Objective:To explore the genetic diversity of Mycobacterium tuberculosis (MTB) in different types of tuberculosis and its association with clinical features, providing evidence for precise diagnosis and treatment of tuberculosis. Methods:This cross-sectional study included 103 cases of tuberculosis (38 with simple pulmonary tuberculosis, 43 with tuberculous pleurisy, and 22 with pulmonary combined with extrapulmonary tuberculosis) from Shenzhen Third People′s Hospital from 2015 to 2018. Paired bacterial strains from lung and pleural effusion/extrapulmonary sites were collected. Whole-genome sequencing (WGS) was used for drug resistance prediction, and genetic diversity (π value) was calculated as well as differential genes screening. Statistical analysis included paired t-tests and χ2 tests to compare clinical, bacteriological and genetic diversity features among groups.Results:The simple pulmonary tuberculosis group exhibited significantly higher rates of retreatment (71.7%, 27/38), cavitation (70.4%, 19/27), and multidrug-resistant or rifampicin-resistant (MDR/RR) (60.5%, 23/38) compared to the tuberculous pleurisy group (retreatment 11.9%, 5/42; cavitation 11.9%, 5/42; MDR/RR 16.3%, 7/43) and extrapulmonary tuberculosis group (retreatment 9.1%, 2/22; cavitation 18.2%, 4/22; MDR/RR 13.6%, 3/22) ( P<0.05). The overall π values of the MTB strain genomes in lung [(5.94±3.93)×10 ?5], pleural effusion[(6.22±3.51)×10 ?5], and extrapulmonary tissues [(5.83±3.54)×10 ?5] showed no significant differences ( H=0.10, P=0.94). Differential gene diversity analysis revealed that π value alternating genes related to respiration and intermediate metabolism were prominently high [tuberculous pleurisy 32.4% (11/34) and extrapulmonary tuberculosis groups 31.4% (32/102)], while cell wall-associated genes dominated in the simple pulmonary tuberculosis group (42.9%, 6/14). Drug resistance profiles and mutation spectra were identical across isolates from different sites within the same patient. Conclusion:WGS revealed the MTB diversity among different types of tuberculosis. Difference between pulmonary and extrapulmonary environments may impel the adaptive alternations of the bacterial strains to maintain survival with higher overall genome stability. Drug resistance testing of lung-derived isolates may provide references on extrapulmonary tuberculosis treatment.

Objective To establish a purification,enrichment and test method of tetrodotoxin in blood.Methods Through the investigation of various hydrophilic chromatographic columns,the comparison of extraction effects of different types of solid phase extraction columns and the interference analysis of mixed peaks on qualitative ion pairs,the matrix influence of tetrodotoxin was reduced,and the detection sensitivity and qualitative accuracy were improved.Results Tetrodotoxin is highly polar and easily inhibited by the matrix,while conventional precipitation protein method has low sensitivity and isomer double peaks,and the C18 column is not reserved.After comprehensive comparative analysis,the weak cation exchange column PWC column is finally used for purification and enrichment.Complete elution was achieved using 0.5 mL 10%formic acid and 50%acetonitrile aqueous solution.Seperation was performed on an Atlantis HILIC column,with qualitative ion pairs set at m/z,320.10>162.15 and 284.15.The detection limit of the method was 0.061 ng/mL.Conclusion The established PWC solid-phase extraction-LC/MS detection method demonstrates significant purification efficacy,minimal matrix influence,unobstructed chromatographic peaks,markedly improved detection sensitivity.This approach is operationally simple,and applicable to forensic casework.

