[Association study between haplotypes of WNT signaling pathway genes and nonsyndromic oral clefts among Chinese Han populations].

OBJECTIVE: To explore whether WNT signaling pathway genes were associated with non-syndromic oral clefts (NSOC) based on haplotypes analyses among 1 008 Chinese NSOC case-parent trios. METHODS: The genome-wide association study (GWAS) data of 806 Chinese non-syndromic cleft lip with or without cleft palate (NSCL/P) trios and 202 Chinese non-syndromic cleft palate (NSCP) case-parent trios were drawn from the International Consortium to Identify Genes and Interactions Controlling Oral Clefts (ICOCs) study GWAS data set, whose Chinese study population were recruited from four provinces in China, namely Taiwan, Shandong, Hubei, and Sichuan provinces. The process of DNA genotyping was conducted by the Center for Inherited Disease Research in the Johns Hopkins University, using Illumina Human610-Quad v.1_B Bead Chip. The method of sliding windows was used to determine the haplotypes for analyses, including 2 SNPs haplotypes and 3 SNPs haplotypes. Haplotypes with a frequency lower than 1% were excluded for further analyses. To further assess the association between haplotypes and NSOC risks, and the transmission disequilibrium test (TDT) was performed. The Bonferroni method was adopted to correct multiple tests in the study, with which the threshold of statistical significance level was set as P < 0.05 divided by the number of tests, e.g P < 3.47×10-4 in the current stu-dy. All the statistical analyses were performed by using plink (v1.07). RESULTS: After quality control, a total of 144 single nucleotide polymorphisms (SNPs) mapped in seven genes in WNT signaling pathway were included for the analyses among the 806 Chinese NSCL/P trios and 202 Chinese NSCP trios. A total of 1 042 haplotypes with frequency higher than 1% were included for NSCL/P analyses and another 1 057 haplotypes with frequency higher than 1% were included for NSCP analyses. Results from the TDT analyses showed that a total of 69 haplotypes were nominally associated with the NSCL/P risk among Chinese (P < 0.05). Another 34 haplotypes showed nominal significant association with the NSCP risk among Chinese (P < 0.05). However, none of these haplotypes reached pre-defined statistical significance level after Bonferroni correction (P>3.47×10-4). CONCLUSION: This study failed to observe any statistically significant associations between haplotypes of seven WNT signaling pathway genes and the risk of NSOC among Chinese. Further studies are warranted to replicate the findings here.

[Exploring the association between de novo mutations and non-syndromic cleft lip with or without palate based on whole exome sequencing of case-parent trios].

OBJECTIVE: To explore the association between de novo mutations (DNM) and non-syndromic cleft lip with or without palate (NSCL/P) using case-parent trio design. METHODS: Whole-exome sequencing was conducted for twenty-two NSCL/P trios and Genome Analysis ToolKit (GATK) was used to identify DNM by comparing the alleles of the cases and their parents. Information of predictable functions was annotated to the locus with SnpEff. Enrichment analysis for DNM was conducted to test the difference between the actual number and the expected number of DNM, and to explore whether there were genes with more DNM than expected. NSCL/P-related genes indicated by previous studies with solid evidence were selected by literature reviewing. Protein-protein interactions analysis was conducted among the genes with protein-altering DNM and NSCL/P-related genes. R package "denovolyzeR" was used for the enrichment analysis (Bonferroni correction: P=0.05/n, n is the number of genes in the whole genome range). Protein-protein interactions among genes with DNM and genes with solid evidence on the risk factors of NSCL/P were predicted depending on the information provided by STRING database. RESULTS: A total of 339 908 SNPs were qualified for the subsequent analysis after quality control. The number of high confident DNM identified by GATK was 345. Among those DNM, forty-four DNM were missense mutations, one DNM was nonsense mutation, two DNM were splicing site mutations, twenty DNM were synonymous mutations and others were located in intron or intergenic regions. The results of enrichment analysis showed that the number of protein-altering DNM on the exome regions was larger than expected (P < 0.05), and five genes (KRTCAP2, HMCN2, ANKRD36C, ADGRL2 and DIPK2A) had more DNM than expected (P < 0.05/(2×19 618)). Protein-protein interaction analysis was conducted among forty-six genes with protein-altering DNM and thirteen genes associated with NSCL/P selected by literature reviewing. Six pairs of interactions occurred between the genes with DNM and known NSCL/P-related genes. The score measuring the confidence level of the predicted interaction between RGPD4 and SUMO1 was 0.868, which was higher than the scores for other pairs of genes. CONCLUSION: Our study provided novel insights into the development of NSCL/P and demonstrated that functional analyses of genes carrying DNM were warranted to understand the genetic architecture of complex diseases.

