Amplified rDNA Restriction Analysis Research Articles

Association of Pseudomonas syringae pv. apii with the Seed of Ajwain (Trachyspermum ammi L.)

Present study aimed to isolate, identify and characterize the bacterial pathogens associated with ajwain seeds using biochemical and molecular methods. One hundred thirty-nine seed samples of ajwain were collected from various districts of Rajasthan. Twenty-four bacterial isolates were isolated and purified from collected seed samples. All strains were characterised for their biochemical activity. All isolates were tested for genetic diversity by using amplified rDNA restriction analysis (ARDRA). Selective strains were subjected for 16S rDNA sequencing and molecular phylogeny study. Most strains were positive for levan production, tobacco hypersensitivity, gelatine liquefaction and aesculin hydrolysis but showed negative reactions for activities of oxidase, tartrate utilisation, potato soft rot, tyrosinase and arginine dihydrolase. These isolates were distinguished into three groups based on the ARDRA pattern. Selected isolates (KANRJ 1507, KANRJ 1562 and KANRJ 1637) were subjected for 16S rRNA gene sequencing. In phylogenetic analysis all three isolates showed 99.85% similarities to each other. In addition, KANRJ 1507 showed 99.93% sequence similarity to P. syringae pv. apii strain BS426 while KANRJ 1562 and KANRJ 1637 showed 99.85% sequence similarity with P. syringae pv. apii strain BS426 isolated from Petroselinum crispum in California, USA. It was also observed that KANRJ 1507 and other two strains (KANRJ 1562 and 1637) showed 99.61% and 99.54% sequence similarities respectively with P. syringae pv. maculicola isolated from radish in the USA.

Beneficial traits of root endophytes and rhizobacteria associated with plants growing in phytomanaged soils with mixed trace metal-polycyclic aromatic hydrocarbon contamination

The diversity of cultivable bacteria associated with plants from phytomanaged soils with mixed trace metal (TM) and polycyclic aromatic hydrocarbon (PAH) contamination in Pierrelaye (France) was evaluated. The emphasis was on the cultivable bacterial community since the overall objective is to obtain inoculants to improve the remediation of this type of contaminated site. Root endophytic and rhizosphere soil bacterial counts were determined, and isolates were pooled by amplified rDNA restriction analysis and identified by 16S rDNA sequencing. Isolates were further characterized for the production of plant growth-promoting (PGP) substances, and resistance to TM. The selected strains were evaluated for their ability to degrade PAHs. The potential of cell-free microbial supernatant to increase the mobilisation of PAHs from the polluted soil of Pierrelaye was also evaluated. Proteobacteria and Actinobacteria dominated the collection of isolates, and differences in taxonomic diversity were observed between plant species (Populus or Zea mays) and depending on the remediation treatment (Populus inoculation with mycorrhizae or Populus intercropping with Alnus). The majority of isolates exhibited at least one of the tested PGP traits, as well as resistance to more than one TM. Several rhizosphere, endophyte and even one bulk soil isolate showed high rates of fluoranthene and pyrene reduction. The endophyte Rhizobium strain MR28 isolated from maize and degrading pyrene produced bioemulsifying molecules capable of improving the availability of PAHs from the soil of Pierrelaye. A selection of the most interesting strains was made for further re-inoculation experiments in order to assess their potential in rhizoremediation processes.

English

Using phylogenetic analysis of the 16S rRNA gene 43 Gram-positive, spore-forming bacteria of the phylum Firmicutes were isolated, cultured and identified from five hot water springs in South Africa. Thirty-nine isolates belonged to the family Bacillaceae, genus Bacillus (n = 31) and genus Anoxybacillus (n = 8), while four isolates belonged to the family Paenibacillaceae, genus Brevibacillus. The majority of isolates fell into the Bacillus Bergey’s Group A together with Bacillus subtilis and Bacillus licheniformis. One isolate matched Bacillus panaciterrae which has not previously been described as a hot-spring isolate. Three unknown isolates from this study (BLAST <95% match) and three “uncultured Bacillus” clones of isolates from hot springs in India, China and Indonesia listed in NCBI Genbank, were included in the analysis. When bioinformatic tools: Basic Local Alignment Search Tool (BLAST), in silico amplified rDNA restriction analysis (ARDRA), guanine-cytosine (GC) percentage and phylogenetic analysis are used in combination, but not independently, differentiation between the complex Bacillus and closely related species was possible. Identification that relies solely on BLAST of the 16S rRNA sequence can be misleading. Key words: Bacillus, phylogeny, ARDRA, 16S rDNA, South Africa

