Objective:To develop a validation method for microbial targeted next generation sequencing (tNGS) detection for respiratorypathogens, and to evaluate the performance of the pathogen-targeted high-throughput sequencing test implemented in local hospital.Methods:Cross-sectional study. A total of 14 patients with severe pulmonary infections were admitted to Huangshi Central Hospital from December 2023 to January 2024. Samples were collected as follows:Bronchoalveolar lavage fluid (BALF) samples ( n=7) subjected to culture, fluorescent PCR, and tNGS testing. Sputum samples ( n=2) analyzed via sputum culture, fluorescent PCR, and tNGS. Throat swab samples ( n=5) tested using fluorescent PCR-capillary electrophoresis and tNGS. Reference samples were prepared using representative species such as Influenza A virus, Adenovirus C, Klebsiella pneumoniae, Aspergillus fumigatus, Staphylococcus aureus, Staphylococcus epidermidis, Candida parapsilosis, and Candida albicans. Jurkat cells at different concentrations were used as a source of human cells. Traditional detection methods such as fluorescent PCR-capillary electrophoresis and culture methods were used as reference methods. The detection performance of tNGS was evaluated by assessing the detection limit, precision, human cell impact, stability, cross-reactivity, and accuracy of metagenomic next-generation sequencing for pathogen detection. Results:The detection limits for Klebsiella pneumoniae, Human Adenovirus C, and Influenza A virus were 2×10 2 copies/ml, and for Aspergillus fumigatus, Staphylococcus aureus, Staphylococcus epidermidis, Candida parapsilosis, and Candida albicans, the detection limits were 4×10 2 copies/ml. The consistency rate of repeated detection results for all pathogens in the reference samples was 100%. The impact assessment experiment of human cells showed that when the concentration of Jurkat cells reached 1×10 6 cells/ml, Influenza A virus, Adenovirus C, Klebsiella pneumoniae, and Aspergillus fumigatus could all be detected. Stability experiments showed that there was no significant change in the number of pathogen sequences after the specimens were stored at 4 ℃ and -20 ℃ for 1 day, 4 days, and 7 days, respectively. Cross-reactivity experiments showed that when the concentration ratios of Staphylococcus aureus, Staphylococcus epidermidis, Candida parapsilosis, and Candida albicans were (5∶1∶1∶5), (1∶5∶5∶1), and (1∶1∶1∶1), respectively, the detection rate of closely related microbial species was 3/3. Accuracy assessment showed that the accuracy of 19 clinical specimens was 18/19 cases. Conclusion:Compared with traditional detection methods as the reference, tNGS demonstrates high sensitivity and a high positive concordance rate, underscoring its significant clinical value in the detection of respiratory pathogens.

Objective:To develop a validation method for microbial targeted next generation sequencing (tNGS) detection for respiratorypathogens, and to evaluate the performance of the pathogen-targeted high-throughput sequencing test implemented in local hospital.Methods:Cross-sectional study. A total of 14 patients with severe pulmonary infections were admitted to Huangshi Central Hospital from December 2023 to January 2024. Samples were collected as follows:Bronchoalveolar lavage fluid (BALF) samples ( n=7) subjected to culture, fluorescent PCR, and tNGS testing. Sputum samples ( n=2) analyzed via sputum culture, fluorescent PCR, and tNGS. Throat swab samples ( n=5) tested using fluorescent PCR-capillary electrophoresis and tNGS. Reference samples were prepared using representative species such as Influenza A virus, Adenovirus C, Klebsiella pneumoniae, Aspergillus fumigatus, Staphylococcus aureus, Staphylococcus epidermidis, Candida parapsilosis, and Candida albicans. Jurkat cells at different concentrations were used as a source of human cells. Traditional detection methods such as fluorescent PCR-capillary electrophoresis and culture methods were used as reference methods. The detection performance of tNGS was evaluated by assessing the detection limit, precision, human cell impact, stability, cross-reactivity, and accuracy of metagenomic next-generation sequencing for pathogen detection. Results:The detection limits for Klebsiella pneumoniae, Human Adenovirus C, and Influenza A virus were 2×10 2 copies/ml, and for Aspergillus fumigatus, Staphylococcus aureus, Staphylococcus epidermidis, Candida parapsilosis, and Candida albicans, the detection limits were 4×10 2 copies/ml. The consistency rate of repeated detection results for all pathogens in the reference samples was 100%. The impact assessment experiment of human cells showed that when the concentration of Jurkat cells reached 1×10 6 cells/ml, Influenza A virus, Adenovirus C, Klebsiella pneumoniae, and Aspergillus fumigatus could all be detected. Stability experiments showed that there was no significant change in the number of pathogen sequences after the specimens were stored at 4 ℃ and -20 ℃ for 1 day, 4 days, and 7 days, respectively. Cross-reactivity experiments showed that when the concentration ratios of Staphylococcus aureus, Staphylococcus epidermidis, Candida parapsilosis, and Candida albicans were (5∶1∶1∶5), (1∶5∶5∶1), and (1∶1∶1∶1), respectively, the detection rate of closely related microbial species was 3/3. Accuracy assessment showed that the accuracy of 19 clinical specimens was 18/19 cases. Conclusion:Compared with traditional detection methods as the reference, tNGS demonstrates high sensitivity and a high positive concordance rate, underscoring its significant clinical value in the detection of respiratory pathogens.

