Local aquatic microflora as a potential source of probionts in biofloc technology for whiteleg shrimp, Penaeus vannamei
DOI:
https://doi.org/10.22364/eeb.23.01Keywords:
C:N ratio, Pseudoalteromonas, Vibrio, water qualityAbstract
The Philippine shrimp aquaculture industry, a key supplier of Penaeus vannamei to domestic and international markets, faces significant disease challenges, particularly from pathogenic Vibrio species. Current disease management strategies often employ terrestrially-sourced probiotics, whose efficacy can be limited by environmental incompatibility with the aquatic host. This study aimed to address this limitation by isolating and characterizing putative probiotic microorganisms from the local aquatic environment of P. vannamei. Water samples, collected from a shrimp farm outlet pond in Negros Occidental, Philippines, were enriched with brown sugar to promote biofloc formation. Bacterial colonies were isolated on Nutrient Agar supplemented with 1% NaCl. In vitro antagonistic activity against the shrimp pathogen Vibrio harveyi was assessed via spot-on-lawn and cross-streak assays. The spot-on-lawn assay revealed a 45% inhibition rate against V. harveyi. Subsequent cross-streak assays confirmed inhibitory effects in five of nine isolates. These five isolates underwent morphological, biochemical, and molecular characterization using 16S rRNA gene sequencing. Three isolates were identified as putative Vibrio alginolyticus, while the remaining two closely matched Pseudoalteromonas species. Given the established use of non-pathogenic Vibrio and Pseudoalteromonas species as probiotics in aquaculture, these findings highlight the potential of local microflora as a source of probionts for biofloc-based shrimp culture. This approach may reduce reliance on external probiotic sources, contributing to enhanced industry sustainability.
References
Aguilera-Rivera D., Prieto-Davó A., Rodríguez-Fuentes G., Escalante-Herrera K.S., Gaxiola G. 2019. A vibriosis outbreak in the Pacific white shrimp, Litopenaeus vannamei reared in biofloc and clear seawater. J. Invertebr. Pathol. 167: 107246. DOI: https://doi.org/10.1016/j.jip.2019.107246
Amenyogbe E., Droepenu E.K., Ayisi C. L., Boamah G.A., Duker R.Q., Abarike E.D., Huang J. 2024. Impact of probiotics, prebiotics, and synbiotics on digestive enzymes, oxidative stress, and antioxidant defense in fish farming: current insights and future perspectives. Front. Mar. Sci. 11: 1368436. DOI: https://doi.org/10.3389/fmars.2024.1368436
Anee I.J., Alam S., Begum R.A., Shahjahan R.M., Khandaker A.M. 2021. The role of probiotics on animal health and nutrition. J. Basic Appl. Zool. 82: 52. DOI: https://doi.org/10.1186/s41936-021-00250-x
Austin B., Stuckey L.F., Robertson P.A.W., Effendi I., Griffith D.R.W. 1995. A probiotic strain of Vibrio alginolyticus effective in reducing diseases caused by Aeromonas salmonicida, Vibrio anguillarum and Vibrio ordalii. J. Fish Dis. 18: 93–96. DOI: https://doi.org/10.1111/j.1365-2761.1995.tb01271.x
Ben-David A., Davidson C.E. 2014. Estimation method for serial dilution experiments. J. Microbiol. Meth. 107: 214–221. DOI: https://doi.org/10.1016/j.mimet.2014.08.023
Caipang, C.M.A., Brinchmann M.F., Kiron V. 2010. Antagonistic activity of bacterial isolates from intestinal microbiota of Atlantic cod, Gadus morhua, and an investigation of their immunomodulatory capabilities. Aquac. Res. 41: 249–256. DOI: https://doi.org/10.1111/j.1365-2109.2009.02327.x
Caipang C.M., Trebol K.M.P., Fagutao F., Pakingking R.V.Jr., Deocampo J.E.Jr. 2022. Biofloc-based nursery production system: Heeding the call towards a sustainable shrimp culture industry in the Philippines. Int. J. Biosci. 20 : 250–259.
