Over-expression of the <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> harpin-encoding gene <Emphasis Type="Italic">hrpZm</Emphasis> confers enhanced tolerance to Phytophthora root and stem rot in transgenic soybean

Phytophthora root and stem rot (PRR) caused by Phytophthora sojae is one of the most devastating diseases reducing soybean (Glycine max) production all over the world. Harpin proteins in many plant pathogenic bacteria were confirmed to enhance disease and insect resistance in crop plants. Here, a harpin protein-encoding gene hrpZpsta from the P. syringae pv. tabaci strain Psta218 was codon-optimized (renamed hrpZm) and introduced into soybean cultivars Williams 82 and Shennong 9 by Agrobacterium-mediated transformation. Three independent transgenic lines over-expressing hrpZm were obtained and exhibited stable and enhanced tolerance to P. sojae infection in T₂–T₄ generations compared to the non-transformed (NT) and empty vector (EV)-transformed plants. Quantitative real-time PCR (qRT-PCR) analysis revealed that the expression of salicylic acid-dependent genes PR1, PR12, and PAL, jasmonic acid-dependent gene PPO, and hypersensitive response (HR)-related genes GmNPR1 and RAR was significantly up-regulated after P. sojae inoculation. Moreover, the activities of defense-related enzymes such as phenylalanine ammonia lyase (PAL), polyphenoloxidase (PPO), peroxidase, and superoxide dismutase also increased significantly in the transgenic lines compared to the NT and EV-transformed plants after inoculation. Our results suggest that over-expression of the hrpZm gene significantly enhances PRR tolerance in soybean by eliciting resistance responses mediated by multiple defense signaling pathways, thus providing an alternative approach for development of soybean varieties with improved tolerance against the soil-borne pathogen PRR.     

Genetic mapping and development of co-segregating markers of <Emphasis Type="Italic">RpsQ</Emphasis>, which provides resistance to <Emphasis Type="Italic">Phytophthora sojae</Emphasis> in soybean

Key message

The RpsQ Phytophthora resistance locus was finely mapped to a 118-kb region on soybean chromosome 3. A best candidate gene was predicted and three co-segregating gene markers were developed.

Abstract

Phytophthora root rot (PRR), caused by Phytophthora sojae, is a major threat to sustainable soybean production. The use of genetically resistant cultivars is considered the most effective way to control this disease. The Chinese soybean cultivar Qichadou 1 exhibited a broad spectrum resistance, with a distinct resistance phenotype, following inoculation with 36 Chinese P. sojae isolates. Genetic analyses indicated that the disease resistance in Qichadou 1 is controlled by a single dominant gene. This gene locus was designated as RpsQ and mapped to a 118-kb region between BARCSOYSSR_03_0165 and InDel281 on soybean chromosome 3, and co-segregated with Insert11, Insert144 and SNP276. Within this region, there was only one gene Glyma.03g27200 encoding a protein with a typical serine/threonine protein kinase structure, and the expression pattern analysis showed that this gene induced by P. sojae infection, which was suggested as a best candidate gene of RpsQ. Candidate gene specific marker Insert144 was used to distinguish RpsQ from the other known Rps genes on chromosome 3. Identical polymerase chain reaction amplification products were produced for cultivars Qichadou 1 (RpsQ) and Ludou 4 (Rps9). All other cultivars carrying Rps genes on chromosome 3 produced different PCR products, which all lacked a 144-bp fragment present in Qichadou 1 and Ludou 4. The phenotypes of the analyzed cultivars combined with the physical position of the PRR resistance locus, candidate gene analyses, and the candidate gene marker test revealed RpsQ and Rps9 are likely the same gene, and confer resistance to P. sojae.

     

Next-generation sequencing to identify candidate genes and develop diagnostic markers for a novel Phytophthora resistance gene, <Emphasis Type="Italic">RpsHC18</Emphasis>, in soybean

Key message

A novel Phytophthora sojae resistance gene RpsHC18 was identified and finely mapped on soybean chromosome 3. Two NBS–LRR candidate genes were identified and two diagnostic markers of RpsHC18 were developed.

