The complex polyploid genome architecture of sugarcane.

Sugarcane, the world's most harvested crop by tonnage, has shaped global history, trade and geopolitics, and is currently responsible for 80% of sugar production worldwide1. While traditional sugarcane breeding methods have effectively generated cultivars adapted to new environments and pathogens, sugar yield improvements have recently plateaued2. The cessation of yield gains may be due to limited genetic diversity within breeding populations, long breeding cycles and the complexity of its genome, the latter preventing breeders from taking advantage of the recent explosion of whole-genome sequencing that has benefited many other crops. Thus, modern sugarcane hybrids are the last remaining major crop without a reference-quality genome. Here we take a major step towards advancing sugarcane biotechnology by generating a polyploid reference genome for R570, a typical modern cultivar derived from interspecific hybridization between the domesticated species (Saccharum officinarum) and the wild species (Saccharum spontaneum). In contrast to the existing single haplotype ('monoploid') representation of R570, our 8.7 billion base assembly contains a complete representation of unique DNA sequences across the approximately 12 chromosome copies in this polyploid genome. Using this highly contiguous genome assembly, we filled a previously unsized gap within an R570 physical genetic map to describe the likely causal genes underlying the single-copy Bru1 brown rust resistance locus. This polyploid genome assembly with fine-grain descriptions of genome architecture and molecular targets for biotechnology will help accelerate molecular and transgenic breeding and adaptation of sugarcane to future environmental conditions.

Clinically node-positive bladder cancer: oncological results of induction chemotherapy and consolidative surgery.

Patients with clinically node-positive bladder cancer have a poor prognosis, with many receiving only palliative chemo- therapy. We evaluated oncological results in bladder cancer patients with clinically regional and supraregional lymph- adenopathy treated with induction chemotherapy (IC) and consolidative cystectomy. Twenty-five patients with clinically node-positive bladder cancer (including pelvic and retroperitoneal nodes) were treated with 2-4 cycles of IC followed by consolidative cystectomy between 2010 and 2016. Pathologic complete response (pCR) was defined as no residual tumor in the final specimen (ypT0N0).The 3-year cancer-specific (CSS) and recurrence-free survival (RFS) for the whole cohort were 52% and 39%, respectively. The 3-year RFS differed according to volume of nodal metastases, the rates were 56% for minimal nodal disease (cN1) versus 33% for cN2-3 and 0% for cM1 disease (p<0.001). pCR was seen in 7 (28%) patients; 50% in cN1 versus 13% in cN3-M1. pCR associated with 3-year CSS of 80% versus 45% in patients with persistent disease after IC. In conclusion, a multimodal approach to patients with clinically node-positive bladder cancer, consisting of IC followed by consolidative surgery, may achieve long-term survival in selected patients. Better results may be expected in patients with initially minimal nodal burden and complete pathologic response to chemotherapy. Further studies are warranted to improve patient selection for consolidative surgery, especially with supra-regional metastases.

[Urinary MicroRNAs as Potential Biomarkers of Bladder Cancer]. / Mocové mikroRNA jako potenciální biomarkery karcinomu mocového mechýre.

BACKGROUND: Currently, there are no urinary-based tumour markers with sufficient sensitivity and specificity to replace cystoscopy in the detection of bladder cancer (BCA). Urinary microRNAs are emerging as clinically useful class of biomarkers for early and non-invasive detection of urologic malignancies. PATIENTS AND METHODS: In this study, 155 patients with BCA and 83 healthy controls were enrolled. Expression profiles of urinary miRNAs were obtained using Affymetrix miRNA microarrays and candidate miRNAs further validated in independent cohort using specific TaqMan assays and quantitative real-time polymerase chain reaction method. RESULTS: Whole-genome profiling identified miRNA signature with significantly different concentrations in urine of BCA compared to controls (p < 0.01). In the independent validation phase of the study, three miRNAs were confirmed to have significantly higher levels in urine of patients with BCA in comparison with control groups (p < 0.0001). In addition, we observed significant decrease in two miRNAs (p < 0.01) concentrations in the urinary samples collected 3 months after surgery compared to pre-operative samples. CONCLUSION: We identified and validated miRNAs to have significantly higher concentrations in urine of patients with BCA in comparison with controls. Our data have shown that urinary miRNAs could serve as sensitive and specific biomarkers enabling non-invasive detection of BCA.Key words: urinary microRNAs - biomarkers - bladder cancer The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study. The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical papers. This study was supported by Ministry of Health of the Czech Republic, grant No. 15-31071A. All rights reserved.Submitted: 19. 3. 2018Accepted: 20. 3. 2018.

