Objective To report the diagnosis of a canine distemper virus outbreak among a colony of cynomolgus monkeys at an experimental monkey farm in 2019. MethodsA total of 46 samples were collected from 21 diseased cynomolgus monkeys (exhibiting symptoms such as facial rash, skin scurf, runny nose, and diarrhea) and from one deceased monkey at an experimental monkey breeding farm in South China in late 2019, including serum, skin rash swabs, and anticoagulated whole blood, liver, lung, and skin tissues were submitted for testing. All submitted samples were tested for canine distemper virus gene fragments using real-time quantitative PCR, while immunohistochemical staining was performed to detect canine distemper virus nucleoprotein in lung tissues. The skin tissue of the deceased monkey was ground and sieved. The filtrate was inoculated into a monolayer MDCK cell line for virus isolation. Then, whole-genome sequencing was performed to identify the isolated virus. The Clustal Omega tool was used to align and analyze the homology of different Asian canine distemper virus isolates. A phylogenetic tree was constructed, followed by genetic evolutionary analysis. ResultsClinical retrospective analysis revealed that the diseased cynomolgus monkeys exhibited symptoms similar to those observed in cynomolgus monkeys infected with measles virus. Necropsy findings showed red lesions in the lungs and significant hemorrhage in the colonic mucosa. Real-time quantitative PCR detected canine distemper virus nucleic acid in the serum, skin rash swabs of the infected monkeys, and various tissue samples of the deceased monkey, all of which tested positive. Calculation based on the standard curve formula indicated the viral load was highest in the skin tissue. Immunohistochemical staining of the deceased monkey's lung tissue demonstrated aggregation of CDV nucleoprotein in alveolar epithelial cells, bronchi, and bronchioles. A CDV strain was isolated from the skin tissue of the deceased monkey. Phylogenetic analysis indicated that this strain shares the closest relationship (98.86%) with the Asian-1 type canine distemper virus strain CDV/dog/HCM/33/140816, previously identified in dogs in Vietnam. ConclusionBased on comprehensive analysis of clinical symptoms, nucleic acid detection, viral protein immunohistochemistry, and whole-genome sequencing results, the diagnosis confirms that the cynomolgus monkeys in this facility are infected with canine distemper virus. It is recommended to include canine distemper virus as a routine surveillance target in captive monkey populations. Additionally, this study provides a foundation for further research on the molecular biological characteristics of canine distemper virus.

Objective To report the diagnosis of a canine distemper virus outbreak among a colony of cynomolgus monkeys at an experimental monkey farm in 2019. MethodsA total of 46 samples were collected from 21 diseased cynomolgus monkeys (exhibiting symptoms such as facial rash, skin scurf, runny nose, and diarrhea) and from one deceased monkey at an experimental monkey breeding farm in South China in late 2019, including serum, skin rash swabs, and anticoagulated whole blood, liver, lung, and skin tissues were submitted for testing. All submitted samples were tested for canine distemper virus gene fragments using real-time quantitative PCR, while immunohistochemical staining was performed to detect canine distemper virus nucleoprotein in lung tissues. The skin tissue of the deceased monkey was ground and sieved. The filtrate was inoculated into a monolayer MDCK cell line for virus isolation. Then, whole-genome sequencing was performed to identify the isolated virus. The Clustal Omega tool was used to align and analyze the homology of different Asian canine distemper virus isolates. A phylogenetic tree was constructed, followed by genetic evolutionary analysis. ResultsClinical retrospective analysis revealed that the diseased cynomolgus monkeys exhibited symptoms similar to those observed in cynomolgus monkeys infected with measles virus. Necropsy findings showed red lesions in the lungs and significant hemorrhage in the colonic mucosa. Real-time quantitative PCR detected canine distemper virus nucleic acid in the serum, skin rash swabs of the infected monkeys, and various tissue samples of the deceased monkey, all of which tested positive. Calculation based on the standard curve formula indicated the viral load was highest in the skin tissue. Immunohistochemical staining of the deceased monkey's lung tissue demonstrated aggregation of CDV nucleoprotein in alveolar epithelial cells, bronchi, and bronchioles. A CDV strain was isolated from the skin tissue of the deceased monkey. Phylogenetic analysis indicated that this strain shares the closest relationship (98.86%) with the Asian-1 type canine distemper virus strain CDV/dog/HCM/33/140816, previously identified in dogs in Vietnam. ConclusionBased on comprehensive analysis of clinical symptoms, nucleic acid detection, viral protein immunohistochemistry, and whole-genome sequencing results, the diagnosis confirms that the cynomolgus monkeys in this facility are infected with canine distemper virus. It is recommended to include canine distemper virus as a routine surveillance target in captive monkey populations. Additionally, this study provides a foundation for further research on the molecular biological characteristics of canine distemper virus.

