To investigate the clinical characteristics, drug resistance types and risk factors of Klebsiella pneumoniae infection in elderly patients. The clinical data from 322 elderly patients with community-acquired pneumonia (CAP) who admitted to Renhe Hospital of Baoshan District Shanghai from 2021 to 2023 were collected, and the patients were divided into a Klebsiella pneumoniae-infected group (KP group, 56 patients) and a non-Klebsiella pneumoniae-infected group (NKP group, 266 patients). Multifactorial logistic regression analysis was used to identify risk factors and the receiver operating characteristic curve (ROC) was assessed to the clinical value of indicators for Klebsiella pneumoniae infection. The results showed that Klebsiella pneumoniae exhibited high rates of resistance to ampicillin, cefuroxime, ceftazidime and piperacillin. Multifactorial analysis showed that males, respiratory failure, solid shadows in the lungs, and high fasting glucose levels were independent risk factors for Klebsiella pneumoniae infection. ROC curve showed that the area under the curve (AUC) for assessing the risk of infection for males, respiratory failure, and high fasting glucose levels exceeded 0.6, and the AUC of the combined indicator of the four independent risk factors reached 0.762. The elderly CAP population with Klebsiella pneumoniae demonstrates specific clinical characteristics and faces a high rate of drug resistance. Being male, high fasting glucose, respiratory failure and having solid shadows in the lungs were independent risk factors for infection with Klebsiella pneumoniae. Therefore, early identification and implementation of effective treatment and management strategies for these high-risk groups is essential in clinical practice.

Abstract: The purpose of this study is to understand the molecular characteristics of Salmonella, reveal the genetic relationship among the strains, and provide a certain basis for the origin of Salmonella outbreak. Virulence and drug resistance genes of 105 Salmonella strains isolated from Changchun in recent years had been screened, by whole genome sequencing and bioinformatics methods. The wgSNP, MLST and cgMLST genotyping were performed for 105 Salmonella. 144 virulence genes were analyzed on this study. The results showed that the gene of Salmonella Pathogenicity Island 1 (SPI) was relatively stable.The carrying rate of the virulence factors outside the secretion varied greatly with serotype.69 drug resistance genes of Classes were screened, with high carrying rate of Aminoglycosides (100%) , β-lactams(73.33%) , Sulfonamides (72.38%), and low carrying rate of Macrolides (6.67%) and Lincosamides (0.95%) . The cgST and MLST types of 105 strains of salmonella were identical and divided into 13 species. The major prevalent strains, Salmonella enteritidis (ST11) and Salmonella I 4, [5] , 12: I:-(ST34) , were essentially in the same branch as the international prevalent strains. This study confirmed that Salmonella carries stable virulence genes and multi-drug resistance genes in Changchun. The major prevalent strains were distributed in different time and source. The molecular characteristics of the two strains were consistent with those of the foreign prevalent strains. The serotypes were closely related to ST and cgST types.

B lymphocyte response patterns vary in different infection scenarios, such as acute or chronic infections. This study aimed to construct recombinant Armstrong and Clone13 strains of lymphocytic choriomeningitis virus (LCMV) to induce the immune response of KL25HL B cells carrying the transgenic B cell receptor (BCR) specific to the glycoprotein (GP) of the WE strain, thereby establishing a model for comparing host B cell immune response differences between acute and chronic infection processes. Firstly, by co-transfecting cells with plasmids to transcribe LCMV genome fragments and express viral proteins, recombinant LCMV-Armstrong (rARM) and recombinant LCMV-Clone13 (rCL13) were successfully rescued. Sequencing analysis results indicated that the GP sequences of these two recombinant viruses were accurately replaced with the GP sequence of the LCMV-WE strain, and they were confirmed to have plaque-forming ability after infecting cells. For further characterizing the properties of rARM and rCL13, they were used to infect C57BL/6J mice. The results suggested that these two recombinant viruses retained the acute or chronic infection properties of wild type LCMV-Armstrong (wtARM) or wild type LCMV-Clone13 (wtCL13) strains. Specifically, after establishing infection, rARM would be rapidly cleared by the host immune system within a week, while rCL13 would persist in the host for more than two months. Finally, wild-type mice adoptively transferred with KL25HL B cells were infected with different LCMV strains to study the immune response. The results showed that, compared with wtARM or wtCL13, infection with rARM or rCL13 could significantly induce the activation and proliferation of KL25HL B cells. The rARM and rCL13 constructed in this study, after infecting mice that were adoptively transferred with KL25HL B cells, could serve as a powerful tool for comparative research on B lymphocyte response patterns in acute and chronic infections, which is of great significance for deepening understanding of humoral immune response mechanisms.