[Ureterointestinal anastomosis in urinary diversion - current opinion]. / Soucasný pohled na ureterointestinální anastomózu v mocové derivaci.

Uretero-intestinal anastomosis (UIA) is one of the most critical points for the occurrence of severe complications of intestinal urinary diversion. Perfection of the current techniques and development of new UIA methods are motivated by the effort to reduce both early and late post-surgical complications. The necessity of using antireflux UIA, especially with an orthotopic neobladder, is controversial: according to some authors, antireflux UIA of any type generally increases, in comparison with reflux UIA, the risk of obstruction that endangers renal function more than the urine reflux itself. ICUD-EAU International Consultation on Bladder Cancer 2012 does not recommend applying antireflux UIA in orthotopic bladder substitutions. In ileal conduit diversions, most surgeons use only refluxing UIA. Studer and Turner recommend antireflux UIA only in cases where urine diversion can generate great intraluminal pressure and/or where there is a high risk of permanent bacterial colonization. The flap-and-trough technique (FT) of nonrefluxing UIA represents another flap-valve method for implanting normal as well as dilated ureters into various types of urinary diversion, with a low risk of stenotic complications. CONCLUSIONS: A simple refluxing end-to-side UIA to a low-pressure orthotopic reservoir is a procedure of choice with low complication rates. On the other hand, the need for reflux prevention with a conduit, continent cutaneous diversion or anal diversion is well-founded.Key words: ureterointestinal anastomosis.

[Salvage lymph node dissection in radical prostatectomy failure - a case study]. / Záchranná pánevní lymfadenektomie po selhání chirurgické lécby karcinomu prostaty - kazuistika.

Salvage lymph node dissection is a therapeutic option in patients with nodal-only recurrence after radical prostatectomy. Although this operation does not achieve a complete cure in the majority of cases, it can prolong survival and help postpone hormonal therapy in selected patients while being a relatively safe procedure. The authors describe a case study of a radical prostatectomy patient with radiologically proven recurrence of prostate cancer limited to an internal iliac node. He underwent salvage lymph node dissection with a complete biochemical response. Remission lasts 13 months following surgery and the patient is currently without the need for androgen deprivation therapy.Key words: salvage lymph node dissection prostate cancer radical prostatectomy neoplasm recurrence.

The expansion of heterochromatin blocks in rye reflects the co-amplification of tandem repeats and adjacent transposable elements.

BACKGROUND: A prominent and distinctive feature of the rye (Secale cereale) chromosomes is the presence of massive blocks of subtelomeric heterochromatin, the size of which is correlated with the copy number of tandem arrays. The rapidity with which these regions have formed over the period of speciation remains unexplained. RESULTS: Using a BAC library created from the short arm telosome of rye chromosome 1R we uncovered numerous arrays of the pSc200 and pSc250 tandem repeat families which are concentrated in subtelomeric heterochromatin and identified the adjacent DNA sequences. The arrays show significant heterogeneity in monomer organization. 454 reads were used to gain a representation of the expansion of these tandem repeats across the whole rye genome. The presence of multiple, relatively short monomer arrays, coupled with the mainly star-like topology of the monomer phylogenetic trees, was taken as indicative of a rapid expansion of the pSc200 and pSc250 arrays. The evolution of subtelomeric heterochromatin appears to have included a significant contribution of illegitimate recombination. The composition of transposable elements (TEs) within the regions flanking the pSc200 and pSc250 arrays differed markedly from that in the genome a whole. Solo-LTRs were strongly enriched, suggestive of a history of active ectopic exchange. Several DNA motifs were over-represented within the LTR sequences. CONCLUSION: The large blocks of subtelomeric heterochromatin have arisen from the combined activity of TEs and the expansion of the tandem repeats. The expansion was likely based on a highly complex network of recombination mechanisms.