[Exploring parent-of-origin effects for non-syndromic cleft lip with or without cleft palate on PTCH1, PTCH2, SHH, SMO genes in Chinese case-parent trios].

OBJECTIVE: Non-syndromic cleft lip with or without cleft palate (NSCL/P) is a common birth defect, affecting 1.4 per 1 000 live births, and multiple genetic and environmental risk factors influencing its risk. All the known genetic risk factors accounted for a small proportion of the heritability. Several authors have suggested parent-of-origin effects (PoO) may play an important role in the etiology of this complex and heterogeneous malformation. To clarify the genetic association between PTCH1, PTCH2, SHH and SMO in hedgehog (HH) pathway and NSCL/P, as well as testing for potential PoO effects in Chinese case-parent trios. METHODS: We tested for transmission disequilibrium tests (TDT) and PoO effects using 83 common single nucleotide polymorphic (SNP) markers of HH pathway genes from 806 NSCL/P case-parent trios. These trios were drawn from an international consortium established for a genome-wide association studies (GWAS) of non-syndromic oral clefts of multiple ethnicities. DNA samples were collected from each trio. Single marker and haplotype based analysis were performed both in TDT tests and PoO effects. SNPs were excluded if they (ⅰ) had a call rate of < 95%, (ⅱ) had a minor allele frequency (MAF) of < 0.05, (ⅲ) had Mendelian errors over all trios of >5%, (ⅳ) had a genotype distribution in the parents that deviated from the Hardy-Weinberg equilibrium (HWE) (P < 0.000 1). The process was done using Plink (version 1.07, http://pngu.mgh.harvard.edu/~purcell/plink/data.shtml). TDT test was performed in Plink v1.07. A log-linear model was used to explore PoO effects using Haplin v6.2.1 as implemented in R package v3.4.2. Significance level was assessed using the Bonferroni correction. RESULTS: A total of 18 SNPs were dropped due to low MAF, thus leaving 65 SNPs available for the analysis. Thus the Bonferroni threshold was 7.7×10-4 (0.05/65). Nominal significant association with NSCL/P was found at a SNP (rs4448343 in PTCH1, P=0.023) and six haplotypes (rs10512249-rs4448343, rs1461208-rs7786445, rs10512249-rs4448343, rs16909865-rs10512249-rs4448343, rs1461208-rs7786445-rs12698335, and rs288756-rs288758-rs1151790, P < 0.05). A total of six haplotypes (rs288765-rs1233563, rs12537550-rs11765352, rs872723-rs288765-rs1233563, rs288765-rs1233563-rs288756, rs6459952-rs12537550-rs11765352, and rs12537550-rs11765352-rs6971211) showed PoO effect (P < 0.05). None of the results remained significant after the Bonferroni correction (P>7.7×10-4). CONCLUSION: Neither significant association between SNPs within HH pathway and the risk of NSCL/P nor PoO effects was seen in this study.

[Evaluating the effect of WNT pathway genes considering interactions on the risk of non-syndromic oral clefts among Chinese populations].

