Determination of the Median Lethal Dose of Botulinum Serotype E in Channel Catfish Fingerlings

Abstract

The median lethal dose of botulinum serotype E in 5.3-g channel catfish Ictalurus punctatus fingerlings was determined. Five tanks (five fish/tank) were assigned to each of the following treatment groups: 70, 50, 35, 25, or 15 pg of purified botulinum serotype E. Fish were injected intracoelomically and observed for 96 h. Administration of the toxin resulted in initial hyperactivity followed by erratic swimming, paresis, and death. The cumulative mortality by treatment group was 100% at 70 pg, 96% at 50 pg, 100% at 35 pg, 88% at 25 pg, and 56% at 15 pg. The median lethal dose was calculated as 13.7 pg/fish (equivalent to a 0.81 median lethal dose for mice Mus musculus) using a logistic regression model. All fish were necropsied; lesions included exophthalmia, ascites, splenic congestion, intussusception of the intestines, congested spleen, and blanching of the intestinal tract. The resultant clinical signs and lesions were similar to those noted in the syndrome of visceral toxicosis of catfish. This study indicates that channel catfish are more sensitive to the effects of botulinum serotype E than laboratory mice, and the signs and lesions of visceral toxicosis of catfish were replicated by injecting catfish with the toxin.

Received October 13, 2011; accepted January 14, 2011     

Detection of botulinum type E toxin in channel catfish with visceral toxicosis syndrome using catfish bioassay and endopep mass spectrometry.

Visceral toxicosis of catfish (VTC) is a syndrome characterized by sudden mortality in apparently healthy market- and brooder-sized catfish (Ictalurus punctatus). This paper reports the design of a catfish neutralization assay to detect botulinum in catfish with VTC and verification by Endopep mass spectrometry (Endopep-MS). Sera from 6 affected catfish were incubated with botulinum antitoxin serotypes A, B, C, D, E, or F. For each serum sample, 3 experimental fingerlings were injected intracoelomically with each serotype-serum mixture and placed separately in an aquarium. Three fish were injected with VTC-affected serum only, and 3 fish were injected with unaffected serum only and also placed in separate aquaria. Signs of morbidity and mortality were seen in fish injected with sera combined with serotype A, B, C, or D, as well as in positive controls. No morbidity or mortality was seen in fish injected with sera combined with antitoxin serotypes E or F or negative control serum. Sera from affected and unaffected catfish were sent to Centers for Disease Control and Prevention for detection and differentiation of botulinum neurotoxin. Aliquots of 0.5 ml of sera were incubated with magnetic beads coated with antibodies to botulinum, and the beads were subjected to the Endopep-MS reaction. Sera from affected catfish tested positive for botulinum E. Sera from 34 unaffected catfish tested negative for botulinum. Although there was not enough botulinum present in affected samples to obtain exact quantification, the estimated quantity of botulinum E in these sera samples was between 0.01 and 0.5 mouse LD50/ml.     

Determination of the median toxic dose of type C botulinum toxin in lactating dairy cows.

Because of the difficulty in identifying botulinum toxin in cattle, it is hypothesized that cattle are sensitive to levels of toxin below the detection limits of current diagnostic techniques (the mouse protection bioassay and the immunostick enzyme-linked immunosorbent assay [ELISA] for type C botulinum toxin). Using an up-down method for toxicologic testing, the median toxic dose (MTD50) for cattle was determined. Four lactating Holstein cows were dosed at 0.125 or 0.25 ng/kg with Clostridium botulinum type C toxin and failed to develop clinical signs of botulism during the 7-day observation period. Three cows given 0.50 ng/kg of toxin developed clinical signs of botulism. From these results, the MTD50 was calculated at 0.388 ng/kg (3.88 mouse lethal doses/kg) using the trim-logit method. These results suggest that cattle are 12.88 times more sensitive to type C botulinum toxin than a mouse on a per kilogram weight basis. The mouse protection bioassay and the immunostick ELISA for type C botulinum toxin failed to identify the presence of the toxin in the serum, blood, and milk samples taken from all 7 animals.     

Effects of Escherichia coli Shiga-like toxins (verotoxins) in pigs.