Objective:To analyze the effects of SARS-CoV-2 infection and vaccination on virus mutation.Methods:The whole genome sequencing sequences of 2 659 local SARS-CoV-2 specimens from Jiangsu Province in 2023 were selected for analysis, and relevant information such as demographic and clinical characteristics were collected, and the effects of infection and vaccination on the genome-wide mutation rate and S gene′s selective pressure of the virus were analyzed by univariate and multivariate linear regression models.Results:The average age of these infected patients was 55.0 (31.0, 74.0) years, 1 150 cases (43.2%) in the age group of ≥60 years, 1 367 cases (51.4%) were males, 2 044 cases (76.9%) had a history of COVID-19 vaccination, and 1 629 cases (61.3%) had the first-time infection. The clinical symptoms of the infected patients were mainly mild, with a total of 2434 cases (91.5%), and 29 cases (1.1%) with severe symptoms or more. The average substitution rate of SARS-CoV-2 was 9.69 (9.38, 9.98)×10 -4 subs/site/year, and the dN/dS value of the S gene was 6.08 (5.56, 8.66), which was significantly greater than that of 1 ( P<0.001), indicating positive selection. The result of univariate and multivariate linear regression model analysis showed that the SARS-CoV-2 substitution rate was higher in those with vaccination history and reinfection, aged 20-30 years, ≥60 years, and the SARS-CoV-2 substitution rate was lower in males with moderate clinical symptoms and severe disease and above. Those with a history of vaccination and reinfection, aged 50-60 years old, ≥60 years old have smaller S gene dN/dS. Conclusions:Under the immune pressure exerted by vaccination and infection, the genome-wide mutation of SARS-COV-2 accelerated, but the non-synonymous mutation rate of the S gene decreased. The mechanism causing these phenomena needs further study.

In recent years, global outbreaks of infectious diseases, such as COVID-19, have triggered great concern about emerging infectious diseases. With the rapid development of next-generation sequencing technology and bioinformatic tools for viral metagenomics, there is now a widespread capability to detect and identify various known and unknown pathogenic microorganisms within both environmental and biological contexts. Furthermore, the continuous evolution of machine learning methods has led to the development and application of multiple rapid and highly accurate approaches for virus identification. Concurrently, owing to the continual progress in machine learning methods, several rapid and accurate virus identification techniques have been widely developed and applied. Therefore, this review aims to systematically summarize the key methodologies, frameworks, and the scope of applicability within the field of viral metagenomics, with a specific focus on virus identification and prediction. It could facilitate a deeper understanding of viral characteristics, identify potential novel pathogens, and provide technical support for the early prevention and control of infectious diseases.

In recent years, global outbreaks of infectious diseases, such as COVID-19, have triggered great concern about emerging infectious diseases. With the rapid development of next-generation sequencing technology and bioinformatic tools for viral metagenomics, there is now a widespread capability to detect and identify various known and unknown pathogenic microorganisms within both environmental and biological contexts. Furthermore, the continuous evolution of machine learning methods has led to the development and application of multiple rapid and highly accurate approaches for virus identification. Concurrently, owing to the continual progress in machine learning methods, several rapid and accurate virus identification techniques have been widely developed and applied. Therefore, this review aims to systematically summarize the key methodologies, frameworks, and the scope of applicability within the field of viral metagenomics, with a specific focus on virus identification and prediction. It could facilitate a deeper understanding of viral characteristics, identify potential novel pathogens, and provide technical support for the early prevention and control of infectious diseases.

