Production and Screening of High Yield Avermectin B1b Mutant of Streptomyces avermitilis 41445 Through Mutagenesis

Abstract

Background: Secondary metabolite production from wild strains is very low for economical purpose therefore certain strain improvement strategies are required to achieve hundred times greater yield of metabolites. Most important strain improvement techniques include physical and chemical mutagenesis. Broad spectrum mutagenesis through UV irradiation is the most important and convenient physical method.

Objectives: The present study was conducted for enhanced production of avermectin B1b from Streptomyces avermitilis 41445 by mutagenesis using ultraviolet (UV) radiation, ethidium bromide (EB), and ethyl methanesulfonate (EMS) as mutagens.

Materials and methods: S. avermitilis DSM 41445 maintained on yeast extract malt extract glucose medium (YMG) was used as inoculum for SM2 fermentation medium. Spores of S. avermitilis DSM 41445 were exposed to UV radiation for physical broad spectrum mutagenesis and to EMS and EB for chemical mutagenesis. For each mutagen, the lethality rate and mutation rate were calculated along with positive mutation rate.

Results: Avermectin B1b-hyper-producing mutant, produced using these three different methods, was selected according to the HPLC results. The mutant obtained after 45 minutes of UV radiation to the spores of S. avermitilis 41445, was found to be the best mutant for the enhanced production of avermectin B1b component (254.14 mg/L). Other avermectin B1b-hyper-producing mutants, were obtained from EMS (1 µL/mL) and EB (30 µL/mL) treatments, and yielded 202.63 mg/L and 199.30 mg/L of B1b, respectively.

Conclusions: The hereditary stability analysis of the UV mentioning 45 minutes revealed the UV exposure time for mutants and 3 represented the colony taken from the plate irradiated for 45 minutes mutant showed that the production of avermectin B1b remained constant and no reverse mutation occurred after 15 generations.

Keywords: Avermectin B1b; Hereditary Stability; Mutagenesis; Streptomyces avermitilis; Submerged Fermentation.

Figure 1.. Avermectin Chemical Structure

Figure 2.. Effects of Different UV Radiation Times of on the Survival and Lethality Rates

Figure 3.. HPLC Chromatogram

(A) Standard solution of Abamectin, (B) B1b produced from the EB 30 strain (minutes) 2, (C) B1b produced from the UV 45 strain (minutes) 3, (D) B1b produced from the EMS 50 strain (minutes) 1

Figure 4.. Effects of Different EMS (1 µL/mL) Treatment Times on the Survival and Lethality Rates

Figure 5.. Comparative Analysis of Production of Avermectin B1b by Different Mutants With the Parent Original Strain, S. avermitilis 41445