Cloning, sequencing, expression, and purification of aspartic proteases isolated from two human Demodex species.

Aspartic proteases (ASPs) are important hydrolases for parasitic invasion of host tissues or cells. This was the first study on Demodex ASP. First, the complete coding sequence (CDS) was amplified, cloned and sequenced. Then, the protein physical and chemical properties was analysed. Finally, the recombinant plasmid, expression and purification system was established. Results showed that the lengths of CDS of Demodex folliculorum and D. brevis were 1161 and 1173 bp, respectively. The molecular weight of the protein was approximately 40 KDa. It contained an aspartic acid residue, a substrate-binding site and signal peptide, yet lacked a transmembrane domain and was located in the membrane or extracellular matrix. The phylogenetic and conserved motif analyses showed that D. folliculorum and D. brevis clustered separately and then formed a single branch, which finally clustered with other Acariformes species. The prokaryotic expression systems for recombinant ASP with His-tag (rASP-His) and GST-tag (rASP-GST) were constructed. The inclusion bodies of rASP-His were renaturated by gradient urea and purified using NI beads, while those of rASP-GST were renaturated by sarkosyl and Triton X-100 and purified using GST beads. Conclusively, the prokaryotic expression and purification system of Demodex rASP was successfully established for further pathogenic mechanism research.

Classification and identification of mosquitoes in China based on rDNA 28S D5 region.

Accurate classification and identification of mosquitoes are essential for the prevention and control of mosquito-borne diseases. In this study, adult mosquitoes were collected from 15 cities across 14 provinces in China. They were identified morphologically with the dominant species determined. Furthermore, representative samples were identified at the molecular level based on rDNA 28S D5. In total, 880 adult mosquitoes were collected belonging to Culex (266), Aedes (473), Armigeres (13), and Anopheles (5). Aedes albopictus and "C. pipiens subgroup" were the dominant species. A total of 140 sequences of 28S D5 region (68 for "C. pipiens subgroup", 51 for Ae. albopictus, 18 for Ar. subalbatus, and three for An. sinensis) ranging from 148 to 161 bp were obtained, with 100 % success of amplification and sequencing. Molecular identification were consistent with morphological classification. Sequence analysis showed that "C. pipiens subgroup" was identified into three clades: the traditional C. pipiens subgroup (Clade I), the newly discovered C. cf. perexiguus (Clade II), and C. new sp. (Clade III). Clade I contained the most abundant haplotypes (16) widely distributed without geographical differences. Clade II included six haplotypes that were aggregately distributed south of the Yangtze River. Only three sequences in Clade III showed two haplotypes with no geographical differences. Further morphological comparisons demonstrated differences in body color, beaks, and abdomens among the three clades. In conclusion, the rDNA 28S D5 region could effectively distinguish Culex, Aedes, Armigeres, and Anopheles species at the lower category level, demonstrating its potential as a mini-DNA barcode for mosquito identification.

Analysis of pressure pulsation and energy characteristics of guide vane for axial flow pump based on Hilbert-Huang transform considering impeller-guide vane interaction.