Escherichia coli K12 strains TB1(pCG5), with the genes for Shiga-like toxin IIv from an edema disease isolate of E. coli and TB1(pCG5-1), with the toxin genes inactivated by transposon mutagenesis, were used to test the hypothesis that Shiga-like toxin IIv was the same as edema disease principle. Ammonium sulfate precipitated culture supernatants from the pair of E. coli K12 strains and from a wild edema disease isolate of E. coli (E145) were tested for their ability to induce signs and lesions of edema disease in intravenously inoculated weaned pigs. Similar preparations from E. coli which produce Shiga-like toxins I and II were also tested. Preparations from E. coli TB1 (pCG5) and E145 contained high levels of Shiga-like toxin IIv and induced signs and lesions similar to those seen in edema disease, whereas preparations from E. coli TB1 (pCG5-1) failed to induce signs or lesions of edema disease. All Shiga-like toxin preparations produced delayed neurological signs, fibrinoid necrosis of arterioles and hemorrhages in the cerebellum of pigs. High doses of Shiga-like toxin IIv were associated with superficial necrosis of the colonic epithelium and vasculitis. Shiga-like toxins I and II resulted in kidney lesions but no enteric pathology. Shiga-like toxin II preparations had the lowest median lethal dose for pigs, Shiga-like toxin IIv was intermediate and Shiga-like toxin I was the least toxic.     

Effects of variable periods of food deprivation on the development of enteric septicemia in channel catfish

Enteric septicemia of catfish (ESC), caused by the bacterium Edwardsiella ictaluri, is the most significant bacterial disease affecting channel catfish Ictalurus punctatus. Withholding feed during outbreaks of ESC is a widely accepted industry practice used to control losses from the disease. Scientific evidence concerning the validity of the practice is contradictory. Two studies were conducted to further evaluate the survival of channel catfish fingerlings following variable periods of feed deprivation before and after exposure to E. ictaluri in controlled aquarium experiments. In the first study, feed was withheld for varying time periods before bacterial challenge. After bacterial challenge, feed was either withheld or fish were fed daily. The second study utilized fish fed daily or fish deprived of feed 7 d before bacterial challenge. Daily feeding was resumed 4, 48, and 96 h after fish were exposed to E. ictaluri. In both experiments, the prechallenge feed treatments did not affect mortality. In contrast, withholding feed after bacterial challenge reduced mortalities by 52% in experiment 1 and by 45% in experiment 2. The highest mortality was observed when fish were fed immediately after immersion exposure and the lowest when fish were completely denied feed or fed daily starting 96 h after challenge. This reduction in mortality occurred when the concentration of E. ictaluri in aquarium water was negligible. These data suggest that when E. ictaluri is present in the water, feeding fish increases mortality by enhancing oral exposure to the pathogen.     

Transmission of Edwardsiella ictaluri from Infected,Dead to Noninfected Channel Catfish

Abstract

Enteric septicemia of catfish (ESC) was transmitted horizontally from channel catfish Icialurus punctatus that had died from Edwardsiella ictaluri infection to contact channel catfish during 2 d of habitation in a tank. The contact channel catfish became positive for E. ictaluri antibody, became infected with this bacterium, and had signs of ESC and died within 12 d postexposure. Edwardsiella ictaluri was recovered from 24 of the 30 contact channel catfish that died from ESC, as well as from 9 of the 25 tested contact survivors. The cannibalizing of E. ictaluri-infected fish, or the shedding of E. ictaluri from dead fish, or both, were shown to be mechanisms of horizontal transmission of ESC among channel catfish.     

The occurrence of Clostridium botulinum type E in Finnish trout farms and the prevention of toxin formation in fresh-salted vacuum-packed trout fillets.

The occurrence of C. botulinum on two Finnish rainbow trout farms were studied. C. botulinum type E toxin was detected from samples of fish intestines in 10% and 4% of the samples and in 0% and 100% of dam bottom sediments, respectively. The toxin formation of inoculated C. botulinum type E in three different brands of commercial fresh-salted vacuum-packed trout fillets was also investigated. In the brand with a salt concentration of 2.7% (a w = 0.96) and no nitrate, the toxin was formed in two weeks at 10 and 20 degrees C. If the pH of the product was lowed to 4.90 and NaCl concentration increased to 5.67% (a w = 0.94%) no toxin was formed even without the use of nitrate. By adding 0.1% of sodium nitrate to the curing solution (0.046% nitrate in the product) the toxin formation was brought to an end. According to the results of the study C. botulinum type E presents a potential health risk for man as well as for fish also in Finland. Trout products must be manufactured and stored in such a way that the possibility of toxin formation is eliminated.     

