Seasonal dynamics of microbial community in an aquaculture system for Nile tilapia (Oreochromis niloticus)

Studies on bacterial abundance and diversity will improve the understanding of the microbial ecology to optimize aquaculture production, water quality, disease control and environmental impact of effluents. We comparatively evaluated an aquaculture system in dry and rainy seasons by Fluorescent in situ Hybridization (FISH) and culture‐dependent methodology. Overall, a negative correlation between seasonality in bacterial and flagellates abundance was observed. Alpha‐, beta‐, gamma‐proteobacteria and Cytophaga‐Flavobacter were observed with seasonal variation. Putative pathogenic bacteria such as coagulase‐negative Staphylococcus, non‐fermenter Gram‐negative rods (Pseudomonas sp. and Burkholderia sp.), and Enterobacteriaceae were also detected in this study with significant seasonal variation. Focusing on medically important bacteria, our data show that microbial diversity in the environment associated with aquaculture, as it is practiced, may be altered in growth ponds used for fish cultivation. As an ecological consequence, potentially pathogenic bacteria might be released in high concentrations to the downstream environments posing potential threats to human and animal health.     

Antagonism against fish pathogens by cellular components and verification of probiotic properties in autochthonous bacteria isolated from the gut of an Indian major carp,Catla catla (Hamilton)

Probiotic potential of the autochthonous bacteria in catla, Catla catla has been evaluated through determination of antagonistic activity (in vitro) of the cellular components of gut bacteria against seven fish pathogens. Altogether 208 strains were isolated, inhibitory activity of the isolates was evaluated through cross‐streaking and 16 primarily selected antagonistic strains were confirmed using the double‐layer method. Four bacteria that showed antagonism against ≥4 pathogens were selected as putative probiotics. The intracellular, extracellular, whole‐cell and heat‐killed cell components exhibited bactericidal activity against the pathogens. In addition, the selected strains were capable of producing different extracellular enzymes, competent to grow in intestinal mucus and could tolerate diluted bile juice. Analysis of 16SrRNA partial gene sequence revealed that both the strains CC1FG2 and CC1FG4 were Bacillus methylotrophicus (KF559344 and KF559345), while the isolates CC1HG5 and CC2HG7 were Bacillus subtilis subsp. spizizenii (KF559346) and Enterobacter hormaechei (KF559347) respectively. Bio‐safety evaluation through intra‐peritoneal injection of the isolates did not induce any pathological signs or mortalities in C. catla. The study confirmed probiotic properties of autochthonous gut bacteria in C. catla and demonstrated potential for using them as bio‐control agents. However, in vivo studies are essential to explore their efficacy in the commercial aquaculture.     

Prevalence and antimicrobial resistance of vibrios of human health significance in inland saline aquaculture areas

This study was conducted to evaluate the prevalence, potential pathogenicity and antimicrobial resistance of Vibrio isolates from 65 soil/water/fish samples collected from inland saline aquaculture areas. Depending on the sample type, presumptive Vibrio counts ranged from 2.50 to 6.16 log₁₀ CFU/ml (or/g). Among the 119 confirmed Vibrio isolates, Vibrio cholerae was found to most dominant (91.6%) and it was detected in all the samples from inland saline areas. Seven other Vibrio spp. including Vibrio parahaemolyticus and Vibrio vulnificus were also detected. Except one O139 serotype, rest of the V. cholerae isolates were found belonging to non‐O1/non‐O139 serogroups. None of the V. cholerae isolate was found positive for ctx gene. Antimicrobial susceptibility testing against 7 commonly used antibiotics revealed highest resistance (50.4%) against ampicillin. Very high intermediate resistance (87.4%) was also observed against erythromycin. Contrary to previous studies, high susceptibility (>70%) to chloramphenicol, nalidixic acid, tetracycline and trimethoprim was observed in Vibrio isolates obtained in present study. Almost 20% of Vibrio isolates were resistant to two or more antibiotic classes with multiple antibiotic resistance (MAR) index value of ≥0.28. Presence of V. cholerae isolates with very high MAR index value of 0.85 also suggested that these multidrug‐resistant environment isolates could serve as reservoir of antibiotic‐resistant genes in aquatic systems. The presence of multiple drug resistance vibrios in emerging inland saline aquaculture systems emphasizes the need for their routine monitoring for developing the risk assessment and mitigation strategies.     