In order to explore the characteristics of pressure pulsation signals and energy distribution of water flow at the guide vane considering impeller-guide vane interaction. The numerical simulation of the vertical axial flow pump device's steady and unsteady three-dimensional flow fields was carried out. The Hilbert-Huang method was used to conduct empirical mode decomposition decomposition and Hilbert spectrum analysis of pressure pulsation signal at each monitoring point in the inlet and outlet regions of the guide vane. The results show: Under the condition of 0.3Qbep, the internal pressure of the guide vane is obviously affected by the impeller, and there are large block-shaped vortex structures in the guide vane. Under the operating conditions of 1.0Qbep and 1.2Qbep, the size of the pressure area in the guide vane is basically not affected by the impeller, and the vortex structures in the guide vane are concentrated near the outlet of the guide vanes, and there are long strip-shaped vortex structures at the edge of the guide vane. The size and number of vortex structures decrease with the increase in flow rate. The pressure pulsation signal at the inlet of the guide vane is affected by the rotation of the impeller and exhibits good periodicity, with the main frequency centered around 146 Hz, and the energy ratio of the main frequency is up to 97.7%. There are low-frequency signals below 100 Hz and high-frequency signals fluctuating around 146 Hz in all three flow conditions. When the flow rate increases, the fluctuation amplitude of the high-frequency signal increases. The flow rate has a significant impact on the water flow at the outlet of the guide vane. At 0.3Qbep, its frequency is distributed in the range of 0-500 Hz, mainly concentrated in the area below 400 Hz. At 1.0Qbep, the frequency of pressure pulsation is distributed below 250 Hz after the guiding function of the guide vane. At 1.2Qbep, the water flow is mainly controlled by the rotation of the impeller, and after the energy recovery of the guide vane, its main frequency is still concentrated around 150 Hz, which is 337.2% and 268.5% of 0.3Qbep and 1.0Qbep. Under the working condition of 0.3Qbep, the proportion of intrinsic mode function energy corresponding to the dominant frequency at the center of the guide vane inlet is as high as 95.9%, and the proportion of intrinsic mode function energy corresponding to the dominant frequency at the shroud side and hub side of the guide vane is rather low. If the flow rate rises from 0.3Qbep to 1.2Qbep, the proportion of intrinsic mode function energy increases by more than 42%. Under the working conditions of 0.3Qbep and 1.0Qbep, the main frequency of pressure pulsation signal of water flow at the guide vane outlet is less affected by the impeller and the corresponding energy proportion is low. Under the working condition of 1.2Qbep, the main frequency of pressure pulsation signal is 4 times the rotational frequency and the corresponding energy proportion is higher than 60%.

[Cervical spondylosis of nerve root type with qi stagnation and blood stasis treated with warming needle with different lengths of moxa stick: a randomized controlled trial].

OBJECTIVE: To compare the clinical efficacy on cervical spondylosis of nerve root type with qi stagnation and blood stasis treated with warming needle with different lengths of moxa stick. METHODS: Six hundred patients with cervical spondylosis of nerve root type with qi stagnation and blood stasis were randomly divided into 4 groups: a 4 cm length group (150 cases, 5 cases dropped off, 2 cases suspended), a 3 cm length group (150 cases, 6 cases dropped off, 2 cases suspended), a 2 cm length group (150 cases, 6 cases dropped off), and a routine acupuncture group (150 cases, 6 cases dropped off). Warming needle with moxa stick in the length of 4 cm, 3 cm and 2 cm was delivered in the 4 cm length group, the 3 cm length group and the 2 cm length group, respectively. In the routine acupuncture group, simple acupuncture was applied. The acupoints selected in the above groups included Dazhui (GV 14) and bilateral Jiaji (EX-B 2) of C5 and C7, Fengchi (GB 20), Jianzhen (SI 9), Quchi (LI 11), Zhongzhu (TE 3), etc. In each group, the intervention was delivered once daily and 5 times a week. One course of intervention was composed of 2 weeks and 2 courses were required. The TCM syndrome score, the score of clinical assessment scale for cervical spondylosis (CASCS), the score of the brachial plexus traction test of the affected upper limb, F wave occurrence rate and conduction velocity of the ulnar nerve, the median nerve and the radial nerve of the affected upper limb were compared before and after treatment in the patients of each group. The levels of serum inflammatory factors, i.e. interleukin-1ß (IL-1ß), interleukin-6 (IL-6), tumor necrosis factor α (TNF-α) and hypersensitive C-reactive protein (hs-CRP), were measured before and after treatment in the patients of each group. The clinical cfficacy was evaluated in the 4 groups. RESULTS: After treatment, the results of TCM syndrome evaluation, i.e. the scores of neck pain, activity limitation and upper limb numbness and pain, as well as the total scores; and the scores of brachial plexus traction test were reduced when compared with those before treatment in each group (P<0.01, P<0.05). The scores of subjective symptoms and adaptability, and the total scores of CASCS were elevated in comparison with those before treatment in each group (P<0.01, P<0.05). In the 4 cm length group, compared with the other 3 groups, the scores of neck pain and activity limitation for TCM syndrome evaluation, and its total score were lower (P<0.05, P<0.01); and the scores of subjective symptoms and adaptability, and the total score of CASCS were higher (P<0.05, P<0.01). The score of the brachial plexus traction test in the 4 cm length group was lower than that of the routine acupuncture group (P<0.05). After treatment, F wave occurrence rates and conduction velocity of median nerve and radial nerve were increased when compared with those before treatment in each group (P<0.05, P<0.01). F wave occurrence rate and conduction velocity of the radial nerve in the 4 cm length group were higher than those of the other 3 groups (P<0.05), and those of the median nerve were higher when compared with the routine acupuncture group (P<0.05). After treatment, the levels of serum IL-1ß, IL-6 and TNF-α were all reduced when compared with those before treatment in each group (P<0.01, P<0.05); the level of serum IL-6 in the 4 cm length group was lower than those of the other 3 groups and serum level of TNF-α was lower compared with that in the routine acupuncture group (P<0.05). The total effective rate of the 4 cm length group was 78.3% (112/143), which was higher when compared with the 3 cm length group (67.6%, 96/142), the 2 cm length group (65.3%, 94/144) and the routine acupuncture group (53.5%, 77/144), respectively (P<0.05). CONCLUSION: Warming needle with moxa stick of 4 cm in length effectively relieves the clinical symptoms of cervical spondylosis of nerve root type with qi stagnation and blood stasis, improves the nerve function of the upper limbs, and reduces the inflammatory responses caused by nerve compression. The clinical efficacy of this therapy with moxa stick of 4 cm in length is superior to the warming needle with moxa sticks of 3 cm and 2 cm, as well as the routine acupuncture.

