Different microbial biofilm formation on polymethylmethacrylate (PMMA) bone cement loaded with gentamicin and vancomycin

We studied the in vitro effects of gentamicin and vancomycin alone and in combination added to polymethylmethacrylate (PMMA) cement specimens on the bacterial adhesion of multiresistant clinical isolates.The PMMA specimens (discs) loaded with gentamicin (1.9%) or vancomycin (1.9%) or with a combination of the two were placed in Mueller-Hinton Broth inoculated with bacterial strains. After incubation, bacterial growth was determined by optical density (OD₅₄₀) and sub-cultures. The biofilm PMMA-associated dye (crystal violet) was measured. Antibiotic concentrations in broth were determined by fluorescence polarisation immunoassay.All antibiotic-loaded PMMA cement specimens released high, inhibitory concentrations of gentamicin and vancomycin. However, differences in strain growth and adhesion were recorded. The clinical isolates Met-R/Gent-R CoNS showed no adhesion to gentamicin-loaded specimens for 24 h; strains with Gent-Intermediate susceptibility exhibited growth after 48 h but reduced adhesion. Some Gent-R strains exhibited growth and adhesion to antibiotic-loaded specimens similar to controls (plain discs). Only the VRSA strain (Staphylococcus aureus 5/7) and Escherichia coli were able to grow and adhere to vancomycin-loaded specimens after 24 h of incubation. The specimens loaded with the gentamicin + vancomycin combination showed a synergistic inhibitory effect against all tested strains (no bacterial growth). The degree of bacterial adhesion to PMMA cement loaded with gentamicin or vancomycin may be reduced in spite of a normal growth rate and is different for the tested strains.The effect of gentamicin and vancomycin on bacterial growth and adhesion to PMMA bone cement depends on the antibiotic concentrations, on the characteristics of each specific strain and on its ability to produce biofilm and adhere to antibiotic-loaded PMMA bone cement.     

Characterization of co-culture of Aeromonas and Pseudomonas bacterial biofilm and spoilage potential on refrigerated grass carp (Ctenopharyngodon idellus)

Aeromonas and Pseudomonas are important bacterial species involved in spoilage of refrigerated freshwater fish. In this study, 10 Aeromonas and seven Pseudomonas bacterial strains were isolated from spoiled grass carp and identified. Twelve of seventeen bacterial strains showed high potential of biofilm formation and 14 of 17 can produce extracellular protease. In order to explore the spoilage capacity of dual-species, the sterile grass carp fillets were inoculated with mono- and dual-species of Aeromonas salmonicida and Pseudomonas azotoformans strains. The results revealed significantly higher levels of the total viable count and total volatile basic nitrogen in dual-species as compared to mono-species from day 6. The higher contents of histamine, cadaverine and serious degradation in muscles tissue were also observed in dual-species after 10 days of storage. Results of in vitro experiments showed that the co-culture of A. salmonicida and P. azotoformans significantly increased the bacterial maximum growth rate, promoted the biofilm formation and improved the spoilage capacity of bacterial strains. This study has revealed that the co-culture of Aeromonas and Pseudomonas bacterial strains accelerated spoilage process of grass carp and increased biofilm formation. It indicates that the mixed-cultures of spoilage micro-organisms pose a huge threat to food industry.     

BACTERIA THAT INDUCE MORPHOGENESIS IN ULVA PERTUSA (CHLOROPHYTA) GROWN UNDER AXENIC CONDITIONS1

Marine foliaceous green macroalgae such as Ulva lose their typical morphology when cultured aseptically in defined synthetic media. However, after reinfection by certain marine bacteria (isolated from unialgal cultures of Ulva pertusa Kjellman), the organisms regain their typical foliaceous or tubular morphology. To investigate the morphogenesis (MG) induced in U. pertusa by bacteria, we isolated and identified bacteria with MG activity on U. pertusa and studied the distribution of such bacteria in seawater and on various marine macroalgae. We isolated 1555 bacterial strains from 18 species of marine macroalgae (six Chlorophyta, five Phaeophyta, and seven Rhodophyta), from seawater and from sediment collected at the beach at Omaezaki, Shizuoka Prefecture; Japan. Of these, 676 bacterial strains (43.5%) showed MG activity. They were classified into six bacterial groups, Flavobacterium, Vibrio, Pseudomonas, Deleya, Escherichia, and gram-positive cocci. These bacteria were ubiquitous among the samples and were not specific to U. pertusa. Several plant growth regulators had no MG activity. Filter-sterilized supernatants of culture media of MG-active bacteria strains did not induce MG. Cocultivation of Ulva with active bacterial strains is so far the only way to induce the MG effect, which suggests that for MG direct contact between Ulva and the bacterial strain is necessary.     

