The Polytene Chromosomes of Cnesia dissimilis (Edwards) and Three Species of Gigantodax Enderlein (Diptera: Simuliidae) from Lanin National Park (Argentina)

Cytological studies were made on larvae of Gigantodax marginalis, G. chilensis, G. fulvescens and Cnesia dissimilis from four creeks in Lanin National Park, Neuquen province, Argentina. Chromosome maps and idiograms of these species are presented. The following inversions were observed: G. marginalis: IL-1 (X-linked inversion), IL-2 (Y-linked inversion), IIS-1.2, IIL-1, IIIL-4,5; G. chilensis: IL-4 (X-linked inversion), IIS-1.2, IIIL-4,5; G. fulvescens:IL-1 (X-linked inversion), IL-3 (Y-linked inversion), IIS-1.2, IIL-1, IIIL-4,5; C. dissimilis: IL-1, IL-5, IIIL-1. Karyological information was used to construct a cladogram and Cnesia sp. Was found to show close resemblance to the three Gigantodax spp.

Malpighian tubule polytene chromosomes of Culex quinquefasciatus (Diptera, Culicinae)

Dipteran polytene chromosomes provide an excellent model for understanding in species complexes, as well as for structural and functional cytogenetics. The status of species in the Culex pipiens complex is controversial and the use of polytene chromosomes for cytogenetic analysis in the subfamily Culicinae has been difficult because of methodological problems. In this study, Malpighian tubule polytene chromosomes were obtained from young (0 to 12 h, 20ºC) and old (20 to 42 h, 28ºC) laboratory-bred C. pipiens quinquefasciatus pupae. The chromosome maps for this species were constructed and compared with published data for C. pipiens pipiens and C. p. quinquefasciatus. Although the banding patterns were conserved between subspecies, analysis of the structural variations in the bands and interbands revealed differences apparently related to the physiological stage and ecogeographical strain. The organization of the centromeric regions in larval and pupal chromosomes showed greater similarity to each other than did those of pupal and adult chromosomes. The use of pupal polytene chromosomes for in situ hybridization with vector competence probes is discussed.

A technique for preparing polytene chromosomes from Aedes aegypti (Diptera, Culicinae)

Polytene chromosome preparations were obtained from larval, pupal and adult female Malpighian tubules of Aedes aegypti. The Malpighian tubules of the pupae (0-4 h old) from larvae reared at 20ºC provided the best cytogenetic analysis. The interaction of nucleic acids and proteins that influence the spreading of the chromosomes could be reduced with the preparation technique of the sheets submitted to a stronger treatment starting with the hypotony of tissue and successive bathings with acetic acid. A simple technique should facilitate molecular cytogenetics used in the location of resistance and vector competence genes.

Anopheles darlingi polytene chromosomes: revised maps including newly described inversions and evidence for population structure in Manaus

Salivary gland polytene chromosomes of 4th instar Anopheles darlingi Root were examined from multiple locations in the Brazilian Amazon. Minor modifications were made to existing polytene photomaps. These included changes to the breakpoint positions of several previously described paracentric inversions and descriptions of four new paracentric inversions, two on the right arm of chromosome 3 and two on the left arm of chromosome 3 that were found in multiple locations. A total of 18 inversions on the X (n = 1) chromosome, chromosome 2 (n = 7) and 3 (n = 11) were scored for 83 individuals from Manaus, Macapá and Porto Velho municipalities. The frequency of 2Ra inversion karyotypes in Manaus shows significant deficiency of heterozygotes (p < 0.0009). No significant linkage disequilibrium was found between inversions on chromosome 2 and 3. We hypothesize that at least two sympatric subpopulations exist within the An. darlingi population at Manaus based on inversion frequencies.

The polytene chromosomes of the mosquito Anopheles bellator compared with those of Anopheles cruzii

A photographic map was made of polytene chromosomes of ovarian nurse cells of Anopheles bellator females. The chromosomes of this species have complete or partial homology with those of A. cruzii, mainly in the telomeric and centromeric regions. Variability at the single band level was observed as asymmetric bands at seven different positions. One inversion (3Ra) was detected in the 3R arm.

Polytene chromosome map and inversion polymorphism in Drosophila mediopunctata

Drosophila mediopunctata belongs to the tripunctata group, and is one of the commonest Drosophila species collected in some places in Brazil, especially in the winter. A standard map of the polytene chromosomes is presented. The breakpoints of the naturally occurring chromosomal rearrangements are marked on the map. The distribution of breaking points through the chromosomes of D. mediopunctata is apparently non-random. Chromosomes X, II and IV show inversion polymorphisms. Chromosome II is the most polymorphic, with 17 inversions, 8 inversions in the distal region and 9 in the proximal region. Chromosome X has four different gene arrangements, while chromosome IV has only two.

