Improving the sensitivity of long read overlap detection using grouped short k-mer matches

Background

Single-molecule, real-time sequencing (SMRT) developed by Pacific BioSciences produces longer reads than second-generation sequencing technologies such as Illumina. The increased read length enables PacBio sequencing to close gaps in genome assembly, reveal structural variations, and characterize the intra-species variations. It also holds the promise to decipher the community structure in complex microbial communities because long reads help metagenomic assembly. One key step in genome assembly using long reads is to quickly identify reads forming overlaps. Because PacBio data has higher sequencing error rate and lower coverage than popular short read sequencing technologies (such as Illumina), efficient detection of true overlaps requires specially designed algorithms. In particular, there is still a need to improve the sensitivity of detecting small overlaps or overlaps with high error rates in both reads. Addressing this need will enable better assembly for metagenomic data produced by third-generation sequencing technologies.

Results

In this work, we designed and implemented an overlap detection program named GroupK, for third-generation sequencing reads based on grouped k-mer hits. While using k-mer hits for detecting reads’ overlaps has been adopted by several existing programs, our method uses a group of short k-mer hits satisfying statistically derived distance constraints to increase the sensitivity of small overlap detection. Grouped k-mer hit was originally designed for homology search. We are the first to apply group hit for long read overlap detection. The experimental results of applying our pipeline to both simulated and real third-generation sequencing data showed that GroupK enables more sensitive overlap detection, especially for datasets of low sequencing coverage.

Conclusions

GroupK is best used for detecting small overlaps for third-generation sequencing data. It provides a useful supplementary tool to existing ones for more sensitive and accurate overlap detection. The source code is freely available at https://github.com/Strideradu/GroupK.

     

双歧杆菌三联活菌对混合痔选择性痔上黏膜吻合术术后患者便秘的预防效果及对肠道菌群的影响

探讨双歧杆菌三联活菌在混合痔选择性痔上黏膜吻合术（TST）术后患者中的应用价值,为临床预防术后便秘提供参考。

选取2021年8月至2023年2月我院收治的100例混合痔TST术后患者为研究对象,并分为对照组（n=46,接受莫沙必利治疗）和研究组（n=54,接受莫沙必利联合双歧杆菌三联活菌治疗）。持续治疗2周后,比较两组患者术后便秘发生情况、术后恢复情况（排气时间、排便时间、住院时间）、排便困难程度、粪便性状、肠道菌群数量、血清P物质（SP）、血管活性肠肽（VIP）水平。

研究组患者便秘发生率低于对照组（χ²=4.617,P=0.032）。研究组患者首次排气时间、排便时间及住院时间均短于对照组（t=15.103、13.358、8.463,均P＜0.001）。治疗2周后,研究组患者排便困难程度及粪便性状评分均低于对照组（t=5.221、4.001,均P＜0.001）。研究组患者肠道乳杆菌、双歧杆菌数量高于对照组,肠杆菌、球杆菌、酵母菌数量均低于对照组（t=4.646、3.688、2.454、4.818、8.681,均P＜0.001）。研究组患者血清SP水平高于对照组,VIP水平低于对照组（t=3.328、8.999,均P＜0.001）。

双歧杆菌三联活菌可预防混合痔TST术后患者便秘的发生,同时优化患者肠道菌群及脑肠肽类激素分泌。

     

Controls over leaf litter decomposition in a mixed evergreen and deciduous broad-leaved forest,Central China

Plant and Soil - Our understanding of the determinants of leaf litter decomposition is lacking for mixed evergreen and deciduous broad-leaved forests compared with tropical and temperate forests....     

Blockage of macrophage migration inhibitory factor (MIF) suppressed uric acid-induced vascular inflammation,smooth muscle cell de-differentiation,and remodeling

Hyperuricemia contributes to vascular injury and dysfunction, yet the potential mechanisms are not well understood. Uric acid (UA) has been found to stimulate macrophage migration inhibitory factor (MIF) up-regulation in renal tubules from rats subjected to UA-induced nephropathy. Given that MIF is able to induce vascular smooth muscle cell (VSMC) de-differentiation (from contractile state to a secretory state), we thus hypothesized that UA-induced vascular injury is via up-regulating of MIF in VSMCs, which enhancing vascular inflammation and VSMC transition. Within a mouse model of UA injection (500?mg/kg, twice/day, 14 days), we measured circulating and vascular MIF levels under UA stimulation at 6?h, day 1, and 14. We tested the efficacy of MIF inhibitor (10?mg/kg, twice/day, 14 days) on UA-induced vascular inflammation and remodeling. High plasma level of UA induced vascular MIF release into the plasma at acute phase. In the chronic phase, the protein level of MIF is up-regulated in the vessels. MIF inhibitor suppressed vascular inflammatory responses, repressed VSMC de-differentiation, and attenuated vascular remodeling and dysfunction following UA stimulation. Knockdown of MIF in cultured VSMCs repressed UA-induced de-differentiation. Our results provided a novel mechanism for MIF-mediated vascular injury in response to UA stimulation, and suggested that anti-MIF interventions may be of therapeutic value in hyperuricemic patients.     

