Vibrio Vaccine Research Articles

Advancements in Vibrio vaccines for aquaculture

Advancements in Vibrio vaccines for aquaculture

Use of probiotics, mangrove leaf extract, and bacterin for alternative disease prevention in tiger shrimp culture

Research has been carried out to determine the effect of probiotics, mangrove leaf extracts, and bacterin as an alternative to disease prevention in shrimp culture. There are 4 treatments and 5 replications. WSSV-free tiger shrimp juveniles with a density of 5 pcs/m2 were applied in each pond. A) used of RICA probiotic (1,2,3) 20g/pond; B), used of mangrove extracts resulting from wet leaf boiling (Sonneratia alba) (300 g in 5 L of cooking water to 2.5 L) as much as 2 L/pond; C) used of vibrio vaccine formalin-inactivated 1% with dose 2.4 mL /kg feed; and D) Control. Administration of probiotics and mangrove leaf extract is carried out every week while bacterin is carried out before culture with immersion method at a dose of 2.4 mL/L for one hour and booster through feed at a dose of 2.4 mL/kg of feed given on days 13, 14, 15, 28, 29 and 30 every month during the culture period. The results showed that the use of mangrove leaf extracts gave the best survival rate and production of the shrimp. TBV/TPC ratio in the pond water ranged from 1.3-8.9%, while in the pond sediment ranged from 0.33-0.97%.

This Month's Highlights.

<h3>Summary</h3> 1.The placental transmission of cholera antibodies from mother to infant rabbits has been demonstrated. 2.Ten-day-old rabbits derived from mothers immunized with live vibrio vaccine were immune to cholera infection, but those from mothers immunized with heat-killed or formolized vaccines were not. 3.The agglutinin titer gave no indication of the protective power against vibrio challenge. 4.The technique of placental transmission of antibodies could be used for the standardization of cholera vaccine. 5.The use of live vibrio vaccine as a cholera prophylactic is suggested.

Development of Recombinant Dihydrolipoamide Dehydrogenase Subunit Vaccine against Vibrio Infection in Large Yellow Croaker

Large yellow croaker (Larimichthys crocea), an economically important marine fish in China, has suffered from serious vibriosis, which has resulted in great economic losses for the large yellow croaker industry. Vaccination has been considered to be a safe and effective method to prevent and control vibriosis. However, due to the complex diversity and serotypes of the Vibrio genus, the progress of Vibrio vaccine development is still slow. In this study, we prepared recombinant Vibrio dihydrolipoamide dehydrogenase (rDLD) protein and investigated its potential as a candidate to be a subunit vaccine against Vibrio. The lysozyme activity and the rDLD-specific antibody level in sera of large yellow croakers immunized with rDLD were significantly higher than those in the control group, and the transcript levels of proinflammatory cytokines (IL-6, IL-8, IL-1β), MHC IIα/β, CD40, CD8α, IL-4/13A, and IL-4/13B were significantly up-regulated in the spleen and head kidney of large yellow croakers immunized with rDLD, suggesting that rDLD could induce both specific and nonspecific immune responses in this species. In addition, rDLD protein increased the survival rate of large yellow croakers against Vibrio alginolyticus and Vibrio parahaemolyticus, with the relative percent of survival (RPS) being 74.5% and 66.9%, respectively. These results will facilitate the development of a potential subunit vaccine against Vibrio in large yellow croaker aquaculture.

Laboratory and Field Assessments of Oral Vibrio Vaccine Indicate the Potential for Protection against Vibriosis in Cultured Marine Fishes

