Characidae is one of the predominant fish groups in the Neotropical region with approximately 30 highly diversified subfamilies, probably not representing a monophyletic group. Three Characidae species from the Iguatemi River Basin (Mundo Novo, MS, Brazil) were analyzed: Astyanax bockmanni with 2n=50 chromosomes (6m+20sm+8st+16a), Bryconamericus stramineus with 2n=52 chromosomes (6m+10sm+16st+20a) and Piabina argentea with 2n=52 chromosomes (6m+24sm+12st+10a), all the species showing no differences between male and female karyotypes. The Ag-NORs were located in only a single chromosome pair in the three species. The chromosomal data obtained may be useful for the taxonomy and phylogenetic studies in these fish groups.
Chromosome numbers were counted in root tip cells of 6 cyperaceous species growing in wetlands in the Mekong Delta, Vietnam. The results were as follows: Cyperus haspan, 2n=26; Eleocharis dulcis, 2n=ca. 212; Eleocharis spiralis, 2n=42; Fimbristylis ferruginea, 2n=20; Fimbristylis polytrichoides, 2n=10; Lepironia articulata, 2n=34. They were reported here for the first time using plant materials from Vietnam. Furthermore, the results of 2 species, Eleocharis spiralis and Lepironia articulata, were the first to be reported for these species. In addition, the dimorphism of Eleocharis dulcis shoots representing emerged and submerged culms was recorded.
High salinity is one of the most serious threats to crop production. The primitive red alga, Cyanidioschyzon merolae, inhabits an extreme environment (42°C, pH 2.5, high salt, metal ion). We have utilized the ability of C. merolae cells to adapt to 0.3 M sodium salt, as well as information from its fully sequenced genome, to produce salt-tolerant transgenic higher plants. To reveal the mechanisms of high salt tolerance, we analyzed, by RT-PCR, genes that were expressed at high levels after salt stress (0.3 M NaCl). The C. merolaeS-adenosylmethionine synthetase (CmSAMS) gene that codes for an enzyme in the polyamine biosynthesis pathway was expressed at high levels (4 to 5 expression ratio). Our results are in accordance with our previously reported DNA microarray data. The CmSAMS gene codes for a 393-aa protein contain 3 conserved domains at the N-terminal and a semi-conserved domain at the C-terminal. The particle bombardment method revealed that the recombinant CmSAMS-green fluorescent protein was localized in the cytoplasm and the nuclei of the plant cells. To further investigate tolerance to salt stress, we produced, by Agrobacterium-mediated transformation, 4 transgenic Arabidopsis thaliana plant lines expressing CmSAMS. Compared to wild-type plants, the CmSAMS transgenic plants were more tolerant to salt stress, clearly defining a role for the CmSAMS gene in conferring salt-stress tolerance.
In oogamous brown algae, mitochondria and mitochondrial DNA (mtDNA) are maternally inherited. In this study, we attempted to clarify the timing of this process through morphological observations and molecular experiments using Undaria pinnatifida (Harvey) Suringar. Immunoelectron microscopy using an anti-DNA antibody indicated that mitochondria have their own mtDNA. The strain-specific single-germling PCR method showed that sperm mtDNA disappeared during the latter part of the one-cell zygote stage; digestion of sperm mitochondria, probably by autophagy, was observed in the one-celled zygote. These data show that in oogamous brown algae, sperm mtDNA is introduced into the cytoplasm of the egg with the mitochondria, and that sperm mitochondria and mtDNA are digested simultaneously, until initiation of the first division of the zygote.
Male meiosis, microsporogenesis and pollen grain studies have been made on 11 accessions of Inula cuspidata C. B. Clarke from the cold desert regions of the Kinnaur district, Himachal Pradesh, India. All the accessions share the same meiotic chromosome number n=10. Out of these, 10 accessions show regular chromosome pairing resulting into normal 10 bivalents formation. The meiotic analysis in these accessions reveals that 2 bivalents are of comparatively large size. These large-sized bivalents in 6 accessions show abnormalities in their disjunctions resulting into laggards/bridges during anaphases/telophases and micronuclei in tetrads and consequently some pollen sterility. In one of the accessions of I. cuspidata scored from Tapri, meiotic course is irregular due to the presence of a high frequency of univalents during the first meiosis. These univalent chromosomes during subsequent stages of meiosis fail to organize at the spindle plate and remain scattered in the cytoplasm. Consequently, the majority of the PMCs in the accession depict irregular anaphases/telophases, formation of multiple nuclei and abnormal sporads (dyads, triads, polyads and tetrads with micronuclei). The accession forms a large number of malformed pollen grains consisting of unstained/sterile and heterogeneous sized stained/fertile pollen grains. The accession showing such a abnormal meiotic behaviour could be attributed to its hybrid nature where the chromosome of the 2 putative parents is left unpaired during earlier stages of prophase I.