Antibacterial Co(II), Ni(II), Cu(II) and Zn(II) Complexes of Schiff bases Derived from Fluorobenzaldehyde and Triazoles

Antibacterial Schiff bases derived from 1,2,4-triazoles as well as their metal complexes incorporating cobalt(II), nickel(II), copper(II) and zinc(II) have been synthesized and characterized. Physico-chemical studies suggest that an octahedral geometry for the cobalt(II), nickel(II) and zinc(II)and square-planer geometry for the copper(II) complexes. These complexes have been screened for antibacterial activity against three Gram-positive (Staphylococcus aureus, Staphylococcus epidermidis and Bacillus subtilis) and two Gram-negative (Salmonella typhi and Pseudomonas aeruginosa) bacterial strains, and results compared with the activity of the free ligands. The metal complexes were found to be more potent against one or more bacterial strains than the free ligands.     

Production of Industrial Enzymes (Amylase,Carboxymethylcellulase and Protease) by Bacteria Isolated from Marine Sedentary Organisms

The abilities of bacteria isolated from eight marine sedentary organisms, six marine sponges (Spirastrella sp., Phyllospongia sp., Ircinia sp., Aaptos sp., Azorica sp. and Axinella sp.), one soft coral (Lobophytum sp.) and one alga (Sargassum sp.) to produce industrial enzymes (amylase, carboxymethylcellulase and protease) were examined. The mean total viable counts of the bacterial isolates ranged from 8.7 × 10⁴ to 8.4 × 10⁵ cfu/g wet weight of the organism. All eight organisms harboured amylase (0.05–0.5 IU/ml), carboxymethylcellulase (0.05–0.5 IU/ml) and protease (0.1–0.5 IU/ml) producing bacteria. Of 56 bacterial strains tested, as many as 60 to 83% of the strains produced at least one of the three enzymes, and 47% of strains were able to produce all three enzymes. High activities (> 0.5 IU/ml) of the three enzymes were recorded in bacterial strains belonging to the genera Alcaligenes and Bacillus. From the results of this study, it appears that bacteria associated with marine sedentary organisms are the novel source of industrial enzymes for possible commercial applications and may play an important role in enzyme‐catalysed organic matter cycling in marine environments.     

Isolation and characterization of novel Serratia marcescens (AY927692) for pentachlorophenol degradation from pulp and paper mill waste

Seven aerobic bacterial strains were isolated from pulp paper mill waste and screened for pentachlorophenol (PCP) tolerance on PCP containing mineral salt agar medium (MSM). The organism was characterized by 16S rDNA sequencing which showed 99.7% sequence similarity with Serratia marcescens. PCP degradation was routinely monitored with spectrophotometric analysis and further confirmed by HPLC analysis. Among seven strains, ITRC S₇ was found to degrade up to 90.33% of 1.127 mM (300 mg/l) of PCP and simultaneous release of chloride ion (2.435 mM) emphasized the bacterial dechlorination in the medium in presence of glucose as an additional carbon and energy source under optimized condition within 168 h incubation. In absence of glucose bacterium was unable to utilize PCP indicating the phenomenon of co-metabolism. Bacterium was identified as S. marcescens (AY927692), was a novel and potential aerobic bacterial strain capable of degrading PCP in axenic condition. Further, this strain may be used for bioremediation of PCP containing pulp paper mill waste in the environment.     

Evaluation of rhizosphere bacterial antagonists for their potential to bioprotect potato (Solanum tuberosum) against bacterial wilt (Ralstonia solanacearum)