Abstract

Phytophthora root rot caused by Phytophthora sojae is a destructive disease of soybean. The most effective disease-control strategy is to deploy resistant cultivars carrying Phytophthora-resistant Rps genes. The soybean cultivar Huachun 18 has a broad and distinct resistance spectrum to 12 P. sojae isolates. Quantitative trait loci sequencing (QTL-seq), based on the whole-genome resequencing (WGRS) of two extreme resistant and susceptible phenotype bulks from an F_2:3 population, was performed, and one 767-kb genomic region with ΔSNP-index ≥ 0.9 on chromosome 3 was identified as the RpsHC18 candidate region in Huachun 18. The candidate region was reduced to a 146-kb region by fine mapping. Nonsynonymous SNP and haplotype analyses were carried out in the 146-kb region among ten soybean genotypes using WGRS. Four specific nonsynonymous SNPs were identified in two nucleotide-binding sites–leucine-rich repeat (NBS–LRR) genes, RpsHC18-NBL1 and RpsHC18-NBL2, which were considered to be the candidate genes. Finally, one specific SNP marker in each candidate gene was successfully developed using a tetra-primer ARMS-PCR assay, and the two markers were verified to be specific for RpsHC18 and to effectively distinguish other known Rps genes. In this study, we applied an integrated genomic-based strategy combining WGRS with traditional genetic mapping to identify RpsHC18 candidate genes and develop diagnostic markers. These results suggest that next-generation sequencing is a precise, rapid and cost-effective way to identify candidate genes and develop diagnostic markers, and it can accelerate Rps gene cloning and marker-assisted selection for breeding of P. sojae-resistant soybean cultivars.

     

First report of an atypical new <Emphasis Type="Italic">Aspergillus parasiticus</Emphasis> isolates with nucleotide insertion in <Emphasis Type="Italic">aflR</Emphasis> gene resembling to <Emphasis Type="Italic">A. sojae</Emphasis>

Aflatoxins are toxic and carcinogenic secondary metabolites produced primarily by the filamentous fungi Aspergillus flavus and Aspergillus parasiticus and cause toxin contamination in food chain worldwide. Aspergillus oryzae and Aspergillus sojae are highly valued as koji molds in the traditional preparation of fermented foods, such as miso, sake, and shoyu. Koji mold species are generally perceived of as being nontoxigenic and are generally recognized as safe (GRAS). Fungal isolates were collected from a California orchard and a few were initially identified to be A. sojae using β-tubulin gene sequences blasted against NCBI data base. These new isolates all produced aflatoxins B₁, B₂, G₁, and G₂ and were named as Pistachio Winter Experiment (PWE) strains. Thus, it is very important to further characterize these strains for food safety purposes. The full length of aflR gene of these new isolates was sequenced. Comparison of aflR DNA sequences of PWE, A. parasiticus and A. sojae, showed that the aflatoxigenic PWE strains had the six base insertion (CTCATG) similar to domesticated A. sojae, but a pre-termination codon TGA at nucleotide positions 1153–1155 was absent due to a nucleotide codon change from T to C. Colony morphology and scanning microscopic imaging of spore surfaces showed similarity of PWE strains to both A. parasiticus and A. sojae. Concordance analysis of multi locus DNA sequences indicated that PWE strains were closely linked between A. parasiticus and A. sojae. The finding documented the first report that such unique strains have been found in North America and in the world.     

Isolation,characterization and virulence of bacteria from <Emphasis Type="Italic">Ostrinia nubilalis</Emphasis> (Lepidoptera: Pyralidae)

