Differential characterization of holocentric chromosomes in triatomines (Heteroptera, Triatominae) using different staining techniques and fluorescent in situ hybridization

A comparative study of holocentric chromosomes in the triatomine species Panstrongylus megistus, Rhodnius pallescens and Triatoma infestans was carried out in order to characterize heterochromatin, rDNA active sites and nucleolar proteins. Cytological preparations of seminiferous tubules were stained by silver impregnation, C banding, fluorochromes cma3/da and dapi/da, and fluorescent in situ hybridization (FISH) with Drosophila melanogaster 28S rDNA probe. Our results showed interesting aspects of the organization of chromatin and chromosomes in the meiotic cells of these insects. In R. pallescens, sex chromosomes (X, Y) were distinct from autosomes, when submitted to silver impregnation, C banding, CMA3 staining, and FISH, confirming that these chromosomes bear nucleolar organizer regions (NORs). In P. megistus, two of the three sex chromosomes were CMA3/DAPI-; at early meiotic prophase and at diakinesis, silver impregnation corresponded with FISH signals, indicating that in this species, two chromosomes (probably a sex chromosome and an autosome) bear NORs. In T. infestans, silver nitrate and FISH also stained corresponding areas on meiotic chromosomes. Our data suggest that in triatomines, in general, the number and location of NORs are species-specific. These regions may be considered important chromosome markers for comparative studies to improve the understanding of evolutionary mechanisms in these hematophagous insects.     

Karyomorphology of six taxa in Chrysanthemum sensu lato (Anthemideae) in Egypt and their genetic relationships by Giemsa C‐banding

Abstract Giemsa C‐banding was applied to the chromosome complements of six diploid species belonging to six genera in Chrysanthemum sensu lato (Anthemideae) distributed in Egypt. Four types of C‐banding distribution were observed in the taxa as follows: (i) negative C‐banding in Anacyclus monanthos (L.) Thell.; (ii) all bands in terminal regions in Achillea fragrantissima (Forssk.) Sch. Bip, which showed 32 bands on 18 chromosomes; (iii) all eight bands at centromeric regions on eight chromosomes in Matricaria recutita L.; and (iv) bands at terminal and centromeric regions in Brocchia cinerea Vis. (12 terminal and six centromeric bands on 12 chromosomes), Cotula barbata DC. (four terminal, six centromeric, and eight short arm bands on 16 chromosomes), and Glebionis coronaria (L.) Cass. ex Spach. (eight terminal on the short arms and four large bands in centromeric regions on 12 chromosomes).     

Banding differences between tiger salamander and axolotl chromosomes

The Hoechst 33258 - Giemsa banding patterns were compared on axolotl (Ambystoma mexicanum Shaw) and axolotl - tiger salamander (Ambystoma tigrinum Green) species hybrid prophase chromosomes. Approximately 369 bands per haploid chromosome set were seen in the axolotl and about 344 bands in the tiger salamander. In the haploid set of 14 chromosomes, chromosome 3 has a constant short or q-arm terminal constriction at the location of the nucleolar organizer. Chromosomes 14 Z and W carry the sex determinants, the female being the heterogametic sex (ZW). The banding patterns of chromosomes 1, 6, 11, and 14 Z of the two species are apparently indistinguishable by our banding method. In the axolotl, chromosome 9 has a small long or p-arm terminal deletion. In the tiger salamander, the remaining 10 chromosomes have terminal or internal deletions. No translocations or inversions seem to have occurred since the gene pool separation of the two closely related species.     

牡丹染色体的Ag-NORs和Giemsa C带的研究

本文首次报道了紫斑牡丹和栽培牡丹5个重要品种的Ag-NORs和Giemsa C带带型。所有材料的核型组成均为2n=2x=10=6m+2sm+2st。多数材料具6个Ag-NORs,但在染色体上的分布位点不同。C带的差别主要表现在第1、2、3对染色体上。以上两种核学特征,具有品种的特异性。分带所显示的端(T)带和Ag-NORs的数目及分布基本一致。牡丹染色体的端带,本质上是N带;常规染色所显示的所谓随体,实际上是瑞部核仁组成区(NORs)。此外,对牡丹的细胞学研究技术进行了讨论。     

alpha-glycerophosphate dehydrogenase activity in flight muscles of triatomine bugs Panstrongylus megistus and Triatoma sordida

The alpha-glycerophosphate dehydrogenase (alpha-GPDH) activity in flight muscles of Panstrongylus megistus and Triatoma sordida, vectors of Chagas disease in Brazil, was studied. Both species showed higher enzymatic activities in fliers than in non-fliers insects. T. sordida exhibited a higher proportion of flier insects than P. megistus. A possible role of alpha-GPDH on triatomines flight is discussed.     

