Study on diversity of endophytic bacterial communities in seeds of hybrid maize and their parental lines

The seeds of plants are carriers of a variety of beneficial bacteria and pathogens. Using the non-culture methods of building 16S rDNA libraries, we investigated the endophytic bacterial communities of seeds of four hybrid maize offspring and their respective parents. The results of this study show that the hybrid offspring Yuyu 23, Zhengdan958, Jingdan 28 and Jingyu 11 had 3, 33, 38 and 2 OTUs of bacteria, respectively. The parents Ye 478, Chang 7-2, Zheng 58, Jing 24 and Jing 89 had 12, 36, 6, 12 and 2 OTUs, respectively. In the hybrid Yuyu 23, the dominant bacterium Pantoea (73.38?%) was detected in its female parent Ye 478, and the second dominant bacterium of Sphingomonas (26.62?%) was detected in both its female (Ye 478) and male (Chang 7-2) parent. In the hybrid Zhengdan 958, the first dominant bacterium Stenotrophomonas (41.67?%) was detected in both the female (Zheng 58) and male (Chang 7-2) parent. The second dominant bacterium Acinetobacter (9.26?%) was also the second dominant bacterium of its male parent. In the hybrid Jingdan 28, the second dominant bacterium Pseudomonas (12.78?%) was also the second dominant bacterium of its female parent, and its third dominant bacterium Sphingomonas (9.90?%) was the second dominant bacterium of its male parent and detected in its female parent. In the hybrid Jingyu 11, the first dominant bacterium Leclercia (73.85?%) was the third dominant bacterium of its male parent, and the second dominant bacterium Enterobacter (26.15?%) was detected in its male parent. As far as we know, this was the first research reported in China on the diversity of the endophytic bacterial communities of the seeds of various maize hybrids with different genotypes.     

Effects of Bacterial Prey Species and Their Concentration on Growth of the Amoebae Acanthamoeba castellanii and Hartmannella vermiformis

Two amoebae were presented with six bacterial prey at a range of concentrations, and the growth parameters of the amoebae were deduced. All but one bacterium (Synechococcus) resulted in a positive growth response, but the gram-positive bacterium Staphylococcus aureus proved to be difficult to digest and the heavily pigmented bacterium Klebsiella ozaenae induced unusual amoebic behavior prior to ingestion.     

Identification and Characterization of a Symbiotic Bacterium Associated with Steinernema carpocapsae in Korea

A symbiotic bacterium was identified from the entomopathogenic nematode, Steinernema carpocapsae, collected in Korea and its biological significance was examined for insecticidal and antibiotic effects. The symbiotic bacterium was isolated from the nematode-infected hemolymph of the fifth instar larvae of Spodoptera exigua. The intra-hemocoelic injection of the bacterial isolates killed the insect hosts within 24h. Several biochemical and morphological characters of the bacterium were identical to those of Xenorhabdus nematophilus. The bacterial growth was not inhibited on the media containing 4,000 ppm of streptomycin sulfate or 2,000 ppm of penicillin. They showed antibacterial effects on Escherichia coli, Ralstonia solanacearum and Pseudomonas aeruginosa, but not on Pseudomonas putida and Pseudomonas fluorescens.     

A preliminary report of vibriosis in cultured American lobsters,Homarus americanus

A vibrio-like bacterium, designated Vibrio sp. (BML 79-078), was isolated from moribund juvenile Amercan lobsters, Homarus americanus. This is the first report of a vibrio-like bacterium being associated with a disease of lobsters. Koch's postulates have been satisfied for this bacterium. In this study, Vibrio sp. (BML 79-078) and Vibrio anguillarum (ATCC 19264) were found to be pathogenic when injected into juvenile lobsters held at 20°C, while Enterobacter aerogenes was not. An endotoxin was found to be associated with the pathogenesis of the disease syndrome caused by Vibrio sp. (BML 79-087).     

