(-3). Mineralization of (14)C-ring-labelled atrazine was monitored continuously over a period of 59 d in the first experiment. The CO(2) production was monitored for 62 d in the second experiment and microbial biomass determined at the end of the incubation period. The addition of 1000 µg g(-1) sucrose reduced atrazine mineralization to 43.5% compared to 51.7% of the applied amount for the treatment without sucrose. The addition of 1000 µg g(-1) sucrose modified the transformation products to 1.08 µg g(-1) deisopropylatrazine (DIA), 0.32 µg g(-1) desethylatrazine (DEA) and 0.18 µg g(-1) deisopropyl-2-hydroxyatrazine (OH-DIA). Treatment without sucrose resulted in formation of 0.64 µg g(-1) hydroxyatrazine (HA), 0.28 µg g(-1) DIA and 0.20 µg g(-1) OH-DIA. Atrazine dealkylation was enhanced in treatments with 100 and 1000 µg g(-1) of sucrose added. HA metabolite was formed in the control (no sucrose) and in the presence of 10 µg g(-1) of sucrose, whereas DEA was only detected in treatment with 1000 µg g(-1) sucrose. Results indicate that total microbial biomass increased significantly (P < 0.001) with the addition of 1000 µg g(-1) sucrose.

Molecular epidemiology of geographically dispersed Vibrio cholerae, Kenya, January 2009-May 2010.

Numerous outbreaks of cholera have occurred in Kenya since 1971. To more fully understand the epidemiology of cholera in Kenya, we analyzed the genetic relationships among 170 Vibrio cholerae O1 isolates at 5 loci containing variable tandem repeats. The isolates were collected during January 2009-May 2010 from various geographic areas throughout the country. The isolates grouped genetically into 5 clonal complexes, each comprising a series of genotypes that differed by an allelic change at a single locus. No obvious correlation between the geographic locations of the isolates and their genotypes was observed. Nevertheless, geographic differentiation of the clonal complexes occurred. Our analyses showed that multiple genetic lineages of V. cholerae were simultaneously infecting persons in Kenya. This finding is consistent with the simultaneous emergence of multiple distinct genetic lineages of V. cholerae from endemic environmental reservoirs rather than recent introduction and spread by travelers.

Biodegradation of s-triazine herbicide atrazine by Enterobacter cloacae and Burkholderia cepacia sp. from long-term treated sugarcane-cultivated soils in Kenya.

In this study soils from sugarcane-cultivated fields were screened for bacterial species capable of atrazine (6-chloro-N²-ethyl-N4-isopropyl-1,3,5-triazine-2,4-diamine) degradation due to long exposure of the soils to this herbicide. To enrich for atrazine degraders, Minimal Salt Medium containing atrazine as the sole N source and glucose as the C source was inoculated with soils impacted with this herbicide and incubated. Bacterial growth was monitored by measuring optical density. The degradation of atrazine was followed by measuring residual atrazine in liquid cultures over a given time period by high performance liquid chromatography. Bacterial strains isolated from the enrichment cultures were characterized by biochemical tests and identified by 16S rRNA gene sequencing. Two bacterial strains coded ISL 8 and ISL 15 isolated from two different fields were shown to have 94 and 96% 16S rRNA gene sequence similarity to Burkholderia cepacia respectively. Another bacterial sp., ISL 14 was closely related to Enterobacter cloacae with a 96% 16S rRNA gene sequence similarity. There was not much difference between the extents of atrazine degradation by the enrichment cultures with communities (79-82% applied amount) from which pure strains were isolated and the pure strains themselves in liquid cultures that showed a degradation of 53-83% of applied amount. The study showed existence of bacterial strains in different sugarcane-cultivated fields which can use atrazine as a nitrogen source. The bacterial strains isolated can be used to enhance the degradation of atrazine in contaminated soils where atrazine is still considered to be recalcitrant.

Prevalence and genetic characteristics of Shigatoxigenic Escherichia coli from patients with diarrhoea in Maasailand, Kenya.