Survival and Antibody Response of Channel Catfish,Blue Catfish,and Channel Catfish Female × Blue Catfish Male Hybrids after Exposure to Edwardsiella ictaluri

Abstract

Juvenile Norris strain channel catfish Ictalurus punctatus, blue catfish I. furcatus, and Norris strain channel catfish female × blue catfish male hybrids were challenged with Edwardsiella ictaluri by bath immersion or intraperitoneal injection (high or low dose) in aquaria. Survival (%) after bath immersion was highest for blue catfish (89.5 ± 2.8), intermediate for hybrids (73.8 ± 6.7), and lowest for channel catfish (62.0 ± 4.2). Prechallenge antibody levels to E. ictaluri, measured by enzyme-linked immunosorbent assay, were negative (mean ± SE optical density [OD] = 0.010 ± 0.003). Postchallenge antibody response for blue catfish (OD = 0.132 ± 0.045) was significantly lower than that of channel catfish (OD = 0.350 ± 0.045), whereas the response of the channel × blue catfish F₁ hybrids (OD = 0.263 ± 0.051) was intermediate and not significantly different from either parental species. Intraperitoneal injections of E. ictaluri resulted in significant mortality only in channel catfish (88.3 ± 2.6% survival) and were sublethal to hybrid catfish and blue catfish with 100.0% and 99.3 ± 0.4% survival, respectively. Antibody responses after the injection challenge were significantly different among catfish groups and injection dose with no group × dose interaction. Antibody responses after the injection challenge were consistent with the immersion challenge, and means of high and low challenge doses were lowest in blue catfish (OD = 0.061 ± 0.014), intermediate in hybrids (OD = 0.187 ± 0.014), and highest in channel catfish (OD = 0.272 ± 0.014). For all fish groups combined, the high injection challenge dose resulted in higher antibody levels (OD = 0.206 ± 0.011) than low injection challenge dose (OD = 0.140 ± 0.012). Overall results indicate greater resistance to E. ictaluri and lower antibody response in blue catfish, and show the potential to identify molecular markers linked with disease resistance and introgression of resistance genes from blue catfish into channel catfish.     

Use of a Mini-Transposon to Study Chondroitinase Activity Associated with Edwardsiella ictaluri

Abstract

Edwardsiella ictaluri, the etiological agent of enteric septicemia of catfish (ESC), is the leading cause of bacterial disease in commercially raised channel catfish Ictalurus punctatus. Little work has been conducted at a genotypic level to determine potential virulence characteristics, but the production of chondroitin sulfatase is a suspected virulence factor. Using transpositional mutagenesis, we created stable E. ictaluri mutants that are deficient in chondroitinase activity. Channel catfish were challenged by injection with E. ictaluri transposon mutant MI15. None of the catfish challenged with the mutant died or showed signs of ESC. These fish were held for 2 weeks and then challenged by injection with the known virulent parent strain of E. ictaluri. The challenged naive control fish showed clinical signs of and a mortality rate consistent with ESC, whereas catfish that had been injected with MI15 prior to challenge with the parent strain were resistant to disease. This work represents a preliminary study to suggest a possible role of chondroitin sulfatase activity in the virulence of E. ictaluri.     

Enhanced susceptibility of channel catfish to the bacterium Edwardsiella ictaluri after parasitism by Ichthyophthirius multifiliis

Bacterium Edwardsiella ictaluri and parasite Ichthyophthirius multifiliis (Ich) are two common pathogens of cultured fish. The objective of this study was to evaluate the susceptibility of channel catfish Ictalurus punctatus to E. ictaluri and determine bacterial loads in different fish organs after parasitism by Ich. Fish received the following treatments: (1) infected by I. multifiliis at 5000 theronts/fish and exposed to E. ictaluri; (2) infected by I. multifiliis alone; (3) exposed to E. ictaluri alone; and (4) non-infected control. E. ictaluri in fish organs were quantified by quantitative real-time polymerase chain reaction and reported as genome equivalents per mg of tissue (GEs/mg). The results demonstrated that the Ich-parasitized catfish showed significantly (P<0.05) higher mortality (91.7%) when exposed to E. ictaluri than non-parasitized fish (10%). The bacterial loads in fish infected by 5000 theronts/fish ranged from 6497 to 163,898 GEs/mg which was between 40 and 2000 fold higher than non-parasitized fish (49-141 GEs/mg). Ich infection enhanced the susceptibility of channel catfish to bacterial invasion and increased fish mortality.     

