The occurrence of killer character in yeasts of various genera

Species of 7 of the 28 yeast genera in the National Collection of Yeast Cultures exhibited killing activity againstSaccharomyces cerevisiae. The highest incidence of killer yeasts was found in the genusHansenula (12 of the 29 strains examined).Saccharomyces, the best represented genus in the Collection, showed a low incidence of killer activity and many of the killer strains are hybrids with a commonS. cerevisiae parent. The activities of culture filtrates of the 59 killer yeast isolated responded differently to pH and four types of response were recognised.     

A comparison of the killer character in different yeasts and its classification

The interactions between 20 killer yeasts of various genera and species were examined. Ten distinct groups were recognised with respect to killer activity and 10 distinct groups with respect to resistance to killer action. Using both killing and resistance phenotypes, 13 classes of killer yeast were found. With the exception of Torulopsis glabrata NCYC 388, non-Saccharomyces strains of yeast were not killed by a member of the genus Saccharomyces.The killer character of the 3 killing groups of Saccharomyces identified could be cured by treatment with cycloheximide or incubation at elevated temperature and the effectiveness of these procedures was indicative of the category of killer yeast examined. Killer yeasts not belonging to the genus Saccharomyces could not be cured of their activity. Double-stranded ribonucleic acids were extracted only from Saccharomyces spp. and the molecular weights of the species present were a function of the killer class to which a strain belonged.By an analysis of the effects of proteolytic enzymes, temperature and pH on killer activity and by gel chromatography of crude preparations of killer factors, the toxins of different killer classes were shown to be biochemically distinct. However all toxins had certain properties in common consistent with there being a protein component essential to killer action.     

The ecological role of killer yeasts in natural communities of yeasts

The killer phenomenon of yeasts was investigated in naturally occurring yeast communities. Yeast species from communities associated with the decaying stems and fruits of cactus and the slime fluxes of trees were studied for production of killer toxins and sensitivity to killer toxins produced by other yeasts. Yeasts found in decaying fruits showed the highest incidence of killing activity (30/112), while yeasts isolated from cactus necroses and tree fluxes showed lower activity (70/699 and 11/140, respectively). Cross-reaction studies indicated that few killer-sensitive interactions occur within the same habitat at a particular time and locality, but that killer-sensitive reactions occur more frequently among yeasts from different localities and habitats. The conditions that should be optimal for killer activity were found in fruits and young rots of Opuntia cladodes where the pH is low. The fruit habitat appears to favor the establishment of killer species. Killer toxin may affect the natural distribution of the killer yeast Pichia kluyveri and the sensitive yeast Cryptococcus cereanus. Their distributions indicate that the toxin produced by P. kluyveri limits the occurrence of Cr. cereanus in fruit and Opuntia pads. In general most communities have only one killer species. Sensitive strains are more widespread than killer strains and few species appear to be immune to all toxins. Genetic study of the killer yeast P. kluyveri indicates that the mode of inheritance of killer toxin production is nuclear and not cytoplasmic as is found in Saccharomyces cerevisiae and Kluyveromyces lactis.     

Biology of killer yeasts

Killer yeasts secrete proteinaceous killer toxins lethal to susceptible yeast strains. These toxins have no activity against microorganisms other than yeasts, and the killer strains are insensitive to their own toxins. Killer toxins differ between species or strains, showing diverse characteristics in terms of structural genes, molecular size, mature structure and immunity. The mechanisms of recognizing and killing sensitive cells differ for each toxin. Killer yeasts and their toxins have many potential applications in environmental, medical and industrial biotechnology. They are also suitable to study the mechanisms of protein processing and secretion, and toxin interaction with sensitive cells. This review focuses on the biological diversity of the killer toxins described up to now and their potential biotechnological applications. Electronic Publication     

The occurrence of multinucleate giant cells in yeasts

Summary The spontaneous occurrence of giant cells has been observed in young cultures ofLipomyces lipofer and three different genotypes ofSaccharomyces. Stained preparations of all abnormal cultures revealed that the giant cells characteristically contained more than one nucleus, the number ranging from one to six. In both genera the phenomenon was found to be transient, for rapidlygrowing cultures arising from isolated giant cells reverted, at varying rates, to populations of small uninucleate cells which appeared normal in all respects.     