Isolation, characterization and virulence of the culturable bacteria from entire tissues of larval Ostrinia nubilalis (Hübner) (Lepidoptera: Pyralidae) were studied to obtain new microbes for biological control. A total of 16 bacteria were isolated from living and dead larvae collected from different maize fields in the Eastern Black Sea Region of Turkey. The bacterial microbiota of O. nubilalis were identified as Pseudomonas aeruginosa (On1), Brevundimonas aurantiaca (On2), Chryseobacterium formosense (On3), Acinetobacter sp. (On4), Microbacterium thalassium (On5), Bacillus megaterium (On6), Serratia sp. (On7), Ochrobactrum sp. (On8), Variovorax paradoxus (On9), Corynebacterium glutamicum (On10), Paenibacillus sp. (On11), Alcaligenes faecalis (On12), Microbacterium testaceum (On13), Leucobacter sp. (On14), Leucobacter sp. (On15) and Serratia marcescens (On16) based on their morphological and biochemical characteristics. A partial sequence of the 16S rRNA gene was also determined to confirm strain identification. The highest insecticidal activities were obtained from P. aeruginosa On1 (80%), Serratia sp. On7 (60%), V. paradoxus On9 (50%) and S. marcescens On16 (50%) against larvae 14 days after treatment (p < 0.05). Also, the highest activity from previously isolated Bacillus species was observed from Bacillus thuringiensis subsp. tenebrionis Xd3 with 80% mortality within the same period (p < 0.05). Our results indicate that P. aeruginosa On1, Serratia sp. On7, V. paradoxus On9, S. marcescens On16 and B. thuringiensis subsp. tenebrionis Xd3 show potential for biocontrol of O. nubilalis.     

Genome-wide analysis of the PHB gene family in <Emphasis Type="Italic">Glycine max</Emphasis> (L.) Merr.

Prohibitins (PHBs) have one SPFH domain in common and present in species ranging from prokaryotes to eukaryotes. Although a number of researches on PHBs were performed in different plant species, a systematic analysis of the PHB family in soybean is still remains uncharacterized. In the present study, 24 putative PHB genes have been first systemically identified in soybean. According to phylogenetic analysis, these GmPHBs could be classified into four groups. Gene structures and motif patterns showed high levels of conservation within the phylogenetic subgroups. Several members of this family have undergone purifying selection based on Ka/Ks analysis on duplicated PHB genes in soybean. We performed microsynteny analysis across four legume species based on the comparisons among the specific regions contained in PHB genes. As a result, numerous microsyntenic gene pairs among soybean, Medicago, Lotus and Phaseolus were identified. Most soybean PHB genes exhibited different expression levels in various tissues and developmental stages through expression analysis using publicly available RNA-seq datasets. The 11 GmPHB genes from III_B subgroup were examined by qPCR for their expression in two soybean cultivar after infection by Phytophthora sojae. Besides three GmPHB genes previous reported by us, here other four genes also were rapidly induced by P. sojae infection in the resistant genotype, while induction was very weak in the susceptible genotype. The comprehensive overview of the PHB gene family in soybean genome will provide useful information for further functional analysis of the PHB gene family in soybean.     

Diversity of indigenous endophytic bacteria associated with the roots of Chinese cabbage (<Emphasis Type="Italic">Brassica campestris</Emphasis> L.) cultivars and their antagonism towards pathogens

The study aimed to reveal the diversity of endophytic bacteria in the roots of Chinese cabbage (CC) cultivated in two areas in Korea, namely, Seosang-gun (SS) and Haenam-gun (HN), and also in a transgenic plant (TP) from the laboratory. A total of 653 colonies were isolated from the interior of CC roots, comprising 118, 302, and 233 isolates from SS, HN, and TP samples, respectively. Based on 16S rRNA gene sequence analysis, the isolates belonged to four major phylogenetic groups: high-G+C Gram-positive bacteria (HGC-GPB), low-G+C Gram-positive bacteria (LGC-GPB), Proteobacteria, and Bacteriodetes. The most dominant groups in the roots of the SS, HN, and TP cultivars were LGC-GPB (48.3%), Proteobacteria (50.2%), and HGC-GPB (38.2%), respectively. Importantly, most of the isolates that produced cell-walldegrading enzymes belonged to the genus Bacillus. Bacillus sp. (HNR03, TPR06), Bacillus pumilus (SSR07, HNR11, TPR07), and Bacillus subtilis (TPR03) showed high antagonism against the tested food-borne pathogenic bacteria. In addition, Bacillus sp. (HNR03, TPR06), Bacillus pumilus (SSR07, HNR11, HNR17, TPR11), Microbacterium oxidans (SSR09, TPR04), Bacillus cereus HNR10, Pseudomonas sp. HNR13, and Bacillus subtilis (TPR02, TPR03) showed strong antagonistic activity against the fungi Phythium ultimum, Phytophthora capsici, Fusarium oxysporum, and Rhizoctonia solani. The endophytes isolated from the TP cultivar showed the strongest antagonistic reactions against pathogens. This study is the first report on endophytic bacteria from Chinese cabbage roots.     

