A Locked Nucleic Acid (LNA)-Based Real-Time PCR Assay for the Rapid Detection of Multiple Bacterial Antibiotic Resistance Genes Directly from Positive Blood Culture

Bacterial strains resistant to various antibiotic drugs are frequently encountered in clinical infections, and the rapid identification of drug-resistant strains is highly essential for clinical treatment. We developed a locked nucleic acid (LNA)-based quantitative real-time PCR (LNA-qPCR) method for the rapid detection of 13 antibiotic resistance genes and successfully used it to distinguish drug-resistant bacterial strains from positive blood culture samples. A sequence-specific primer-probe set was designed, and the specificity of the assays was assessed using 27 ATCC bacterial strains and 77 negative blood culture samples. No cross-reaction was identified among bacterial strains and in negative samples, indicating 100% specificity. The sensitivity of the assays was determined by spiking each bacterial strain into negative blood samples, and the detection limit was 1–10 colony forming units (CFU) per reaction. The LNA-qPCR assays were first applied to 72 clinical bacterial isolates for the identification of known drug resistance genes, and the results were verified by the direct sequencing of PCR products. Finally, the LNA-qPCR assays were used for the detection in 47 positive blood culture samples, 19 of which (40.4%) were positive for antibiotic resistance genes, showing 91.5% consistency with phenotypic susceptibility results. In conclusion, LNA-qPCR is a reliable method for the rapid detection of bacterial antibiotic resistance genes and can be used as a supplement to phenotypic susceptibility testing for the early detection of antimicrobial resistance to allow the selection of appropriate antimicrobial treatment and to prevent the spread of resistant isolates.     

干细胞向骨骼肌细胞的诱导分化及其在治疗骨骼肌病变中的应用

随着干细胞技术、生物材料及组织工程技术的日趋发展,专家和学者的研究不仅强调对细胞生长的调控及机制,而且越来越注重其临床应用的潜能。运用组织工程和干细胞等技术重建和再生骨骼肌可以为肌肉相关疾病的治疗带来新的希望。干细胞是一类具有自我复制和多向分化潜能的细胞,在合适的培养条件或给予合适的信号时,干细胞可以分化为多种不同形态和功能的成熟细胞,也因此在骨骼肌重建和疾病治疗中得到广泛研究。该文就干细胞向骨骼肌细胞的分化及机制、干细胞作为骨骼肌组织工程的种子细胞以及在骨骼肌疾病治疗中的应用和前景作一综述。     

Genetic Analysis Identifies DDR2 as a Novel Gene Affecting Bone Mineral Density and Osteoporotic Fractures in Chinese Population

DDR2 gene, playing an essential role in regulating osteoblast differentiation and chondrocyte maturation, may influence bone mineral density (BMD) and osteoporosis, but the genetic variations actually leading to the association remain to be elucidated. Therefore, the aim of this study was to investigate whether the genetic variants in DDR2 are associated with BMD and fracture risk. This study was performed in three samples from two ethnicities, including 1,300 Chinese Han subjects, 700 Chinese Han subjects (350 with osteoporotic hip fractures and 350 healthy controls) and 2,286 US white subjects. Twenty-eight SNPs in DDR2 were genotyped and tested for associations with hip BMD and fractures. We identified 3 SNPs in DDR2 significantly associated with hip BMD in the Chinese population after multiple testing adjustments, which were rs7521233 (P = 1.06×10⁻⁴, β: −0.018 for allele C), rs7553831 (P = 1.30×10⁻⁴, β: −0.018 for allele T), and rs6697469 (P = 1.59×10⁻³, β: −0.015 for allele C), separately. These three SNPs were in high linkage disequilibrium. Haplotype analyses detected two significantly associated haplotypes, including one haplotype in block 2 (P = 9.54×10⁻⁴, β: −0.016) where these three SNPs located. SNP rs6697469 was also associated with hip fractures (P = 0.043, OR: 1.42) in the Chinese population. The effect on fracture risk was consistent with its association with lower BMD. However, in the white population, we didn’t observe significant associations with hip BMD. eQTL analyses revealed that SNPs associated with BMD also affected DDR2 mRNA expression levels in Chinese. Our findings, together with the prior biological evidence, suggest that DDR2 could be a new candidate for osteoporosis in Chinese population. Our results also reveal an ethnic difference, which highlights the need for further genetic studies in each ethnic group.     

A Weighted Polygenic Risk Score Using 14 Known Susceptibility Variants to Estimate Risk and Age Onset of Psoriasis in Han Chinese

With numbers of common variants identified mainly through genome-wide association studies (GWASs), there is great interest in incorporating the findings into screening individuals at high risk of psoriasis. The purpose of this study is to establish genetic prediction models and evaluate its discriminatory ability in psoriasis in Han Chinese population. We built the genetic prediction models through weighted polygenic risk score (PRS) using 14 susceptibility variants in 8,819 samples. We found the risk of psoriasis among individuals in the top quartile of PRS was significantly larger than those in the lowest quartile of PRS (OR = 28.20, P < 2.0×10^-16). We also observed statistically significant associations between the PRS, family history and early age onset of psoriasis. We also built a predictive model with all 14 known susceptibility variants and alcohol consumption, which achieved an area under the curve statistic of ~ 0.88. Our study suggests that 14 psoriasis known susceptibility loci have the discriminating potential, as is also associated with family history and age of onset. This is the genetic predictive model in psoriasis with the largest accuracy to date.     

