A genetic linkage map for the apicomplexan protozoan parasite Eimeria maxima and comparison with Eimeria tenella

Eimeria maxima is one of the seven Eimeria spp. that infect the chicken and cause the disease coccidiosis. The well characterised immunogenicity and genetic diversity associated with E. maxima promote its use in genetics-led studies on avian coccidiosis. The development of a genetic map for E. maxima, presented here based upon 647 amplified fragment length polymorphism markers typed from 22 clonal hybrid lines and assembled into 13 major linkage groups, is a major new resource for work with this parasite. Comparison with genetic maps produced for other coccidial parasites indicates relatively high levels of genetic recombination. Conversion of ∼14% of the markers representing the major linkage groups to sequence characterised amplified region markers can provide a scaffold for the assembly of future genomic sequences as well as providing a foundation for more detailed genetic maps. Comparison with the Eimeria tenella genetic map produced 10 years ago has revealed a less biased marker distribution, with no more than nine markers mapped within any unresolved heritable unit. Nonetheless, preliminary bioinformatic characterisation of the three largest publicly available genomic E. maxima sequences suggest that the feature-poor/feature-rich structure which has previously been found to define the first sequenced E. tenella chromosome also defines the E. maxima genome. The significance of such a segmented genome and the apparent potential for variation in genetic recombination will be relevant to haplotype stability and the longevity of future anticoccidial strategies based upon multiple loci targeted by novel chemotherapeutic drugs or recombinant subunit vaccines.     

Synaptonemal complex karyotype of Eimeria tenella

In most organisms, biological variability rests on the behaviour of the chromosomes in the meiotic context. Despite the importance of meiosis, very little is known about the meiotic behaviour of the Eimeria chromosomes. The aim of the present study is to describe the standard synaptonemal complex karyotype from Eimeria tenella oocyst spreads by electron microscopy. For that purpose, complete sets of pachytene synaptonemal complexes were obtained and the morphological pachytene karyotype was determined. The authors used a previously reported method that overcomes the difficulty of the extreme resistance of protozoan oocysts to disruption and permits the release of intact meiotic chromosomes. The chromosomes were selected under a light microscope and those selected were stained with phosphotungtic acid and studied by transmission electron microscopy. The authors confirmed 14 chromosomes, which were observed as synaptonemal complexes, and the karyotype was constructed by arranging synaptonemal complexes according to their relative lengths and kinetochore position. Components of the synaptonemal complex, lateral elements, central element, recombination nodules and kinetochore were observed. Measures of the kynetochore, width of the synaptonemal complex, diameter of the recombination nodule and length of the telomeres are given. Minimal and no significant differences were found between measures of chromosomes isolated from different Eimeria tenella strains. To the best of our knowledge, the present investigation for the first time identifies and describes the morphological characteristics of the synaptonemal complex of Eimeria tenella during the meiosis that occurs within the oocysts. In addition, the authors provide evidence of the presence of recombination nodules, suggesting that the recombination process may play an important role in the molecular evolution of this parasite.     

Mannitol metabolism in Eimeria tenella.

, and 1992. Mannitol metabolism in Eimeria tenella. International Journal for Parasitology 22: 1157–1163. Unsporulated oocysts of Eimeria tenella contain large quantities of carbohydrates, namely amylopectin, mannitol and glucose. Analysis of the carbohydrate content of sporulating oocysts revealed that mannitol content increased markedly during early stages of sporogony (first 4–6 h) but slowly diminished during the next 40 h of sporulation. Accumulation of mannitol was accompanied by a rapid decrease in amylopectin and free glucose, suggesting that mannitol might be synthesized from glucose released from amylopectin. Mannitol was also detected in sporozoite and merozoite extracts. All four mannitol cycle enzymes were detected in oocysts. Sporozoites excysted in vitro had lower activities of all four enzymes. Mannitol-1 -phosphatase and mannitol dehydrogenase activity was also detected in merozoites obtained from the second stage schizonts. Sporozoites incubated with ¹⁴C-glucose accumulated radioactively labelled precursor continuously for over 12 h and some of the ¹⁴C-glucose was converted into ¹⁴C-mannitol. These results indicate that mannitol plays an important role in the metabolism and development of the intracellular stages of the parasite.     

