[Epidemiological characteristics of tick-borne infectious diseases in Hubei Province, 2016-2021].

Objective: To understand the epidemiological characteristics of tick-borne infectious diseases (TBID) and the risk factors of severe illness and death in Hubei Province from 2016 to 2021. Methods: Based on the incidence data of fever with thrombocytopenia syndrome (SFTS), tsutsugamushi disease, typhus and other TBID reported during 2016-2021, the epidemiological analysis was conducted. Field investigation results of TBID in areas with high incidence in 2021, logistic regression analysis of population characteristics, epidemiological history and other factors were used to explore the risk factors of severe and fatal cases. In the field vector investigation, free ticks and surface ticks of the host animals in the cases' home and surrounding grassland were monitored and detected. Results: A total of 3 826 TBID cases were reported in Hubei from 2016 to 2021, of which 71.30% (2 728/3 826) were SFTS, 13.04% (499/3 826) were tsutsugamushi disease and 15.66% (599/3 826) were typhus. A total of 44 cases died in 6 years; the fatality rate was 1.15% (44/3 826). In the peak seasons of incidence from May to July, the cases in people engaged in agriculture related work accounted for 84.61% (3 237/3 826). The incidence rate in women was higher than that in men, and the cases aged ≥50 years accounted for 81.02% of the total (3 100/3 826), and the incidence rate increased with age (P<0.001). The TBID cases were distributed in 86 counties and districts in 16 prefectures (municipality). The incidence rates of different areas had significant differences (P<0.05), and there was a certain spatial-temporal clustering and expasion. Bovis microplus and Haemaphysalis longicornis were captured in the field, and the positive rates in host animals and grassland ticks were 10.94% (7/64) and 40.00% (2/5), respectively. Univariate logistic regression analysis results showed that age ≥50 years and leukocyte <2.0×109/L were risk factors for severe illness and death. Conclusions: The TBID reported in Hubei were mainly SFTS, tsutsugamushi disease and typhus. In order to reduce the incidence of TBID, it is necessary to strengthen the prevention and control in women aged ≥50 years and reduce field exposure and tick bites during the epidemic period.

[Hand, foot and mouth disease in Hubei province, 2009-2015: an epidemiological and etiological study].

Objective: To clarify the age patterns and types of differences so as to provide reference on prevention and interventions of hand, foot and mouth disease (HFMD) cases, in Hubei province. Methods: We collected the HFMD case information of Hubei province from the Chinese National Notifiable Infectious Disease Reporting System in 2009-2015 while the information on pathogens from the laboratory monitoring system of Center for Disease Control and Prevention at all levels in Hubei province. All the data were stratified by age, disease severity, laboratory confirmation status, and serotypes of enterovirus. Results: There were 495 783 reported HFMD cases from 2009 to 2015, in Hubei province, of which 1 045 were severe with 99 fatal. The annual notification rate was 1 231.0/10(6). HFMD cases were concentrated mainly in 0.5-5 year olds, with highest severity and mortality seen in 6-11 month-olds. The predominated pathogen in mild laboratory-confirmed cases each year, in order during 2009-2015 as: EV71, Cox A16, Cox A16, Cox A16, EV71, Cox A16 and other EV. HFMD showed semiannual peaks in April-June, November-December, and with more cases in the even years than in the odd years. Conclusions: Children aged 0.5 to 5 years with 6 to 11 month-olds in particular, were the focused groups of attention in Hubei province. Our findings provided evidence for the improvement on monitoring program. Targeted intervention approaches should be strengthened to reduce the mortality and morbidity of HFMD in the province.

[Risk factors for lower extremity amputation in patients with diabetic foot].