INTRODUCTION: Shigatoxigenic Escherichia coli strains are food-borne bacterial pathogens that may cause haemorrhagic colitis (HC) in humans which can lead to life-threatening systemic complication, including haemolytic uremic syndrome (HUS). This study aimed to characterize and analyze virulence properties of pathogenic E. coli isolates among patients with diarrhoea from a Maasai community in Kenya. METHODOLOGY: Stool samples from 380 patients of all ages from the Kajiado and Narok districts of Kenya were investigated for the presence of enteric bacterial pathogens by conventional and molecular methods. RESULTS: Bacterial diarrhoea was diagnosed in 141/380 (37.1%) cases, of which enterotoxigenic E. coli (ETEC) compromised 29.8%, shigatoxigenic E. coli (STEC) 24.1%, enteroaggregative E. coli (EAEC) 14.2%, enteroinvasive E. coli (EIEC) 12.8% and enteropathogenic E. coli (EPEC) 3.5%. Gene analysis for STEC virulence factors showed that 52.9% isolates carried stx1, 29.4% possessed stx2, 14.7% carried both stx1 and stx2, and 2.9% had stx2e. 23.5% isolates carried enterohaemolysin and 20.5% isolates possessed the Intimin gene. From 9 strains that exhibited adherence, 7 contained both Intimin and Haemolysin genes. Infections with Intimin-positive STEC strains (46%) were more frequent in patients with bloody diarrhoea, especially in children under 5 years of age, whereas Intimin-negative STEC infections dominated in adults. CONCLUSION: Although STEC infection as a cause of bloody diarrhoea has not attracted much attention as a medical problem in Kenya, our findings indicate that this is a problem that must be investigated. The 24.1% isolation rate of STEC among the Maasai is one of the highest reported rates worldwide.

Archaeal diversity in the haloalkaline Lake Elmenteita in Kenya.

A non-culture approach was used to study the archaeal diversity in Lake Elmenteita, Kenya. Five different sampling points were selected randomly within the lake. Wet sediments and water samples were collected from each sampling point. In addition, dry mud cake was collected from three points where the lake had dried. DNA was extracted from these samples and the 16S rRNA genes were amplified using primers described to be Domain-specific for Archaea. Eleven clone libraries were constructed using PCR-amplified 16S rRNA genes. A total of 1,399 clones were picked and analysed via ARDRA. 170 ARDRA patterns were unique and the respective clones were selected for sequencing. 149 clones gave analysable sequences. BLAST analysis showed that 49 belong to the Domain Archaea while the others were either chimera or affiliated to eukaryotic taxa. Comparative sequence analysis of archaeal clones affiliated them to a wide range of genera. The order Halobacteriales was represented by members of the genera Natronococcus, Halovivax, Halobiforma, Halorubrum, and Halalkalicoccus. The highest percentage (46%) of the clones, however, belonged to uncultured members of the Domain Archaea in the order Halobacteriales. The results show that the archaeal diversity in the lake could be higher than previously reported.

Effect of temperature on virulence of Beauveria bassiana and Metarhizium anisopliae isolates to Tetranychus evansi.

The virulence of three isolates of Beauveria bassiana (Bals.) Vuill. and 23 isolates of Metarhizium anisopliae (Metschnik.) Sorok. (Ascomycota: Hypocreales) against the tomato spider mite, Tetranychus evansi Baker and Pritchard (Acari: Tetranychidae), was assessed in the laboratory. The effect of temperature on germination, radial growth and virulence of selected isolates (two isolates of B. bassiana and nine of M. anisopliae) on T. evansi was also investigated in the laboratory. All the fungal isolates tested were pathogenic to the adult females of T. evansi, and there were significant differences in mortality between fungal isolates. The lethal time to 50% mortality (LT(50)) values ranged from 4.2 to 8.1 days and the LT(90) values from 5.6 to 15.1 days. Temperature had significant effects on germination, radial growth and virulence of the various isolates. The best fungal germination was observed at 25 and 30 degrees C, while for the fungal radial growth it was 30 degrees C. All the isolates germinated and grew at all temperatures, but germination and radial growth varied with isolate and temperature. The selected isolates were all virulent to T. evansi, but virulence varied also with isolate and temperature.

High incidence of enteroaggregative Escherichia coli among food handlers in three areas of Kenya: a possible transmission route of travelers' diarrhea.