Screening and characterization of siderophore producing endophytic bacteria from Cicer arietinum and Pisum sativum plants

Siderophores are low molecular weight iron chelating secondary metabolites synthesized by various groups of microorganisms help in scavenging iron-limited conditions. Siderophores produced by endophytic bacteria facilitate the plant growth by providing iron to plants. The objective of this study was to isolate and screen the siderophore producing endophytes from nodules and roots of Cicer arietinum and Pisum sativum plants. Out of total eighty four isolates, only 14 endophytes produced siderophore and quantitative analysis was also done. Ten best siderophore producers (above 65% siderophore units) were characterized for the type of siderophore produced. Most of them were producing hydroxamate and carboxylate type of siderophores. These ten isolates were evaluated for other plant growth promoting (PGP) traits invitro. All of them were producing ammonia and IAA. Isolate CPFR10 was found to be positive for all the PGP traits viz. ammonia, organic acid, HCN and IAA production. Diversity analysis of these ten isolates using ARDRA (Amplified rDNA Restriction Analysis) profile revealed nine haplogroups at 90% similarity.

Extracellular hydrolytic enzymes produced by halophilic bacteria and archaea isolated from hypersaline lake

The screening of bacteria and archaea from Chott El Jerid, a hypersaline lake in the south of Tunisia, led to the isolation of 68 extremely halophilic prokaryotes growing in media with 15-25% of salt. Assessment of 68 partial 16S rRNA analyzed by amplified rDNA restriction analysis (ARDRA) revealed 15 different bacterial and archaeal taxonomic groups. Based on ARDRA results, phenotypic and hydrolytic activity tests, 20 archaeal and 6 bacterial isolates were selected for sequencing. The halophilic isolates were identified as members of the genera: Salicola, Bacillus, Halorubrum, Natrinema and Haloterrigena. Most of these isolates are able to produce hydrolytic enzymes such as amylase, protease, lipase, cellulase, xylanase, pectinase and some of them showed combined activities. Natrinema genus is an excellent candidate for lipase production. These results indicated that the extremely halophilic archaea and bacteria from Chott El Jerid are a potential source of hydrolytic enzymes and may possess commercial value.

Combination of amplified rDNA restriction analysis and high-throughput sequencing revealed the negative effect of colistin sulfate on the diversity of soil microorganisms

Colistin sulfate is widely used in both human and veterinary medicine. However, its effect on the microbial ecologyis unknown. In this study, we determined the effect of colistin sulfate on the diversity of soil microorganisms by amplified rDNA restriction analysis (ARDRA) and high-throughput sequencing.ARDRAshowed that the diversity of DNA from soil microorganisms was reduced after soil was treated with colistin sulfate, with the most dramatic reductionobserved after 35days of treatment. High-throughput sequencing showed that the Chao1 and abundance-based coverage estimators (ACE) were reduced in the soils treated with colistin sulfate for 35 dayscompared to those treated with colistin sulfate for 7days. Furthermore, Chao1 and ACE tended to be lower when higher concentration of colistin sulfate was used, suggesting that the microbial abundance is reduced by colistin sulfate in a dose-dependent manner. Shannon index showed that the diversity of soil microorganism was reduced upon treatment with colistin sulfate compared to the untreated control group. Following 7days of treatment, Bacillus, Clostridiumand Sphingomonas were sensitive to all the concentration of colistin sulfate used in this study. Following 35days of treatment, the abundance of Choroplast, Haliangium, Pseudomonas, Lactococcus, and Clostridium was significantly decreased. Our results demonstrated that colistin sulfate especially at high concentration (≥5mg/kg) could alter the population structure of microorganisms and consequently the microbial community function in soil.