Bacterial wilt (Ralstonia solanacearum) is one of the production constraints of potato (Solanum tuberosum). The intent of the study was to evaluate potential of bacterial antagonists to suppress bacterial wilt disease development and evaluate the role of the strains as plant growth-promoting rhizobacteria (PGPR) in potato. One hundred-twenty rhizosphere bacterial isolates were screened against virulent strain of Ralstonia solanacearum PPRC-Rs. After in vitro screening, six antagonistic strains (PFMRI, BS-DFS, PF9, PF20, BC, and BS-wly) with inhibition diameter >11 mm were selected and studied further in the greenhouse, in vivo. During in vivo study, the strains were evaluated for their effect in suppressing disease development in terms of area under disease progress curve (AUDPC) and increasing biomass (plant height and dry weight) of potato. Accordingly, PFMRI, BS-DFS, and PF9, significantly reduced AUDPC by 78.6, 66, and 64.3%, and wilt incidence by 82.7, 66.2, and 65.7%, respectively, compared to the control. During the sole application, the strains significantly (P < 0.0001) increased plant height by 35.6, 45.9, and 45%, and dry matter by 111, 130.4, and 129%, respectively compared to non-bacterized control. In the presence of the pathogen strain PFMRI, BS-DFS, and PF9 increased plant height by 66, 50, and 48.2%, and dry matter by 153.8, 96.8, and 92.5%, respectively compared to the pathogen treated control. Hence, the study shows that PFMRI, BS-DFS, and PF9 strains have potential use in potato bioprotection, as PGPR or in an integrated bacterial wilt management; whose effectiveness under a variety of field conditions should be investigated.     

Phylogenetic analysis and screening of antimicrobial and cytotoxic activities of moderately halophilic bacteria isolated from the Weihai Solar Saltern (China)

A total of 45 moderately halophilic bacteria was isolated from sediment and saline water collected from the Weihai Solar Saltern (China). The phylogenetic position of all the isolated strains was determined by 16S rRNA sequencing. The halophilic strains were tested for their antimicrobial activity. Cytotoxicity assay was performed to determine which of the halophilic strains could inhibit proliferation of human hepatocellular carcinoma Bel 7402 cells. Our results showed that all of the isolated 45 strains displayed moderately halophilic characteristics. Phylogenetic analysis indicated that 17 of the isolated strains were related to the phylum Firmicutes and belonged to four genera, Bacillus, Halobacillus, Planococcus and Salinicoccus. The other strains identified as genus of Halomonas belonged to phylum γ-Proteobacteria. Most of the halophilic bacterial strains showed potent activities against Gram-positive bacteria, human pathogenic fungi and plant pathogenic fungi. In addition, the crude extracts from 14 halophilic bacterial strains showed cytotoxic activity against tumor cells Bel 7402, and five of them showed remarkable activities with IC₅₀ less than 40 μg ml⁻¹. Our results suggest that the moderately halophilic bacteria may be developed as promising sources for the discovery of novel bioactive substances.     

Antibacterial activity of the leaf essential oil of <Emphasis Type="Italic">Blumea mollis</Emphasis> (D. Don) Merr.

The antibacterial activity of the leaf essential oil of Blumea mollis was assayed against 14 clinically isolated bacterial strains on Muller–Hinton Agar medium and Muller–Hinton Agar medium with 5% sheep blood. The essential oil had promising antibacterial activity against all the bacterial strains tested. The highest mean zone of inhibition and lowest values of minimum inhibitory concentration were recorded against methicillin-resistant Staphylococcus aureus followed by beta hemolytic Streptococcus pyogenes. The Gram-positive bacteria were more sensitive than Gram-negative bacteria. Among the bacterial strains tested, Psudomonas aeruginosa was resistant to the essential oil. The results of the present study suggest that the essential oil of B. mollis is one of the new medicinal resources as an antibacterial agent against the bacterial strains tested.     

Application of endophytic bacteria for the biocontrol of Rhizoctonia solani (Cantharellales: ceratobasidiaceae) damping-off disease in cotton seedlings

The antifungal potentialities of three endophytic bacterial strains, Stenotrophomonas maltophila H8 (Xanthomonadales: Xanthomonadaceae), Pseudomonas aeruginosa H40 (Pseudomonadales: Pseudomonadaceae) and Bacillus subtilis H18 (Bacillales: Bacillaceae) were evaluated against the phytopathogenic fungus Rhizoctonia solani in cotton seedlings under greenhouse conditions. The bacterial strains were applied as a soil drench or talc-based bioformulation in R. solani-infested soil and non-infested soil. Results indicated that the soil drench treatment was more efficient than talc-based bioformulation. A significant increase of seed emergence and seedling survival with a clear reduction of disease severity was achieved with the endophytic bacterial treatments. At the same time, the fresh weight, dry weight, shoot length and root length of the treated plants were markedly enhanced. Moreover, there was an apparent induction of the antioxidant enzymes (peroxidase, polyphenol oxidase and catalase) of the treated seedlings. Gas chromatography–mass spectrometry revealed the presence of various bioactive compounds in the bacterial supernatant. The antagonistic activity of the bacterial strains against R. solani was attributed to their capability to produce a broad spectrum of antifungal compounds in addition to bioactive molecules that can trigger the systemic resistance in the infected seedlings.     