[Genetic analysis of a Chinese pedigree with Lesch-Nyhan syndrome].

OBJECTIVE: To explore the genetic etiology for a Chinese pedigree affected with Lesch-Nyhan syndrome. METHODS: Members of the pedigree who had visited the Genetic Counseling Clinic of Linyi People's Hospital on February 10, 2022 were selected as the study subjects. Clinical data and family history of the proband were collected, and trio-whole exome sequencing (trio-WES) was carried out for the proband and his parents. Candidate variants were verified by Sanger sequencing. RESULTS: Trio-WES revealed that both the proband and his cousin brother had harbored a hemizygous c.385-1G>C variant in intron 4 of the HPRT1 gene, which was unreported previously. A heterozygous c.385-1G>C variant of the HPRT1 gene was also found in the proband's mother, grandmother, two aunts, and a female cousin, whilst all phenotypically normal males in his pedigree were found to have a wild type for the locus, which has conformed to an X-linked recessive inheritance. CONCLUSION: The heterozygous c.385-1G>C variant of the HPRT1 gene probably underlay the Lesch-Nyhan syndrome in this pedigree.

Benign prostatic hyperplasia associated with white matter hyperintensities in men.

PURPOSE: Benign prostatic hyperplasia (BPH) describes common noncancerous prostate enlargement. BPH is usually associated with lower urinary tract symptoms and an increased risk of cerebrovascular diseases, such as stroke and its recurrence. White matter hyperintensities (WMHs), markers of cerebral injury, increase the risk of stroke, cognitive impairment, dementia, and death. The relationship between BPH and WMHs remains unclear. This study aimed to determine the association between BPH and WMHs. METHODS: A total of 788 male patients from the First Affiliated Hospital of Kunming Medical University from July 2019 to September 2021 were enrolled in this cross-sectional study. BPH was assessed by abdominal ultrasound, and three independent neuroradiologists rated the presence or absence of WMHs. Multiple imputations of chained equations were used to handle missing data. Logistic regression was used to assess the relationship between BPH and WMHs. RESULTS: Patients with BPH presented an increased risk of WMHs with a crude odds ratio (OR) of 2.76 (95% CI, 2.02-3.79) and an adjusted OR of 1.75 (95% CI, 1.24-2.48) after controlling for potential confounding factors in the multivariate logistic regression. CONCLUSION: We found that BPH was closely associated with WMHs in male Chinese individuals.