OBJECTIVE: In this study, we used genome-wide association study (GWAS) data to explore whether WNT pathway genes were associated with non-syndromic oral clefts (NSOC) considering gene-gene interaction and gene-environment interaction. METHODS: We conducted the analysis using 806 non-syndromic cleft lip with or without cleft palate (NSCL/P) case-parent trios and 202 non-syndromic cleft palate (NSCP) case-parent trios among Chinese populations selected from an international consortium established for a GWAS of non-syndromic oral clefts. Genotype data and maternal environmental exposures were collected through DNA samples and questionnaires. Conditional Logistic regression models were adopted to explore gene-gene interaction and gene-environment in teraction using trio package in R software. The threshold of significance level was set as 3.47×10-4 using Bonferroni correction. RESULTS: A total of 144 single nucleotide polymorphisms (SNPs) in seven genes passed the quality control process in NSCL/P trios and NSCP trios, respectively. Totally six pairs of SNPs interactions showed statistically significant SNP-SNP interaction (P < 3.47×10-4) after Bonferroni correction, which were rs7618735 (WNT5A) and rs10848543 (WNT5B), rs631948 (WNT11) and rs556874 (WNT5A), and rs631948 (WNT11) and rs472631 (WNT5A) among NSCL/P trios; rs589149 (WNT11) and rs4765834 (WNT5B), rs1402704 (WNT11) and rs358792 (WNT5A), and rs1402704 (WNT11) and rs358793 (WNT5A) among NSCP trios, respectively. In addition, no significant result was found for gene-environment interaction analysis in both of the NSCL/P trios and NSCP trios. CONCLUSION: Though this study failed to detect significant association based on gene-environment interactions of seven WNT pathway genes and the risk of NSOC, WNT pathway genes may influence the risk of NSOC through potential gene-gene interaction.

[Exploring the association between SPRY gene family and non-syndromic oral clefts among Chinese populations using data of a next-generation sequencing study].

OBJECTIVE: To explore the association between SPRY gene family and the risk of non-syndromic oral clefts among Chinese populations, in respect of single nucleotide polymorphisms (SNPs) association and parent-of-origin effects. METHODS: Based on case-parent design, this study used the data of SPRY gene family in a next generation sequencing study of 183 non-syndromic cleft lip with or without cleft palate (NSCL/P) case-parent trios (549 participants) recruited from 2016 to 2018, to analyze the effects of SNP association and parent-of-origin. The sequencing study adopted a two-stage design. In the first stage, whole exome sequencing was conducted among 24 NSCL/P trios with family history to explore potential signals. Then in the second stage, another 159 NSCL/P trios were used as validation samples to verify the signals found in the first stage. The data of general information, disease features and parental environmental exposures for participants were collected through questionnaires. Blood samples were collected from each participant for DNA extraction and sequencing. Transmission disequilibrium tests (TDT) were conducted to test for the association between SNPs and NSCL/P, while Z score tests were applied to analyze parent-of-origin effects. The analyses were performed using Plink (v1.07). TRIO package in R (v3.5.1). Besides, famSKAT analyses were conducted in the first stage to combine the effect of SNPs located on the same gene, using famSKAT package in R(V3.5.1). Bonferroni method was adopted to correct multiple tests in the second stage. RESULTS: Twenty-two SNPs in SPRY gene family were included for analyses after the quality control process in the first stage. Based on the variants annotation, functional prediction and statistical analysis, rs1298215244 (SPRY1) and rs504122 (SPRY2) were included in the second verification stage. TDTs in the verification stage revealed that rs1298215244: T>C, rs504122: G>C and rs504122: G>T were associated with the risk of NSCL/P after Bonferroni corrections, where rs504122: G>T was a rare variation. Although the test for parent-of-origin effect of rs1298215244: T>C reached nominal significance level, no SNP showed significant association with NSCL/P through parent-of-origin effect after Bonferroni corrections. CONCLUSION: This study found that SNPs (including both common and rare variants) among the SPRY gene family were associated with the risk of NSCL/P among Chinese populations. This study failed to detect parent-of-origin effects among the SPRY gene family.

[The self-injury status and relevant factors of disabled children and adolescents in Beijing].

From September 2017 to February 2018, 650 disabled children and adolescents aged 6-to 17-year-old in Beijing were matched 1∶1 to those in normal physical condition with same age, gender and place of residence. All children and adolescents were investigated for self-injury status in the last year to understand the difference of self-injury incidence between groups. Multivariate unconditional logistic regression model was applied for exploring relevant factors of self-injury of children and adolescents. This study found that the disability, insufficient sleep, difficulty falling asleep, and sleeping late were associated with self-injury of children and adolescents.

Evidence of interaction between genes in the folate/homocysteine metabolic pathway in controlling risk of non-syndromic oral cleft.