Caipang C.M.A., Trebol K.M.P., Suharman I., Pakingking R.V.Jr., Deocampo J.E.Jr. 2023. Isolation of potential probionts from brackishwater enriched with high levels of carbon source. J. Microbiol. Biotechnol. Food Sci. 13: e9819. DOI: https://doi.org/10.55251/jmbfs.9819
Chauhan A., Singh R. 2018. Probiotics in aquaculture: a promising emerging alternative approach. Symbiosis 77: 99–113. DOI: https://doi.org/10.1007/s13199-018-0580-1
Clapano M.B, Diuyan J.M.T., Rapiz F.G.B., Macusi E.D. 2022. Typology of smallholder and commercial shrimp (Penaeus vannamei) farms, including threats and challenges in Davao Region, Philippines. Sustainability 14: 5713. DOI: https://doi.org/10.3390/su14095713
de la Peña L.D., Lavilla-Pitogo C.R., Paner M.G. 2001. Luminescent Vibrios associated with mortality in pond-cultured shrimp Penaeus monodon in the Philippines: species composition. Fish Pathol. 36: 133–138. DOI: https://doi.org/10.3147/jsfp.36.133
De Schryver P., Defoirdt T., Sorgeloos, P. 2014. Early mortality syndrome outbreaks: a microbial management issue in shrimp farming? PLoS Pathogens 10: e1003919. DOI: https://doi.org/10.1371/journal.ppat.1003919
El‐Sayed A. M. 2020. Use of biofloc technology in shrimp aquaculture: a comprehensive review, with emphasis on the last decade. Rev. Aquacult. 13: 676–705. DOI: https://doi.org/10.1111/raq.12494
Eze O.C., Berebon D.P., Emencheta S.C., Evurani S.A., Okorie C.N., Balcão V.M., Maria M. 2023. Therapeutic potential of marine probiotics: a survey on the anticancer and antibacterial effects of Pseudoalteromonas spp. Pharmaceuticals 16: 1091. DOI: https://doi.org/10.3390/ph16081091
García-Márquez J., Vizcaíno A.J., Barany A., Galafat A., Acién G., Figueroa F.L., Alarcón F.J., Mancera J.M., Martos-Sitcha J.A., Arijo S., Abdala-Diaz R.T. 2023. Evaluation of the combined administration of Chlorella fusca and Vibrio proteolyticus in diets for Chelon labrosus: effects on growth, metabolism, and digestive functionality. Animals 13: 589–589. DOI: https://doi.org/10.3390/ani13040589
Gomez-Gil B., Roque A., Velasco-Blanco G. 2002. Culture of Vibrio alginolyticus C7b, a potential probiotic bacterium, with the microalga Chaetoceros muelleri. Aquaculture 211: 43–48. DOI: https://doi.org/10.1016/S0044-8486(02)00004-2
Gustilatov M., Widanarni W., Ekasari J., Julyantoro P.G.S., Waturangi D.E. 2024. Biofloc system supplemented by Pseudoalteromonas piscicida 1Ub protects the Pacific white shrimp Penaeus vannamei from Vibrio parahaemolyticus infection. Aquacult. Fisher. 9: 967–974. DOI: https://doi.org/10.1016/j.aaf.2023.05.003
Hai N.V. 2015. The use of probiotics in aquaculture. J. Appl. Microbiol. 119: 917–935. DOI: https://doi.org/10.1111/jam.12886
Holt G.J., Krieg N.R., Sneath P.H.A., Staley J.T., Williams S.T. 2000. Bergey’s Manual of Determinative Bacteriology. 9th Ed. Lippincott Williams and Wilkins, Philadelphia, USA.
