When F− cells are mixed with an excess of Hfr cells there is a lethal event which results in a decrease in the number of F− survivors. We have described and discussed the parameters affecting this phenomenon of lethal zygosis, and these include the cultural conditions of both donor and recipient cells prior to mixing and the use of aeration throughout the period of the experiment. The absence of lethal zygosis with filtrates and supernatant fluids from donors suggests a dependence on direct cell-cell contact as found in conjugation. The phenomenon, which is normally observed in liquid media, also occurs on solid media, and use of these two methods has allowed examination of strains of different mating types. Whereas most Hfr strains capable of producing normal yields of recombinants showed killing activity, no F+ and only one F′ donor produced lethal zygosis. Only F− strains were sensitive to this phenomenon. The relationship between lethal zygosis and the various stages of conjugation is discussed. Images

Characterization of Lethal Zygosis Associated with Conjugation in Escherichia coli K-12

A new prophylactic and therapeutic antimicrobial strategy based on a specific physiological target that is effectively used by killer yeasts in their natural ecological competition is theorized. The natural system exploited is the yeast killer phenomenon previously adopted as an epidemiological marker for intraspecific differentiation of opportunistic yeasts, hyphomycetes, and bacteria. Pathogenic microorganisms (Candida albicans) may be susceptible to the activity of yeast killer toxins due to the presence of specific cell wall receptors. On the basis of the idiotypic network, we report that antiidiotypic antibodies, produced against a monoclonal antibody bearing the receptor-like idiotype, are in vivo protecting animals immunized through idiotypic vaccination and in vitro mimicking the antimicrobial activity of yeast killer toxins, thus acting as antibiotics.

Interfaces of the Yeast Killer Phenomenon

The advantage in using membrane filters for crosses between P+ (donor) and P~ (recipient) strains of Vibrio cholerae (Bhaskaran, 1964; Bhaskaran, Sinha & Iyer, 1967) is probably due to more frequent and effective contacts between the cells on the surface of the membrane than is possible in fluid media with actively motile cells of this species. The transfer of the fertility (P) factor itself, as well as the yield of genetic recombinants, was considerably improved by this procedure. It seemed worthwhile, therefore, to utilize this technique for crosses between strains of V. cholerae and V. El Tor which have given irregular and often negative results in the past. In particular, P+ strains of V. El Tor were never shown to yield recombinants with P~ strains, either of V. cholerae or V. El Tor, when tested by conventional procedures such as plating mixtures of washed suspensions of parental strains on selective minimal media. As a preliminary to this, mixtures of different P+ and P~ strains (in pairs, containing c. 2-5 x 10 cells of each strain derived from 4 h nutrient broth cultures) were fixed by nitration on membrane filters ('Metricel'; pore size, 0-45/*; diam., 47 mm; Gelman Instrument Co., Ann Arbor, Michigan, U.S.A.) and the proportion of P~ cells acquiring the P factor was determined, at intervals of time, when these membranes were incubated at 37 °C on the surface of sterile, moist nutrient agar plates. This was done by resuspending the culture (on the surface of the membrane) in fluid minimal medium and plating its dilutions on nutrient agar plates for the isolation of discrete colonies of the original P~ strain. When this strain was streptomycin resistant, as it was in most cases, nutrient agar containing streptomycin (500 /ig/ml) was used. Twenty randomly selected colonies, which were the progeny of the original P~ parent, were tested for the presence of the P factor which may have been acquired by conjugation with the P + partner on the membrane. The test consisted in placing a loopful of these cultures (in nutrient broth) on a semi-solid nutrient agar seeded with an indicator (P~) strain of V. cholerae (162/p.) After overnight incubation at 37 °C, P+ cultures were identified by a hazy clearing around the periphery of their growth (Bhaskaran, 1964). In our experience, this activity (possibly due to the production of a bacteriocine) is invariably associated with donor ability in V. cholerae (but not in V. El Tor). The results are recorded in Table 1. I t is clear that the transfer of P factor was less efficient when one or the other partner was V. El Tor, whereas, between

K. Bhaskaran

Papers

A lethal factor in a strain of Vibrio El Tor.