Objective To analyze the clinical drug-sensitivity results, types of drug resistance Staphylococcus aureus  (S.aureus)  isolated from the patients of nosocomial infection and risk factors for nosocomial infection over two years, and to explore its reference value in clinical control of the occurrence of drug-resistant S. aureus, hospital infection and rational application of antimicrobial drugs.  Methods  The clinical characteristics and drug resistance of drug-resistant S. aureus isolated in 2020-2021 were analyzed, and independent risk factors of nosocomial infection caused by drug-resistant S. aureus were explored using univariate and logistic regression.  Results  Multi-drug resistant S. aureus accounted for 49.7% (69/139), MRSA (methicillin-resistant Staphylococcus aureus) strains accounted for 41.0% (57/139), β-lactamase-producing strains accounted for 72.7% (101/139), and clindamycin-induced resistant strains accounted for 15.1% (21/139). S. aureus was mainly isolated from respiratory and wound infection sites (36.0% and 48.9%), and it had a high rate of resistance to a variety of antimicrobial drugs, with 100% sensitivity to vancomycin and tigecycline, the sensitivity rate to linezolid and chloramphenicol was 99.28% and 93.85%. Regression analysis showed an OR of 3.184 for invasive manipulation, with a P value < 0.05.  Conclusion   S. aureus was mainly isolated from wound site specimens, with a high occupancy of clinical multi-drug resistant bacteria, mostly MRSA and β-lactamase-producing strains, and a high prevalence of common antibiotic resistance. Invasive manipulation was an independent risk factor for drug-resistant S. aureus infection.

Tuberculosis remains a chronic infectious disease that poses a serious risk to human health and the work to control tuberculosis remains very challenging. Host-directed therapy may be a new anti-tuberculosis treatment method worthy of attention and exploration. The purpose of this study was to investigate the regulatory role of GPR84, a member of the G protein-coupled receptors (GPCRs) family, in mycobacterium bovis Bacillus Calmette-Guerin (BCG) infection. In this study, wild-type C57BL/6 mice (WT Group) and Gpr84 gene-deficient mice (Gpr84-/- group) were used as controls to establish a high-dose BCG infection mouse model, the differences in the amount of bacteria in lungs, spleen, liver, kidney and other organs and the severity of lung pathological damage between Gpr84 gene-deficient mice and wild-type mice were evaluated by CFU experiment, lung tissue HE staining and acid-fast staining, the changes of body weight and general condition of mice in the two groups were observed. The results showed that the body weight of Gpr84 gene-deficient mice infected with BCG was significantly lower than that of uninfected mice. The CFU results and antacid staining showed that bacterial load in lung tissues of Gpr84-/- group was higher than that of WT group, and the differences of bacterial load in spleen and liver were consistent with those in lung. HE staining showed that the normal alveolar structure of lung tissues of Gpr84-/- group was significantly more damaged than that of WT group after BCG infection, and the number of lesions was more than that of WT group. Flow cytometric analysis of lung cells of mice showed that in the absence of infection, the knockout of Gpr84 had no significant effect on the number of resident alveolar macrophages (AMs) and interstitial macrophages (IMs) in the lungs, and in acute infection, the proportion of AMs in the lungs of mice in Gpr84-/- group was significantly higher than in WT group, while the proportion of IMs was significantly lower than in WT group, the proportion of mononuclear phagocytes (MNPs) Ly6Chi subpopulation was increased, the number of Ly6Clo subpopulation was decreased, and neutrophils (Neuts) infiltration was increased. The results revealed that the aggravated bacterial load and pathological damage after Gpr84 knockout may be related to the reduction of the relative ratio of IMs/AMs and the increase of the Ly6Chi MNPs subgroup, affecting its transformation to the Ly6Clo subgroup and increasing the infiltration of Neuts. Therefore, Gpr84 has the potential to be one of the candidate targets for host-directed therapy of tuberculosis by regulating immune function.

