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The failure to reduce the Campylobacter contamination of intensively reared poultry may be partially due to Campylobacter resisting disinfection in water after their internalization by waterborne protozoa. Campylobacter jejuni and a variety of waterborne protozoa, including ciliates, flagellates, and alveolates, were detected in the drinking water of intensively reared poultry by a combination of culture and molecular techniques. An in vitro assay showed that C. jejuni remained viable when internalized by Tetrahymena pyriformis and Acanthamoeba castellanii for significantly longer (up to 36 h) than when they were in purely a planktonic state. The internalized Campylobacter were also significantly more resistant to disinfection than planktonic organisms. Collectively, our results strongly suggest that protozoa in broiler drinking water systems can delay the decline of Campylobacter viability and increase Campylobacter disinfection resistance, thus increasing the potential of Campylobacter to colonize broilers.

The sequence of a third member of the Tetrahymena pyriformis chaperonin CCT ('chaperonin containing TCP1') subunit gene family is presented. This gene, designated TpCCT alpha, is the orthologue of the mouse chaperonin gene TCP1/CCT alpha. To characterize the CCT complex in this ciliate, we have produced polyclonal antibodies against synthetic peptides based on C-terminal sequences deduced from the primary sequences of the TpCCT alpha, TpCCT gamma and TpCCT eta subunits. We have also used polyclonal antibodies produced against recombinant yeast CCT alpha and CCT beta subunits. Using these antibodies, we show that Tetrahymena cells contain a hetero-oligomeric CCT chaperonin comprising at least seven distinct subunits. Three of these were assigned to specific TpCCT genes, whereas a fourth was recognized by the polyclonal antibody against yeast CCT beta, suggesting that this gene is also present in the ciliate. The CCT complex also contains other unidentified proteins that were recognized by the polyclonal antibody UM-1, raised against the putative ATP binding domain of the chaperonin proteins. TpCCT alpha gene expression was shown in exponentially growing cells and cells regenerating their cilia for different periods to have a similar pattern to the previously identified genes TpCCT gamma and TpCCT eta, and also to tubulin genes.

Genomic exclusion, an unusual cytogenetic sequence during mating in Tetrahymena pyriformis, results in the production of homozygous germinal nuclei by the diploidization of haploid nuclei following meiosis. A method is presented for selecting cells that have made new somatic nuclei from these homozygous germinal nuclei, a step necessary for phenotypic expression of new mutations; this variation of the normal set of events is termed short-circuit genomic exclusion. The utility of thisapproach for obtaining induced mutations is demonstrated by the isolation and analysis of a strain homozygous for a recessive mutation conferring resistance to 2-fluoroadenosine. Occurring in about 5% of the unmutagenized pairs in specific crosses, short-circuit genomic exclusion should be of general use for the isolation of dominant or recessive induced mutations in this protozoan.

A critical comparison of Coulter, viable, and microscope counts for several mixed cultures of microorganisms has been made. This investigation shows that Coulter counting can provide reliable estimates of microbial numbers in mixed cultures. Precautions and limitations of Coulter counting in mixed cultures are discussed.