Halide peroxidase in tissues that interact with bacteria in the host squid Euprymna scolopes

An enzyme with similarities to myeloperoxidase, the antimicrobial halide peroxidase in mammalian neutrophils, occurs abundantly in the light organ tissue of Euprymna scolopes, a squid that maintains a beneficial association with the luminous bacterium Vibrio fischeri. Using three independent assays typically applied to the analysis of halide peroxidase enzymes, we directly compared the activity of the squid enzyme with that of human myeloperoxidase. One of these methods, the diethanolamine assay, confirmed that the squid peroxidase requires halide ions for its activity. The identification of a halide peroxidase in a cooperative bacterial association suggested that this type of enzyme can function not only to control pathogens, but also to modulate the interactions of host animals with their beneficial partners. To determine whether the squid peroxidase functions under both circumstances, we examined its distribution in a variety of host tissues, including those that typically interact with bacteria and those that do not. Tissues interacting with bacteria included those that have specific cooperative associations with bacteria (i.e., the light organ and accessory nidamental gland) and those that have transient nonspecific interactions with bacteria (i.e., the gills, which clear the cephalopod circulatory system of invading microorganisms). These bacteria-associated tissues were compared with the eye, digestive gland, white body, and ink-producing tissues, which do not typically interact directly with bacteria. Peroxidase enzyme assays, immunocytochemical localization, and DNA-RNA hybridizations showed that the halide-dependent peroxidase is consistently expressed in high concentration in tissues that interact bacteria. Elevated levels of the peroxidase were also found in the ink-producing tissues, which are known to have enzymatic pathways associated with antimicrobial activity. Taken together, these data suggest that the host uses a common biochemical response to the variety of types of associations that it forms with microorganisms.     

Genome sequence of the probiotic bacterium Sporolactobacillus vineae SL153T

The novel Sporolactobacillus vineae SL153(T) strain has excellent intestinal adherence and growth inhibitory effect on pathogenic microorganisms, including Vibrio genus microorganisms, and therefore can be effectively used for the prevention and treatment of disease caused by pathogenic microorganisms. Here, we first report the draft genome sequence of a novel species in the genus Sporolactobacillus.     

Stable isotopes as tracers of trophic interactions in marine mutualistic symbioses

Mutualistic nutritional symbioses are widespread in marine ecosystems. They involve the association of a host organism (algae, protists, or marine invertebrates) with symbiotic microorganisms, such as bacteria, cyanobacteria, or dinoflagellates. Nutritional interactions between the partners are difficult to identify in symbioses because they only occur in intact associations. Stable isotope analysis (SIA) has proven to be a useful tool to highlight original nutrient sources and to trace nutrients acquired by and exchanged between the different partners of the association. However, although SIA has been extensively applied to study different marine symbiotic associations, there is no review taking into account of the different types of symbiotic associations, how they have been studied via SIA, methodological issues common among symbiotic associations, and solutions that can be transferred from one type of association with another. The present review aims to fill such gaps in the scientific literature by summarizing the current knowledge of how isotopes have been applied to key marine symbioses to unravel nutrient exchanges between partners, and by describing the difficulties in interpreting the isotopic signal. This review also focuses on the use of compound‐specific stable isotope analysis and on statistical advances to analyze stable isotope data. It also highlights the knowledge gaps that would benefit from future research.     

Oxygen-utilizing reactions and symbiotic colonization of the squid light organ by Vibrio fischeri.

A major goal in microbiology is to understand the processes by which bacteria successfully colonize host tissue. Although a wealth of studies focusing on pathogenic microorganisms has revealed much about the rare interactions that result in disease, far less is known about the regulation of the ubiquitous, long-term, cooperative associations of bacteria with their animal hosts.     

