The mechanism of adventive embryogenesis was studied at ultrastructural level. Three Citrus cultivars were used as material, two poly-embryonic and one mono-embryonic. The adventive embryos in Citrus are initiated in the nucellar tissue adjacent to the embryo sac in the micropylar half and incidentally from the chalazal end. The nucellar embryoids derive from initial cells. Initial cells are present already at the time of flowering in the poly-embryonic cultivars: in the mono-embryonic cultivar initial cells are absent. At the mature embryo sac stage the initial cells have a « meristematic » character. Before division they develop new cell walls and become « zygote-resembling » cells. During this development they are isolated and their original cell walls are disintegrated. Simultaneously the surrounding nucellar cells degenerate and disappear and the initial cells become embedded in the endosperm.
Karyotypes of three barbels belonging to the group of «small» African species of Barbus sensu lato, B. bigornei, B. ablabes and B. macrops from the Republic of Guinea (Western Africa), were investigated. Diploid chromosome (2n) and chromosome arm (NF) numbers were for B. bigornei 2n = 48 and NF = 96, for B. ablabes 2n= 50 and NF = 98, and for B. macrops 2n = 50 and NF = 92, respectively. The first pair of metacentric chromosomes in all karyotypes was remarkably larger, and it can be considered as a «marker» element for these 3 species. The karyotype characteristics of Barbus species under study demonstrate that they belong to the diploid group of African barbels and they are, in fact, not related to the genus Barbus sensu stricto which is of a distinct evolutionary polyploid origin. Karyology of this poorly studied African cyprinid group is reviewed and discussed.
In contrast to the enormous variability of Neotropical fish species, genetic information about many groups is not yet available. Chromosomal studies have greatly contributed to the characterization of several species, providing important data on these animals. We performed the first cytogenetic characterization of the Amazonian fish species Brachyplatystoma filamentosum (Pimelodidae), popularly known as “piraíba”. The results revealed a diploid number of 2n = 56 composed of 24 metacentric, 12 submetacentric, and 10 acrocentric chromosomes in both females and males. After silver nitrate treatment (AgNORs) it was possible to visualize the nucleolus organizer region located in the terminal portion of the short arms of subtelocentric chromosome pair 22, presenting size polymorphism. Hybridization with 18S and 5S rDNA probes confirmed the number and location of 18S marks rDNA in pair 22 and differences in the size of signals among homologs. The 5S rDNA genes were localized near the centromere on the short arms of chromosome pair 19. Constitutive heterochromatin (C-bands) were localized predominantly in the terminal regions of the chromosomes, and also occurred in some interstitial and centromeric positions. The chromosomal genetic data obtained in this study contribute to the biological characterization of B. filamentosum, which has economic and ecological importance as the largest freshwater catfish occurring in many rivers of the Amazon and Tocantins-Araguaia basins in Brazil. These results may also be used in to infer relationships among Pimelodidae species.
The somatic chromosome numbers and detailed morphometric properties of eight of the nine annual Turkish Carduus L. taxa were examined. The somatic chromosome numbers were 2n = 2x = 22 in C. rechingerianus Kazmi; 2n = 2x = 28 in C. argentatus L., C. acicularis Bertol. and C. pycnocephalus L. subsp. albidus (M. Bieb.) Kazmi; 2n = 2x = 34 in C. pycnocephalus subsp. arabicus (Jacq. ex Murray) Nyman and subsp. breviphyllarius P. H. Davis; 2n = 2x = 50 in C. nervosus K. Koch.; and 2n = 2x = 62 in C. pycnocephalus subsp. pycnocephalus. Karyotype analysis indicated that chromosomes of the Carduus taxa generally have centromere median region (m), submedian region (sm), and rarely median point (M) of the chromosome. A pair of satellite chromosomes (sat-chromosome) was observed in C. pycnocephalus subsp. breviphyllarius. The findings for each of the analysed taxa are compared with the results of previous studies. The chromosome number and morphology of C. nervosus, C. rechingerianus, C. argentatus, C. pycnocephalus subsp. arabicus and subsp. breviphyllarius are newly reported here.
