Targeted phasing of 2-200 kilobase DNA fragments with a short-read sequencer and a single-tube linked-read library method.

In the human genome, heterozygous sites refer to genomic positions with a different allele or nucleotide variant on the maternal and paternal chromosomes. Resolving these allelic differences by chromosomal copy, also known as phasing, is achievable on a short-read sequencer when using a library preparation method that captures long-range genomic information. TELL-Seq is a library preparation that captures long-range genomic information with the aid of molecular identifiers (barcodes). The same barcode is used to tag the reads derived from the same long DNA fragment within a range of up to 200 kilobases (kb), generating linked-reads. This strategy can be used to phase an entire genome. Here, we introduce a TELL-Seq protocol developed for targeted applications, enabling the phasing of enriched loci of varying sizes, purity levels, and heterozygosity. To validate this protocol, we phased 2-200 kb loci enriched with different methods: CRISPR/Cas9-mediated excision coupled with pulse-field electrophoresis for the longest fragments, CRISPR/Cas9-mediated protection from exonuclease digestion for mid-size fragments, and long PCR for the shortest fragments. All selected loci have known clinical relevance: BRCA1, BRCA2, MLH1, MSH2, MSH6, APC, PMS2, SCN5A-SCN10A, and PKI3CA. Collectively, the analyses show that TELL-Seq can accurately phase 2-200 kb targets using a short-read sequencer.

The effect of carbon nanoparticles vs. immune colloidal gold technique test strips on parathyroid protection in total thyroidectomy: A randomized clinical trial study.

BACKGROUND: The long-term effect of intraoperative usage of carbon nanoparticles (CN) and parathyroid hormone (PTH) test strip using immune colloidal gold technique (ICGT) is unclear. This study aims to compare the effect of intraoperative usage of CN and ICGT test strips on PG function. METHODS: This randomized clinical study involved adult patients who underwent total thyroidectomy. They were randomly allocated into three groups (control, CN, and ICGT group). Clinical data were analyzed. RESULTS: Each group involved 98 patients. Serum calcium and PTH concentrations at 24 h postoperatively (PTH24h) were higher in CN group. The parathyroid function recovered quicker in CN group. Use of CN increased in situ PG preservation and PTH24h. Mediation analysis indicated that 23.05% of the total effect of CN on PTH24h was attributed to PGRIS. CONCLUSION: CN holds promise to improve in situ PG preservation and protect PG vasculature, thereby reducing the incidence of early hypoparathyroidism. The value of ICGT test strips for PG protection is dubious.

Targeted Phasing of 2-200 Kilobase DNA Fragments with a Short-Read Sequencer and a Single-Tube Linked-Read Library Method.

In the human genome, heterozygous sites are genomic positions with different alleles inherited from each parent. On average, there is a heterozygous site every 1-2 kilobases (kb). Resolving whether two alleles in neighboring heterozygous positions are physically linked-that is, phased-is possible with a short-read sequencer if the sequencing library captures long-range information. TELL-Seq is a library preparation method based on millions of barcoded micro-sized beads that enables instrument-free phasing of a whole human genome in a single PCR tube. TELL-Seq incorporates a unique molecular identifier (barcode) to the short reads generated from the same high-molecular-weight (HMW) DNA fragment (known as 'linked-reads'). However, genome-scale TELL-Seq is not cost-effective for applications focusing on a single locus or a few loci. Here, we present an optimized TELL-Seq protocol that enables the cost-effective phasing of enriched loci (targets) of varying sizes, purity levels, and heterozygosity. Targeted TELL-Seq maximizes linked-read efficiency and library yield while minimizing input requirements, fragment collisions on microbeads, and sequencing burden. To validate the targeted protocol, we phased seven 180-200 kb loci enriched by CRISPR/Cas9-mediated excision coupled with pulse-field electrophoresis, four 20 kb loci enriched by CRISPR/Cas9-mediated protection from exonuclease digestion, and six 2-13 kb loci amplified by PCR. The selected targets have clinical and research relevance (BRCA1, BRCA2, MLH1, MSH2, MSH6, APC, PMS2, SCN5A-SCN10A, and PKI3CA). These analyses reveal that targeted TELL-Seq provides a reliable way of phasing allelic variants within targets (2-200 kb in length) with the low cost and high accuracy of short-read sequencing.