Expression of plant antimicrobial peptide <Emphasis Type="Italic">pro-SmAMP2</Emphasis> gene increases resistance of transgenic potato plants to <Emphasis Type="Italic">Alternaria</Emphasis> and <Emphasis Type="Italic">Fusarium</Emphasis> pathogens

The chickweed (Stellaria media L.) pro-SmAMP2 gene encodes the hevein-like peptides that have in vitro antimicrobial activity against certain harmful microorganisms. These peptides play an important role in protecting the chickweed plants from infection, and the pro-SmAMP2 gene was previously used to protect transgenic tobacco and Arabidopsis plants from phytopathogens. In this study, the pro-SmAMP2 gene under control of viral CaMV35S promoter or under control of its own pro-SmAMP2 promoter was transformed into cultivated potato plants of two cultivars, differing in the resistance to Alternaria: Yubiley Zhukova (resistant) and Skoroplodny (susceptible). With the help of quantitative real-time PCR, it was demonstrated that transgenic potato plants expressed the pro-SmAMP2 gene under control of both promoters at the level comparable to or exceeding the level of the potato actin gene. Assessment of the immune status of the transformants demonstrated that expression of antimicrobial peptide pro-SmAMP2 gene was able to increase the resistance to a complex of Alternaria sp. and Fusarium sp. phytopathogens only in potato plants of the Yubiley Zhukova cultivar. The possible role of the pro-SmAMP2 products in protecting potatoes from Alternaria sp. and Fusarium sp. is discussed.     

New species of the genus <Emphasis Type="Italic">Spathius</Emphasis> Nees (Hymenoptera,Braconidae) from Japan and neighboring territories

Eight new species of the genus Spathius Nees from Japan and the Russian Far East are described and illustrated: S. enigmatus sp. n., S. ishigakus sp. n., S. konishii sp. n., S. multigranulatus sp. n., S. omotodakus sp. n., S. pseudaspersus sp. n., S. pumilio sp. n., and S. tsushimus sp. n.     

Identification of the di-<Emphasis Type="Italic">n</Emphasis>-butyl phthalate-biodegrading strains and the biodegradation pathway in strain LMB-1

DNA isolated from a greenhouse soil (Nanjing, Jiangsu Province, China) was suitable for PCR amplification of gene segment coding for the 16S rRNA. Diverse PCR products were characterized by cloning and sequencing, and analysis of bacterial colonies showed the presence over 26 phyla. The most bacteria belonged to Proteobacteria, Actinobacteria, Gemmatimonadetes, Acidobacteria and Planctomycetes. Furthermore, after the enrichment procedure of DBP-degrading microorganisms, 4 strains were isolated from the soil sample with di-n-butyl phthalate (DBP) biodegradability, and they were identified to be Rhizobium sp., Streptomyces sp., Pseudomonas sp. and Acinetobacter sp. Analysis of the degradation products by LC-MS led to identification of metabolites of DBP in strain LMB-1 (identified as Rhizobium sp.) which suggests that DBP was degraded through β-oxidation, demethylation, de-esterification and cleavage of aromatic ring.     