Alterations in soil microbial communities caused by treatments with penicillin or neomycin

Antibiotic residues in soils can lead to serious health risk and ecological hazards. In this study, the effects of penicillin and neomycin, two antibiotics widely used in animal production, were investigated on soil bacterial communities. Changes in the community structure were monitored using three 16S ribosomal DNA (rDNA) polymerase chain reaction-based approaches, including denaturing gradient gel electrophoresis (DGGE), amplified rDNA restriction analysis (ARDRA), and terminal-restriction fragment length polymorphism (T-RFLP) analysis. The prominent DGGE bands were excised from gels and sequenced, and the data indicated the prevalence of Gammaproteobacteria in the soils. The total soil bacterial community, including uncultured bacteria, exhibited a higher diversity than that of cultured bacteria. Some microbial strains were capable of surviving and even subsisting on penicillin or neomycin. We also observed toxic effects of the antibiotics on the indigenous soil bacterial communities since some genotypes disappeared after the treatments (e.g., Pseudomonas sp., Stenotrophomonas sp., Salinimonas, and uncultured Acinetobacter sp.). The implications of these findings are that the functions of soil bacterial communities may be negatively affected if key microbial community members are lost.

Gut microbial diversity in health and disease: experience of healthy Indian subjects, and colon carcinoma and inflammatory bowel disease patients.

ABSTRACT Background: The intestinal microbiota, through complex interactions with the gut mucosa, play a key role in the pathogenesis of colon carcinoma and inflammatory bowel disease (IBD). The disease condition and dietary habits both influence gut microbial diversity. Objective: The aim of this study was to assess the gut microbial profile of healthy subjects and patients with colon carcinoma and IBD. Healthy subjects included ‘Indian vegetarians/lactovegetarians’, who eat plant produce, milk and milk products, and ‘Indian non-vegetarians’, who eat plant produce, milk and milk products, certain meats and fish, and the eggs of certain birds and fish. ‘Indian vegetarians’ are different from ‘vegans’, who do not eat any foods derived wholly or partly from animals, including milk products. Design: Stool samples were collected from healthy Indian vegetarians/lactovegetarians and non-vegetarians, and colon cancer and IBD patients. Clonal libraries of 16S ribosomal DNA (rDNA) of bacteria were created from each sample. Clones were sequenced from one representative sample of each group. Approximately 500 white colonies were picked at random from each sample and 100 colonies were sequenced after amplified rDNA restriction analysis. Results: The dominant phylum from the healthy vegetarian was Firmicutes (34%), followed by Bacteroidetes (15%). The balance was reversed in the healthy non-vegetarian (Bacteroidetes 84%, Firmicutes 4%; ratio 21:1). The colon cancer and IBD patients had higher percentages of Bacteroidetes (55% in both) than Firmicutes (26% and 12%, respectively) but lower Bacteroidetes:Firmicutes ratios (3.8:1 and 2.4:1, respectively) than the healthy non-vegetarian. Bacterial phyla of Verrucomicrobiota and Actinobacteria were detected in 23% and 5% of IBD and colon patients, respectively. Conclusions: Ribosomal Database Project profiling of gut flora in this study population showed remarkable differences, with unique diversity attributed to different diets and disease conditions.

Identification of bacterial agents causing mortality in postlarvae of giant freshwater prawn (Macrobrachium rosenbergii ) in south-west coastal districts of Bangladesh

Following an incidence of freshwater prawn Macrobrachium rosenbergii postlarvae (PL) mortality in hatcheries in summer 2012, samples from dead PL, rearing water and prawn feed from two south-west coastal districts of Bangladesh were collected to isolate, identify and characterize the agents causing PL mortality. Antibiogram profile of sixteen randomly selected bacteria, isolated from dead PL, that grew on TCBS, to 20 different antibiotics belonging to 12 major groups revealed that the drug resistance pattern varied from moderate (56% to the drugs: ciprofloxacin, cefotaxime, nitrofurantoin, kanamycin) to complete (to penicillin, ceftazidime and oxacillin) level. To identify the isolates, amplified rDNA restriction analysis (ARDRA) classified them in to four groups, and RAPD (randomly amplified polymorphic DNA) typing yielded nine different types of isolates within these four ARDRA groups. The 16S rDNA gene sequences identified that the groups were genotypically diverse belonging to the bacterial species: Enterobacter cloacae, Klebsiella pneumoniae, Exiguobacterium profundum and Enterococcus casseliflavus, respectively, that all demonstrated their killing potential to PLs in a simulated environment. The study therefore identified four different bacterial pathogens, one of which, Exiguobacterium profundum is reported for the first here in Bangladesh, that demand special consideration for disease management strategy.

