Bacterial Colonization of Particles: Growth and Interactions

Marine particles in the ocean are exposed to diverse bacterial communities, and colonization and growth of attached bacteria are important processes in the degradation and transformation of the particles. In an earlier study, we showed that the initial colonization of model particles by individual bacterial strains isolated from marine aggregates was a function of attachment and detachment. In the present study, we have investigated how this colonization process was further affected by growth and interspecific interactions among the bacteria. Long-term incubation experiments showed that growth dominated over attachment and detachment after a few hours in controlling the bacterial population density on agar particles. In the absence of grazing mortality, this growth led to an equilibrium population density consistent with the theoretical limit due to oxygen diffusion. Interspecific interaction experiments showed that the presence of some bacterial strains (“residents”) on the agar particles either increased or decreased the colonization rate of other strains (“newcomers”). Comparison between an antibiotic-producing strain and its antibiotic-free mutant showed no inhibitory effect on the newcomers due to antibiotic production. On the contrary, hydrolytic activity of the antibiotic-producing strain appeared to benefit the newcomers and enhance their colonization rate. These results show that growth- and species-specific interactions have to be taken into account to adequately describe bacterial colonization of marine particles. Changes in colonization pattern due to such small-scale processes may have profound effects on the transformation and fluxes of particulate matter in the ocean.     

Iron effects on colonization behavior, motility, and enzymatic activity of marine bacteria

Iron availability in the ocean has been shown to affect the growth and production of phytoplankton and free-living bacteria. A large fraction of marine bacteria are specialized in colonizing and living on particles and aggregates, but the effects of iron limitation on these bacteria are not fully known. We conducted laboratory experiments to study the effects of iron availability on particle colonization behavior, motility, and enzymatic activities of 4 strains of marine bacteria. Iron depletion reduced the bacterial particle colonization rate by 1.7%-43.1%, which could be attributed to reduced swimming speeds in 2 of the 4 strains. Protease activity was not affected by iron availability. However, attached bacteria did show higher protease activities than their free counterparts. Our results suggest that iron limitation in the ocean could in some cases reduce bacteria-particle interactions by reducing bacterial motility and colonization rate.     

In vitro suppression of fungi caused by combinations of apparently non-antagonistic soil bacteria

We hypothesized that apparently non-antagonistic soil bacteria may contribute to suppression of fungi during competitive interactions with other bacteria. Four soil bacteria (Brevundimonas sp., Luteibacter sp., Pedobacter sp. and Pseudomonas sp.) that exhibited little or no visible antifungal activity on different agar media were prescribed. Single and mixed strains of these species were tested for antagonism on a nutrient-poor agar medium against the plant pathogenic fungi Fusarium culmorum and Rhizoctonia solani and the saprotrophic fungus Trichoderma harzianum. Single bacterial strains caused little to moderate growth reduction of fungi (quantified as ergosterol), most probably due to nutrient withdrawal from the media. Growth reduction of fungi by the bacterial mixture was much stronger than that by the single strains. This appeared to be mostly due to competitive interactions between the Pseudomonas and Pedobacter strains. We argue that cohabitation of these strains triggered antibiotic production via interspecific interactions and that the growth reduction of fungi was a side-effect caused by the sensitivity of the fungi to bacterial secondary metabolites. Induction of gliding behavior in the Pedobacter strain by other strains was also observed. Our results indicate that apparently non-antagonistic soil bacteria may be important contributors to soil suppressiveness and fungistasis when in a community context.     

Dynamics of microbial communities on marine snow aggregates: colonization,growth, detachment,and grazing mortality of attached bacteria

We studied the dynamics of microbial communities attached to model aggregates (4-mm-diameter agar spheres) and the component processes of colonization, detachment, growth, and grazing mortality. Agar spheres incubated in raw seawater were rapidly colonized by bacteria, followed by flagellates and ciliates. Colonization can be described as a diffusion process, and encounter volume rates were estimated at about 0.01 and 0.1 cm(3) h(-1) for bacteria and flagellates, respectively. After initial colonization, the abundances of flagellates and ciliates remained approximately constant at 10(3) to 10(4) and approximately 10(2) cells sphere(-1), respectively, whereas bacterial populations increased at a declining rate to >10(7) cells sphere(-1). Attached microorganisms initially detached at high specific rates of approximately 10(-2) min(-1), but the bacteria gradually became irreversibly attached to the spheres. Bacterial growth (0 to 2 day(-1)) was density dependent and declined hyperbolically when cell density exceeded a threshold. Bacterivorous flagellates grazed on the sphere surface at an average saturated rate of 15 bacteria flagellate(-1) h(-1). At low bacterial densities, the flagellate surface clearance rate was approximately 5 x 10(-7) cm(2) min(-1), but it declined hyperbolically with increasing bacterial density. Using the experimentally estimated process rates and integrating the component processes in a simple model reproduces the main features of the observed microbial population dynamics. Differences between observed and predicted population dynamics suggest, however, that other factors, e.g., antagonistic interactions between bacteria, are of importance in shaping marine snow microbial communities.     