海洋固氮生物多样性及其对海洋生产力的氮、碳贡献

固氮生物及其共生体在自然生态系统的氮素供给和生产力的持续发展中有重要作用。是贫营养盐海域和海洋生态系统中新生产力的主要贡献者。就海洋固氮生物研究的历史和现状,以及在世界不同海域和海洋生态环境条件下,海洋固氮生物的物种多样性和在海洋生态环境中的生存方式、共生互作关系多样化,作较全面的概述。综合分析了海洋固氮生物的固氮能力（活性）、在光合固碳中的作用和对初级生产力的贡献。介绍了固氮生物在海洋生物链和海洋不同生态系统和生物群落中的作用及其对环境的影响的最新研究概况。     

海洋养殖环境微生物多样性及其作用

微生物是海洋养殖环境和海洋牧场生态系统中的关键环境因素之一，在牧场环境的物质循环、水质保障、饵料供应、水产健康等方面起重要作用，是营造海洋牧场环境且影响其可持续发展的重要因素。主要围绕海洋养殖环境中的病原微生物以及有益微生物两方面，概述了我国海洋养殖环境常见的病原性细菌、真菌、病毒以及相应的病害防治措施等，并介绍了海洋养殖环境中能够改善海水养殖环境，包括降低水体氨氮含量、降解水体硫化物、抑制赤潮藻类的生长等以及直接促进海水养殖生物生长的有益功能微生物类群。目前，我国关于海洋牧场环境营造过程中微生物的研究仍很少，而与海洋牧场环境有一定相似性的海水养殖环境中微生物的研究较为成熟，其对海洋牧场环境健康营造有很好的启示。海洋水产养殖环境中微生物的新功能菌种的发现、作用机制以及功能菌应用等方面的深入研究将会对我国大力推进海洋牧场建设起到重要的作用。     

Construction of a Vibrio splendidus mutant lacking the metalloprotease gene vsm by use of a novel counterselectable suicide vector

Vibrio splendidus is a dominant culturable Vibrio in seawater, and strains related to this species are also associated with mortality in a variety of marine animals. The determinants encoding the pathogenic properties of these strains are still poorly understood; however, the recent sequencing of the genome of V. splendidus LGP32, an oyster pathogen, provides an opportunity to decipher the basis of the virulence properties by disruption of candidate genes. We developed a novel suicide vector based on the pir-dependent R6K replicative origin, which potentially can be transferred by RP4-based conjugation to any Vibrio strain and which also carries the plasmid F toxin ccdB gene under control of the PBAD promoter. We demonstrated that this genetic system allows efficient counterselection of integrated plasmids in the presence of arabinose in both V. splendidus and Vibrio cholerae and thus permits efficient markerless allelic replacement in these species. We used this technique to construct several mutants of V. splendidus LGP32, including a derivative with a secreted metalloprotease gene, vsm, deleted. We found that this gene is essential for LGP32 extracellular product toxicity when the extracellular products are injected into oysters but is not necessary for virulence of bacteria in the oyster infection model when bacteria are injected.     

Denaturing gradient gel electrophoresis analyses of the vertical distribution and diversity of Vibrio spp. populations in the Cariaco Basin

The Cariaco system is the second largest permanently anoxic marine water body in the world. Its water column is characterized by a pronounced vertical layering of microbial communities. The goal of our study was to investigate the vertical distribution and diversity of Vibrio spp. present in the Cariaco Basin waters using denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA fragments. Representatives of the Vibrio genus were detected by nested and direct PCR in seawater at 10 depths. Sequence analyses of 55 DGGE bands revealed that only 11 different operational taxonomic units (OTU) are identified as Vibrio species. Between one and five OTUs were detected at each depth and the most common OTUs were OTU 1 and OTU 2, which phylogenetically clustered with Vibrio chagasii and Vibrio fortis, respectively. OTUs 3 and 4 were only found in the anoxic zone and were identified as Vibrio orientalis and Vibrio neptunius, respectively. Several Vibrio species detected are potentially pathogenic to human, prawns and corals such as Vibrio parahaemolyticus, Vibrio fischeri and Vibrio shilonii. In the Cariaco Basin, different Vibrio species were found to be specific to specific depths strata, suggesting that this genus is a natural component of the microbial communities in this marine redox environment.     