Bacteriocins and their applications for the treatment of bacterial diseases in aquaculture: a review

Aquaculture is one of the fastest growing food producing sectors in the world. The increase in the world population requires faster growth in aquaculture for better food availability and to overcome malnutrition. But this worldwide growth of aquaculture is overwhelmed by catastrophic fish diseases and spoilage during cultivation and preservation problems caused by pathogenic bacteria. Various remedies are available for food preservation and also from the bacterial diseases in the fish production. Due to the emergence of antibiotic resistance and adverse effects, an alternative to antibiotic is the need of the hour. The proteins such as bacteriocins, i.e. ribosomally synthesized antimicrobial peptides (AMPs) and possess antagonistic against closely related and other bacteria. These proteins are produced by most lineages of bacteria which are playing key roles in recognition and possess a cognate immunity system for self‐protection as well as host protection from infections. These proteins are potent immunomodulators with broad spectrum inhibition properties which are further used as novel therapeutic agents. In this review, we have tried to summarize the bacteriocins on the basis of their classifications, structural and functional attributes, mode of actions, bacteriocins isolated from fish and gut microbiota and presence of beneficiary bacteria in the fish gut. Further, this study highlighted where further research is a prerequisite to increase our basic understanding and search for novel bacteriocins to elucidate the proteins/peptides having antimicrobial properties for disease control in aquaculture as an alternative to antibiotics.     

Diversity of siderophore‐producing bacteria isolated from the intestinal tracts of fish along the Japanese coast

This study examined the diversity of siderophore‐producing bacteria in the intestinal tracts of coastal fish in Japan and screened candidate bacterial strains for probiotic use in aquaculture. Of the 2637 bacteria isolated from the 27 fish specimens (13 species) and six environmental samples collected in this study, 266 isolates exhibited the ability to produce siderophores. Siderophore producers were detected in the intestines of 18 of the fish specimens caught (68%) at densities of 2.3 × 10⁴–2.3 × 10⁸ CFU g^?1, in all three seawater samples at 2.0 × 10²–1.3 × 10³ CFU mL^?1 and in all three sand samples at 2.6 × 10¹–2.8 × 10⁴ CFU g^?1. These findings suggest that siderophore‐producing bacteria are widely distributed in the intestinal tracts of coastal fish and their environments. Analysis of 16S rRNA gene sequences revealed that the siderophore producers belonged to 38 species, of which Vibrio splendidus, Vibrio ichthyoenteri and Vibrio crassostreae accounted for 32.7%, 19.5% and 11.3% of the 266 isolates, respectively, suggesting that these bacteria are indigenous to the intestinal tract of coastal fish. Six bacterial species, Enterovibri norvegicus, Photobacterium leiognathi, Photobacterium phosphoreum, Photobacterium rosenbergii, V. crassostrea and Vibrio scophthalmi were identified as possible candidates for use as probionts in fish aquaculture.     

Identification and characterization of lactic acid bacteria isolated from rainbow trout, Oncorhynchus mykiss (Walbaum), with inhibitory activity against Lactococcus garvieae

A study was conducted to evaluate the probiotic properties of endogenous rainbow trout microbiota against pathogenic Lactococcus garvieae. A total of 335 bacterial strains were isolated from rainbow trout and screened for antagonistic activity against L. garvieae using an agar spot assay. Antagonistic strains were grouped by PCR amplification of repetitive bacterial DNA elements (rep‐PCR) and identified by 16S rRNA gene sequence analysis. The results revealed that the antagonistic strains belonged to the genera Lactobacillus, Lactococcus and Leuconostoc. Further probiotic characteristics, such as specific growth rate, doubling time, resistance to biological barriers, antibiotic resistance, hydrophobicity and production of antimicrobial substances, were also studied. These strains were able to survive low pH and high bile concentrations, showed good adherence characteristics and a broad spectrum of antibiotic resistance. The antagonistic efficacy was maintained after sterile filtration and was sensitive to proteinase K, indicating that proteinaceous extracellular inhibitory compounds were at least partially responsible for pathogen antagonism. Based on these results, these strains should be further studied to explore their probiotic effects in challenge experiments in vivo. This study shows clear evidence that the indigenous trout‐associated microbiota may provide a defensive barrier against L. garvieae.     