C-banding analysis of eight species ofKengyilia (Poaceae: Triticeae)

To characterize chromosomes and the interspecific relationships within the genus Kengyilia, 8 species were used for Giemsa C-banding analysis. Results indicated that the species differed in C-banding patterns. K. gobicola, K. alatavica and K. batalinii had distinct centromeric bands and no banded chromosomes, while K. hirsuta, K. longiglumis, K. melanthera, K. rigidula and K. thoroldiana had more abundant and diagnostic C-bands with interstitial and terminal bands.     

Comparison of the karyotypes ofPsathyrostachys juncea andP. huashanica (Poaceae) studied by banding techniques

The karyotypes ofP. juncea (Elymus junceus) andP. huashanica (both outbreeders) were investigated by Feulgen-staining and by C-, N-, and Agbanding, based on a single plant in cach case. Both species have 2n=2x=14 and large chromosomes, possibly a generic character. The karyotype ofP. juncea has 8 metacentrics and 6 SAT-chromosomes with minute, heterochromatic satellites while that ofP. huashanica has 9 metacentrics and 5 SAT-chromosomes only, 2 of which with small, heterochromatic satellites. The C-banding patterns ofP. juncea chromosomes comprise from one to five, mostly small, bands at distal, and terminal positions, while those ofP. huashanica chromosomes are characterized by large telomeric bands in most arms. Banding patterns and chromosome morphology allow identification of the homologues of the seven chromosome pairs inP. juncea, but of two pairs inP. huashanica only. The patterns of both taxa are polymorphic, supporting that both taxa are outbreeders. The karyotypic characters suggest thatP. juncea is more closely related toP. fragilis than either is toP. huashanica. N-banding stains weakly. Silver nitrate staining demonstrates that nucleolus organizers of both species have different nucleolus forming capacities. The presence of micronucleoli suggests that both species have an extra unidentified chromosome with nucleolus forming capacity.     

Aspects of classification of Hemiptera hemocytes from six triatomine species.

The objective of this work was to characterize, and compare different morphological types of hemocytes of Rhodnius prolixus, Rhodnius robustus, Rhodnius neglectus, Triatoma infestans, Panstrongylus megistus, and Dipetalogaster maximus. This information provides the basis for studying the cellular immune systems of these insects. Seven morphological hemocyte types were identified by phase-contrast microscopy: prohemocytes, plasmatocytes, granular cells, cystocytes, oenocytoids, adipohemocytes and giant cells. All seven types of hemocytes are not present in every species. For example, adipohemocytes and oenocytoids were not observed in P. megistus and P. infestans, and giant cells were rarely found in any of the species studied. The hemocytes of Rhodnius and Dipetalogaster are more similar to each other than those from Triatoma and Panstronglus which in turn closely resemble each other. Emphasis is placed on methodological problems arising in this work which are discussed in detail.     

C- and G-bands of the opossum chromosomes: terminal sequences of DNA replication.

The sex chromosomes of the opossum, Didelphys virginiana, are the only elements that exhibit C-banding. In contrast, the sex chromosomes as well as the autosomes bear specific G-Bands. However, unlike other mammalian species different types of G-banding are observed if the chromosomes are pretreated with trypsin and SSC solution The SSC-pretreated chromosomes show discrete bands only when stained with Giemsa at certain pH values. An asynchronous pattern of terminal DNA replication is observed among the three C-banding regions of the X-chromosome. The inter- and intrapositive G-banding areas of the chromosomes are not always late in DNA replication in comparison to those negatively stained G-banding areas.     

Cytogenetics of Manihot esculenta Crantz (cassava) and eight related species

Thirty-nine cultivars of cassava and eight related wild species of Manihot were analyzed in this work for number, morphology and size of chromosomes, prophase condensation pattern and the structure of the interphase nucleus. In four accessions, the chromosome size was measured and in some others, the number of secondary constrictions, meiotic behavior, C-band pattern, CMA/DAPI bands, nucleoli number and the location of 5S and 18S-5.8S-28S rDNA sites were also observed. All investigated accessions showed a similar karyotype with 2n = 36, small metacentric to submetacentric chromosomes. Two pairs of terminal secondary constrictions were observed in the chromosome complement of each accession except Manihot sp. 1, which presented two proximal secondary constrictions. The prophase chromosome condensation pattern was proximal and the interphase nuclei structure was areticulate to semi-reticulate. The meiosis, investigated in seven cultivars and four wild species, was regular, displaying 18 bivalents. C-banding revealed heterochromatin in 9 or 10 chromosomes. The analysis with fluorochromes frequently showed four chromosome pairs with a single CMA+ terminal or subterminal band and a few other chromosomes with DAPI+ unstable bands. Six 45S rDNA sites were revealed by FISH, which seemed to colocalize with six CMA+ bands. Only one chromosome pair presented a 5S rDNA site. The maximum nucleoli number observed per nucleus was also six. These data suggest that all Manihot species present a very similar chromosome complement.     