Objective: To explore the risk factors for lower extremity amputation in patients with diabetic foot. Methods: The clinical data of 1 771 patients with diabetic foot at the Air Force General Hospital of PLA from November 2001 to April 2015 were retrospectively analyzed. The patients were divided into the non-amputation and amputation groups. Within the amputation group, subjects were further divided into the minor and major amputation subgroups. Binary logistic regression analyses were used to assess the association between risk factors and lower extremity amputation. Results: Among 1 771 patients with diabetic foot, 323 of them (18.24%) were in the amputation group (major amputation: 41; minor amputation: 282) and 1 448 (81.76%) in the non-amputation group. Compared with non-amputation patients, those in the amputation group had a longer hospital stay and higher estimated glomerular filtration rate(eGFR)levels. Fasting plasma glucose (FPG), glycosylated hemoglobin (HbA1c), C-reaction protein (CRP), ESR, ferritin, fibrinogen and WBC levels of the amputation group were higher, while hemoglobin albumin, transferrin, TC, TG, HDL-C and LDL-C were lower than those of the non-amputation group (all P<0.05). The proportion of hypertension(52.48% vs 59.98%), peripheral vascular disease (PAD)(68.11% vs 25.04%), and coronary heart disease(21.33% vs 28.71%)were different between the amputation and non-amputation groups (all P<0.05). Multivariable logistic regression analyses showed that Wagner's grade, PAD and CRP were the independent risk factors associated with lower extremity amputation in hospitalized patients with diabetic foot. Conclusion: Wagner's grade, ischemia of lower limbs and infection are closely associated with amputation of diabetic foot patients.

Insilico Approaches in Anesthetic Drug Development: Computer Aided Drug Designing.

OBJECTIVE: Computer Aided Drug DESIGNing is fast becoming an important tool in Drug discovery, and in the field of anesthetic drug development we are the first to use in silico approaches to look for novel anesthetic compounds. DESIGN: The approach of molecular modeling, Virtual screening, Drug-likeness, molecular docking and molecular dynamics simulations (MDS) was employed for this study. RESULT: Our approach of virtual screening Drug-likeness, adsorption, distribution, metabolism, excretion and toxicity analysis of around 50 000 compounds from Inter Bio Screen (IBS) Database have given us top 5 Lead compounds against ASN289 of γ-aminobutyric acid (GABAA) receptor, a common target of known anesthetic compounds. Out of the top 5 Lead compounds one (Lead 5) was selected for further MDS analysis based on its Binding free energy and number of physical interactions with GABAA. CONCLUSION: The MDS analysis of Lead 5 reveals the complex to be stable and thus suitable for further in vitro and in vivo analysis.

The Imbalance of Th17 cells and CD4(+) CD25(high) Foxp3(+) Treg cells in patients with atopic dermatitis.

BACKGROUND: Th17/Treg imbalance is involved in several autoimmune, inflammatory and allergic reactions. Nevertheless, the possible contribution of Th17/Treg imbalance in atopic dermatitis (AD) remains unknown. OBJECTIVE: To explore the possible role of Th17/Treg imbalance in AD. METHODS: Th17 and Treg cells percentage in peripheral blood mononuclear cells (PBMCs) and skin specimens, specific transcription factor retinoic acid-related orphan receptor (ROR)γt and Foxp3 mRNA levels in PBMCs, as well as Th17- and Treg-related cytokines mRNA levels in PBMCs, serum concentrations, and expression levels in PBMCs culture supernatant after recombinant Dermatophagoides pteronyssinus antigen stimulation were detected in AD patients. Controls included patients with psoriasis, allergic contact dermatitis (ACD) and healthy donors. RESULTS: Th17 cells percentage, RORγt, IL-17 and IL-23 levels in peripheral circulation of AD patients were significantly higher than those in ACD patients and healthy controls, but lower than those of psoriasis patients. Treg cells percentage, Foxp3 and TGF-ß mRNA levels were reduced in AD patients compared with healthy controls, while there were no significant differences among AD, ACD and psoriasis patients. Th17 cells percentage, IL-17 and IL-23 levels were increased, while Treg cells percentage and TGF-ß level were decreased in AD lesion and PBMCs culture supernatant respectively. There was a negative association between Th17 and Treg cells percentage in AD patients. AD severity score positively correlated with Th17 cells percentage and Th17/Treg ratio, while negatively correlated with Treg cells percentage. Serum IgE levels positively correlated with Th17/Treg ratio. CONCLUSION: In AD, there exists an immune imbalance in Th17 and Treg cells, which may contribute to its pathogenesis and development.