Cytological studies have been made of 12 species of tribe Senecioneae (Asteraceae) from north and central India. Of these, 3 species, namely S. grahami Hook. f. (n=10), S. kunthianus Wall ex DC. (n=20) and S. sexatilis Sensu Hook. f. (n=10), are counted for the first time. Additional and/or variable cytotypes are recorded for 2 species, namely S. nudicaulis Buch.-Ham D. Don. (n=5, 10) and S. rufinervis DC. (n=10). Existence of B-chromosomes has been recorded in S. krascheninnikovii Schischk. (n=10+0−2B). Two species, S. vulgaris L. (n=20) and S. krascheninnikovii Schischk. (n=10), are explored for the first time from India. Besides these, cytomorphological variabilities have been observed in S. rufinervis (2n=20, 40) and S. nudicaulis Buch-Ham ex D. Don (2n=10, 20 and 40). Great variation in morphological characteristic have been noticed in different accessions of S. laetus Edgew, however all the accessions are found to exist at 2n=40. Meiotic configurations in the form of multivalents and/or secondary association of bivalents are found in Gynura nepalensis DC., Senecio laetus Edgew, S. kunthianus Wall ex DC., S. krascheninnikovii Schischk. and S. nudicaulis Buch-Ham ex D. Don.
The genus Impatiens Linn. belongs to the family Balsaminaceae and includes mostly wild as well as commonly cultivated ornamental plants. Nearly 91% of Indian species of Impatiens are reported to be endemic. To generate basic information on genetic diversity required for the improvement of germplasm, the present study has been carried out from the different selected parts of Western Himalayas such as Kashmir (J&K) and the Kangra and Sirmaur districts (H.P). During this study, 23 accessions belonging to 9 species of the genus Impatiens have been cytomorphologically observed. The species being cytologically worked out for the first time on a worldwide basis include 2 species as I.laxiflora (n=7, 8) and I. reidii (n=7). Six aneuploid cytotypes have been reported for the first time for the species I. arguta (n=7), I. bicornuta (n=7), I. brachycentra (n=8), I. glandulifera (n=6), I. scabrida (n=6) and I. sulcata (n=8) on a worldwide basis. The meiotic course in most of these accessions has been observed to be normal except for some of the accessions of I. brachycentra, I. glandulifera, I. scabrida and I. sulcata marked with abnormal meiosis. Out of 4 species (6 accessions) marked with cytomixis, in 2 accessions, one for each of I. scabrida and I. sulcata, the percentage of cytomixis has been seen to be relatively higher with production of heterogenous sized fertile pollen grains. The presence of B-chromosomes in I. balsamina is in conformity with the earlier reports for this species. Morphological comparison at intraspecific level has also been made for the different aneuploid cytotypes of I. brachycentra (n=7 and n=8), I. glandulifera (n=9 and n=6), I. laxiflora (n=7 and n=8) and I. scabrida (n=6 and n=7) highlighting the morphological changes coupled with chromosomal changes.
We developed a novel model system for hypersensitive cell death, in which the elicitor induced cell death in suspension-cultured cells with 100% efficiency. The 100% efficient cell death was achieved when tobacco BY-2 cells at stationary-phase were diluted to 1×105 cells/ml with conditioned medium and treated with 1.5 μM of cryptogein, an oomycete-derived proteinaceous elicitor. During the cell-death process, nuclear chromatin fragmentation, activation of several genes associated with hypersensitive response (HR), and induction of several genes related to antimicrobial activity were observed. The results strongly suggest that the cells undergoing HR cell death activated antimicrobial mechanism while executing cell-death program.
Physarum plasmodium living as a slimy mass of protoplast in the dark is fragmented into small multinucleated microplasmodia (mPL) in a liquid medium. When mPL were exposed to several unfavorable environments, they transformed into “spherules” with the cell wall structure. We established a synchronous spherule-induction system for mPL consisting of 3 steps: Step 1, subculture in the 1% medium (1% sucrose and 1% soytone peptone) for 84 h at 25°C; Step 2, preculture in the 2% medium (2% sucrose and 2% soytone peptone) for 72 h at 25°C; and Step 3, dark-starvation treatment in the spherulation medium for 96 h at 25°C. Approximately 100% spherulation was observed within 48 h in Step 3. By fluorescence microscopy, we confirmed for the first time that cellulose was the major component of the cell wall in the Physarum spherule. From the results of experiments using the synchronous spherule-induction system and those of experiments on the inhibitory effect of 2,6-dichlorobenzonitrile (DCB) on cellulose synthesis in mPL in the preculture period, we presumed that the nourished medium in the preculture period was essential for mPL prior to spherulation to attain 100% of spherule. Our in vivo labeling experiments revealed that mPL of multinucleated protoplasts produced cellulose in the preculture period by de novo synthesis. We conclude that the Physarum plasmodia always supply themselves with materials to adapt to unfavorable environmental conditions such as starvation as rapidly as they can, even when they are under excellent conditions.