Diversity of endophytic fungal and bacterial communities in <Emphasis Type="Italic">Ilex paraguariensis</Emphasis> grown under field conditions

The composition and diversity of the endophytic community associated with yerba mate (Ilex paraguariensis) was investigated using culture-depending methods. Fungi were identified based on their micromorphological characteristics and internal transcribed spacer rDNA sequence analysis; for bacteria 16S rDNA sequence analysis was used. Fungal and bacterial diversity did not show significant differences between organ age. The highest fungal diversity was registered during fall season and the lowest in winter. Bacterial diversity was higher in stems and increased from summer to winter, in contrast with leaves, which decreased. The most frequently isolated fungus was Fusarium, followed by Colletotrichum; they were both present in all the sampling seasons and organ types assayed. Actinobacteria represented 57.5 % of all bacterial isolates. The most dominant bacterial taxa were Curtobacterium and Microbacterium. Other bacteria frequently found were Methylobacterium, Sphingomonas, Herbiconiux and Bacillus. Nitrogen fixation and phosphate solubilization activity, ACC deaminase production and antagonism against plant fungal pathogens were assayed in endophytic bacterial strains. In the case of fungi, strains of Trichoderma, Penicillium and Aspergillus were assayed for antagonism against pathogenic Fusarium sp. All microbial isolates assayed showed at least one growth promoting activity. Strains of Bacillus, Pantoea, Curtobacterium, Methylobacterium, Brevundimonas and Paenibacillus had at least two growth-promoting activities, and Bacillus, Paenibacillus and the three endophytic fungi showed high antagonistic activity against Fusarium sp. In this work we have made a wide study of the culturable endophytic community within yerba mate plants and found that several microbial isolates could be considered as potential inoculants useful for improving yerba mate production.     

Analysis of antimicrobial and immunomodulatory substances produced by heterofermentative <Emphasis Type="Italic">Lactobacillus reuteri</Emphasis>

Antimicrobial and immunomodulatory potential of various Lactobacillus reuteri strains is closely connected to their metabolite production profile under given cultivation conditions. We determined the in vitro production of antimicrobial substances such as organic acids, ethanol, and reuterin by four strains of L. reuteri (L. reuteri E, L. reuteri KO5, L. reuteri CCM 3625, and L. reuteri ATCC 55730). All studied L. reuteri strains showed the ability to produce lactic acid, acetic acid, and ethanol with concominant consumption of glucose and together with phenyllactic acid—a potent antifungal compound—with concominant consumption of phenylalanine. The reuterin production from glycerol was confirmed for all analyzed lactobacilli strains except L. reuteri CCM 3625. Production of organic acids, ethanol, and reuterin is significantly involved in antimicrobial activity of lactobacilli which was determined using the dual-culture overlay diffusion method against six indicator bacteria and five indicator moulds. In comparison to the referential L. reuteri ATCC 55730, the highest inhibition potential was observed against Escherichia coli CCM 3988 and Pseudomonas aeruginosa CCM 3955. Among analyzed indicators of moulds, the growth of Alternaria alternata CCM F-128 was the most inhibited by all four analyzed L. reuteri strains. Finally, the immunomodulatory potential of analyzed lactobacilli were proven by the determination of the in vitro production of biogenic amines histamine and tyramine. L. reuteri CCM 3625 was able to produce tyramine, and L. reuteri E and L. reuteri KO5 were able to produce histamine under given cultivation conditions.     

Role of bacterial pyrroloquinoline quinone in phosphate solubilizing ability and in plant growth promotion on strain <Emphasis Type="Italic">Serratia</Emphasis> sp. S119

The aim of this study was to analyze if cofactor pyrroquinoline quinone from Serratia sp. S119 is involved in the inorganic phosphate solubilization mechanism and in its ability to promote the plant growth. Site directed mutagenesis was performed to obtain a pqqE- minus mutant of strain Serratia sp. S119. The phosphate solubilization ability, gluconate and PQQ production of the mutant Serratia sp. RSL (pqqE-) was analyzed. Mutant RSL (pqqE-) showed significant decrease in P soluble and gluconic acid levels produced and undetectable levels of PQQ cofactor compared with wild-type strain. Complementation with synthetic PQQ cofactor restored P solubilization and gluconate production reaching the levels produced by wild-type strain. PqqE gene sequence indicated that it is highly conserved within Serratia strains and its product shows conserved motifs found in other PqqE proteins of several bacteria. The effect of the inoculation of the PQQ- mutant on peanut and maize plants was evaluated in pot assays. Plants growth parameters showed no differences among the different treatments indicating that PQQ from Serratia sp. S119 is not involved in the growth promotion of these plants. PQQ cofactor is essential for phosphate solubilization ability of Serratia sp. S119 but is not required for growth promotion of peanut and maize plants.     