BACKGROUND: Contaminated food and water are acknowledged vehicles for the transmission of travelers' diarrhea (TD). Importance of food handlers as reservoirs of enteroaggregative Escherichia coli (EAEC), enteropathogenic E coli (EPEC), and Shiga toxin-producing E coli (STEC) causing TD has not been clearly demonstrated. METHODS: We undertook a 1-year prospective study to determine the presence and selected risk factors of carriage of EAEC, EPEC, and STEC by 1,399 food handlers working in tourist hotels in three popular tourist destinations of Kenya. Enterotoxigenic E coli (ETEC) was not sought in this study. RESULTS: During the period April 2003 to May 2004, EAEC harboring the aggR gene were detected from 29 (2.1%) subjects and EPEC harboring the eaeA gene and STEC harboring the stx2 gene were detected from 11 (0.8%) and 2 (0.1%) of the study subjects, respectively. Mean age of subjects with EAEC was significantly lower (24.6 y) than the rest of the study population (28.2 y) (p < 0.05). Pit latrines usage was significantly associated with the isolation of EAEC (<0.001) but not with EPEC and STEC. Four of the 29 EAEC isolates were sensitive to all antibiotics tested, and 19 (65.5%) were multidrug resistant (MDR). Antibiotic resistance varied from 6.9% for cefuroxime to 72.4% for co-trimoxazole. Six EPEC isolates (6/13, 46.2%) showed multidrug resistance. Cluster analysis of the pulsed-field gel electrophoresis (PFGE) profiles showed that the EAEC isolates belonged to two clonally unrelated genotypes. CONCLUSIONS: We conclude that food handlers working in tourist hotels are important carriers of EAEC that could cause TD and a high proportion of the EAEC are MDR. The isolation of MDR EAEC from food handlers working in tourist hotels is of potential public health importance. There is a need for a study employing molecular methods including PFGE to examine carriage of similar pathogens in food handlers, processed foods, and travelers consuming the food who develop diarrhea.

Sporotalea propionica gen. nov. sp. nov., a hydrogen-oxidizing, oxygen-reducing, propionigenic firmicute from the intestinal tract of a soil-feeding termite.

An unusual propionigenic bacterium was isolated from the intestinal tract of the soil-feeding termite Thoracotermes macrothorax. Strain TmPN3 is a motile, long rod that stains gram-positive, but reacts gram-negative in the KOH test. It forms terminal endospores and ferments lactate, glucose, lactose, fructose, and pyruvate to propionate and acetate via the methyl-malonyl-CoA pathway. Propionate and acetate are formed at a ratio of 2:1, typical of most propionigenic bacteria. Under a H(2)/CO(2) atmosphere, the fermentation product pattern of glucose, fructose, and pyruvate shifts towards propionate formation at the expense of acetate. Cell suspensions reduce oxygen with lactate, glucose, glycerol, or hydrogen as electron donor. In the presence of oxygen, the product pattern of lactate fermentation shifts from propionate to acetate production. 16S rRNA gene sequence analysis showed that strain TmPN3 is a firmicute that clusters among the Acidaminococcaceae, a subgroup of the Clostridiales comprising obligately anaerobic, often endospore-forming bacteria that possess an outer membrane. Based on phenotypic differences and less than 92% sequence similarity to the 16S rRNA gene sequence of its closest relative, the termite hindgut isolate Acetonema longum, strain TmPN3(T) is proposed as the type species of a new genus, Sporotalea propionica gen. nov. sp. nov. (DSM 13327(T), ATCC BAA-626(T)).

Pathogenicity of Beauveria bassiana and Metarhizium anisopliae to the tobacco spider mite Tetranychus evansi.

Seventeen isolates of Metarhizium anisopliae (Metschnikoff) Sorokin and two isolates of Beauveria bassiana (Balsamo) Vuillemin were evaluated for their pathogenicity against the tobacco spider mite, Tetranychus evansi Baker & Pritchard. In the laboratory all the fungal isolates were pathogenic to the adult female mites, causing mortality between 22.1 and 82.6%. Isolates causing more than 70% mortality were subjected to dose-response mortality bioassays. The lethal concentration causing 50% mortality (LC5o) values ranged between 0.7 x 10(7) and 2.5 x 10(7) conidia ml(-1). The lethal time to 50% mortality (LT50) values of the most active isolates of B. bassiana and M. anisopliae strains varied between 4.6 and 5.8 days. Potted tomato plants were artificially infested with T. evansi and treated with B. bassiana isolate GPK and M. anisopliae isolate ICIPE78. Both fungal isolates reduced the population density of mites as compared to untreated controls. However, conidia formulated in oil outperformed the ones formulated in water. This study demonstrates the prospects of pathogenic fungi for the management of T. evansi.