Objective To observe the effect of transcutaneous electrical acupoint stimulation(TEAS)on circulation depression in patients underwent thoracoscopic radical resection of lung cancer under general anesthesia combined with thoracic paravertebral block(TPVB).Methods A total of 150 patients from Octomber 2021 to May 2022,58 males and 92 females,aged 19-64 years,BMI 18-30 kg/m2,ASA physical status Ⅰ or Ⅱ,underwent thoracoscopic radical resection of lung cancer under general anesthesia combined with TPVB were enrolled.According to random number table method,the patients were divided into two groups:the TEAS group and the control group,75 patients in each group.In the TEAS group,transcutaneous electrical acupoint stimulation was performed at Hegu,Neiguan,and Zusanli 30 minutes be-fore induction until the end of operation.In the control group,the electrodes were only connected at the same time point without electrical stimulation.HR,SBP,DBP,MAP,and BIS were recorded before stimu-lation(T0),10 minutes after TPVB(T1),the time of skin incision(T2),30 minutes after operation star-ted(T3),60 minutes after operation started(T4),the end of operation(T5),and 30 minutes after opera-tion(T6).The incidences of bradycardia,tachycardia,hypotension,and hypertension,and the usages of vasoactive drugs during operation were recorded.The dosages of propofol,sufentanil,and remifentanil in the operation were recorded.The VAS pain score 1,2,and 7 days after operation,the usages of analgesics used within 7 days after operation,postoperative adverse effects such as nausea and vomiting,dizziness,chest tightness,and shortness of breath,and the length of hospital stay were recorded.Results Compared with the control group,intraoperative infusion volume,incidence of hypotension,hypertension,and circulation depression,the usages of deoxyepinephrine,ephedrine,norepinephrine,and urapidil intraoperation,VAS pain scores 1 and 2 days after operation,and the usage of analgesics within 7 days after operation were sig-nificantly decreased(P＜0.05),length of hospital stay was significantly shortened(P＜0.05),SBP,DBP,and MAP were significantly increased at T1(P＜0.05),the dosagesof propofol,sufentanil,and remifentanil were significantly decreased in the TEAS group(P＜0.05).There were no significantly differ-ences of nausea and vomiting,dizziness,and shortness of breath between the two groups.Conclusion TEAS can improve the circulation depression,and reduce the incidences of intraoperative hypotension and hypertension,decrease the dosages of anesthetics and the rate of using vasoactive drugs during operation,improve early postoperative acute pain and shorten the length of hospital stay in patients undergoing thoraco-scopic radical resection of lung cancer under general anesthesia combined with TPVB.

Objective:To investigate the clinical effects of the modified posteromedial approach combined with the anterolateral approach in the treatment of posterior pilon fractures in the supine position.Methods:A retrospective was conducted to analyze the clinical data of 54 patients [45 males and 9 females with an age of (47.7 ± 13.1) years] who had been treated surgically for posterior pilon fractures from January 2016 to December 2020 at Department of Orthopedics, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine. The patients were divided into 2 groups according to their surgical positions: a supine group of 24 patients (the modified posteromedial approach combined with the anterolateral approach in the supine position) and a prone group of 30 patients (the posteromedial approach combined with the anterolateral approach in the prone position). The 2 groups were compared in terms of operation time, hospitalization time, radiographic outcomes (bone union time and ratio of congruent articular reduction), range of ankle motion, and postoperative complications. The post-operative function was evaluated using the Manchester Oxford Foot Questionnaire (MOXFQ) and the visual analogue scale (VAS).Results:There was no statistically significant difference between the 2 groups in the general clinical data before operation, showing comparability ( P>0.05). The mean follow-up time was (19.4 ± 4.4) months for the supine group and (17.8 ± 4.2) months for the prone group. The operation time, hospitalization time, bone union time, rate of fixation of syndesmosis and ratio of congruent articular reduction were (90.8 ± 9.9) min, (9.5 ± 2.4) d, (8.4 ± 1.4) weeks, 33.3% (8/24) and 95.8% (23/24) in the supine group, and (89.1 ± 10.8) min, (9.5 ± 2.5) d, (8.1 ± 1.4) weeks, 53.3% (16/30) and 96.6% (29/30) in the prone group, showing no significant differences (all P>0.05). At the last follow-up, the dorsiflexion and plantar flexion of the ankle, VAS, and MOXFQ scores for pain, walking and social capability were, respectively, 15.0° ± 2.1°, 26.1° ± 4.2°, (1.0 ± 0.5) points, 20.0(0, 30.0) points, (16.5 ± 13.2) points and 12.5(0, 18.8) points in the supine group, and 15.7° ± 1.6°, 27° ± 4.0°, (1.3 ± 0.7) points, 12.5(10.0, 30.0) points, (19.0 ± 11.5) points and 15.6(6.3, 25.0) points in the prone group, showing no significant differences ( P>0.05). The total incidence of complications was 8.3% (2/24) in the supine group and 3.3% (1/30) in the prone group, showing no significant difference either ( P>0.05). Conclusion:In the treatment of posterior pilon fractures, as the modified posteromedial approach combined with the anterolateral approach in the supine position is equivalent to the posteromedial and the posterolateral approaches in the prone position in terms of reduction quality, bone union time, functional outcomes and complications, it can be used as an alternative choice.