Early effect of standalone oblique lateral interbody fusion vs. combined with lateral screw fixation of the vertebral body on single-level lumbar degenerative disc disease: A pilot study.

BACKGROUND: For the treatment of single-level lumbar degenerative disc disease (DDD), oblique lateral interbody fusion (OLIF) has clinical advantages. Whether internal fixation needs to be combined for treatment has been the subject of debate. OBJECTIVE: To compare the early clinical effects of standalone oblique lateral interbody fusion (S-OLIF) versus OLIF combined with lateral screw fixation of the vertebral body (F-OLIF) on single-level lumbar DDD. METHODS: A retrospective analysis was performed on the data of 34 patients for whom the OLIF technique was applied to treat single-level lumbar DDD from August 2018 to May 2021. Patients were divided into the S-OLIF (n= 18) and F-OLIF groups (n= 16). Intraoperative blood loss, operative time, and length of hospital stay were recorded. The pain visual analogue scale (VAS) and Oswestry disability index (ODI) before and after the operation were evaluated. The disc height (DH), foraminal height (FH), fused segment lordosis (FSL), lumbar lordosis (LL), cage subsidence, and fusion by CT examination were measured before and after the operation. RESULTS: The S-OLIF group experienced a shorter operative time and less intraoperative blood loss than the F-OLIF group, and the differences were statistically significant (p< 0.05), but the difference in the length of hospital stay was not statistically significant. The postoperative VAS score and ODI of the two groups were significantly lower than those before the operation, but the postoperative differences between the two groups were not statistically significant. Differences were not statistically significant in postoperative FH, DH, FSL and LL of the two groups. Both groups were followed up for no less than 12 months. In the two groups, fusion was achieved at the last follow-up visit. CONCLUSION: According to short-term follow-up results, both S-OLIF and F-OLIF can achieve reliable and stable fusion and good clinical effect in the treatment of single-level lumbar DDD.

The effect of preperitoneal pelvic packing for hemodynamically unstable patients with pelvic fractures.

PURPOSE: Hemodynamically unstable patients with pelvic fractures still represent a challenge to trauma surgeons and have a very high mortality. This study was designed to explore the effect of the interventions of direct preperitoneal pelvic packing for the hemodynamically unstable pelvic fractures. METHODS: This retrospective study enrolled 67 cases of severe pelvic fractures with unstable hemodynamics from October 2011 to December 2019. All patients presented in our emergency center and received preperitoneal pelvic packing were included in this study. The indication was persistent systolic blood pressure ≤90 mmHg during initial resuscitation and after transfusion of two units of red blood cells. Patients with hemodynamic stability who need no preperitoneal pelvic packing to control bleeding were excluded. Their demographic characteristics, clinical features, laboratory results, therapeutic interventions, adverse events, and prognostic outcomes were collected from digital information system of electronic medical records. Statistics were described as mean ± standard deviation or medium and analyzed using pair sample t-test or Mann-Whitney U-test. RESULTS: The patients' average age was 41.6 years, ranging from 10 to 88 years. Among them, 45 cases were male (67.2%) and 22 cases were female (32.8%). Significant difference was found regarding the systolic blood pressure (mmHg) in the emergency department (78.4 ± 13.9) and after preperitoneal pelvic packing in the surgery intensive care unit (100.1 ± 17.6) (p < 0.05). Simultaneously, the arterial base deficit (mmol/L) were significantly lower in the surgery intensive care unit (median -6, interquartile range -8 to -2) than in the emergency department (median -10, interquartile range -14 to -8) (p < 0.05). After preperitoneal pelvic packing, 15 patients (22.4%) underwent pelvic angiography for persistent hypotension or suspected ongoing haemorrhage. The overall mortality rate was 29.5% (20 of 67). CONCLUSIONS: Preperitoneal pelvic packing, as a useful surgical technique, is less invasive and can be very efficient in early intra-pelvic bleed control.

The genome of opportunistic fungal pathogen Fusarium oxysporum carries a unique set of lineage-specific chromosomes.