OBJECTIVE: Little consistent evidence is available for the association between the risk of non-syndromic cleft lip with or without cleft palate (NSCL/P) and any of the individual genes in the folate/homocysteine metabolic pathway. We investigated the genes in the folate pathway to further clarify its potential influence on the risk of NSCL/P considering gene-gene (G×G) interaction. SUBJECTS AND METHODS: We selected markers in 18 genes from the pathway and applied Cordell's method to test for G×G interaction using 1,908 NSCL/P case-parent trios ascertained in an international consortium where a genomewide association study (GWAS) of oral clefts was conducted. RESULTS: We found intriguing signals among Asian and European ancestry groups for G×G interaction between markers in betaine-homocysteine methyltransferase gene (BHMT/BHMT2) and dimethylglycine dehydrogenase gene (DMGDH) attaining genomewide significance. In the pooled data, the top significant interaction was found between rs13158309 (BHMT) and rs10514154 (DMGDH, p = 1.45 × 10-12 ). CONCLUSIONS: Our study illustrated the importance of taking into account potential G×G interaction for genetic association analysis in NSCL/P, and this study suggested both BHMT/BHMT2 and DMGDH should be considered as candidate genes for NSCL/P in future studies.

Epidemiological investigation of a tap water-mediated hepatitis E virus genotype 4 outbreak in Zhejiang Province, China.

An outbreak of acute hepatitis recently occurred in a nursing home in Zhejiang Province, China. The objectives of this study were to confirm the outbreak and identify the aetiology, source and transmission patterns. All residents and staff in or near the nursing home during the period from 1 October 2014 to 21 May 2015 were investigated regarding hygiene and for epidemiological information including water and food (eating meat especially pork products). Serum and stool specimens were collected for detection of hepatitis E virus (HEV) antibodies using ELISA and RNA using RT-PCR. Samples that were RNA positive were genotyped. Of 185 senior residents and 24 staff in the nursing home, there were 37 laboratory-confirmed cases during the outbreak. Of these cases, 12 patients (three deaths) were symptomatic with jaundice, a common clinical symptom for hepatitis E infection. HEV strains were isolated from three cases and they formed a single cluster within genotype 4d. A case-control study was conducted to investigate potential risk factors for the outbreak and the results revealed that cases more often washed their dishes and rinsed their mouths using tap water than the controls (P < 0·05). Based on hygiene investigation and meteorological information, it is likely that HEV-infected sewage and faeces contaminated the water network on rainy days. Collectively, these results suggest that the outbreak of HEV genotype 4 infection was most likely caused by contaminated tap water rather than food.

Screening and identification of microsatellite markers associated with cold tolerance in Nile tilapia Oreochromis niloticus.

Tilapia is an important fish cultured in tropical and subtropical areas. Cold sensitivity limits the expansion of tilapia culture into colder regions of the world, and mass mortalities of cultured tilapia have been reported due to severe cold currents in winter. Since the late 1990s, several strains of Nile tilapia have been domesticated to improve the ability to adapt to low temperatures. Previous studies revealed that these varieties were more cold-tolerant than the founder population and overwintered naturally well in ponds in the west-south area of Guangdong Province. In this study, to develop tilapia strains with improved cold tolerance for breeding programs through marker-assisted selection, two microsatellite markers, UNH916 and UNH999, showed complete co-segregation with cold tolerance among the polymorphic microsatellite primers. Our results provide a foundation for identifying resistant gene(s) linked with these markers, as well as identifying simple sequence repeat markers associated with cold tolerance that can be used for maker-assisted selection programs in tilapia breeding to increase the growing range and productivity of tilapia aquaculture.

[Progress in research of risk prediction of non-syndromic oral clefts using genetic information].

Non-syndromic oral cleft (NSOC), a common birth defect, remains to be a critical public health problem in China. In the context of adjustment of childbearing policy for two times in China and the increase of pregnancy at older childbearing age, NSOC risk prediction will provide evidence for high-risk population identification and prenatal counseling. Genome-wide association study and second generation sequencing have identified multiple loci associated with NSOC, facilitating the development of genetic risk prediction of NSOC. Despite the marked progress, risk prediction models of NSOC still faces multiple challenges. This paper summarizes the recent progress in research of NSOC risk prediction models based on the results of extensive literature retrieval to provide some insights for the model development regarding research design, variable selection, model-build strategy and evaluation methods.

Numerical investigation on effect of particle aspect ratio on the dynamical behaviour of ellipsoidal particle flow.

