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Biomedical subjects

R Prichard

Publications and source records attributed to R Prichard.

At least 19 recordsLinked to original sources
PubMed1998

Ivermectin resistance in nematodes may be caused by alteration of P-glycoprotein homolog.

Resistance to ivermectin and related drugs is an increasing problem for parasite control. The mechanism of ivermectin resistance in nematode parasites is currently unknown. Some P-glycoproteins and multidrug resistance proteins have been found to act as membrane transporters which pump drugs from the cell. A disruption of the mdrla gene, which encodes a P-glycoprotein in mice, results in hypersensitivity to ivermectin. Genes encoding members of the P-glycoprotein family are known to exist in nematodes but the involvement of P-glycoprotein in nematode ivermectin-resistance has not been described. Our data suggest that a P-glycoprotein may play a role in ivermectin resistance in the sheep nematode parasite Haemonchus contortus. A full length P-glycoprotein cDNA from H. contortus has been cloned and sequenced. Analysis of the sequence showed 61-65% homology to other P-glycoprotein/multidrug resistant protein sequences, such as mice, human and Caenorhabditis elegans. Expression of P-glycoprotein mRNA was higher in ivermectin-selected than unselected strains of H. contortus. An alteration in the restriction pattern was also found for the genomic locus of P-glycoprotein derived from ivermectin-selected strains of H. contortus compared with unselected strains. P-glycoprotein gene structure and/or its transcription are altered in ivermectin-selected H. contortus. The multidrug resistance reversing agent, verapamil, increased the efficacy of ivermectin and moxidectin against a moxidectin-selected strain of this nematode in jirds (Meriones unguiculatus). These data indicate that a P-glycoprotein may be involved in resistance to ivermectin and other macrocyclic lactones in H. contortus.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

Application of molecular biology in veterinary parasitology.

The number of applications of molecular biology in veterinary parasitology is increasing rapidly. The techniques used with eukaryotic cells are generally applicable to the study of parasites and their hosts. The polymerase chain reaction is particularly important for identification and diagnosis of parasites, as well as for many other applications. With species and type specific probes or primers, sensitivities and specificities unheard of with conventional techniques can be achieved. The accumulation of more information on the DNA sequences of parasites will reveal many more unique sequences which can be used for identification, diagnosis, molecular epidemiology, vaccine development and for studying the evolutionary biology and the physiology of parasites and the host-parasite relationship. Similarly, the completion of genome projects on host organisms will greatly assist efforts to select for hosts that are genetically resistant to parasite infection. The study of the molecular biology of antiparasitic drug receptors, potential targets for chemotherapy, and the molecular genetics of drug resistance will allow molecular screens to be used with combinatorial chemistry in the search for new antiparasitic drugs, improvements to existing chemotherapeutic families and better diagnosis and monitoring of drug resistance. While there is a proliferation of molecular biology techniques, the availability of simple kits and of automated techniques and services for sequencing, library construction and oligonucleotide synthesis and other procedures is making it easier for non-specialists to apply many of the common techniques of molecular biology. Molecular biology and the benefits from its application are relevant for veterinary parasitologists in developing countries as well as developed countries and we should introduce aspects of molecular biology to the teaching and training of veterinary parasitologists.

Animals↗
PubMed1994

Anthelmintic resistance.

Anthelmintic resistance is widespread in nematode parasites of sheep, goats and horses. Resistance is also developing in nematode parasites of cattle and has been detected in pig parasites. Benzimidazole, levamisole/morantel and ivermectin resistances occur in nematodes of sheep and goats and closantel resistance has been found in Haemonchus contortus. Anthelmintic resistance is likely to develop wherever anthelmintics are frequently used and be detected if it is investigated. Worm count or egg count reduction after treatment are useful for the detection of all types of anthelmintic resistances. More economical, faster and more sensitive in vitro assays for the detection of anthelmintic resistance have been developed. Some, such as the egg hatch assay are specific for a particular class of anthelmintic, whilst others such as larval development assays can be used with most anthelmintics. Improvements in our understanding of the biochemistry and molecular genetics of anthelmintic actions should lead to the development of more sensitive assays for the detection of anthelmintic resistance in individual nematodes. Levamisole/morantel resistance appears to be associated with alterations in cholinergic receptors in resistant nematodes. Ivermectin appears to act by binding to a glutamate receptor of a membrane chloride channel. This receptor has been expressed in vitro so that further studies of the interaction of ivermectin with this receptor and its possible alteration in ivermectin resistance will be feasible. Benzimidazole resistance in nematodes and fungi appears to be associated with an alteration in beta-tubulin genes which reduces or abolishes the high affinity binding of benzimidazoles for tubulin in these organisms. This knowledge can be exploited for DNA probes for benzimidazole resistance/susceptibility in individual organisms.

Animals↗
PubMed1993

Characterization and biological activities of anti-Brugia pahangi tubulin monoclonal antibodies.

Three monoclonal antibodies (mAb) specific to beta-tubulin were used to investigate the heterogeneity of tubulins from nematodes and mammals. Western blot analysis of one-dimensional SDS-PAGE showed that anti-Brugia pahangi tubulin mAb 1B6 and P3D react with epitope(s) specific to nematode beta-tubulin and recognize tubulin from adults and microfilariae of B. pahangi, adult B. malayi and Dirofilaria immitis, eggs of Haemonchus contortus and adult Ascaris suum. However, the same mAb did not recognize tubulin from trophozoites of Giardia lamblia, pig brain or 3T3 mouse fibroblast cells. In two-dimensional SDS-PAGE, mAb 1B6 recognized one isoform of beta-tubulin and mAb P3D recognized two beta-tubulin isoforms. Limited proteolysis showed that mAb 1B6 reacted with the amino-terminal fragments of beta-tubulin. In contrast, mAb P3D recognized the carboxy-terminal fragments of beta-tubulin. In ELISA, mAb P3D reacted with an 18 amino acid peptide corresponding to residues 430-448 of B. pahangi beta-tubulin. These observations confirm that the epitope of mAb P3D is located on the extreme carboxy-terminal region. Immunogold labelling of adult B. pahangi sections with mAb P3D revealed the presence of beta-tubulin isoforms in the cuticle, hypodermal layer and somatic muscle blocks of B. pahangi. Under in vitro conditions, mAb P3D caused 80% reduction in worm viability, during exposure over 48 h.

Animals↗