Enterotoxigenic Escherichia coli（ETEC）infection can induce watery diarrhea，leading to dehydration，electrolyte disturbance，and even death in severe cases. Recombinant B subunit/inactivated whole-cell cholera（rBS/WC）vaccine is effective in preventing ETEC infectious diarrhea. On the basis of the latest evidence on etiology and epidemiology of ETEC，as well as the effectiveness，safety，and health economics of rBS/WC vaccine，National Clinical Research Center for Child Health（The Children’s Hospital，Zhejiang University School of Medicine）and Zhejiang Provincial Center for Disease Control and Prevention invited experts to develop expert consensus on rBS/WC vaccine in prevention of ETEC infectious diarrhea. It aims to provide the clinicians and vaccination professionals with guidelines on using rBS/WC vaccine to reduce the incidence of ETEC infectious diarrhea.

ObjectiveTo investigate the infection of mouse norovirus (MNV) in experimental mice raised under natural conditions from 19 biological companies in Beijing. MethodsThe mice used in this study were randomly selected from mice produced by 19 companies, and 14 mice of each strain and batch were combined into one cage, totaling 1 396 cages of 19 544 mice. The fecal samples from BALB/c, C57BL/6, ICR, KM, and BALB/c-nude mice were collected. TaqMan probe fluorescence quantitative PCR method was used to detect MNV infection of mice with MNV-1 primer, and whether the mice were infected with MNV was determined according to cycle threshold (Ct value). The chi-square test was used to analyze the difference of positive rate among the fecal samples from the five types of mice. The Ct values of the positive samples were statistically described; the non-parametric test was used to analyze the differences in Ct values among the five types of mice. Results A total of 1 396 fecal samples were collected. The positive rates of fecal MNV detection in BALB/c, C57BL/6, ICR, KM, and BALB/c-nude mice were 17.65%, 39.33%, 10.57%, 18.32% and 27.4%, respectively. According to the chi-square test results, the positive rate of fecal in C57BL/6 mice was higher than that in BALB/c, ICR, and KM mice (all P<0.05), and the positive rate of BALB/c-nude mice was higher than that in ICR and BALB/c mice (P<0.001, P＜0.05) . The viral load of BALB/c-nude or C57BL/6 mice was generally greater than that of KM mice (P<0.05). ConclusionMNV-1 primers can be applied to the detection of MNV infection in mice. The positive rate of MNV in five types of experimental mice in Beijing ranges from 10% to 40%, among which C57BL/6 mice and BALB/c-nude mice have higher positive rates of MNV than the others.

Both cholinergic dysfunction and protein citrullination are the hallmarks of rheumatoid arthritis (RA), but the relationship between the two phenomena remains unclear. We explored whether and how cholinergic dysfunction accelerates protein citrullination and consequently drives the development of RA. Cholinergic function and protein citrullination levels in patients with RA and collagen-induced arthritis (CIA) mice were collected. In both neuron-macrophage coculture system and CIA mice, the effect of cholinergic dysfunction on protein citrullination and expression of peptidylarginine deiminases (PADs) was assessed by immunofluorescence. The key transcription factors for PAD4 expression were predicted and validated. Cholinergic dysfunction in the patients with RA and CIA mice negatively correlated with the degree of protein citrullination in synovial tissues. The cholinergic or alpha7 nicotinic acetylcholine receptor (α7nAChR) deactivation and activation resulted in the promotion and reduction of protein citrullination in vitro and in vivo, respectively. Especially, the activation deficiency of α7nAChR induced the earlier onset and aggravation of CIA. Furthermore, deactivation of α7nAChR increased the expression of PAD4 and specificity protein-3 (SP3) in vitro and in vivo. Our results suggest that cholinergic dysfunction-induced deficient α7nAChR activation, which induces the expression of SP3 and its downstream molecule PAD4, accelerating protein citrullination and the development of RA.