Objective To propose a set of referable and replicable standardized management system of sensory control technology, so as to provide reference for promoting the rational application of antibiotics in hospitals and curbing bacterial drug resistance. Methods With the establishment of a microbiological monitoring management system and rational application management system of antibiotics as the starting point and the cultivation of professional talents as the carrier, the management strategy based on the linkage of hospital sense, medical, clinical, pharmaceutical, laboratory and nursing departments was formed and continuously optimized. The detection situations of multi-drug resistant bacteria, occurrence of nosocomial infection, relevant indicators of rational use of antibiotics in our hospital from January 2018 to December 2022 were retrospectively analyzed to evaluate the implementation effect of this strategy. Results In the process of constructing and gradually improving the multi-department linkage management strategy, the pre-application detection rate of inpatient antibiotics increased from 61.32% to 64.17% from 2019-2022 compared with 2017-2019, and the infection rate of multi-drug resistant bacteria MRSA, CRKPN, CRAB and CRPA in hospitals showed a significant decline. The intensity of antibiotic use during hospitalization decreased from 43.32 (2018) to 40.39 (2022). Conclusion The multi-department linkage management system based on precision infection control proposed in this study can effectively promote the rational application of antibiotics and curb bacterial drug resistance, and this system can be used as a reference for hospital infection control departments to promote and replicate in medical institutions.

Abstract：Objective To investigate the risk factors and clinical characteristics of lung abscess patients with high virulence Klebsiella pneumoniae infection to provide more reference for early identification of high virulence infection high-risk groups. Methods 35 lung abscess patients with Klebsiella pneumoniae infection were retrospectively chosen in the period from January 2019 to January 2022 and grouped according to the types of Klebsiella pneumoniae infection into high virulence group (23 cases) and classical group (12 cases). The clinical characteristics, imaging data, drug resistance and clinical characteristics of 2 groups were compared. The independent risk factors of highly virulent Klebsiella pneumoniae infection in lung abscess were evaluated by Logistic regression model multivariate method. Results The proportion of high virulence combination with diabetes mellitus, extrapulmonary abscess, multiple infections, respiratory tract infection as the first symptom and multiple abscess as indicated by imaging were significantly higher than classical group（P<0.05）. The proportion of indignant catheters and invasive procedures before infection in the high virulence group were significantly lower than classical group（P<0.05）. Multivariate analysis of Logistic regression model showed that combined diabetes were independent risk factors of highly virulent Klebsiella pneumoniae infection in lung abscess（P<0.05）. The drug resistance rates of cefuroxime, ceftriaxone, ceftazidime, cefepime, amoxicillin/clavulanate, piperacillin/tazobactam, cefoperazone/sulbactam and levofloxacin in high virulence group were significantly lower than classic group（P<0.05）. The proportions of virulence genotype aero and rmpA+aero in high virulence group were significantly higher than classic group（P<0.05）. Conclusion High virulence Klebsiella pneumoniae infection in lung abscess was closely related to diabetes mellitus. Meanwhile, the main virulence genotype of this infection was aero.

In this study, a strain of Getah virus (GETV, GenBank:OM363683) was successfully rescued using a reverse genetics approach. First, the optimized viral sequence was cloned into the low-copy plasmid pFK containing either the T7 or CMV promoter, named T7-GETV and CMV-GETV, respectively. Viral RNA was synthesized via in vitro transcription (IVT) using T7-GETV as a template, followed by RNA transfection into BHK-21 cells via liposome-mediated transfection. The viral titer was determined using 50% tissue culture infectious dose (TCID50) method and the expression of viral protein E1 was detected through Western blot (WB), validating the successful rescue of GETV. Further, the cytopathic effect (CPE) was observed via light microscope, and the growth characteristics of GETV were systematically analyzed using the TCID50 assay, real-time quantitative polymerase chain reaction (RT-qPCR), and WB. Additionally, based on the structure of GETV replicon, two replicative expression vectors were constructed using CMV-GETV as a template by retaining the non-structural protein genes and replacing the structural protein genes with the reporter gene mCherry or Renilla luciferase (Rluc), named pFK-GETV-mCherry and pFK-GETV-Rluc, respectively. Two control plasmids were also constructed by inserting only the reporter gene mCherry or Rluc downstream of the CMV promoter in the pFK plasmid, named pFK-mCherry and pFK-Rluc, respectively. Analysis with Image J software revealed that the average fluorescence intensity per cell of cells transfected with pFK-GETV-mCherry was approximately 5 times higher than that of cells transfected with pFK-mCherry. RT-qPCR results indicated that mCherry gene transcription level in cells transfected with pFK-GETV-mCherry was also about 5 times higher than that in cells transfected with pFK-mCherry, and both results were highly consistent. WB results indicated that mCherry protein expression level in cells transfected with pFK-GETV-mCherry was also higher than that in cells transfected with pFK-mCherry at the same time point. Renilla luciferase assay showed that Rluc expression efficiency in cells transfected with pFK-GETV-Rluc was about three orders of magnitude greater than that in cells transfected with pFK-Rluc. This study provides a robust reverse genetics tool for GETV research and demonstrates the ability of the GETV replicative expression vector to enhance exogenous gene expression, establishing a foundation for the development of replicative DNA vaccines.

