Genome fragment of Wolbachia endosymbiont transferred to X chromosome of host insect - PubMed (original) (raw)

Genome fragment of Wolbachia endosymbiont transferred to X chromosome of host insect

Natsuko Kondo et al. Proc Natl Acad Sci U S A. 2002.

Abstract

The adzuki bean beetle, Callosobruchus chinensis, is triple-infected with distinct lineages of Wolbachia endosymbiont, wBruCon, wBruOri, and wBruAus, which were identified by their wsp (Wolbachia surface protein) gene sequences. Whereas wBruCon and wBruOri caused cytoplasmic incompatibility of the host insect, wBruAus did not. Although wBruCon and wBruOri were easily eliminated by antibiotic treatments, wBruAus persisted over five treated generations and could not be eliminated. The inheritance pattern of wBruAus was, surprisingly, explained by sex-linked inheritance in male-heterozygotic organisms, which agreed with the karyotype of C. chinensis (2n = 20, XY). Quantitative PCR analysis demonstrated that females contain around twice as much wsp titer as males, which is concordant with an X chromosome linkage. Specific PCR and Southern blot analyses indicated that the wBruAus-bearing strain of C. chinensis contains only a fraction of the Wolbachia gene repertoire. Several genome fragments of wBruAus were isolated using an inverse PCR technique. The fragments exhibited a bacterial genome structure containing a number of ORFs typical of the alpha-proteobacteria, although some of the ORFs contained disruptive mutations. In the flanking region of ftsZ gene, a non-long terminal repeat (non-LTR) retrotransposon sequence, which is typical of insects but not found from bacteria, was present. These results strongly suggest that wBruAus has no microbial entity but is a genome fragment of Wolbachia endosymbiont transferred to the X chromosome of the host insect.

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Figures

Fig 1.

Fig 1.

Specific PCR detection of Wolbachia before and after tetracycline treatment. wBruCon, wBruOri, and wBruAus were detected by PCR using specific primers. 1, untreated; 2, after treatment for 1 generation; 3, after treatment for 5 generations; 4, 10 generations after treatment for 1 generation. C, wBruCon; O, wBruOri; A, wBruAus; M, DNA size markers (1,000, 700, 500, 400, 300, 200, and 100 bp from top to bottom). Each sample contains whole DNA from an adult female. Although data from only one insect for each treatment were shown, reproducibility of the results was confirmed for more than 10 individuals.

Fig 2.

Fig 2.

Inheritance of wBruAus. (A) Cross between wBruAus-bearing (COA) males and wBruAus-free (CO) females. (B) Cross between wBruAus-free (CO) males and wBruAus-bearing (COA) females. Shade indicates presence of wBruAus. The inheritance patterns are in agreement with X-linked inheritance. Sex chromosome types deduced are shown in rectangles. XA means the X chromosome carrying wBruAus. Numbers beneath the rectangles are the number of offspring obtained.

Fig 3.

Fig 3.

Sex-linked difference in titer of wBruAus. Titers of wBruAus in adult males and females were quantified in terms of wsp gene copies per nanogram of total insect DNA. The difference was statistically significant (Mann–Whitney U test, P < 0.001).

Fig 4.

Fig 4.

Specific PCR detection of Wolbachia genes from the original triple-infected strain jC and the tetracycline-treated, wBruAus-bearing strain jCAus. Lanes 1, jC; lanes 2, jCAus; lanes 3, no template control; lanes M, DNA size markers (1,500, 1,000, 700, 500, 400, 300, and 200 bp from top to bottom). A faint band in lane 2 of the 16S rDNA panel is due to a nonspecific PCR product.

Fig 5.

Fig 5.

Structure of the genome fragments of wBruAus obtained by inverse PCR, aligned with genome sequences of wMel, a strain of Wolbachia from D. melanogaster. (A) A 4.1-kb fragment containing wsp gene. (B) An 11.4-kb fragment containing ftsZ gene. Arrows indicate the position of intermittent stop codons. Arrowheads show the position of frame-shift substitutions. Filled ORFs contain either stop codons or frame-shift substitutions, whereas shaded ORFs are structurally intact. A non-LTR retrotransposon-like sequence was located upstream of the ftsZ gene. All ORFs on wMel genome fragments are structurally intact. Question marks indicate unidentified ORFs.

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