Visual Sorting of Express Packages Based on the Multi-Dimensional Fusion Method under Complex Logistics Sorting.

Visual sorting of express packages is faced with many problems such as the various types, complex status, and the changeable detection environment, resulting in low sorting efficiency. In order to improve the sorting efficiency of packages under complex logistics sorting, a multi-dimensional fusion method (MDFM) for visual sorting in actual complex scenes is proposed. In MDFM, the Mask R-CNN is designed and applied to detect and recognize different kinds of express packages in complex scenes. Combined with the boundary information of 2D instance segmentation from Mask R-CNN, the 3D point cloud data of grasping surface is accurately filtered and fitted to determining the optimal grasping position and sorting vector. The images of box, bag, and envelope, which are the most common types of express packages in logistics transportation, are collected and the dataset is made. The experiments with Mask R-CNN and robot sorting were carried out. The results show that Mask R-CNN achieves better results in object detection and instance segmentation on the express packages, and the robot sorting success rate by the MDFM reaches 97.2%, improving 2.9, 7.5, and 8.0 percentage points, respectively, compared to baseline methods. The MDFM is suitable for complex and diverse actual logistics sorting scenes, and improves the efficiency of logistics sorting, which has great application value.

[Association of gene expression in ovarian granulosa cells with embryonic development potential among patients with polycystic ovary syndrome].

OBJECTIVE: To assess the association of gene expression with development potential of early embryos derived from patients with polycystic ovary syndrome (PCOS). METHODS: Three pairs of infertile patients with respectively matched age, body mass index, ovarian reserve and treatment with gonadotrophin-releasing hormone (GnRH) antagonists were selected. Patients with fewer embryos were assigned as the case group (n = 3), whilst the remainders were assigned as the control group (n = 3). Ovarian granulosa cells from patients were collected for the extraction of total RNA and subjected to RNA sequencing. The results were subjected to differential gene expression and functional enrichment analyses. RESULTS: Compared with the control group, 76 genes were up-regulated and 110 genes were down-regulated in the case group. The level of estradiol (E2) was significantly higher in the control group on the trigger day with the injection of human chorionic gonadotrophin (HCG). Compared with the control group, the KRT7 gene was most significantly up-regulated, whilst the CCNYL2 gene was most significantly down-regulated in the case group. Gene ontology (GO) entries enrichment has found those associated with chromosome segregation, cell cycle regulation, and fatty acid metabolism to be significantly enriched. The genes participating in the regulation of cell assembly, differentiation, negative regulation of cell cycle, negative regulation of development, extracellular regulated protein kinases (ERK), ERK1 and ERK2 signaling pathways to be significantly down-regulated. KEGG enrichment analysis of cell signaling pathways revealed that steroid hormone biosynthesis-related genes were enriched. CONCLUSION: Among patients treated with GnRH antagonists, the significant difference in the number of oocytes fertilized in vitro and the number of available embryos are associated with the difference in the expression of genes of ovarian granulosa cells.

Glomerular filtration Rate, urine Albumin/ creatinine ratio and current perception threshold in patients with diabetic kidney disease.

BACKGROUND: Diabetic microvascular complications, including diabetic kidney disease (DKD), retinopathy (DR), and neuropathy (DN), were major causes of morbidity and mortality in diabetic patients worldwide. It has been suggested that urinary albumin-to-creatinine ratio (UACR) and estimated glomerular filtration rate (eGFR) were not the only indicators of renal function impairment in DKD and that they were also associated with diabetic peripheral neuropathy (DPN) which might affect nerve conduction velocity (NCV). As 30-40% of DPN patients had no subjective symptoms, while current perception threshold (CPT) could detect sensory nerve damage at an early stage. As a result, we aimed to investigate correlation between UACR, eGFR and CPT in DKD patients. METHODS: A total of 273 DKD patients from the First Affiliated Hospital of Kunming Medical University from January 2018 to June 2020 were enrolled to complete the CPT test. CPT values of the bilateral median nerve and superficial and deep peroneal nerves at 2000 Hz, 250 Hz, and 5 Hz were collected. RESULTS: In normoesthesia and hypaesthesia patients with DKD, MDRD-eGFR correlated negatively with TC (r = -0.135, P = 0.037), left superficial peroneal and deep peroneal nerve 2000 Hz CPT (r = -0.205, P = 0.001) and right superficial peroneal and deep peroneal nerve 2000 Hz CPT (r = -0.154, P = 0.017). Besides, left and right superficial peroneal and deep peroneal nerve 2000 Hz CPT correlated with CKD-EPI-eGFR and UACR. Multivariate logistic regression analysis found left superficial peroneal and deep peroneal nerve 2000 Hz CPT was independently associated with both MDRD-eGFR and CKD-EPI-eGFR. CONCLUSION: Decreased MDRD-eGFR and CKD-EPI-eGFR were expected to be a predictor of peripheral nerve injury in normoesthesia and hypaesthesia patients with DKD.