Fusarium oxysporum is a cross-kingdom fungal pathogen that infects plants and humans. Horizontally transferred lineage-specific (LS) chromosomes were reported to determine host-specific pathogenicity among phytopathogenic F. oxysporum. However, the existence and functional importance of LS chromosomes among human pathogenic isolates are unknown. Here we report four unique LS chromosomes in a human pathogenic strain NRRL 32931, isolated from a leukemia patient. These LS chromosomes were devoid of housekeeping genes, but were significantly enriched in genes encoding metal ion transporters and cation transporters. Homologs of NRRL 32931 LS genes, including a homolog of ceruloplasmin and the genes that contribute to the expansion of the alkaline pH-responsive transcription factor PacC/Rim1p, were also present in the genome of NRRL 47514, a strain associated with Fusarium keratitis outbreak. This study provides the first evidence, to our knowledge, for genomic compartmentalization in two human pathogenic fungal genomes and suggests an important role of LS chromosomes in niche adaptation.

Electrostatic confinement and manipulation of DNA molecules for genome analysis.

Very large DNA molecules enable comprehensive analysis of complex genomes, such as human, cancer, and plants because they span across sequence repeats and complex somatic events. When physically manipulated, or analyzed as single molecules, long polyelectrolytes are problematic because of mechanical considerations that include shear-mediated breakage, dealing with the massive size of these coils, or the length of stretched DNAs using common experimental techniques and fluidic devices. Accordingly, we harness analyte "issues" as exploitable advantages by our invention and characterization of the "molecular gate," which controls and synchronizes formation of stretched DNA molecules as DNA dumbbells within nanoslit geometries. Molecular gate geometries comprise micro- and nanoscale features designed to synergize very low ionic strength conditions in ways we show effectively create an "electrostatic bottle." This effect greatly enhances molecular confinement within large slit geometries and supports facile, synchronized electrokinetic loading of nanoslits, even without dumbbell formation. Device geometries were considered at the molecular and continuum scales through computer simulations, which also guided our efforts to optimize design and functionalities. In addition, we show that the molecular gate may govern DNA separations because DNA molecules can be electrokinetically triggered, by varying applied voltage, to enter slits in a size-dependent manner. Lastly, mapping the Mesoplasmaflorum genome, via synchronized dumbbell formation, validates our nascent approach as a viable starting point for advanced development that will build an integrated system capable of large-scale genome analysis.

Gapless genome assembly of Colletotrichum higginsianum reveals chromosome structure and association of transposable elements with secondary metabolite gene clusters.

BACKGROUND: The ascomycete fungus Colletotrichum higginsianum causes anthracnose disease of brassica crops and the model plant Arabidopsis thaliana. Previous versions of the genome sequence were highly fragmented, causing errors in the prediction of protein-coding genes and preventing the analysis of repetitive sequences and genome architecture. RESULTS: Here, we re-sequenced the genome using single-molecule real-time (SMRT) sequencing technology and, in combination with optical map data, this provided a gapless assembly of all twelve chromosomes except for the ribosomal DNA repeat cluster on chromosome 7. The more accurate gene annotation made possible by this new assembly revealed a large repertoire of secondary metabolism (SM) key genes (89) and putative biosynthetic pathways (77 SM gene clusters). The two mini-chromosomes differed from the ten core chromosomes in being repeat- and AT-rich and gene-poor but were significantly enriched with genes encoding putative secreted effector proteins. Transposable elements (TEs) were found to occupy 7% of the genome by length. Certain TE families showed a statistically significant association with effector genes and SM cluster genes and were transcriptionally active at particular stages of fungal development. All 24 subtelomeres were found to contain one of three highly-conserved repeat elements which, by providing sites for homologous recombination, were probably instrumental in four segmental duplications. CONCLUSION: The gapless genome of C. higginsianum provides access to repeat-rich regions that were previously poorly assembled, notably the mini-chromosomes and subtelomeres, and allowed prediction of the complete SM gene repertoire. It also provides insights into the potential role of TEs in gene and genome evolution and host adaptation in this asexual pathogen.

A gapless genome sequence of the fungus Botrytis cinerea.