Possible pathogenic role of T helper type 9 cells and interleukin (IL)-9 in atopic dermatitis.

T helper type 9 (Th9) cells are a novel identified subset of CD4(+) T helper cells, which could partly contribute to allergic inflammation, while the precise contribution of Th9 cells in atopic dermatitis (AD) remains unknown. We aimed to explore the possible role of Th9 cells in AD pathogenesis. The Th9 cell percentage, transcription factor PU.1 and cytokine interleukin (IL)-9 mRNA levels, as well as IL-9 serum concentration in peripheral circulation, were measured in AD patients, psoriasis patients and healthy controls. The Th9 cell percentage, PU.1 and IL-9 expression levels of AD patients were all increased significantly compared with the other two control groups (P < 0·01), and correlated positively with SCORing Atopic Dermatitis index, serum immunoglobulin (Ig)E and thymus- and activation-regulated chemokine (TARC) levels (P < 0·05). In simple AD patients and AD patients complicated by allergic rhinitis or asthma, there were no significant differences in the Th9 cell percentage, PU.1 and IL-9 expression levels between them. At the same time, IL-9 and vascular endothelial growth factor (VEGF) mRNA levels were detected in AD lesions and normal skin samples, which were both distinctly elevated in AD lesions, and there was a positive association between them (P < 0·01). Keratinocytes were cultured with IL-9 stimulation and the secretion of VEGF was detected. IL-9 can promote the secretion of VEGF by keratinocytes in a time- and dose-dependent manner. In conclusion, the expansion of the Th9 cell subset, up-regulation of the PU.1 transcription factor and increased secretion of the IL-9 cytokine may contribute to the pathogenesis of AD, which may be supported by the increased release of VEGF by keratinocyes after IL-9 stimulation.

Association of variable number of tandem repeat polymorphism in the IL-4 gene with ischemic stroke in the Chinese Uyghur population.

Ischemic stroke (IS) is a multifactorial disorder, and genetic factors act as important contributors to its onset and progression. Inflammation is a key event that is closely associated with the pathophysiology of IS. The association of genetic polymorphisms of inflammatory cytokines with IS remains poorly understood. We investigated the relationship between the variable number of tandem repeats (VNTR) for IL-4, which is an important biomarker of inflammation, and the risk of IS. To assess the nature of the VNTR polymorphism in IL-4 and identify any links with IS, we recruited 200 subjects from a unique population that has 60% European and 40% East Asian ancestry. The subjects comprised 100 IS patients diagnosed using magnetic resonance imaging within 24 h of symptom onset and 100 age-, gender- and ethnicity-matched normal healthy controls. VNTR was identified using high-performance capillary electrophoresis with specially designed tailed primers. The IL-4 VNTR polymorphism was significantly associated with IS after adjustment for cardiovascular risk factors (OR = 0.571, 95%CI = 0.330-0.949, P < 0.05). Our data indicate that IL-4 VNTR polymorphism may affect susceptibility to IS in the Chinese Uyghur population. Moreover, total cholesterol, fasting blood glucose, waist-to-hip ratio, hypertension, history of heart diseases, and negative events may increase the risk of IS, with a trend for HDL to be a protective factor for IS in the Uyghur population.

[Gene cloning, construction and expression of single-chain Fv (scFv) against the membrane protein of Schistosoma japonicum].

OBJECTIVE: To construct single chain antibody specific to membrane protein of Schistosoma japonicum by genetic engineering technique. METHODS: The VH (heavy-chain variable region) and VL(light-chain variable region) genes were amplified by PCR from the genomic DNA of NP11-4 cell line, and sequenced by Sanger's method. The ScFv was constructed in pTHA90 vector using VH and VL genes, then expressed by IPTG. RESULTS: The VH and VL genes were obtained through PCR. The DNA sequences showed that VH and VL were new variable region genes of antibody. They were registered by GenBank. A ScFv gene with (Gly4Ser) 3 intralinker in the pTHA90 vector was successfully constructed. The ScFv was expressed as thioredoxin-fused proteins about 36.2 kDa. CONCLUSION: A specific ScFv against the membrane protein of Schistosoma japonicum was constructed and expressed.

