Development of Antibiofilm Biosurfactants from Marine Bacteria Against Shrimp Vibrio pathogens
Keywords:
Biosurfactants, vibriosis, shrimp aquaculture, biofilm disruptionAbstract
Vibrio disease is being described as a major bacterial disease obviously known as penaeid bacterial septicaemia,
penaeid Vibriosis, luminescent Vibriosis or red leg diseases. Signs of Vibrio disease include lethargy, tissue and
appendage necrosis, slow growth, slow larval metamorphosis, body malformation, bioluminescence in shrimp
particularly produced in floc systems, muscle opacity, melanization, empty midgut and anorexia. In Asia, V.
alginolyticus and V. harveyi were considered as the most significant pathogens in the grow-out ponds of giant black
tiger shrimp Penaeus monodon. Survival and pathogenicity of Vibrio was associated with the biofilm formation and
quorum sensing. Therefore, disruption of biofilm formation and/or quorum sensing would be an effective management
strategy in aquatic systems instead of killing the pathogens which obviously leads to the development of resistant
strains. Biosurfactants are surface active smart biomolecules showed strong antibiofilm activity against Vibrio
pathogens. In this report, biofilm producing Vibrio pathogens include V. harveyi VB1, V. alginolyticus VB2, V.
vulnificus VB3, V. fischeri VB4, V. parahaemolyticus VB5 and Photobacterium damselae VB6 were isolated from
the moribund shrimp samples collected from farms located southeast coast of India. Based on their surface-active
properties, we hypothesized that biosurfactants could disrupt biofilms of Vibrio pathogens. To test the hypothesis, we
examined the effects of the lipopeptides extracted from marine bacteria MSI-A 07 and MSI-A 08, on the biofilmforming
capacity of biofilm infection causing pathogenic Vibrio spp. (V. harveyi VB1, V. alginolyticus VB2, V.
vulnificus VB3, V. fischeri VB4, V. parahaemolyticus VB5 and Photobacterium damselae VB6). The both lipopeptide
biosurfactants potentially disrupted biofilm formation under dynamic conditions. The biofilm disruption potential of
the lipopeptide biosurfactants was consistent against all shrimp pathogens. Based on this finding, biosurfactant
incorporated feed can be formulated to contain Vibrio outbreaks in shrimp aquaculture.
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References
Alvarez, J. D., Austin, B., Alvarez, A. M. and Reyes, H. (1998) ‘Vibrio harveyi: a pathogen of penaeid shrimps and
fish in Venezuela’, Journal of Fish Diseases. Wiley Online Library, 21(4), pp. 313–316.
Alcaráz, L. E., Satorres, S. E., Lucero, R. M. and Centorbi, O. N. (2003) ‘Species identification, slime production and
oxacillin susceptibility in coagulase-negative staphylococci isolated from nosocomial specimens’, Brazilian
Journal of Microbiology. SciELO Brasil, 34(1), pp. 45–51.
Banat, I. M. (1993) ‘The isolation of a thermophilic biosurfactant producing Bacillus sp’, Biotechnology letters.
Springer, 15(6), pp. 591–594.
Busscher, H. J. and Van der Mei, H. C. (1997) ‘Physico-chemical interactions in initial microbial adhesion and
relevance for biofilm formation’, Advances in dental research. SAGE Publications Sage CA: Los Angeles,
CA, 11(1), pp. 24–32.
Christensen, B. T. and Sørensen, L. H. (1985) ‘The distribution of native and labelled carbon between soil particle
size fractions isolated from long‐term incubation experiments’, European Journal of Soil Science. Wiley
Online Library, 36(2), pp. 219–229.
Costerton, J. W., Lewandowski, Z., Caldwell, D. E., Korber, D. R. and Lappin-Scott, H. M. (1995) ‘Microbial
biofilms’, Annual Reviews in Microbiology. Annual Reviews 4139 El Camino Way, PO Box 10139, Palo
Alto, CA 94303-0139, USA, 49(1), pp. 711–745.
Dalton, H. M., Poulsen, L. K., Halasz, P., Angles, M. L., Goodman, A. E. and Marshall, K. C. (1994) ‘Substratuminduced
morphological changes in a marine bacterium and their relevance to biofilm structure.’, Journal of
bacteriology. Am Soc Microbiol, 176(22), pp. 6900–6906.