Hoseinifar S.H., Sun Y.-Z., Wang A., Zhou Z. 2018. Probiotics as means of diseases control in aquaculture, a review of current knowledge and future perspectives. Front. Microbiol. 9: 2429.. DOI: https://doi.org/10.3389/fmicb.2018.02429
Huang Y., Li M., Yu Z., Qian P. 2011. Correlation between pigmentation and larval settlement deterrence by Pseudoalteromonas sp. sf57. Biofouling 27: 287–293. DOI: https://doi.org/10.1080/08927014.2011.562978
Jamal M.T., Abdulrahman I.A., Al Harbi M., Chithambaran S. 2019. Probiotics as alternative control measures in shrimp aquaculture: a review. J. Appl. Biol. Biotechnol. 7: 69–77. DOI: https://doi.org/10.7324/JABB.2019.70313
Karunasagar I. 2018. Ecology, virulence factors and global spread of Vibrio parahaemolyticus. Asian Fisher. Sci. 31S: 29–58. DOI: https://doi.org/10.33997/j.afs.2018.31.S1.002
Kobayashi K., Ikemoto Y. 2019. Biofilm-associated toxin and extracellular protease cooperatively suppress competitors in Bacillus subtilis biofilms. PLOS Genetics 15: e1008232. DOI: https://doi.org/10.1371/journal.pgen.1008232
Kopermsub P., Yunchalard S. 2010. Identification of lactic acid bacteria associated with the production of plaa-som, a traditional fermented fish product of Thailand. Int. J. Food Microbiol. 138: 200–204. DOI: https://doi.org/10.1016/j.ijfoodmicro.2010.01.024
Krummenauer D., Peixoto, S. Cavalli, R. O., Poersch L. H., Wasielesky W. 2011. Superintensive culture of white shrimp, Litopenaeus vannamei, in a biofloc technology system in Southern Brazil at different stocking densities. J. World Aquacult. Soc. 42: 726–733. DOI: https://doi.org/10.1111/j.1749-7345.2011.00507.x
Lazado C. C., Caipang C. M. A. 2014. Atlantic cod in the dynamic probiotics research in aquaculture. Aquaculture 424-425: 53–62. DOI: https://doi.org/10.1016/j.aquaculture.2013.12.040
Lazado C.C., Caipang C.M.A., Brinchmann M.F., Kiron V. 2011. In vitro adherence of two candidate probiotics from Atlantic cod and their interference with the adhesion of two pathogenic bacteria. Vet. Microbiol. 148: 252–259. DOI: https://doi.org/10.1016/j.vetmic.2010.08.024
Mardani T., Khiabani M.S., Mokarram R.R., Hamishehkar H. 2018. Immobilization of α-amylase on chitosan-montmorillonite nanocomposite beads. Int. J. Biol. Macromolec. 120: 354–360. DOI: https://doi.org/10.1016/j.ijbiomac.2018.08.065
Neves H.I., Machado G.T., Ramos T.C.S., Yang H.M., Yagil E., Spira B. 2020. Competition for nutritional resources masks the true frequency of bacterial mutants. BMC Biology 18: 194. DOI: https://doi.org/10.1186/s12915-020-00913-1
Ninawe A.S., Selvin J. 2009. Probiotics in shrimp aquaculture: avenues and challenges. Crit. Rev. Microbiol. 35: 43–66. DOI: https://doi.org/10.1080/10408410802667202
Nguyen T.D.T., Kang J.H., Lee M.S. 2007. Characterization of Lactobacillus plantarum PH04, a potential probiotic bacterium with cholesterol-lowering effects. Int. J. Food Microbiol. 113: 358–361. DOI: https://doi.org/10.1016/j.ijfoodmicro.2006.08.015
Rosario W.R., Lopez N.A. 2005. Status of P. vannamei aquaculture in the Philippines. In: Aquaculture of P. vannamei and Other Exotic Shrimps. Annex: Philippines. SEAFDEC Aqaculture Department Institutional Repository, pp. 62–68.
Sharma S., Shah E., Davla D., Dixit G., Patel A., Kumar A.K. 2019. Effect of microalga-based diet on oxidative stress enzymes of African catfish, Clarias gariepinus. Int. Aquat. Res. 11: 377–387. DOI: https://doi.org/10.1007/s40071-019-00245-z
Shinn A.P., Pratoomyot J., Griffiths D., Trong T.Q., Vu N.T., Jiravanichpaisal P., Briggs M. 2018. Asian shrimp production and the economic costs of disease. Asian Fisher. Sci. 31S: 29–58. DOI: https://doi.org/10.33997/j.afs.2018.31.S1.003
Siladan M., Failaman A., Ferriols V., Traifalgar R. 2013. Isolation and screening of gut microflora from Siganus guttatus (Bloch, 1787) for potential antivibrio agents. Eur. J. Exp. Biol. 3: 236–240.