Karyotypes and meiosis of five spider species belonging to the families Gnaphosidae, Miturgidae and Philodromidae were studied using standard Giemsa staining. The male diploid chromosome numbers (2n) and the sex chromosome systems were as follows: Drassodes lutescens: 2n = 21 (X0), Micaria albovittata: 2n = 22 (X1X20), Cheiracanthium mildei: 2n = 26 (X1X20), Cheiracanthium pennyi: 2n = 26 (X1X20) and Philodromus lividus: 2n = 28 (X1X20). Chromosomes of all specimens were telocentric. During the first meiotic division, 10, 10, 12, 12 and 13 autosomal bivalents, respectively, and two sex chromosomes were obtained, with the exception of Drassodes lutescens, which had a different type of sex chromosome system (♂X0/♀XX). During the second meiotic division, two types of nuclei were determined, with or without the sex chromosomes. These karyological data are useful for a better understanding of the chromosomal evolution of araneomorph spiders.
The Callichthyinae subfamily is composed of five genera with a small number of described species. Molecular cytogenetics studies show the scarcity of records for this subfamily. The aim of the present study was to employ cytogenetic parameters in the analysis of three species belonging to the subfamily Callichthyinae sampled from the Paraná River (Brazil). Callichthys callichthys had 2n = 56 chromosomes (26 m-sm + 30 st-a); Lepthoplosternum pectorale had 2n = 64 chromosomes (8 m-sm + 56 st-a); and Hoplosternum littorale had 2n = 60 chromosomes (8 m-sm + 52 st-a). Regarding the location of the nucleolar organizer regions (NORs) (Ag-staining and 18S rDNA-FISH), different situations were found: single NORs in H. littorale and C. callichthys; multiple NORs in L. pectorale, with intra-individual and inter-individual variation. Heterochromatin was observed in the centromeric region in the chromosomes of the three species. Equilocal, interstitial bands were also found in H. littorale. Physical mapping of 5S ribosomal genes by FISH shows eight 5S rDNA sites in C. callichthys; four 5S rDNA sites in H. littorale and six 5S rDNA sites in L. pectorale. Using cytogenetic markers (diploid number, chromosome formula, NORs, heterochromatin distribution pattern and 5S and 18S rDNA sites), the chromosomal evolution in Callichthyinae is presented: callichthys shows mostly basal chromosomal conditions, with mostly derived chromosomal conditions verified for the Hoplosternum–Dianema clade and Megalechis–Lepthoplosternum clade.
Asparagus officinalis L. is a dioecious plant. A region called the M-locus located on a pair of homomorphic L5 chromosomes controls sexual dimorphism in asparagus. This study aims to conduct microdissection and DOP-PCR on the L5 chromosome, and then physically locate the DOP-PCR products of the L5 single chromosome on the metaphase chromosome of asparagus by chromosome painting. The micromanipulator results indicated that the L5 chromosome was successfully microdissected and its DNA was amplified by DOP-PCR. The molecular weight of its PCR products ranged from 250 to 3000 bp according to 1% agarose gel electrophoresis. The second round DOP-PCR products of the L5 chromosome were labeled with Alexa Fluor-488 by using the nick translation method and the metaphase chromosomes of male asparagus were painted, showing that the fluorescence signals were distributed on the centromeres and secondary constriction areas of all chromosomes. This study has provided a basis for the construction of L5 chromosome DNA library, which will facilitate specific probe screening, molecular mapping, gene cloning, and DNA sequencing for this chromosome.