Davey, M. E., Caiazza, N. C. and O’Toole, G. A. (2003) ‘Rhamnolipid surfactant production affects biofilm
architecture in Pseudomonas aeruginosa PAO1’, Journal of bacteriology. Am Soc Microbiol, 185(3), pp.
–1036.Dusane, D. H., Nancharaiah, Y. V., Zinjarde, S. S. and Venugopalan, V. P. (2010) ‘Rhamnolipid
mediated disruption of marine Bacillus pumilus biofilms’, Colloids and Surfaces B: Biointerfaces. Elsevier,
(1), pp. 242–248.
Davies, D. G., Parsek, M. R., Pearson, J. P., Iglewski, B. H., Costerton, J. W. t and Greenberg, E. P. (1998) ‘The
involvement of cell-to-cell signals in the development of a bacterial biofilm’, Science. American Association
for the Advancement of Science, 280(5361), pp. 295–298.
Donlan, R. M. and Costerton, J. W. (2002) ‘Biofilms: survival mechanisms of clinically relevant microorganisms’,
Clinical microbiology reviews. Am Soc Microbiol, 15(2), pp. 167–193.
Faruque, S. M., Biswas, K., Udden, S. M. N., Ahmad, Q. S., Sack, D. A., Nair, G. B. and Mekalanos, J. J. (2006)
‘Transmissibility of cholera: in vivo-formed biofilms and their relationship to infectivity and persistence in
the environment’, Proceedings of the National Academy of Sciences. National Acad Sciences, 103(16), pp.
–6355.
Gandhimathi, R., Kiran, G. S., Hema, T. A., Selvin, J., Raviji, T. R. and Shanmughapriya, S. (2009) ‘Production and
characterization of lipopeptide biosurfactant by a sponge-associated marine actinomycetes Nocardiopsis alba
MSA10’, Bioprocess and biosystems engineering. Springer, 32(6), pp. 825–835.
Hall-Stoodley, L., Costerton, J. W. and Stoodley, P. (2004) ‘Bacterial biofilms: from the natural environment to
infectious diseases’, Nature reviews. Microbiology. Nature Publishing Group, 2(2), p. 95.
Høiby, N., Johansen, H. K., Moser, C., Song, Z., Ciofu, O. and Kharazmi, A. (2001) ‘Pseudomonas aeruginosa and
the in vitroand in vivo biofilm mode of growth’, Microbes and Infection. Elsevier, 3(1), pp. 23–35.
Irie, Y., O’toole, G. A. and Yuk, M. H. (2005) ‘Pseudomonas aeruginosa rhamnolipids disperse Bordetella
bronchiseptica biofilms’, FEMS microbiology letters. Blackwell Publishing Ltd Oxford, UK, 250(2), pp.
–243.
Karanth, N. G. K., Deo, P. G. and Veenanadig, N. K. (1999) ‘Microbial production of biosurfactants and their
importance’, Current Science. JSTOR, pp. 116–126
Karunasagar, I., Pai, R., Malathi, G. R. and Karunasagar, I. (1994) ‘Mass mortality of Penaeus monodon larvae due
to antibiotic-resistant Vibrio harveyi infection’, Aquaculture. Elsevier, 128(3), pp. 203–209.
Karunasagar, I., Otta, S. K. and Karunasagar, I. (1996) ‘Biofilm formation by Vibrio harveyi on surfaces’,
Aquaculture. Elsevier, 140(3), pp. 241–245.
Karunasagar, I., Otta, S. K. and Karunasagar, I. (1997) ‘Histopathological and bacteriological study of white spot
syndrome of Penaeus monodon along the west coast of India’, Aquaculture. Elsevier, 153(1–2), pp. 9–13.
Kiran, G. S., Hema, T. A., Gandhimathi, R., Selvin, J., Manilal, A., Sujith, S. and Natarajaseenivasan, K. (2009)
‘Optimization and production of a biosurfactant from the sponge-associated marine fungus Aspergillus ustus
MSF3’, Coll Surf B: Biointerf, 73. doi: 10.1016/j.colsurfb.2009.05.025.
Kiran, G. S., Sabarathnam, B. and Selvin, J. (2010) ‘Biofilm disruption potential of a glycolipid biosurfactant from
marine Brevibacterium casei’, FEMS Immunology & Medical Microbiology. Blackwell Publishing Ltd
Oxford, UK, 59(3), pp. 432–438.
