Biology of two lysogenic phages from Bacillus thuringiensis MZ1

Abstract:

A Bacillus thuringiensis (Bt) fermentative strain MZ1 (subsp.kurstaki), from a company in Meixian County of Guangdong Province, produce toxins during sporulation and are extensively used in the field to control pest insects (Lepidoptera) in China. But some unknown or random factors that inhibited or stopped B.t growth in the fermentation can be regarded as reflecting the exist of lysogenic phage. Therefore, strain MZ1 and its lysogenic phages were studied in this paper. With indicator strain ZK1, two kind of phage plaques, one with about 3mm diameter and the other with about 1mm diameter, can be observed after strain MZ1 cultured in plates or flasks was induced by mitomycin C. Then, two lysogenic phages, namely MZTP01 and MZTP02, were isolated and characterized in biology. They belonged to family Siphoviridae, which had icosahedral heads (MZTP01:82nm×85nm; MZTP02: 75nm×55nm) and long tails (MZTP01: 220nm×18nm; MZTP02:183nm×12nm) without flexibility. Host range examination showed that six and seven (including indicator strain ZK1) out of 113 B.t strains saved in our laboratory were sensitive to MZTP01 and MZTP02, respectively. MZTP01 was more stable than MZTP02 against pH value, ultraviolet and heat treatment, but contrary against organic solvents. For MZTP01 and MZTP02, K values in the neutralization reactions were 45 and 326, respectively. Both phages had no relationship in their antigenicity. Burst size of phage MZTP02 was 175, two times more than that of MZTP01. Latent time of MZTP02 was 40min, one times shorter than that of MZTP01. It was suggested that both phages DNA be linear dsDNA through the typical absorption curves, reaction with diphenylamine, DNase sensitivity and acridine orange staining. And this was in good accord with the previous findings that all tailed phages being dsDNA moleculars. The genomic DNA and their restriction maps showed that both molecular weights should be between 9.4～23kb. Both phage genomic DNAs were digested by HindⅢ, producing eight and nine bands, respectively. The results above show that strain MZ1 is proved a double lysogen. It is considered to be the main reason of great losses in the production. The biological information of the two lysogenic phages belonging to strain MZ1 is provided to help solve the problem and the two lysogenic phages is prepared for complete DNA sequencing in the next step.