Cytogenetic studies were carried out in three species from genus Astyanax: Astyanax paranae (Tagaçaba Stream and Tauá Stream), Astyanax altiparanae (Iguaçu River and Maringá Stream) and Astyanax fasciatus (Maringá Stream). Both populations of A. paranae featured 2n = 50 chromosomes, although with different karyotype formulae and Ag-nucleolar organizer regions systems. Using the FISH technique in the Tagaçaba population, it was possible to detect rDNA genes in 14 chromosomes and bitelomeric marking in one chromosome. The constitutive heterochromatin pattern also showed differences between the two populations. Moreover, the Tagaçaba population showed macrochromosome B in 50% of analyzed females. Both A. altiparanae populations featured 2n = 50 chromosomes, but also differed in karyotype formula. Both populations featured multiple-NOR systems and quite similar constitutive heterochromatin patterns. For the A. fasciatus population from Maringá Stream, the diploid number was 2n = 46 chromosomes with karyotype formulae 14m, 10sm, 12st and 10a (fundamental number = 82), with single-NOR system and heterochromatin pattern showing evident markers in the telomeric regions of several subtelocentric and acrocentric chromosomes. The data presented for this species differ from some previously analyzed populations with regard to diploid number (2n = 48 chromosomes) and karyotype formula for those with the same diploid number. The present data strengthen the accentuated cytogenetical variability of the genus Astyanax.
This study presents results of histopathologic, cytogenetics and flow cytometry analyses performed on Macoma balthica collected from the Gulf of Gdansk (Baltic Sea) in 2003 in order to compare the techniques for diagnosis of neoplasia. The proportion of affected clams gave a crude prevalence of 15.7 %. The four stages of the disease defined by histology and three stages of neoplasia defined by flow cytometry were reported. Stage I defined by flow cytometry corresponded to stages I and II defined by histology. Chromosome analysis did not lead to a staging of neoplastic progression. Both cytogenetics and flow cytometry indicated a difference in the DNA content of non-neoplastic and neoplastic cells. Cytogenetics examination marked that the range of chromosome numbers scored in abnormal mitosis corresponded to pentaploid-like cells (2.37 x diploid) and was similar to the mean DNA quantity identified using flow cytometry (2.36 x diploid). These methods generally have lower diagnosis sensitivity because with both techniques only a part of an animal can be studied. Thus, histology examination appeared to be the most sensitive tool for detection of the possible foci of neoplastic cells, metastasis and rare tumour cells freely circulating in the hemolymph in the early stages of the disease. Cytogenetic analysis has been considered as an important tool for the evaluation of aquatic environment quality as well as for the ecological risk assessment. Flow cytometry provided a rapid and easy method for discrimination of the aneuploid cells within thousands of cells per individual. Thus, in diagnosis of early stages of the cancer as well as early metastasis histology analyses should be performed. Chromosomes analysis and flow cytometry examination are important techniques for detection abnormalities in cell division, cell viability and DNA quantity. They appear to be very important in diagnosis of tumors based on high aneuploidy level.
Chromosomes of the short-tailed shrew Blarina brevicauda, which display the numerical polymorphism arisen from Robertsonian rearrangements, were analyzed with conventional and silver staining and G- and C-banding techniques. With respect to all specimens examined in the present study, the diploid chromosome number (2n) and fundamental autosomal arm number (FN) were 50 and 48, respectively. The karyotype consisted of 24 pairs of acrocentric autosomes, a large-sized metacentric X chromosome, and a small-sized submetacentric Y chromosome. The comparison with previous findings suggested the geographic polymorphism of Y chromosome in this species. All autosomes and the X chromosome carried slight centromeric constitutive heterochromatin, whereas the Y chromosome was entirely heterochromatic. On the satellites of short arms of two autosomal pairs, the nucleolus organizer regions (NORs) were recognized. The G- and C-banded and Ag-NOR-stained karyotypes presented in the present study could be useful cytogenetic characteristics for specification of chromosomes participating in Robertsonian rearrangements within this species and for karyo-systematic study of genus Blarina.
C-banding patterns were analyzed in six taxa of Paris: P. polyphylla var. chinensis, P. marmorata, P. luquanensis, P. thibetica, P. polyphylla var. yunnanensis and P. polyphylla var. alba. All plants had a diploid chromosome number of 10. The basic banding consisted of three metacentric and two teloblastic chromosomes. All taxa showed interspecific variation in karyotype structure. P. thibetica had a unique C-banding pattern, having six bands on the long arm of the middle median centromeric chromosomes. P. polyphylla var. yunnanensis had mostly faint C-bands. Chromosome morphology and C-banding patterns have been used for chromosome identification and population divergence detection.
