Sand fly transmitted Leishmania species are responsible for severe, wide ranging, visceral and cutaneous leishmaniases. Genetic exchange can occur among natural Leishmania populations and hybrids can now be produced experimentally, with limitations. Feeding Phlebotomus orientalis or Phlebotomus argentipes on two strains of Leishmania donovani yielded hybrid progeny, selected using double drug resistance and fluorescence markers. Fluorescence activated cell sorting of cultured clones derived from these hybrids indicated diploid progeny. Multilocus sequence typing of the clones showed hybridisation and nuclear heterozygosity, although with inheritance of single haplotypes in a kinetoplastid target. Comparative genomics showed diversity of clonal progeny between single chromosomes, and extraordinary heterozygosity across all 36 chromosomes. Diversity between progeny was seen for the HASPB antigen, which has been noted previously as having implications for design of a therapeutic vaccine. Genomic diversity seen among Leishmania strains and hybrid progeny is of great importance in understanding the epidemiology and control of leishmaniasis. As an outcome of this study we strongly recommend that wider biological archives of different Leishmania species from endemic regions should be established and made available for comparative genomics. However, in parallel, performance of genetic crosses and genomic comparisons should give fundamental insight into the specificity, diversity and limitations of candidate diagnostics, vaccines and drugs, for targeted control of leishmaniasis.
- MeSH
- Genomics MeSH
- Crosses, Genetic MeSH
- Leishmania donovani * genetics MeSH
- Leishmaniasis, Cutaneous * MeSH
- Leishmaniasis, Visceral * diagnosis prevention & control epidemiology MeSH
- Phlebotomus * genetics MeSH
- Psychodidae * genetics MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
The classical hypothesis proposes that the lack of recombination on sex chromosomes arises due to selection for linkage between a sex-determining locus and sexually antagonistic loci, primarily facilitated by inversions. However, cessation of recombination on sex chromosomes could be attributed also to neutral processes, connected with other chromosome rearrangements or can reflect sex-specific recombination patterns existing already before sex chromosome differentiation. Three Coleonyx gecko species share a complex X1X1X2X2/X1X2Y system of sex chromosomes evolved via a fusion of the Y chromosome with an autosome. We analyzed synaptonemal complexes and sequenced flow-sorted sex chromosomes to investigate the effect of chromosomal rearrangement on recombination and differentiation of these sex chromosomes. The gecko sex chromosomes evolved from syntenic regions that were also co-opted also for sex chromosomes in other reptiles. We showed that in male geckos, recombination is less prevalent in the proximal regions of chromosomes and is even further drastically reduced around the centromere of the neo-Y chromosome. We highlight that pre-existing recombination patterns and Robertsonian fusions can be responsible for the cessation of recombination on sex chromosomes and that such processes can be largely neutral.
- MeSH
- Y Chromosome genetics MeSH
- Lizards * genetics MeSH
- Sex Chromosomes genetics MeSH
- Cell Movement MeSH
- Recombination, Genetic MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
OBJECTIVE: The aim of this study was to analyse the long-term radiation effects on human sperm. METHODS: In total, 104 samples of male donors from 2 regions of Ukraine were tested. Group 1 consisted of 32 donors from the Ivano-Frankivsk region, group 2 included 72 volunteers from the Zhytomyr region. The average age of donors in both groups was 35 ± 6 years (range 24-49). To assess the level of apoptosis, membrane mitochondrial potential, concentration of reactive oxygen species, and ploidy of sperm, flow cytometry was performed. RESULTS: The individual equivalent dose of group 1 was < 0.4 mSv and of group 2 ≥ 0.4 mSv. Live spermatozoa with signs of apoptosis were significantly higher (p < 0.05) in group 2 in comparison to group 1 (15.6% and 11.2%, respectively). Spermatozoa without violating integrity were 73.2% in group 1 and approximately 16% higher than the indices of group 2. The percentage of dead necrotic spermatozoa was twice as high in men with a predicted equivalent dose of ≥ 0.4 mSv than in comparison group. A higher percentage of spermatozoa with low mitochondrial membrane potential, di- and tetraploid was found in group 2. CONCLUSIONS: An equivalent individual dose of ≥ 0.4 mSv can cause a decrease in mitochondrial potential, an increase in the production of spermatozoa with pathological ploidy, as well as to provoke increasing apoptosis in cells.