Analysis of Plant Growth-Promoting Effects of Fluorescent Pseudomonas Strains Isolated from Mentha piperita Rhizosphere and Effects of Their Volatile Organic Compounds on Essential Oil Composition.

Many species or strains of the genus Pseudomonas have been characterized as plant growth promoting rhizobacteria (PGPR). We used a combination of phenotypic and genotypic techniques to analyze the community of fluorescent Pseudomonas strains in the rhizosphere of commercially grown Mentha piperita (peppermint). Biochemical techniques, Amplified rDNA Restriction Analysis (ARDRA), and 16S rRNA gene sequence analysis revealed that the majority of the isolated native fluorescent strains were P. putida. Use of two Repetitive Sequence-based PCR (rep-PCR) techniques, BOX-PCR and ERIC-PCR, allowed us to evaluate diversity among the native strains and to more effectively distinguish among them. PGPR activity was tested for the native strains and reference strain P. fluorescens WCS417r. Micropropagated M. piperita plantlets were exposed to microbial volatile organic compounds (mVOCs) emitted by the bacterial strains, and plant biomass parameters and production of essential oils (EOs) were measured. mVOCs from 11 of the native strains caused an increase in shoot fresh weight. mVOCs from three native strains (SJ04, SJ25, SJ48) induced changes in M. pierita EO composition. The mVOCs caused a reduction of metabolites in the monoterpene pathway, for example menthofuran, and an increase in menthol production. Menthol production is the primary indicator of EO quality. The mVOCs produced by native strains SJ04, SJ25, SJ48, and strain WCS417r were analyzed. The obtained mVOC chromatographic profiles were unique for each of the three native strains analyzed, containing varying hydrocarbon, aromatic, and alogenic compounds. The differential effects of the strains were most likely due to the specific mixtures of mVOCs emitted by each strain, suggesting a synergistic effect occurs among the compounds present.

Microbiological investigations on the water of a thermal bath at Budapest.

Thermal baths are unique aquatic environments combining a wide variety of natural and anthropogenic ecological factors, which also appear in their microbiological state. There is limited information on the microbiology of thermal baths in their complexity, tracking community shifts from the thermal wells to the pools. In the present study, the natural microbial community of well and pool waters in Gellért bath was studied in detail by cultivation-based techniques. To isolate bacteria, 10%R2A and minimal synthetic media (with "bath water") with agar-agar and gellan gum were used after prolonged incubation time; moreover, polyurethane blocks covered with media were also applied. Strains were identified by sequencing their 16S rRNA gene after grouping them by amplified rDNA restriction analysis. From each sample, the dominance of Alphaproteobacteria was characteristic though their diversity differed among samples. Members of Actinobacteria, Firmicutes, Beta- and Gamma-proteobacteria, Deinococcus-Thermus, and Bacteroidetes were also identified. Representatives of Deinococcus-Thermus phylum appeared only in the pool water. The largest groups in the pool water belonged to the Tistrella and Chelatococcus genera. The most dominant member in the well water was a new taxon, its similarity to Hartmannibacter diazotrophicus as closest relative was 93.93%.

In vitro antagonistic activity, plant growth promoting traits and phylogenetic affiliation of rhizobacteria associated with wild plants grown in arid soil

The role of plant growth-promoting rhizobacteria (PGPR) in adaptation of plants in extreme environments is not yet completely understood. For this study native bacteria were isolated from rhizospeheric arid soils and evaluated for both growth-promoting abilities and antagonistic potential against phytopathogenic fungi and nematodes. The phylogentic affiliation of these representative isolates was also characterized. Rhizobacteria associated with 11 wild plant species from the arid soil of Almadinah Almunawarah, Kingdom of Saudi Arabia (KSA) were investigated. From a total of 531 isolates, only 66 bacterial isolates were selected based on their ability to inhibit Fusarium oxysporum, and Sclerotinia sclerotiorum. The selected isolates were screened in vitro for activities related to plant nutrition and plant growth regulation as well as for antifungal and nematicidal traits. Isolated bacteria were found to exhibit capabilities in fix atmospheric nitrogen, produce ammonia, indoleacetic acid (IAA), siderophores, solubilize phosphate and zinc, and showed an antagonistic potential against some phytopathogenic fungi and one nematode species (Meloidogyne incognita) to various extent. Isolates were ranked by their potential ability to function as PGPR. The 66 isolates were genotyped using amplified rDNA restriction analysis (ARDRA) and 16S rRNA gene sequence analysis. The taxonomic composition of the representative genotypes from both rhizosphere and rhizoplane comprised Bacillus, Enterobacter and Pseudomonas. Out of the 10 genotypes, three strains designated as PHP03, CCP05, and TAP02 might be regarded as novel strains based on their low similarity percentages and high bootstrap values. The present study clearly identified specific traits in the isolated rhizobacteria, which make them good candidates as PGPR and might contribute to plant adaption to arid environments. Application of such results in agricultural fields may improve and enhance plant growth in arid soils.

