Variations in the sensitivity of different primers for detecting Wolbachia in Anastrepha (diptera: tephritidae)

Wolbachia are endosymbiont bacteria of the family Rickettsiacea that are widespread in invertebrates and occur between 20 percent and 60 percent of Neotropical insects. These bacteria are responsible for reproductive phenomena such as cytoplasmic incompatibility, male killing, feminization and parthenogenesis. Supergroups A and B of Wolbachia are common in insects and can be identified using primers for 16S rDNA, ftsZ and wsp; these primers vary in their ability to detect Wolbachia. The ftsZ primer was the first primer used to detect Wolbachia in Anastrepha fruit flies. The primers for 16S rDNA, ftsZ and wsp and the corresponding PCR conditions have been optimized to study the distribution of Wolbachia and their effect on the biology of Anastrepha in Brazil. In this work, we examined the ability of these primers to detect Wolbachia in Anastrepha populations from three regions in the State of São Paulo, southeastern Brazil. All of the samples were positive for Wolbachia supergroup A when screened with primers for 16S A rDNA and wsp A; the wsp B primer also gave a positive result, indicating cross-reactivity. The ftsZ primer showed a poor ability to detect Wolbachia in Anastrepha and generated false negatives in 44.9 percent of the samples. These findings indicate that reliable PCR detection of Wolbachia requires the use of primers for 16S rDNA and wsp to avoid cross-reactions and false negatives, and that the ftsZ primer needs to be redesigned to improve its selectivity.

Karyotype relationships among Anastrepha bistrigata, A. striata and A. serpentina (Diptera, Tephritidae)

The species of Anastrepha are arranged into 17 intrageneric groups. Recently, it was proposed that two species of the striata group, Anastrepha striata and A. bistrigata, might be realocated to serpentina group. Anastrepha bistrigata and A. serpentina have an X1X2Y/X1X1 X2X2 sex chromosome system while A. striata has a XY/XX system. It was previously proposed that the karyotype of A. bistrigata could be derived from that of A. striata by an Y:A fusion, and that the karyotype of A. serpentina would be derived from another, hypothetical karyotype. In the present report sequential staining with DAPI and chromomycin A3 (CMA3), followed by C-banding, revealed that the C-banded heterochromatic blocks of the sex chromosomes of A. bistrigata have different affinities to fluorochromes in comparison to the chromosomes of A. striata, from which they have hypothetically derived. The chromosomes of A. serpentina show substantial differences in their cytochemical properties compared to their A. bistrigata and A. striata counterparts. The FISH technique showed that the ribosomal gene sequences are located in DAPI- or DAPI/CMA3-positive heterochromatic blocks of the sex chromosomes, one site on the Y chromosome and one site on the X chromosome (X1 in A. bistrigata and A. serpentina). The data suggest that the karyotype of A. striata and A. bistrigata could be derived from a common ancestral karyotype, while the A. serpentina karyotype probably has a distinct origin.

Asymmetry of frontal bristles and postocular setae in species and hybrids of the Anastrepha fraterculus complex (Diptera, Tephritidae)

Asymmetry of the frontal bristles and postocular setae was studied in samples from natural populations and laboratory colonies of Anastrepha sp. 1 aff. fraterculus, of A. sp. 2 aff. fraterculus, and in F1 hybrids obtained from laboratory reciprocal crosses. Natural populations were sampled in a zone of sympatry and in two geographically distant regions with different climatic conditions. Asymmetry was scored as the differences between the number of bristles and of setae on the right and left sides of the head, males and females analyzed independently. The two traits exhibited variability according to the model of fluctuating asymmetry (FA). No significant differences among samples were found in the FA of frontal bristles. A significant FA was observed for the postocular setae of A. sp. 1 males from a southern population (Vacaria, RS) as compared to the asymmetry exhibited by males and females of some other samples. No significant differences in FA were observed among the interspecific hybrids and the laboratory samples of both parental species. The higher FA found in the males from Vacaria was attributed to climatic conditions prevailing in that region. The absence of a higher FA in hybrids may be related to the relatively recent evolutionary history of the two species.

Occurrence of Wolbachia in Brazilian samples of Ceratitis capitata (Wiedemann) (Diptera: Tephritidae)

Wolbachia bacteria were detected by PCR followed by sequencing of a fragment of the 16S ribosomal gene, in a natural population sample and in two laboratory colonies of the medfly, C. capitata (Wied.), from Brazil. Sequencing revealed that the fragment was compatible with the Wolbachia type A group found in several insect species. This is the first description of a natural Wolbachia infection in C. capitata, since several other samples from different regions so far analyzed seemed to be free of infection.