C-BANDING KARYOTYPE AND MALE MEIOSIS OF ACROPHYLLA WUELFINGI (REDTENBACHER) (PHASMATODEA: PHASMATIDAE)

Abstract
The C-Banded karyotype and the male meiosis of the Australian species Acrophylla wuelfingi are described and compared with other members of the subfamily Phasmatinae. The chromosomes of A. wuelfingi are rather large and their number is 2n ♂= 45, XO, 2n ♀= 46, XX; the metacentric X being the largest element of the set. Karyotype analysis shows a predominance of heterobrachial chromosomes and centromeric heterochromatin bands or blocks in all elements. Male meiosis is chiasmate, with mainly terminal chiasmata.     

Comparative karyotype analysis ofCeratozamia mexicana andMicrocycas calocoma (Zamiaceae) using fluorochrome banding (CMA/DAPI) and fluorescence in situ hybridization of ribosomal DNA

Orcein staining, differential staining with CMA and DAPI, and FISH with an rDNA probe were used to compare somatic chromosomes ofCeratozamia mexicana andMicrocycas calocoma. CMA-positive dots and hybridization signals appeared on chromosomes at early interphase and mitotic prophase, but in significantly different number in the two species. InCeratozamia mexicana, the CMA-positive and DAPI-negative bands and the hybridization signal were located at the terminal region of the long arm of three median-centromeric chromosomes, the terminal region of the short arm of two median-centromeric chromosomes and the terminal region of the long arm of two subterminal-centromeric chromosomes. InMicrocycas calocoma, they were located at the pericentric region of two median-centromeric chromosomes. These chromosome data suggested thatMicrocycas has no simple Robertsonian relationship toCeratozamia.     

甘肃紫斑牡丹品种与中原牡丹品种银带和Giemsa C带的研究

分别对甘肃紫斑牡丹品种和中原牡丹品种进行了核型,Ａｇｇｋｐｈｔ Ｇｉｅｎｓａ Ｃ带的研究。发现紫斑牡丹品种核型组成为２ ＝１０＝８ｍ＋２ｓｔ;中原牡丹品种核型组成为２ｎ＝１０＝６ｍ＋２ｓｍ＋２ｓｔ。ＧｉｅｍｓａＣ带带型显示,供试品种均能显示染色体端带,但天染色体端带的数目及分布位置上具品种特异性。     

Cytological characterization of heterochromatin and rDNA inPinus radiata andP. taeda

Fluorochrome C-banding ofPinus radiata andP. taeda metaphase chromosomes showed many pericentromeric DAPI bands and interstitial CMA bands inP. radiata, and centromeric and interstitial CMA bands inP. taeda. Giemsa C-band patterns differed between the species with centromeric bands inP. radiata but no consistent bands inP. taeda. A karyotype ofP. radiata was developed based on banding patterns that distinguished all but two of the 12 pairs of chromosomes. In situ hybridization (ISH) using probes for high-copy ribosomal DNA (rDNA) showed 10 pairs of 18S–25S sites and two pairs of 5S sites in both species. Most of the sites were interstitial or centromeric.     

Randomly Amplified Polymorphic DNA (RAPD) and Isoenzyme Analysis of Trypanosoma rangeli Strains

ABSTRACT. Sixteen Trypanosoma rangeli strains were compared by isoenzyme and randomly amplified polymorphic DNA (RAPD) analysis. Eight strains were isolated from either Rhodnius prolixus or Homo sapiens from Honduras, Colombia and Venezuela. Another eight strains were isolated from either Panstrongylus megistus or the rodent Echimys dasythrix from the State of Santa Catarina, southern Brazil. All six T. rangeli strains isolated from P. megistus were co-infections with Trypanosoma cruzi , demonstrating an overlap of the sylvatic cycles of these parasites and that the accurate identification of species is of utmost importance. Both isoenzyme and RAPD analysis revealed two distinct groups of T. rangeli strains, one formed by the strains from Santa Catarina and the other, by the strains from Honduras, Colombia and Venezuela. With the five enzymes used, all the strains from Santa Catarina had identical profiles which overlapped with those of the other regions only in the pattern obtained with malic enzyme. Analysis of 138 RAPD bands by means of an unweighted pair group method analysis (UPGMA) phenogram using the Dice similarity coefficient allowed the separation of the two groups based on their divergence at a lower level of similarity than the phenon line. We show that the identification of T. cruzi and T. rangeli in naturally mixed infections is readily achieved by either RAPD or isoenzyme analysis.     

