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The use of probiotics in shrimp aquaculture.

Shrimp aquaculture, as well as other industries, constantly requires new techniques in order to increase production yield. Modern technologies and other sciences such as biotechnology and microbiology are important tools that could lead to a higher quality and greater quantity of products. Feeding and new practices in farming usually play an important role in aquaculture, and the addition of various additives to a balanced feed formula to achieve better growth is a common practice of many fish and shrimp feed manufacturers and farmers. Probiotics, as 'bio-friendly agents' such as lactic acid bacteria and Bacillus spp., can be introduced into the culture environment to control and compete with pathogenic bacteria as well as to promote the growth of the cultured organisms. In addition, probiotics are nonpathogenic and nontoxic microorganisms without undesirable side-effects when administered to aquatic organisms. These strains of bacteria have many other positive effects, which are described in this article.

Animals↗

Draft genome sequence of Bacillus thuringiensis GIFSPR-111, a putative probiotic isolated from shrimp farm soil in Bangladesh.

Bacillus thuringiensis GIFSPR-111, isolated from shrimp farm soil in Shamnagar, Bangladesh, inhibits Vibrio parahaemolyticus, the causative agent of acute hepatopancreatic necrosis disease in shrimp and exhibits probiotic potential. The 5,608,442 bp draft genome contains multiple biosynthetic gene clusters, with potential to synthesize diverse bioactive metabolites.

Bacillus thuringiensis↗

Role of probiotics on the environment of shrimp pond.

Recent disease outbreak in shrimp farming caused mainly by bacteria, virus, fungi or a combination of these etiologic agents is attributed to disturbance in the environment of pond. To combat this, different antibiotics and chemicals are being used which are reported to be not environment friendly. Of late, a new and unique biotechnological product called "Probiotics " is being used widely by all the shrimp farmers worldwide, which is found to be more effective and environmentally safe also. In the present study 2 probiotics were used in a small 0.7 ha shrimp farm near Pattukottai in Tamil Nadu State for one culture period for the management of pond environment and also the gut ecology of Penaeus monodon. The environmental parameters analysed were within the acceptable limits. It was evident from the results that the production was better in the experimental pond where the probiotics were used. The biological parameters such as the average body weight, FCR and total harvest achieved were better in the experimental pond than the control pond, all due to congenial environment, which obtained in the former mainly due to the use of probiotics.

Animals↗

Control of pathogenic Vibrio spp. by Bacillus subtilis BT23, a possible probiotic treatment for black tiger shrimp Penaeus monodon.

AIMS: The present study evaluated the in vitro and in vivo antagonistic effect of Bacillus against the pathogenic vibrios. METHODS AND RESULTS: Cell-free extracts of Bacillus subtilis BT23 showed greater inhibitory effects against the growth of Vibrio harveyi isolated by agar antagonism assay from Penaeus monodon with black gill disease. The probiotic effect of Bacillus was tested by exposing shrimp to B. subtilis BT23 at a density of 106-108 cfu ml-1 for 6 d before a challenge with V. harveyi at 103-104 cfu ml-1 for 1 h infection. The combined results of long- and short-term probiotic treatment of B. subtilis BT23 showed a 90% reduction in accumulated mortality. CONCLUSIONS: This study reports that pathogenic vibrios were controlled by Bacillus under in vitro and in vivo conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: Results indicated that probiotic treatment offers a promising alternative to the use of antibiotics in shrimp aquaculture.

Animals↗

Effect of probiotics on bacterial population and health status of shrimp in culture pond ecosystem.

The artificially manufactured probiotics having beneficial bacteria, Bacillus spp. was applied regularly in a modified extensive shrimp (Penaeus monodon) culture pond, located on the bank of Vellar estuary, Parangipettai. The populations of total heterotrophic bacteria (THB), beneficial bacteria (Bacillus spp.) and pathogenic bacteria (vibrios) were monitored in water and sediment of the pond. The results were compared with a control pond, situated in the same location having same water spread area, stocking density, species managed with same technologies and optimum environmental parameters in which no probiotic was applied. The populations of THB and Bacillus spp. in the experimental pond increased and the vibrios decreased after each application of probiotics. But the result of the control pond showed an increasing trend of the populations of THB, Bacillus spp. and vibrios towards days of culture. The control pond had lower levels of THB and Bacillus spp. and higher levels of vibrios than the probiotic applied (experimental) pond. Also the probiotics maintained optimum transparency and low organic load in the experimental pond as compared to control. In general, water and sediment had almost equal number of Bacillus spp. and vibrios, but sediment had higher THB load than water. The applications of probiotics lesser pathogenic vibrios and enhance beneficial bacilli in the culture leading to improved water quality, promoted growth and survival rates and increased the health status of the shrimp without stress and disease outbreaks. Thus the application of probiotics could lead to disease-free and profitable shrimp culture operations which will be helpful for shrimp farmers as most of them are now-a-days severely affected by microbial diseases.