Cytological and isoenzyme analysis of the Bucay and Quevedo cytotypes of the onchocerciasis vector Simulium exiguum (Diptera: Simuliidae) in Ecuador

Four cytotypes of Simulium exiguum occur in Ecuador, where this morphospecies is the primary vector of onchocerciasis. In this paper, we give the first full description of the banding pattern of the larval polytene chromosomes of the Quevedo cytotypes differ from the chromosomal standard sequence (of the Cayapa cytotype) by the fixed inversions IIL-5 and IIL-6. The Quevedo cytotype additionally differs from the standard and Bucay cytotypes by processing a differentiated X chromosome, wich is indicated by the inversion IIS-A. As the degree of reproductive isolation between the Bucay and Quevedo cytotypes has not yet been estabilished, they must be regarded as intraspecific variants of the same species. In fact, isoenzyme characterizations showed that the Bucay and Quevedo cytotypes are differentiated only to the extent expected of incipient species or geographical populations. Moreover, the sibiling species status previously given to the Bucay cytotype needs be reassessed, there being inadequate analysis from areas in Ecuador where Bucay occurs in sympatry with the standard Cayapa cytotype. No isoenzyme electromorphs were discovered that identified all or mostadult females of any one (cytotype-pure) collection.

The Potential for Dispersal of Onchocerciasis in Ecuador in Relation to the Distribution of the Vector Simulium exiguum (Diptera:Simuliidae)

The future dispersal of onchocerciasis in Ecuador is dependent on the distribution of cytotypes of the vector species complex Simulium exiguum. Over the last 14 years, collections of larvae have been made from over 25 rivers, between 80-1600 m altitude, from various sites on both sides of the Andes. Analysis of larval polytene chromosomes was used to determine the distributions of each cytotype. On the western side of the Andes, the Cayapa cytotype (the only cytotype directly incriminated as a vector) has a distribution from Santo Domingo de los Colorados northwards. The Quevedo and Bucay cytotypes occur from Santo Domingo de los Colorados southwards. On the eastern side of the Andes, the Aguarico cytotype occurs in the Rio Aguarico and a new cytotype is present in the tributaries of the Rio Napo. Whether the disease will spread south of Santo Domingo and on the eastern side of the Andes depends on vector capacity of the cytotypes and the dispersal patterns of individuals infected with onchocerciasis. At present the Aguarico, Bucay and Quevedo cytotypes are known to be efficient hosts, but their biting preferences and biting densities have not yet been evaluated

Chromosomal comparisons among and within populations of Simulium (Chirostilbia) pertinax (Diptera, Simuliidae)

Chromosomal studies were carried on six larval populations of Simulium (Chirostilbia) pertinax from different locations in Brazil. Larvae were collected in the states of Paraná, Rio Grande do Sul, Rio de Janeiro and São Paulo. Polytene chromosome map comparisons within and among populations showed no differences in banding pattern, except for some limited polymorphism (secondary NOR and four band polymorphisms). There were no chromosomal variations associated with the resistance or susceptibility of the larvae to temephos. The chromosomal homosequentiality found among the six populations suggests that S. pertinax may be a monomorphic species.

Black flies (Diptera: Simuliidae) of French Guiana: cytotaxonomy and a preliminary list of species

The objectives of the present study were to broaden the survey of simuliid species in French Guiana and to cytologically analyze the species in the Simulium perflavum species group. Twelve species of Simulium were collected from which S. goeldii, S. quadrifidum, S. trombetense, S. near incrustatum, S. metallicum (s.l.) sp1, S. metallicum (s.l.) sp2 and S. ochraceum (s.l.) are reported for the first time for this region. The only species collected in the S. perflavum group was S. rorotaense; 34 larvae of this species were cytologically analyzed, all of which had the standard sequence. S. metallicum (s.l.), S. ochraceum (s.l.), S. guianense (s.l.) and S. oyapockense (s.l.) are involved with transmission of onchocerciasis in Central and South America, however, in French Guiana these species were not found biting humans during the sampling period. With the few collections made during this study, we increase the number of simulid species known in French Guiana from 6 to 13. It is clear that more simuliid species can be expected to be found when more sampling is done, including collections in other ecoregions in French Guiana.

Cytotaxonomy of Simulium cauchense Floch & Abonnenc and Simulium quadrifidum Lutz (Diptera: Simuliidae) in Brazilian Amazonia

Simulium cauchense Floch & Abonnenc and Simulium quadrifidum Lutz are widely distributed in the Amazon region and are morphologically similar at the larval and pupal stages. Chromosomally, these species are readily distinguished by the position of the nucleolar organizer, which is in the short arm of chromosome I in S. cauchense and in the long arm of chromosomes III in S. quadrifidum. They also differ by three fixed inversions. Sex chromosomes are undifferentiated in both species. Chromosomal resolution of the two species allowed us to evaluate four structural features previously used as diagnostic aids at the larval stage. Characters that distinguish larvae of the two species are the number of branches and branching patterns of the dorsal abdominal setae and the dark band on each primary fan. Branching patterns of the gill histoblasts were often diagnostic, with S. quadrifidum exhibiting more proximal branching and S. cauchense more distal branching. Sites where both species occurred sometimes had larvae with one petiole branching proximally and the other distally; in these cases examination of the chromosomes permitted assignment of the specimen to species. Pigmentation patterns of larvae, on the other hand, are highly variable. Color typically is sex linked in both species.