DArT whole genome profiling provides insights on the evolution and taxonomy of edible Banana (Musa spp.).

BACKGROUND AND AIMS: Dessert and cooking bananas are vegetatively propagated crops of great importance for both the subsistence and the livelihood of people in developing countries. A wide diversity of diploid and triploid cultivars including AA, AB, AS, AT, AAA, AAB, ABB, AAS and AAT genomic constitutions exists. Within each of this genome groups, cultivars are classified into subgroups that are reported to correspond to varieties clonally derived from each other after a single sexual event. The number of those founding events at the basis of the diversity of bananas is a matter of debate. METHODS: We analysed a large panel of 575 accessions, 94 wild relatives and 481 cultivated accessions belonging to the section Musa with a set of 498 DArT markers previously developed. KEY RESULTS: DArT appeared successful and accurate to describe Musa diversity and help in the resolution of cultivated banana genome constitution and taxonomy, and highlighted discrepancies in the acknowledged classification of some accessions. This study also argues for at least two centres of domestication corresponding to South-East Asia and New Guinea, respectively. Banana domestication in New Guinea probably followed different schemes that those previously reported where hybridization underpins the emergence of edible banana. In addition, our results suggest that not all wild ancestors of bananas are known, especially in M. acuminata subspecies. We also estimate the extent of the two consecutive bottlenecks in edible bananas by evaluating the number of sexual founding events underlying our sets of edible diploids and triploids, respectively. CONCLUSIONS: The attribution of clone identity to each sample of the sets allowed the detection of subgroups represented by several sets of clones. Although morphological characterization of some of the accessions is needed to correct potentially erroneous classifications, some of the subgroups seem polyclonal.

[Rare discovery of metastasis of renal cell carcinoma in location of struma polynodosa]. / Raritní nález metastázy svetlobunecného karcinomu z renálních bunek v terénu polynodózní strumy.

Renal carcinoma metastasis to the thyroid gland is not particularly rare; the interval since the initial diagnosis in the case presented, that is, nephrectomy, is more of a rarity. We present the case of a 58-year-old man admitted to our hospital with the diagnosis of polynodular retrosternal goitre. Complete thyroidectomy by cervical approach was performed. Histological examination showed multiple metastases of conventional renal carcinoma located within colloid struma polynodosa. Postoperative progress was uneventful, the surgical wound healed per primam. Renal carcinoma metastasing to the thyroid gland is rather unusual and is often found only postoperatively.Key words: renal carcinoma - metastasis - thyroid gland.

Mapping nonrecombining regions in barley using multicolor FISH.

Fluorescence in situ hybridization (FISH) is a widely used method to localize DNA sequences on chromosomes. Out of the many uses, FISH facilitates construction of physical maps by ordering contigs of large-insert DNA clones, typically bacterial artificial chromosome (BAC) and establishing their orientation. This is important in genomic regions with low recombination frequency where genetic maps suffer from poor resolution. While BAC clones can be mapped directly by FISH in plants with small genomes, excess of repetitive DNA hampers this application in species with large genomes. Mapping single-copy sequences such as complementary DNA (cDNA) is an attractive alternative. Unfortunately, localization of single-copy sequences shorter than 10 kb remains a challenging task in plants. Here, we present a highly efficient FISH technique that enables unambiguous localization of single copy genes. We demonstrated its utility by mapping 13 out of 15 full-length cDNAs of variable length (2,127-3,400 bp), which were genetically defined to centromeric and pericentromeric regions of barley chromosome 7H. We showed that a region of 1.2 cM (0.7 %) on genetic map represented more than 40 % of the physical length of the chromosome. Surprisingly, all cDNA probes occasionally revealed hybridization signals on other chromosomes, indicating the presence of partially homologous sequences. We confirmed the order of 10 cDNA clones and suggested a different position for three cDNAs as compared to published genetic order. These results underline the need for alternative approaches such as FISH, which can resolve the order of markers in genomic regions where genetic mapping fails.