Animals↗

Enhanced growth and resistance to Vibrio challenge in pond-reared black tiger shrimp Penaeus monodon fed a Bacillus probiotic.

The bacterial probiont Bacillus S11 (BS11) was used as a supplement in feed (PF) for black tiger shrimp Penaeus monodon in 2 earthen pond field-trials carried out for 100 d during 2 different seasons in Thailand. Growth and survival were compared with those of shrimp receiving an unsupplemented feed (UF). In the hot and cool seasons, respectively, shrimp fed PF grew significantly larger and had significantly higher survival than shrimp fed UF (p < 0.05). Projected yields on an annual basis (two 100 d crops) were 49% greater with PF-fed shrimp. In 8 d challenge tests using the luminescent bacterium Vibrio harveyi 1526, shrimp fed UF all died within 6 d while survival for shrimp fed PF was 5 and 9%.

Animal Feed↗

A new bacterial white spot syndrome (BWSS) in cultured tiger shrimp Penaeus monodon and its comparison with white spot syndrome (WSS) caused by virus.

This paper describes a new bacterial white spot syndrome (BWSS) in cultured tiger shrimp Penaeus monodon. The affected shrimp showed white spots similar to those caused by white spot syndrome virus (WSSV), but the shrimp remained active and grew normally without significant mortalities. The study revealed no evidence of WSSV infection using electron microscopy, histopathology and nested polymerase chain reaction. Electron microscopy indicated bacteria associated with white spot formation, and with degeneration and discoloration of the cuticle as a result of erosion of the epicuticle and underlying cuticular layers. Grossly the white spots in BWSS and WSS look similar but showed different profiles under wet mount microscopy. The bacterial white spots were lichen-like, having perforated centers unlike the melanized dots in WSSV-induced white spots. Bacteriological examination showed that the dominant isolate in the lesions was Bacillus subtilis. The occurrence of BWSS may be associated with the regular use of probiotics containing B. subtilis in shrimp ponds. The externally induced white spot lesions were localized at the integumental tissues, i.e., cuticle and epidermis, and connective tissues. Damage to the deeper tissues was limited. The BWS lesions are non-fatal in the absence of other complications and are usually shed through molting.

Animals↗

The functional property of Bacillus for shrimp feeds.

In shrimp cultures, feed represents the most expensive production cost. The quantity and quality of diets are primary factors influencing shrimp growth, nitrogen loading of the culture system and disease proliferation. For these reasons there is an interest in developing 'environmentally friendly' feeds for optimal growth. In this study, Bacillus strains were isolated from marine environments. The isolates were qualitatively assayed for proteases, carbohydrolases and lipases using selective media. The selected strains (9b, 31 and 33) were molecularly identified as Bacillus subtilis, B. megaterium and B. megaterium, respectively. Subsequently the strains were grown in an inexpensive culture medium (soybean mineral medium (MMS)). The information generated from the present investigation may contribute towards better feed formulations for shrimp at low cost, including bacterial strains as probiotics.

Animal Feed↗

Evaluation of Pseudomonas sp. PM 11 and Vibrio fluvialis PM 17 on immune indices of tiger shrimp, Penaeus monodon.

Occurrence of widespread epizootics among cultured stock of shrimp has put research programmes on preventive approaches such as application of probiotics on a high priority in aquaculture. In the present study two bacteria, Pseudomonas sp. PM 11 and Vibrio fluvialis PM 17 were selected as candidate probionts from a pool of bacteria isolated from gut of farm reared sub-adult shrimp and tested for their effect on the immunity indicators of tiger shrimp. Sub-adult shrimp, weighing 14 to 22 g were treated in separate experiments with Pseudomonas sp. PM 11 and V. fluvialis PM 17 at 10(3) bacterial cells ml(-1) in the experimental shrimp culture tanks. One set of experimental animals was treated every 3 days and another set of animals every 7 days with each of the candidate probionts. Estimation of immunological indicators such as haemocyte counts, phenol oxidase and antibacterial activity showed declining trends. The haemocyte counts dropped from 31 x 10(3) to 65 x 10(3) ml(-1) on the first day to 4-16 x 10(3) ml(-1) on the 45th day. Similarly, the phenol oxidase activity declined from 12-32 units on the first day to 11-14 units on 45th day of the experiment. Antibacterial activity of haemolymph reduced to 46-67 percent on the 45th day of the experiment. The results of the study suggest that, the criteria used for the selection of putative probiotic strains in the present study, such as predominant growth on primary isolation media, ability to produce extracellular enzymes and siderophores, did not bring about the desired effect in vivo and improve the immune system in shrimp. Hence, new protocols have to be evolved for selection of microbe(s) as putative probiotics and that, detailed understanding of proven probiotics, employed presently on empirical basis may provide a clue on the selection procedure.