Simple SummaryThe application of chemicals and drugs in fish culture has increased the existence of antimicrobial-resistant bacteria and drug residues in fish. Developing an effective vaccine to reduce specific disease-related losses and avoid antibiotic use has become an increasingly important part of aquaculture. Our current study found that the oral inactivated V. harveyi strain VH1 vaccine confers high protection in fish post-infection with virulent Vibrio spp. under experimental conditions. The oral vaccine was further tested for field efficacy and showed a higher survival and growth performance in vaccinated fish than unvaccinated fish. This current vaccine was shown to potentially provide sufficient protection to the host in both controlled and field environments against vibriosis.Vibriosis is one of the most common threats to farmed grouper; thus, substantial efforts are underway to control the disease. This study presents an oral vaccination against multiple Vibrio spp. in a marine fish with double booster immunisation. The Vibrio harveyi strain VH1 vaccine candidate was selected from infected groupers Epinephelus sp. in a local farm and was formalin inactivated and combined with commercial feed at a 10% ratio (v/w). A laboratory vaccination trial was conducted for seventy days. The induction of IgM antibody responses in the serum of Asian seabass Lates calcarifer immunised with the oral Vibrio harveyi strain VH1 was significantly (p < 0.05) increased as early as week one post-primary vaccination. Subsequent administration of the first and second booster for 5 consecutive days, starting on days 14 and 42, respectively, improved the specific antibody level and reached a highly significant (p < 0.05) value at days 35 and 49 before slightly decreasing from day 56 onwards. Antibody titres of the control unvaccinated group remained relatively stable and low throughout the experimental period. At the end of the 70-day vaccination trial, 23 days post final boost, an intraperitoneal challenge with a field strain of Vibrio harveyi, V. alginolyticus, and V. parahaemolyticus was carried out. Our challenge study showed that oral Vibrio harveyi strain VH1 vaccine candidate could induce significant protection, with an RPS of 70–80% against different Vibrio species. Thereafter, a field trial was conducted in a mariculture farm to study the effect of field vaccination using the oral Vibrio harveyi strain VH1 vaccine candidate. A total of 3000 hybrid grouper juveniles were divided into two groups in triplicate. Fish of Group 1 were not vaccinated, while Group 2 were vaccinated with the feed-based vaccine. Vaccinations were carried out on days 0, 14, and 42 via feeding the fish with the vaccine at 4% body weight for 5 consecutive days. At the end of the study period, the fish survival rate was 80% for the vaccinated group, significantly (p < 0.05) higher than the 65% seen in the control unvaccinated group. Furthermore, the vaccinated fish showed significantly (p < 0.05) better growth performances. Therefore, the oral Vibrio vaccine from the inactivated Vibrio harveyi strain VH1 is a potential versatile vaccine candidate that could stimulate good immune responses and confer high protection in both Asian seabass, Lates calcarifer, and farm hybrid grouper Epinephelus fuscoguttatus × Epinephelus lanceolatus.

Study on Immune Response of Organs of Epinephelus coioides and Carassius auratus After Immersion Vaccination With Inactivated Vibrio harveyi Vaccine.

Immersion vaccination relies on the response of fish mucosa-associated lymphoid tissues, the Crucian carp (Carassius auratus) and Grouper (Epinephelus coioides) were researched in this paper to examine local mucosal immune responses and associated humoral system responses following immersion vaccination. We administered 1.5 × 107 CFU/ml formalin-inactivated Vibrio harveyi cells and measured mucus and serum antibody titers as well as IgM, MHC II mRNA levels in immune organs. The mucosal antibody response preceded the serum response indicating a role for local mucosal immunity in immersion vaccination. IgM and MHC II mRNA levels were relatively greater for the spleen and head kidney indicating the importance and central position of systemic immunity. Expression levels were also high for the gills while skin levels were the lowest. IgM and MHC II mRNA levels were altered over time following vaccination and the hindgut, liver and spleen were similar indicating a close relationship, so the absolute value of r is used to analyze the correlation among different organs immunized. It can be inferred the existence of an internal immune molecular mechanism for Immune synergy hindgut-liver-spleen, from the peak time (14th day), the relative ratio of genes expression in the same tissues between the immunized grouper and the control group (26 times), and Pearson correlation coefficient (0.8<|r|<1). Injection challenges with live V. harveyi indicated that the relative protection rates for the crucian carp and Grouper was basically the same at 44.4% and 47.4%, respectively. It is believe that crucian carp may be used as a substitute for the valuable grouper in immunity experiment, just from aspect of the relative percent survival (RPS) and how it changes with time. But they were not consistent about the IgM mRNA expression between that of crucian carp and grouper after immersion the Vibrio vaccine.