In-vitro antibacterial and cytotoxic activity of cobalt (ii), copper (ii), nickel (ii) and zinc (ii) complexes of the antibiotic drug cephalothin (Keflin)

Keflin (kefl) interacts with Co(II), Cu(II), Ni(II) and Zn(II) metal ions leading to complexes of the type M(kefl)₂Cl₂ and M(kefl)Cl₂, which have been characterized by physicochemical and spectroscopic methods. Magnetic moment, IR, electronic spectral and elemental analyses data suggest that keflin behaves tridentately forming octahedral or trigonal bipyramidal complexes with the metal ions mentioned above. The new compounds have been screened in-vitro for antibacterial and cytotoxic activity against Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa, Salmonella typhi, Shigella dysentriae, Bacillus cereus, Corynebacterium diphtheriae, Staphylococcus aureus and Streptococcus pyogenes bacterial strains. Compounds, 4 and 8 showed promising activity (90%) against seven, compound 6 showed significant activity (52%) against four and, compounds 1 and 5 showed activity (40%) against three test bacterial strains at concentration of 10 μM.     

Bacterial Reduction of Cr(VI) and Fe(III) in In Vitro Conditions

ABSTRACT Chemical reduction of Cr(VI) can be a strategy to detoxify toxic metals in oxidized states, whereas reduction of Fe(III) could enhance the availability of Fe in the form of Fe(II) to boost plant growth. However, it creates another problem of chemical sludge disposal. Hence, microbial conversion of Cr(VI) to Cr(III) and Fe(III) to Fe(II) is preferred over the chemical method. Out of 11 bacterial strains isolated from the rhizospheric zone of Typha latifolia growing on fly ash dump sites, four isolates were selected for the reduction of Cr(VI) and Fe(III) and were identified as Micrococcus roseus NBRFT2 (MTCC 9018), Bacillus endophyticus NBRFT4 (MTCC 9021), Paenibacillus macerans NBRFT5 (MTCC 8912), and Bacillus pumilus NBRFT9 (MTCC 8913). These strains were individually tested for survival at different concentrations of Cr(VI) and Fe(III), pH, and temperature, and then, their ability for reduction of both metals was evaluated at optimum pH 8.0 and temperature 35°C. The results indicated that NBRFT5 was able to reduce the maximum amount, 99% Cr(VI) and 98% Fe(III). Other strains also reduced these metals to different levels, but less than NBRFT5. Hence, these strains may be used for decontamination of metal-contaminated sites, particularly with Cr(VI) and Fe(III) through the reduction process.     

Comparative analysis of oyster (Crassostrea gigas) immune responses under challenge by different Vibrio strains and conditions

Screening a virulent Vibrio strain to study the interactions between bacterial infection and host immune defence is important to resolve large-scale summer oyster mortalities. Using adductor muscle injection we tested the oyster responses to phosphate-buffered saline (PBS), lipopolysaccharide (LPS), or Vibrio strains. Mortalities and stress gene expressions were used to characterize oyster immune responses. At 5 days post injection, the cumulative mortality rates in PBS, LPS, Vibrio tubiashii, Vibrio anguillarum, Vibrio alginolyticus and Vibrio aestuarianus groups were 8%, 16%, 36%, 40%, 40% and 76%, respectively, indicating that V. aestuarianus induced the highest death rate. Two-factor analyses of variance revealed that expression of SOD, CAT, GPX and HSP70 was influenced by bacterial injections in a time-dependent manner. The expression of all genes increased and reached their peak 3 or 12 h after bacterial injection and then decreased. These genes could be applied as immune responsive biomarkers to monitor early changes in oysters in response to bacterial infection. The greatest changes were observed in the V. aestuarianus-injected group, which may indicate that V. aestuarianus could be used as one of the more virulent strains for experimental infections.     