Objective To analyze the risk factors affecting the prognosis of patients with advanced gastric cancer,so as to provide evidence for identifying high-risk populations and guiding clinical treatment.Methods The clinical data of 282 patients with advanced gastric cancer who admitted to Zhoushan Hospital of Traditional Chinese Medicine from January 2017 to December 2021 were retrospectively analyzed.The survival outcome of patients was obtained by telephone follow-up and regular review.Kaplan-Meier method was used to analyze the overall survival(OS).Univariate and multivariate COX regression models were used to analyze the effects of clinical features on OS in patients with advanced gastric cancer.Results The median OS of 282 patients with advanced gastric cancer was 10.0 months.Univariate analysis showed that first-line chemotherapy,traditional Chinese medicine treatment,body mass index(BMI),Karnofsky performance status(KPS)score,hemoglobin(Hb)and albumin(Alb)were significantly correlated with the survival of patients with advanced gastric cancer(P<0.05).Multivariate COX regression models analysis showed that first-line chemotherapy,BMI,KPS score and Alb were independent risk factors affecting the survival of patients with advanced gastric cancer(P<0.05).Conclusion BMI,Alb,KPS score and first-line chemotherapy are independent risk factors for predicting the prognosis of patients with advanced gastric cancer,indicating nutrition management may be an important way to improve the outcome of patients with advanced gastric cancer.

Objective We evaluated the molecular epidemiology ofStenotrophomonas maltophilia strains in adult patients in Renji Hospital to Shanghai Jiao Tong University School of Medicine for better control ofS.maltophilia infections.Methods Nonduplicate clinical isolates of S.maltophilia were collected from Renji Hospital from January 2014 to September 2014.We examined the clonality among the S.maltophilia isolates by multilocus sequence typing (MLST) and pulsed field gel electrophoresis (PFGE).Antimicrobial resistance pattern was investigated by Kirby-Bauer method and prevalence of toxin genes (Stmpr1,Stmpr2,stmr-1,Smlt3773locus) by PCR.We also studied the biofilm formation of S.maltophilia by semiquantitative biofilm formation test.Results A total of 78 nonduplicate S.maltophilia isolates were analyzed,of which 26 were isolated from surgical intensive care unit,and 53 strains were from male patients.All patients infected by S.maltophilia had received antibiotic therapy before the strains were isolated.At least three kinds of antibiotics were used in 62.8％ of the patients.MLST analysis indicated that 49 isolates were assigned novel STs(STnewl-STnew38),with new combinations of allelic profiles.The largest cluster of isolates was ST23 (6 strains).PFGE showed that there was weak genetic linkage between S.maltophilia strains.The 78 isolates were divided into 58 groups.About 2.6％ (2/78) and 10.3％ (8/78) of these strains were resistant to levofloxacin and trimethoprim-sulfamethoxazole,respectively.All the strains were susceptible to minocycline.The prevalence of virulence genes Stmprl,Stmpr2,snf-1 and Smlt3773 locus was 79.5％ (62/78),93.6％ (73/78),94.9％ (74/78) and 48.7％ (38/78),respectively.Biofilm formation test indicated that the mean ability of biofilm formation was 0.51±0.44 in terms of D492.There was no significant difference between males and females.Conclusions All patients with Stenotrophomonas maltophilia infection had a history of antibiotic use and male patients were susceptible population.Stenotrophomonas maltophilia isolates showed high prevalence of virulence genes.No clonal dissemination was found in the same department of hospital.

Objective To compare the impacts of sustained and intermittent gradient pressure on perfusion disorders and myofiber injuries and to explore the mechanism by which intermittent gradient pressure helps treat deep tissue injury (DTI). MethodsTwenty Sprague-Dawley rats 10-12 weeks old were randomly divided into an experimental group and a control group.The tibialis anterior muscles of the experimental group received intermittent fluctuating pressure at 8.0-21.3 kPa while the control group received sustained pressure at 13.3 kPa.The contralateral notpressed muscles served as healthy controls.The experiment was terminated after 3 cycles of compression where each cycle involved 2 h of compression and 30 min of conpression release.The general condition of the rats'skin and muscles in the compressed region were observed and laser doppler perfusion imaging was used to detect blood perfusion of the skin of the compressed leg after 3 cycles of compression.Any pathomorphological changes in the tibialis anterior muscles of the compressed region were observed by phosphotungstic acid hematoxylin (PTAH) staining. Results Blood perfusion of the skin and muscle in the control group was significantly less than in the experimental group after compression,and blood flow velocity increased with time in both groups.However,perfusion was significantly more extensive in the experimental group.PTAH staining showed that muscle injury in the experimental group was significantly less severe than in the control group. ConclusionCompared to the sustained pressure,intermittent pressure cause significantly less severe DTI by reducing compression-induced ischemia and hypoxia.

Objective To explore mood disorders after left and right cerebral hemisphere injury,to search for possible mechanisms and to provide the basis for designing rehabilitation protocols and assessing prognosis.Methods Fifty-one cases with brain injury were recruited from a rehabilitation center and divided into left and right hemisphere injury groups with 26 and 25 cases,respectively.Hamilton rating scale for depression (HRSD) scores were analyzed. Results The left hemisphere patients had significantly higher HRSD total scores and higher scores indicating 15 kinds of mood disorders (depression,feelings of guilt,suicidal thoughts etc.).Right hemisphere injury patients had significantly higher diurnal variation,depersonalization or derealization and paranoid symptoms. Conclusions The lateralization of the cerebral hemispheres may display itself in mood and emotion.After left or right hemisphere injury,the depression presentation is different,so treatment,prognosis assessment and psychological intervention should be different for left and right hemisphere injuries.

With the development of Chinese education system, academic students in medical university can not meet the demand of current social development. The combination of teaching and research ideas should run through the whole teaching process, which is an important approach to bring up innovative talents. This article focuses on teaching process of rehabilitation medicine, and how to run research ideas through each sections of rehabilitation medicine. Research ideas begins with teaching, shaped in clinical practice, and being applied for clinical work. That is just the right thinking for cultivating outstanding medical specialists in the 21st century.