Comparative cytogenetics between the species Passiflora edulis and Passiflora cacaoensis

Passiflora edulis Sims is the most economically important species of the genus Passiflora. A new species was described recently, Passiflora cacaoensis Bernacci & Souza, which displayed morphologic characteristics very similar to P. edulis. Due to the need for delimitation of the two species, karyomorphological and banding analyses were carried out. Both species have 2n = 18, with the same karyotype formula 16 m + 2sm. There was variation between the species regarding the location of satellites and the width of chromosome pairs 2, 4 and 8. C banding revealed the presence of constitutive heterochromatin in the centromeric and telomeric regions of all chromosomes in both species. However, only in P. cacaoensis did chromosomes 3 and 9 have a large quantity of heterochromatin. Fluorochrome banding revealed CMA⁺ bands only in the satellites, but no DAPI⁺ bands. Fluorescence in situ hybridisation (FISH) showed that in P. cacaoensis the rDNA 5S probe is located in a single site in the subterminal position of the long arm of chromosome 5. However, for the rDNA 45S probe, two sites were detected in terminal positions of the long arms of chromosome 7, with a bigger and stronger signal, and of chromosome 9. According to the asymmetry index and the quantity of heterochromatin, P. cacaoensis is a more basal species than P. edulis. The cytogenetic data indicate that P. cacaoensis is closely related to P. edulis, but is a different species.     

The threat of reintroduction of natural transmission of Chagas' disease in Bambuí, Minas Gerais State, Brazil, due to Panstrongylus megistus.

In the Epidemiological Surveillance Program in the county of Bambuí, Minas Gerais, between August 1986 and December 1988, 154 Panstrongylus megistus were captured by the local population in both peridomicile and intradomicile environments. Fifteen (9.8%) of the P. megistus harboured Trypanosoma cruzi. Precipitin tests showed that the most frequent triatomine blood meal sources were birds, but other sources were dogs, men and cats. The isoenzyme characterization of 13 T. cruzi strains showed that six belonged to zymodeme Z1, corresponding to the wild cycle parasites, and seven belonged to zymodeme Z2, corresponding to parasites isolated from chronic chagasic patients (domestic cycle). As P. megistus were found to be naturally infected by parasites from both cycles. They are clearly able to transmit T. cruzi from the wild cycle to the domestic cycle. Furthermore the capacity of P. megistus in colonizing houses was observed in one residence, vacant for several years, in which 153 triatomines were captured. The data show the possibility of P. megistus reintroducing the natural transmission of Chagas' disease in the county if Epidemiological Surveillance is interrupted.     

Chromatin differentiation between Theobroma cacao L. and T. grandiflorum Schum

A comparative analysis of mitotic chromosomes of Theobroma cacao (cacao) and T. grandiflorum (cupuaçu) was performed aiming to identify cytological differences between the two most important species of this genus. Both species have symmetric karyotypes, with 2n = 20 metacentric chromosomes ranging in size from 2.00 to 1.19 μm (cacao) and from 2.21 to 1.15 μm (cupuaçu). The interphase nuclei of both species were of the arreticulate type, displaying up to 20 chromocentres, which were more regularly shaped in cacao than in cupuaçu. Prophase chromosomes of both species were more condensed in the proximal region, sometimes including the whole short arm. Both species exhibited only one pair of terminal heterochromatic bands, positively stained with chromomycin A ₃ , which co-localized with the single 45S rDNA site. Each karyotype displayed a single 5S rDNA site in the proximal region of another chromosome pair. Heterochromatic bands were also observed on the centromeric/pericentromeric regions of all 20 chromosomes of cacao after C-banding followed by Giemsa or DAPI staining, whereas in cupuaçu they were never detected. These data suggest that the chromosomes of both species have been largely conserved and their pericentromeric chromatin is the only citologically differentiated region.     

Fluorescence analysis of late DNA replication in human metaphase chromosomes.

Thymidine incorporated as a terminal pulse into chromosomes otherwise substituted with 5-bromodeoxyuridine can be detected by associated bright 33258 Hoechst fluorescence. The location of metaphase chromosome regions identified by this method as last to complete DNA synthesis is consistent with the results of autoradiographic analyses with tritiated thymidine. The very late-replicating regions correspond to a subset of those which appear as bands after chromosomes are stained by quinacrine or modified Giemsa techniques. The high resolution of the 33258 Hoechst fluorescence pattern within individual cells is especially useful for revealing variations in the order of terminal replication. Both homolog asynchrony and fluctuations in the distribution of bright 33258 Hoechst fluorescence within chromosomes from different cells are apparent and localized to individual bands. The results are consistent with the possibility that these bands constitute units of chromosome replication as well as structure.