Common rust of the coffee-shrub (Coffea sp.): study of the fungus « Hemileia vastatrix » Berk, et Br. in leaf-tissues. After the germ filaments of uredospores of Hemileia vastatrix penetrated by way of stomata, we have studied the development of the intercellular mycelium of this fungus, its localization in the mesophyll, its branches and its walled and dicaryotic structure. We have described the relations between hyphae and leaf-cells of the coffee-shrub such as rounding and close contact between the walls of both organisms which is sometimes maintained over long distances, emission of short and pointed oranches which go through the wall and formation of « internal appressorium ». We have studied the first developmental stages of a haustorium, while stressing the modifications suffered by the walls of both organisms and we have observed the differentiation of the protective coverings of the maturing haustorium such as the collar, the encapsulation, the extra-haustorial membrane and the sheath. We have studied the distribution of the haustoria in the mesophyll, their localization in relation to the host cell and their mother cell. We have observed the differentiation of intercellular hyphae into fruiting hyphae producing uredospores and their clustered discharge through stomata.
The fine structure of the haustorium of the pycnio-aecial stage of Melampsora pinitorqua in infected cells of cotyledons of Pinus pinea was studied. Some features were found to be different from those of the uredial-telial stage of the same rust on Populus tremula. The mother cell wall lacked thichening at the penetration site; the penetration site was found to be very wide; the mother cell wall was continuous with the haustorial wall and the fungal wall was clearly distinct from the host wall; the haustorium had a hypha-like shape and a septum in its proximal region. The interface between the haustorium and the host cell, and the relationship of some host organelles with the haustorium appeared similar to those of the uredial-telial stage of Melampsora pinitorqua on aspen.
Karyotypes of Sorex caecutiens from Cheju Island of Korea were examined with conventional staining and G-banding by trypsin treatment stained with Giemsa (GTG). The diploid and fundamental autosomal arm numbers were 42 and 66, respectively. The autosomal complement in the karyotype comprised six pairs of metacentrics, seven pairs of submetacentrics or subtelocentrics, and seven pairs of acrocentrics. The X was a largest acrocentric chromosome, and the Y was a small subtelocentric chromosome. Chromosomal constitutions and G-banding pattern of S. caecutiens from Cheju Island were essentially identical to those reported for S. shinto from Honshu Island, Japan. This similarity may indicate the ancestral character of the monophyletic S. caecutiens/shinto group.
Karyomorphological analysis of four species of Schoenoplectus (Cyperaceae) from north-central Mexico were carried out. Chromosome numbers ranged from 2n = 38 to 2n = 84. New records of counting are given for Schoenoplectus acutus var. occidentalis (2n = 38 and 2n = 84) and S. americanus (2n = 66). Intra-individual variation in chromosome number is reported for the first time for S. acutus, with a rare polyploid mixoploidy with a prevalence of cells with 2n = 38 (36 small + 2 compound, larger chromosomes) and a few cells with 2n = 84 small, dot-shaped chromosomes, this being the first record of polyploid mixoploidy for Cyperaceae. Mean length of the diploid set ranged from 51.5 μm (S. tabernaemontani) to 79.5 μm (S. acutus). The lowest average chromosome length for the dot-shaped chromosomes was 0.69 μm (S. acutus) and the highest 1.62 μm (S. tabernaemontani); the pair of large chromosomes in S. acutus reached 3.17 μm. A low interchromosomal asymmetry index (A2), 0.11 to 0.14 was found, very similar among all the species except for S. acutus (A2 = 0.30). Absence of primary constrictions was confirmed. The most common mechanism of karyotype variation in the studied species is dysploidy, followed by polyploidy. A comparison of chromosome numbers between Schoenoplectus and the recently segregated Schoenoplectiella based on the literature reveals that Schoenoplectus has higher numbers (n = 18 to 64; 2n = 36 to 84) than Schoenoplectiella (n = 5 to 44; 2n = 18 to 76) as well as a higher prevalence of disploids.