Carbon Isotope Fractionation Characteristics of Normally Pressured Shale Gas from the Southeastern Margin of the Sichuan Basin; Insights into Shale Gas Storage Mechanisms.

Since the development of shale gas in the Wufeng-Longmaxi Formation in the Sichuan Basin, China's shale gas production and reserves have increased rapidly. The southeastern margin of the Sichuan Basin is located in a normally pressured transition zone, where single well gas production varies greatly under complex geological structures. In order to reveal the shale gas enrichment mechanism and favorable shale gas regions, shale gas samples from production wells were collected from different structures, with the formation pressure coefficient ranging between 0.98 and 1.35. The gas components and carbon isotope characteristics of normally pressured shale gas were investigated. The carbon isotope characteristics of the Wufeng-Longmaxi shale gas from the basin scale was mainly controlled using thermal maturity; as the thermal maturity increased, heavier carbon isotopes were found, in addition to drier shale gas. For normally pressured shale gas, the composition of δ13C1 and δ13C2 becomes heavier, and the dryness coefficient decreases with the decreasing pressure coefficient; this is not consistent with the results from thermal evolution. By comparing possible influencing factors, it is evident that the change in geological structure destroys the original shale gas reservoir, which leads to the escape of some gases, and it may be the main factor that contributes to the gas geochemical characteristics of the normally pressured shale gas. Compared with the geological parameters of the shale samples, such as mineral composition, organic abundance, organic pore distribution, and gas content, the carbon isotope characteristics of normally pressured shale gas show a higher efficiency, thus indicating favorable sweet spot evaluations for shale gas in the studied areas.

Analysis of Differentially Expressed Novel MicroRNAs as Potential Biomarkers in Dengue Virus Type 1 Infection.

BACKGROUND: Host-derived miRNAs are reported to play diverse roles in dengue virus (DENV) infection, but DENV-derived miRNAs have been rarely studied. METHODS: Here, we used serum samples from three patients infected with dengue virus type 1 (DENV1) and three healthy volunteers to detect and profile novel microRNAs in dengue virus-infected human serum. MicroRNAs in serum samples were sequenced using an Illumina HiSeq 2000 system, and clean reads were trimmed, aligned, normalized, and analyzed to identify differentially expressed and novel microRNAs. Four microRNAs were selected as DENV1 infection biomarkers and verified through 1:2 paired case-control study, using serum from 15 DENV1-infected patients and 30 healthy volunteers. RESULTS: We identified 182 potential novel miRNAs, with 20 novel miRNAs found to have miRDeep2 score ≥ 4.0. Fifty-eight known and 11 novel miRNAs were upregulated at least 2-fold in DENV1-infected serum. Twenty-two known and 4 novel miRNAs were downregulated at least 0.5-fold. The AUCs of four selected miRNAs, hsa-miR-106b-3p, hsa-miR-122-5p, hsa-miR-novel-chr17_35150, and hsa-miR-novel-chr10_24390, were respectively: 0.962 (95% CI: 0.913 - 1.000, p > 0.05), 0.924 (95% CI: 0.851 - 0.998, p < 0.05), 0.941 (95% CI:0.877 - 1.000, p > 0.05), and 0.991 (95% CI: 0.973 - 1.000, p > 0.05). CONCLUSIONS: In summary, our study demonstrates that serum miRNA levels are affected by DENV1 infection, identifies novel DENV1-associated miRNAs, and suggests that Hsa-miR-122-5p may be a potential biomarker for DENV1 infection.