Masking of antibiotic-resistance upon recovery of endophytic bacteria

During studies on internal plant colonization by rhizosphere bacteria and endophytic bacteria over several years, we frequently observed lack of growth of rifampicin-resistant mutants (rif+) on tryptic soy agar amended with rifampicin (RTSA). Following seed treatment of cucumber with 6 species of rif+ rhizosphere bacteria in one experiment, all strains were recoverable on RTSA when external root colonization was monitored. Following trituration of surface-disinfested roots, only one strain grew directly on RTSA; however colonies isolated on tryptic soy agar (TSA) grew within 18 h after transfer to RTSA. We term this temporary loss of the antibiotic-resistant phenotype ‘antibiotic masking’. Antibiotic masking was also observed with isolation of 7 rif+ endophytic bacterial strains from inside stems of cotton and with isolation of mutants of bacterial endophytes resistant to polymyxin B sulfate from cotton plants. Rifampicin-masking was not accounted for in vitro by inhibitory compounds from cotton plant extracts, by bacterial growth on low nutrient agar, or by competition with other bacteria. Collectively, these results suggest that expression of antibiotic-resistance may be altered in planta, although causes for this antibiotic-masking remain to be elucidated, methods for quantifying internal plant colonization by rif+ bacteria should account for this possibility. ei]Section editor: R O D Dixon     

Role of alginate lyase in cell detachment of Pseudomonas aeruginosa.

The exopolysaccharide alginate of Pseudomonas aeruginosa was shown to be important in determining the degree of cell detachment from an agar surface. Nonmucoid strain 8822 gave rise to 50-fold more sloughed cells than mucoid strains 8821 and 8830. Alginate anchors the bacteria to the agar surface, thereby influencing the extent of detachment. The role of the P. aeruginosa alginate lyase in the process of cell sloughing was investigated. Increased expression of the alginate lyase in mucoid strain 8830 led to alginate degradation and increased cell detachment. Similar effects were seen both when the alginate lyase was induced at the initial stage of cell inoculation and when it was induced at a later stage of growth. It appears that high-molecular-weight alginate polymers are required to efficiently retain the bacteria within the growth film. When expressed from a regulated promoter, the alginate lyase can induce enhanced sloughing of cells because of degradation of the alginate. This suggests a possible role for the lyase in the development of bacterial growth films.     

Effect of Motility on Surface Colonization and Reproductive Success of Pseudomonas fluorescens in Dual-Dilution Continuous Culture and Batch Culture Systems

The colonization of glass surfaces by motile and nonmotile strains of Pseudomonas fluorescens was evaluated by using dual-dilution continuous culture (DDCC), competitive and noncompetitive attachment assays, and continuous-flow slide culture. Both strains possessed identical growth rates whether in the attached or planktonic state. Results of attachment assays using radiolabeled bacteria indicated that both strains obeyed first-order (monolayer) adsorption kinetics in pure culture. However, the motile strain attached about four times more rapidly and achieved higher final cell densities on surfaces than did the nonmotile strain (2.03 × 10⁸ versus 5.57 × 10⁷ cells vial^-1) whether evaluated alone or in cocultures containing motile and nonmotile P. fluorescens. These kinetics were attributed to the increased transport of motile cells from the bulk aqueous phase to the hydrodynamic boundary layer where bacterial attachment, growth, and recolonization could occur. First-order attachment kinetics were also observed for both strains by using continuous-flow slide culture assays analyzed by image analysis. The DDCC system contained both aqueous and particulate phases which could be diluted independently. DDCC results indicated that when cocultures containing motile and nonmotile P. fluorescens colonized solid particles, the motile strain replaced the nonmotile strain in the system over time. Increasing the aqueous-phase rates of dilution decreased the time required for extinction of the nonmotile strain while concurrently decreasing the overall carrying capacity of the DDCC system for both strains. These results confirmed that bacterial motility conveyed a selective advantage during surface colonization even in aqueous-phase systems not dominated by laminar flow.     