This review aims to provide a comprehensive overview of the advancements in Nipah virus animal models, enhancing the understanding of Nipah virus infection pathways, pathogenic mechanisms, and the targeted development and evaluation of vaccines and antibodies. Nipah virus is a zoonotic paramyxovirus with a high fatality rate and is classified as a Biosafety Level 4 pathogen. Since its first outbreak in Malaysia in 1998, it has continued to cause epidemics in Southeast and South Asia, leading to severe respiratory diseases and encephalitis in humans and animals, with no approved vaccines or therapeutic drugs currently on the market. The World Health Organization has prioritized Nipah virus disease for research and development. This review summarizes the construction and development of Nipah virus infection animal models, with multiple species having been utilized in the development of Nipah virus infection models, such as golden hamsters, ferrets, pigs, African green monkeys, and guinea pigs, which reflect the symptoms and pathological characteristics of Nipah virus infection to varying degrees, as well as the advantages and limitations of the models. The review also discusses the establishment and application of pseudovirus animal infection models and looks forward to the future direction of Nipah virus animal model research.

The Type VII secretion system (T7SS) is the only secretion system exclusively found in Gram-positive bacteria. It closely links to bacterial growth and virulence, which plays an important role in the interaction between bacteria-bacteria, bacteria-host, and bacteria-environment. Overall, the functional diversity of T7SS is associated with its effector proteins. Therefore, the article will discuss the effector proteins secreted by T7SS, providing a comprehensive review of their structure and function, secretion mechanisms, and the regulatory modes of gene transcription, with the aim to offer new perspectives for further research into bacterial T7SS.

Zika virus (ZIKV) is a kind of mosquito-borne pathogen. Its infection can cause Zika virus disease. There is currently no specific treatment. Autophagy is a process in which cells maintain their own homeostasis and promote the reuse of intracellular substances and energy, but it is often hijacked by some viruses to promote their own replication. In this study, human hepatoma cell line (Huh7) was used as the target cell, and the mTagRFP-mWasabi-LC3 dual fluorescent reporter system was used to detect the formation of autophagosomes. Western blotting was used to detect the expression of autophagy markers LC3 and P62 proteins after ZIKV infection at different time points. Cells were pretreated with autophagy inhibitors (wortmannin and chloroquine), Beclin-1 specific short hairpin RNA (shRNA) and autophagy inducer rapamycin. Then their effects on viral protein expression and extracellular progeny virus titer were detected by Western blotting and virus plaque assay. The signaling pathways involved in the regulation of autophagy in ZIKV infection were detected by Western blotting. The results showed that the number of LC3-positive autophagosomes increased significantly in Huh7 cells after infection by ZIKV. The expression of LC3-II increased significantly from 12 h post infection, and P62 protein was significantly down-regulated at 48 h post infection. Both autophagy inhibitor treatments and interference of Beclin-1 expression could effectively inhibited ZIKV infection and the production of progeny viruses. Rapamycin treatment increased the content of extracellular progeny viruses (P<0.05). ZIKV infection enhanced the phosphorylation of adenosine 5’-monophosphate-activated protein kinase (AMPK) and TSC2, and reduced the phosphorylation of mammalian target of rapamycin (mTOR). The results demonstrate that ZIKV infection induces autophagy by activating the AMPK/TSC2/mTOR signaling pathway in Huh7 cells, and utilizes autophagy to promote virus replication and progeny virus production. It provides theoretical and experimental basis for the development of anti-ZIKV drugs based on autophagy-related molecules.

Yellow fever (YF) is an acute hemorrhagic infectious disease transmitted by Aedes mosquitoes, and is endemic mainly in tropical regions such as West Africa and South America. The clinical manifestations of YF include high fever, nausea, vomiting, jaundice, hemorrhage, etc. The etiological pathogen of YF is yellow fever virus (YFV), which is a positive-sense single-stranded RNA virus and belongs to the flavivirus genus of Flaviviridae family. At present, there is no specific antiviral drug for YF, and the general treatment is mainly based on symptomatic and supportive care. Vaccination of an live attenuated yellow fever vaccine (YF-17D) is the most effective way of prevention. Although, YF-17D vaccine has been widely applied and recognized globally, some adverse reactions have been reported occasionally. This review summarizes the epidemic situation, vaccine research progress and application of YFV in current years.