The chromosome morphology of four populations of Dugesia gonoce-phala s.l. from various European localities is described. Three populations (S1, V26 and V35) showed diploid complements of 16 metacentric chromosomes, whereas one population (V28) has a triploid complement of 24 meta- and submetacentric chromosomes. The karyotypes of the populations show small variation of chromosome length and centromere position, which do not enable the identification of individual chromosomes. For a statistical approach of this problem, the 95% confidence limits of relative length and centromeric index were calculated for each chromosome and set out in two dimensional scatter diagram. It is argued that in cases where chromosomes decrease gradually in relative length and show similar centromeric indices, matching of chromosomes in pairs is statistically unsound. Chromosome differentiation techniques were introduced to obtain characteristic banding patterns for chromosome recognition. Tentative results obtained with the BSG-technique showed clear C-bands at the telomeres and centromere regions of some chromosomes. Silver (Ag-As) staining demonstrated the presence of Ag-NORs in two chromosomes of one of the populations. Quinacrine staining of squashed blastemas revealed rather uniform fluorescence intensity at all chromosomes.
Somatic chromosome numbers and karyotype of Hippocrepis unisiliquosa ssp. unisiliquosa L. and Hippocrepis ciliata L. (Leguminosae) collected from Turkey were examined. Chromosome numbers were determined as 2n = 2x = 14 in Hippocrepis unisiliquosa ssp. unisiliquosa L. and Hippocrepis ciliata. The mitotic chromosome number and karyotype of the taxa were reported here for the first time.
The generative cell divides into two irregular shaped sperm cells, with a sharp angle between them. A thin space separates the membranes of the two sperm cells from the membrane of the vegetative cell. This space is interrupted by cytoplasmic channels. The sperm cell cytoplasm contains small mitochondria with evident cristae, Golgi bodies with few plated cisternae, many free ribosomes, some vesicles and some small vacuoles. The nuclei at mature stage are multilobate and have many pores. Plastids are not present. At the sides of the pores the intine consists of an electrondense layer of compact fibrils. In the vegetative cytoplasm two types of RER are present. Around the vegetative nucleus and the sperm cells the cisternae are long. Short and sacculated cisternae surround the plastids. In the early stages of development also short RER cisternae surround the drops of lipid. At anthesis lipids are absent.
Endopolyploidy is a common feature in seed plants. This phenomenon occurs during early development. Flow cytometry of 54 individuals of the species Trifolium pratense variety Manuela, in the vegetative organs during four ontogenetic stages is described. Fresh plant material grown from seeds was used for the analysis. The calculation made on the basis of the mean cycle value revealed that the level of endopolyploidy is different in various organs during the ontogeny. The highest endopolyploidy was recorded in the cotyledons of the first stage (1.0) and the lowest in the third leaf of the fourth ontogenetic stage (0.18). It was noticed that the degree of endopolyploidy decreased during the ontogeny in most of the plants and thus the ontogenetically oldest organs had higher endopolyploidy than the younger ones.
Teliospores of Melampsora pinitorqua Rostr. were collected in the formation, overwintering and germination stages and observed by transmission electron microscope. Some information about their general ultrastructure is given, particularly concerning the cell wall, lipid bodies, mitochondria, ER, cytoplasmic vesicles, vacuoles, unidentified membrane formations, nucleus and germ tube wall. Some organelle changes occurring from the maturation to germination stage were observed.RIASSUNTOTeleutoconidi di Melampsora pinitorqua Rostr. su foglie di Populus tremula sono stati raccolti in località Luriano (Monticiano, Siena) nei periodi della loro formazione, svernamento e germinabilità, e osservati al microscopio elettronico.Essi presentano: una parete bistratificata non particolarmente complessa, un nucleo cospicuo con nucleolo ben visibile, un caratteristico accumulo di globuli lipidici, numerosi mitocondri, diversi piccoli vacuoli. Nel periodo di germinabilità si rendono evidenti anche i ribosomi, cisterne di ER e vescicole citoplasmatiche. La parete del tubetto germinativo appare formarsi come uno strato exnovo interno alla parete del teleutoconidio. I teleutoconidi fissati nel periodo di germinabilità rispetto a quelli svernanti mostrano: una minore quantità di globuli lipidici; un aumento del contenuto citoplasmatico in generale accompagnato dall'evidenziazione delle membrane quali il plasmalemma, ER, membrana nucleare, membrana limitante i mitocondri, dall'aumento del numero dei vacuoli e dalla comparsa di vescicole citoplasmatiche.