Cellular interactions between arbuscular mycorrhizal fungi and rhizosphere bacteria

Summary We have investigated whether direct physical interactions occur between arbuscular mycorrhizal (AM) fungi and plant growth promoting rhizobacteria (PGPRs), some of which are used as biocontrol agents. Attachment of rhizobia and pseudomonads to the spores and fungal mycelium ofGigaspora margarita has been assessed in vitro and visualized by a combination of electron and confocal microscopy. The results showed that both rhizobia and pseudomonads adhere to spores and hyphae of AM fungi germinated under sterile conditions, although the degree of attachment depended upon the strain.Pseudomonas fluorescens strain WCS 365 andRhizobium leguminosarum strains B556 and 3841 were the most effective colonizers. Extracellular material of bacterial origin containing cellulose produced around the attached bacteria may mediate fungal/bacterial interactions. These results suggest that antagonistic and synergistic interactions between AM fungi and rhizosphere bacteria may be mediated by soluble factors or physical contact. They also support the view that AM fungi are a vehicle for the colonization of plant roots by soil rhizobacteria.Abbreviations AM arbuscular mycorrhiza - PGPR plant growth promoting rhizobacteria - CBH cellobiohydrolase - DAPG 2,4-(diacetyl-phloroglucinol - TY triptone-yeast - LB Lauria-Bertani Dedicated to Prof. Dr. Dr. h.c. Eberhard Schnepf on the occasion of his retirement     

Antibiotic production in a spatially structured environment

Ecological factors influencing the effects of antibiotic production were explored experimentally and theoretically. A spatially structured model was used to model the dynamics of antibiotic-producing and nonproducing bacteria in which growth of the nonproducers was reduced by neighbouring antibiotic producers. Various factors affecting spatial interactions between the bacteria were examined for their impact on antibiotic producers. Spatial clustering had a positive impact on the effect of antibiotic production, as measured by the decline in growth of the nonproducing strain, while increasing the initial density of the nonproducing strain had a negative impact. Experiments examined the growth of antibiotic-producing Streptomyces species and a nonproducing, antibiotic-sensitive strain of Bacillus subtilis that were coinoculated on surface media. There was an effect of the Streptomyces on Bacillus growth in some experiments but not in others. In light of the predictions from the model, unintentional clustering of cells is a more likely explanation for this finding than different initial Bacillus densities. The importance of spatial structure seen in this study is consistent with a terrestrial rather than an aquatic distribution of antibiotic-producing bacteria, and may have implications in the search for novel antibiotics.     

Distinct glycoconjugate cell surface structures make the pelagic diatom Thalassiosira rotula an attractive habitat for bacteria

Interactions between marine diatoms and bacteria have been studied for decades. However, the visualization of physical interactions between these diatoms and their colonizers is still limited. To enhance our understanding of these specific interactions, a new Thalassiosira rotula isolate from the North Sea (strain 8673) was characterized by scanning electron microscopy and confocal laser scanning microscopy (CLSM) after staining with fluorescently labeled lectins targeting specific glycoconjugates. To investigate defined interactions of this strain with bacteria the new strain was made axenic and co-cultivated with a natural bacterial community and in two- or three-partner consortia with different bacteria of the Roseobacter group, Gammaproteobacteria and Bacteroidetes. The CLSM analysis of the consortia identified six out of 78 different lectins as very suitable to characterize glycoconjugates of T. rotula. The resulting images show that fucose-containing threads were the dominant glycoconjugates secreted by the T. rotula cells but chitin and to a lesser extent other glycoconjugates were also identified. Bacteria attached predominantly to the fucose glycoconjugates. The colonizing bacteria showed various attachment patterns such as adhering to the diatom threads in aggregates only or attaching to both the surfaces and the threads of the diatom. Interestingly the colonization patterns of single bacteria differed strikingly from those of bacterial co-cultures, indicating that interactions between two bacterial species impacted the colonization of the diatom. Our observations help to better understand physical interactions and specific colonization patterns of distinct bacterial mono- and co-cultures with an abundant diatom of costal seas.     