- MeSH
- Chernobyl Nuclear Accident * MeSH
- Adult MeSH
- Middle Aged MeSH
- Humans MeSH
- Young Adult MeSH
- Flow Cytometry MeSH
- Reactive Oxygen Species pharmacology MeSH
- Semen MeSH
- Spermatozoa pathology radiation effects MeSH
- Check Tag
- Adult MeSH
- Middle Aged MeSH
- Humans MeSH
- Young Adult MeSH
- Male MeSH
- Publication type
- Journal Article MeSH
The novel wheat powdery mildew and stripe rust resistance genes Pm5V/Yr5V are introgressed from Dasypyrum villosum and fine mapped to a narrowed region in 5VS, and their effects on yield-related traits were characterized. The powdery mildew and stripe rust seriously threaten wheat production worldwide. Dasypyrum villosum (2n = 2x = 14, VV), a relative of wheat, is a valuable resource of resistance genes for wheat improvement. Here, we describe a platform for rapid introgression of the resistance genes from D. villosum into the wheat D genome. A complete set of new wheat-D. villosum V (D) disomic substitution lines and 11 D/V Robertsonian translocation lines are developed and characterized by molecular cytogenetic method. A new T5DL·5V#5S line NAU1908 shows resistance to both powdery mildew and stripe rust, and the resistances associated with 5VS are confirmed to be conferred by seedling resistance gene Pm5V and adult-plant resistance gene Yr5V, respectively. We flow-sort chromosome arm 5VS and sequence it using the Illumina NovaSeq 6000 system that allows us to generate 5VS-specific markers for genetic mapping of Pm5V/Yr5V. Fine mapping shows that Pm5V and Yr5V are closely linked and the location is narrowed to an approximately 0.9 Mb region referencing the sequence of Chinese Spring 5DS. In this region, a NLR gene in scaffold 24,874 of 5VS orthologous to TraesCS5D02G044300 is the most likely candidate gene for Pm5V. Soft- and hard-grained T5DL·5V#5S introgressions confer resistance to both powdery mildew and stripe rust in diverse wheat genetic backgrounds without yield penalty. Meanwhile, significant decrease in plant height and increase in yield were observed in NIL-5DL·5V#5S compared with that in NIL-5DL·5DS. These results indicate that Pm5V/Yr5V lines might have the potential value to facilitate wheat breeding for disease resistance.
- MeSH
- Basidiomycota * MeSH
- Poaceae genetics MeSH
- Plant Diseases genetics MeSH
- Disease Resistance genetics MeSH
- Triticum * genetics MeSH
- Plant Breeding MeSH
- Publication type
- Journal Article MeSH
The identification of causal genomic loci and their interactions underlying various traits in plants has been greatly aided by progress in understanding the organization of the nuclear genome. This provides clues to the responses of plants to environmental stimuli at the molecular level. Apart from other uses, these insights are needed to fully explore the potential of new breeding techniques that rely on genome editing. However, genome analysis and sequencing is not straightforward in the many agricultural crops and their wild relatives that possess large and complex genomes. Chromosome genomics streamlines this task by dissecting the genome to single chromosomes whose DNA is then used instead of nuclear DNA. This results in a massive and lossless reduction in DNA sample complexity, reduces the time and cost of the experiment, and simplifies data interpretation. Flow cytometric sorting of condensed mitotic chromosomes makes it possible to purify single chromosomes in large quantities, and as the DNA remains intact this process can be coupled successfully with many techniques in molecular biology and genomics. Since the first experiments with flow cytometric sorting in the late 1980s, numerous applications have been developed, and chromosome genomics has been having a significant impact in many areas of research, including the sequencing of complex genomes of important crops and gene cloning. This review discusses these applications, describes their contribution to advancements in plant genome analysis and gene cloning, and outlines future directions.
DivIVA is a protein initially identified as a spatial regulator of cell division in the model organism Bacillus subtilis, but its homologues are present in many other Gram-positive bacteria, including Clostridia species. Besides its role as topological regulator of the Min system during bacterial cell division, DivIVA is involved in chromosome segregation during sporulation, genetic competence, and cell wall synthesis. DivIVA localizes to regions of high membrane curvature, such as the cell poles and cell division site, where it recruits distinct binding partners. Previously, it was suggested that negative curvature sensing is the main mechanism by which DivIVA binds to these specific regions. Here, we show that Clostridioides difficile DivIVA binds preferably to membranes containing negatively charged phospholipids, especially cardiolipin. Strikingly, we observed that upon binding, DivIVA modifies the lipid distribution and induces changes to lipid bilayers containing cardiolipin. Our observations indicate that DivIVA might play a more complex and so far unknown active role during the formation of the cell division septal membrane.