Insects as alternative hosts for phytopathogenic bacteria

Phytopathogens have evolved specialized pathogenicity determinants that enable them to colonize their specific plant hosts and cause disease, but their intimate associations with plants also predispose them to frequent encounters with herbivorous insects, providing these phytopathogens with ample opportunity to colonize and eventually evolve alternative associations with insects. Decades of research have revealed that these associations have resulted in the formation of bacterial-vector relationships, in which the insect mediates dissemination of the plant pathogen. Emerging research, however, has highlighted the ability of plant pathogenic bacteria to use insects as alternative hosts, exploiting them as they would their primary plant host. The identification of specific bacterial genetic determinants that mediate the interaction between bacterium and insect suggests that these interactions are not incidental, but have likely arisen following the repeated association of microorganisms with particular insects over evolutionary time. This review will address the biology and ecology of phytopathogenic bacteria that interact with insects, including the traditional role of insects as vectors, as well as the newly emerging paradigm of insects serving as alternative primary hosts. Also discussed is one case where an insect serves as both host and vector, which may represent a transitionary stage in the evolution of insect-phytopathogen associations.     

MALDI-TOF MS与16S rDNA方法对 弧菌科微生物的鉴定与系统分类学分析

目的比较基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)与16S rDNA方法对弧菌科微生物的鉴定及系统分类学分析能力。方法对19株弧菌科微生物,采用MALDI-TOF MS进行蛋白质图谱采集,通过对特征峰的分析,实现对微生物的鉴定和系统分类学分析;同时对19株微生物进行16S rDNA测序,用邻接法对16S rDNA序列进行鉴定和系统分类学分析,比较两种方法在弧菌科微生物鉴定和系统分类学分析中的异同。结果两种方法对19株弧菌科微生物的种属鉴定结果一致。系统分类学分析中,多株同种属的弧菌的两种方法分析结果一致,但对拟态弧菌和霍乱弧菌在树状图中的位置和亲缘关系,两种方法差异较大。结论 MALDI-TOF MS与16S rDNA均能够快速准确地鉴定弧菌科微生物,但利用MALDI-TOF MS进行系统分类学分析还有待数据库的扩大及算法的优化。     

Inhibition of <Emphasis Type="Italic">Vibrio</Emphasis> biofilm formation by a marine actinomycete strain A66

China remains by far the largest aquaculture producer in the world. However, biofilms formed by pathogenic Vibrio strains pose serious problems to marine aquaculture. To provide a strategy for biofilm prevention, control, and eradication, extracts from 88 marine actinomycetes were screened. Thirty-five inhibited the biofilm formation of Vibrio harveyi, Vibrio vulnificus, and Vibrio anguillarum at a concentration of 2.5% (v/v). Thirty-three of the actinomycete extracts dispersed the mature biofilm. Six extracts inhibited the quorum-sensing system of V. harveyi by attenuating the signal molecules N-acylated homoserine lactones’ activity. Strain A66, which was identified as Steptomyces albus, both attenuated the biofilms and inhibited their quorum-sensing system. It is suggested that strain A66 is a promising candidate to be used in future marine aquaculture.     

Screening and evaluation of probiotics as a biocontrol agent against pathogenic Vibrios in marine aquaculture

AIMS: The present work aims at finding potential probionts from marine sources as a biocontrol agent against pathogenic Vibrio species in shrimp larval culture. METHODS AND RESULTS: A total of 109 bacterial strains were isolated from seawater, sediment and marine fish-gut samples, and were screened for their antagonistic activity against Vibrio species. Three strains (Q, Q1 and M) isolated from the marine sediment were found antagonistic against Vibrio strains. Based on 16S ribosomal DNA gene sequence analysis, the strain Q was identified as Paenibacillus spp. (EF012164); Q1 as Bacillus cereus (DQ915582); and the M as Paenibacillus polymyxa (DQ915580). Further, the two bacterial species, Paenibacillus spp. and B. cereus were challenged separately at two different concentrations of 10(4) and 10(5) CFU ml(-1) for probiotic activity in the postlarvae of Penaeus monodon against pathogenic Vibrio harveyi and Vibrio spp. CONCLUSIONS: The present study identified the probiotic activity of Paenibacillus spp., B. cereus and Pa. polymyxa against the pathogenic Vibrios in the postlarvae of P. monodon. SIGNIFICANCE AND IMPACT OF THE STUDY: In vivo study reveals that the marine bacterial species can be used as probionts against pathogenic Vibrios in shrimp larval culture practices.     