Animals↗

The environmental impact of shrimp aquaculture: causes, effects, and mitigating alternatives.

Attracted by the demand for shrimp in the developed countries, shrimp aquaculture has expanded rapidly, mainly in the subtropical and tropical lowlands of America and Asia. This work provides a global review and viewpoint on the environmental impacts of shrimp aquaculture, considering the causes and effects of the siting and operation of shrimp ponds and abandonment of farm facilities. Additionally, mitigating alternatives are discussed. To date, approximately 1-1.5 million ha of coastal lowlands have been converted into shrimp ponds, comprising mainly salt flats, mangrove areas, marshes, and agricultural lands. The impact of shrimp farming of most concern is the destruction of mangroves and salt marshes for pond construction. Compatibility with other users, the presence of buffer zones, maintaining an acceptable balance between mangroves and shrimp pond area, improved pond design, reduction of water exchange, and an improved residence time of water, size and capacity to assimilate effluents of the water body, are examples of ways to mitigate the adverse effects. The use of mangroves and halophytes as biofilters of shrimp pond effluents offers an attractive tool for reducing the impact in those regions where mangrove wetlands and appropriate conditions for halophyte plantations exist. Healthy seed supply, good feed with the use of prophylactic agents (including probiotics), good water quality, and lower stocking densities are examples of actions suggested to control disease in shrimp farming. Finally, in the context of integrated management, research priorities are suggested.

Agrochemicals↗

[Effects of probiotics on Penaeus vannamei pond sediments].

This paper studied the effects of probiotics on the sediment of Penaeus vannamei pond during 117 days of culture period. The results showed that probiotics application significantly decreased the concentrations of total nitrogen, total phosphorous, and sulfide in sediment, but no significant difference was observed in total plate count (TPC) of microbes between treated and control ponds. The final average presumptive vibrio count (PVC) of treated pond sediment (3.65 x 10(3) cfu x g(-1)) was significantly lower than that of the control (1.16 x 10(5) cfu x g(-1)), while the average number of BS (Bacillus), AB (ammonifying bacteria), PSOB (presumptive sulphur oxidizing bacteria) and SRB (sulphur reducing bacteria) in treated pond sediment was higher than that of the control. These data showed that probiotics could decrease the nutrients (nitrogen, phosphate and sulfur) accumulation and improve the composition of bacterial populations in pond sediment, and thus, supply a good sediment environment for the healthily culture of the shrimp.

Animals↗

Genetic heterogeneity among Vibrio alginolyticus isolated from shrimp farms by PCR fingerprinting.

AIMS: To study the strain variability among Vibrio alginolyticus isolates from different sources by insertion sequence-targeted PCR fingerprinting and whole cell protein profile analysis. METHODS AND RESULTS: Eleven strains of V. alginolyticus were isolated from seven different sources including healthy, infected, farm-reared and wild shrimps. Following biochemical characterization, the isolates were analysed by PCR fingerprinting and whole cell protein analysis by SDS-PAGE. The strains were genetically different irrespective of the sources of isolation. CONCLUSIONS: Strain variation exists in V. alginolyticus isolates obtained even from the same source, and PCR fingerprinting is a simple and efficient method in identifying strain-specific variations among the different isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: Vibrio alginolyticus is implicated in severe vibriosis of marine aquaculture systems although many strains are avirulent and could be used as probiotic strains. As a wide variation exists among this species, differentiating the harmful and beneficial strains would help in finding ways of controlling the infections by eliminating harmful shrimp pathogenic vibrios.

Animals↗

From dysbiosis to resilience: Microbiome engineering for sustainable shrimp aquaculture.

