David Mencher

Eimeria maxima: identification of gametocyte protein antigens.

The antigenicity of Eimeria maxima gametocyte proteins during the course of an infection and when injected into mice and rabbits was demonstrated using the Western blotting technique. Serum taken from chickens at various times postinfection reacted to a few gametocyte proteins, with the strongest reactivity seen with serum taken 14-days postinfection. Two major antigens having molecular weights of 56,000 and 82,000 were consistently detected by these sera. Using immune rabbit or mouse sera to whole gametocyte detergent extracts, the 56,000 and 82,000 molecular weight proteins were again the immunodominant antigens, despite their representing only a small proportion of the extract which was used to immunize the animals. These results, together with those obtained by Rose (1971) using recovered chicken serum to passively immunize chickens, indicate that these two gametocyte antigens may play a role in protective immunity to E. maxima.

Developmental gene expression of a 230-kilodalton macrogamete-specific protein of the avian coccidial parasite, Eimeria maxima

We prepared a cDNA library from gametocytes of Eimeria maxima and screened it using antibodies raised against an 82-kDa gametocyte antigen. One cDNA clone designated pEM230 was isolated and characterized. It encodes a portion of a 230-kDa gametocyte protein and its DNA sequence shows the presence of several tandem repeats of 42 bp. In order to determine the stage and sex specificity of the mRNA for the 230-kDa protein, Northern blotting and in situ hybridization studies were performed. The 230-kDa protein is encoded for by a 7 kb mRNA, which is expressed exclusively during the macrogamete stage with no detectable expression seen in any other stage of parasite development.

Eimeria maxima: Isolation of gametocytes and their immunogenicity in mice, rabbits, and chickens

Eimeria maxima gametocytes were isolated from infected chicken intestinal tissue by treatment with hyaluronidase and subsequent filtration through polymon filters. The isolated gametocytes were analyzed by microscopical and biochemical methods and shown to be highly enriched. The antigenicity of the gametocytes was analyzed in mice, rabbits, and chickens by ELISA and indirect immunofluorescence. Contrary to published results, we have found gametocytes to be highly immunogenic in all animals tested.

Antigenic proteins of Eimeria maxima gametocytes: Cell-free translation and detection with recovered chicken serum

RNA was extracted from isolated Eimeria maxima gametocytes and translated in a rabbit reticulocyte cell-free protein synthesis system. The major cell-free translation products from E. maxima gametocyte RNA ranged from 225 to 50 kDa, distinct and different from uninfected chicken intestine cell-free translation products. Rabbit antiserum to E. maxima gametocytes as well as recovered chicken sera specifically precipitated some of the major gametocyte cell-free products. A time course of infected intestine RNA indicated that these cell-free synthesized gametocyte antigens appear at 130 to 138 hr postinfection.

Conservation from rat to human of cytosolic phosphoenolpyruvate carboxykinase and the control of its gene expression

Structural conservation of cytosolic phosphoenolpyruvate carboxykinase protein and mRNA sequence was found in all species examined from rodents to human. The mitochondrial isoenzyme, in all species tested, represents a distinct protein. Moreover, irrespective of the ratio of cytosolic to mitochondrial isoenzyme, cytosolic phosphoenolpyruvate carboxykinase activity in the human as in the rat is controlled at the level of gene expression and through the same multiple hormonal stimulation. This evolutionary conservation of the cytosolic phosphoenolpyruvate carboxykinase structure and mode of regulation supports the enzymes physiological importance in mammals.