Antibacterial activity of oxytetracycline on microbial ecology of Nile tilapia (Oreochromis niloticus) gastrointestinal tract under laboratory condition

The antibacterial effect of oxytetracycline (OTC) on bacterial load, pathogen and possible development of antibiotic resistance in intestinal flora of apparently healthy Nile tilapia (Oreochromis niloticus) was investigated under laboratory condition. OTC, a broad spectrum approved antibiotic widely used in aquaculture, was fed to Nile tilapia through medicated diet at a rate of 2 g/ kg of feed for consecutive 7 days in treatment 1 (T₁) and 30 days in treatment 2 (T₂). Changes in physico‐chemical parameters of water were also recorded where pH and dissolved oxygen influenced bacterial load and coliform count respectively. At the start of the experiment, initial bacterial load in fish intestine was 1.61 ± 2.25 × 10¹⁰ cfu/g, which after OTC treatment decreased significantly (p < .05) to 3.06 ± 2.08 × 10⁸ cfu/g on day 7 and 3.45 ± 4.46 × 10⁷ cfu/g on day 30 when compared with the control group. Among the 10 bacterial genera identified in the intestine, 4 potential public health concern bacteria viz., Salmonella, Escherichia, Enterobacter and Staphylococcaus were predominant. There was a sharp increase in bacterial load on day 18 in T₂ that may be attributed to the development of antibiotic resistance in intestinal bacteria. Antibiotic susceptibility test for isolates against seven antibiotics: amoxycillin, ciprofloxacin, co‐trimoxazole, erythromycin, gentamicin, nitrofurantoin and tetracycline using disc diffusion method revealed significantly increased resistance of Gram‐negative rods (p < .05) that possibly caused increased frequency of OTC‐resistant microorganisms. In conclusion, short‐ and long‐term exposure to OTC treatment affected the distribution of bacterial genera including pathogens in the Nile tilapia gastrointestinal tract and concomitantly influenced their antimicrobial resistance.     

Isolation and evaluation of putative probiotic strains from different teleost to prevent Pseudomonas aeruginosa infection in Cyprinus carpio

Probiotics renowned as valuable microbes serve as a potential alternative to control diseases in aquaculture and are considered as an efficient and environment‐friendly approach to reduce the use of antibiotics. The present study aims at the isolation of putative probiotic bacteria from the intestinal tract of different fish species from the Doaba region of Punjab, India. In this study, isolated bacterial strains were characterized based on their morphological, biochemical and molecular characterization by 16S rRNA gene sequencing, followed by in vitro evaluation of different selection parameters described in FAO/WHO guidelines. A total of 169 different bacterial strains were isolated from the gastrointestinal tract of 52 different fish species. After in vitro evaluation, out of 169 bacterial strains only five bacteria (S3, S7, BDK2', BDK7 and BDK9) identified as Enterococcus and Bacillus species showed antagonistic activity against the fish pathogen Pseudomonas aeruginosa (MTCC 4 673). These isolates were screened based on their response towards bile tolerance, pH tolerance, adhesion and drug susceptibility to different antibiotic discs. And, the in vivo results indicated improved growth and survival against the infection (P. aeruginosa) after oral administration of the probiotics. The observations of in vitro and in vivo evaluation indicate that these isolated probiotic strains serve as effective probiotics and can be used as a novel and safe treatment to cure current issues in aquaculture.     

In vitro selection and characterization of putative probiotics isolated from the gut of Acipenser baerii (Brandt, 1869)

To select and characterize potential probiotic bacteria from the gut microbiota of Siberian sturgeon (Acipenser baerii), 129 strains isolated from the hindgut were screened for antagonistic activity against five fish pathogens. Ten isolates showed antagonism towards three or more pathogens. Nine of these isolates were Gram‐positive, belonging to Lactococcus (seven) and Bacillus (two), and a single strain belonging to the Gram‐negative Citrobacter. These inhibitory isolates were identified using genetic, phentotypic and biochemical traits, and further characterized by in vitro tests assessing the adhesion and growth in mucus and resistance to gastric and intestinal fluids. The candidate probiotics were determined to be non‐pathogenic through an in vivo study. Based on these assays, Lactococcus lactis ssp. lactis STG45 and STG81 showed the broadest inhibitory potential, a high viability in simulated gastrointestinal juice and the highest adhesion capacity to mucus. They were therefore selected as the most promising candidate probiotics. This is the first study screening probiotics among the gut microflora of Siberian sturgeon.     