Efficiency of monovalent and polyvalent Vibrio alginolyticus and Vibrio Parahaemolyticus vaccines on the immune response and protection in gilthead sea bream, Sparus aurata (L.) against vibriosis

This experimental studies investigated the protective efficiencies and the potential immune mechanisms of vibrio monovalent and polyvalent autogenous formalin-inactivated whole-cell bacterins (FKC) in Gilthead sea bream (Sparus aurata) cultured in Egypt. Two months post-vaccination, the relative percentage survival (RPS) was estimated after challenge with the vaccine's homologues pathogenic strains. The survival values were 100% and 83.3% in groups immunized with monovalent V. alginolyticus or V. parahaemolyticus FKC bacterins, respectively. On the other hand, survival values were 91.75% and 75% in fish groups subjected to polyvalent (V. parahaemolyticus O11: K40 &V. alginolyticus) and (V. parahaemolyticus O3: K6 &V. alginolyticus) FKC bacterins, respectively. Overall, the tested vaccine preparations were significantly increased (P < 0.05) the agglutination antibody titer, phagocytic activity, respiratory burst activity, when compared to the non-immunized control group.The current results conclude that, autogenous Vibrio vaccines provoked a promising protection against vibriosis in Gilthead sea bream cultured in Egypt, it was superior in monovalent FKC V. alginolyticus vaccine and polyvalent FKC of V. parahaemolyticus O11: K40 with V. alginolyticus vaccine that could be useful means of prevention and control of vibriosis.

Global mapping of cholera Vibrio and outbreaks in the Pre-Millennium Development Goals (MDG)/Sustainable Development Goals (SDG) and MDGs/SDGs era of 1990–2019

Vibrio species and cholera outbreak yet remain a frequent health emergency despite progress made in integrated implementation of the MDGs/SDGs/WASH worldwide. Hence, this study aimed at appraising the impacts of MDGs/SDGs/WASH campaigns on the mitigation of cholera outbreak and associated consequences. The study mapped scientific production related to Vibrio outbreak from 1990 to 2019, identified trend, institutional/international concerted efforts toward outbreak research/response and gaps for future preparedness. Relevant documents were identified from the Web of Science database using an optimised title-field specific search Boolean that accommodated all pre-set inclusion criteria for the study. A total of 901 documents were identified including 869 available abstracts were retrieved for content-review of human incidence cases, mortality, culprit Vibrio species, strains, and biotypes. Explanatory analysis showed that the trend of outbreak documents approximately increased in 6th order quadratic relationship (R2 = 0.7948) from 1990 to 2019 with an annual growth rate of 3.21% and a mean value of 30.0 ± 18.0 per year. Other details revealed an increased and undulating case report/mortality rate of cholera outbreaks especially in the MDGs/SDGs era. Decadal comparison of Vibrio outbreak during the period showed significant variation in documents distribution (Kruskal-Wallis p = 0.00077). based on countries’ efforts, the USA, ranked first in terms of article numbers (191), publication frequency (24.6(%) and total citations (5962). Four prevailing conceptual frameworks were identified in the outbreak documents with global community interest revealed as the largest topical coverage. All conceptual frameworks consisted in Vibrio characterisation, methodology-related, intervention-related, geographic-related concepts and some replete with health and climate-change depicting concepts. Also, the study observed high mortality in Vibrio outbreaks during 1990–1999 (29080 deaths), and 2010–2019 (386606 deaths) compared to 2000–2009 (7705 deaths) (Kruskal-Wallis, p < 0.05). High number of outbreaks due to V. cholerae and V. parahaemolyticus and a limited outbreaks attributed to emerging strains. In conclusion, vibrio outbreak has not lived up to various investment put into its control from various programme evolutions. The broad spectrum Vibrio vaccines that could cater for outbreak caused by common and emerging strains is inevitable and a significant thrust for future research.