2-n-Pentyl-4-quinolinol produced by a marine Alteromonas sp. and its potential ecological and biogeochemical roles

Bacterium-bacterium interactions occur at intimate spatial scales on the order of micrometers, but our knowledge of interactions at this level is rudimentary. Antagonism is a potential interaction in such microenvironments. To study the ecological role of antibiosis, we developed a model system involving an antibiotic-producing isolate (SWAT5) derived from a marine particle and its dominant antibiotic product, 2-n-pentyl-4-quinolinol (PQ). This system was used to address questions about the significance of this antibiotic for microbial ecology and carbon cycling on particles. We characterized the chemical and inhibitory properties of PQ in relation to the mechanisms used by particle-associated bacteria in interacting with particles and with other attached bacteria. PQ was produced by SWAT5 only on surfaces. When SWAT5 was grown in polysaccharide matrices, PQ diffused within the matrices but not into the surrounding seawater. SWAT5 might thus be able to generate a localized zone of high antibiotic concentration on particles suspended or sinking through seawater. Target bacterial respiration was most sensitive to PQ (75 nM), while inhibition of DNA synthesis, protein synthesis, and bacterial motility required higher (micromolar) PQ levels. The presence of PQ altered the composition of the bacterial community that colonized and developed in a model particle system. PQ also inhibited Synechococcus and phytoplankton growth. Our results suggest that antibiosis may significantly influence community composition and activities of attached bacterial and thus regulate the biogeochemical fate of particulate organic matter in the ocean.     

Scanning electron microscopy and epifluorescence investigation of bacterial colonization of marine sand sediments

Scanning electron microscopy (SEM) was employed for the investigation of microorganisms living in marine sand sediments. Epifluorescence, as well as sediment analyses, gave further data on the parameters of the sediment samples.SEM revealed a correlation between the site and density of bacterial colonization and the microtopography of the individual sand grains.Sand grains with a medium roundness showed the greatest density of bacterial colonization. Protected surface sites were favored in the colonization process. The mode of bacterial attachment varied; mostly the barren sand grain surface was colonized. However, bacteria were also observed close to or within detritus or attached to diatoms. Many of the attaching bacteria observed were found to produce polymer strands.In some cases special structures were discovered which could serve bacterial attachment. Entire colonies attached by means of polymer nets, and disc-shaped bacteria were observed.     

Antibiotic activity of epiphytic bacteria isolated from intertidal seaweeds

A survey of antibiotic-producing bacteria from the microbial flora attached to seaweeds and the study of their antibiotic capacities were carried out. From 5 species of green and brown marine algae, 224 bacterial strains were isolated and tested for antibiotic production. A total of 38 strains displayed antibiotic activity, withEnteromorpha intestinalis being the source of the highest number of producer strains. All epiphytic bacteria with antibiotic activity were assigned to thePseudomonas-Alteromonas group. Antagonism assays among the isolates demonstrated that each producer strain inhibits the growth of the other producers, as well as of some nonproducer strains also isolated from seaweeds. Likewise, an autoinhibitory effect was observed in all antibiotic-producing strains. Antibacterial spectra of all the strains include activity againstStaphylococcus, Alcaligenes, Pseudomonas, Vibrio, Pasteurella, andAchromobacter. A preliminary characterization of the antibiotic substances produced by these epiphytic bacteria demonstrated that they are low molecular weight compounds, thermolabile, and anionic and are not affected by proteolytic enzymes. The role that these inhibitory substances can play in the natural environment is discussed.     

Adhesion of bacteria to epithelial cell surfaces within the reticulo-rumen of cattle.

Blocks of tissue were removed from various locations in the bovine digestive tract and fixed and processed for transmission and scanning electron microscopy by techniques that retained adherent bacteria. The distribution of bacteria on the surface of epithelial cells was examined by scanning electron microscopy. This showed intermittent colonization of the epithelia with the formation of occasional microcolonies of morphologically similar bacterial cells. Transmission electron microscopy of ruthenium red-stained material showed the presence of both the glycocalyx of the bovine epithelial cells and fibrous carbohydrate coats surrounding adherent bacteria. The carbohydrate coats appeared to mediate the attachment of bacteria to the epithelium, to food particles, and to each other so that microcolonies were formed. Careful examination of the bacterial colonization of keratinized cells in the process of being sloughed from the surface of the stratified squamous epithelium of the rumen showed that these dead cells were digested by adherent bacteria of a limited number of morphological types. The spatial relationship of this mixed, adherent, microbial population to living and dead epithelial cells and to food particles indicates that digestive processes of some importance may be accomplished by this stationary component of the microbial flora of the digestive tract.     