Following earlier incomplete and fragmented versions of a genome sequence for the grey mould Botrytis cinerea, a gapless, near-finished genome sequence for B. cinerea strain B05.10 is reported. The assembly comprised 18 chromosomes and was confirmed by an optical map and a genetic map based on approximately 75 000 single nucleotide polymorphism (SNP) markers. All chromosomes contained fully assembled centromeric regions, and 10 chromosomes had telomeres on both ends. The genetic map consisted of 4153 cM and a comparison of the genetic distances with the physical distances identified 40 recombination hotspots. The linkage map also identified two mutations, located in the previously described genes Bos1 and BcsdhB, that conferred resistance to the fungicides boscalid and iprodione. The genome was predicted to encode 11 701 proteins. RNAseq data from >20 different samples were used to validate and improve gene models. Manual curation of chromosome 1 revealed interesting features, such as the occurrence of a dicistronic transcript and fully overlapping genes in opposite orientations, as well as many spliced antisense transcripts. Manual curation also revealed that the untranslated regions (UTRs) of genes can be complex and long, with many UTRs exceeding lengths of 1 kb and possessing multiple introns. Community annotation is in progress.

Genome Sequence and Annotation of Colletotrichum higginsianum, a Causal Agent of Crucifer Anthracnose Disease.

Colletotrichum higginsianum is an ascomycete fungus causing anthracnose disease on numerous cultivated plants in the family Brassicaceae, as well as the model plant Arabidopsis thaliana We report an assembly of the nuclear genome and gene annotation of this pathogen, which was obtained using a combination of PacBio long-read sequencing and optical mapping.

Allele-Specific Quantification of Structural Variations in Cancer Genomes.

Aneuploidy and structural variations (SVs) generate cancer genomes containing a mixture of rearranged genomic segments with extensive somatic copy number alterations. However, existing methods can identify either SVs or allele-specific copy number alterations but not both simultaneously, which provides a limited view of cancer genome structure. Here, we introduce Weaver, an algorithm for the quantification and analysis of allele-specific copy numbers of SVs. Weaver uses a Markov random field to estimate joint probabilities of allele-specific copy numbers of SVs and their inter-connectivity based on paired-end whole-genome sequencing data. Weaver also predicts the timing of SVs relative to chromosome amplifications. We demonstrate the accuracy of Weaver using simulations and findings from whole-genome optical mapping. We apply Weaver to generate allele-specific copy numbers of SVs for MCF-7 and HeLa cell lines and identify recurrent SV patterns in 44 TCGA ovarian cancer whole-genome sequencing datasets. Our approach provides a more complete assessment of the complex genomic architectures inherent to many cancer genomes.

Who are the Best Candidates for Decompressive Surgery and Spine Stabilization in Patients With Metastatic Spinal Cord Compression?: A New Scoring System.

STUDY DESIGN: A retrospective study. OBJECTIVE: This study aims to develop a new scoring system that can guild surgeons to select the best candidates for decompressive surgery in patients with metastatic spinal cord compression (MSCC). SUMMARY OF BACKGROUND DATA: Predicting survival and functional outcome is essential when selecting the individual treatment for patients with MSCC. The criteria for identifying MSCC patients who are most likely to benefit from decompressive surgery remain unclear. METHODS: We retrospectively analyzed 12 preoperative characteristics for postoperative survival in a series of 206 patients with MSCC who were operated with decompressive surgery and spine stabilization. Characteristics significantly associated with survival in the multivariate analysis were included in the scoring system. Postoperative function outcome was also analyzed on the basis of the scoring system. RESULTS: According to the multivariate analysis, primary site (P < 0.01), preoperative ambulatory status (P < 0.01), visceral metastases (P < 0.01), preoperative chemotherapy (P = 0.02), and bone metastasis at cancer diagnosis (P = 0.03) had a significant impact on postoperative survival and were included in the scoring system. According to the prognostic scores, which ranged from 0 to 10 points, three risk groups were designed: 0 to 2, 3 to 5, and 6 to 10 points. The corresponding 6 months survival rates were 8.2%, 56.5%, and 91.5%, respectively (P < 0.01), and postoperative ambulatory rates were 35.7%, 73.3%, and 95.9%, respectively (P < 0.01). CONCLUSION: We present a new scoring system for predicting survival and function outcome of MSCC patients after surgical decompression and spine stabilization. This new scoring system can help surgeons select the best candidates for surgical treatment. LEVEL OF EVIDENCE: 4.

Tantalum rod implantation for femoral head osteonecrosis: survivorship analysis and determination of prognostic factors for total hip arthroplasty.