Molecular Characterization of the Nonphotosynthetic Partner Bacterium in the Consortium “Chlorochromatium aggregatum”

Phototrophic consortia represent valuable model systems for the study of signal transduction and coevolution between different bacteria. The phototrophic consortium “Chlorochromatium aggregatum” consists of a colorless central rod-shaped bacterium surrounded by about 20 green-pigmented epibionts. Although the epibiont was identified as a member of the green sulfur bacteria, and recently isolated and characterized in pure culture, the central colorless bacterium has been identified as a member of the β-Proteobacteria but so far could not be characterized further. In the present study, “C. aggregatum” was enriched chemotactically, and the 16S rRNA gene sequence of the central bacterium was elucidated. Based on the sequence information, fluorescence in situ hybridization probes targeting four different regions of the 16S rRNA were designed and shown to hybridize exclusively to cells of the central bacterium. Phylogenetic analyses of the 1,437-bp-long sequence revealed that the central bacterium of “C. aggregatum” represents a so far isolated phylogenetic lineage related to Rhodoferax spp., Polaromonas vacuolata, and Variovorax paradoxus within the family Comamonadaceae. The majority of relatives of this lineage are not yet cultured and were found in low-temperature aquatic environments or aquatic environments containing xenobiotica or hydrocarbons. In CsCl-bisbenzimidazole equilibrium density gradients, genomic DNA of the central bacterium of “Chlorochromatium aggregatum” formed a distinct band which could be detected by quantitative PCR using specific primers. Using this method, the G+C content of the central bacterium was determined to be 55.6 mol%.     

Brevibacillus laterosporus strain BPM3, a potential biocontrol agent isolated from a natural hot water spring of Assam, India

A bacterial strain designated as BPM3 isolated from mud of a natural hot water spring of Nambar Wild Life Sanctuary, Assam, India, strongly inhibited growth of phytopathogenic fungi (Fusarium oxysporum f. sp. ciceri, F. semitectum, Magnaporthe grisea and Rhizoctonia oryzae) and gram-positive bacterium (Staphylococcus aureus). The maximum growth and antagonistic activity was recorded at 30 °C, pH 8.5 when starch and peptone were amended as carbon and nitrogen sources, respectively. In greenhouse experiment, this bacterium (BPM3) suppressed blast disease of rice by 30-67% and protected the weight loss by 35-56.5%. The maximum disease protection (67%) and weight loss protection (56.5%) were recorded when the bacterium was applied before 2 days of the pathogen inoculation. Antifungal and antibacterial compounds were isolated from the bacterium which also inhibited the growth of these targeted pathogens. The compounds were purified and on spectroscopic analysis of a purified fraction having R_f 0.22 which showed strong antifungal and antibacterial activity indicated the presence of C-H, carbonyl group, dimethyl group, -CH₂ and methyl group. The bacterium was characterized by morphological, biochemical and molecular approaches and confirmed that the strain BPM3 is Brevibacillus laterosporus.     

Isolation and characterization of protective anti-LPS nanobody against V. cholerae O1 recognizing Inaba and Ogawa serotypes

Vibrio cholerae is considered one of the major health threats in developing countries. Lack of efficient vaccine, short incubating time of the disease, and bacterium ability to survive in aquatic environment have made cholera one of the most epidemic diseases yet known. The lipopolysaccharide is one of the bacterium key antigens used to classify V. cholerae into 206 serogroups. V. cholerae serogroup O1 is a causative agent of all cholera pandemics. Research has shown that anti-lipopolysaccharide (LPS) antibodies could provide protective immunity in cholera cases. In this research, we used N-terminal fragments of the camel's heavy-chain antibodies called VHH or nanobodies and produced a phagemid library. The obtained library was panned against V. cholerae O1 LPS, and four monoclonal nanobodies were isolated. Isolated nanobodies were tested in LPS ELISA and bacterial ELISA. The nanobody with the highest affinity toward the bacterium was used in an in vivo challenge and successfully neutralized the bacterium infection. The isolated nanobody showed high thermostability and proteolytic resistance in characterization tests.     