PURPOSE OF THE STUDY The study aimed to evaluate the intraoperative and early postoperative response to simultaneous bilateral femoral osteotomy usually accompanied by soft tissue release of hip joints, or open reduction, capsuloplasty, pelvic osteotomy or extraarticular shelf procedure. MATERIAL AND METHODS A bilateral surgery was performed in 16 children. Twelve children suffered from (spastic) cerebral palsy and there was one case of paralytic dislocation in a patient with myelomeningocele, while the remaining patients suffered from chromosome I aberrations, Dandy-Walker syndrome and merosin-deficient muscular dystrophy. GMFCS Level IV and V prevailed. The patients with femoral head deformity or severe adduction contracture were removed from the study group. In all cases the LCP Pediatric Hip Plate 3.5 or 5.0 (Synthes) was used for osteosynthesis. The postoperative fixation by a hip spica cast was applied for 6 weeks, after which in most cases SWASH orthosis was used at night. The age of the patient, the hip joint finding, the GMFCS level and the type of procedure were recorded. RESULTS The evaluation took into account the use of general anaesthesia only or a combination of general and epidural anaesthesia, most often through caudal block, duration of surgery, time when blood transfusion was necessary and the volumes of blood needed, duration of stay in the Anaesthesiology and Resuscitation Unit, or Intensive Care Unit. As a response to surgery, the changes in haemoglobin levels in g/l and VAS pain score were studied. In four patients only the operative time exceeded 3 hours. Blood transfusion was necessary in 13 patients, with one blood unit being always sufficient. Two patients were admitted to the Anaesthesiology and Resuscitation Unit, the remaining patients spent 1-3 days after surgery in the ICU. The average length of hospital stay did not exceed a week. The postoperative decrease in haemoglobin levels quickly improved. The pain intensity was regularly recorded postoperatively and on day 3-4 it was evaluated as moderate, with patients responding well to common analgesics (VAS 4-7). DISCUSSION The evaluation of duration of simultaneous bilateral procedure, postoperative recovery based on the need for blood transfusion, changes in blood count and VAS scores indicated that the procedure performed on both hip joints simultaneously does not significantly exceed the reasonable limits in terms of the patient s burden. In literature, we found only a single article on a topic of this sort, the conclusions of which are very similar. CONCLUSIONS The simultaneous bilateral femoral osteotomy can be considered a fairly safe procedure. Key words: hip joint instability, simultaneous femoral osteotomy, cerebral palsy.
- MeSH
- Child MeSH
- Femur MeSH
- Hip Joint surgery MeSH
- Humans MeSH
- Hip Dislocation * surgery MeSH
- Cerebral Palsy * MeSH
- Osteotomy MeSH
- Retrospective Studies MeSH
- Feasibility Studies MeSH
- Treatment Outcome MeSH
- Check Tag
- Child MeSH
- Humans MeSH
- Publication type
- Journal Article MeSH
The study investigated the effects of sperm sorting, capacitation treatment and co-cultivation on sexed bovine in vitro embryo production. The effect of treatment and co-culture on production of embryos of the preferred sex from unsorted sperm was also studied. Sperm from five breeding bulls was used for fertilization of mature oocytes as follows: Experiment 1, sorted and unsorted sperm (bulls A-E) treated only with heparin in standard co-cultures; Experiment 2, sorted sperm (bulls A-E) treated with heparin-PHE (penicillamine, hypotaurine, and epinephrine) or heparin-caffeine in drop co-cultures; and Experiment 3, unsorted sperm (bull E) treated with either heparin-PHE or heparin-caffeine in both standard and drop co-cultures. In all bulls, treatment with heparin resulted in significantly (p < .05) reduced cleavage and blastocyst rates from sorted sperm, as compared with those from unsorted sperm. In bulls A, B, D and E, treatment of sorted sperm with heparin-PHE in drops significantly increased the blastocyst rate (p < .05). In unsorted sperm of bull E, heparin-PHE treatment in drops resulted in the XX/XY sex ratio inverse to that obtained by heparin-caffeine treatment in standard co-cultures (32.3%/67.7% and 66.7%/33.3%, respectively). In conclusion, the treatment of sorted sperm with heparin-PHE in modified drop co-cultures can be recommended for production of in vitro sexed embryos. The use of unsorted sperm for production of embryos of the preferred sex by selected capacitation treatment and co-culture can be the method of choice in bulls with low IVF yields from sorted sperm.