Objective To establish a purification,enrichment and test method of tetrodotoxin in blood.Methods Through the investigation of various hydrophilic chromatographic columns,the comparison of extraction effects of different types of solid phase extraction columns and the interference analysis of mixed peaks on qualitative ion pairs,the matrix influence of tetrodotoxin was reduced,and the detection sensitivity and qualitative accuracy were improved.Results Tetrodotoxin is highly polar and easily inhibited by the matrix,while conventional precipitation protein method has low sensitivity and isomer double peaks,and the C18 column is not reserved.After comprehensive comparative analysis,the weak cation exchange column PWC column is finally used for purification and enrichment.Complete elution was achieved using 0.5 mL 10%formic acid and 50%acetonitrile aqueous solution.Seperation was performed on an Atlantis HILIC column,with qualitative ion pairs set at m/z,320.10>162.15 and 284.15.The detection limit of the method was 0.061 ng/mL.Conclusion The established PWC solid-phase extraction-LC/MS detection method demonstrates significant purification efficacy,minimal matrix influence,unobstructed chromatographic peaks,markedly improved detection sensitivity.This approach is operationally simple,and applicable to forensic casework.

Objective:To explore the genetic diversity of Mycobacterium tuberculosis (MTB) in different types of tuberculosis and its association with clinical features, providing evidence for precise diagnosis and treatment of tuberculosis. Methods:This cross-sectional study included 103 cases of tuberculosis (38 with simple pulmonary tuberculosis, 43 with tuberculous pleurisy, and 22 with pulmonary combined with extrapulmonary tuberculosis) from Shenzhen Third People′s Hospital from 2015 to 2018. Paired bacterial strains from lung and pleural effusion/extrapulmonary sites were collected. Whole-genome sequencing (WGS) was used for drug resistance prediction, and genetic diversity (π value) was calculated as well as differential genes screening. Statistical analysis included paired t-tests and χ2 tests to compare clinical, bacteriological and genetic diversity features among groups.Results:The simple pulmonary tuberculosis group exhibited significantly higher rates of retreatment (71.7%, 27/38), cavitation (70.4%, 19/27), and multidrug-resistant or rifampicin-resistant (MDR/RR) (60.5%, 23/38) compared to the tuberculous pleurisy group (retreatment 11.9%, 5/42; cavitation 11.9%, 5/42; MDR/RR 16.3%, 7/43) and extrapulmonary tuberculosis group (retreatment 9.1%, 2/22; cavitation 18.2%, 4/22; MDR/RR 13.6%, 3/22) ( P<0.05). The overall π values of the MTB strain genomes in lung [(5.94±3.93)×10 ?5], pleural effusion[(6.22±3.51)×10 ?5], and extrapulmonary tissues [(5.83±3.54)×10 ?5] showed no significant differences ( H=0.10, P=0.94). Differential gene diversity analysis revealed that π value alternating genes related to respiration and intermediate metabolism were prominently high [tuberculous pleurisy 32.4% (11/34) and extrapulmonary tuberculosis groups 31.4% (32/102)], while cell wall-associated genes dominated in the simple pulmonary tuberculosis group (42.9%, 6/14). Drug resistance profiles and mutation spectra were identical across isolates from different sites within the same patient. Conclusion:WGS revealed the MTB diversity among different types of tuberculosis. Difference between pulmonary and extrapulmonary environments may impel the adaptive alternations of the bacterial strains to maintain survival with higher overall genome stability. Drug resistance testing of lung-derived isolates may provide references on extrapulmonary tuberculosis treatment.