The intensification of shrimp aquaculture has increased exposure to disease, environmental perturbations, and antimicrobial pressure, making microbial stability increasingly relevant to sustainable production. Microbiome stability-encompassing resistance to disturbance and resilience of functional recovery-provides an ecological framework for understanding how shrimp and culture-environment microbial communities respond to intensive farming. This review examines the transition from microbial homeostasis to dysbiosis and evaluates how microbiome engineering could redirect disrupted communities towards resilient states. Evidence is integrated across the intestine, hepatopancreas, rearing water, sediment and biofloc to assess how host genetics, ontogeny, diet, culture conditions, antibiotics and pollutants shape microbiome assembly and destabilization. Disease-associated changes in acute hepatopancreatic necrosis disease, white faeces syndrome, Enterocytozoon hepatopenaei infection, and white spot syndrome virus infection are critically evaluated, with explicit separation of associations, pathogen-induced dysbiosis, and community-level causality. Established and emerging interventions-including probiotics, prebiotics, synbiotics, functional diets, biofloc management, phages, postbiotics, microbiota transplantation and synthetic microbial communities-are assessed according to their capacity to modify microbial function, persistence and recovery rather than taxonomic change alone. We further examine how multi-omics, microbiome-informed breeding, and environmental monitoring could support biomarker development, predictive decision support and context-specific intervention. We argue that progress requires a shift from taxonomic description to function-guided engineering, from endpoint comparisons to direct measurement of resilience, and from laboratory efficacy to reproducible farm-scale validation. Overall, microbiome management may contribute to more disease-resilient and sustainable shrimp production, provided that its effectiveness can be validated under commercial farming conditions.

Dysbiosis↗

The choice of disease control strategies to secure international market access for aquaculture products.

Since production from capture fisheries cannot meet the demands of exports, aquaculture has subsequently played a major role in securing the raw materials for the world's food industries. Aquaculture has rapidly developed from extensive systems to semi-intensive, intensive and super-intensive systems. This has introduced the use of chemicals and drugs into the systems, which cause residual problems in the products. In the developed world, food safety has become a major issue of concern. The world market now demands healthy aquaculture products from farm to table. To achieve these requirements and to keep their markets, countries involved in aquaculture have implemented control measures such as farm licensing, code of conduct for sustainable aquaculture, hazard analysis and critical control point (HACCP) and good aquaculture practice. However, infectious diseases in aquaculture are of major concern to the industry and are typically controlled by eradication of the pathogen, treatment with antibiotic or chemotherapeutics, and/or by preventative measures such as the use of probiotics or vaccines. To limit the use of chemicals and antibiotics, good farm management is highly recommended. In terms of treatment, chemicals and antibiotics should be evaluated to establish recommended doses and withdrawal periods, otherwise alternative treatments should be developed. Environmentally-friendly probiotics have been introduced to aquaculture practice in the last decade to replace pathogenic bacteria with beneficial bacteria transient in the gut. Micro-organisms have also been prepared for the purpose of biocontrol and bioremediation. The application of probiotic, biocontrol and bioremediation seem promising; however considerable efforts of further research in terms of food and environmental safety are needed. Vaccination has proved highly effective in controlling diseases in the salmon industry mainly in Europe, America and Japan. In other Asian countries, this practice seem to be slower to develop due to differences in the aquaculture systems used and the economic value of the fish species farmed. Because shrimp farming is a large industry in this region, much effort has been put into developing a vaccine for viral diseases of shrimp. The efficacy of those vaccines remains inconclusive. Immunostimulants are another option developed for use in the shrimp industry in the region; however their efficacy also remains unclear.

Animals↗

Vibrios associated with Litopenaeus vannamei larvae, postlarvae, broodstock, and hatchery probionts.

Several bacteriological surveys were performed from 1994 to 1996 at different Litopenaeus vannamei hatcheries (in Ecuador) and shrimp farms (in Mexico). Samples were taken from routine productions of healthy and diseased L. vannamei larvae, postlarvae, and their culture environment and from healthy and diseased juveniles and broodstock. In Ecuador, the dominant bacterial flora associated with shrimp larvae showing symptoms of zoea 2 syndrome, mysis mold syndrome, and bolitas syndrome has been determined. Strains were characterized by Biolog metabolic fingerprinting and identified by comparison to a database of 850 Vibrio type and reference strains. A selection of strains was further genotypically fine typed by AFLP. Vibrio alginolyticus is predominantly present in all larval stages and is associated with healthy nauplius and zoea stages. AFLP genetic fingerprinting shows high genetic heterogeneity among V. alginolyticus strains, and the results suggest that putative probiotic and pathogenic strains each have specific genotypes. V. alginolyticus was found to be associated with larvae with the zoea 2 syndrome and the mysis mold syndrome, while different Vibrio species (V. alginolyticus and V. harveyi) are associated with the bolitas syndrome. V. harveyi is associated with diseased postlarvae, juveniles, and broodstock. The identities of the strains identified as V. harveyi by the Biolog system could not be unambiguously confirmed by AFLP genomic fingerprinting. Vibrio strain STD3-988 and one unidentified strain (STD3-959) are suspected pathogens of only juvenile and adult stages. V. parahaemolyticus, Photobacterium damselae, and V. mimicus are associated with juvenile and adult stages.