Suspected caffeine and ephedrine toxicosis resulting from ingestion of an herbal supplement containing guarana and ma huang in dogs: 47 cases (1997-1999)

OBJECTIVE: To describe the clinical signs following ingestion of an herbal supplement containing guarana and ma huang in dogs, estimate minimum dose at which clinical signs of toxicosis and death were reported, and evaluate treatment options. DESIGN: Retrospective study. ANIMALS: 47 dogs with evidence of ingestion of an herbal supplement containing primarily guarana and ma huang. PROCEDURE: Records of dogs that had ingested an herbal supplement containing ma huang and guarana between July 1997 and October 1999 were retrieved from the National Animal Poison Control Center database. Data were retrieved and reviewed regarding signalment, dose ingested, clinical signs, laboratory test results, treatment, and final outcome. Cases were assessed by staff veterinarians as toxicosis or suspected toxicosis on the basis of strength of evidence supporting a diagnosis. RESULTS: Most dogs (80%) developed clinical signs of toxicosis within 8 hours of ingestion, and clinical signs persisted for up to 48 hours. Hyperactivity, tremors, seizures, and behavior changes were reported in 83% of dogs; other signs included vomiting (47%), tachycardia (30%), and hyperthermia (28%). Seventeen percent of the dogs died or were euthanatized. Estimated doses of guarana and ma huang ranged from 4.4 to 296.2 mg/kg (1.98 to 133.2 mg/lb) and 1.3 to 88.9 mg/kg (0.58 to 40.0 mg/lb) of body weight, respectively; minimum dose at which death was reported was 19.1 mg of guarana/kg (8.7 mg/lb) and 5.8 mg of ma huang/kg (2.6 mg/lb). CONCLUSIONS AND CLINICAL RELEVANCE: Accidental ingestion of herbal supplements containing primarily guarana and ma huang in dogs can lead to a potentially lethal condition that may require prompt detoxification and supportive treatment for several days. Most dogs recovered with supportive treatment.     

Outer membrane protein profiles of Edwardsiella ictaluri from fish.

Outer membrane proteins (OMP) prepared with sodium N-lauroyl sarcocinate (SLS) from 33 Edwardsiella ictaluri isolates from fish were examined by electrophoresis. Twenty-eight isolates from channel catfish (Ictalurus punctatus) had similar OMP profiles. Ten bands (71 kilodaltons [kD] to 19.5 kD) were identified in all isolates from channel catfish. One major 35-kD protein comprised most of the protein content of the outer membrane of isolates from channel catfish. Differences existed among isolates in the amount of protein within minor OMP bands. Edwardsiella ictaluri ATCC 33202 contained larger quantities of the 38.5- and 37-kD proteins than did the other isolates. Outer membrane protein profiles of E ictaluri derived from Bengal danio (Danio devario) and walking catfish (Clarias batrachus) were identical to OMP profiles of isolates from channel catfish. In contrast, OMP profiles from single isolates from green knife fish (Eigemannia virescens) and white catfish (Ictalurus catus) were different. Variations in incubation time, SLS extraction time, SLS extraction number, and in vivo and in vitro passage had no effect on the OMP profile of E ictaluri ATCC 33202. An increase in duration of sample solubilization did affect the OMP profile of E ictaluri ATCC 33202 by decreasing the amount of protein in 52-, 46-, and 43.5-kD bands. Accompanying the decrease were increased staining intensity in the 31.5- and 28.5-kD bands and the appearance of 4 new bands (34, 33, 25.5, and 22.5 kD). Edwardsiella ictaluri, a gram-negative bacterium in the family Enterobacteriaceae, is the cause of enteric septicemia of catfish.(ABSTRACT TRUNCATED AT 250 WORDS)     