A test of the effects of androgens on immunity: No relationship between 11-ketotestosterone and immune performance in bluegill (Lepomis macrochirus)

The immunosuppressive effects of androgens are a key component of the immunocompetence handicap hypothesis (ICHH). Here, we use bluegill sunfish (Lepomis macrochirus) to test two predictions arising from this hypothesis: (1) natural circulating concentrations of the androgen 11-ketotestosterone (11-KT) will be negatively related with measures of immunity, and (2) immune stimulation will lower circulating 11-KT concentration. We found no evidence for a relationship between natural circulating 11-KT concentration and measures of immunity (lymphocyte and granulocyte counts, respiratory burst, cytokine mRNA levels), and an immune stimulation with Vibrio vaccine did not affect circulating 11-KT concentration. We also performed a meta-analysis of immune stimulation studies to help interpret our results, and report evidence suggesting that immune stimulation has weaker effects on androgen levels in fishes compared to other vertebrates. These results suggest that the ICHH may not apply to all vertebrates, although it remains premature to state what factors account for the weaker evidence in fishes that androgens are immunosuppressive.

Modulation of innate immune responses in Atlantic salmon by chronic hypoxia-induced stress

Atlantic salmon post-smolts were exposed to either chronic hypoxic (Hy) or normal oxygen (No) conditions in seawater tanks for 58 days, mimicking conditions typical of sea cages for farmed salmon at some periods of the year. By day 29 head kidney macrophages were isolated and subjected to in vitro poly I:C stimulation to simulate viral infection, and samples were collected over 48 h. By day 58 fish were subjected to in vivo stimulation using poly I:C or a Vibrio water-based vaccine to simulate viral or bacterial infection, respectively. The fish were monitored for stress responses and expression of several pro-inflammatory genes in head kidney and intestinal tissue up to five days post-injection. Stress load was monitored by plasma cortisol estimation at days 29 and 58, and on days 1, 2, 3 and 5 post-injection in the in vivo trial. Hy exposure resulted in elevated plasma cortisol levels on day 29 compared to No, while on day 58 cortisol levels were higher in the control group. Additionally, both poly I:C and the Vibrio vaccine gave significantly increased cortisol levels one day post-injection compared to PBS treated controls, irrespective of previous oxygen exposure. In vitro stimulation of macrophages with poly I:C revealed higher IFNα mRNA levels at 6, 12 and 24 h and for Mx at 12 and 24 h post-stimulation, for both No and Hy individuals. Moreover, IFNα levels were higher in No than in Hy individuals at all time points, and a similar difference was seen in Mx at 48 h. In vivo stimulation with poly I:C elicited strong elevation of the IL-1β, IFNγ, Mx and IP10 mRNA transcripts in head kidney, while TNFα1 and IFNα were found unaffected. The Vibrio vaccine elicited a strong up regulation of IL-1β, IFNγ and IP10 mRNA, whereas Mx, TNFα1 and IFNα appeared unchanged. Significant differences in expression between different oxygen exposure groups were found for all genes and both stimuli. The overall trend suggests that long-term hypoxia either reduces or delays the expression of these genes in head kidney. Expression of IFNγ and Mx in intestinal tissues also showed a strong up regulation of the genes following poly I:C stimulation, and also here the overall trend suggests that chronic hypoxia results in a lower or delayed expression of the measured genes.In summary, our results indicate that chronic hypoxia modulates the expression of important immune related genes putatively altering the immune response. As the effect is present in isolated macrophages as well as head kidney and intestinal tissue the modulation appears to be affecting local as well as systemic responses.