Genome Sequences of the Primary Endosymbiont “Candidatus Portiera aleyrodidarum” in the Whitefly Bemisia tabaci B and Q Biotypes

“Candidatus Portiera aleyrodidarum” is the obligate primary endosymbiotic bacterium of whiteflies, including the sweet potato whitefly Bemisia tabaci, and provides essential nutrients to its host. Here we report two complete genome sequences of this bacterium from the B and Q biotypes of B. tabaci.     

Bionomics of a Phoretic Association Between Paenibacillus sp. and the Entomopathogenic Nematode Steinernema diaprepesi

Spores of an unidentified bacterium were discovered adhering to cuticles of third-stage infective juvenile (IJ) Steinernema diaprepesi endemic in a central Florida citrus orchard. The spores were cup-shaped, 5 to 6 mm in length, and contained a central endospore. Based on 16S rDNA gene sequencing, the bacterium is closely related to the insect pathogens Paenibacillus popilliae and P. lentimorbus. However, unlike the latter bacteria, the Paenibacillus sp. is non-fastidious and grew readily on several standard media. The bacterium did not attach to cuticles of several entomopathogenic or plant-parasitic nematodes tested, suggesting host specificity to S. diaprepesi. Attachment of Paenibacillus sp. to the third-stage cuticle of S. diaprepesi differed from Paenibacillus spp. associated with heterorhabditid entomopathogenic nematodes, which attach to the IJ sheath (second-stage cuticle). The inability to detect endospores within the body of S. diaprepesi indicates that the bacterial association with the nematode is phoretic. The Paenibacillus sp. showed limited virulence to Diaprepes abbreviatus, requiring inoculation of larvae with 108 spores to achieve death of the insect and reproduction of the bacterium. The effect of the bacterium on the nematode population biology was studied in 25-cm-long vertical sand columns. A single D. abbreviatus larva was confined below 15-cm depth, and the soil surface was inoculated with either spore-free or spore-encumbered IJ nematodes. After 7 days, the proportion of IJ below 5-cm depth was seven-fold greater for spore-free IJ than for spore-encumbered nematodes. Mortality of D. abbreviatus larvae was 72% greater (P <= 0.01) for spore-free compared to spore-encumbered S. diaprepesi. More than 5 times as many progeny IJs (P <= 0.01) were produced by spore-free compared to spore-encumbered nematodes. These data suggest that the bacterium is a component of the D. abbreviatus food web with some potential to regulate a natural enemy of the insect.     

13C-NMR study of the glucose synthesis pathways in the bacterium Chlorobium thiosulfatophilum

Photoassimilation of ¹³CO₂ and acetate by the photosynthetic bacterium Chlorobium thiosulfatophilum was investigated using ¹³C-NMR as the method for determination of the labelling pattern of the glucose synthesized by the bacterium. Metabolic pathways functioning in the bacterium were identified by analysis of the multiplet structure of the spectra of tightly coupled systems. The labelling pattern showed that glucose was synthesized in C. thiosulfatophilum mainly by the gluconeogenesis pathway. In agreement with previous investigations, the reserve polysaccharide of C. thiosulfatophilum was shown to be polyglucose, with the glucose units linked predominantly by (1→4)α-glucosidic linkages.     

Isolation from the Sorghum bicolor Mycorrhizosphere of a Bacterium Compatible with Arbuscular Mycorrhiza Development and Antagonistic towards Soilborne Fungal Pathogens

A gram-positive bacterium with antagonistic activity towards soilborne fungal pathogens has been isolated from the mycorrhizosphere of Sorghum bicolor inoculated with Glomus mosseae. It has been identified as Paenibacillus sp. strain B2 based on its analytical profile index and on 16S ribosomal DNA analysis. Besides having antagonistic activity, this bacterium stimulates mycorrhization.     