Objective:To investigate the protective effect and related mechanisms of Huatuo Zaizao pills (HT) on white matter injury and cognitive impairment induced by chronic cerebral hypoperfusion in mice. Methods:Forty adult male C57BL/6J mice were randomly divided into sham-operation group, bilateral carotid artery stenosis (BCAS) model group, and HT group. An animal model of BCAS was constructed using the spring loop into the bilateral common carotid arteries. After continuous treatment with 5 g/kg HT (or an equal amount of purified water) for 4 weeks, cognitive function was evaluated using the novel object recognition test. Morphological and structural changes in myelin sheath were evaluated by LFB myelin staining. White matter damage and glial cell expression were detected by myelin associated glycoprotein (MAG) in the corpus callosum, ionized calcium-binding adapter molecule 1 (IBA-1), and glial fibrillar acidic protein (GFAP) in corpus callosum and hippocampus through immunofluorescence staining. Real time quantitative polymerase chain reaction (qPCR) was used to detect mRNA expressions of myelin-associated proteins, Janus kinase 2 (JAK2), signal transducer and activator of the transcription 3 (STAT3) in corpus callosum, as well as brain-derived neurotrophic factor (BDNF), glutathione peroxidase 1 (GPx-1), and various inflammatory factors in hippocampus.Results:The novel object recognition test showed that mice had significant working memory impairment at 4 weeks after BCAS ( P<0.01), while the HT group showed significant improvement in working memory impairment compared to the BCAS group ( P<0.01). LFB myelin staining showed significant myelin damage in the BCAS group ( P<0.001), while the degree of myelin damage in the HT group was significantly improved compared to the BCAS group. Immunofluorescence staining showed that both the BCAS and HT groups had proliferation of microglia in the corpus callosum and hippocampus, and there was no significant difference between the two groups. In contrast, the activation of astrocytes in the corpus callosum was significantly improved in the HT group compared to the BCAS group ( P<0.05). qPCR showed upregulation of myelin-associated proteins as well as JAK2 and STAT3 mRNA expression in the BCAS group. Compared with the BCAS group, the expressions of JAK2 and STAT3 mRNA were decreased in the HT group (all P<0.05), while the expression of myelin-associated proteins were upregulated (all P<0.05). There were no significant difference in the expressions of inflammatory factors, BDNF, and GPX1 mRNA in the hippocampal tissue between the BCAS group and the HT group. Conclusion:HT may improve cognitive impairment and white matter damage in mice with chronic cerebral hypoperfusion, and the JAK2-STAT3 pathway may be one of its effect pathways.

Objective:To investigate the expression and cellular provenance of interleukin 17A (IL-17A) in non-eosinophilic chronic rhinosinusitis with nasal polyps (nECRSwNP), and to analyze the possible reasons for its different expression.Methods:Samples were collected from 14 patients with eosinophilic chronic rhinosinusitis with nasal polyps (ECRSwNP) and 28 patients with nECRSwNP, who underwent functional endoscopic sinus surgery in the Third Affiliated Hospital of Sun Yat-sen University from January 2017 to May 2018, including 33 males and 9 females, with the age ranging from 18 to 65 years old. Enzyme linked immune sorbent assay (ELISA) and flow cytometry were used to investigate the expression and cellular origins of IL-17A in the nasal tissue of ECRSwNP and nECRSwNP groups. Then the difference of quantity and differentiation ability of the major cells producing IL-17A between ECRSwNP and nECRSwNP groups were analyzed by flow cytometry. Finally, the expressions of IL-6, transforming growth factor-β(TGF-β), and IL-23, which were considered as the important factors in promoting Th17/Tc17 differentiation in CRSwNP and their correlation with IL-17A, were analyzed by ELISA. Statistical analysis was performed using IBM SPSS 20.Results:The IL-17A protein levels and IL-17A +lymphocyte percentages were higher in nECRSwNP group compared with that of the ECRSwNP group (158.56 (167.76) pg/ml ( M( QR)) vs. 9.42 (11.33) pg/ml, 10.21%±1.54% ( ± s) vs. 3.93%±0.80%, Z=2.95, t=3.62, all P<0.01). Tc17 cells (CD8 +T cells producing IL-17A) and Th17 cells (CD4 +T cells producing IL-17A) were major IL-17A producers in both ECRSwNP and nECRSwNP group. Further analysis revealed that there was no significant difference in quantity of CD8 +and CD4 +T cells between ECRSwNP and nECRSwNP group, but the differentiation ability about CD8 +and CD4 +T cells differentiating into Tc17 and Th17 in nECRSwNP group was stronger than that in ECRSwNP. The high expressions of IL-6 and TGF-β, which were considered as the important factors in promoting Th17/Tc17 differentiation were also found in nECRSwNP group compared with ECRSwNP (56.07 (234.25) pg/ml vs. 8.27 (12.51) pg/ml, (5.44±0.34) pg/ml vs. (4.17±0.22) pg/ml, Z=2.426, t=2.29, all P<0.05). But the difference in expression of IL-23 was not significant difference between the two groups. Moreover, the expression of IL-17A showed significantly positive correlation with IL-6 ( r=0.615, P=0.009). No positive correlation between IL-17A and TGF-β or IL-23 was observed. Conclusions:The expression of IL-17A in nasal mucosa of nECRSwNP patients is significantly higher than that of ECRSwNP, which is due to the increase of expression and differentiation of Tc17/Th17 cells. IL-17A shows positive correlation with IL-6 in CRSwNP, which is the important factor in promoting Th17/Tc17 differentiation.