Mutagen-induced polyploidy can be considered to be a system of increasing genotypic plasticity so that aberrations at the genic and chromosomal level are easily tolerated by the plants. A tetraploid and a mixoploid mutant was induced in Trigonella foenum-graecum by sodium azide. Healthy seeds of Trigonella were treated with different doses of sodium azide. In M2 screening, we found variants at 0.15% treatment for 4 h and at 0.25% treatment for 2 h and they were studied for morphological characters, meiotic behaviour, pollen sterility and stomata frequency. They were confirmed as tetraploid and mixoploid respectively and their surviving M3 progeny was further studied on the same line. Meiotic studies showed different types of abnormalities for both. In mixoploid, the RAR increased from M2 to M3 but in tetraploid plant it decreased in M3 generation. The number of chiasma per chromosome was reduced in M3 generation for both the mutants. A comparison of the different anomalies in 2 generations of the tetraploid and mixoploid helps us to understand the reason for very high pollen sterility and negligible seed set in the tetraploid. Stickiness was increased in both, but the relative increase was less in tetraploid. Stickiness has been considered as a natural method of ameliorating stress and hence may be responsible for better performance of mixoploid. Laggards and spindle disorientation was significantly higher in the tetraploid as compared to the mixoploid.
The phenomenon of cytomixis involving chromatin transfer and spindle abnormalities were recorded for the first time in the diploid (n=6) accession of Plantago lanceolata scored from Parvati Valley, Kullu District, Himachal Pradesh (India). Cytomixis with chromatin transfer involved 2–3 PMCs (pollen mother cells) at different stages of meiosis from early prophase-I to T-II, but the maximum frequency of PMCs involved in cytomixis was recorded during the early prophase stages of meiosis-I (11.03%). The transfer of chromatin material among meiocytes was observed to be both partial and complete resulting into PMCs with hypo- and hyperploid chromosome numbers (2n=14, 15). During cytomixis, nucleolus also transmigrated to neighbouring PMCs and the PMCs with supernumerary nucleoli also resulted. Other meiotic abnormalities associated with cytomixis included chromatin stickiness, out of plate bivalents, laggards, bridges, unoriented chromosomes during A-I/T-I and A-II/T-II, and micronuclei. The effect of chromatin transfer, spindle abnormalities and associated meiotic anomalies on meiotic behaviour, pollen fertility and pollen size has been discussed. The cytomixis in P. lanceolata seems to be a natural phenomenon under genetic control and certainly have played a role in the origin of aneuploids and polyploids.
The coenocytic thallus of Bryopsis gametophytes is composed of a main axis and side branches, both of which have a dome-like growing tip with a clear zone of protoplasm and a subsequent cylindrical, basal cell region. Pinnate thallus is formed by regular development of side branches lateral to the main axis. It has been reported that both cytoplasmic streaming and tip growth in the alga are controlled by the microtubule cytoskeleton. We examined the physiology of the tip-growing region in B. plumosa by observing changes in cytoplasmic streaming during 2 different modes of growing tip formation—the germination of spherical cells regenerated from protoplasm aggregates and the formation of side branches. In the main axes, longitudinal cytoplasmic streaming, indicating chloroplast movement, was evaluated using time-lapse videomicroscopy. In the spherical cells regenerated from protoplast aggregates, chloroplasts showed active movement before and after their germination, but showed just small wriggling motion during germination. Chloroplast motility was unchanged during the early stages of side branch formation, during which a clear zone developed in cell surface protrusion. Thus cytoplasmic streaming appeared not to be involved in the growing tip formation in B. plumosa and hence microtubules may play a role in the growing-tip morphogenesis in this alga via mechanisms not directly related to the cytoplasmic streaming.
The cytoplasmic tail (CT) of the influenza virus hemagglutinin (HA) consists of an amino acid sequence that is highly conserved among various subtypes of the influenza A virus. To study the effect of CT size on HA-mediated membrane fusion, we constructed a cDNA encoding a chimeric protein in which the size of CT of influenza virus HA (from A/Hong Kong/1/68; subtype H3) was enlarged by tagging with enhanced green fluorescent protein (EGFP). The EGFP-tagged HA (HA-EGFP) or wild-type HA was individually expressed in HeLa cells, and cell membrane fusion activities were examined using dual-labeled chicken red blood cells. The red blood cells were labeled with both lipidic and aqueous dyes to verify the fusion activity of the HA-EGFP at different steps of the fusion process: 1) lipid-mixing, 2) content-mixing and 3) fusion pore formation. Our data showed that the binding activity of HA towards red blood cells was not affected by enlargement of the CT of HA with EGFP. However, its lipid-mixing, subsequent content-mixing, and fusion pore formation activities were drastically impaired by the enlargement. Therefore, we conclude that CT size is crucial for progression from the binding step to the lipid-mixing step in the process of HA-mediated membrane fusion.