Additive, non-additive and maternal effects of cytokine transcription in response to immunostimulation with Vibrio vaccine in Chinook salmon (Oncorhynchus tshawytscha)

Estimation of quantitative genetic parameters is important for improving salmonid broodstock management in commercial and government hatcheries. Using a replicated 2 × 2 factorial breeding design (48 families and 192 individuals), we partitioned early immune response transcription variation into additive genetic, non-additive genetic, and maternal components in juvenile Chinook salmon (Oncorhynchus tshawytscha). Transcription of four cytokine genes (IL1, TNF-α, IL-8, IL8-R) and two control genes (IgM and RPS-11) was measured relative to an endogenous control (EF1a) before and 24 h after immune stimulation with Vibrio vaccine. Additive genetic variation was not significant for cytokine transcription and heritability ranged from 0.44 (in pre-challenge IL1) to 0.04 (in post-challenge TNF-α). Non-additive genetic variance was significant in post-challenge IL1 (18 %) and TNF-α (12 %) while maternal effects contributed to pre-challenge cytokine transcription. Cytokine transcription co-expressed within but not between pre- and post-challenge states. The lack of additive genetic effects indicates that cytokine transcription is not a likely candidate for selection programs to improve immune function in Chinook salmon. Our results add to the growing evidence that non-additivity in salmon is common and contributes to our understanding of the genetic architecture of transcription. This indicates that transcription variation may act to maintain genetic variation and facilitate rapid adaptive response in salmonids.

Enhanced immune defences in Pacific white shrimp (Litopenaeus vannamei) post-exposure to a vibrio vaccine

This study was conducted to determine if exposure of shrimp, Litopenaeus vannamei, to a commercial anti-vibrio vaccine caused changes in antibacterial and cellular (phagocytosis) defences. Shrimp post-larvae were administered either Vibromax™ vaccine or a blank preparation. Whole body homogenates were prepared before (day 0), during (day 10) and after (day 20) vaccination and incubated with a selection of pathogenic vibrios. Homogenate from day 0 animals showed natural antibacterial activity towards Vibrioanguillarum which was significantly enhanced for bacteria-exposed shrimp at 10days post-challenge. This effect of the vaccine was short-term in its duration. No antibacterial activity was observed in day 0 shrimp homogenate against Vibrio alginolyticus but it was significantly enhanced for both vaccinated and blank-vaccinated shrimp by day 10. No natural or inducible antibacterial activity was observed against Vibrio harveyi at 0, 10 or 20days post-challenge. To determine if prior exposure of shrimp to inactivated vibrios results in elevated hemocyte phagocytic activity, juveniles were injected with either a mixture of formalin-inactivated vibrios or saline. Hemocyte monolayers made from these shrimp were overlaid with a 1:1 mix of Bacillus subtilis and these vibrios. Hemocytes from vibrio-exposed animals showed elevated levels of internalised vibrios compared with those from the saline injected group. These studies show selectively enhanced cellular defences of shrimp following ‘vaccination’.

Variability among Vibrio anguillarum O2 isolates from Atlantic cod ( Gadus morhua L.): Characterisation and vaccination studies

Farmed Atlantic cod, Gadus morhua L., in Norway are vaccinated against Vibrio anguillarum serotypes O1, O2a and O2b, but outbreaks of classical vibriosis remain relatively common. In the present study, V. anguillarum isolated from vaccinated and unvaccinated cod between 2002 and 2005, were shown to differ in biochemical, serological and genetic characteristics compared to V. anguillarum serotypes O2a and O2b. They did not utilize sorbitol and some were shown to be lysine decarboxylase positive or did not produce gelatinase. Furthermore, they displayed varying reaction patterns with monoclonal antibodies, and appeared in a separate cluster in a similarity dendrogram based on amplified fragment length polymorphisms (AFLP) analysis, distant from the cluster containing both serotypes O2a and O2b. The results indicate that these V. anguillarum isolates belong to a sero-subtype other than O2a or O2b. The efficacy of ALPHA MARINE™ Vibrio vaccine in cod varied depending on the V. anguillarum O2 isolate used for challenge. The vaccine elicited good protection against bath challenge with isolates of V. anguillarum O2b and O2a (RPS > 80) comparable to the vaccine strains, but protected poorly (RPS of 32) or not at all when challenged with two recent and differing isolates from diseased vaccinated cod.