Flow of ellipsoidal particles in a modal shear cell was investigated at the microdynamic level based on discrete element method simulations. In a stress-controlled double-shear condition, the flow was studied by varying the aspect ratio of ellipsoidal particles and comparing with the flow of spherical particle assembly in terms of some key properties, including particle alignment, linear velocity, angular velocity, porosity, contact force and contact energy. It was found that particle elongation impacts the rotational displacement around the axis perpendicular to the shear direction, which causes that the ellipsoidal particles with higher elongation are more aligned with the direction of the shear velocity, with more uniform force network. This then affects other particle properties. The fluctuation of linear velocity and the angular velocity decreases with an increase in particle aspect ratio, although the particle elongation does not significantly affect the flow velocity gradient. There is a reduction in both normal and tangential forces per contact with an increase of particle elongation. Due to the variation of the particle alignment with elongation, the standard deviation of the contact energies increases and then reduces when an increase in particle aspect ratio occurs, and on contrary, the porosity has an opposite variation trend.

[Correlation between immune reconstitution and chronic graft-versus-host disease after unrelated cord blood transplantation and sibling peripheral blood stem cell transplantation].

Objective: To explore the relationship between the reconstitution of immune cells in patients with hematological malignancies and the occurrence of chronic graft-versus-host disease (cGVHD) after treatment with unrelated cord blood transplantation (UCBT) and sibling peripheral blood stem cell transplantation (PBSCT) . Methods: A total of 124 patients undergoing allogenic hematopoietic stem cell transplantation (allo-HSCT) in the First Affiliated Hospital of University of Science and Technology of China from March 2018 to August 2019, including 96 patients with UCBT and 28 patients with PBSCT. Peripheral blood immune cells of patients with UCBT and PBSCT were detected at 1, 3, 6, 9, and 12 months after transplantation using flow cytometry, and both UCBT and PBSCT patients were divided into cGVHD and non-cGVHD groups based on whether cGVHD occurred to explore the correlation between the immune cells reconstitution of the two types of transplantation and cGVHD. Results: ①The cumulative incidence of the moderate to severe cGVHD in the UCBT group was significantly lower than that in the PBSCT group[9.38% (95% CI 3.35%-15.02%) vs 28.57% (95% CI 9.72%-43.50%) , P=0.008]; the 2-year cumulative incidence of cGVHD and moderate to severe cGVHD in the UCBT group was lower than that in the PBSCT group[15.60% (95% CI 9.20%-23.60%) vs 32.10% (95% CI 15.80%-49.70%) , P=0.047; 10.40% (95% CI 5.30%-17.50%) vs 28.60% (95% CI 13.30%-46.00%) , P=0.014]. ②The absolute counts of CD4(+)T cells in the UCBT group were higher than those in the PBSCT group at 6, 9, and 12 months after transplantation[59.00 (36.70-89.65) ×10(7)/L vs 31.40 (18.10-44.00) ×10(7)/L, P<0.001; 71.30 (49.60-101.45) ×10(7)/L vs 41.60 (25.82-56.27) ×10(7)/L, P<0.001; 83.00 (50.17-121.55) ×10(7)/L vs 44.85 (31.62-62.10) ×10(7)/L, P<0.001]; the proportions of CD4(+)T cells in the UCBT group were always higher than those in the PBSCT group (P<0.05) . The absolute counts and proportions of B cells in the PBSCT group were higher than those in the UCBT group at the first month after transplantation[0.70 (0.30-1.70) ×10(7)/L vs 0.10 (0-0.30) ×10(7)/L, P<0.001; 0.45% (0.30%-2.20%) vs 0.20% (0.10%-0.40%) , P=0.002]; the absolute counts and proportions of B cells in the UCBT group were higher than those in the PBSCT group at 9 and 12 months after transplantation[53.80 (28.00-103.20) ×10(7)/L vs 23.35 (5.07-35.00) ×10(7)/L, P<0.001; 21.45 (11.80-30.45) % vs 9.00% (3.08%-16.73%) , P<0.001. 66.70 (36.97-98.72) ×10(7)/L vs 20.85 (7.72-39.40) ×10(7)/L, P<0.001; 22.20% (14.93%-29.68%) vs 8.75% (5.80%-18.93%) , P<0.001]. The absolute counts and proportions of regulatory B (Breg) cells in the UCBT group were higher than those in the PBSCT group at 6, 9, and 12 months after transplantation[1.23 (0.38-3.52) ×10(7)/L vs 0.05 (0-0.84) ×10(7)/L, P<0.001; 5.35% (1.90%-12.20%) vs 1.45% (0-7.78%) , P=0.002. 2.25 (1.07-6.71) ×10(7)/L vs 0.12 (0-0.77) ×10(7)/L, P<0.001; 6.25% (2.00%-12.33%) vs 0.80% (0-5.25%) , P<0.001. 3.69 (0.83-8.66) ×10(7)/L vs 0.46 (0-0.93) ×10(7)/L, P<0.001; 6.15% (1.63%-11.75%) vs 1.40% (0.18%-5.85%) , P<0.001].The absolute counts and proportions of CD3(+)T cells, CD8(+)T cells, and Treg cells in the UCBT group were not significantly different from those in the PBSCT group. ③The absolute counts of B cells in the non-cGVHD group of UCBT patients were higher than those in the moderate to severe cGVHD group at 6 and 12 months after transplantation (P=0.038, P=0.043) ; the proportions of B cells in the non-cGVHD group were higher than those in the moderate to severe cGVHD group at 6 months after transplantation (P=0.049) . The absolute counts of Breg cells in the non-cGVHD group of patients with UCBT were higher than those in the moderate to severe cGVHD group at 6, 9, and 12 months after transplantation (P=0.006, P=0.028, P=0.050) ; the proportions of Breg cells in the non-cGVHD group were higher than those in the moderate to severe cGVHD group at 9 months after transplantation (P=0.038) . ④The absolute counts and proportions of B and Breg cells in the non-cGVHD group of patients with PBSCT were not statistically different than those in the moderate to severe cGVHD group. Conclusion: In the process of immune cell reconstitution, the Breg cells in the UCBT group were higher than those in the PBSCT group, and the Breg cells in the non-cGVHD group of the two types of transplantation were always higher than those in the moderate to severe cGVHD group, indicating that Breg cells can reduce the occurrence of cGVHD, revealing the possible reason for the lower incidence of cGVHD in the UCBT group.