[Systematic mediastinal lymphadenectomy in surgery for sarcomatous pulmonary metastases]. / Význam systematické mediastinální lymfadenektomie v chirurgii plicních metastáz sarkomového puvodu.

PURPOSE OF THE STUDY: Systematic lymph node dissection is performed as standard curative resection for non-small cell lung cancer. Its role in lung metastasectomy is unknown. The aim of our study was to find out the frequency of lymph node metastases, the survival of patients with and without lymph node involvement, and to consider if routine lymph node dissection should be recommended. MATERIAL AND METHODS: The study was undertaken at three departments of surgery and included 14 patients undergoing lung metastasectomy between July 2008 and December 2011. In all patients systematic lymph node dissection was also performed. When wedge resection was done, N1 nodes were removed only as part of a local procedure at that anatomical site. Patients with mediastinal lymph node involvement detected by pre-operative CT or PET scans were not included in the study. RESULTS: Lung metastasectomy for metastatic sarcoma was performed in 14 patients. Nine patients had bilateral lung metastases. Solitary lung metastasis was found in only one case. The mean number of removed mediastinal lymph nodes was 14.8 (7-32). Mediastinal lymph node metastases were found in two patients (14.3%). The average disease free interval (DFI) was 79.6 months (median, 25.5 months). Using the Kaplan-Meier method, the 3-year survival rate was 46% (0.46±0.15). The Cox-Mantel test for comparing the survival curves showed, at a 0.05 level of significance, better survival rates for the patients with no metastatic lymph node involvement (p=0.01). DISCUSSION: The frequency of mediastinal lymph node involvement in our study was 14.3% and this was in agreement with the data reported in the literature. The 3-year survival rate was 46% in our patients; the published 5-year survival is 15-50%. A systematic mediastinal lymphadenectomy during lung metastasectomy for metastatic sarcoma has been recommended, but also argued against because of a low incidence of mediastinal lymph node involvement in sarcomatous metastases reported by some authors. We showed that mediastinal lymph node involvement was a negative prognostic factor. Systematic mediastinal lymphadenectomy as a routine procedure provides for a better staging. This is important in association with the development of adjuvant modalities, such as monoclonal antibodies, at present or a gene therapy in the future. CONCLUSIONS: Even in a carefully selected group of patients, incidence of mediastinal lymph node metastases is high. Since no relevant data based on large patient groups are available, we recommend routine nodal dissection for all patients indicated for lung metastasectomy. Drawing a definite conclusion will require analyses of large numbers of data from multi-institutional studies and cooperation with the international database, if possible.

[Repeated resection for recurrent pulmonary metastases]. / Opakované plicní resekce v chirurgii plicních metastáz.

INTRODUCTION: Repeated pulmonary resections are predominantly used in sarcomas, colorectal cancer, Grawitz tumours, and at a young age in general. Patients with the recurrence of pulmonary metastases within a period of six months have a poor prognosis. MATERIAL AND METHODS: During the period from I/1997 to XII/2011, a total of 165 operations were performed in 149 patients. 11patients had multiple pulmonary resections. According to histological origin, sarcomas were found in 6 cases, Grawitz tumours in 2 cases, Schwannoma malignum in 1 case and laryngeal carcinoma in 1 case. There was synovialosarcoma, osteosarcoma, rhabdomyosarcoma, alveolar sarcoma and uterine sarcoma in the group of sarcomas. RESULTS: In our retrospective study, the most commonly used approach was muscle-sparing vertical thoracotomy in 14 patients, posterolateral thoracotomy in 8 patients, VATS in 3 patients and clamshell thoracotomy in 3 patients. We performed 22 extraanatomic resections - of which 3 times VATS, 6 times lobectomy (of which 1 completion pneumonectomy). We observed complications in 6 cases (small air leak in 3 patients, wound infection in 3 patients) in the postoperative period. No rethoracotomy for complications was needed, lethality was 0. The overall 5-year survival that we observed was 45.4%. CONCLUSION: Surgery is an integral part of complex oncological care. What is extremely important is strict selection of patients eligible for surgery for repeated resections and, of course, an experienced team of thoracic surgeons, but also other physicians. When performed by experienced specialists, surgical treatment of lung metastases is a safe and very useful procedure.