광양만 해수의 세균 군집의 계절적 변화

광양만 해수의 세균 군집의 계절적 변화를 배양법과 비배양법을 사용하여 분석하였다. 200개의 분리 균주에 대해 Amplified rDNA restriction 방법을 적용한 경우 분리 균은 Firmicutes, Proteobacteria, Actinobacteria, Bacteroidetes의 4개의 문에 속함을 확인하였다. 비 배양방법으로는 해수로부터 직접 추출한 DNA를 사용하여 pyrosequencing과 변성 농도구배 전기영동(DGGE)을 실시하였다. Pyrosequencing에 의한 16S rRNA 유전자의 염기서열 분석 결과 세균 군집은 춘계와 하계에 각각 24개, 추계에 39개 그리고 동계에 32개의 문으로 구성되었다. 다양도 지수는 추계에 높았으며 춘계에는 우점도 지수가 높았다. Firmicutes 문의 세균이 춘계에 예외적으로 많은 비율을 차지하였으며 나머지 계절에는 Proteobacteria 문의 세균이 우점하였다. 차 우점 분류군은 춘계에는 Proteobacteria 문의 세균인 반면 하계에는 Firmicutes 문, 추계와 동계에는 Bacteroidetes 문의 세균이 차지하였다. 과 수준에서의 우점 분류군은 Bacilliaceae가 춘계에, Rhodobacteraceae와 Bacilliaceae가 하계에, Rhodobacteraceae가 동계에 나타났으나 추계에는 우점 분류군이 없었다. DGGE 에서 확인된 27개의 DNA 절편을 추출하여 계통분석을 실시한 결과 춘계에는 Firmicutes 문에 이어 Proteobacteria 문이 우점하였으며 다른 계절에는 Proteobacteria 문이 우점하였다. 두 가지의 비배양법에 의한 군집 분석 결과 문 수준에서의 세균 군집의 계절적 변화는 유사한 경향이 나타났다. Seasonal variations in the bacterial community of Gwangyang Bay seawater were analyzed using both isolation and cultivation-independent methods. Amplified rDNA restriction analysis was applied to 200 bacterial isolates. Bacterial isolates were composed of four phyla: Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes. Pyrosequencing was conducted, in addition to denaturing gradient gel electrophoresis (DGGE) of genomic DNA extracted directly from the water samples. The bacterial sequences obtained by pyrosequencing of 16S rRNA genes consisted of 24 phyla in the spring and summer, 39 in the fall, and 32 in the winter. The diversity index was high in the fall, whereas the dominancy index was high in the spring. In the spring, phylum Firmicutes was dominant, whereas phylum Proteobacteria dominated in the other three seasons. The second most dominant phyla were Proteobacteria in the spring, Firmicutes in the summer, and Bacteroidetes both in the fall and winter. Bacilliaceae was the most predominant family in the spring. Rhodobacteraceae and Bacilliaceae dominated in the summer, and Rhodobacteraceae dominated in the winter. Neither was dominant in the fall Twenty-seven bands purified from DGGE profiles were cloned and analyzed phylogenetically. In the spring, phylum Firmicutes dominated, followed by Proteobacteria. Proteobacteria dominated in all other seasons. Thus, two cultivation-independent methods for determination of seasonal variation patterns at the phylum level were in accordance with each other.

