Potential antimicrobial activity of marine molluscs from tuticorin, southeast coast of India against 40 biofilm bacteria

Journal of Shellfisheries Research, Jan, 2005 by M. Santhana Ramasamy, A. Murugan

ABSTRACT Methanol: water (1:1), methanol: dichloromethane (1:1) and acetone extracts of molluscs comprising 77 whole body, four inks, four opercula, 10 egg masses, and 10 digestive glands were screened for antimicrobial activity on marine biofilm bacteria. The methanol: water (1:1) whole body extracts of Nerita albicilla and Nerita oryzarum showed broad spectral inhibitory activity against 93% and 95% of the 40 biofilm bacteria. The egg masses from 10 gastropods showed activity against more than a quarter of the biofilm bacteria. The methanolic extract of Chicoreus virgineus, Chicoreus ramosus egg masses, and acetone extract of the egg mass of Rapana rapiformis showed broad-spectrum antibacterial activity against all the 40 biofilm bacterial strains. The activity in gastropod egg masses was localized to their internal matrix. Overall screening showed activity in 38.1% of the methanolic extracts followed by 13.3% of methanol: water, 12.4% of methanol: dichloromethane, and 3.8% of acetone extracts. Gastropods showed good activity when compared with bivalves and cephalopods.

KEY WORDS: antifouling, bacteria, biofilm, egg mass, mollusc

INTRODUCTION

The efforts to control marine biofouling are ongoing since humans started venturing into the sea. Among the antifoulants developed to date, organotin compounds are considered as the most effective and have been widely used since the 1970s. It is well established that TBT compounds cause imposex universally (Barroso et al. 2002) and has been documented in over 118 species in 63 genera (Bettin et al. 1996). Organotin pollution has also raised concerns over the accumulation of organotin compounds in the food chain and the associated risks related to the presence of contaminants (Belfroid et al. 2000). International Maritime Organization (IMO) has proposed a complete phasing out of TBT use by the year 2008. The alternative tin free coatings are not only costly, but also their metal constituents may pose environmental problems (Hellio et al. 2004). So, a great deal of research has been focused on finding an alternate nontoxic and eco-friendly antifoulant (De Nys & Steinburg 2002, Fusetani 2004). In this context, marine organisms, especially the sedentary forms have interested researchers very much because they exhibit characteristic chemical defense against the epizooic organisms. More than 100 species of marine organisms have been shown to exhibit antimicrobial activity, as well as the ability to prevent settlement of fouling organisms (Clare 1996, Koh 1997).

Most currently available antifouling compounds are active against barnacles and algae and not against microbes like biofilm bacteria, which cause initial fouling of a submerged surface (Tang & Cooney 1998). Bacteria are the first organisms to foul submerged surfaces in the marine environment. Their subsequent multiplication and production of exopolymers leads to the formation of bacterial biofilms, which attract a large number of secondary fouling organisms (Steinberg et al. 2002). The interaction between bacteria and macrofoulers are dynamic and varied. The resultant biofouling community leads to economic and ecologic implications on both human-made structures and living surfaces.

However, bacteria do not colonize all submerged surfaces in a uniform manner and are influenced by many physical and chemical factors (Fletcher & Marshall 1982). The colonization process of the bacteria in the living marine surfaces may also be influenced by the secondary metabolites produced by the epibiont host. There exists an inverse correlation between macrofouling and antimicrobial activity of the epibiont host (Koh 1997, Santhana Ramasamy & Murugan 2003). This has been proposed as evidence that secondary metabolites are involved in the control of epibiosis (Slattery et al. 1995, Newbold et al. 1999). So, the natural mechanism of upsetting the development of the microfouling bacterial community by nontoxic marine natural compounds produced by the foul free host organisms may be taken as the most desirable way for breaking the fouling chain to reduce macrofouling settlement.

The cnidarians, sponges, ascidians, bryozoans, marine plant, and the like have received much attention and widely studied for antifouling activity (Clare 1996, Fusetani 2004). The study on the bioactivity of marine mollusc metabolites is scanty except for the nudibranchs. Rinehart et al. (1981) opined that marine molluscs might provide potential for isolating compounds with specific activity against certain microorganisms or cell types. Molluscan forms do exhibit adaptations to avoid fouling of their surface. The rare incidence of fouling on the Helgoland snails of Littorina littorea is due to the density dependent mutual grazing (Wahl et al. 1998). The periostracum of the mussel Mytilus edulis has been shown to possess some repellents to avoid fouling of their surface (Wahl et al. 1998).

The gelatinous egg masses of marine molluscs also have no physical protection from predation. So, many marine molluscs might have evolved mechanisms to avoid predation and surface fouling on their egg masses. Lord (1986) has demonstrated that the content of the egg capsules in Nucella lapillus was free from bacterial contamination and the capsule wall was impermeable to bacteria. Benkendorff et al. (2001) indicated that antibacterial compounds are common in both tough egg capsules and gelatinous egg masses, suggesting that chemical defense is the primary mechanism used to maintain an axenic environment for embryonic development. The egg masses of some marine molluscs have been the source of several compounds with antibacterial activity (Kamiya et al. 1984, Matsunaga et al. 1986, Yamazaki 1993, Benkendorff et al. 2000a, Pawlik et al. 1988). But, study on the antimicrobial properties of the marine molluscs against marine bacteria is scanty except a few works (Kamiya et al. 1984, Benkendorff et al. 2000b, Mitta et al. 2000, Benkendorff et al. 2001) and no previous studies have tested for activity against biofilm bacteria. So, in this study antimicrobial activity of a broad range of marine molluscs was evaluated against marine biofilm bacteria.