Lavilla-Pitogo, C. R. (1995) ‘Bacterial diseases of penaeid shrimps: an Asian view’, in Diseases in Asian Aquaculture
II: Proceedings of the Second Symposium on Diseases in Asian Aquaculture, 25-29 October 1993, Phuket,
Thailand. Fish Health Section, Asian Fisheries Society, pp. 107–121.
Lightner, D. V and Redman, R. M. (1994) ‘An epizootic of necrotizing hepatopancreatitis in cultured penaeid shrimp
(Crustacea: Decapoda) in northwestern Peru’, Aquaculture. Elsevier, 122(1), pp. 9–18.
Lightner, D. V (1996) ‘A handbook of shrimp pathology and diagnostic procedures for diseases of cultured penaeid
shrimp’. Baton Rouge, LA (USA) World Aquaculture Society.
Manilal, A., Sujith, S., Selvin, J., Kiran, G. S., Shakir, C. and Lipton, A. P. (2010) ‘Antimicrobial potential of marine
organisms collected from the southwest coast of India against multiresistant human and shrimp pathogens’,
Scientia marina, 74(2), pp. 287–296.
McLean, R. J. C., Pierson, L. S. and Fuqua, C. (2004) ‘A simple screening protocol for the identification of quorum
signal antagonists’, Journal of Microbiological Methods. Elsevier, 58(3), pp. 351–360.
Meylheuc, T., Van Oss, C. J. and Bellon‐Fontaine, M. (2001) ‘Adsorption of biosurfactant on solid surfaces and
consequences regarding the bioadhesion of Listeria monocytogenes LO28’, Journal of Applied
Microbiology. Wiley Online Library, 91(5), pp. 822–832.
Meylheuc, T., Methivier, C., Renault, M., Herry, J.-M., Pradier, C.-M. and Bellon-Fontaine, M. N. (2006) ‘Adsorption
on stainless steel surfaces of biosurfactants produced by gram-negative and gram-positive bacteria:
consequence on the bioadhesive behavior of Listeria monocytogenes’, Colloids and Surfaces B:
Biointerfaces. Elsevier, 52(2), pp. 128–137.
Mireles, J. R., Toguchi, A. and Harshey, R. M. (2001) ‘Salmonella enterica serovar Typhimurium swarming mutants
with altered biofilm-forming abilities: surfactin inhibits biofilm formation’, Journal of Bacteriology. Am Soc
Microbiol, 183(20), pp. 5848–5854.
Ophir, T. and Gutnick, D. L. (1994) ‘A role for exopolysaccharides in the protection of microorganisms from
desiccation’, Applied and Environmental Microbiology. Am Soc Microbiol, 60(2), pp. 740–745.
O’toole, G. A. and Kolter, R. (1998) ‘Flagellar and twitching motility are necessary for Pseudomonas aeruginosa
biofilm development’, Molecular microbiology. Wiley Online Library, 30(2), pp. 295–304.
O’Toole, G., Kaplan, H. B. and Kolter, R. (2000) ‘Biofilm formation as microbial development’, Annual Reviews in
Microbiology. Annual Reviews 4139 El Camino Way, PO Box 10139, Palo Alto, CA 94303-0139, USA,
(1), pp. 49–79
Parsek, M. R. and Fuqua, C. (2004) ‘Biofilms 2003: emerging themes and challenges in studies of surface-associated
microbial life’, Journal of bacteriology. Am Soc Microbiol, 186(14), pp. 4427–4440.
Quinn, G. A., Maloy, A. P., McClean, S., Carney, B. and Slater, J. W. (2012) ‘Lipopeptide biosurfactants from
Paenibacillus polymyxa inhibit single and mixed species biofilms’, Biofouling. Taylor & Francis, 28(10), pp.
–1166. doi: 10.1080/08927014.2012.738292.
Rodrigues, L., Banat, I. M., Teixeira, J. and Oliveira, R. (2006) ‘Biosurfactants: potential applications in medicine’,
Journal of Antimicrobial Chemotherapy. Oxford University Press, 57(4), pp. 609–618.
Ruangpan, L. and Kitao, T. (1991) ‘Vibrio bacteria isolated from black tiger shrimp, Penaeus monodon Fabricius’,
Journal of Fish diseases. Wiley Online Library, 14(3), pp. 383–388.
Selvin, J. and Lipton, A. P. (2003) ‘Vibrio alginolyticus associated with white spot disease of Penaeus monodon’,
Dis Aquat Org, 57. doi: 10.3354/dao057147.