Genetic compatibility between Anopheles lesteri from Korea and Anopheles paraliae from Thailand

To assess differentiation and relationships between Anopheles lesteri and Anopheles paraliae we established three and five iso-female lines of An. lesteri from Korea and An. paraliae from Thailand, respectively. These isolines were used to investigate the genetic relationships between the two taxa by crossing experiments and by comparing DNA sequences of ribosomal DNA second internal transcribed spacer (ITS2) and mitochondrial DNA cytochrome c oxidase subunit I (COI) and subunit II (COII). Results of reciprocal and F1-hybrid crosses between An. lesteri and An. paraliae indicated that they were compatible genetically producing viable progenies and complete synaptic salivary gland polytene chromosomes without inversion loops in all chromosome arms. The pairwise genetic distances of ITS2, COI and COII between these morphological species were 0.040, 0.007-0.017 and 0.008-0.011, respectively. The specific species status of An. paraliae in Thailand and/or other parts of the continent are discussed.

Concanavalin A-reactive nuclear matrix glycoprotein

The binding capacity of concanavalin A (Con A) to condensed euchromatin and heterochromatin was investigated in chicken erythrocyte nuclei (CEN), mouse liver cells, Zea mays mays meristematic cells and Drosophila melanogaster polytene chromosomes after 4 N HCl hydrolysis to determine whether binding was preferentially occurring in bands and heterochromatin. Dry mass (DM) variation was investigated in CEN by interference microscopy. Feulgen and Con A reactions were employed for all materials to correlate the loci of the two reactions. Quantifications and topological verifications were carried out by video image analysis (high performance cytometry). It was observed that 4 N HCl hydrolysis caused an important DM loss in CEN leaving a level corresponding to the average DNA DM content. In this material, Con A binding was restricted to the nuclear envelope, which reinforces the idea of the absence of a nuclear matrix in these cells. The other cell types exhibited a correspondence of Feulgen-positive and Con A-reactive areas. The Con A reaction was highly positive in the condensed chromatin areas and heterochromatin. This fact led us to speculate that Con A-positive proteins may play a role in the chromatin condensation mechanism, endowing this structure with physico-chemical stability towards acid hydrolysis and contributing to its rheological properties.

Bradysia hygida (Diptera, Sciaridae) presents two eukaryotic Elongation Factor 1A gene homologues: partial characterization of the eukaryotic Elongation Factor 1A-F1 gene

Elongation factor 1A is a highly conserved protein that participates in translation. We report the occurrence of two genes homologous to the eukaryotic Elongation Factor 1A in Bradysia hygida and describe the partial cloning and characterization of the B. hygida eukaryotic Elongation Factor 1A-F1 (BheEF1A-F1) gene. The pattern of BheEF1A-F1 expression in the salivary gland at the end of the fourth larval instar was investigated using real-time PCR. The results showed that BheEF1A-F1 expression levels are relatively constant at the time when rapid changes in protein synthesis occur in this tissue. In situ hybridization experiments coupled to Southern blot analyses showed that the BheEF1A-F1 gene is located at position 3d of the A chromosome and a second gene homologous to eEF1A is located at position 6a of the X chromosome. Southern blot analyses showed that both the BheEF1A-F1 gene and the second gene homologous to eEF1A constitute non-amplified genes. The present results contribute to the molecular characterization of a sciarid eEF1A gene.

5-Bromo-2’-deoxyuridine induces visible morphological alteration in the DNA puffs of the anterior salivary gland region of Bradysia hygida (Diptera, Sciaridae)

5-Bromo-2’-deoxyuridine (BrdUrd) has long been known to interfere with cell differentiation. We found that treatment ofBradysia hygida larvae with BrdUrd during DNA puff anlage formation in the polytene chromosomes of the salivary gland S1 region noticeably affects anlage morphology. However, it does not affect subsequent metamorphosis to the adult stage. The chromatin of the chromosomal sites that would normally form DNA puffs remains very compact and DNA puff expansion does not occur with administration of 4 to 8 mM BrdUrd. Injection of BrdUrd at different ages provoked a gradient of compaction of the DNA puff chromatin, leading to the formation of very small to almost normal puffs. By immunodetection, we show that the analogue is preferentially incorporated into the DNA puff anlages. When BrdUrd is injected in a mixture with thymidine, it is not incorporated into the DNA, and normal DNA puffs form. Therefore, incorporation of this analogue into the amplified DNA seems to be the cause of this extreme compaction. Autoradiographic experiments and silver grains counting showed that this treatment decreases the efficiency of RNA synthesis at DNA puff anlages.