Multiple Antibacterial Activities of Proteinaceous Compounds in Crude Extract from the Eastern Subterranean Termite, Reticulitermes flavipes Kollar (Blattodea: Isoptera: Rhinotermitidae)

Aims: To assess the presence of antibacterial activities in Reticulitermes flavipes against a common Gram-positive soil bacterium Bacillus subtilis; to determine the nature of antibacterial compounds of crude extracts; and to analyze the size profile of active compounds.

Place and Duration of Study: Department of Entomology and Plant Pathology and Department of Biological Sciences in Auburn University between August 2012 and July 2013.

Methodology: The cell free crude extracts were obtained from R. flavipes workers. Five size-fractionated solutions (≥100, 30-100, 10-30, 3-10, and ≤3 kDa) were achieved by sequentially size fractionating the crude extract with MicrosepTM Advance Centrifugal Devices. Heat-stable fraction was acquired by subjecting the crude extract to heat-treatment to denature proteins. Activities of the crude extract, heat-treated extract, and size-fractionated extracts against B. subtilis were determined using the inhibition zone assay. Anti-bacterial activities were determined by the measurements of the diameters (mm) of growth inhibition zones. The significances among the seven treatments, in comparison to Ampicillin (positive control) and Tris-NaCl buffer (negative control), were determined using repeated measures ANOVA.

Results: The activity against B. subtilis was evidenced in all but the heat-treated solutions, indicating the presence of antibacterial activities, the existence of multiple active compounds in the crude extracts, and the protein nature of the active compounds. The active compounds, with the molecular sizes ranging from <3 to >100 kDa, demonstrated different levels of antibacterial activity (P = 0.017). The greatest activity was observed in the fraction of 3-10 kDa and Ampicillin, followed by the fractions of ≤3 kDa and ≥100 kDa, and the lowest in the fraction of 10-30 kDa.

Conclusion: Crude extracts from R. flavipes workers contain multiple proteins with various antibacterial activities against a common Gram-positive soil bacterium B. subtilis.