[Wound microbial sampling methods in surgical practice, imprint techniques]. / Metody odberu vzorku na kultivacní vysetrení v chirurgické praxi, otiskové metody.

The wound is a damage of tissue. The process of healing is influenced by many systemic and local factors. The most crucial and the most discussed local factor of wound healing is infection. Surgical site infection in the wound is caused by micro-organisms. This information is known for many years, however the conditions leading to an infection occurrence have not been sufficiently described yet. Correct sampling technique, correct storage, transportation, evaluation, and valid interpretation of these data are very important in clinical practice. There are many methods for microbiological sampling, but the best one has not been yet identified and validated. We aim to discuss the problem with the focus on the imprint technique.

An accurate DNA marker assay for stem rust resistance gene Sr2 in wheat.

The stem rust resistance gene Sr2 has provided broad-spectrum protection against stem rust (Puccinia graminis Pers. f. sp. tritici) since its wide spread deployment in wheat from the 1940s. Because Sr2 confers partial resistance which is difficult to select under field conditions, a DNA marker is desirable that accurately predicts Sr2 in diverse wheat germplasm. Using DNA sequence derived from the vicinity of the Sr2 locus, we developed a cleaved amplified polymorphic sequence (CAPS) marker that is associated with the presence or absence of the gene in 115 of 122 (95%) diverse wheat lines. The marker genotype predicted the absence of the gene in 100% of lines which were considered to lack Sr2. Discrepancies were observed in lines that were predicted to carry Sr2 but failed to show the CAPS marker. Given the high level of accuracy observed, the marker provides breeders with a selection tool for one of the most important disease resistance genes of wheat.

Characterization of a new 4BS.7HL wheat-barley translocation line using GISH, FISH, and SSR markers and its effect on the ß-glucan content of wheat.

A spontaneous interspecific Robertsonian translocation was revealed by genomic in situ hybridization (GISH) in the progenies of a monosomic 7H addition line originating from a new wheat 'Asakaze komugi' × barley 'Manas' hybrid. Fluorescence in situ hybridization (FISH) with repetitive DNA sequences (Afa family, pSc119.2, and pTa71) allowed identification of all wheat chromosomes, including wheat chromosome arm 4BS involved in the translocation. FISH using barley telomere- and centromere-specific repetitive DNA probes (HvT01 and (AGGGAG)(n)) confirmed that one of the arms of barley chromosome 7H was involved in the translocation. Simple sequence repeat (SSR) markers specific to the long (L) and short (S) arms of barley chromosome 7H identified the translocated chromosome segment as 7HL. Further analysis of the translocation chromosome clarified the physical position of genetically mapped SSRs within 7H, with a special focus on its centromeric region. The presence of the HvCslF6 gene, responsible for (1,3;1,4)-ß-D-glucan production, was revealed in the centromeric region of 7HL. An increased (1,3;1,4)-ß-D-glucan level was also detected in the translocation line, demonstrating that the HvCslF6 gene is of potential relevance for the manipulation of wheat (1,3;1,4)-ß-D-glucan levels.

[Is pneumonectomy justified in the management of lung metastases?]. / Je oprávnené provedení pneumonektomie v lécbe plicních metastáz?

The authors discuss about questions of possibility pneumonectomy in concept of lung metastasis therapy. They analysed in detail indications, surgery, complications, survival and factors of survival. The authors also introduce case report of one patient with pneumonectomy in your group of patients.

[Spontaneous pneumomediastinum--a case review]. / Spontánní pneumomediastinum--kazuistika.

The authors present in the following case review a patient with spontaneous pneumomediastinum which can be on of the causes of acute chest pain and it is necessary to think about it in the differential diagnosis of this condition.

[Rates of metastatic affections of mediastinal nodes during pulmonary metastasectomy]. / Cetnost metatastického postizení mediastinálních uzlin behem plicní metastasektomie.