Culturable bacterial diversity associated with cysts of <Emphasis Type="Italic">Eurhizococcus brasiliensis</Emphasis> (Hempel) (Hemiptera: Margarodidae)

The ground-pearl Eurhizococcus brasiliensis is an important insect pest of grapes. Nowadays, its biology is still barely known and studies related to its secondary symbionts are virtually non-existent. Our main goal was to evaluate the bacterial diversity associated with cysts of E. brasiliensis using a culture-dependent approach. Six different isolation media were used and shown to be suitable for culturing bacteria associated with E. brasiliensis. A total of 39 bacteria strains were isolated and classified into 10 different morphotypes. The ISP-4 medium was the most suitable for bacteria isolation from cysts of the ground-pearl, providing the highest number of morphotypes. 16S rDNA gene analysis demonstrated a high diversity of bacteria associated with cysts, with six Pseudomonas chlororaphis isolates (the most predominant morphotype) and three different morphotypes of Bacillus spp. as the most representative genera. The phylogenetic analysis showed close affinity between the isolated morphotypes and bacterial strains usually isolated from plant and soil samples. Sphingopyxis and Stenotrophomonas were isolated for the first time from arthropods. Although it was not possible to determine the primary source of infection by these bacteria, our data suggests that these microorganisms may be transitory and acquired from the environment.     

Helicobacter pylori outer inflammatory protein A (OipA) suppresses apoptosis of AGS gastric cells in vitro

Outer inflammatory protein A (OipA) is an important virulence factor associated with gastric cancer and ulcer development; however, the results have not been well established and turned out to be controversial. This study aims to elucidate the role of OipA in Helicobacter pylori infection using clinical strains harbouring oipA “on” and “off” motifs. Proteomics analysis was performed on AGS cell pre‐infection and postinfection with H. pylori oipA “on” and “off” strains, using liquid chromatography/mass spectrometry. AGS apoptosis and cell cycle assays were performed. Moreover, expression of vacuolating cytotoxin A (VacA) was screened using Western blotting. AGS proteins that have been suggested previously to play a role or associated with gastric disease were down‐regulated postinfection with oipA “off” strains comparing to oipA “on” strains. Furthermore, oipA “off” and ΔoipA cause higher level of AGS cells apoptosis and G0/G1 cell‐cycle arrest than oipA “on” strains. Interestingly, deletion of oipA increased bacterial VacA production. The capability of H. pylori to induce apoptosis and suppress expression of proteins having roles in human disease in the absence of oipA suggests that strains not expressing OipA may be less virulent or may even be protective against carcinogenesis compared those expressing OipA. This potentially explains the higher incidence of gastric cancer in East Asia where oipA “on” strains predominates.     

THE TOXIC DINOFLAGELLATE GYMNODINIUM CATENATUM (DINOPHYCEAE) REQUIRES MARINE BACTERIA FOR GROWTH1

Interactions with the bacterial community are increasingly considered to have a significant influence on marine phytoplankton populations. Here we used a simplified dinoflagellate‐bacterium experimental culture model to conclusively demonstrate that the toxic dinoflagellate Gymnodinium catenatum H. W. Graham requires growth‐stimulatory marine bacteria for postgermination survival and growth, from the point of resting cyst germination through to vegetative growth at bloom concentrations (10³ cells · mL^?1). Cysts of G. catenatum were germinated and grown in unibacterial coculture with antibiotic‐resistant or antibiotic‐sensitive Marinobacter sp. DG879 or Brachybacterium sp., and with mixtures of these two bacteria. Addition of antibiotics to cultures grown with antibiotic‐sensitive strains of bacteria resulted in death of the dinoflagellate culture, whereas cultures grown with antibiotic‐resistant bacteria survived antibiotic addition and continued to grow beyond the 21 d experiment. Removal of either bacterial type from mixed‐bacterial dinoflagellate cultures (using an antibiotic) resulted in cessation of dinoflagellate growth until bacterial concentration recovered to preaddition concentrations, suggesting that the bacterial growth factors are used for dinoflagellate growth or are labile. Examination of published reports of axenic dinoflagellate culture indicate that a requirement for bacteria is not universal among dinoflagellates, but rather that species may vary in their relative reliance on, and relationship with, the bacterial community. The experimental model approach described here solves a number of inherent and logical problems plaguing studies of algal‐bacterium interactions and provides a flexible and tractable tool that can be extended to examine bacterial interactions with other phytoplankton species.     