Objective To investigate the nursing of sedation and analgesia using low-dose fentanyl with midazolam during intra-arterial treatment with mechanical thrombectomy by means of stent for patients with anterior-circulation acute ischemic stroke.Methods All of 61 patients who underwent intra-arterial treatment with anterior-circulation acute ischemic stroke were retrospectively collected.They were divided into two groups according to the different program of individual conscious sedation:the control group(30 cases)used midazolam for intravenous maintain sedation,experimental group(31 cases)added the low-dose fentanyl on the basis of the control group.The thrombectomy was performed by the same group of interventional radiologists,while the effect of sedative,vital signs,respiratory curve and SpO2(blood oxygen saturation),assessment of Ramsay's sedation score,restlessness and other sedation adverse reactions were respectively observed in both groups.Nursing intervention was performed aiming at different states of patients during operation.Results All 61 patients were implemented with good medical care with no intracranial hemorrhage or intraoperative death.In the control group,there were 4 patients with Ramsay 1 point of sedation,resulted motion artifacts in digital subtraction angiography images in 3 of them,1 patient with slight reduction of SpO2,and vomiting in 4 patients.1 patient with decreased mean arterial pressure.In the experimental group,there was no Ramsay 1 point of sedative patient.The SpO2 was decreased in 5 patients.Respiratory inhibition occurred in 3 patients and the mean arterial pressure decreased slightly in the other 3 patients.Conclusions Application of low-dose fentanyl and midazolam is safe and reliable in patients who underwent intra-arterial treatment with anterior-circulation acute ischemic stroke.While closely observation to the changes of breath frequency curve and SpO2 in operation are very essential to actively prevent the occurrence of respiratory depression.

Objective:The aim of the current study is to understand the sleep disorders in children with autism and related factors.Methods:A case-control study method was conducted in this study which included 198 children autism who met the criteria of the Diagnostic and Statistical Manual of Mental Disorders,Fourth Edition (DSM-Ⅳ) and 233 cases of gender-and age-matched normal children.Their general demographic and sleep data were collected and the children with autism were assessed with Autism Behavior Checklist,Child Behavior Checklist and Autism Spectrum Screening Questionnaire.Correlation analysis and regression analysis were performed to compare sleep quality,sleep problems and the related factors of the two groups.Results:The scores of difficulty falling asleep,sleep restlessness,taking hold or shaking,restless sleep,interrupted sleep,nightmares,sleepwalking,night terrors,sleep snoring,nocturnal enuresis,sleep apnea,mouth breathing,sweating and sleep scores were higher in the autism group compared to the normal control group before falling asleep (P ＜ 0.05),while the proportion of daytime sleep time in the autism group was lower than the normal control group (P ＜0.05).With sleep status as dependent variable,linear regression analysis was conducted with children's birth weight,walking month age,speaking month age,father age,and autism spectrum as independent variables.The multiple linear regression analysis showed the scores of,sensory behaviors,body and object use behaviors,and self-help behaviors in Autism Behavior Checklist for autistic children were positively associated with their sleep (β =0.21,0.21,0.29,P ＜ 0.05).However,paternal age at birth in the autism group had negative associatin with the sleep of autistic children (β =-0.16,P ＜0.05).In the autism group,their total scores of Autism Behavior Checklist and the scores of language and social behavior in Autism Behavior Checklist were not associated with their sleep status score (P ＞ 0.05).Conclusion:It suggests that children with autism may have more sleep problems,and their sleep problems are related to autism symptoms including the scores of,sensory behaviors,body and object use behaviors,and self-help behaviors in Autism Behavior Checklist for autistic and their paternal age at birth.