OBJECT: The aim is the evaluation of lymph node metastasis rate during pulmonary metastasectomy harvested by technique of systematic mediastinal lymphadenectomy in year 2009-2010. METHODS: We performed systematic mediastinal lymphadenectomy (the same technique as in non-small cell lung cancer) during pulmonary metastasectomy. RESULTS: There was found lymph node metastasis in four cases from 70 patients. The rate of metastatic involved mediastinal lymph node 5.7% harvested during lung metastasectomy is on lower limit of published interval. We expected higher lymph node involvement in lung metastasis of colorectal origin.

Development of chromosome-specific BAC resources for genomics of bread wheat.

The large bread wheat genome (1C approximately 17 Gbp) contains a preponderance of repetitive DNA and the species is polyploid. These characteristics together serve to hamper the molecular analysis of the wheat genome. Its complexity can, however, be reduced by using flow cytometry to isolate individual chromosomes, and these can be exploited to construct chromosome-specific BAC libraries. Such libraries simplify the task of physical map construction, positional cloning and the targeted development of genetic markers. Rapid improvements in the efficiency and cost of DNA sequencing provide an opportunity to contemplate sequencing the wheat genome by preparing sequence-ready physical maps for each chromosome or chromosome arm in turn. The quality of the chromosome-specific libraries depends on their chromosome coverage and the mean insert size. First-generation libraries suffered from a relatively low mean insert size, but improvements to the protocol have generated a second wave of libraries with a significantly increased mean insert size and better chromosome coverage. Each chromosome (arm)-specific library is composed of a manageable number of clones, and so represents a practical tool in the area of wheat genomics.

Physical distribution of homoeologous recombination in individual chromosomes of Festuca pratensis in Lolium multiflorum.

Crossing over-based recombination is a powerful tool for generating new allelic combinations during sexual reproduction. It usually occurs between homologous chromosomes. However, under some conditions, homoeologues may also be capable of crossing over. Whether homologous and homoeologous crossovers are equivalent and governed by the same rules has never been established. Here we report on chromosome distribution of homoeologous crossovers in a unique system of Festuca x Lolium hybrids. Unlike in most other hybrids, in these intergeneric hybrids, homoeologous chromosomes are capable of pairing and crossing over with frequencies approaching that of homologues. At the same time, genome divergence makes cytological detection of chromosome recombination feasible. We analyzed the distribution of homoeologous recombination along individual chromosomes in a complete set of intergeneric single chromosome substitutions from F. pratensis into tetraploid L. multiflorum. Homoeologous recombination sites were not evenly distributed along the chromosomes, being concentrated in intercalary regions of the arms and reduced in proximal and distal regions. Several recombination hotspots and cold spots were found along individual chromosomes and the recombination was not affected by the presence of a secondary constriction. Our results indicate that despite the uneven distribution of homoeologous recombination, introgression of any part of the F. pratensis genome into L. multiflorum is feasible.

Chromosome pairing of individual genomes in tall fescue (Festuca arundinacea Schreb.), its progenitors, and hybrids with Italian ryegrass (Lolium multiflorum Lam.).

A diploid-like pairing system prevents meiotic irregularities and improves the efficiency of gamete production in allopolyploid species. While the nature of the system is known in some polyploid crops including wheat, little is known about the control of chromosome pairing in polyploid fescues (Festuca spp.). In this work we studied chromosome pairing in allohexaploid F. arundinacea, its progenitors F. pratensis and F. glaucescens, and two intergeneric hybrids Lolium multiflorum (2x) x F. arundinacea (6x) and L. multiflorum (4x) x F. glaucescens (4x). The use of genomic in situ hybridization (GISH) permitted the analysis of homoeologous chromosome pairing and recombination of different genomes involved. We detected a diploid-like pairing system in polyploid fescues F. arundinacea and F. glaucescens, the latter being one of the progenitors of F. arundinacea. The pairing control system was absent in the second progenitor F. pratensis. Detailed analysis of intergeneric hybrids confirmed the presumed haploinsufficiency of the fescue system, which resulted in homoeologous pairing between all component genomes. This indicates that introgression of any specific chromosome segment from one genome to another is possible in all genome combinations. Our results not only contribute to the quest to discover the nature of the system controlling chromosome pairing in polyploid fescues, but may also have serious implications for design of hybrid breeding schemes in forage grasses.