The influence of dietary ß-glucans on the adaptive and innate immune responses of European sea bass (Dicentrarchus labrax) vaccinated against vibriosis

The effects of feeding 1,3/1,6 β-glucans on the innate and the adaptive immune responses of European sea bass (Dicentrarchus labrax) was investigated. Two experiments were carried out during the study. In the first, a number of non-specific immune parameters were examined at 4, 7, 10, 14, 21 and 25 days of feeding fish with a semipurified diet containing Macrogard ©, a commercially available form of 1,3/1,6 β-glucans. The respiratory burst activity of head kidney macrophages isolated from the different groups of fish fed the immunostimulant peaked and subsequently decreased at different times during the experiment. Head kidney macrophages from fish fed 250 ppm β-glucans had a statistically higher level of respiratory burst activity at Day 21 of feeding compared with fish fed no immunostimulant. No statistical differences were observed in lyzozyme activity during this trial.In the second experiment, the effect of feeding 1,3/1,6 β-glucans on the immune response of fish to an alginate-encapsulated Vibrio vaccine administered orally was examined. Respiratory burst of head kidney macrophages and serum lysozyme activity decreased in all fish over the course of the trial, while serum lysozyme activity was considerably lower than values obtained in the first experiment. Fish vaccinated orally had significant increases in antibody response by Week 2 post-vaccination, but β-glucans did not appear to affect these levels. Vaccination may have resulted in activating the immune system as a whole, thus masking any difference in immunostimulation by the β-glucans. It may be that the optimal doses and timing of β-glucans administration is different when the immunostimulant is administered alone or in combination with the vaccine. In conclusion, European sea bass can be immunomodulated with oral administration of β-glucan. Optimal doses and administration times have been established when β-glucans are fed alone, although further studies are needed to establish ideal feeding regimes for glucans administrated in combination with vaccination.

EFEKTIVITAS VAKSIN POLIVALEN UNTUK PENGENDALIAN VIBRIOSIS PADA KERAPU TIKUS (Cromileptes altivelis)

The objective of this research was to know the effectiveness of polyvalen Vibrio vaccine to control vibriosis in humpback grouper (Cromileptes altivelis). The effectiveness of vaccine was evaluated by the survival rate (SR), relative percent survival (RPS), mean time to death (MTD) as well as growth rate of vaccinated fish. This research consisted of 4 treatments (control, injection, immersion, and oral vaccinations) in quadruplicates. Injection vaccination was conducted by intraperitoneal injection of polyvalen vaccine at 107 cells/fish. Immersion vaccination was done by immersing the fishes at 107 cells/ml for 30 minutes. Oral administration of vaccine was also carried out at 107 cells/fish. One week after the first vaccination, second vaccination (booster) was carried out at the same dosage and by the same administration. One week after the second vaccination, fishes were challenged with 3.16x104 cells/fish of Vibrio ordalii 3J by intraperitoneal injection, and reared for 20 days post infection. Results indicated that polyvalen Vibrio vaccine increased SR (P&lt;0.01) up to 100%. Vaccination was also able to delay MTD of fishes. However, the vaccination was not influence the growth rate of fish.

Age related changes in humoral immune response of hybrid striped bass (Morone chrysops x Morone saxatilis).

A number of factors can influence immune function affecting the magnitude and duration of specific immune responses. One factor which has been noted to affect immune responses is age of animal. In mammals, juveniles have a lower immune response when compared to adults. In fish, fry have a lower immune response than adults; however, few studies have investigated the immune response in juveniles, the age when most fish are reared in aquaculture settings. The humoral immune responses of hybrid striped bass (Morone saxatilis x Morone chrysops) at five different ages were compared to determine any maturational changes. Fifty fish were bath immunized with a commercial Vibrio vaccine at 4, 6, 9, 15, and 19 months of age. The antibody response in these fish was monitored by an enzyme-linked immunosorbent assay (ELISA) for 106 days post-vaccination. The magnitude, duration, and time to peak level were compared to controls and between the different ages using analysis of variance (ANOVA). Younger fish exhibited significantly lower antibody levels indicating that juvenile fish may not be able to mount as effective an immune response as young adult fish.

