Detection of aphid remains in predatory insects and spiders by ELISA

An ELISA was developed which would detect and quantify ingested aphids in predators found in and around cereal crops. The detection limit of the assay was less than one hundredth of an homogenised adult aphid. Tests with 13 species of aphid showed that those which had been used as the principal immunogens reacted most strongly in the assay. Nearly a hundred species of invertebrates, both predators and alternative prey, have been tested in the assay and no evidence of significant cross-reaction was found with any of these species or with a number of samples of plant material on which aphids may be found. Aphid material could still be detected in predators which had been stored for up to 7 days in 4% formalin or 70% ethanol.     

Progress in Taxonomy of the Apicomplexan Protozoa

In 1987, 4516 species and 339 genera of the phylum Apicomplexa had been named. They consisted of the gregarines (subclass Gregarinasida) (1624 named species and 231 named genera), the hemogregarines (family Haemogregarinidae) (399 species and 4 genera), the eimeriorins (order Eimeriorida) (1771 species and 43 genera), the hemospororids (order Haemospororida) (444 species and 9 genera), the piroplasmids (order Piroplasmorida) (173 species and 20 genera), and a few others (105 species and 32 genera). The first apicomplexan protozoon was seen by Antony van Leeuwenhoek; in 1674 he saw oocysts of Eimeria stiedai in the gall bladder of a rabbit. The first member of the phylum to be named (by Dufour in 1828) was Gregarina ovata in earwigs. During the quarter century 1826–1850, 41 species and 6 genera of Apicomplexa were named. These numbers increased progressively. In the quarter century 1951–1975, 1873 new species and 83 new genera were named. Data are given for the numbers of named species and genera of apicomplexan protozoa of each group known in 1850, 1875, 1900, 1925, 1950, 1975, and 1987.     

A Comparison of Endogenous Development of Three Isolates of Cryptosporidium in Suckling Mice1

ABSTRACT. Suckling mice were used as a model host to compare the endogenous development of three different isolates of Cryptosporidium: one from a naturally infected calf, one from an immunocompetent human with a short-term diarrheal illness, and one from a patient with acquired immune deficiency syndrome (AIDS) and persistent, life-threatening, gastrointestinal cryptosporidiosis. After oral inoculation of mice with oocysts, no differences were noted among developmental stages of the three isolates in their sites of infection, times of appearance, and duration, morphology, and fine structure. Sporozoites excysted within the lumen of the duodenum and ileum, penetrated into the microvillous region of villous enterocytes, and developed into type I meronts with six or eight merozoites. Type I merozoites penetrated enterocytes and underwent cyclic development as type I meronts or they became type II meronts with four merozoites. Type II merozoites did not exhibit cyclic development but developed directly into sexual forms. Microgamonts produced £16 small, bullet-shaped microgametes, which were observed attaching to and penetrating macrogametes. Approximately 80% of the oocysts observed in enterocytes had a thick, two-layered wall. After sporulating within the parasitophorous vacuole, these thick-walled oocysts passed through the gut unaltered and were the resistant forms that transmitted the infection to a new host. Approximately 20% of the oocysts in enterocytes consisted of four sporozoites and a residuum surrounded only by a single oocyst membrane that ruptured soon after the parasite was released from the host cell. The presence of thin-walled, autoinfective oocysts and recycling of type I meronts may explain why a small oral inoculum can produce an overwhelming infection in a suitable host and why immune deficient persons can have persistent, life-threatening cryptosporidiosis in the absence of repeated oral exposure to thick-walled oocysts.     

Development of Eimeria auburnensis in Cell Cultures

SYNOPSIS. Monolayer established cell line cultures of bovine kidney (Madin-Darby) and human intestine (Intestine 407), as well as embryonic bovine tracheal and embryonic spleen cell line cultures were inoculated with E. auburnensis sporozoites and observed for a maximum of 22 days. Mature 1st generation schizonts developed in the kidney, tracheal and spleen cells. In the intestine cells, trophozoites were seen in 3 of 4 experiments, but schizonts were not found. Sporozoites penetrated cells, beginning within a few minutes after inoculation. Penetration was usually accomplished within 10 seconds, and the body of the sporozoite underwent a slight constriction as it passed thru the host cell membrane. Some sporozoites left cells. Numerous intracellular sporozoites were observed in kidney, tracheal and spleen cultures. Crescent bodies were seen in the parasitophorous vacuole as early as 1 day after inoculation. At this time, the nuclei of most intracellular sporozoites had changed from vesicular to compact. Beginning 4 days after inoculation, enlarged sporozoites and parasites having a sporozoite shape, but with 2-5 nuclei, were frequently seen. These enlarged sporozoites and sporozoite-shaped schizonts evidently transformed into trophozoites and spheroidal schizonts by means of lateral outpocketings. Few trophozoites were seen. More immature schizonts developed in kidney cells than in the other cell types. The numbers of mature schizonts observed in kidney and tracheal cells were similar, but development occurred less consistently in the latter. Few immature and mature schizonts developed in spleen cells. Mature schizonts, first seen 9 days after inoculation, were considerably smaller than those reported from calves. Some motile merozoites were seen; evidently no development beyond these occurred. The nucleus and nucleolus of host cells were enlarged; this enlargement was not as pronounced as in infections in calves. Multiple host cell nuclei were frequently observed. Degenerative changes in the cultured cells and in the parasites usually occurred, beginning 9-17 days after inoculation; these were more pronounced in the spleen cells than in the others.     

Further Studies on in vitro Development of Eimeria bovis and Attempts to Obtain Second-Generation Schizonts*

SYNOPSIS. Eimeria bovis merozoites occurred in tissue culture medium removed from Leighton tube cultures of embryonic bovine tracheal cells beginning 12-14 days after inoculation with 270,000-369,000 sporozoites per tube. The number of merozoites produced in these cultures increased daily until a peak was reached 18-21 days after inoculation. In 3 experiments an average of 2.0–15.6 million merozoites per tube was produced during the 20-day observation period. When such merozoites were frozen in liquid nitrogen and stored 26–42 days, some were motile upon thawing. These merozoites as well as others freshly obtained from cell cultures and from calves were inoculated into 11 different types of cultured mammalian cells including primary, cell line and established cell line cultures. Some merozoites were exposed to substances normally found in the lumen of the gut, before or at the time of inoculation. Altho small numbers of intracellular merozoites were found, no further development was observed. Gametocytes were observed in the cecum of a calf 4 days after merozoites from cell cultures were introduced into a ligated cecum of the calf.