Longevity of Bolbophorus damnificus infections in channel catfish

The digenean Bolbophorus damnificus infects commercial channel catfish Ictalurus punctatus, causing mortality, lower feed consumption, and reduced growth in surviving fish. The purpose of this study was to determine the length of time for which B. damnificus prodiplostomulum metacercariae (juvenile trematode stage that infects fish) would remain viable (parasite appearing to be intact or exhibiting movement) in channel catfish. Fish (n = 210) were infected with molecularly confirmed B. damnificus cercariae harvested from naturally infected marsh rams-horn snails Planorbella trivolvis. During the first sampling (at 20 d postinfection), 8.3 +/- 3.6 metacercariae/fish (mean +/- SD) were found in the host muscle and visceral organs. The channel catfish were then acclimated to a water temperature of either 18 degrees C or 28 degrees C. After 11 months, 6.8 +/- 3.5 and 5.9 +/- 3.0 metacercariae/fish were found in groups held at 18 degrees C and 28 degrees C, respectively. The mean number of parasites per fish did not significantly differ between fish held at the two temperatures and did not significantly decline over time at either temperature. Fish examined from 13 to 30 months postinfection all contained viable metacercariae that were morphologically and molecularly identified as B. damnificus. At 18 months, 12 metacercariae (of which 11 were intact and 10 displayed movement) were found in the one fish sampled; at 30 months, the last fish sampled contained three intact metacercariae (one displayed slight movement). Our results indicate that B. damnificus metacercariae can remain viable in channel catfish for at least an 18-30-month production cycle during which they have the potential to affect fish growth; in addition, infected fish may serve as intermediate hosts for these metacercariae for at least 2.5 years postinfection.     

肉毒梭菌毒素灭鼠研究进展

就肉毒梭菌毒素用于灭鼠的研究和应用情况进行了综述，充分肯定了C型肉毒梭菌毒素灭鼠制剂毒力强，适口性好，对非靶动物毒性低，作用缓慢，无2次中毒，易降解和适合于规模灭鼠的特点，为鼠害防治提供了新的思路。     

Effect of water temperature on the clinical outcome of infection with Edwardsiella ictaluri in channel catfish

Channel catfish fingerlings (mean body weight, 19 +/- 3 g each) were given intraperitoneal inoculations of Edwardsiella ictaluri suspensions of 10(4), 10(5), or 10(6) bacteria. Control fish were inoculated intraperitoneally with sterile 0.85% NaCl solution. Ten-day trials were conducted at water temperatures of 17, 21, 23, 25, 28, and 32 C. Differences in mortality between E ictaluri-infected fish and fish used as controls were observed at water temperatures of 23, 25, and 28 C, but not at temperatures of 17, 21, and 32 C. Clinical signs and lesions induced by intraperitoneal inoculation of E ictaluri were comparable with those found with the intestinal form of the natural disease. The characteristic erosion of skin and muscle overlying the skull, known as the "hole-in-the-head" lesion, was not observed. A given fish may be susceptible to infection at any water temperature, but a population is at risk when water temperatures are in the 22 to 28 C range.     

Olfactory Organ of Channel Catfish as a Site of Experimental Edwardsiella ictaluri Infection

Abstract

Edwardsiella ictaluri, the causative agent of enteric septicemia of catfish (ESC), is one of the most important pathogens to infect channel catfish Ictalurus punctatus. Although the full pathogenesis of E. ictaluri is unclear, the olfactory organ is thought to be a site of entry. We have examined the effects of applying E. ictaluri directly into the olfactory capsule of channel catfish. Olfactory organs of 30 experimental fish were exposed to E. ictaluri for 1 h (1 mL, 1 × 10⁶ colony-forming units/mL). Live fish were sampled at 1, 24, 48, and 72 h, and days 5 and 14 postinfection, and their olfactory organs were examined by light and electron microscopy. Damage, including loss of sensory cilia and microvilli from the olfactory mucosal surface, was observed at 1 h postinfection. Degeneration of olfactory receptors and supporting cells was evident by 24 h postinfection. The nonsensory region also showed signs of degeneration, such as columnar cells lacking cilia. Electron microscopic immunocytochemistry confirmed the presence of E. ictaluri on the mucosal surface and within the epithelium. Host leukocytes responded to bacteria by migrating through the olfactory epithelium into the interlamellar lumen and phagocytosing organisms, but phagocytosed E. ictaluri did not appear to be destroyed. Our results indicate that during initial stages of infection channel catfish olfactory epithelium is vulnerable, and E. ictaluri can enter the host through the olfactory organ. It is also possible that host phagocytic cells serve as a vehicle for the systemic dissemination of E. ictaluri     

Cool Weather Feeding Influences Responses of Channel Catfish to Edwardsiella ictaluri Challenge