<Emphasis Type="Italic">Paralepidapedon variabile</Emphasis> sp. n. (Trematoda,Lepocreadioidea, Lepidapedidae) and other representatives of the genus <Emphasis Type="Italic">Paralepidapedon</Emphasis> from Antarctic fish

In the Ross Sea and Amundsen Sea, four representatives of the genus Paralepidapedon—Paralepidapedon cf. dubium Prudhoe et Bray 1973 sensu Sokolov et Gordeev 2013, P. lepidum (Gaevskaya et Rodyuk 1988), Paralepidapedon sp., and P. variabile sp. n.—were found in demersal fishes Muraenolepis marmorata and Macrourus whitsoni. Paralepidapedon variabile sp. n. is described from Muraenolepis marmorata in the Amundsen Sea. Paralepidapedon variabile sp. n. differs from other species of the genus Paralepidapedon by the position of the anterior border of the vitellarium at the level of the anterior edge of the ventral sucker or genital pore and by the highly variable shape of the testes: from roundish with a smooth edge to sinuate–lobate. Paralepidapedon lepidum was found for the first time in the Antarctic.     

Bacterial diversity of the outflows of a Polichnitos (Lesvos,Greece) hot spring,laboratory studies of a <Emphasis Type="Italic">Cyanobacterium</Emphasis> sp. strain and potential medical applications

The bacterial diversity of the outflows of Polichnitos (Lesvos, Greece) hot spring has been investigated. Cyanobacteria showing high sequence homologies with Phormidium sp. and Cyanobacterium aponinum were found. Members of the Alphaproteobacteria closely related to Rhodobium sp. Albidovulum sp., Rhodobacter sp., Microvigra sp., Nitratireductor sp. and Phaeobacter sp. Gammaproteobacteria, Betaproteobacteria and Firmicutes were represented by members of Idiomarina sp., Marinobacter sp., Shinella sp., Bacillus sp. and Clostridium sp. with sequence homologies ranging from 92% to 100%. Members of the Bacteroidetes and Planctomycetes were represented by sequences of novel phylogenetic linkages exhibiting 87–90% sequence homology with type strains. When the hot spring consortium was cultivated in bioreactor repeated batch culture under photo-autotrophic growth conditions at temperature < 30 °C, Cyanobacterium sp. dominated over Phormidium sp. Cyanobacterium sp. seems to have biotechnological potential since its extracellular broth exhibited a strong insecticidal activity against larvae of Aedes aegypti (a vector of important human diseases) and significant anti-cancer activity against the PC3 human prostate cancer cell line, while its toxicity against human endothelial cells was relatively low.     

<Emphasis Type="SmallCaps">d</Emphasis>-Xylose fermentation,xylitol production and xylanase activities by seven new species of <Emphasis Type="Italic">Sugiyamaella</Emphasis>