Isolation and differentiation of methanogenic Archaea from mesophilic corn-fed on-farm biogas plants with special emphasis on the genus Methanobacterium

In this study, methanogenic Archaea were isolated from five full-scale agricultural biogas plants (BGPs) located in Rhineland-Palatinate and Saarland, Germany, digesting maize silage and cattle manure. According to partial 16S rRNA gene sequences, the strains isolated from enrichment cultures were related to Methanoculleus bourgensis, Methanosarcina mazei, Methanosaeta concilii, and Methanobacterium formicicum. The 16S rRNA gene libraries of two representative BGPs screened with the direct amplified rDNA restriction analysis approach also revealed these Archaea to be present. Comparative phylogenetic analyses of reference strains and the isolates of genus Methanobacterium based on 16S and 23S rRNA gene sequences suggest two major groups of isolates, with both of them closely associated with Methanobacterium formicicum strain MF(T). The affiliation of Methanobacterium isolates is further supported by denaturating gradient gel electrophoresis of 16S rRNA gene amplificates, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and specifically amplified polymorphic DNA-PCR (SAPD-PCR), a novel fingerprint approach applied to methanogenic Archaea for the first time. Signature sequence 03Mbf derived from the application of SAPD-PCR was subsequently used to develop a PCR-based primer system for the detection of Methanobacterium formicicum-related isolates and the reference strain in BGP samples. Amplification of 03Mbf fragments down to a minimal titer of 10(3) cells of Methanobacterium formicicum-related isolate Mb9 was possible under BGP fermenter-comparable conditions.

Clavibacter michiganensis subsp. phaseoli subsp. nov., pathogenic in bean

A yellow Gram-reaction-positive bacterium isolated from bean seeds (Phaseolus vulgaris L.) was identified as Clavibacter michiganensis by 16S rRNA gene sequencing. Molecular methods were employed in order to identify the subspecies. Such methods included the amplification of specific sequences by PCR, 16S amplified rDNA restriction analysis (ARDRA), RFLP and multilocus sequence analysis as well as the analysis of biochemical and phenotypic traits including API 50CH and API ZYM results. The results showed that strain LPPA 982T did not represent any known subspecies of C. michiganensis. Pathogenicity tests revealed that the strain is a bean pathogen causing a newly identified bacterial disease that we name bacterial bean leaf yellowing. On the basis of these results, strain LPPA 982T is regarded as representing a novel subspecies for which the name Clavibacter michiganensis subsp. phaseoli subsp. nov. is proposed. The type strain is LPPA 982T (=CECT 8144T=LMG 27667T).

Biochemical Characterization and Molecular Fingerprinting of Plant Growth Promoting Rhizobacteria

Plant growth promoting rhizobacteria (PGPR) are a heterogeneous group of bacteria that can be found in the rhizosphere, which can improve the extent or quality of plant growth directly or indirectly. However, screening strategies for selecting the best rhizobacterial strain for rhizosphere competence with other microbial species in the plant rhizosphere will require more comprehensive knowledge. In present investigation nine different strains were tested for their PGPR properties by using RFLP analysis on 16S rRNA gene or amplified rDNA restriction analysis (ARDRA). 16 S rDNA amplification was done and restriction profiling was done using two endonuclease i.e. msp1 and taq1. Depending upon banding pattern of all the nine strains dendogram was created using NTsys software. A clear cut difference was seen in genetic diversity among the strains. Pseudomonas was found to be the most effective strain among all. Depending upon the outcome we can conclude that ARDRA can be effective tool for analyzing the genetic diversity among different bacteria and PGPR starin e.g. Pseudomonas, Bacillus can be used as a potent biofertilizer.

Diversity of endophytic bacteria in Caragana microphylla grown in the desert grassland of the Ningxia Hui autonomous region of China.

The diversity of endophytic bacteria in the sand-fixation plant Caragana microphylla was investigated by amplified rDNA restriction analysis and by sequence and phylogenetic comparisons of the 16S rRNA genes. A total of 24, 19, and 17 operational taxonomic units were identified from 16S rDNA libraries of the plant roots, stems and leaves, respectively. Homology analysis revealed a 92-100% identity of bacterial 16S rDNA sequences compared with those in the GenBank database. The bacteria identified by sequence homology fell into the following groups: α-Proteobacteria, β-Proteobacteria, γ-Proteobacteria, bacilli, and uncultured bacterium. Sequence analysis demonstrated that the roots were colonized predominantly by Bradyrhizobiaceae, while bacteria from Burkholderiaceae and Sphingomonadaceae were predominant in the stems and leaves, respectively. Additionally, the endophytic bacterial community in leaves was more diverse than those in the roots and stems. Overall, the most abundant bacteria in all three tissues analyzed were from the Sphingomonadaceae and Burkholderiaceae families, although many bacterial populations were found in only a single tissue. These results suggest that the bacterial population of C. microphylla is diverse.

