Characterization of antibacterial activity of a novel environmental isolate of Serratia plymuthica

Abstract

Serratia species are an important source of antimicrobial compounds with numerous studies concerning their production of secondary metabolites with applications in medicine, pharmaceutics and biotechnology. This study continued investigations on the antibacterial activity of an environmental isolate of Serratia plymuthica. This activity could be detected on agar plates against Gram-positive bacteria. A previous study reported the isolation of KmR transposon mutants deficient in the activity identified on agar plates but appeared to possess another activity secreted in liquid media. Those mutants had Tn5 insertions into genes encoding polyketide synthases (PKSs). The multiple antibacterial activities of S. plymuthica indicate the production of antimicrobials that can diffuse into aqueous environments in addition to the ones closely bound to the outer cell surface and secreted into solid mediums. This raised the question about the relationship between the two activities, were they distinct or related? This study investigated the secreted antibacterial activity of S. plymuthica using transposon mutagenesis to create mutants deficient in the secreted antibacterial activity and to identify the genes involved in antibacterial production. Cultivation parameters such as; culture medium, aeration and temperature had a clear impact on the production of the secreted antibacterial activity. Growing S. plymuthica in MRS broth at 16oC for 48h-72h produced the highest amount of inhibitory activity detected in the CFCS. The secreted antibacterial activity was thermo-stable and remained active even after sterilization (121°C/15mins) and was insensitive to methanol, Proteinase K, Tween and pH. However, the antibacterial activity faded after few days even when stored at (4oC). The antibacterial activity was bactericidal and targeted a sub-set of Gram-postive bacteria especially Bacillaceae. The CFCS precipitated in the presences of solvents such as methanol, Tris-buffer and Tween. Structural characterization of the antibacterial compound(s) was confirmed via NMR and COSY spectrometry and showed the presence of two polyketide compounds resembling the macrolide antibiotics erythromycin and rapamycin produced by type I PKSs. Erythromycin and rapamycin are anti-bacterial and immune-suppressant compounds. Results of this study indicate that the secreted antibacterial activity of S. plymuthica is synthesized by PKSs that are distinct from other known Serratia PKSs. Results are also consistent with the possibility of S. plymuthica possessing separate biochemical pathways leading to two distinct antibacterial activities or a branched biochemical pathway. However, future studies are needed to reveal if the genes responsible for the two activities are in separate genomic locations and regulated in different manners to prove that the strain possesses two separate biochemical pathways leading to the two distinct activities. Further work is needed to unlock these unanswered questions. This includes genomic studies in order to identify new secondary metabolites with future importance that might be produced by this novel strain of S. plymuthica.