Fourteen open-pollinated seedling progenies of a high yielding, released variety of turmeric, namely ‘IISR Kedaram’, have been characterized for chromosome number, plant morphology, fresh rhizome yield and rhizome quality, in comparison to the mother. The progenies showed significant variation from the mother variety with respect to chromosome number and most of the other characteristics studied. Chromosome number in the mother plant was 2n=63 while in the progenies it ranged from 2n=78 to 2n=84, the latter being the most frequent. For the morphological characteristics namely plant height, number of tillers, leaf length, leaf breadth, petiole length and internode length of primary rhizomes, several progenies excelled the mother. Five progenies produced a significantly higher fresh rhizome yield than the mother variety. Among the quality parameters studied, curcumin and oleoresin content was found to be lower in all the progenies while oil content was found to be slightly higher in 2 progenies compared to the mother. The role of triploid segregation in generating the variation among seedling progenies and the usefulness of the variation in the improvement of turmeric is discussed.
Muskmelon is one of the important vegetable crops in Cucurbitaceae. Karyotypic analysis and flow cytometric (FCM) measurements of DNA C-values of 20 Iranian endemic muskmelon populations are being reported for the first time. Root tips were examined for karyological studies and fresh young leaves of the muskmelon samples and the standard reference (Parsley, Petroselinum crispum, 2C DNA=4.45 pg) stained with propidium iodide (PI) for FCM measurements. All populations examined were diploids (2n=2x=24), having a mean chromosome length (TL) of 1.36 μm (1.15–1.60 μm), volume (TCV) of 1.03 μm3 (0.73–1.63 μm3) and chromatin length (X) of 32.77 μm (27.56–38.43 μm). FCM measurements showed the mean nuclear DNA content (2C-value) of 0.64 pg (0.53–0.71 pg) and monoploid genome size of 313.4 Mbp (262–350 Mbp). Significant positive linear regression was found between 2C DNA amount and latitude.
The genus Eigenmannia belongs to the family Sternopygidae, a monophyletic group of Neotropical electrogenic fish. Eigenmannia trilineata from Iguatemi River basin were analyzed using classical cytogenetic techniques. Chromosomes of this species of Eigenmannia presented a X1X1X2X2 : X1X2Y sex chromosome system, with 2n=31chromosomes for the males and 2n=32 for the females, where the Y is metacentric chromosome originated from Robertsonian translocation. The NORs are located interstitially on the long arm of chromosome pair 10. Constitutive heterochromatin is distributed in very conspicuous blocks at the juxtacentromeric regions acrocentrics chromosomes in the males and females, including the unique large metacentric neo-Y in the males also presented a juxtacentromeric block. The chromosomal data obtained may be useful for the taxonomy and phylogenetic studies in this fish group.
Chromosome features and karyotypes of male meiosis of 2 species of pill millipedes belonging to the genus Arthrosphaera (Arthrosphaera dalyi and Arthrosphaera sp.) endemic to the Western Ghats revealed interesting facts. The Arthrosphaera dalyi has diploid chromosome number (2n) 30 with relative lengths ranging from 39–195. It showed 3–4 distinct subtelocentric chromosome pairs with discernible short arms and the rest are acrocentrics and/or telocentrics. In Arthrosphaera sp. the diploid chromosome number (2n) is 26 and all chromosomes are telocentrics with relative lengths ranging from 36–178. The first pair of chromosome is the largest of all the chromosomes in both the species and is homomorphic in Arthrosphaera sp. and hetromorphic in A. dalyi, wherein one of the chromosomes is the largest and subtelocentric with a clear short arm and the other homologue is a telocentric chromosome. The sex determination mechanism is XX/XY type. A synopsis of the diplopod chromosomes is presented, compared and the possible evolutionary significance have been discussed.
The species of E. spinidens, E. marschalliana, E. bungei, E. buhsei and E. szovitsii showed 2n=2x=20 while E. orientalis had 2n=2x=18 and E. helioscopia showed 2n=4x=42. The chromosome numbers obtained for E. szovitsii and E. helioscopia support the earlier reports, while the chromosome number of E. spinidens, E. marschalliana, E. bungei and E. buhsei are new to science. ANOVA and LSD tests revealed a significant difference for the size of the chromosomes, indicating the role of quantitative genomic changes in the Euphorbia species diversification. Euphorbia species differ in their karyotype formulae indicating the occurrence of structural changes in their chromosomes supported by meiotic chromosome pairing as quadrivalents were formed in diploid species due to heterozygote translocations. The occurrence of unreduced pollen grains was observed in the species showing tripolar cell formation.