[Progress in genetic epidemiology of non-syndromic cleft palate only].

One of the most common birth defects is cleft palate only (CPO) of which non-syndromic cleft palate only (NSCPO) accounts for 50%. NSCPO is a complex disease where multiple genes and environmental factors contribute to its risk. Unlike non-syndromic cleft lip with or without cleft palate (NSCL/P), previous genome-wide association studies only identified a few common genetic variations achieving genome-wide significance. This review summarizes the recent findings on genetic epidemiology of NSCPO. According to the current evidence, the candidate genes are divided into three categories: candidate genes with strong evidence, candidate genes with suggestive evidence, and candidate genes with inadequate evidence. The findings of epigenetic studies, the next generation sequencing studies, interaction analysis on NSCPO are also reviewed.

Silence of HOTAIR inhibits insulin secretion and proliferation in pancreatic ß cells.

OBJECTIVE: To examine the expression level of HOTAIR in pancreatic ß cells. Moreover, regulatory effects of HOTAIR on insulin secretion, proliferation, cell cycle, and apoptosis in ß cells are determined. MATERIALS AND METHODS: HOTAIR levels in mouse primary pancreatic cells and MIN6 cell line were determined by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Its level was significantly observed in MIN6 cells treated with different doses of glucose. After the knockdown of HOTAIR, insulin secretion, cell cycle distribution, proliferation, and apoptosis in pancreatic ß cells were assessed. RESULTS: HOTAIR was abundantly expressed in pancreatic islets. HOTAIR level in pancreatic tissues of db/db mice was downregulated and influenced by glucose level. Knockdown of HOTAIR attenuated insulin secretion and synthesis capacities in both MIN6 cells and primary pancreatic cells, which may be related by the downregulation of MafA, Pdx1, and NeuroD. Moreover, the silence of HOTAIR suppressed proliferation, arrested cell cycle, and stimulated apoptosis in pancreatic ß cells. CONCLUSIONS: HOTAIR is highly expressed in pancreatic tissues. The silence of HOTAIR inhibits insulin secretion by downregulating insulin transcription-related genes. In addition, the silence of HOTAIR suppresses proliferation, arrests cell cycle progression, and induces apoptosis in pancreatic ß cells.

[Genotype and evolution of hantavirus in Tiantai of Zhejiang province, 2011-2018].