Isolation and use of beneficial microbiota from the digestive tract of lions‐paw scallop Nodipecten subnodosus and winged pearl oyster Pteria sterna in oyster aquaculture

We isolated microbiota from the digestive tract of Nodipecten subnodosus and Pteria sterna and determined in vitro their haemolytic activity, antagonism against Vibrio spp., bacterial hydrophobicity, production of extracellular enzymes and molecular identification. Five bacterial strains were selected: RL5 and C3 (Lactobacillus spp.) and PB1‐1, PB1‐5 and PB1‐6 (Bacillus spp.). The RL5 and C3 isolates showed antimicrobial activity against Vibrio spp. and the PB1‐1, PB1‐5 and PB1‐6 isolates showed enzymatic activity for amylase, protease, lipase and cellulose; the C3 and PB1‐5 isolates were highly hydrophobic. The selected strains of bacteria were tested in vivo as probiotics, together with a treatment of ampicillin and a control without bacteria on juvenile Kumamoto oysters Crassostrea sikamea. The strains were provided individually and as mixes of isolates. Survival, growth and biochemical composition of the juveniles were determined as in vivo indicators. Juveniles grew significantly larger and faster when treated with a specific mix of isolates (MIX‐B), compared with the control. The protein, lipid and carbohydrate concentrations were also significantly higher in oysters exposed to probiotic treatments, compared with the control and the antibiotic treatment. The selected microbiota showed probiotic proprieties for cultivating C. sikamea juveniles.     

Incidence of class 1 integron and other antibiotic resistance determinants in Aeromonas spp. from rainbow trout farms in Australia

There is limited information on antibiotic resistance determinants present in bacteria of aquaculture origin in Australia. The presence of integron and other resistance determinants was investigated in 90 Aeromonas isolates derived from nine freshwater trout farms in Victoria (Australia). Polymerase chain reaction was carried out for the detection of integrase genes Int1, Int2 and Int3, gene cassette array, integron‐associated aadA, sul1 and qac1 genes, streptomycin resistance genes strA‐strB, β‐lactamase resistance genes bla_TEM and bla_SHV, and tetracycline resistance genes tetA‐E and tetM. Clonal analysis was performed by pulsed‐field gel electrophoresis (PFGE). Class 1 integrons were detected in 28/90 (31%) and class 2 and class 3 in none of the strains, aadA gene in 19/27 (70%) streptomycin‐resistant strains, sul1 in 13/15 (86.7%) sulphonamide‐resistant strains and qac1 gene in 8/28 (28.6%) integron‐bearing strains. Five strains from two different farms carried gene cassettes of 1000 bp each containing the aadA2 gene and PFGE analysis revealed genetic relatedness. tetC was detected in all and tetA in 9/18 (50%) tetracycline‐resistant strains. The strA‐strB, bla_TEM or bla_SHV genes were not detected in any of the strains. Aeromonas spp. carrying integrons and other resistance genes are present in farm‐raised fish and sediments even though no antibiotics were licensed for use in Australian aquaculture at the time of the study.     

Occurrence of multiple antibiotic resistance and genotypic characterization in Edwardsiella tarda isolated from cage‐cultured hybrid red tilapia (Oreochromis sp.) in the Ping River,Northern Thailand

Edwardsiella tarda is a pathogen that causes edwardsiellosis in aquatic animals. The emergence of multiple antibiotic‐resistant strains makes antibiotic treatment difficult. This study aimed to investigate the antibiotic susceptibility patterns and the genotypic characterization of E. tarda isolated from cage‐cultured red tilapia in Thailand. A total of 30 isolates were identified as E. tarda using biochemical and molecular analysis. The disc diffusion method for testing antibiotic susceptibility showed all the isolates were resistant to colistin sulphate and oxolinic acid. High levels of resistance to amoxicillin, ampicillin, ceftazidime, oxytetracycline and sulphamethoxazole/trimethoprim were observed as well. The multiple antibiotic resistance index ranged from 0.25 to 0.92, indicating that these isolates had been exposed to high risk sources of contamination where antibiotics were commonly used. All the isolates carried the bla_TEM gene based on polymerase chain reaction (PCR). The tetA and sul3 genes were detected in 90% (27/30) and 26.7% (8/30) of the isolates respectively. Nine different genetic groups of isolates were obtained using enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC‐PCR). A correlation between genetic types and multiple antibiotic‐resistant patterns was found. These results highlight the potential risks of multiple antibiotic‐resistant isolates for humans and the environment.     