Bactérias Wolbachia foram detectadas por PCR seguido de seqüenciamento de um segmento do gene ribossomal 16S em uma amostra de população natural e em duas colônias de laboratório de Ceratitis capitata (Wied.) do Brasil. O seqüenciamento do fragmento amplificado mostrou que este é compatível com os de Wolbachia tipo A, encontrada em numerosas espécies de insetos. Esta é a primeira descrição de uma infecção natural desse hospedeiro por Wolbachia desde que, em linhagens de C. capitata provenientes de outras regiões, não foram encontradas essas bactérias.

Karyotype characterization of Anastrepha fruit flies (Diptera: Tephritidae)

The study comprises the description of the C-banded mitotic chromosomes of five species of Anastrepha, A. amita Zucchi, A. turpiniae Stone, A. zenildae Zucchi, A. grandis (Macquart) and A. leptozona Hendel , and a reanalysis of the chromosomes of three other species, A. distincta Greene, A. obliqua (Macquart) and A. sororcula Zucchi. The species have a diploid number of 12, while A. leptozona has 2n = 10. Heteromorphism of the sex chromosomes is present in all of the species, which have a XY/XX sex chromosome system. A. grandis and A. leptozona have meta and submetacentric autosomes while all of the chromosomes in the other species are acrocentric. The species can be characterized unequivocally by the length and the pattern of blocks of heterochromatin in the sex chromosomes.

O estudo apresenta a descrição dos cromossomos mitóticos, através do bandamento-C, de cinco espécies de Anastrepha, A. amita Zucchi, A. turpiniae Stone, A. zenildae Zucchi, A. grandis (Macquart) and A. leptozona Hendel e uma reanálise dos carótipos de três outras espécies, A. distincta Greene, A. obliqua (Macquart) and A. sororcula Zucchi, segundo a mesma técnica. As espécies apresentam número diplóide igual a 12 cromossomos, exceto A. leptozona com 2n = 10. Os cromossomos sexuais são heteromórficos e com sistema XX/XY. Os autossomos são meta ou submetacêntricos em A. grandis and A. leptozona e acrocêntricos nas demais espécies. Os cromossomos sexuais apresentam comprimento e padrão de blocos de heterocromatina característicos para cada espécie.

Evaluation of egg morphology from ethanol preserved females of Anastrepha sp.2 aff. fraterculus (Diptera: Tephritidae) / Avaliação da morfologia dos ovos de fêmeas de Anastrepha sp.2 aff. fraterculus (Diptera: Tephritidae) preservadas em etanol

A morfologia dos ovos de Anastrepha sp.2 aff. fraterculus foi avaliada em amostras de ovaríolos maduros obtidos de fêmeas preservadas em etanol, de fêmeas recém-mortas e comparadacom a de ovos recém-depositados. As características da ornamentação do córion, assim como da estrutura interna dos ovos recém-depositados também puderam ser observadas nos ovos dissecados das fêmeas. Fêmeas preservadas em etanol podem ser utilizadas para análise da morfologia dos ovos, possibilitando a inclusão de espécies, cujos ovos não são obtidos facilmente, nesse tipo de análise.

Features of eggshell morphology in Anastrepha sp.2 aff. fraterculus were evaluated in mature ovariole ova dissected from females preserved in ethanol, and from ovaries of fresh killedfemales in comparison to that of freshly laid eggs. The characteristics of sculpturing as well as of internal structure of the chorion in oviposited eggs could be observed in both samples of mature ovariole ova. Ethanol preserved females may be used as a source of eggs for morphological studies allowing this kind of analysis to species whose eggs are not readily obtained.

Haldane's rule and other aspects of reproductive isolation observed in the Anastrepha fraterculus complex (diptera: tephritidae)

Neste trabalho foram avaliados alguns aspectos do isolamento reprodutivo entre populaçöes alopátricas de duas espécies do complexo Anastrepha fraterculus (A. fraterculus sp. 1 e sp. 2) em condiçöes de laboratório. A maioria dos cruzamentos intraespecíficos, assim como aqueles entre fêmeas da sp. 2 e machos de sp. 1, foram férteis. No cruzamento recíproco, somente 41.7 por cento resultaram em progênies viáveis. A taxa de eclosäo das larvas foi similar para as duas espécies, mas significativamente menor para os cruzamentos intraespecíficos. A proporçäo sexual da progênie adulta foi de 1:1 tanto nos cruzamentos intraespecíficos quanto naqueles de fêmeas de sp. 1 com machos de sp. 2. Entretanto, nos cruzamentos entre fêmeas de sp. 2 com machos de sp. 1 houve um desvio significativo na proporçäo sexual das progênies, com um excesso de fêmeas. Esta observaçäo está de acordo com a regra de Haldane, cuja ocorrência reflete a existência de isolamento reprodutivo pós-zigótico. Os resultados reforçam a sugestäo da existência de dois taxa distintos em A. fraterculus.