PURPOSE: The purpose of this study was to evaluate the moderate survival data of porous tantalum rod implants for the treatment of osteonecrosis of the femoral head (ONFH). Additionally, some independent prognostic factors for conversion to total hip arthroplasty (THA) were identified. METHODS: The porous tantalum rod population was obtained from a prospective, consecutive group of patients treated for Steinberg stage I and II osteonecrosis from April 2009 through July 2011. The historical core decompression and impaction of bone filling particle subjects underwent surgery from April 2007 through March 2009. Surgical data including time of surgery, blood loss, and cell transfusions were recorded. Post-operative values were measured for hospitalization length as well as days requiring a patient-controlled analgesia (PCA) pump. Primary outcomes were Harris hip score and survivorship analysis. Demographics and baseline characteristics included age, sex, etiology, bilateral disease, associated chronic systemic disease, Steinberg stage, Harris hip score, accompanied with bone marrow edema of femoral head, and osteonecrotic lesion size. RESULTS: Demographic/baseline characteristics were similar between two groups. At the post-operative follow-up of 62 months, Harris hip scores were significantly increased (P < 0.0001) when compared to that before surgery in both groups. The magnitude of increase in the tantalum rod implant group was significantly greater than that in the control group (P = 0.0426). With an average follow-up of 48 months (range, 38-62 months), the tantalum rod group had an 84.6 % survival rate. With an average follow-up of 72 months (range, 67-85 months), the control group had a 63.3 % survival rate. A comparison of Kaplan-Meier curves showed significantly higher cumulative survival rates (P = 0.048) for hips with implantation of the porous tantalum rod (74.1 % at 62 months) than for those with impaction composite bone material (49.9 % at 62 months). The Cox proportional-hazard model revealed that implantation of tantalum rod (P = 0.012), bone marrow edema (P = 0.003), corticosteroids intake (P = 0.007), and age less than 50 years (P = 0.014) were the independent prognostic factors related to conversion into THA. CONCLUSIONS: Compared with the traditional impaction composite bone material technique, implantation of tantalum rod in the treatment of Steinberg stages I and II ONFH can obtain better clinical results and higher cumulative survival rates. For patients without the use of corticosteroids, and especially for hips without bone marrow oedema, the clinical results from our study show highly encouraging survival rates and a delay in or prevention of conversion into THA.

Prediction of survival prognosis after surgery in patients with symptomatic metastatic spinal cord compression from non-small cell lung cancer.

BACKGROUND: The aim of this study was to develop a scoring system for prediction of survival prognosis after surgery in patients with symptomatic metastatic spinal cord compression (MSCC) from non-small cell lung cancer (NSCLC). METHODS: We retrospectively analyzed nine preoperative characteristics for survival in a series of 64 patients with NSCLC who were operated with posterior decompression and spine stabilization for MSCC. Characteristics significantly associated with survival on multivariate analysis were included in the scoring system. The scoring point for each significant characteristic was derived from the hazard ratios on Cox proportional hazards model. The total score for each patient was obtained by adding the scoring points of all significant characteristics. RESULTS: Eastern Cooperative Oncology Group (ECOG) performance status, number of involved vertebrae, visceral metastases, and time developing motor deficits had significant impact on survival on multivariate analysis and were included in the scoring system. According to the prognostic scores, which ranged from 4 to 10 points, three prognostic groups were designed: 4-5 points (n = 22), 6-7 points (n = 23), and 8-10 points (n = 19). The corresponding 6-month survival rates were 95, 47 and 11%, respectively (P < 0.0001). In addition, the functional outcome was worse in the group of patients with 8-10 points compared with other two prognostic groups. CONCLUSIONS: The new scoring system will enable physicians to identify patient with MSCC from NSCLC who may be a candidate for decompression and spine stabilization, more radical surgery, or supportive care alone. Patients with scores of 4-5, who have the most favorable survival prognosis and functional outcome, can be treated with more radical surgery in order to realize better local control of disease and prevent the occurrence of local disease. Patients with scores of 6-7 points should be surgical candidates, because survival prognosis and functional outcome are acceptable after surgery, while patients with scores of 8-10 points, who have the shortest survival time and poorest functional outcome after surgery, appear to be best treated with radiotherapy or best supportive care.

Association between VEGF -634G/C polymorphism and osteonecrosis of the femoral head susceptibility: a meta analysis.