Objective: By investigating the genotype and evolutionary variation of hantavirus (HV) in Tiantai county, a national surveillance site for hemorrhagic fever with renal syndrome (HFRS) was set in Zhejiang province, from 2011 to 2018, to reveal the molecular epidemiological characteristics of hantavirus (HV) in Tiantai. Methods: Total RNA was extracted from ultrasound treated HV antigen- positive rat lung samples in Tiantai from 2011 to 2018. After cDNA was prepared, nested PCR was used to amplify partial sequence of M fragments by using specific primers of HV. The sequences of HV in Tiantai from 2011 to 2018 were compared with other known HV sequences in order to identify the genotype and analyze the evolution and variation of the virus. Results: In 67 HV antigen-positive lung specimens, 31 were positive in nested PCR amplification with type-specific primers, including 30 Hantaan virus (HTNV) positive samples, 1 Seoul virus (SEOV) positive sample, and all the 31 samples were from Apodemus agrarius. The phylogenetic tree based on partial M segment was divided into monophyletic group, 30 strains were distributed in HTNV group and 1 was in SEOV group. The HTNV strain Tiantai T2018-130 was independently in one branch, sharing 84.8%-87.9% homology with other strains both at home and abroad, including 29 strains in HTNV group in Tiantai. The other 29 HTNV strains in Tiantai showed closer relationship. The SEOV strain T2016-31 from Apodemus agrarius showed closer relationship with previous strains of SEOV, Tiantai ZT71, ZT10 and Z37 strains of Wenzhou, Zhejiang province. Conclusions: HTNV, the main genotype of HV in Tiantai of Zhejiang province, showed obvious geographic clustering, but the strain T2018-130 was distinct from the others in Tiantai. Meanwhile, by sequence analysis, we confirmed that The SEOV strain T2016-31 existed in in Apodemus agrarius, indicating there was a phenomenon of "spillover" between virus and host in SEOV evolution.

[Study regarding the parent-of-origin effect of WNT pathway genes on non-syndromic cleft lip with or without cleft palate among the Chinese population].

Objective: Non-syndromic cleft lip with or without cleft palate (NSCL/P) is a common birth defect with its genetic evidence widely explored. This study explored the potential the parent-of-origin (PoO) effect of WNT pathway on the risks of NSCL/P, using a case-parent trio design. Methods: Data on the single nucleotide polymorphism (SNP) of WNT genes were selected from a genome-wide association study (GWAS). A total of 806 Chinese non-syndromic cleft lip patients, with or without cleft palate (NSCL/P) case-parent trios, were gathered from an international consortium. PoO effect of WNT pathway genes and its haplotypes were explored by log-linear models. Additional Wald tests were performed to assess: a) the heterogeneity of PoO effect between different maternal exposures, b) the interaction between PoO effect, c) maternal exposure to environmental tobacco smoke (ETS), and d) multivitamin supplementation during pregnancy. The threshold for statistical significance was adjusted as 3.47×10(-4), according to Bonferroni correction. Results: After quality control, a total of 144 SNPs within seven genes were included for analyses, among which 8 SNPs were of potential PoO effect (P<0.05). However, none of them achieved the statistical significance after Bonferroni correction. The haplotype rs4074668-rs12725747 (T-A) on WNT9A showed significant PoO effect, based on the haplotype test for PoO (P=2.74×10(-4)). In addition, no statistically significant interaction was found in further exploration of this haplotype under environmental exposures as ETS or multivitamin supplementation. Conclusions: Genes in the WNT pathway may influence the NSCL/P risks through the potential PoO effect. Particularly, the haplotype rs4074668-rs12725747 (T-A) on WNT9A presented significant PoO effect on NSCL/P, statistically. From this current study, findings on WNT pathway related risks among the NSCL/P, need to be further validated by independent samples in the future.

Field-induced magnetic transitions and strong anisotropy in α-CoV2O6 single crystal.

