Acriflavine-Resistant Mutant of Streptococcus cremoris

Abstract

Selection for resistance to acriflavine in Streptococcus cremoris resulted in cross-resistance to the drugs neomycin, streptomycin, ethidium bromide, mitomycin C, and proflavine. Furthermore, the mutants showed resistance to lytic bacteriophages to which the parental strain was sensitive, and, unlike the parent, the mutants grew well at higher temperatures (40°C). Revertants selected independently either for temperature sensitivity or for acriflavine sensitivity lost resistance to all the drugs and dyes but retained the bacteriophage resistance phenotype. The acriflavine-resistant mutation resulted in an increase in resistance by the bacterial cells to sodium dodecyl sulfate, a potent solvent of lipopolysaccharide and lipoprotein. It is suggested that the acriflavine resistance mutation determines the synthesis of a membrane substance resistant to higher temperatures.

Isolation and Characterization of Plasmid DNA in Streptococcus cremoris

“…In any event, this phenotype did not resemble Lac-Prtmutants previously described from S. lactis C2 (7,21). Sinha (30) has recently selected mutants of ML1, in elevated concentrations of the curing agent acriflavine which show altered phage and temperature sensitivities. The nearly unchanged size of a satellite peak of DNA in a CsCl-EB gradient from his isolate led to his conclusion that plasmids were neither eliminated nor amplified.…”

“…The supernatant (cleared lysate) containing the supercoiled DNA was further purified by the dye-buoyant density procedure of Radloff et al (29). CsCl-EB gradients contained 2 to 3 ml of cleared lysate, 4.5 ml of TES buffer [50 mM NaCl, 5 mM ethylenediaminetetraacetic acid (EDTA), 30 mM tris(hydroxymethyl)aminomethane (Tris)], pH 8.0, 7 g of CsCl, and 250 /g of EB per ml and were adjusted with TES buffer to obtain a refractive index of 1.3885 to 1.3914. Gradients were formed in Beckman Type 40 or 50 Ti fixed-angle rotors at 37,000 to 41,000 rpm at 15°C for 48 to 62 h.…”

“…McKay (1) to contain only two plasmids, and one was linked to lactose fermentation. Sinha (30) recently reported a satellite peak of covalently closed circular (CCC) DNA in a cesium chloride-ethidium bromide (CsCl-EB) density gradient of cleared lysate material from S. cremoris ML1. Several investigators (20,28) have also speculated that the appearance of slow acid-producing variants of S. cremoris, resulting from deficiencies in the cell wall-associated proteinase, might be linked to the loss of plasmid DNA.…”

Alteration of Host Specificity to Lytic Bacteriophages in Streptococcus cremoris

“…Effect of AF resistance mutation on bacteriophage infection. It was reported that the AF-resistant mutant strain ML1-11 was resistant to all of the phages to which the parent ML1 was sensitive (23). The question was then raised as to whether the mutant would be resistant to all other known phages of lactic acid bacteria.…”

“…The spontaneous and induced revertants that had lost AF resistance (15 ytg of AF per ml) were obtained by replica plating as described earlier (23). Preparation of phage stock.…”