[Screening the mimic antigen epitopes of triosephosphate isomerase of Schistosoma japonicum Chinese strain (Sjc-Tpi) with random phage peptide library].

OBJECTIVE: To screen the mimic antigen epitopes of the triose phosphate isomerase of Schistosoma japonicum Chinese strain (SjC-TPI) and investigate their immunogenicity. METHODS: The random phage peptide library (PH.D.-12) was screened with the purified antibody(IgG) against SjC-TPI to get the positive phage which contained the mimic antigen epitopes of SjC-TPI, and the immuno-characterization of the mimic antigen epitopes were investigated. RESULTS: Two mimic antigen epitopes (M1, M2) of SjC-TPI were obtained. The immuno-sera of mice (Kunming strain) against the positive phages could recognize both the SjC-TPI and the protein of the positive phages. The DNA sequencing data showed no homology between the sequences of the deduced amino acid of the two mimic antigen peptides and the amino acid of SjC-TPI. CONCLUSION: The two mimic antigen epitopes of SjC-TPI obtained are imitative epitopes of the configuration antigen of SjC-TPI.

[Effects of anti-idiotypic antibody NP30 on modulation of egg granuloma formation and hepatic fibrosis of schistosomiasis].

OBJECTIVE: To study the effects of the monoclonal anti-idiotypic antibody NP30 active immunization on egg granuloma formation and hepatic fibrosis in Schistosoma japonicum infection. METHODS: ICR mice were actively immunized with NP30 100 micrograms x 3 i.p. every 10 days while the mice in control group were injected with SP2/0 ascites i.p. simultaneously. After cercariae challenging, the mice were killed at the 4th, 8th, 12th, 16th, 20th and 24th week, respectively. Mouse livers were removed and stained histochemically with VG and subjected to immunohistochemical assay of collagen type I, III and fibronectin(FN). The volume of egg granulomas and the content of collagen type I, III and FN were determined quantitatively by NYD-1000 Image Analysis System. RESULTS: The volume of egg granulomas in NP30 immunized group was much smaller than that of control group from the 12th week after cercariae challenge. The cellular components of egg granulomas in NP30 immunized group were significantly different from those of the control group, exhibiting two types of atypical egg granulomas were found. VG stain revealed that the average optical density of collagen in hepatic granulomas of experimental group was lower than that of control group. Immunohistochemical assay revealed that the contents of collagen type I, III and fibronectin in egg granulomas of experimental group were lower than those of control group. CONCLUSION: NP30 vaccination may induce both cellular and humoral protective immunity to modulate egg granulomas and suppress liver fibrosis of schistosomiasis japonica.

[Cloning and sequence analysis of the light chain variable region gene of monoclonal anti-idiotypic antibody NP30 of Schistosoma japonicum].

OBJECTIVE: To amplify and sequence the light chain of anti-idiotypic monoclonal antibody NP30 of Schistosoma japonicum. METHODS: By comparing the conserved regions at each end of the nucleotide sequences of murine germ-line genes encoding FR1 and FR4 regions of immunoglobulin light chain variable regions, we designed a set of primers for amplification of VL gene. The hybridoma cells secreting anti-idiotypic monoclonal antibody NP30 of Schistosoma japonicum were cultured and their genome DNAs were extracted and used as templates for PCR. The PCR product was then cloned into pUC19 vector. The recombinants were sequenced by Sanger's method. The VL gene was compared with GenBank and published mouse VL genes. RESULTS: The full-length of VL gene was 318 bp. The VL gene was a member of mouse Ig kappa light chain subgroup IV and generated from rearrangement of germ line V and J kappa 4 genes. The VL gene sequence has been registered by GenBank(accession No. AF206720). CONCLUSION: The obtained VL gene was a potentially functional gene of anti-idiotypic monoclonal antibody NP30 of Schistosoma japonicum.

Immunodiagnosis of Schistosoma japonicum infection in China.