BACKGROUND: VEGF plays an important role in bone formation and repair. However, the effects of VEGF -634G/C polymorphisms on the pathogenesis of osteonecrosis of the femoral head (ONFH) were not conclusive. Our research was aimed to further analyze the association of VEGF -634G/C polymorphism with ONFH risk. METHODS: The relevant articles were searched in PubMed, Elsevier, EMBASE, Web of Science and Chinese National Knowledge Infrastructure (CNKI) database. And a total of 692 cases and 875 controls were included. Pooled odds ratios (ORs) with 95% confidence intervals (CIs) were calculated to assess the correlation of VEGF -634G/C polymorphism and ONFH susceptibility. Chi-square based Q-statistic test was used to evaluate heterogeneity among the studies. The random-effects model or fixed-effects model was used depending on heterogeneity. RESULTS: The sensitivity analysis and publication bias test indicated that our results were stable and credible. And the results suggested that VEGF -634G/C polymorphism was significantly related with increased risk for ONFH in Asian population (CC versus GG: OR=1.34, 95% CI=1.02-1.76). CONCLUSION: The results indicated that VEGF -634G/C polymorphism might serve as genetic-susceptibility factor for ONFH.

A clone-free, single molecule map of the domestic cow (Bos taurus) genome.

BACKGROUND: The cattle (Bos taurus) genome was originally selected for sequencing due to its economic importance and unique biology as a model organism for understanding other ruminants, or mammals. Currently, there are two cattle genome sequence assemblies (UMD3.1 and Btau4.6) from groups using dissimilar assembly algorithms, which were complemented by genetic and physical map resources. However, past comparisons between these assemblies revealed substantial differences. Consequently, such discordances have engendered ambiguities when using reference sequence data, impacting genomic studies in cattle and motivating construction of a new optical map resource--BtOM1.0--to guide comparisons and improvements to the current sequence builds. Accordingly, our comprehensive comparisons of BtOM1.0 against the UMD3.1 and Btau4.6 sequence builds tabulate large-to-immediate scale discordances requiring mediation. RESULTS: The optical map, BtOM1.0, spanning the B. taurus genome (Hereford breed, L1 Dominette 01449) was assembled from an optical map dataset consisting of 2,973,315 (439 X; raw dataset size before assembly) single molecule optical maps (Rmaps; 1 Rmap = 1 restriction mapped DNA molecule) generated by the Optical Mapping System. The BamHI map spans 2,575.30 Mb and comprises 78 optical contigs assembled by a combination of iterative (using the reference sequence: UMD3.1) and de novo assembly techniques. BtOM1.0 is a high-resolution physical map featuring an average restriction fragment size of 8.91 Kb. Comparisons of BtOM1.0 vs. UMD3.1, or Btau4.6, revealed that Btau4.6 presented far more discordances (7,463) vs. UMD3.1 (4,754). Overall, we found that Btau4.6 presented almost double the number of discordances than UMD3.1 across most of the 6 categories of sequence vs. map discrepancies, which are: COMPLEX (misassembly), DELs (extraneous sequences), INSs (missing sequences), ITs (Inverted/Translocated sequences), ECs (extra restriction cuts) and MCs (missing restriction cuts). CONCLUSION: Alignments of UMD3.1 and Btau4.6 to BtOM1.0 reveal discordances commensurate with previous reports, and affirm the NCBI's current designation of UMD3.1 sequence assembly as the "reference assembly" and the Btau4.6 as the "alternate assembly." The cattle genome optical map, BtOM1.0, when used as a comprehensive and largely independent guide, will greatly assist improvements to existing sequence builds, and later serve as an accurate physical scaffold for studies concerning the comparative genomics of cattle breeds.

Single-molecule analysis reveals widespread structural variation in multiple myeloma.

Multiple myeloma (MM), a malignancy of plasma cells, is characterized by widespread genomic heterogeneity and, consequently, differences in disease progression and drug response. Although recent large-scale sequencing studies have greatly improved our understanding of MM genomes, our knowledge about genomic structural variation in MM is attenuated due to the limitations of commonly used sequencing approaches. In this study, we present the application of optical mapping, a single-molecule, whole-genome analysis system, to discover new structural variants in a primary MM genome. Through our analysis, we have identified and characterized widespread structural variation in this tumor genome. Additionally, we describe our efforts toward comprehensive characterization of genome structure and variation by integrating our findings from optical mapping with those from DNA sequencing-based genomic analysis. Finally, by studying this MM genome at two time points during tumor progression, we have demonstrated an increase in mutational burden with tumor progression at all length scales of variation.

Combined effect of tnf-α polymorphisms and hypoxia on steroid-induced osteonecrosis of femoral head.