Sohel A.M., Shahjahan M., Hossain M.K., Sumi K.R., Hossain M.S., Kari Z.A., Tahiluddin A., Téllez-Isaías G. 2023. Effects of multispecies probiotics on growth, hematology, and gut health of stinging catfish (Heteropneustes fossilis) in biofloc system. Water 15: 2519–2519. DOI: https://doi.org/10.3390/w15142519
Soltani M., Ghosh K., Hoseinifar S.H., Kumar V., Lymbery A.J., Roy S., Ringø E. 2019. Genus Bacillus, promising probiotics in aquaculture: aquatic animal origin, bio-active components, bioremediation and efficacy in fish and shellfish. Rev. Fisher. Sci. Aquacult. 27: 331–379. DOI: https://doi.org/10.1080/23308249.2019.1597010
Stubbendieck R.M., May D.S., Chevrette M.G., Temkin M.I., Wendt-Pienkowski E., Cagnazzo J., Carlson C.M., Gern J.E., Currie C.R. 2019. Competition among nasal bacteria suggests a role for siderophore-mediated interactions in shaping the human nasal microbiota. Appl. Environ. Microbiol. 85: e02406-18. DOI: https://doi.org/10.1128/AEM.02406-18
Sumon M.S., Ahmmed F., Khushi S.S., Ahmmed M.K., Rouf M.A., Chisty M.A.H., Sarower M.G. 2018. Growth performance, digestive enzyme activity and immune response of Macrobrachium rosenbergii fed with probiotic Clostridium butyricum incorporated diets. J. King Saud Univ. Sci. 30: 21–28. DOI: https://doi.org/10.1016/j.jksus.2016.11.003
Thompson J., Gregory S., Plummer S., Shields R.J., Rowley A.F. 2010. An in vitro and in vivo assessment of the potential of Vibrio spp. as probiotics for the Pacific White shrimp, Litopenaeus vannamei. J. Appl. Microbiol. 109: 1177–1187. DOI: https://doi.org/10.1111/j.1365-2672.2010.04743.x
Usakova N.A., Nekrasov R.V., Pravdin I.V., Sverchkova N.V., Kolomiyets E.I., Pavlov D.S. 2015. Mechanisms of the effects of probiotics on symbiotic digestion. Biol. Bull. 42: 394–400. DOI: https://doi.org/10.1134/S1062359015050131
Van Spaendonk H., Ceuleers H., Witters L., Patteet E., Joossens J., Augustyns K., Lambeir A. -M., De Meester I., De Man J.G., De Winter B.Y. 2017. Regulation of intestinal permeability: The role of proteases. World J. Gastroenterol. 23: 2106–2123. DOI: https://doi.org/10.3748/wjg.v23.i12.2106
Vergel J.C.V., Cabawatan L.D.P., Madrona V.A.C., Rosario A.F.T., Ana J.B.M.S., Tare M.V.R., Maningas M.B.B. 2019. Detection of Taura Syndrome Virus (TSV) in Litopenaeus vannamei in the Philippines. Philippine J. Fisher. 26: 8–14. DOI: https://doi.org/10.31398/tpjf/25.2.2018-0003
Vergel J. C. V. 2017. Current trends in the Philippines’ shrimp aquaculture industry: a booming blue economy in the Pacific. Oceanogr. Fisher. Open Access J. 5: 555668. DOI: https://doi.org/10.19080/OFOAJ.2017.05.555668
Verschuere L., Rombaut G., Huys G., Dhont J., Sorgeloos P., Verstraete W. 1999. Microbial control of the culture of Artemia juveniles through preemptive colonization by selected bacterial strains. Appl. Environ. Microbiol. 65: 2527–2533. DOI: https://doi.org/10.1128/AEM.65.6.2527-2533.1999
Wang H., Sun B., Xie G., Wan X., Huang J., Song X. 2021. Spotlight on a novel bactericidal mechanism and a novel SXT/R391-like integrative and conjugative element, carrying multiple antibiotic resistance genes, in Pseudoalteromonas flavipulchra strain CDM8. Microbiol. Res. 242: 126598–126598. DOI: https://doi.org/10.1016/j.micres.2020.126598
Wang H., Wang C., Tang Y., Sun B., Huang J., Song X. 2018. Pseudoalteromonas probiotics as potential biocontrol agents improve the survival of Penaeus vannamei challenged with acute hepatopancreatic necrosis disease (AHPND)-causing Vibrio parahaemolyticus. Aquaculture 494: 30–36. DOI: https://doi.org/10.1016/j.aquaculture.2018.05.020
Zorriehzahra M. J., Banaederakhshan, R. 2015. Early mortality syndrome (EMS) as new emerging threat in shrimp industry. Adv. Anim. Vet. Sci. 3: 64–72. DOI: https://doi.org/10.14737/journal.aavs/2015/3.2s.64.72
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