Objective: To investigate the clinical features of intracranial Castleman's disease, and to improve the understanding of the disease. Methods: The clinical features of one case of intracranial mixed type Castleman's disease, who was diagnosed in Department of Neurosurgery, Zhujiang Hospital, Southern Medical University in February 9, 2017, were retrospectively analyzed, and the relevant literatures about intracranial Castleman's disease were reviewed. The PubMed, Embase, Ovid, Cochrane, Wanfang Database and China Journal Full-text Database were searched using the keywords including "Castleman", "Intracranial"and "Central nervous system" for related literatures published between January 1, 1980 and January 31, 2017. Results: A 32-year old female was admitted because of headache for one month. CT examination showed an irregular and well-defined hyperdense lesion in the right temporo-parietal area with surrounding extensive edema and this mass was markedly contrast enhanced. Laboratory examination revealed anemia and hyperimmunoglobulinemia. The mass was removed completely by surgical resection. Postoperative pathology examination revealed mixed type Castleman's disease in the right temporo-parietal area. MRI was performed 3 months after the surgery. No residual tumor and recurrence were demonstrated. Seventeen related case reports were retrieved and 20 cases of intracranial Castleman's disease were reported. Among total 21 cases, there were 7 males and 14 females, ranging from 8 to 82 years old, and the median age was 47 years old. The mass arised from the dura matter or leptomeninges, and the most common sites of origin were located at the cerebral convexity and the parafalcine. The most common clinical features were seizures and headache. Radiographically, the previously reported cases presented as a homogeneously enhanced extra-axial well-defined mass with a marked peritumoral edema. Conclusion: Intracranial Castleman's disease is a rare clinical condition. The diagnosis can be made in combination with clinical findings and the laboratory examination.

Objective]In order to study the impact of adenoid hypertrophy(AH)with allergic rhinitis(AR)in the otitis media with effusion(OME)in children,as well as to discuss the risk factors on the inducement of OME in patients with AH.[Methods]The clinical materials were collected and analyzed from 205 children with AH who admitted for surgical treatment between 2013 and 2015, including medical history and signs,acoustic immittance measurement,allergy screen as well as blood routine,and to evaluate the situation of OME and AR in patients with AH. All the data were analyzed by SPSS 20.0.[Results]Among the 205 AH children,66 cases(32.20%)were accompanying with OME. The AH patients aged 3-5 years had the highest incidence of OME,which decreased with age(P=0.018). It is significant on the prevalence of the OME in patients with AH between the two groups with and without AR (P = 0.010). In the logistic model investigating the risk factors of inducing OME among patients with AH,the third-degree AH, Type-three and the accompanying with AR were significant. The third-degree and third-type AH children were 2.729 and 6.390-folds higher than others respectively (P = 0.047,P = 0.001). The incidence of OME appeared to be 1.212-folds higher among patients with AR(P = 0.010).[Conclusion]Mechanical obstruction and AR played a superimposed role in the inducement of OME among patients with AH,which was effected by multiple factors. We should pay attention to the“lateral respiratory”allergic diseases on eustachian tube and middle ear.