Whole-Genome Sequence of Enterobacter sp. Strain SST3, an Endophyte Isolated from Jamaican Sugarcane (Saccharum sp.) Stalk Tissue

Enterobacter sp. strain SST3 is an endophytic bacterium isolated from Saccharum spp. Here we present its annotated draft genome that may shed light on its role as a bacterial endophyte of sugarcane. To our knowledge, this is the first genome announcement of a sugarcane-associated bacterium from the genus Enterobacter.     

Pseudomonas stutzeri YPL-1 Genetic Transformation and Antifungal Mechanism against Fusarium solani, an Agent of Plant Root Rot

An actively antagonistic bacterium that could be used as a biocontrol agent against Fusarium solani, which causes root rots with considerable losses in many important crops, was isolated from a ginseng rhizosphere and identified as a strain of Pseudomonas stutzeri. In several biochemical tests with culture filtrates of P. stutzeri YPL-1 and in mutational analyses of antifungal activities of reinforced or defective mutants, we found that the anti-F. solani mechanism of the bacterium may involve a lytic enzyme rather than a toxic substance or antibiotic. P. stutzeri YPL-1 produced extracellular chitinase and laminarinase when grown on different polymers such as chitin, laminarin, or F. solani mycelium. These lytic extracellular enzymes markedly inhibited mycelial growth rather than spore germination and also caused lysis of F. solani mycelia and germ tubes. Scanning electron microscopy revealed degradation of the F. solani mycelium. Abnormal hyphal swelling and retreating were caused by the lysing agents from P. stutzeri YPL-1, and a penetration hole was formed on the hyphae in the region of interaction with the bacterium; the walls of this region were rapidly lysed, causing leakage of protoplasm. Genetically bred P. stutzeri YPL-1 was obtained by transformation of the bacterium with a broad-host-range vector, pKT230. Also, the best conditions for the transformation were investigated.     

“Candidatus Contubernalis alkalaceticum,” an Obligately Syntrophic Alkaliphilic Bacterium Capable of Anaerobic Acetate Oxidation in a Coculture with Desulfonatronum cooperativum

From the silty sediments of the Khadyn soda lake (Tuva), a binary sulfidogenic bacterial association capable of syntrophic acetate oxidation at pH 10.0 was isolated. An obligately syntrophic, gram-positive, spore-forming alkaliphilic rod-shaped bacterium performs acetate oxidation in a syntrophic association with a hydrogenotrophic, alkaliphilic sulfate-reducing bacterium; the latter organism was previously isolated and characterized as the new species Desulfonatronum cooperativum. Other sulfate-reducing bacteria of the genera Desulfonatronum and Desulfonatronovibrio can also act as the hydrogenotrophic partner. Apart from acetate, the syntrophic culture can oxidize ethanol, propanol, isopropanol, serine, fructose, and isobutyric acid. Selective amplification of 16S rRNA gene fragments of the acetate-utilizing syntrophic component of the binary culture was performed; it was found to cluster with clones of uncultured gram-positive bacteria within the family Syntrophomonadaceae. The acetate-oxidizing bacterium is thus the first representative of this cluster obtained in a laboratory culture. Based on its phylogenetic position, the new acetate-oxidizing syntrophic bacterium is proposed in the Candidatus status for a new genus and species: “Candidatus Contubernalis alkalaceticum.”     

Fate of Escherichia coli O157:H7 on Fresh-Cut Apple Tissue and Its Potential for Transmission by Fruit Flies