- MeSH
- Epinephrine pharmacology MeSH
- Fertilization in Vitro methods veterinary MeSH
- Heparin pharmacology MeSH
- Coculture Techniques methods veterinary MeSH
- Embryo Culture Techniques veterinary MeSH
- Oocytes MeSH
- Penicillamine pharmacology MeSH
- Sex Preselection methods veterinary MeSH
- Cattle MeSH
- Spermatozoa drug effects MeSH
- Taurine analogs & derivatives pharmacology MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Cattle MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
S neplodností, tedy s neschopností otěhotnět v průběhu jednoho roku pravidelného nechráněného pohlavního styku, se potýká cca 15 % párů ve vyspělých zemích. Jelikož se jedná o diagnózu konkrétního páru, a nikoli jedince, je namístě přistupovat k páru jako celku, ne odděleně k ženě či muži. Pro posouzení fertilního potenciálu muže je za zlatý standard již několik desetiletí považován spermiogram, tedy nativní vyšetření ejakulátu. Spermiogram je ve své základní podobě pouze morfologickým, nikoli funkčním vyšetřením, proto nemusí vždy spolehlivě informovat o skutečném fertilním potenciálu daného muže. Z tohoto důvodu jsou do praxe zaváděny nové metody pro zlepšení diagnostiky a následné terapie. Uvádíme možnosti vyšetření ejakulátu průtokovou cytometrií a vliv asymptomatických urogenitálních infekcí na plodnost.
In developed countries, approximately 15% of couples suffer from infertility, i.e. they do not conceive within one year of a regular unprotected sexual intercourse. Since infertility is the only one diagnosis of a couple, and not of an individual, it is essential to examine the couple as the unit. Sperm analysis, i.e. native microscopic evaluation, has been used for decades as a golden standard for male fertile potential assessment. Sperm analysis, in its fundamental form, has been only morphological, and not functional evaluation of ejaculate, thus it might not give us reliable information about actual fertile potential of an individual male. On that account, new methods are being introduced to the clinical practice with a goal to improve diagnostics and subsequent treatment. The article presents these new methods, namely flow cytometry, and the impact of asymptomatic urogenital infections on fertility.
- Keywords
- antispermatické protilátky,
- MeSH
- Semen Analysis * MeSH
- Apoptosis MeSH
- DNA Fragmentation MeSH
- Urinary Tract Infections drug therapy pathology MeSH
- Humans MeSH
- Infertility, Male * diagnosis MeSH
- Flow Cytometry MeSH
- Spermatogenesis MeSH
- Spermatozoa abnormalities pathology MeSH
- Check Tag
- Humans MeSH
- Male MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
- Review MeSH
Flow cytometric analysis and sorting of plant mitotic chromosomes has been mastered by only a few laboratories worldwide. Yet, it has been contributing significantly to progress in plant genetics, including the production of genome assemblies and the cloning of important genes. The dissection of complex genomes by flow sorting into the individual chromosomes that represent small parts of the genome reduces DNA sample complexity and streamlines projects relying on molecular and genomic techniques. Whereas flow cytometric analysis, that is, chromosome classification according to fluorescence and light scatter properties, is an integral part of any chromosome sorting project, it has rarely been used on its own due to lower resolution and sensitivity as compared to other cytogenetic methods. To perform chromosome analysis and sorting, commercially available electrostatic droplet sorters are suitable. However, in order to resolve and purify chromosomes of interest the instrument must offer high resolution of optical signals as well as stability during long runs. The challenge is thus not the instrumentation, but the adequate sample preparation. The sample must be a suspension of intact mitotic metaphase chromosomes and the protocol, which includes the induction of cell cycle synchrony, accumulation of dividing cells at metaphase, and release of undamaged chromosomes, is time consuming and laborious and needs to be performed very carefully. Moreover, in addition to fluorescent staining chromosomal DNA, the protocol may include specific labelling of DNA repeats to facilitate discrimination of particular chromosomes. This review introduces the applications of chromosome sorting in plants, and discusses in detail sample preparation, chromosome analysis and sorting to achieve the highest purity in flow-sorted fractions, and their suitability for downstream applications.
