Article

Internal Repetition and Intraindividual Variation in the rDNA ITS1 of the Anopheles punctulatus Group (Diptera: Culicidae): Multiple Units and Rates of Turnover

CSIRO, Long Pocket Laboratories Indooroopilly QLD 4068 Australia
Journal of Molecular Evolution (Impact Factor: 1.86). 12/2008; 68(1):66-79. DOI: 10.1007/s00239-008-9188-z

ABSTRACT The rapid divergence of repetitive sequences makes them desirable markers for phylogenetic studies of closely related groups,
provided that a high level of sequence homogeneity has been maintained within species. Intraspecific polymorphisms are found
in an increasing number of studies now, and this highlights the need to determine why these occur. In this study we examined
intraindividual variation present in the first ribosomal internal transcribed spacer (ITS1) from a group of cryptic mosquito
species. Individuals of the Anopheles punctulatus group contained multiple ITS1 length variants that ranged from 1.2 to 8.0kb. Nucleotide and copy number variation for several
homologous internal repeats is common, yet the intraspecific sequence divergence of cloned PCR isolates is comparable to that
of other mosquito species (~0.2–1.5%). Most of the length variation is comprised of a 5′-ITS1 repeat that was identified as
a duplication of a conserved ITS2 region. Secondary structure conservation for this repeat is pronounced and several repeat
types that are highly homogenized have formed. Significant interspecific divergence indicates a high rate of evolutionary
change for this spacer. A maximum likelihood tree constructed here was congruent with previous phylogenetic hypotheses and
suggests that concerted evolution is also accompanied by interpopulation divergence. The lack of interindividual differences
and the presence of homogenized internal repeats suggest that a high rate of turnover has reduced the overall level of variation.
However, the intraindividual variation also appears to be maintained by the absence of a single turnover rate and the complex
dynamics of ongoing recombination within the spacer.

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    • "The division of populations into arrangements (1), (2) and (3) is marked by slashes *P<0.01, significant percentage values a High values but nonsignificant possibly due to low df species, such as fagaceans (Mayol and Rosselló 2001), pinaceans (Wei et al. 2003; Wei and Wang 2004), boxes (Roselló et al. 2007), mosquitoes (Fairley et al. 2005; Bower et al. 2009), lice (Leo and Barker 2002), etc. In cestodes, intragenomic ITS sequence variation has been detected in the cyclophyllideans Echinococcus spp. "
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    • "The ITS2 PCR-RFLP patterns were better defined and more clearly distinguishable among species than those for the ITS1. Also, various studies have found greater levels of sequence and length variation in the ITS1 compared with the ITS2 (Fairley et al., 2005; Bower et al., 2008; 2009). The Zapata et al. (2007) assay was subsequently optimized to include other species of the Oswaldoi Group reported in the country (Cienfuegos et al., 2008). "
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    • "Technical artifacts such as non-specific amplification can be ruled out by our sequence data, which unambiguously demonstrate the presence of both ITS-2 Groups 2 and 3 haplotypes in single individuals. Distinct ITS-2 sequence variants may be explained by either an incomplete concerted evolution of the rDNA array or heterozygocity, i.e., hybridization of individuals with divergent ITS-2 sequences (Bower et al., 2009; Koch et al., 2003; Mukabayire et al., 2001; Onyabe and Conn, 1999). Incomplete concerted evolution appears highly unlikely for several reasons. "
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