ABSTRACT
For the first time we used amplified fragment length polymorphism on individual nematode parasites to analyse the genetic diversity between and within isolates during consecutive stages of increased benzimidazole resistance and of increased levamisole resistance of Haemonchus contortus. The genetic diversity of the H. contortus genome turned out to be unusually high, within and between the isolates. The difference between individuals of an isolate could be as high as between individuals of two different mammalian species that do not interbreed. During benzimidazole selection the genetic constitution of the population was changed, but did not lead to a decrease in the genetic diversity. The selection for levamisole resistance resulted in a limited reduction of the genetic diversity only after the first selection step. The extensive genetic diversity apparently has allowed a fast and flexible response of H. contortus to drug selection as shown by the appearance of drug resistant isolates. This selection however has little or no effect on the extent of the genetic diversity of these resistant isolates. Implications for more sustainable control methods are discussed.
Subject(s)
Anthelmintics/pharmacology , Haemonchus/drug effects , Haemonchus/genetics , Animals , Benzimidazoles/pharmacology , Cluster Analysis , DNA Fingerprinting , DNA, Helminth/chemistry , DNA, Helminth/isolation & purification , Drug Resistance/genetics , Genetic Variation/drug effects , Genetic Variation/genetics , Haemonchiasis/drug therapy , Haemonchiasis/parasitology , Haemonchus/isolation & purification , Levamisole/pharmacology , Multivariate Analysis , Polymerase Chain Reaction/veterinary , Polymorphism, Genetic/drug effects , Polymorphism, Genetic/genetics , Sheep , Sheep Diseases/drug therapy , Sheep Diseases/parasitologyABSTRACT
The alarming development of anthelmintic resistance in important gastrointestinal nematode parasites of man and live-stock is caused by selection for specific genotypes. In order to provide genetic tools to study the nematode populations and the consequences of anthelmintic treatment, we isolated and sequenced 59 microsatellites of the sheep and goat parasite Haemonchus contortus. These microsatellites consist typically of 2-10 tandems CA/GT repeats that are interrupted by sequences of 1-10 bp. A predominant cause of the imperfect structure of the microsatellites appeared mutations of G/C bp in the tandem repeat. About 44% of the microsatellites were associated with the HcREP1 direct repeat, and it was demonstrated that a generic HcREP1 primer could be used to amplify HcREP1-associated microsatellites. Thirty microsatellites could be typed by polymerase chain reaction (PCR) of which 27 were polymorphic. A number of these markers were used to detect genetic contamination of an experimental inbred population. The microsatellites may also contribute to the genetic mapping of drug resistance genes.
Subject(s)
Dinucleotide Repeats/genetics , Genetic Variation , Haemonchus/genetics , Animals , Antinematodal Agents/pharmacology , Benzimidazoles/pharmacology , DNA, Helminth/genetics , Drug Resistance/genetics , Genetic Markers , Genotype , Haemonchus/classification , Haemonchus/growth & development , Haemonchus/isolation & purification , Molecular Sequence Data , Sequence Analysis, DNASubject(s)
Genetic Markers , Haemonchus/genetics , Introns/genetics , Animals , Base Sequence , Conserved Sequence , DNA Primers/chemistry , DNA, Complementary/chemistry , DNA, Helminth/chemistry , Drug Resistance/genetics , Haemonchus/drug effects , Mutation , Polymerase Chain Reaction , Polymorphism, Restriction Fragment LengthABSTRACT
Several animal species such as cattle, goats, sheep, and water buffalo provide milk for dairy products. We describe a simple procedure for detecting the species origin of milk used for cheese production. DNA was isolated from Italian mozzarella or Greek feta by sequential organic extractions and resin purification. This DNA was analyzed by polymerase chain reaction-restriction fragment length polymorphism as described previously for meat samples. This procedure differentiated mozzarella made from water buffalo milk and from less expensive bovine milk and also feta cheeses made from bovine, ovine, and caprine milk.
Subject(s)
Cheese , Food Contamination , Food Inspection/methods , Milk , Animals , Cattle , Cytochrome b Group/genetics , DNA/analysis , DNA/isolation & purification , Food Handling/standards , Goats , Polymerase Chain Reaction/methods , Polymorphism, Restriction Fragment Length , Sheep , Species SpecificityABSTRACT
Polymorphic molecular markers are being identified to characterize the genomes of parasitic nematodes. The aim is to construct a map with markers evenly spread over the six chromosomes. With such a map, regions can be identified that are under selection pressure when attempts are being made to eradicate worms, be it by drugs, vaccines or genetic resistance in the sheep. Several types of markers have been developed, microsatellites, transposon-associated markers, amplified fragment length polymorphism (AFLP) and expressed sequence tag (EST) markers. Linkage groups can be constructed using several genetic crosses between inbred and drug resistant strains. EST markers will be especially important for comparative mapping with the genome of Caenorhabditis elegans, and therefore localization of the linkage group on a chromosome. It will then be possible to identify functional genes close to markers that have changed allele frequencies under selection pressure and identify the mechanisms of resistance to parasite control.