Occurrence of killer yeasts in Argentine wineries

Fifty-six wine must samples, from wineries in various regions of Argentina, were examined at different fermentation stages for the presence of killer yeast strains. The distribution of isolated killer strains was markedly different from one region to another.The authors are with the Instituto de Biotecnología, Facultad de Ingeniería, Universidad Nacional de San Juan, Av. Lib, Gral. San Martín 1109, Oeste 5400, San Juan, Argentina;     

Occurrence of killer yeasts in leaf-cutting ant nests

Killer activity was screened in 99 yeast strains isolated from the nests of the leaf-cutting antAtta sexdens against 6 standard sensitive strains, as well as against each other. Among this yeast community killer activity was widespread since 77, strains (78%) were able to kill or inhibit the growth of at least one standard strain or nest strain. Toxin production was observed in representatives of all the studied genera includingAureobasidium, Rhodotorula, Tremella andTrichosporon, whose killer activity has not yet been described.     

The occurrence of vascular tracheids inBetula and some otherBetulaceae genera

The occurrence of vascular tracheids inBetula, Alnus, Carpinus andCorylus is reported. It is established that the cells are a consistent feature in early and late wood where they are associated with vessel elements in radial multiples. The possible function and taxonomic importance of these cells are briefly considered.     

The production of 2,3-butanediol as a differentiating character in wine yeasts

The capacity to produce 2,3-butanediol by 90 strains of four different species of wine yeasts (Kloeckera apiculata, Saccharomyces cerevisiae, Saccharomycodes ludwigii, Zygosaccharomyces bailii) was tested in grape must by automated multiple development HPTLC. The total amount of 2,3-butanediol produced varied between 23mg l–1 and 857.7mg l–1 according to the yeast species. S. cerevisiae and Z. bailii behaved similarly, producing elevated amounts of 2,3-butanediol. K. apiculata and Sc. ludwigii, in contrast, were low producers. When considerable amounts of 2,3-butanediol were found, little acetoin was present; the amounts of butanediol and acetoin were characteristic of the individual species.     

The occurrence of killer,sensitive, and neutral yeasts in Brazilian Riesling Italico grape must and the effect of neutral strains on killing behaviour

 The occurrence of killer toxins amongst yeasts in Brazilian Riesling Italico grape must was investigated by using the sensitive strain EMBRAPA-26B as a reference strain at 18°C and 28°C. From a total of 85 previously isolated yeasts, 21 strains showed ability to kill the sensitive strain on unbuffered grape must/agar (MA-MB) and 0.1 M citrate/phosphate-buffered yeast extract/peptone/dextrose/agar (YEPD-MB) media both supplemented with 30 mg/l methylene blue. The killer activity of only four yeasts depended on the incubation temperature rather than the medium used. At 28°C, the strains 11B and 53B were not able to show killer action. On the other hand, strains 49B and 84B did not kill the sensitive yeast at 18°C. The killer strain EMBRAPA-91B and a commercial wine killer yeast K-1 were employed to examine the sensitivity of the isolated yeasts on YEPD-MB and MA-MB at 18°C. The sensitivity and neutral characteristics of yeasts were shown to be dependent on the medium and the killer strain. Interactions, including K^- R^-, K^- R⁺ and K⁺ R⁺ strains, simultaneously, have revealed that some K^-R⁺ strains appear to protect the K^- R^- strain against the killer toxin. Sensitive dead cells, although to a less extent, also exhibited similar protection. Kinetic studies have shown that the maximum specific growth rates were higher for the 20B YEPD-MB-sensitive strain (μ_max=0.517 h^-1) than for both the 91B (μ_max=0.428 h^-1) and K-1 (μ_max= 0.466 h^-1) killer strains. The protective capacity of neutral or sensitive cells that contaminate a fermentation, as well as the higher maximum specific growth rate of sensitive yeasts, besides other factors, may preclude the dominance of a killer strain. This protective capacity may also reduce the risk of a sensitive inoculum being killed by wild-type killer yeasts in open non-sterile fermentation. Received: 3 November 1995/Received revision: 11 March 1996/Accepted: 15 April 1996     

Rapid method for estimation of killer toxin activity in yeasts

Summary Conditions have been established for a rapid assay which enables to determine killer toxin activity in a few hours. The procedure is based upon recognition of rhodamine B-stained cells using fluorescence microscope.