OBJECTIVE: Tumor necrosis factor (TNF)-α is a proinflammatory cytokine, some studies reported that TNF-α gene plays important role in the pathogenesis of SONFH. And the polymorphisms of TNF-α were presented as risk factors for steroid-induced osteonecrosis of the femoral head (SONFH). Meanwhile, various environment factors involve in the pathogenesis of SONFH. Our study aimed to investigate the interaction effect of TNF-α polymorphisms and hypoxia factor on SONFH. METHODS: 120 patients with SONFH and 100 healthy people, matched with the cases on age and sex, participated in this study. DNA was extracted from all participants. According to previous studies, genotyping of TNF-α polymorphisms (rs1800629, rs1799964 and rs1800630) was tested with the method of PCR-RDB (Reverse Dot Blot). Environmental factors were also chose. Logistic regression analysis was used to analyze the interaction between TNF-α polymorphisms and environment factors on SONFH. RESULTS: The polymorphisms of rs1800629 and rs1800630 were significantly associated with SONFH (OR: 3.70, 9.93). Patients with hypoxia history were found higher (65.00%) compared with the healthy controls (43.00%). For the person with hypoxic history, GG and AG+AA genotypes of rs1800629 could increase their risk to suffer SONFH (OR: 2.12, 3.78). If the patients with the variant genotypes of rs1800630 experienced hypoxia state, then the risk for SONFH increased 2.41 folds. CONCLUSION: We concluded that the onset of SONFH was influenced by TNF-α and hypoxia history. There existed strong interaction between TNF-α and hypoxia history.

Chromosome-level genome map provides insights into diverse defense mechanisms in the medicinal fungus Ganoderma sinense.

Fungi have evolved powerful genomic and chemical defense systems to protect themselves against genetic destabilization and other organisms. However, the precise molecular basis involved in fungal defense remain largely unknown in Basidiomycetes. Here the complete genome sequence, as well as DNA methylation patterns and small RNA transcriptomes, was analyzed to provide a holistic overview of secondary metabolism and defense processes in the model medicinal fungus, Ganoderma sinense. We reported the 48.96 Mb genome sequence of G. sinense, consisting of 12 chromosomes and encoding 15,688 genes. More than thirty gene clusters involved in the biosynthesis of secondary metabolites, as well as a large array of genes responsible for their transport and regulation were highlighted. In addition, components of genome defense mechanisms, namely repeat-induced point mutation (RIP), DNA methylation and small RNA-mediated gene silencing, were revealed in G. sinense. Systematic bioinformatic investigation of the genome and methylome suggested that RIP and DNA methylation combinatorially maintain G. sinense genome stability by inactivating invasive genetic material and transposable elements. The elucidation of the G. sinense genome and epigenome provides an unparalleled opportunity to advance our understanding of secondary metabolism and fungal defense mechanisms.

A modified porous tantalum implant technique for osteonecrosis of the femoral head: survivorship analysis and prognostic factors for radiographic progression and conversion to total hip arthroplasty.

Tantalum rod implant following core decompression is reported to be effective in early stage of osteonecrosis of the femoral head (ONFH). The purpose of this study was to assess the survivorship and prognostic factors for radiographic progression and conversion to total hip arthroplasty (THA) after treatment with a modified tantalum implant technology. 59 consecutive hips (45 patients) in whom ONFH was treated with core decompression, impaction bone grafting of 2 mm-composite bone filling material, and insertion of a porous tantalum implant. 57 hips (44 patients, mean age 43 years, range 21 to 70 years) with Steinberg Stage I-IVA ONFH were available for follow-up at a mean of 44.8 months (rang, 11 to 62 months). Outcome measures included HHS (Harris Hip Score), radiographic outcome, and survivorship analysis with reversion to THA. Radiographic progression occurred in 17 hips (17/57, 29.82%). 11 hips (11/57, 19.30%) were converted to THA. The overall survival rate was 72.49% at 60 months post-operatively. After logistic regression analysis, corticosteroid use and bone marrow edema were found to be predictors of radiographic progression. The Cox proportional-hazard model revealed that bone marrow edema was an independent prognostic factor for conversion to THA. This modified technology may make patients avoid the use of corticosteroid, especially those without bone marrow edema, and obtains encouraging survival rates and a delay in or prevention of THA.