DNA Methylation in Cosmc Promoter Region and Aberrantly Glycosylated IgA1 Associated with Pediatric IgA Nephropathy

IgA nephropathy (IgAN) is one of the most common glomerular diseases leading to end-stage renal failure. Elevation of aberrantly glycosylated IgA1 is a key feature of it. The expression of the specific molecular chaperone of core1ß1, 3galactosyl transferase (Cosmc) is known to be reduced in IgAN. We aimed to investigate whether the methylation of CpG islands of Cosmc gene promoter region could act as a possible mechanism responsible for down-regulation of Cosmc and related higher secretion of aberrantly glycosylated IgA1in lymphocytes from children with IgA nephropathy. Three groups were included: IgAN children (n = 26), other renal diseases (n = 11) and healthy children (n = 13). B-lymphocytes were isolated and cultured, treated or not with IL-4 or 5-Aza-2’-deoxycytidine (AZA). The levels of DNA methylation of Cosmc promotor region were not significantly different between the lymphocytes of the three children populations (P = 0.113), but there were significant differences between IgAN lymphocytes and lymphocytes of the other two children populations after IL-4 (P<0.0001) or AZA (P<0.0001). Cosmc mRNA expression was low in IgAN lymphocytes compared to the other two groups (P<0.0001). The level of aberrantly glycosylated IgA1 was markedly higher in IgAN group compared to the other groups (P<0.0001). After treatment with IL-4, the levels of Cosmc DNA methylation and aberrantly glycosylated IgA1 in IgAN lymphocytes were remarkably higher than the other two groups (P<0.0001) with more markedly decreased Cosmc mRNA content (P<0.0001). After treatment with AZA, the levels in IgAN lymphocytes were decreased, but was still remarkably higher than the other two groups (P<0.0001), while Cosmc mRNA content in IgAN lymphocytes were more markedly increased than the other two groups (P<0.0001). The alteration of DNA methylation by IL-4 or AZA specifically correlates in IgAN lymphocytes with alterations in Cosmc mRNA expression and with the level of aberrantly glycosylated IgA1 (r = −0.948, r = 0. 707). Our results suggested that hypermethylation of Cosmc promoter region could be a key mechanism for the reduction of Cosmc mRNA expression in IgAN lymphocytes with associated increase in aberrantly glycosylated IgA1.     

Body Mass Index Is Associated with Physical Performance in Suburb-Dwelling Older Chinese: A Cross-Sectional Study

Background

Physical performance is reported to have various beneficial effects on human health, especially in older individuals. Although such effects are associated with body mass index (BMI), the relationship between BMI and physical performance has not been clarified.

Design

We conducted a cross-sectional study of 966 suburb-dwelling Tianjin individuals aged ≥ 60 years (average age 67.5±6.02, men 435, women 531). Mobility, balance, and muscle strength were assessed by walking speed, timed up-and-go test (TUGT), and grip strength, respectively. The subjects were categorized into three groups based on BMI (kg/m²) as follows: normal weight, 18.5 ≤ BMI ≤ 23.9; overweight, 24.0 ≤ BMI ≤ 27.9; and obese, BMI ≥ 28.0.

Result

After adjusting for all other variables, relative grip strength decreased when BMI increased in both men and women (P for trend <0.001 and <0.001, respectively). BMI may be negatively associated with TUGT performance in the women only. There was no apparent association between walking speed and BMI in either sex, but after adjusting for age, walking speed was faster when BMI increased in women (P for trend= 0.0162).

Conclusion

This study suggests that in older individuals, higher BMI is associated with poor muscle strength in both sexes.     

Multi-Agent Patrolling under Uncertainty and Threats

We investigate a multi-agent patrolling problem where information is distributed alongside threats in environments with uncertainties. Specifically, the information and threat at each location are independently modelled as multi-state Markov chains, whose states are not observed until the location is visited by an agent. While agents will obtain information at a location, they may also suffer damage from the threat at that location. Therefore, the goal of the agents is to gather as much information as possible while mitigating the damage incurred. To address this challenge, we formulate the single-agent patrolling problem as a Partially Observable Markov Decision Process (POMDP) and propose a computationally efficient algorithm to solve this model. Building upon this, to compute patrols for multiple agents, the single-agent algorithm is extended for each agent with the aim of maximising its marginal contribution to the team. We empirically evaluate our algorithm on problems of multi-agent patrolling and show that it outperforms a baseline algorithm up to 44% for 10 agents and by 21% for 15 agents in large domains.     

The Effect of Long-Term Continuous Cropping of Black Pepper on Soil Bacterial Communities as Determined by 454 Pyrosequencing

In the present study, 3 replanted black pepper orchards with continuously cropping histories for 10, 21, and 55 years in tropical China, were selected for investigating the effect of monoculture on soil physiochemical properties, enzyme activities, bacterial abundance, and bacterial community structures. Results showed long-term continuous cropping led to a significant decline in soil pH, organic matter contents, enzymatic activities, and resulted in a decrease in soil bacterial abundance. 454 pyrosequencing analysis of 16S rRNA genes revealed that the Acidobacteria and Proteobacteria were the main phyla in the replanted black pepper orchard soils, comprising up to 73.82% of the total sequences; the relative abundances of Bacteroidetes and Firmicutes phyla decreased with long-term continuous cropping; and at genus level, the Pseudomonas abundance significantly depleted after 21 years continuous cropping. In addition, bacterial diversity significantly decreased after 55 years black pepper continuous cropping; obvious variations for community structures across the 3 time-scale replanted black pepper orchards were observed, suggesting monoculture duration was the major determinant for bacterial community structure. Overall, continuous cropping during black pepper cultivation led to a significant decline in soil pH, organic matter contents, enzymatic activities, resulted a decrease in soil bacterial abundance, and altered soil microbial community membership and structure, which in turn resulted in black pepper poor growth in the continuous cropping system.