The Ising-like antiferromagnet α-CoV2O6 has received considerable interests because of stabilized 1/3 magnetization plateau around 5 K under magnetic field applied along magnetic easy c-axis. In this work, this magnetization plateau was studied by varying temperature or rotating magnetic field. As temperature decreased, this stabilized plateau collapsed, and additional magnetic transitions were observed. As a result, a rich magnetic phase diagram was constructed and extended to temperature lower than previously reported. When magnetic field moved from the c to b (or a) axis, the magnetization plateau developed with field directions and vanished finally when the field was restricted in the ab plane. An impressive observation is that this 1/3-plateau can be stabilized and remain robust even when magnetic field deviated from the c axis, accompanied by the evolutions of the magnetic moments and the critical transition fields. We suppose that the origins of these temperature and angular dependences of the 1/3 magnetization plateau are related to strong spin-orbital coupling. Indeed, electron spin resonance (ESR) measurement gives large Landé factor of 8.9, evidencing that there exists strong spin-orbital coupling.

[Progress in next-generation sequencing research of non-syndromic oral clefts].

Non-syndromic oral clefts (NSOC) are among the most common birth defects. The prevalence of NSOC is 1.13-1.30 per 1 000 live births in China, which is higher than those in other major ethnic groups. The etiology of NSOC is complex and heterogeneous, which involves both genetic and environmental risk factors. Although genome-wide association studies have identified a number of risk loci, these loci can only account for a small proportion of the heritability of NSOC. The next-generation sequencing research provides new ideas for further exploring the genetic risk factors of NSOC. This paper summaries the progress in the next-generation sequencing research of NSOC.

[Recombinant expression of hantaan virus protein N with application of Western-blot in detecting anti-hantavirus antibody].

Objective: S gene of hantavirus(HV) was expressed in insect cells by genetic engineering technology. The expression product of S gene was used as antigen to detect anti-HV specific antibody IgG in serum. Methods: Gene encoding NP of the strain HV-Z10 was amplified by PCR and then its eukaryotic expression system rBAC-Z10S-TN was constructed by using the routine genetic engineering method. SDS-PAGE was applied to measure the expression of rNP.Ion-exchange plus Ni-NTA-affinity chromatography was performed to purify the recombinant product. Indirect immuno-fluorescence assay (IFA) was used to determine the specific immune-reactivity of rNP. WB assay was established to detect the serum samples from 95 confirmed HFRS patients. Parameters related to the outcomes of detection were compared with the routine HV-IgG IFA method. Results: rBAC-Z10S-TN was able to express rNP with high efficiency. The purified rNP only showed a single protein fragment in the gel after SDS-PAGE. HV IgG could efficiently recognize rNP and hybridize with the recombinant protein. 97.67% of the serum samples from the HFRS patients were positive confirmed by WB. Conclusions: We successfully constructed a high efficient prokaryotic expression system of NP encoding gene from hantavirus strain HV-Z10. WB assay which was established in this study could be used as a new serological test for HFRS diagnosis, thanks to the simplicity, safety, sensitivity and specificity of this method.

Early blood purification therapy of severe acute pancreatitis complicated by acute lung injury.

OBJECTIVE: Severe acute pancreatitis (SAP) can often be complicated by acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), leading to increased mortality. Early blood purification clears inflammatory cytokines and promotes immune function recovery. Here we evaluated the usefulness of this therapy in SAP complicated by ALI. PATIENTS AND METHODS: 32 patients received routine treatment (control group), whereas other 32 patients received routine treatment and early blood purification therapy (study group). We evaluated respiratory indexes (PaO2, PaO2/FiO2, alveolar-arterial oxygen difference, intrapulmonary arteriovenous shunt percentage, and respiratory rate), blood biochemical (creatinine, blood urea nitrogen, alanine aminotransferase, and lactate levels) and inflammatory (CRP, IL-10, TNF-α, and IL-10/TNF-α ratio) markers, and prognostic outcomes (multiple organ dysfunction syndrome [MODS] and APACHE II scores) before and 72 hours after the treatment. We also documented mechanical ventilation use, occurrence of MODS and ARDS, and mortality rates. RESULTS: There were no deaths. Mechanical ventilation was used in a similar percentage of patients in either group. Treatment in study group led to a faster and better recovery of respiratory indexes, and less pronounced changes in the levels of blood urea nitrogen and alanine aminotransferase. Inflammatory markers also normalized better in the study group. Furthermore, MODS and APACHE II scores decreased to a greater extent in the study group, paralleled by a lower occurrence of MPDS and ARDS. CONCLUSIONS: Early blood purification therapy improves respiratory function and inflammatory markers in patients with SAP complicated by ALI, and decreases the occurrence of MODS and ARDS.