凡纳滨对虾不同生长阶段肠道可培养细菌耐药性研究

在凡纳滨对虾(Litopenaeus vannamei)养殖过程中多使用微生态制剂来调节水质,为避免破坏池塘菌群结构,很少使用抗生素.为了解凡纳滨对虾肠道细菌耐药性与不同生长阶段的关系,本研究选取江苏地区4种主要养殖模式凡纳滨对虾成虾和虾苗作为研究对象,利用K-B纸片法和qRT-PCR技术,研究对虾样本肠道可培养细菌对...     

Screening of cultivated seaweeds for antibacterial activity against fish pathogenic bacteria

Because of the evolving resistance of microorganisms to existing antibiotics, there is an increasing need for new antibiotics not only in human but also in veterinary medicine. Competition for space and nutrients led to the evolution of antimicrobial defence strategies in the aquatic environment. Therefore, aquatic organisms, e.g., seaweeds, offer a particularly rich source of potential new drugs. The aim of our studies was to identify seaweeds, which possess activities against fish pathogenic bacteria and could be an alternative to the commonly used antibiotics in aquaculture.Dichloromethane, methanole and water extracts of 26 species of cultivated seaweeds were screened for their antibacterial activities against five fish pathogenic bacteria strains (Aeromonas salmonicida ssp. salmonicida, Aeromonas hydrophila ssp. hydrophila, Pseudomonas anguilliseptica, Vibrio anguillarum, Yersinia ruckeri). The dichloromethane extracts of Asparagopsis armata, Ceramium rubrum, Drachiella minuta, Falkenbergia rufolanosa, Gracilaria cornea and Halopitys incurvus showed strong antibacterial activities. V. anguillarum and P. anguilliseptica were the two most susceptible bacteria strains. The screening results confirm the possible use of seaweeds as a source of antimicrobial compounds or as a health-promoting food for aquaculture.     

育肥饲料中虾青素含量对雄性中华绒螯蟹肠道和鳃部可培养优势细菌数量和组成的影响

在育肥饲料中添加虾青素会影响中华绒螫蟹肠道及鳃内的可培养优势菌群以及机体的免疫力,本实验旨在探讨饲料中虾青素含量及养殖水体对其肠道和鳃部菌群的影响。采用不同虾青素含量的饲料(0.00、26.60、41.62、81.37和75.35 mg/kg,分别对应饲料1#~5#)对雄性中华绒螫蟹进行育肥70 d后,利用基础培养基和选择性培养基对其肠道、鳃及养殖水体中的细菌进行传统分离和纯化,所得菌株进行16S rRNA测序,再进行同源性分析后做系统进化树。结果发现,在分离获得的106株细菌(登录号:KU570293~KU570368,KU570370,KU570372~KU570383,KU601302~KU6013 16,KU720553)中,92株为优势菌株,其中,肠道、鳃部和水体各29、32和31株。肠道中的可培养优势细菌属于柠檬酸杆菌属、假单胞菌属和气单胞菌属;而鳃中可培养优势细菌属于柠檬酸杆菌属、芽孢杆菌属、假单胞菌属和气单胞菌属;养殖水体中可培养优势细菌属于芽孢杆菌属、气单胞菌属、柠檬酸杆菌属、假单胞菌属和黄杆菌属。饲料2#组肠道中可培养细菌总数最高(1.06×10~8cfu/g);饲料5#肠道中的潜在致病菌数量最低,但其可培养细菌总数显著高于其他饲料组;饲料3#组肠道和鳃中潜在致病菌数量相对较高。研究表明,饲料中添加不同含量的虾青素能够显著影响雄性中华绒螫蟹肠道和鳃中的菌群构成,但对水体中可培养细菌数量无显著影响。本研究首次对投喂不同含量虾青素饲料的育肥期雄性中华绒螫蟹肠道、鳃及养殖水体可培养的优势细菌数量和组成进行分析,探究了饲料中虾青素含量以及养殖水体与蟹肠道内可培养细菌之间的联系,为中华绒螫蟹营养免疫和菌群调控提供了理论基础。     

Assessment of Aquatic Aeromonas spp. Isolates' Susceptibility to Cinnamaldehyde,Vanillin, and Crude Kigelia africana Fruit Extracts

