ABSTRACT
Alveolar echinococcosis (AE)--caused by the cestode Echinococcus multilocularis--is a severe zoonotic disease found in temperate and arctic regions of the northern hemisphere. Even though the transmission patterns observed in different geographical areas are heterogeneous, the nuclear and mitochondrial targets usually used for the genotyping of E. multilocularis have shown only a marked genetic homogeneity in this species. We used microsatellite sequences, because of their high typing resolution, to explore the genetic diversity of E. multilocularis. Four microsatellite targets (EmsJ, EmsK, and EmsB, which were designed in our laboratory, and NAK1, selected from the literature) were tested on a panel of 76 E. multilocularis samples (larval and adult stages) obtained from Alaska, Canada, Europe, and Asia. Genetic diversity for each target was assessed by size polymorphism analysis. With the EmsJ and EmsK targets, two alleles were found for each locus, yielding two and three genotypes, respectively, discriminating European isolates from the other groups. With NAK1, five alleles were found, yielding seven genotypes, including those specific to Tibetan and Alaskan isolates. The EmsB target, a tandem repeated multilocus microsatellite, found 17 alleles showing a complex pattern. Hierarchical clustering analyses were performed with the EmsB findings, and 29 genotypes were identified. Due to its higher genetic polymorphism, EmsB exhibited a higher discriminatory power than the other targets. The complex EmsB pattern was able to discriminate isolates on a regional and sectoral level, while avoiding overdistinction. EmsB will be used to assess the putative emergence of E. multilocularis in Europe.
Subject(s)
Echinococcosis, Hepatic/parasitology , Echinococcosis/veterinary , Echinococcus multilocularis/classification , Echinococcus multilocularis/genetics , Microsatellite Repeats/genetics , Polymorphism, Genetic , Alaska , Animals , Animals, Domestic , Animals, Wild , Asia , Canada , Cluster Analysis , Echinococcosis/epidemiology , Echinococcosis/parasitology , Echinococcosis, Hepatic/epidemiology , Europe , Genotype , Geography , Humans , Molecular EpidemiologyABSTRACT
In order to explore the genetic diversity within Echinococcus multilocularis (E. multilocularis), the cestode responsible for the alveolar echinococcosis (AE) in humans, a microsatellite, composed of (CA) and (GA) repeats and designated EmsB, was isolated and characterized in view of its nature and potential field application. PCR-amplification with specific primers exhibited a high degree of size polymorphism between E. multilocularis and Echinococcus granulosus sheep (G1) and camel (G6) strains. Fluorescent-PCR was subsequently performed on a panel of E. multilocularis isolates to assess intra-species polymorphism level. EmsB provided a multi-peak profile, characterized by tandemly repeated microsatellite sequences in the E. multilocularis genome. This "repetition of repeats" feature provided to EmsB a high discriminatory power in that eight clusters, supported by bootstrap p-values larger than 95%, could be defined among the tested E. multilocularis samples. We were able to differentiate not only the Alaskan from the European samples, but also to detect different European isolate clusters. In total, 25 genotypes were defined within 37 E. multilocularis samples. Despite its complexity, this tandem repeated multi-loci microsatellite possesses the three important features for a molecular marker, i.e. sensitivity, repetitiveness and discriminatory power. It will permit assessing the genetic polymorphism of E. multilocularis and to investigate its spatial distribution in detail.
Subject(s)
Echinococcus multilocularis/genetics , Genetic Variation , Microsatellite Repeats , Tandem Repeat Sequences/genetics , Animals , Base Sequence , DNA, Helminth/isolation & purification , Echinococcosis, Hepatic/diagnosis , Echinococcosis, Hepatic/genetics , Echinococcus granulosus/genetics , Echinococcus multilocularis/isolation & purification , Foxes/parasitology , Genomic Instability , Humans , Mice , Molecular Sequence Data , Phylogeny , Polymerase Chain Reaction , Polymorphism, Genetic , Reproducibility of Results , Rodentia/parasitology , Sensitivity and Specificity , Sequence Homology, Nucleic Acid , Sheep/parasitologyABSTRACT
In this study it was investigated whether the "Einsiedler" warmblood horse, a historically old horse population from central Switzerland (Abbey of Einsiedeln), is distinguishable from micellaneous horse breeds, using molecular genetic techniques. The breeding history of Einsiedler horses is characterised by systematic line breeding through the dams. Therefore, two Einsiedler dam lines (N = 28), going back to the middle of the 19th century according to pedigree entries, were the focus of the survey. Random samples of diverse warmblood horse populations, but also samples from more distinct types of horse breeds, served as comparison populations (N = 52). Variation in the mitochondrial genome appeared to be only partially informative to demarcate the studied horses, as horses of distinct breeds may share identical mtDNA sequence fragments. Both dam lines revealed haplotypes commonly found in Iberian horse breeds. This is to take as an indication on the genetic origin of Einsiedler horses. Furthermore, the Klima dam line held a homologous mtDNA sequence fragment with E. ferus przewalskii. Therefore, this seems to be a phylogenetically old haplotype. The analysis of microsatellite loci revealed that horses from the two Einsiedler dam lines were in fact distinguishable from more distinct types of horses, but not from closely related European warmblood horse breeds and English thoroughbred.
Subject(s)
DNA, Mitochondrial/analysis , Horses/genetics , Microsatellite Repeats/genetics , Animals , Breeding , Female , Genetic Variation , Male , Pedigree , Phylogeny , SwitzerlandABSTRACT
The multilocus probe pV47 detected an average of nine bands in cattle between 23 kb and 4 kb. Band sharing was estimated for three groups of unrelated animals. The first group comprised 20 individuals of 12 different breeds, the second group 10 individuals of the Swiss Simmental population and the third group 11 individuals of the Swiss Brown Swiss population. The band sharing probabilities were 33%, 42% and 58% respectively. The DNA fingerprints of 38 offspring with a total of 277 bands revealed no bands that could not be traced to the parents.
Subject(s)
Cattle/genetics , DNA Fingerprinting , DNA Probes , Animals , Breeding , Chromosome Banding , DNA Fingerprinting/methods , Female , Genetic Linkage , MaleABSTRACT
Losses caused by inherited diseases affect not only the breeder's income but also the breeding programs themselves. In Switzerland the cardiomyopathy of cattle and the syndrome of arachnomelia and arthrogryposis are currently being investigated in order to find genetic markers for these diseases. It seems that DNA fingerprinting with multilocus probes is the most efficient approach available at present. Microsatellites analyzed with the polymerase chain reaction technique will provide a usable marker map within the next ten years and will therefore provide a perfect tool to find markers for hereditary diseases.
Subject(s)
Cattle Diseases/diagnosis , DNA Fingerprinting , Genetic Diseases, Inborn/diagnosis , Animals , Arthrogryposis/diagnosis , Arthrogryposis/genetics , Arthrogryposis/veterinary , Cardiomyopathies/diagnosis , Cardiomyopathies/genetics , Cardiomyopathies/veterinary , Cattle , DNA/isolation & purification , DNA Probes , Genetic Markers , Organ Specificity , Polymerase Chain ReactionABSTRACT
The IgD concentrations of eluates of artificial bloodstains and of the corresponding sera from 40 subjects of different ages were measured by single radial immunodiffusion. IgD was found in bloodstains stored up to 53 days, i.e. IgD storage stability is sufficient for forensic purposes. Since serum IgD concentrations of individuals, in particular of adults, are almost invariable and serum levels of different individuals can vary by more than a 1000-fold, the discrimination of bloodstains on the basis of IgD is generally possible. Thus IgD constitutes a further marker in antibody profiling of bloodstains.