New selective and differential medium for Vibrio cholerae and Vibrio vulnificus

Thiosulfate-citrate-bile salts-sucrose agar has been routinely used for the isolation of pathogenic vibrios, although its selectivity for Vibrio cholerae and Vibrio vulnificus is inadequate. Therefore, a new plating medium, cellobiose-polymyxin B-colistin agar, was developed for the isolation of these two species. Cellobiose-polymyxin B-colistin agar demonstrated a significant advantage over other media designed for the isolation or differentiation of vibrios: of both the 136 strains representing 19 Vibrio species and the marine isolates of the genera Pseudomonas, Flavobacterium, and Photobacterium, only V. vulnificus and V. cholerae were able to grow. Furthermore, the fermentation of cellobiose by V. vulnificus allowed for the easy differentiation of these two species. This medium offers significant potential as a selective and differential medium for these two pathogenic vibrios.     

New selective and differential medium for Vibrio cholerae and Vibrio vulnificus.

Thiosulfate-citrate-bile salts-sucrose agar has been routinely used for the isolation of pathogenic vibrios, although its selectivity for Vibrio cholerae and Vibrio vulnificus is inadequate. Therefore, a new plating medium, cellobiose-polymyxin B-colistin agar, was developed for the isolation of these two species. Cellobiose-polymyxin B-colistin agar demonstrated a significant advantage over other media designed for the isolation or differentiation of vibrios: of both the 136 strains representing 19 Vibrio species and the marine isolates of the genera Pseudomonas, Flavobacterium, and Photobacterium, only V. vulnificus and V. cholerae were able to grow. Furthermore, the fermentation of cellobiose by V. vulnificus allowed for the easy differentiation of these two species. This medium offers significant potential as a selective and differential medium for these two pathogenic vibrios.     

Kinetics of adhesion of selected fish-pathogenic Vibrio strains of skin mucus of gilt-head sea bream (Sparus aurata L.).

The kinetics of adhesion of Vibrio strains isolated from diseased fish to skin mucus of gilt-head sea bream was studied. A modified Langmuir adsorption isotherm was calculated, and the results obtained indicate that the strains tested (Vibrio alginolyticus DP1HE4 and Vibrio anguillarum-like DC12R8 and DC12R9) showed a saturation kinetics except for V. alginolyticus (CAN), which showed a proportional adsorption kinetics. The adhesive capability for skin mucus does not seem to be an essential virulence factor of pathogenic strains of Vibrio, since this specific interaction depended on several environmental factors, temperature and salinity being the most important. However, the absence of an inhibitory effect of mucus on the pathogenic microorganisms, and the capability of the Vibrio strains to utilize mucus as a carbon source, could favor their settlement on the skin with a potential for infection of cultured, stressed fish.     

Co‐evolution of marine worms and their chemoautotrophic bacterial symbionts: unexpected host switches explained by ecological fitting?