Travel-related infections in mainland China, 2014-16: an active surveillance study.

BACKGROUND: Transmission of infection through international travel is a growing health issue, and the frequency of imported infection is increasing in China. We aimed to quantify the total number of infections imported into mainland China by arriving travellers. METHODS: We actively surveyed arriving travellers at all 272 international entry-exit ports in mainland China. Suspected cases were detected through fever screening, medical inspection, self-declaration, and reporting by on-board staff. Participants completed a standardised questionnaire with questions about demographics, their travel itinerary (including detailed information about all countries or regions visited), and clinical manifestations. Nasopharyngeal swabs, sputum samples, faecal samples, vomitus, blood, and serum were collected as appropriate for diagnoses. Diagnosis was made by specific laboratory tests according to the national technical guidelines. Infections were classified as respiratory, gastrointestinal, vector-borne, blood-transmitted and sex-transmitted, or mucocutaneous. We divided arriving travellers into two groups: travellers coming from countries other than China, and travellers coming from Hong Kong, Macau, and Taiwan. We integrated surveillance data for 2014-16, calculated incidences of travel-related infections, and compared the frequency of infections among subgroups. FINDINGS: Between Jan 1, 2014, and Dec 31, 2016, 22 797 cases were identified among 805 993 392 arriving travellers-an overall incidence of 28·3 per million. 45 pathogens were detected in participants: 18 respiratory (19 662 cases), ten gastrointestinal (189 cases), seven vector-borne (831 cases), seven blood-transmitted and sex-transmitted (1531 cases), and three mucocutaneous (584 cases). Both the overall number and incidence of infection were more than five times higher in 2016 than in 2014. Case numbers and incidences also varied substantially by province, autonomous region, and municipality. Overall, 17 643 (77%) infections were detected by fever screening, but 753 (49%) blood-transmitted and sex-transmitted infections were identified through medical inspection. 14 305 (73%) cases of respiratory infection and 96 (51%) of gastrointestinal infections were in tourists. Tuberculosis, hepatitis A virus, vector-borne, and blood-transmitted and sex-transmitted infections were common among Chinese labourers who worked abroad. Dengue and malaria were most commonly diagnosed in travellers arriving from Africa. 12 126 (93%) of the 12 985 cases arriving from Hong Kong, Macau, or Taiwan were respiratory infections. Hand, foot, and mouth disease accounted for 2·90% of infections in travellers from Hong Kong, Macau, or Taiwan and 0·31% of infections in international travellers. INTERPRETATION: This report is the first to characterise the profile of travel-related infections among arriving travellers in mainland China. Our findings should increase public awareness of the potential risk of imported infections, and help health-care providers to make evidence-based health recommendations to travellers. FUNDING: The Natural Science Foundation of China.

[Study on the types and water pollution driving forces of the typical and medium-small-sized cities in the southern China based on the analysis of water environment].

According to the major pollution sources of urban water environment, 10 indexes such as industrial sewage quantity were closen to establish evaluation indexes system about the types and influencing factors of the typical and medium-small-sized cities in the southern China. Case studies of 16 typical and medium-small-sized cities were taken in Jiangsu, Zhejiang, Hubei and Anhui provinces. Combined with SPSS 11.0 cluster analysis results, city types were divided in reference to the values of water resources comprehensive pollution indexes and economical development indexes. The driving forces about city water environment pollution were studied by principal component analysis method. The result indicates that the 16 cities belong to two categories and four sub-categories, which are rich economy as well as light pollution of water environment and poor economy as well as heavy pollution of water environment. The influencing factors of water environment pollution are in sequence of industrial water pollution, agricultural no-point source pollution and urban domestic water pollution. The main factors of water environment pollution influenced I category cities, II as well as IV category cities and III category cities are industrial water pollution, urban domestic pollution and agricultural no-point source pollution respectively.