Diversity, biocontrol, and plant growth promoting abilities of xylem residing bacteria from solanaceous crops.

Eggplant (Solanum melongena L.) is one of the solanaceous crops of economic and cultural importance and is widely cultivated in the state of Goa, India. Eggplant cultivation is severely affected by bacterial wilt caused by Ralstonia solanacearum that colonizes the xylem tissue. In this study, 167 bacteria were isolated from the xylem of healthy eggplant, chilli, and Solanum torvum Sw. by vacuum infiltration and maceration. Amplified rDNA restriction analysis (ARDRA) grouped these xylem residing bacteria (XRB) into 38 haplotypes. Twenty-eight strains inhibited growth of R. solanacearum and produced volatile and diffusible antagonistic compounds and plant growth promoting substances in vitro. Antagonistic strains XB86, XB169, XB177, and XB200 recorded a biocontrol efficacy greater than 85% against BW and exhibited 12%–22 % increase in shoot length in eggplant in the greenhouse screening. 16S rRNA based identification revealed the presence of 23 different bacterial genera. XRB with high biocontrol and plant growth promoting activities were identified as strains of Staphylococcus sp., Bacillus sp., Streptomyces sp., Enterobacter sp., and Agrobacterium sp. This study is the first report on identity of bacteria from the xylem of solanaceous crops having traits useful in cultivation of eggplant.

Multifarious plant growth promoting characteristics of chickpea rhizosphere associated Bacilli help to suppress soil-borne pathogens

Wilt and root rot are the major constraints in chickpea production and very difficult to manage through agrochemicals. Hence, for an ecofriendly and biological management, 240 strains of Bacillus and Bacillus derived genera were isolated from chickpea rhizosphere, further narrowed down to 14 strains on the basis of in vitro production of indole acetic acid, siderophore, phosphate solubilization, hydrolytic enzymes and were evaluated for antagonism against chickpea pathogens (Fusarium oxysporum f. sp. ciceri race 1, F. solani and Macrophomina phaseolina). The strains were identified on the basis of physiological characters and 16S RNA gene sequencing. The genotypic comparisons of strains were determined by BOX-polymerase chain reaction profiles and amplified rDNA restriction analysis. These isolates were evaluated in greenhouse assay in which B. subtilis (B-CM191, B-CV235, B-CL-122) proved to be effective in reducing wilt incidence and significant enhancement in growth (root and shoot length) and dry matter of chickpea plants. PCR amplification of bacillomycin (bmyB) and β-glucanase genes suggests that amplified genes from the Bacillus could have a role to further define the diversity, ecology, and biocontrol activities in the suppression of soil-borne pathogens.

소아의 치아 우식 부위별 세균 다양성

Objectives : Molecular biology techniques were employed to assess diversity of bacterial in children`s dental caries. Methods : DNA of germs was extracted and the diversity of the 16S rRNA clones was analyzed by amplified rDNA restriction analysis and sequencing. The experimental samples were pit and fissure caries (PC), deep dentinal caries (DC), smooth surface caries (SC), and supragingival plaque (PQ) from 50 children of age less than 12 years old. The control group was healthy teeth supragingival plaque (HT). Thirty clones from each 16S rRNA clone library of 5 samples were randomly selected, thus a total of 150 clones were analyzed. Results : Amplified rDNA restriction analysis uncovered 18, 20, 11, 17, and 22 phylotypes from healthy teeth, pit and fissure caries, deep dentinal caries, smooth surface caries, and supragingival plaque, respectively. Sequencing analysis found the dominance of Actinomycs naeslundii and Fusobacterium nucleatum in the healthy teeth; Leptotrichia sp. in the pit and fissure caries; Actinomyces sp., Streptococcus mutans, and Rahnella aquatilis in the deep dentinal caries; Streptococcus mutans and Actinomyces sp. in the smooth surface caries; Enterobacter hormaechei and Streptococcus sanguinis in the supragingival plaque. Conclusions : Clonal analysis identified 6 phyla, 20 genera, and 51 species.