Two strains of Pseudomonas fluorescens bacteria differentially affect survivorship of waxworm (Galleria mellonella) larvae exposed to an arthropod fungal pathogen,Beauveria bassiana

Two strains of Pseudomonas fluorescens were found contaminating a biopesticide used in a previous study against Varroa destructor infestations in honey bee hives. In that study, the biopesticide, a formulation of a fungal pathogen of arthropods, Beauveria bassiana, failed to have any negative impact on the mite infestation despite successful results in previous studies using uncontaminated batches of the same biopesticide. The objective of the present research was to determine whether the bacteria may have interfered with the infectivity and/or virulence of B. bassiana in a simplified system; positive results in that system would then provide a rationale for further work under more complex conditions. Galleria mellonella late instar larvae treated topically with both a bacterial suspension of 6.8 to 7.0×10⁷ cfu/ml and a fungal suspension of 2.5×10⁷ or 2.5×10⁸ B. bassiana conidia/ml showed, in the case of one of the bacterial strains, significantly increased survivorship compared to larvae treated with just the B. bassiana suspension. When larvae were immersed in a bacterial suspension prior to application of B. bassiana suspension using a spray tower, a significant positive effect of the same P. fluorescens strain on larval survivorship was observed at 2.5×10⁸ conidia/ml. Neither the bacterial suspensions alone nor blank control solutions had any effect on larval survivorship. These results show that an interaction between the bacteria and the pathogen may explain some of the results from the prior field trial.     

Screening of yeasts for cell-free production of (R)-phenylacetylcarbinol

105 yeast strains from 10 genera and 40 species were evaluated for cell-free production of (R)-phenylacetylcarbinol (PAC), the chiral precursor in the manufacture of the pharmaceuticals ephedrine and pseudoephedrine. Carboligase activity of pyruvate decarboxylase (PDC), forming PAC from benzaldehyde and pyruvate, was found in extracts of 98 strains. PAC was not formed from benzaldehyde and acetaldehyde, an activity of bacterial PDCs from Zymomonas mobilis and Zymobacter palmae. Two interesting groups of candidates were identified in the yeast screening: carboligase activities of Schizosaccharomyces pombe PDCs were very low but showed best resistance to pre-incubation with acetaldehyde and benzaldehyde; and highest carboligase activities combined with medium resistance were found in strains of Candida utilis, C. tropicalis and C. albicans.     

Immunofluorescence detection of the denitrifying strain Pseudomonas stutzeri (ATCC 14405) in seawater and intertidal sediment environments

A strain-specific immunofluorescence assay for enumeration of a marine denitrifying bacterium was developed and applied in the marine environment. The polyclonal antiserum for Pseudomonas stutzeri (ATCC 14405) did not react with other pseudomonads, other heterotrophs, or autotrophic nitrifying strains. The abundance of P. stutzeri in the shallow water column of Monterey Bay was less than 0.1% of the total bacterial abundance and decreased with depth, whereas the total bacterial abundance was variable and nearly constant with depth. P. stutzeri was also detected in the sediments of a microbial mat from Tomales Bay. The relatively low contribution of P. stutzeri to the total bacterial abundance in both environments implies that it is not a major component of the heterotrophic assemblage. This conclusion appears to hold for most other strains for which specific assays have been applied in the marine environment. The isolation of several different denitrifying strains from local marine environments implies that the culturable population is quite diverse, even in the absence of different selective enrichment media. Thus, strain specific immunofluorescence is of limited use in quantifying functional groups of bacteria. Conversely, they provide specific information on the diversity of natural populations and their relation to culturable strains.Offprint requests to: B. Ward     

Identification of a Vibrio strain producing antimicrobial agents in the excretory organs of Nautilus pompilius (Cephalopoda: Nautiloidea)

The aim of the present study was to identify and characterize bacteria producing antimicrobial compounds in the excretory organs of Nautilus pompilius. Culture-dependent and culture-independent complementary approaches were used for bacterial identification such as: culture on selective media, Gram staining, CARD-FISH, direct DNA extraction from host tissue, PCR amplification and sequencing of the bacterial 16S rRNA gene. Results show presence of three bacterial groups: γ-Proteobacteria with three clusters (Pseudomonadales, Vibrionales, Alteromonadales), β-Proteobacteria and spirochetes. In order to screen for active strains, antimicrobial activity was tested by diffusion agar assay against Micrococcus luteus, Escherichia coli, Vibrio harveyi, and Candida albicans. Nautilus isolates showed antimicrobial activities against both Gram-positive and Gram-negative reference strains. Most of the active strains were phylogenetically related to environmental Vibrionaceae. These strains were always abundant in N. pompilius PA but were absent from Nautilus macromphalus from other geographical areas. Therefore, we suggest that antimicrobial active Vibrionaceae infect N. pompilius by environmental transmission.