Sixteen yeast isolates identified as belonging to the genus Sugiyamaella were studied in relation to D-xylose fermentation, xylitol production, and xylanase activities. The yeasts were recovered from rotting wood and sugarcane bagasse samples in different Brazilian regions. Sequence analyses of the internal transcribed spacer (ITS) region and the D1/D2 domains of large subunit rRNA gene showed that these isolates belong to seven new species. The species are described here as Sugiyamaella ayubii f.a., sp. nov. (UFMG-CM-Y607^T = CBS 14108^T), Sugiyamaella bahiana f.a., sp. nov. (UFMG-CM-Y304^T = CBS 13474^T), Sugiyamaella bonitensis f.a., sp. nov. (UFMG-CM-Y608^T = CBS 14270^T), Sugiyamaella carassensis f.a., sp. nov. (UFMG-CM-Y606^T = CBS 14107^T), Sugiyamaella ligni f.a., sp. nov. (UFMG-CM-Y295^T = CBS 13482^T), Sugiyamaella valenteae f.a., sp. nov. (UFMG-CM-Y609^T = CBS 14109^T) and Sugiyamaella xylolytica f.a., sp. nov. (UFMG-CM-Y348^T = CBS 13493^T). Strains of the described species S. boreocaroliniensis, S. lignohabitans, S. novakii and S. xylanicola, isolated from rotting wood of Brazilian ecosystems, were also compared for traits relevant to xylose metabolism. S. valenteae sp. nov., S. xylolytica sp. nov., S. bahiana sp. nov., S. bonitensis sp. nov., S. boreocarolinensis, S. lignohabitans and S. xylanicola were able to ferment d-xylose to ethanol. Xylitol production was observed for all Sugiyamaella species studied, except for S. ayubii sp. nov. All species studied showed xylanolytic activity, with S. xylanicola, S. lignohabitans and S. valenteae sp. nov. having the highest values. Our results suggest these Sugiyamaella species have good potential for biotechnological applications.     

Taxonomic composition of bacteria associated with cultivated mollusks <Emphasis Type="Italic">Crassostrea lugubris</Emphasis> and <Emphasis Type="Italic">Perna viridis</Emphasis> and with the water of the Gulf of Nha Trang lagoon,Vietnam

One hundred and four strains of heterotrophic bacteria have been isolated and characterized from two species of bivalve mollusks cultivated in the Gulf of Nha Trang (Vietnam) and from the water of a mariculture farm. The isolates have been identified on the basis of morphological, physiological, biochemical, and chemotaxonomic properties, as well as by the content of G+C bases in DNA. In the microflora of mollusks, Vibrio alginolyticus was predominant; the pathogenic species V. harveyi and V. splendidus were found as well. Staphylococci and bacilli occupied the second place in abundance after vibrios. In addition, coryneforms and enterobacteria, as well as Pseudomonas spp. and Pseudoalteromonas spp., were revealed. The composition of the water microflora was more diverse as compared with the microflora of mollusks. In the water, Bacillus spp., Vibrio spp., and Pseudomonas spp. were predominant. Brevibacterium spp. and other coryneform bacteria, as well as enterobacteria, occurred in significant amounts. In addition, Pseudoalteromonas spp., Marinococcus sp., Halobacillus sp., Shewanella sp., Sulfitobacter sp., and bacteria of the CFB cluster were noticed. The presence of pathogenic and conditionally pathogenic bacterial species in the water and mollusks is probably the reason for the high death rate of cultivated animals at the mariculture farm.     

Biological control of tomato gray mold caused by <Emphasis Type="Italic">Botrytis cinerea</Emphasis> by using <Emphasis Type="Italic">Streptomyces</Emphasis> spp.

Streptomyces is a genus known for its ability to protect plants against many pathogens and various strains of this bacteria have been used as biological control agents. In this study, the efficacy of Streptomyces philanthi RM-1-138, S. philanthi RL-1-178, and Streptomyce mycarofaciens SS-2-243 to control various strains of Botrytis cinerea was evaluated both in vitro and in vivo. In vitro studies using confrontation tests on PDA plates indicated that the three strains of Streptomyces spp. inhibited the growth of 41 strains of B. cinerea. Volatile compounds produced by Streptomyces spp. had an influence on the growth of ten strains of B. cinerea while its culture filtrate at low concentration (diluted at 10^?3) showed a complete inhibition (100%) of spore germination of B. cinerea strain BC1. A significant protection efficacy of tomato against B. cinerea was observed on both whole plant test (57.4%) and detached leaf test (60.1%) with S. philanti RM-1-138. Moreover, this antagonistic strain had a preventive and a curative effect. These results indicated that S. philanthi RM-1-138 may have the potential to control gray mold caused by B. cinerea on tomato but further work is required to enhance its efficacy and its survival in planta.