Animals↗

Rice-induced enterocolitis in an infant: TH1/TH2 cellular hypersensitivity and absent IgE reactivity.

BACKGROUND: Although food allergy is common in children, rice allergy is unusual in Western cultures. OBJECTIVE: To report a case of T-cell-mediated rice intolerance in an 11-month-old girl. METHODS: To evaluate the intolerance to rice in this patient, a graded rice food challenge was performed. To examine the immunologic reactivity to rice, in vitro lymphoproliferative responses and cytokine synthesis of rice-stimulated peripheral blood lymphocytes (PBLs) was performed. Subsequently, skin patch testing to rice and other foods was performed. RESULTS: Allergy skin prick test results were negative for rice and positive for egg, milk, and soy. Specific IgE antibodies to rice, egg, peanut, wheat, walnut, codfish, milk, soybean, corn, shrimp, scallops, and clams were undetectable. Results of a single-blind rice food challenge were positive, manifested by emesis that persisted for more than an hour and required intravenous hydration. In vitro lymphoproliferation by the patient's PBLs to rice stimulation was positive. In addition, cytokine synthesis of interferon-gamma, interleukin 10 (IL-10), tumor necrosis factor a, and IL-5 by the patient's rice-stimulated PBLs was elevated, indicating a TH1/TH2 cell response to rice. Endoscopy revealed normal esophageal, gastric, and duodenal mucosa; a biopsy specimen revealed mild esophagitis. Duodenal explant T cells were initially established by stimulation with rice and IL-2. After a 2-day rest, the lymphocytes were restimulated with rice for 7 days and revealed increased interferon-gamma and IL-5 synthesis. Twenty billion colony forming units of Lactobacillus GG were added to the patient's diet twice daily. After 6 weeks, rice rechallenge resulted in emesis within 1 hour. Results of patch testing were positive to rice, wheat, and barley but negative to soy, which the patient tolerated on food challenge. CONCLUSIONS: Although this patient did not demonstrate IgE antibody to rice, TH1/TH2 cell-mediated responses to rice were detected, and the patient experienced significant morbidity. Patch testing for gastrointestinal food allergies may be useful when the food specific IgE antibody is negative. Probiotic therapy in this patient did not ameliorate her sensitivity to rice, and food elimination remains the only reliable treatment for TH1/TH2-mediated food hypersensitivity.

Enterocolitis↗

Single-cell RNA sequencing provides further insights into the immunostimulatory action of freeze-dried Lactiplantibacillus plantarum on Penaeus vannamei shrimp.

Immunostimulation through dietary interventions opened new avenues in developing disease control and prevention tools for shrimp aquaculture. We have previously shown that feeding with freeze-dried Lactiplantibacillus plantarum (LAB) increased disease resistance of Penaeus vannamei against both Vibrio parahaemolyticus and white spot syndrome virus (WSSV) based on bulk RNA sequencing of shrimp gills. This tissue participates in ion transport and serves as a first line of defense against environmental stressors and pathogenic infections. However, characterization of their cell composition and functions remains limited. Here, we implemented a single-cell RNA sequencing approach to further gather insights into how feeding with freeze-dried LAB modulates host immunity which may not be evident with bulk RNA sequencing approach. A total of five clusters with unique transcriptional signatures were identified, corresponding to pillar cells, septal cells, and sessile hemocytes. Pseudo-bulk analyses at global- and cluster-levels showed differential expression of genes related to host immunity and metabolism. We further revealed how overall transcriptomic changes are not exclusively caused by gene expression changes but may also be driven by cell population dynamics. This study highlighted how single-cell RNA sequencing approach may shed light on the mechanisms of action of immunostimulants which may be masked in bulk transcriptome analyses.

Animals↗