Mutualistic associations of bacteria and invertebrates are widespread and encompass an enormous diversity on the side of both partners. The advantages gained from the symbiosis favour reciprocal adaptations that increase the stability of the association and can lead to codiversification of symbiont and host. While numerous examples of a strictly vertical transfer of the symbionts from parent to offspring among intracellular associations abound, little is known about the fidelity of the partners in extracellular associations, where symbionts colonize the surface or body cavity of their host. In this issue of Molecular Ecology, Zimmermann et al. ( 2016 ) investigated the evolutionary history of the symbiotic association between a monophyletic clade of sulphur‐oxidizing Gammaproteobacteria and two distantly related lineages of marine worms (nematodes and annelids). The study supports the surprising conclusion that partner fidelity does not necessarily increase with the intimacy of the association. Ectosymbionts on the cuticle of the nematodes seem to be cospeciating with their hosts, whereas endosymbionts housed in the body cavity of the annelids must have originated multiple times, probably by host switching, from ectosymbionts of sympatric nematodes. This excellent case study on the evolutionary history of invertebrate–microbe interactions supports the emerging concept that the co‐evolutionary processes shaping such mutualistic symbioses include both codiversification and ecological fitting.     

Chemical defense in the egg masses of benthic invertebrates: an assessment of antibacterial activity in 39 mollusks and 4 polychaetes.

Many marine invertebrates deposit benthic egg masses that are potentially vulnerable to microbial infection. To help counter this threat these species may have evolved some form of chemical protection for their encapsulated embryos. In this study the egg masses from 7 marine mollusks were tested for antibacterial activity against 4 marine pathogens: Enterococcus sericolicida, Vibrio anguillarum, Vibrio alginolyticus, and Vibrio harveyi. Extracts from all of these egg masses were found to inhibit the growth of at least 1 marine bacterium at concentrations that approximate the natural concentration of extract in the egg masses. The egg masses of 39 mollusks and 4 polychaetes were then tested for antibacterial activity against 3 human pathogenic bacteria; Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa. Activity was detected in the egg masses from 34 species, including 2 polychaetes and mollusks from two classes and 18 families. Antibacterial activity in molluskan egg masses was found to extend across the marine, estuarine, freshwater, and terrestrial environments. Both gelatinous egg masses and tough egg capsules were found to inhibit microbial growth, suggesting that physical protection alone may not be sufficient to protect the eggs. Antimicrobial activity was observed in the fresh egg masses but not in the well-developed egg masses of a subset of species. The results of this study indicate that a wide range of invertebrates use chemical defense to protect their early stage embryos against bacterial infection.     

Negotiations between animals and bacteria: the 'diplomacy' of the squid-vibrio symbiosis

A shared characteristic among animals is their propensity to form stable, beneficial relationships with prokaryotes. Usually these associations occur in the form of consortia, i.e. a diverse assemblage of bacteria interacting with a single animal host. These complex communities, while common, have been difficult to characterize. The two-partner symbiosis between the squid Euprymna scolopes and the marine luminous bacterium Vibrio fischeri offers the opportunity to study the interaction between animal and bacterial cells, because both partners can be cultured in the laboratory and the symbiosis can be manipulated experimentally. This system is being used to characterize the mechanisms by which animals establish, develop and maintain stable alliances with bacteria. This review summarizes the progress to date on the development of this model.     

The Leopoldina international symposium on parasitism,commensalism and symbiosis--common themes,different outcome

The development of new methods, including genomics, which can even be applied to unculturable microorganisms, has significantly increased our knowledge about bacterial pathogenesis and symbiosis and, in consequence, is profoundly modifying our views on the evolution and the genetic and physiological basis of bacteria-host interactions. The presentations at this symposium revealed conceptual links between bacterial pathogenesis and symbiosis. The close co-operation of experts in both fields will result in significant synergy and new insights into basic mechanisms of bacteria-host interactions and their evolution. The meeting provided fascinating news about the genetic and metabolic consequences that the change in their lifestyle had for bacteria that developed from free-living to permanent host-associated organisms exemplified by intracellular pathogens or symbionts. In addition, surprising similarities but also striking differences between the strategies involved in the establishment of a symbiotic versus a parasitic lifestyle can be noted. In the long run, the characterization of such differences might lead to lifestyle prediction or to an evaluation of the pathogenic potential of newly isolated bacteria via the definition of genetic and/or metabolic signatures characteristic for pathogenic or symbiotic organisms. Moreover, it is expected that these investigations will lead to new strategies for the treatment or prevention of bacterial infections, or the avoidance of pathogen transmission.