Phylogenetic diversity and activity screening of cultivable Actinobacteria isolated from marine sponges and associated environments from the western coast of India

Abstract

The phylogenetic diversity of cultivable actinobacteria isolated from sponges (Haliclona spp.) and associated intertidal zone environments along the northern parts of the western coast of India were studied using 16S rRNA gene sequences. A subset of randomly selected actinobacterial cultures were screened for three activities, namely predatory behaviour, antibacterial activity and enzyme inhibition. We recovered 237 isolates from the phylum Actinobacteria belonging to 19 families and 28 genera, which could be attributed to 95 putative species using maximum-likelihood partition and 100 putative species using Bayesian partition in Poisson tree processes. Although the trends in the discovery of actinobacterial genera isolated from sponges were consistent with previous studies from different study areas, we provide the first report of nine actinobacterial species from sponges. We observed widespread non-obligate epibiotic predatory behaviour in eight actinobacterial genera and we provide the first report of predatory activity in Brevibacterium , Glutamicibacter , Micromonospora , Nocardiopsis , Rhodococcus and Rothia . Sponge-associated actinobacteria showed significantly more predatory behaviour than environmental isolates. While antibacterial activity by actinobacterial isolates mainly affected Gram-positive target bacteria with little or no effect on Gram-negative bacteria, predation targeted both Gram-positive and Gram-negative prey with equal propensity. Actinobacterial isolates from both sponges and associated environments produced inhibitors of serine proteases and angiotensin-converting enzyme. Predatory behaviour was strongly associated with inhibition of trypsin and chymotrypsin. Our study suggests that the sponges and associated environments of the western coast of India are rich in actinobacterial diversity, with widespread predatory activity, antibacterial activity and production of enzyme inhibitors. Understanding the diversity and associations among various actinobacterial activities - with each other and the source of isolation - can provide new insights into marine microbial ecology and provide opportunities to isolate novel therapeutic agents.

Keywords: antibiotic production; bacterial predation; enzyme inhibition; molecular phylogeny; secondary metabolites.

Fig. 1.

Maximum-likelihood phylogenetic tree of actinobacterial isolates based on the TIM3+F+I+G4 nucleotide substitution model (lnL of consensus tree: −18 684.58). Firmicutes belonging to genus Bacillus were used as outgroups.

Fig. 2.

Association between source of actinobacterial isolation and predatory behaviour. There was a significant association between the source (sponge or associated environment) of actinobacterial isolation and predation (χ²=5.265, P=0.0218).

Fig. 3.

Venn diagram combination of enzyme inhibitors produced by actinobacterial isolates. Venn diagram is not to scale.

Fig. 4.

Venn diagram of predation, antibiotic production and enzyme inhibition by actinobacterial isolates. Venn diagram is not to scale.

Fig. 5.

Association between enzyme inhibition and predation in actinobacterial isolates. Predation was significantly associated with (a) inhibition of any one of the four enzymes tested (χ²=26.172, P<0.0001), (b) inhibition of trypsin (χ²=23.165, P<0.0001) and (c) inhibition of chymotrypsin (χ²=42.604, P<0.0001).