Monosaccharide transport by Eimeria tenella sporozoites

Eimeria tenella sporozoites were incubated in the presence of 3 different [14C]-labeled sugars; D-glucose, 2-deoxy-D-glucose and 3-O-methyl-D-glucose. The initial velocity, Vi, of uptake of D-glucose and 2-deoxy-D-glucose was similar, 41 micrograms/10(10) sporozoites/min and 46 micrograms/10(10) sporozoites/min, respectively; whereas that for 3-O-methyl-D-glucose was significantly lower, 17 micrograms/10(10) sporozoites/min. Initial velocity studies also revealed that glucose uptake was a saturable event, with an apparent KT of 20 mM and an apparent Vmax of 312 micrograms/10(10) sporozoites/min. Uptake was unaffected by exogenous sodium levels or the presence of ouabain. However, 0.1 mM phloretin significantly inhibited glucose uptake. Thus, it would appear that E. tenella sporozoites possess a Na-independent, phloretin-sensitive, carrier-mediated monosaccharide-transport system.     

Suppression of Eimeria tenella Sporulation by Disinfectants

The disinfectant effects (DEs) of 10 types of chemicals, defined by their ability to destroy or inhibit oocysts and consequently prevent sporulation of Eimeria tenella field isolate, were evaluated in vitro. Correct species assignments and sample purities were confirmed by the singular internal transcribed spacer (ITS)-PCR analysis. A total of 18 treatments were performed, and the disinfection suppression levels were 75.9% for 39% benzene + 22% xylene (1:10 dilution), 85.5% for 30% cresol soup (1:1 dilution), and 91.7% for 99.9% acetic acid (1:2 dilution) group. The results indicate that acetic acid, cresol soup, and benzene+xylene are good candidates for suppression of E. tenella oocyst sporulation.     

A comparison of the immunosuppressive effects of salivary gland extracts from two laboratory strains of Boophilus microplus

This study addresses three questions related to the immune response of cattle to tick salivary gland extracts. Firstly, is there a difference in the inhibition of proliferation of Concanavalin A (ConA) stimulated bovine lymphocytes induced by salivary gland extracts of the N and Y strains of Boophilus microplus? Second, is there a difference in the development rate of the Y and N tick strains? Third, does the host affect the inhibitory effect of salivary gland extract on the proliferation of ConA stimulated lymphocytes from the two tick strains? Salivary gland extract of the Y strain inhibited in vitro proliferation of lymphocytes stimulated by ConA significantly more than that of the N strain, when each strain was raised on different animals. A difference in the development rate was observed between the tick strains when raised on the same animal, with female ticks of the Y strain developing faster and reaching a greater fully engorged weight than ticks of the N strain. The difference in their rate of development did not appear to contribute to a difference in inhibitory effects of the salivary gland extracts and there was no difference between the inhibitory effects of salivary gland extracts from both strains. However, when Y strain ticks were raised on different animals, there was a significant difference in the inhibition of lymphocyte proliferation between the two salivary gland extracts. Therefore, it was concluded that there is no difference between the inhibitory effects of the two tick strains and that the host has an influence on salivary gland extract composition of B. microplus and its inhibitive properties.     

The prokaryotic characteristics of Eimeria tenella ribosomes

• 1.1. Ribosomes were purified from the oocysts of Eimeria tenella and characterized. Those in the unsporulated oocysts were mostly in the form of polysomes which became gradually dissociated during the early phase of sporulation and were mostly in the monomeric form in sporulated oocysts.
• 2.2. The monomeric E. tenella ribosome had the size of about 80S and consisted of 60 and 40S subunits. It was readily and completely dissociated to subunits at high potassium concentrations, and its subunits could not hybridize with those of chicken liver ribosome.
• 3.3. The E. tenella ribosomal RNA was estimated to have sedimentation coefficients of 5, 16 and 23S.
• 4.4. The E. tenella ribosomal proteins had a gel electrophoretic pattern similar to that of E. coli.
• 5.5. The in vitro polypeptide synthesis mediated by E. tenella ribosomes was inhibitable by tetracycline and chloramphenicol which are also active against the parasite in vivo.
• 6.6. Chloramphenicol bound to E. tenella ribosomes with a dissociation constant of about 10⁻⁵ M.
• 7.7. All the evidence demonstrates prokaryotic characteristics of the E. tenella ribosome and suggests that the parasite may be a primitive eukaryote.