Validation of Fermentation and Processing Procedures for the Commercial-Scale Production of a Live,Attenuated Edwardsiella ictaluri Vaccine for Use in Channel Catfish Aquaculture

Abstract

Mortality associated with Edwardsiella ictaluri infection is a serious impediment to the commercial production of fingerling Channel Catfish Ictalurus punctatus. A patented, live, attenuated, orally delivered vaccine has been developed that offers exceptional protection against E. ictaluri infection in both laboratory and small-scale pond trials. Further vaccine development is contingent on the successful completion of large-scale field trials that accurately reflect industry conditions. This current work focuses on the validation of fermentation protocols and the optimization of downstream processing procedures to produce sufficient quantities of vaccine to conduct commercial-scale field trials. Eight vaccine serials were produced from a master seed stock (S97-773-340X2) in a 50-L floor model fermenter over two consecutive years. Following fermentation, cells were harvested, concentrated 10-fold, and cryogenically stored (?74°C). To assess processing protocols and determine shelf life of cryogenically stored vaccine, serials were tested for cell viability and vaccine potency at various intervals over 24 months. There were no significant differences in cell viability between the fresh vaccine and the stored frozen product. All serials provided a high level of protection (77–100% relative percent survival) against E. ictaluri infection in juvenile Channel Catfish and exhibited excellent poststorage viability. This data demonstrates that the live, attenuated, orally delivered vaccine can be stored at ?74°C for at least 2 years with no reduction in cell viability or vaccine potency.

Received May 17, 2016; accepted January 19, 2017 Published online April 4, 2017     

Pulmonary cryptococcosis in slaughtered sheep: anatomopathology and culture

Cryptococcosis is considered a zoonosis but is rare in stock animals. In humans it is imputed to be a mortality cause in immunocompromised patients. This case study reports the anatomopathology of respiratory cryptococcosis in ovine and the recuperation and identification of Cryptococcus neoformans var. neoformans from 1-year frozen lung tissue sample. The findings from haematoxylin/eosin staining microscopy and special staining showed the destruction of the columnar respiratory epithelium architecture associated to suppuration, and the growth of spherical yeast buddings sometimes involved by clear halo produced by unstained capsular substance. The agent was identified by miniApi ID32C, while variety neoformans was further identified by culture in CGB agar. The diagnosis in stock animal suggests the involvement of an exogenous source of the agent as an alert to public health and contributes for epidemiological investigations.     

Teaching veterinary radiography by e-learning versus structured tutorial: a randomized, single-blinded controlled trial

Case-based e-learning may allow effective teaching of veterinary radiology in the field of equine orthopedics. The objective of this study was to investigate the effectiveness of a new case-based e-learning tool, compared with a standard structured tutorial, in altering students' knowledge and skills about interpretation of radiographs of the digit in the horse. It was also designed to assess students' attitudes toward the two educational interventions. A randomized, single-blinded, controlled trial of 96 fourth-year undergraduate veterinary students, involving an educational intervention of either structured tutorial or case-based e-learning, was performed. A multiple-choice examination based on six learning outcomes was carried out in each group after the session, followed by an evaluation of students' attitudes toward their session on a seven-point scale. Text blanks were available to students to allow them to comment on the educational interventions and on their learning outcomes. Students also rated, on a Likert scale from 1 to 7, their performance for each specific learning outcome and their general ability to use a systematic approach in interpreting radiographs. Data were analyzed using the Mann-Whitney test, the t-test, and the equivalence test. There was no significant difference in student achievement on course tests. The results of the survey suggest positive student attitudes toward the e-learning tool and illustrate the difference between objective ratings and subjective assessments by students in testing a new educational intervention.     

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.     

Fusariotoxicosis from barley in British Columbia. II. Analysis and toxicity of syspected barley.

Fusariotoxin T-2, a trichothecene, was tentatively identified in barley samples which caused field outbreaks of mycotoxicosis in British Columbia. Geese died when fed the contaminated barley experimentally but mice were little affected after long term feeding. The methods used in the laboratory for trichothecene extraction and identification of T-2 toxin are described.     

Comparative Susceptibility of Channel Catfish,Blue Catfish,and their Hybrid Cross to Experimental Challenge with Bolbophorus damnificus (Digenea: Bolbophoridae) Cercariae

Abstract

The digenetic trematode Bolbophorus damnificus has been implicated in significant losses in catfish aquaculture since the late 1990s. The complex life cycle sequentially involves the American white pelican Pelecanus erythrorhynchos, the marsh rams horn snail Planorbella trivolvis, and Channel Catfish Ictalurus punctatus. Research supports anecdotal reports from the industry, suggesting that the hybrid of Channel Catfish×Blue Catfish I. furcatus is less susceptible to disease agents that have been historically prohibitive to Channel Catfish production, namely the gram-negative bacteria Edwardsiella ictaluri and Flavobacterium columnare, as well as the myxozoan parasite Henneguya ictaluri. This current research compared the susceptibility of Channel Catfish, Blue Catfish, and their hybrid cross to an experimental challenge by B. damnificus. Fish were exposed to 0, 100, 200, and 400 B. damnificus cercariae per fish, and the numbers of metacercariae per fish were determined 14 d postchallenge. Metacercariae were recovered from all challenged fish. There were no significant differences among fish groups challenged with the same dose, suggesting Channel and Blue Catfish and their hybrid are comparably susceptible to B. damnificus infection. As such, it is recommended that producers raising hybrid catfish remain diligent in controlling populations of the snail intermediate host to prevent production losses attributed to B. damnificus, especially when loafing pelicans have been observed at the aquaculture operation.

Received October 22, 2013; accepted January 5, 2014     

Case control study to investigate risk factors for impaction colic in donkeys in the UK

Impaction colic is the single most common type of colic diagnosed in a large population of donkeys (more than 2000 animals) at The Donkey Sanctuary, UK and the fatality rate from the disease is high. Here we identify risk factors for impaction colic in this population during 2006 using an unmatched case control study. There were 71 cases of impaction colic and multivariable analysis identified a number of variables associated with the disease. Management factors that increased the risk of impaction included paper bedding, feeding of concentrates, limited access to pasture and increasing number of carers. In addition health variables that were associated with an increased risk of impaction colic were weight loss, recent vaccination and a number of dental abnormalities. This study has identified variables which may help to identify donkeys at high risk of impaction colic; those with a history of weight loss and with concurrent dental pathology. Furthermore the study has also identified a number of variables that may be targeted to reduce the incidence of impaction colic in this donkey population, such as modification of concentrate feeding practices and pasture access.     

Comparison of two DNA extractions and nested PCR, real-time PCR, a new commercial PCR assay, and bacterial culture for detection of Mycobacterium avium subsp. paratuberculosis in bovine feces.

In this study, 5 combinations of 2 DNA extractions and 3 polymerase chain reaction (PCR) techniques were compared with culture for the detection of Mycobacterium paratuberculosis directly from bovine feces. These combinations included a new commercial extraction technique combined with a commercial PCR/Southern blot technique, nested PCR (nPCR), or real-time PCR, and a university-developed extraction combined with nPCR or real-time PCR. Four of the 5 combinations had statistically similar sensitivities between 93% and 100% and specificity between 95% and 100%, when compared with culture results from 63 bovine fecal samples. These results indicated that using a commercial extraction with a commercial PCR/Southern blot, nPCR, or real-time PCR, or a university-developed extraction with real-time PCR would result in similar sensitivities to culture for the identification of M. paratuberculosis from bovine feces and are valid alternatives to culture.