Adhesion of bacteria to epithelial cell surfaces within the reticulo-rumen of cattle

Blocks of tissue were removed from various locations in the bovine digestive tract and fixed and processed for transmission and scanning electron microscopy by techniques that retained adherent bacteria. The distribution of bacteria on the surface of epithelial cells was examined by scanning electron microscopy. This showed intermittent colonization of the epithelia with the formation of occasional microcolonies of morphologically similar bacterial cells. Transmission electron microscopy of ruthenium red-stained material showed the presence of both the glycocalyx of the bovine epithelial cells and fibrous carbohydrate coats surrounding adherent bacteria. The carbohydrate coats appeared to mediate the attachment of bacteria to the epithelium, to food particles, and to each other so that microcolonies were formed. Careful examination of the bacterial colonization of keratinized cells in the process of being sloughed from the surface of the stratified squamous epithelium of the rumen showed that these dead cells were digested by adherent bacteria of a limited number of morphological types. The spatial relationship of this mixed, adherent, microbial population to living and dead epithelial cells and to food particles indicates that digestive processes of some importance may be accomplished by this stationary component of the microbial flora of the digestive tract.     

Dynamics of Microbial Communities on Marine Snow Aggregates: Colonization, Growth, Detachment, and Grazing Mortality of Attached Bacteria

We studied the dynamics of microbial communities attached to model aggregates (4-mm-diameter agar spheres) and the component processes of colonization, detachment, growth, and grazing mortality. Agar spheres incubated in raw seawater were rapidly colonized by bacteria, followed by flagellates and ciliates. Colonization can be described as a diffusion process, and encounter volume rates were estimated at about 0.01 and 0.1 cm³ h⁻¹ for bacteria and flagellates, respectively. After initial colonization, the abundances of flagellates and ciliates remained approximately constant at 10³ to 10⁴ and ~10² cells sphere⁻¹, respectively, whereas bacterial populations increased at a declining rate to >10⁷ cells sphere⁻¹. Attached microorganisms initially detached at high specific rates of ~10⁻² min⁻¹, but the bacteria gradually became irreversibly attached to the spheres. Bacterial growth (0 to 2 day⁻¹) was density dependent and declined hyperbolically when cell density exceeded a threshold. Bacterivorous flagellates grazed on the sphere surface at an average saturated rate of 15 bacteria flagellate⁻¹ h⁻¹. At low bacterial densities, the flagellate surface clearance rate was ~5 × 10⁻⁷ cm² min⁻¹, but it declined hyperbolically with increasing bacterial density. Using the experimentally estimated process rates and integrating the component processes in a simple model reproduces the main features of the observed microbial population dynamics. Differences between observed and predicted population dynamics suggest, however, that other factors, e.g., antagonistic interactions between bacteria, are of importance in shaping marine snow microbial communities.     

2-n-Pentyl-4-Quinolinol Produced by a Marine Alteromonas sp. and Its Potential Ecological and Biogeochemical Roles

Bacterium-bacterium interactions occur at intimate spatial scales on the order of micrometers, but our knowledge of interactions at this level is rudimentary. Antagonism is a potential interaction in such microenvironments. To study the ecological role of antibiosis, we developed a model system involving an antibiotic-producing isolate (SWAT5) derived from a marine particle and its dominant antibiotic product, 2-n-pentyl-4-quinolinol (PQ). This system was used to address questions about the significance of this antibiotic for microbial ecology and carbon cycling on particles. We characterized the chemical and inhibitory properties of PQ in relation to the mechanisms used by particle-associated bacteria in interacting with particles and with other attached bacteria. PQ was produced by SWAT5 only on surfaces. When SWAT5 was grown in polysaccharide matrices, PQ diffused within the matrices but not into the surrounding seawater. SWAT5 might thus be able to generate a localized zone of high antibiotic concentration on particles suspended or sinking through seawater. Target bacterial respiration was most sensitive to PQ (75 nM), while inhibition of DNA synthesis, protein synthesis, and bacterial motility required higher (micromolar) PQ levels. The presence of PQ altered the composition of the bacterial community that colonized and developed in a model particle system. PQ also inhibited Synechococcus and phytoplankton growth. Our results suggest that antibiosis may significantly influence community composition and activities of attached bacterial and thus regulate the biogeochemical fate of particulate organic matter in the ocean.     

