Characterization and complete genome sequence analysis of the novel phage RPZH3 infecting Ralstonia solanacearum.

A novel lytic Ralstonia phage, RPZH3, was isolated from the soil of a tobacco field via a double agar overlay plaque assay. The phage has an icosahedral head 75 ± 5 nm in diameter with a short tail about 15 ± 5 nm in length. It was able to infect 18 out of 30 tested strains of R. solanacearum isolated from tobacco, sweet potato, tomato, pepper, and eggplant. The latent period of the phage was 80 min, and the burst period was 60 min with a burst size of about 27 pfu/cell. The phage was stable at pH 4-12 at 28°C, and it was also stable at temperatures from 45°C to 60°C at pH 7.0. The complete genome of phage RPZH3 consists of 65,958 bp, with a GC content of 64.93%. The genome contains 93 open reading frames (ORFs) and encodes a tRNA for cysteine. Nucleotide sequence alignment and phylogenetic analysis indicated that RPZH3 is a new member of the genus Gervaisevirus belonging to the class Caudoviricetes.

[Isolation and identification of a bacterial strain JS018 capable of degrading several kinds of organophosphate pesticides].

Organophosphate pesticides are used widely all over the world and play an important role in plant pest control. However these pesticides are considered as pollutants and harmful to human health. To search for microorganisms that can degrade organophosphate pesticides with high efficiency, a bacterial strain, coded as JS018, was isolated and screened from the soil in the vicinity of Shanming Pesticides Factory, Shanming, Fujian. Laboratory tests showed that the bacterium could degrade several kinds of organophosphate pesticides, such as Parathion-methyl and phoxin. The strain's degrading rates on phoxin, Parathion-methyl, hostathion and dichlorvos in LB liquid fermentation medium for 36 h were 99%, 96%, 80.4% and 69.0% respectively. The bacterial colonies on LB plate appeared shiny and pale-pink in color. The bacteria were Gram-negative coccoids, 0.5 - 0.7 microm in diameter. They grew well at 30 - 38 degrees C and pH 7.0 - 9.0. The optimal temperature and pH for cell growth was 32 degrees C and pH 7.5 - 8.0, respectively. They did not grow in medium containing 6% or more NaCl. The antibiotic susceptibility tests showed that the strain was resistant to ampicillin, penicillin and lincomycin. It was sensitive to kanamycin, tetracycline and gentamicin. Laboratory tests also showed that the strain could ferment D-glucose, trehalose, melezitose and ethanol. It was negative in the production of indole and hydrogen sulfide. It could not liquefy gelatin, utilize citrate, nor ferment L-arabinose, sucrose, D-mannitol, D-xylose, fructose, D-galactose, maltose or lactose. The catalase, urease and nitrate reduction were positive. Based on its morphological, physiological and biochemical properties as well as the 16S rDNA sequence analysis result, the strain was tentatively identified as Roseomonas sp.

The Endo-ß-1,4-Glucanase of Bacillus amyloliquefaciens Is Required for Optimum Endophytic Colonization of Plants.

The eglS gene in Bacillus amyloliquefaciens encodes an endo-ß-1,4-glucanase that belongs to glycosyl hydrolase family 5. In this study, a disruption mutant of gene eglS was constructed to examine its role in bacterial adaptation in plants. The mutant TB2k, eglS gene inactivated bacterial strain, was remarkably impaired in extracellular cellulase activity. When inoculated on Brassica campestris, the TB2k population was reduced by more than 60% compared with the wild-type strain in the root, stem, and leaf tissues. Overexpression of eglS in the wild-type strain increased the bacteria population in the plant tissues. Further studies revealed that the transcription level of eglS was correlated with bacterial population. These data demonstrate that endo-ß-1,4-glucanase of B. amyloliquefaciens is required for its optimal endophytic colonization.

[Colonization and disease control and fruit preservation functions of endophytic bacterial strains in lychee].

By spraying the GFP-marked endophytic bacterial strains BS-2-gfp and TB2-gfp, this paper studied their colonization in lychee organs and the functions of the strains in disease control and fruit preservation. The BS-2-gfp and TB2-gfp could colonize and propagate in lychee leaves, flowers, un-matured fruits, and matured fruits, and transfer from the flowers to un-matured fruits. The colonization of BS-2-gfp and TB2-gfp in lychee leaves varied with season and growth stage, being larger in quantity and longer in duration in spring than in autumn. The colonization quantity and duration of the strains also differed in other organs. Both the BS-2-gfp and the TB2-gfp could be isolated and recovered from lychee leaves after 37 d inoculation, the BS-2-gfp could not be isolated from the flowers after inoculation for 10 d, and the BS-2-gfp and TB2-gfp had the largest colonization quantity in matured fruits. The colonization quantity of TB2-gfp in lychee pericarp reached to the maximum (1.90 x 10(6) CFU x g(-1) FM) when the disease index of litchi downy blight had a sharp increase, and, compared with BS-2-gfp, the TB2-gfp had better fruit preservation efficiency, and its colonization quantity in lychee pericarp was also higher. It was suggested that there was a positive correlation between the colonization quantity of test bacterial strains in lychee pericarp and the disease control and fruit preservation effect.

Isolation of GPD promoter from Tremella fuciformis and driving expression of EGFP gene.

Glyceraldehyde-3-phosphate dehydrogenase (GPD) cDNA was cloned by RT-PCR using total RNA from Tremella fuciformis as template with a pair of degenerate primers. Then, a 500-bp 5'-upstream promoter region of the gene encoding GPD from T. fuciformis genomic DNA was isolated by thermal asymmetric interlaced PCR. The cloned promoter was fused to 5'-upstream of enhanced green fluorescent protein gene to construct T. fuciformis expression vector pCB-TEGFP with hygromycin gene as a selectable marker. Electroporation was performed to transfer plasmid DNA of pCB-TEGFP into yeast-like conidia from T. fuciformis. Molecular evidence, including PCR analysis, fluorescence detection, fluorescence spectra assay, and SDS-PAGE, indicated that the EGFP gene had been integrated into the genome of transgenic T. fuciformis strains and was expressed successfully. The results also showed that this promoter could be used to carry out regulated expression of heterologous gene products in T. fuciformis.