Selvin, J., Ninawe, A. S., Seghal Kiran, G. and Lipton, A. P. (2010) ‘Sponge-microbial interactions: Ecological
implications and bioprospecting avenues’, Critical reviews in microbiology. Taylor & Francis, 36(1), pp. 82–
Singh, P. and Cameotra, S. S. (2004) ‘Potential applications of microbial surfactants in biomedical sciences’, TRENDS
in Biotechnology. Elsevier, 22(3), pp. 142–146.
Smith, P. T. (2000) ‘Diseases of the eye of farmed shrimp Penaeus monodon’, Diseases of aquatic organisms, 43(3),
pp. 159–173.
Snoussi, M., Ouchani, T. and Niazi, S. (2008) ‘Vulnerability assessment of the impact of sea-level rise and flooding
on the Moroccan coast: the case of the Mediterranean eastern zone’, Estuarine, Coastal and Shelf Science.
Elsevier, 77(2), pp. 206–213.
Stoodley, P., Sauer, K., Davies, D. G. and Costerton, J. W. (2002) ‘Biofilms as complex differentiated communities’,
Annual Reviews in Microbiology. Annual Reviews 4139 El Camino Way, PO Box 10139, Palo Alto, CA
-0139, USA, 56(1), pp. 187–209.
Sung, H.-H., Hsu, S.-F., Chen, C.-K., Ting, Y.-Y. and Chao, W.-L. (2001) ‘Relationships between disease outbreak
in cultured tiger shrimp (Penaeus monodon) and the composition of Vibrio communities in pond water and
shrimp hepatopancreas during cultivation’, Aquaculture. Elsevier, 192(2), pp. 101–110.
Velraeds, M. M., Van der Mei, H. C., Reid, G. and Busscher, H. J. (1996) ‘Inhibition of initial adhesion of
uropathogenic Enterococcus faecalis by biosurfactants from Lactobacillus isolates.’, Applied and
environmental microbiology. Am Soc Microbiol, 62(6), pp. 1958–1963.
Velraeds, M. M. C., van de Belt-Gritter, B., Busscher, H. J., Reid, G. and van der Mei, H. C. (2000) ‘Inhibition of
uropathogenic biofilm growth on silicone rubber in human urine by lactobacilli–a teleologic approach’,
World journal of urology. Springer, 18(6), pp. 422–426.
Watnick, P. I. and Kolter, R. (1999) ‘Steps in the development of a Vibrio cholerae El Tor biofilm’, Molecular
microbiology. Wiley Online Library, 34(3), pp. 586–595.
Watnick, P. I., Lauriano, C. M., Klose, K. E., Croal, L. and Kolter, R. (2001) ‘The absence of a flagellum leads to
altered colony morphology, biofilm development and virulence in Vibrio cholerae O139’, Molecular
microbiology. Wiley Online Library, 39(2), pp. 223–235.
Yildiz, F. H. and Schoolnik, G. K. (1999) ‘Vibrio cholerae O1 El Tor: identification of a gene cluster required for the
rugose colony type, exopolysaccharide production, chlorine resistance, and biofilm formation’, Proceedings
of the National Academy of Sciences. National Acad Sciences, 96(7), pp. 4028–4033.
Yildiz, F. H. and Visick, K. L. (2009) ‘Vibrio biofilms: so much the same yet so different’, Trends in microbiology.
Elsevier, 17(3), pp. 109–118.
You, J., Xue, X., Cao, L., Lu, X., Wang, J., Zhang, L. and Zhou, S. (2007) ‘Inhibition of Vibrio biofilm formation by
a marine actinomycete strain A66’, Applied microbiology and biotechnology. Springer, 76(5), pp. 1137–
Zhu, J. and Mekalanos, J. J. (2003) ‘Quorum sensing-dependent biofilms enhance colonization in Vibrio cholerae’,
Developmental cell. Elsevier, 5(4), pp. 647–656.
Zmantar, T., Chaieb, K., Makni, H., Miladi, H., Abdallah, F. Ben, Mahdouani, K. and Bakhrouf, A. (2008)
‘Detection by PCR of adhesins genes and slime production in clinical Staphylococcus aureus’, Journal of
basic microbiology. Wiley Online Library, 48(4), pp. 308–314.
Acknowledgements: Saba Rathnam thankful to MoES for fellowship (JRF). GSK and JS are thankful to
Ministry of Earth Sciences for the funding. This work is a part of MoES funded project.