Qualitative and quantitative agar invasion test based on bacterial colony/biofilm

Invasion of the culture medium is a feature frequently studied in yeasts, in which it has been related to a greater virulence, but it is practically unknown in bacteria. Recently, it has been demonstrated that several clinically relevant bacterial species were also able of invading agar media, so it was necessary to design a microbiological assay to study the expression of this character in bacteria. Accordingly, a bacterial agar invasion test based on colony/biofilm development was designed, which allows qualitative and quantitative characterization of bacterial growth into the agar culture medium. Once the culture conditions were optimized, the test was applied to 90 strains from nine bacterial species, validating its usefulness for differentiating invasive strains (positive) from those non invasive (negative). The test also allows sorting invasive strains according to agar invasion intensity (low, moderate, high) and topographic invasion pattern (peripheral, homogeneous, mixed). Moreover, an image analysis routine to quantify the invasion was developed. Implemented method enables direct measuring of two invasion parameters (invasion area and number of invasion dots), automated calculation of three relative variables (invasion relative area, invasion dots relative density, and invasion dot average area), and the establishment of strain specific frequency histograms.     

A semi-quantitative approach to assess biofilm formation using wrinkled colony development

Biofilms, or surface-attached communities of cells encapsulated in an extracellular matrix, represent a common lifestyle for many bacteria. Within a biofilm, bacterial cells often exhibit altered physiology, including enhanced resistance to antibiotics and other environmental stresses. Additionally, biofilms can play important roles in host-microbe interactions. Biofilms develop when bacteria transition from individual, planktonic cells to form complex, multi-cellular communities. In the laboratory, biofilms are studied by assessing the development of specific biofilm phenotypes. A common biofilm phenotype involves the formation of wrinkled or rugose bacterial colonies on solid agar media. Wrinkled colony formation provides a particularly simple and useful means to identify and characterize bacterial strains exhibiting altered biofilm phenotypes, and to investigate environmental conditions that impact biofilm formation. Wrinkled colony formation serves as an indicator of biofilm formation in a variety of bacteria, including both Gram-positive bacteria, such as Bacillus subtilis, and Gram-negative bacteria, such as Vibrio cholerae, Vibrio parahaemolyticus, Pseudomonas aeruginosa, and Vibrio fischeri. The marine bacterium V. fischeri has become a model for biofilm formation due to the critical role of biofilms during host colonization: biofilms produced by V. fischeri promote its colonization of the Hawaiian bobtail squid Euprymna scolopes. Importantly, biofilm phenotypes observed in vitro correlate with the ability of V. fischeri cells to effectively colonize host animals: strains impaired for biofilm formation in vitro possess a colonization defect, while strains exhibiting increased biofilm phenotypes are enhanced for colonization. V. fischeri therefore provides a simple model system to assess the mechanisms by which bacteria regulate biofilm formation and how biofilms impact host colonization. In this report, we describe a semi-quantitative method to assess biofilm formation using V. fischeri as a model system. This method involves the careful spotting of bacterial cultures at defined concentrations and volumes onto solid agar media; a spotted culture is synonymous to a single bacterial colony. This 'spotted culture' technique can be utilized to compare gross biofilm phenotypes at single, specified time-points (end-point assays), or to identify and characterize subtle biofilm phenotypes through time-course assays of biofilm development and measurements of the colony diameter, which is influenced by biofilm formation. Thus, this technique provides a semi-quantitative analysis of biofilm formation, permitting evaluation of the timing and patterning of wrinkled colony development and the relative size of the developing structure, characteristics that extend beyond the simple overall morphology.     

Inhibitory activity of antibiotic-producing marine bacteria against fish pathogens

The activity of antibiotic-producing marine bacteria was assayed against bacterial fish pathogens belonging to the genera Vibrio, Aeromonas, Pasteurella, Edwardsiella, Yersinia and Pseudomonas with the aim of evaluating the possible use of these marine strains for controlling epizootics in aquaculture. Inhibition tests on solid medium showed that, in general, the majority of fish bacteria were strongly sensitive to the marine bacteria. Only two strains ( Edwardsiella tarda and Pseudomonas aeruginosa ), were resistant to all the antibiotic-producing strains. The results of antagonism assays in sea water, however, varied according to the fish pathogens examined. Experiments conducted using cell-free supernatant fluids of marine bacteria demonstrated the involvement of antibiotic substances in the inhibition of fish pathogens.