地下滴灌下土壤水势对毛白杨纸浆林生长及生理特性的影响

在6、7年生三倍体毛白杨(triploid Populus tomentosa)纸浆林中研究了地下滴灌(SDI)下不同土壤水势(-25、-50、-75kPa,即灌溉起始阈值)对林木生长及生理特征的影响。结果表明:与不灌溉(CK)相比,SDI使6、7年生林分的生产力分别平均提高24%和28%;其中,-25 kPa使6年生林分的生产力达到39.90 m3hm-2a-1,较CK极显著提高43.5%(P<0.01)。各水势处理间,-25 kPa的生产力在林分6年生时分别较-50和-75 kPa极显著提高20%和31%(P<0.01),在7年生时分别提高13%和14%(P>0.05)。能在毛白杨速生期内(4—7月)大幅提高土壤含水率(20和50 cm处分别平均提高35%和27%)、树干日平均液流速率(46%,SFmean)、黎明前叶水势(41%,ψpd)是SDI促进林木生长的重要机制。灌溉起始水势阈值的差异对毛白杨SFmean和ψpd无显著影响(P>0.05)。3个水势处理中,-25 kPa的平均SFmean和ψpd均最高,且其能使根系活动剧烈的表土层(0—20cm)的水分有效性有最大幅的提高,这可能是其对林木生长有最大促进作用的主要原因。综上,应在毛白杨纸浆林培育中大力推广SDI,并在应用时可将距滴头10cm、地下20 cm处的土壤水势达到-25 kPa作为灌溉起始阈值。另外,在与试验地环境相似地区栽植毛白杨时,应于4—7月灌溉,8—10月一般可不灌溉。     

Tumour necrosis factor‐α promotes BMHSC differentiation by increasing P2X7 receptor in oestrogen‐deficient osteoporosis

The exact mechanism of tumour necrosis factor α (TNF‐α) promoting osteoclast differentiation is not completely clear. A variety of P2 purine receptor subtypes have been confirmed to be widely involved in bone metabolism. Thus, the purpose of this study was to explore whether P2 receptor is involved in the differentiation of osteoclasts. Mouse bone marrow haematopoietic stem cells (BMHSCs) were co‐cultured with TNF‐α to explore the effect of TNF‐α on osteoclast differentiation and bone resorption capacity in vitro, and changes in the P2 receptor were detected at the same time. The P2 receptor was silenced and overexpressed to explore the effect on differentiation of BMHSCs into osteoclasts. In an in vivo experiment, the animal model of PMOP was established in ovariectomized mice, and anti‐TNF‐α intervention was used to detect the ability of BMHCs to differentiate into osteoclasts as well as the expression of the P2 receptor. It was confirmed in vitro that TNF‐α at a concentration of 20 ng/mL up‐regulated the P2X7 receptor of BMHSCs through the PI3k/Akt signalling pathway, promoted BMHSCs to differentiate into a large number of osteoclasts and enhanced bone resorption. In vivo experiments showed that more P2X7 receptor positive osteoclasts were produced in postmenopausal osteoporotic mice. Anti‐TNF‐α could significantly delay the progression of PMOP by inhibiting the production of osteoclasts. Overall, our results revealed a novel function of the P2X7 receptor and suggested that suppressing the P2X7 receptor may be an effective strategy to delay bone formation in oestrogen deficiency‐induced osteoporosis.     

Physical Localization of Genes for CaM and Ca2+- ATPase in Rice by in situ Hybridization

CaM and Ca2 + -ATPase genes are important components of signal transduction chains which affect the regulation of' gene expression and development in plants. These two genes are tunctionally closely related. The rice ( Oryza sativa L. )cDNA probes C419 and SSU304 for these two genes, which are small single-copy ones and 0.8 and 0.3 kb in size respectively, were first physically mapped on rice chromosomes by biotin-labeled in situ hybridization. Both probes were detected on chromosome 5. The detection rate was 6.18 %, and the average chromosome ann ratios and standard deviations of detected chromosomes for probes C419 and SSU304 were 1.79 ± 0.06 and 1.91 ± 0.08 respectively. Probe C419 for CaM gene was located at the end of the long ann, and probe SSU304 for Ca2 + -ATPase gene -- on the short arm near the centromere. As it was reported before, they were closely linked in the high density genetic map. This demonstrated that there was a large discrepancy between the results of genetic and physical mapping of genes, and it indicated that the region between the functionally related genes could be the cold spot. What relationship there is between the region and gene expression is to be shudied further. The nonradioactive in situ hybridization technique about the physical mapping of low/single copy, short DNA fragments is also discussed.     

Change in the Green-Up Dates for Quercus mongolica in Northeast China and Its Climate-Driven Mechanism from 1962 to 2012

The currently available studies on the green-up date were mainly based on ground observations and/or satellite data, and few model simulations integrated with wide coverage satellite data have been reported at large scale over a long time period (i.e., > 30 years). In this study, we combined phenology mechanism model, long-term climate data and synoptic scale remote sensing data to investigate the change in the green-up dates for Quercus mongolica over 33 weather stations in Northeast China and its climate-driven mechanism during 1962-2012. The results indicated that the unified phenology model can be well parameterized with the satellite derived green-up dates. The optimal daily mean temperature for chilling effect was between -27°C and 1°C for Q. mongolica in Northeast China, while the optimal daily mean temperature for forcing effect was above -3°C. The green-up dates for Q. mongolica across Northeast China showed a delayed latitudinal gradient of 2.699 days degree^-1, with the earliest date on the Julian day 93 (i.e., 3^th April) in the south and the latest date on the Julian day 129 (i.e., 9^th May) in the north. The green-up date for Q. mongolica in Northeast China has advanced 6.6 days (1.3 days decade^-1) from 1962 to 2012. With the prevailing warming in autumn, winter and spring in Northeast China during the past 51 years, the chilling effect for Q. mongolica has been weakened, while the forcing effect has been enhanced. The advancing trend in the green-up dates for Q. mongolica implied that the enhanced forcing effect to accelerate green-up was stronger than the weakened chilling effect to hold back green-up while the changes of both effects were caused by the warming climate.