Sporomusa aerivorans sp. nov., an oxygen-reducing homoacetogenic bacterium from the gut of a soil-feeding termite.

Previously undescribed, homoacetogenic bacteria were isolated from gut homogenates of the soil-feeding termite Thoracotermes macrothorax. The isolates were slightly curved, banana-shaped rods (0.6-0.7x1.3-7.0 micro m) and were motile by one or more lateral flagella. In older cultures, cells formed club-like sporangia that developed into terminal, heat-resistant endospores. Cells stained Gram-positive but were Gram-negative in the KOH test. The isolates were mesophilic and grew homoacetogenically on H(2)/CO(2) and L-lactate. Strain TmAO3(T), which was characterized further, also grew homoacetogenically on pyruvate, citrate, L-alanine, D-mannitol, ethanol, formate and methanol. Succinate was decarboxylated to propionate; fumarate, L-malate and oxaloacetate were fermented to propionate and acetate. Hexoses were not used as substrates. Resting cells had a large capacity for hydrogen-dependent oxygen reduction [826 nmol min(-1) (mg protein)(-1)], which enabled them to initiate growth in non-reduced basal medium that originally contained up to 1.5 kPa oxygen in the headspace, although growth commenced only after the medium had been rendered anoxic. Redox difference spectra of cell extracts indicated the presence of membrane-bound b-type cytochrome(s). Comparative 16S rRNA gene sequence analysis revealed that strain TmAO3(T) belongs to a subgroup of the phylum of Gram-positive bacteria that is characterized by low DNA G+C content and a Gram-negative cell wall. It is related most closely to representatives of the genus SPOROMUSA: Based on morphological and physiological properties and on 16S rRNA gene sequence similarity of 94-97 % to other Sporomusa species, the isolates are assigned to Sporomusa aerivorans sp. nov. (type strain, TmAO3(T)=DSM 13326(T)=ATCC BAA-625(T)).

Hydrogen-dependent oxygen reduction by homoacetogenic bacteria isolated from termite guts.

Although homoacetogenic bacteria are generally considered to be obligate anaerobes, they colonize the intestinal tracts of termites and other environments that are not entirely anoxic in space or time. In this study, we investigated how homoacetogenic bacteria isolated from the hindguts of various termites respond to the presence of molecular oxygen. All strains investigated formed growth bands in oxygen gradient agar tubes under a headspace of H(2)-CO(2). The position of the bands coincided with the oxic-anoxic interface and depended on the O(2) partial pressure in the headspace; the position of the bands relative to the meniscus remained stable for more than 1 month. Experiments with dense cell suspensions, performed with Clark-type O(2) and H(2) electrodes, revealed a large capacity for H(2)-dependent oxygen reduction in Sporomusa termitida and Sporomusa sp. strain TmAO3 (149 and 826 nmol min(-1) mg of protein(-1), respectively). Both strains also reduced O(2) with endogenous reductants, albeit at lower rates. Only in Acetonema longum did the basal rates exceed the H(2)-dependent rates considerably (181 versus 28 nmol min(-1) mg of protein)(-1)). Addition of organic substrates did not stimulate O(2) consumption in any of the strains. Nevertheless, reductive acetogenesis by cell suspensions of strain TmAO3 was inhibited even at the lowest O(2) fluxes, and growth in nonreduced medium occurred only after the bacteria had rendered the medium anoxic. Similar results were obtained with Acetobacterium woodii, suggesting that the results are not unique to the strains isolated from termites. We concluded that because of their tolerance to temporary exposure to O(2) at low partial pressures (up to 1.5 kPa in the case of strain TmAO3) and because of their large capacity for O(2) reduction, homoacetogens can reestablish conditions favorable for growth by actively removing oxygen from their environment.