Functional assessment of narAa and narAb genes encoding naphthalene dioxygenase in Micromonospora sp. strain IBH using gene-targeted polymerase chain reaction and assessment of dioxygenase activity

Abstract

Actinobacteria, have attracted attention in the field of bioremediation due to their exceptional metabolic versatility and ability to degrade a wide array of complex organic compounds. However, the genus Micromonospora, which is a Gram-positive, filamentous bacterium with highly branched mycelium, has scarcely been attributed to these properties; even though the species of Micromonospora are widely ubiquitous across the assorted environments, such as the soil, marine and mangrove sediment, epiphytic habitat, and hydrocarbon-contaminated sites. This prompted this study to investigate dioxygenase activity and probe for the nar-type naphthalene dioxygenase gene in Micromonospora sp. strain IBH of Irish urban soil origin. Before the dioxygenase assay, lyophilized cells of Micromonospora sp. strain IBH (Accession: PZ027962.1) were recovered, purified, and confirmed via Gram staining and microscopy. The dioxegenase gene activity was determined via the preliminary and confirmatory dioxygensae activity assay using a modified method involving the conversion of indole substrate to indoxyl then to indigo by the isolates. The gene-targeted polymerase chain reaction (PCR) was carried out for the amplification of the 205 bp fragment of narAa and narAb naphthalene dioxygenase (NDO) gene using primers specific for Rhodococcus sp. strain NCIMB 12038, known to harbor the nar-type NDO. Findings from this study revealed the Micromonospora sp. strain IBH exhibited positive dioxygenase gene activity, implying it portray genetic potential for naphthalene degradation. However, PCR probing of the narAa and narAb fragments of NDO produced no specific 205-bp band targeted, suggesting the isolate does not harbor this particular nar-type NDO. The exhibition of the phenotypic activity of naphthalene dioxygenase by the Micromonospora sp. strain IBH is a high prospect for naphthalene metabolism, degradation and bioremediation application. For future research, other variant NDO genes, such as the Psudomonas nah-type NDO genes should be probed to corroborate its gene competence and strain optimized for bioremediation application.