Screening of marine Streptomyces spp. for potential use as probiotics in aquaculture Surajit Das ⁎, Louise R. Ward, Chris Burke National Centre for Marine Conservation and Resource Sustainability, Australian Maritime College, University of Tasmania, Locked Bag 1370, Launceston, Tasmania 7250, Australia abstract article info Article history: Received 7 January 2010 Received in revised form 30 March 2010 Accepted 1 April 2010 Keywords: Artemia Penaeus monodon Probiotics Streptomyces Vibrio Survival Protection Marine Streptomyces strains (CLS-28, CLS-39 and CLS-45) were used to colonise Artemia nauplii (Instar I) and 15 d old adult Artemia prior to challenge with Vibrio harveyi and V. proteolyticus. The LC 50 of V. harveyi and V. proteolyticus was found to be ∼ 10 6 CFU ml -1 . V. proteolyticus was more pathogenic than V. harveyi at 10 6 CFU ml -1 . A significant reduction in mortality (P b 0.001) was found by addition of 1% wet cell mass of Streptomyces strains in nauplii and adult Artemia against both the pathogens. The best protective responses were shown by CLS-39 in both nauplii and adults against V. harveyi and by CLS-39 in nauplii and CLS-28 in adults against V. proteolyticus. Shrimp feeds were supplemented with Streptomyces cell mass at 1% dosage and fed to black tiger shrimp Penaeus monodon postlarvae for 15 d in three treatments with two treatments of commercial probiotic (T1: feed + CLS-28; T2: feed + CLS-39; T3: feed + CLS-45; T4: feed + Sanolife® commercial probiotic and T5: Sanolife® commercial probiotic in water). During this time, ammonia was in the range of 1 to 2 ppm in all the treatments with significant differences between treatments (P b 0.05). Significant differences (P b 0.05) were also found in survival, total length and wet weight of the shrimp postlarvae during the 15 d trial. T5 showed the best gains in terms of length and weight followed by T1, T2, T3 and T4. Streptomyces treatments T1, T2 and T3 showed better survival and higher length and weight than the control and T4. Total heterotrophic bacteria and Vibrio counts were in the range of 10 8 and 10 6 CFU ml -1 respectively in all the treatments. The Vibrio population differed significantly in the treatments (P b 0.05) and the total bacterial counts showed no significant differences in the treatments (P N 0.05). After challenge with V. harveyi at 10 7 CFU ml -1 , highest survival was found in T1 and T5. Among the Streptomyces treatments, T1 showed significantly higher survival compared to the control, followed by T2 and T3. Thus Streptomyces strains show promise as probiotic agents in mariculture. © 2010 Elsevier B.V. All rights reserved. 1. Introduction Probiotics which compete with bacterial pathogens for nutrients and/ or inhibit the growth of pathogens can be a valid alternative to the prophylactic application of antibiotics and biocides. Fuller (1989) defined a probiotic as ‘a live microbial feed supplement which beneficially affects the host animal by improving its intestinal microbial balance’ . A modified and more appropriate definition was proposed by Verschuere et al. (2000a) —‘a live microbial adjunct which has a beneficial effect on the host by modifying the host-associated or ambient microbial community, by ensuring improved use of the feed or enhancing its nutritional value, by enhancing the host response towards disease, or by improving the quality of its ambient environment’ . Actinobacteria is a class with five subclasses that was proposed by Stackebrandt et al. (1997) to group the highly diverse so called actinomycetes based on chemical composition, DNA–DNA reassocia- tion and 16S rRNA gene sequence similarities. Members of the Actinobacteria are prolific sources of secondary metabolites and the vast majority of these compounds are derived from the single genus Streptomyces. Streptomyces is a Gram-positive aerobic genus in the order Actinomycetales, suborder Streptomycineae and family Strepto- mycetaceae (Stackebrandt et al., 1997) and has a DNA G + C content of 69–78 mol%. Marine-derived Streptomyces have been studied for isolation of several novel secondary metabolites (Fenical and Jensen, 2006; Das et al., 2006a). However, to date there have only been a few studies that have considered Actinobacteria for their application as probiotics in aquaculture. We have reported the prospects of using marine Actinobacteria as probiotics in aquaculture (Das et al., 2008a) and began screening marine Actinobacteria for use as new biocontrol agents for aquatic animals. We report here the effect of three marine Streptomyces strains on Artemia and Penaeus monodon. Artemia has long been considered as a model/ test organism to study the mode of action of probiotic bacteria due to its adaptability to wide ranges of salinity and temperature, short life cycle, high adaptability to adverse environmental conditions, high fecundity, parthenogenetic and sexual reproduction strategy (with nauplii or cysts production), small body size, and adaptability to varied nutrient resources (Nunes et al., 2006). There have been several experiments carried out on Artemia in the search for new biocontrol agents for aquaculture Aquaculture 305 (2010) 32–41 ⁎ Corresponding author. Present address: Department of Life Science, National Institute of Technology, Rourkela-769 008, Orissa, India. Tel.: +91 661 2462684; fax: +91 661 2462022. E-mail addresses: surajit@myself.com, surajit@nitrkl.ac.in (S. Das). 0044-8486/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.aquaculture.2010.04.001 Contents lists available at ScienceDirect Aquaculture journal homepage: www.elsevier.com/locate/aqua-online