Biology Reference
In-Depth Information
species,” the biological species concept will define them as two if there is
no gene flow between the populations. In this case morphological
differences and diagnostic markers, which can be used to distinguish
between species, will arise over time. The use of molecular markers to
“tag” and “track” worms is therefore an invaluable tool to explore
whether there is gene flow between Ascaris populations in pig and human
and thereby settle the taxonomic status according to the biological species
concept. This would aim to bring some better taxonomic stability to the
long and often historically confusing list of named species within Ascaris.
APPLICATION OF BIOCHEMICAL
AND MOLECULAR TOOLS
Early studies investigating the genetic diversity of Ascaris, and in
particular differences between worms from human and pig hosts, used
a variety of biochemical and immunological methods including one-
dimensional and two-dimensional electrophoresis of proteins and
isoenzyme analysis, which investigates polymorphism at isoenzyme loci
such as mannose phosphate isomerase and lactate dehydrogenase.
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Molecular techniques were first applied to investigate the genetic diver-
sity of Ascaris and transmission between pigs and humans in the early
1990s. The RAPD (random amplified polymorphic DNA) method, which
involves PCR amplification of genomic DNA using random primers to
produce a profile of amplified fragments that can be compared between
individuals/populations, was employed in a handful of studies but was
soon superseded by other methods.
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PCR amplification of mitochon-
drial and nuclear markers followed by restriction enzyme digestion to
identify polymorphic restriction sites has been widely used.
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Alter-
native methods employed to detect polymorphisms in PCR-amplified
mitochondrial (e.g. cox1 and nad1) and nuclear markers (e.g. ITS-1)
include direct sequencing
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and single-strand conformation poly-
morphism (SSCP),
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which is based on the fact that single-stranded
DNA fragments of different sequence will adopt different conformations
that can be detected by gel electrophoresis. Direct sequencing has become
increasingly popular with dramatic reductions in sequencing costs. In
contrast to these methodologies targeting specific markers, amplified
fragment length polymorphism (AFLP), a technique for whole genome
fingerprinting, has been used to detect genetic diversity in Ascaris from
humans, pigs, and captive chimpanzees in Denmark.
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Recently,
more than 30 microsatellite markers, representing autosomal and sex-
linked loci, have been described in Ascaris.
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Microsatellites are repeat-
ing sequences of 2
6 base pairs in the nuclear genome, which can vary in
length between individuals and populations. The Ascaris microsatellites
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