Symposium “Current Status of Fish Vaccine Development in Japan”

A symposium entitled “Current status of fish vaccine development in Japan” was held by the Japanese Society of Fish Pathology in Tokyo on 29th March, 2001. This was aimed to have an overview of the status quo of fish vaccine developments in Japan, especially those for marine fishes. In Japan, a vibrio vaccine to ayu Plecoglossus altivelis was first developed in 1988 and five vaccines have been totally developed until now. In this symposium, ten subjects were included in five categories : (1) Vaccine-use system, utilization and practical problems, (2) Vaccines developed or being developed, (3) Genetic engineering techniques and adjuvants for vaccines (4) General control methods to prevent fish diseases, and (5) Problems confronting the development of fish vaccines in Japan. Vaccines will contribute to make safe products in aquaculture, which can give “peace of mind” to consumers. Vaccines developed in Japan have been effective to protect fish from the target diseases and it was emphasized that fish condition should be kept healthy to get a good result by vaccination.

Prolonged Immersion Improves the Effectiveness of Dilute Vibrio Vaccine for Rainbow Trout.

In order to clarify the effect of a prolonged exposure period on vaccine effectiveness, juvenile rainbow trout were immersed in 100, 1, 000 and 10, 000 times dilutions of a commercial vibrio bacterin for 3 min (brief immersion, BI) and for 24 h (prolonged immersion, PI). Two weeks and 2, 4 and 6 mo after the vaccination, fish were challenged by waterborne exposure to Vibrio ordalii. Prolonged exposure to dilute bacterin resulted in better protection than the standard BI method. Two weeks after vaccination, the cumulative mortality was 0% in the group vaccinated by PI with the 100, 1, 000 and 10, 000 times diluted vaccine. In contrast, mortality was 58 and 77% in the groups vaccinated by BI with the 100 and 10, 000 times diluted vaccine respectively, and was 100% in the unimmunized control group. Furthermore, 6 mo after vaccination, the mortality was 20% (100 times dilution) to 77% (10, 000 times dilution) in the PI group, while it was 64% (100 times dilution) to 93% (10, 000 times dilution) in the BI group. Mortality in the corresponding control group was 97%. These results suggest that prolonged exposure period improves vaccine effectiveness.

Uptake of Vibrio anguillarum vaccine by Artemia salina as a potential oral delivery system to fish fry

An ELISA to detect Vibrio anguillarum antigen in Artemia salina was developed. This assay was then used to determine the time course of antigen uptake by Artemia during incubations with Vibrio vaccine with a view to using Artemia as a vaccine carrier for oral vaccination of fish fry. The rate of uptake of vaccine by Artemia was shown to be concentration dependent. At a dose of 1·5 × 10 7 cfu ml -1, 1 mg wet weight of Artemia took up 1 × 10 5 cfu within 30 min and reached maximum of 6 × 10 6 cfu after 60 min. Thus each Artemia appears to have taken up approximately 1·2 × 10 5 cfu. Comparison of vaccine formulations indicated that uptake was consistently higher with formalised than with heat-treated vaccine.

Communications: Protective Immunity Induced by a CommercialVibrioVaccine in Hybrid Striped Bass

Abstract Commercial Vibrio anguiliarum-V. ordalii bacterin was used to vaccinate hybrid striped bass (Morone saxatilis ♀ × M. chrysops ♂) to test the vaccine efficacy against vibriosis. Vaccination by direct immersion of fish in diluted Vibrio vaccine for 20 s resulted in increased protective immunity. The relative percent survival of hybrid striped bass challenged 35 d after vaccination was 66.7% for those challenged by 1-h immersion exposure to 7.03 × 107 V. anguillarum cells/mL, 75.0% for those challenged by injection with 3.51 × 105 cells/fish, and 86.7% for those challenged by injection with 3.51 × 104 cells/fish.