Pathogenic Escherichia coli O157:H7, as well as nonpathogenic strains ATCC 11775 and ATCC 23716, grew exponentially in wounds on Golden Delicious apple fruit. The exponential growth occurred over a longer time period on fruit inoculated with a lower concentration of the bacterium than on fruit inoculated with a higher concentration. The bacterium reached the maximum population supported in the wounds regardless of the initial inoculum concentrations. Populations of E. coli O157:H7 in various concentrations of sterilized apple juice and unsterilized cider declined over time and declined more quickly in diluted juice and cider. The decline was greater in the unsterilized cider than in juice, which may have resulted from the interaction of E. coli O157:H7 with natural populations of yeasts that increased with time. Experiments on the transmission of E. coli by fruit flies, collected from a compost pile of decaying apples and peaches, were conducted with strain F-11775, a fluorescent transformant of nonpathogenic E. coli ATCC 11775. Fruit flies were easily contaminated externally and internally with E. coli F-11775 after contact with the bacterium source. The flies transmitted this bacterium to uncontaminated apple wounds, resulting in a high incidence of contaminated wounds. Populations of the bacterium in apple wounds increased significantly during the first 48 h after transmission. Further studies under commercial conditions are necessary to confirm these findings.     

Whole-Genome Sequence of N-Acylhomoserine Lactone-Synthesizing and -Degrading Acinetobacter sp. Strain GG2

Acinetobacter sp. strain GG2 is a quorum-sensing and quorum-quenching bacterium isolated from the ginger rhizosphere. It degrades a broad range of N-acylhomoserine lactone molecules via lactonase. The genome sequence of strain GG2 may provide insights on the regulation of quorum-sensing and quorum-quenching mechanisms in this bacterium.     

Wolbachia infection in the Korean endemic firefly,Luciola unmunsana (Coleoptera: Lampyridae)

Wolbachia is one of the most prevalent endosymbiontic bacteria of arthropods. The bacterium induces sex ratio distortions in various host insects through processes such as cytoplasmic incompatibility, feminization, male killing, and parthenogenesis. We investigated if the Korean endemic firefly, Luciola unmunsana was infected with the bacterium because the species had an abnormal sex ratio in the field. The results show that some individuals are infected with the bacterium. Phylogenetic analyses showed that the bacterial strain infecting the firefly is closely related to strains that infect phylogenetically distant hosts.     

Evidence for two subclasses of methylamine dehydrogenases with distinct large subunits and conserved PQQ-bearing small subunits

Antibodies, which were raised to the isolated subunits of the methylamine dehydrogenases of Paracoccus denitrificans and bacterium W3A1, were used to demonstrate immunological cross-reactivity between the small pyrrolo-quinoline quinone bearing subunits of these two enzymes. The small subunits of these enzymes also exhibited nearly identical pI values. Conversely, the large subunits of the P. denitrificans and bacterium W3A1 enzymes did not cross-react immunologically and exhibited, respectively, very acidic and very basic pI values which reflected the electrostatic properties of their respective holoenzymes. Antibodies specific for the large subunit of the P. denitrificans enzyme, but not the bacterium W3A1 enzyme, cross-reacted with the analogous proteins in cell extracts of Thiobacillus versutus and Methylobacterium sp strain AM1.     

Gut bacterium of Dendrobaena veneta (Annelida: Oligochaeta) possesses antimycobacterial activity

The new bacterial strain with antimycobacterial activity has been isolated from the midgut of Dendrobaena veneta (Annelida). Biochemical and molecular characterization of isolates from 18 individuals identified all as Raoultella ornithinolytica genus with 99% similarity. The bacterium is a possible symbiont of the earthworm D. veneta. The isolated microorganism has shown the activity against four strains of fast-growing mycobacteria: Mycobacterium butiricum, Mycobacterium jucho, Mycobacterium smegmatis and Mycobacterium phlei. The multiplication of the gut bacterium on plates with Sauton medium containing mycobacteria has caused a lytic effect. After the incubation of the cell free extract prepared from the gut bacterium with four strains of mycobacteria in liquid Sauton medium, the cells of all tested strains were deformed and divided to small oval forms and sometimes created long filaments. The effect was observed by the use of light, transmission and scanning microscopy. Viability of all examined species of mycobacteria was significantly decreased. The antimycobacterial effect was probably the result of the antibiotic action produced by the gut bacterium of the earthworm. The application of ultrafiltration procedure allowed to demonstrate that antimicrobial substance with strong antimycobacterial activity from bacterial culture supernatant, is a protein with the molecular mass above 100 kDa.