Aeromonas spp. are primary and opportunistic fish pathogens that are responsible for severe economic losses in aquaculture, and are exacerbated by their increasing incidence of antimicrobial resistance. Phytochemicals are being explored as alternatives to conventional antimicrobial agents since they have destressing, growth‐promoting, immune‐stimulating, and antimicrobial properties. The susceptibility of 93 aquatic Aeromonas spp. and six Plesiomonas shigelloides isolates (from cultured catfish, koi carp, tilapia, and seawater) to three phytochemicals, viz.: cinnamaldehyde (10–1250 µg/mL), vanillin (5–250 µg/mL), and crude Kigelia africana fruit extracts (4–10 mg/mL ethyl acetate, dichloromethane, methanol, and hexane) were assessed using the disk diffusion assay and compared to ampicillin and tetracycline, using activity indices. No zones of inhibition were obtained with 10 µg/mL of cinnamaldehyde, or with 5–250 µg/mL of vanillin. However, varying degrees of inhibition were observed with higher concentrations of cinnamaldehyde, as well as with the K. africana methanol extract. Cinnamaldehyde (≥500 µg/mL) and the K. africana methanol extract displayed better antimicrobial activity against study isolates in comparison to vanillin and ampicillin. They appear to be promising and sustainable phytochemicals that might be useful as alternatives to the antimicrobial agents currently in use in aquaculture.     

近江牡蛎肠道厌氧菌及其产酶能力的研究

为了解近江牡蛎肠道厌氧菌在食饵消化过程中的作用，为微生态饲用添加剂的开发研究打下基础，从近江牡蛎肠道中分离出8株厌氧菌，其中有5株为革兰氏阳性菌。并研究了它们蛋白酶、脂肪酶、淀粉酶、纤维素酶的产酶能力，结果表明仅有3株菌具有产酶能力：2株菌能分泌脂肪酶，1株菌能分泌蛋白酶。由此可见．厌氧菌在分泌消化酶、消化食饵中所起的作用不大。     

The effect of dietary inulin on aerobic bacteria associated with hindgut of Arctic charr (Salvelinus alpinus L.)

The primary aim of the present study was to evaluate the population level of adherent (autochthonous) aerobic and facultative anaerobic bacteria in the hindgut of healthy Arctic charr (Salvelinus alpinus L.) fed dextrin or inulin. This was assessed by the dilution plate technique, and visualized using both transmission and scanning electron microscopy. A population level of 4.8 × 10⁵ adherent bacteria per gram wet mass was found in the hindgut of fish fed a casein‐based diet supplemented with 15% dextrin. However, substituting dextrin with 15% inulin reduced the bacterial population level in the hindgut (3.56 × 10⁴). A total of 217 bacterial isolates were identified by key phenotypical and biochemical characteristics. In addition, 22 strains were also identified by partial sequencing of the 16S rRNA gene. The composition of bacteria colonizing the hindgut of Arctic charr fed dextrin was dominated by the genera Staphylococcus, Pseudomonas, Micrococcus, Psychrobacter glacincola and Streptococcus. However, bacteria colonizing the hindgut of fish fed inulin were dominated by Gram‐positive bacteria of the genera Staphylococcus, Streptococcus, Carnobacterium and Bacillus. While Carnobacterium divergens‐like strains were isolated from charr fed dextrin, Carnobacterium maltaromicus‐like strains were isolated from the hindgut of fish fed inulin. Electron microscopical analysis of hindgut regions confirmed traditional culture‐based microbial analysis as fewer bacterial cells were observed between microvilli and associated with the surfaces of enterocytes of fish fed inulin rather than dextrin.     

Provisional epidemiological cutoff values for standard broth microdilution susceptibility testing of Flavobacterium columnare

The gliding aquatic bacterium Flavobacterium columnare causes columnaris disease, a common problem for wild and farmed freshwater fish worldwide. Recently, a broth microdilution method was standardized to test the susceptibility of F. columnare against antimicrobials commonly used in aquaculture. We used this new method to measure the minimal inhibitory concentrations (MICs) of ten antimicrobials against 120 F. columnare isolates. The resulting MIC frequency distributions for each antimicrobial (1 MIC/isolate) were used to estimate epidemiological cut‐off values (ECVs) which separate isolates with typical wild‐type (WT) susceptibility from isolates with decreased non‐wild‐type (NWT) susceptibility. We identified 22 NWT isolates with elevated MICs relative to the ECV that covered 99.9% of the MIC distribution against one or more of the antimicrobials: ampicillin, enrofloxacin, erythromycin, florfenicol, flumequine, oxolinic acid or oxytetracycline. Ten of the NWT isolates had decreased susceptibility to a single antimicrobial class, six isolates to two antimicrobial classes and six isolates to three or more antimicrobial classes. The MIC frequency distributions and provisional cut‐off values provide data needed to set epidemiological cut‐off values to monitor for the development of antimicrobial resistance among F. columnare.