Three New Temperate Phages of Bacillus subtilis

Abstract

Three temperate bacteriophages of Bacillus subtilis were isolated from soil samples and analysed, together with all the other known temperate phages of this organism, with respect to their host range, immunity, serology and DNA restriction pattern, and by other tests. The results show that the newly isolated phages are new members of the immunity sub-group I of the group I11 of B. subtilis temperate bacteriophages. We named these new phages IGl, IG3 and IG4. We characterized the new phages with respect to host range, serology, DNA restriction pattern, potential for methylation of specific DNA sequences, immunity of lysogens to superinfection, ability to complement thymine auxotrophy of the host, and production of and resistance to betacin. The newly isolated phages behaved differently from the other known B. subtilis temperate bacteriophages. METHODS

Action of 6-(p-hydroxyphenylazo)-uracil on bacteriophage IG 1

“…The media and methods used for growing the cultures and carrying out phage assays were previously described [9]. In order to estimate the effect of HPUra on the growth of the strains CU 1065 and IGCg 225 (lysogenic and non-lysogenic for IG 1, respectively) a single colony from each strain was inoculated into 10ml of M medium.…”

“…subtilis CU 1065 (trp C 2 att SP [3) was provided by Dr. S. Zahler (Cornell University, U.S.A.) and was used as the indicator strain for phage assays and/or the control strain to determine the effect of HPUra and chloramphenicol on bacterial growth and phage multiplication. IG 1 was isolated from soil samples [9] and the strain IGCg 225 is a derivative of CU 1065 lysogenic for IG 1.…”

“…IG 1 is a Bacillus subtilis temperate phage isolated from soil samples and previously characterized [5,[8][9][10].…”

Cloning of the thymidylate synthetase gene (thyPIG 3) from theBacillus subtilis temperate phage IG 3

“…Since according to transformation data [11,20,21] the thyP 3 gene of phage <P3 T is identical to thyPIG 3, the presence of the thyPIG 3 gene in the clones was also assessed by colony hybridization using as a probe the 3z P-Bgl IIHindIII fragment of pCAT 12 [12] (see Fig. 3 C) that contains the thyP3 gene (not shown).…”

“…The thyP 3 as well as the thyP from IG 3 (gene thyPIG 3) has a strong homology to the B. subtilis thyA gene. It can be distinguished phenotypically from the B. subtilis thyB gene, since the lysogens grow well at 46 °C, like the ThyA + ThyB-mutants, and not like the T h y A -ThyB + strains that do not grow at this high temperature [11,16,20,21]. The IG 3 prophage, like the prophage ~p3 T, can be integrated in the thy A region of the host which is distinct from the normal attachment site, at the terminus of the chromosome, between ilvA and kauA [17,20,21; I.…”