3-[(1-methyl-4-nitro-1H-imidazol-5-yl)thio]-4-methyl-1,2,4-triazole (MNITMT) is an immunosuppressive agent used to treat autoimmune disorders. The objectives of this study were to determine the genotoxic and cytotoxic effects of MNITMT on bone marrow cells of mice. Various concentrations of MNITMT were tested (2, 5, 10 and 20 mg ml–1). Bone marrow cells were prepared for mitotic and mitodepressive indices and blood smears for micronuclei. Cell viability of cultured bone marrow cells was determined using an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide)] assay and the proliferation index was calculated. Our results show that MNITMT significantly lowered the mitotic index (MI) at the highest two doses (10 and 20 mg ml–1) with 14.3 % and 14.9 % at 24 h, 19.2 % and 19.6 % at 48 h, 15.4 % and 19.8 % at 72 h, respectively as compared to control. The percentage of micronuclei (MN) increased with increasing the MNITMT doses 2, 5, 10 and 20 mg ml–1 (79, 125, 141 and 145% respectively). Administration of 2, 5 and 10 mg ml–1 of MNITMT did not cause any bone marrow toxicity, while approximately 50% reduction in the rate of proliferation was observed using the highest dose, 20 mg ml–1. These results suggest that high doses of MNITMT cause both genotoxic and cytotoxic effects.
Cytological features and karyomorphology of two endemic giant pill-millipedes (Arthrosphaera fumosa and A. magna) of the Western Ghats were studied based on the meiotic cell divisions using Giemsa staining technique. Different meiotic stages of spermatogonial cell divisions were compared and karyotypes were constructed from the metaphase stages. The diploid chromosome number, 2n = 30, in A. fumosa as well as A. magna with XY sex determination system was seen. Most of the meiotic chromosome behaviors were similar between the millipede species. All meiotic chromosomes in A. fumosa belonged to acrocentric type, while in A. magna 27 were acrocentrics, two (3rd and 10th) were sub-metacentrics and one (5th) was metacentric. The relative lengths of autosomes in A. fumosa and A. magna were 21.9–109.3 and 38.8–90.6, respectively. The X chromosome was longer than the Y chromosome in both millipedes. The relative lengths of the X chromosomes of A. fumosa and A. magna were 130.1 and 103.6, while for the Y chromosome they were 113.6 and 92.6, respectively. Based on this study and earlier cytological and karyological studies on Arthrosphaera, the diploid chromosome number has been predicted to be higher than 30. Further assessment is needed of identified and unidentified species distributed in the Western Ghats of India and other geographic locations to follow the chromosomal evolutionary pattern.
Keywords: Arthrosphaera; chromosomes; Diplopoda; karyotype; giant pill-millipedes; spermatogonial meiosis; Western Ghats
In this study, 15 different accessions of Iranian lettuce cultivars and varieties that were collected from different regions were investigated. Starting from the root apical meristem, we measured the number and size of the chromosomes. The karyotypic formula was determined. The basic chromosome number was n = 9 in all accessions, possibly due to ancient origin, also all accessions had morphological dissimilarity, and heteromorphisms between chromosomes and their chromosomal types were metacentric, submetacentric and subtelocentric. Analysis of variance showed significant differences between chromosomal characteristics. The maximum size of the estimated genome (in terms of chromosome length) was that of the Qom accession (62.45 μm). The minimum size was that of the Babol accession (19.94 μm). Symmetrical and asymmetrical karyotypes were observed between the accessions. The formula of the Ahwaz accession was 12m + 4sm + 2t with the maximum centromic index (40.93%), while the Fasa accession had a 4m + 10sm + 4st karyotypic formula and the minimum value (28.73). There were different numbers of satellites present on the chromosomes in the different accessions. Some of them had two satellites, some one and the others none. In principle component analysis two components had more than 82.75% of the data variations. Based on the first and second components the accessions separated into three groups. Cluster analysis of karyotypic traits based on the unweighted paired group method using arithmetic average (UPGMA) separated the accessions in three groups. The results of this study will be useful for accession classification and identification.