Abstract

A pond study was conducted from November 1 through April 30 with young (age-0; average size, 43 g) and market-size (age-2; average size, 660 g) channel catfish Ictalurus punctatus to compare three management regimens: No feeding, partial feeding (no feeding in December, January, and February), and continuous feeding according to fish size and water temperature. Weight change, feed conversion ratio, and responses to experimental challenge with Edwardsiella ictaluri were evaluated. No significant difference in weight gain occurred between partially fed and continuously fed fish in either age-group; average weight increase for fed age-0 fish was 99% and for fed age-2 fish was 38%. The nonfed age-0 fish lost 12.3% and the nonfed age-2 fish lost 7% of their initial weight. Feed conversion ratios were significantly lower in partially fed fish than in continuously fed fish in both age-groups. Mortality from E. ictaluri challenge among age-0 fish was significantly higher in the nonfed fish, but among the age-2 fish, mortality was significantly lower in the nonfed fish. No difference in mortality rate occurred between partially fed and continuously fed fish in either age-group. Starvation induced lower antibody production against E. ictaluri antigen in the age-0 fish but higher antibody production in the age-2 fish. Phagocytic index was lower in nonfed fish than in fed fish from both age-groups. This study indicates no benefit from feeding age-0 and age-2 channel catfish during December, January, or February if feeding is reintroduced in March and continued through April. Although starvation was immunosuppressive in small channel catfish, it enhanced resistance to bacterial infection in larger fish. However, more research information is needed before reduced feeding can be recommended to enhance resistance of channel catfish to E. ictaluri.     

Parasitism by protozoan Ichthyophthirius multifiliis enhanced invasion of Aeromonas hydrophila in tissues of channel catfish

Protozoan Ichthyophthirius multifiliis Fouquet (Ich) and bacterium Aeromonas hydrophila are two common pathogens of cultured fish, which cause high fish mortality. Currently there is no information available for the effect of parasitism by Ich on survival of channel catfish and invasion of A. hydrophila in fish tissues following exposure to A. hydrophila. A trial was conducted in this study to: (1) determine whether A. hydrophila increased fish mortality in Ich-parasitized channel catfish; and (2) compare the bacterial quantity in different tissues between non-parasitized and Ich-parasitized catfish by real-time polymerase chain reaction (qPCR). The results demonstrated that the Ich-parasitized catfish showed significantly (P<0.05) higher mortality (80%) when exposed to A. hydrophila by immersion than non-parasitized fish (22%). Low mortality was observed in catfish exposed to Ich alone (35%) or A. hydrophila alone (22%). A. hydrophila in fish tissues were quantified by qPCR using a pair of gene-specific primers and reported as genome equivalents per mg of tissue (GEs/mg). Skin, gill, kidney, liver and spleen in Ich-parasitized fish showed significantly higher load of A. hydrophila (9400-188,300 GEs/mg) than non-parasitized fish (4700-42,100 GEs/mg) after exposure to A. hydrophila. This study provides evidence that parasite infections enhance bacterial invasion and cause high fish mortality.     

Distribution of Edwardsiella ictaluri in California

Abstract

Wild and domestic populations of channel catfish Ictalurus punctatus were examined to determine the distribution of the disease called enteric septicemia of catfish (ESC) in California. The causative agent of ESC, Edwardsiella ictaluri, was isolated from five separate sites in California. Two of these isolations were from rectal swabs of asymptomatic fish, confirming that a carrier state may exist. Normal-appearing fish with serum antibody titer to E. ictaluri were commonly found in domestic channel catfish populations, suggesting that many fish become infected but recover. Wild channel catfish with antibody to E. ictaluri were also found in major reservoirs and water distribution canals. Edwardsiella ictaluri appears to be widely distributed within California.     

Communications: Specificity of the Channel Catfish Antibody to Edwardsiella ictaluri

Abstract

The specificity of channel catfish Ictalurus punctatus serum antibody to Edwardsiella ictaluri was characterized by microtiter agglutination assay. There was no correlation between antibody titer to Aeromonas hydrophila and antibody titer to E. ictaluri in wild or feral channel catfish. Anti-E. ictaluri antibodies in naturally infected channel catfish were not removed by adsorption by nine other species of bacteria found in the channel catfish intestine and fish ponds. Channel catfish immunized with nine other species of bacteria did not develop substantial antibody titer to E. ictaluri. The antibody response of channel catfish to E. ictaluri is highly specific, and the microtiter agglutination test is a specific indicator of previous exposure to E. ictaluri