Diagnosis is an essential prerequisite for the control programs of schistosomiasis. Chinese scientists have been making efforts for decades towards the development of immunological approach in an attempt to offer alternatives of or supplements to parasitological diagnosis. The authors outline briefly the main advances in the research of immunodiagnosis for schistosomiasis japonica in China in two catalogues: antibody detection and antigen detection. A number of immunodiagnostic assays based on antibody detection, which have been used widely in the control programs in China, are appraised in terms of sensitivity and specificity, and the achievements in the development of immunodiagnostic assays based on antigen detection are introduced. Readers will find some information concerning the efforts so far made by Chinese scientists in the improvement of immunodiagnosis by use of molecular approach, including search for biochemically and/or immunochemically isolated and/or purified antigens, recombinant antigens and anti-idiotypic reagents of diagnostic interest to construct defined antigen-antibody systems for diagnosis, and development of monoclonal antibody-based assays for antibody or antigen detections.

[A study on the monoclonal anti-idiotypic antibody NP30 for the diagnosis of schistosomiasis japonica].

In this study the ability of the monoclonal anti-idiotypic antibody NP30 was tested as a substitute of diagnostic antigen in detecting antibody of Schistosoma japonicum from human sera by use of ELISA. The results showed that the seropositive rate was 98% with NP30 in the group of acute infection, which was comparable to 94% with gut associated antigens (GAA) and 98% with the soluble egg antigens (SEA); 87% with NP30 in the group of chronic infection which was comparable to 86% with GAA but lower than that of 98% with SEA. The false positive rate was about 3% for all three diagnostic antigens. The results also showed that the geometric mean titer (GMT) of antibody to NP30 was higher than that to GAA but lower than that to SEA in the acute infection group and the GMT of antibody to NP30 was lower than both those to GAA and to SEA in the chronic infection group, suggesting that the antibody to NP30 appeared earlier and decayed more quickly during the process of infection. The authors suggested that NP30 could be used for the diagnosis of schistosomiasis japonica.

Comparison of immune repertoires of Chinese and Philippine patients infected with Schistosoma japonicum.

Chinese and Philippine patients exhibited detectable titers of serum antibodies to Schistosoma japonicum worm and egg antigens. Western blot (immunoblot) data revealed differing antigenic profiles. Antigen inhibition studies showed low and high levels of cross-reactivity with anti-worm and anti-egg antibodies, respectively, derived from both Chinese and Philippine patients. Thus, anti-egg antibodies and egg antigens are more conserved than anti-worm antibodies and worm antigens.

An immunoregulatory human monoclonal antibody in schistosomiasis japonica.

A series of human monoclonal antibodies were generated using splenocytes from a Chinese patient with chronic schistosomiasis who had undergone splenectomy as part of a portacaval decompression operation. Splenocytes were transformed in bulk culture by Epstein Barr virus and transformants fused with the HMMA 2.11 TG/O cell line. Twenty individual IgG antiworm and egg antibody-producing hybridomas were generated and screened for antigen reactivity by Western blot and for suppression of antigen-induced blastogenesis of murine splenocytes from Schistosoma japonicum-infected animals. Only one IgG clone significantly suppressed (56% P less than 0.05) soluble egg antigen (SEA)-induced blastogenesis. This human monoclonal antibody bound a 50 kD carbohydrate antigen on Western blot analysis, binding both the adult worm and egg antigens of this parasite. The non-regulatory monoclonal antibodies bound this same molecule present in adult worms but not the corresponding molecule in a preparation of soluble eggs. Thus, specific immunoregulatory epitopes can be identified by human monoclonal antibodies generated from patients with chronic disease.

[The establishment and characterization of the anti-idiotypic monoclonal antibody-NP30 of Schistosoma japonicum].

A hybridoma cell line secreting an IgM monoclonal antibody designated NP30 was obtained from a fusion of SP2/o and spleen cells of a BALB/c mouse chronically infected with schistosoma japonicum for one and a half year and identified by screening with immunized rabbit sera against gut-associated antigen (GAA) and soluble egg antigen (SEA) of S. japonicum, indicating that the NP30 was an anti-anti-antigen or anti-antibody. NP30 was further determined to be an anti-idiotypic antibody (anti-id) which was serologically and functionally identical to GAA, so that it could be portrayed as the internal image of GAA, which might have the potential to be used as an antigenic reagent in immunodiagnostic assays of schistosomiasis japonica.