To understand the prevalence of rabies, Toxoplasma gondii and influenza A virus in stray cats in Qingpu, Shanghai. Serum samples from 153 stray cats collected from 40 residential community between June 2022 and August 2010 were tested for antibodies to rabies, Toxoplasma gondii and influenza A viruses by ELISA. The H3, H5, H7 and H9 serotypes of influenza A antibody positive samples were identified by HI test. Real-time PCR was used for the detection of Toxoplasma gondii RNA in whole blood samples and influenza virus RNA in throat and anal swab samples. The results showed that the positive rates of rabies, Toxoplasma gondii and influenza A were 14.38% (22/153) , 11.76% (18/153) and 3.92% (6/153) respectively. The difference of positive rate of Toxoplasma gondii antibody between spring (March-May) and summer (June-August) was very significant. The positive rate of H3 was 0.65% (1/153) . No H5, H7 and H9 antibody was detected. The results showed no Toxoplasma gondii antigen was detected in the whole blood samples ,and no influenza antigen was detected in throat swab samples and anal swab samples. The results suggest that the rabies antibody level of stray cats in Qingpu District was low, which was not enough to block the Rabies transmission. Toxoplasma gondii infection exists in stray cats, especially in summer. The presence of influenza A infection in stray cats suggests that stray cats can act as a "Mixer" for human and animal influenza viruses, posing a risk of zoonosis transmission. Therefore, it is necessary to continuously monitor the infection of rabies, Toxoplasma gondii and influenza A virus in stray cats to provide basis for scientific prevention and control.

This study aims to investigate the species distribution and drug resistance of pathogenic bacteria in the children with bloodstream infections in Shanghai Children’s Hospital from 2016 to 2018, and to provide rational drug use basis for clinical prevention and treatment of bloodstream infections. The automatic microbiological analysis system was used to identify the pathogenic bacteria isolated from the blood culture samples and conduct drug sensitivity test to analyze their distribution characteristics and drug resistance. A total of 24 687 samples were sent for examinations, finally 2 092 pathogenic bacteria were isolated with a positive detection rate of 8.5%. Gram-positive bacteria accounted for 84.4%, and Gram-negative bacteria accounted for 15.6%. The top five pathogens were coagulase-negative Staphylococcus (CoNS), Enterococcus, Klebsiella pneumoniae, Streptococcus and Escherichia coli. The positive detection rate of blood culture in Neonatal Department was 34.5%, and the isolation rate of CoNS was significantly higher than that in other departments. The average time to positivity of CoNS was (32.53±12.36) h. No Staphylococcus and Enterococcus resistant to vancomycin and linezolid were found. Imipenem, meropenem, amikacin, cefoperazone-sulbactam and piperacillin-tazobactam had strong antibacterial activity against Escherichia coli, and the drug resistance rate was less than 10%. The drug resistance rate of Klebsiella pneumoniae to third-generation cephalosporins reached more than 80%, and the drug resistance rate to carbapenems increased year by year, reaching 62.5% in 2018. The drug resistance rate of Pseudomonas aeruginosa to carbapenems was 9.1%. The drug resistance rate of Acinetobacter baumannii to most antimicrobial agents was more than 60%, and to carbapenems was 66.7%. It is shown that pathogenic bacteria from blood culture of children with bloodstream infections are complex, and CoNS accounts for a large proportion, which makes the diagnosis of bloodstream infections become more complicated. Therefore, understanding the distribution characteristics and drug resistance of pathogenic bacteria from blood culture of children with bloodstream infections is helpful to guide the rational use of antibiotics in clinical application.

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (Mtb) . With the emergence of multidrug-resistant TB (MDR-TB) and extensively drug-resistant TB (XDR-TB), the treatment of TB has become more severe. In recent years, studies have found that the existence of efflux pumps in Mycobacterium tuberculosis is one of the reasons for its drug resistance. The efflux pumps of major facilitator superfamily (MFS), ATP-Binding Cassette (ABC), resistance-nodulation-division (RND) and small multidrug resistance (SMR) have been found in Mtb. However, the drug resistance mediated by Mtb efflux pumps still remains largely unknown, and there is not much work to develop efflux pump inhibitors from the perspective of new drug discovery. This article reviews the structure and function of the efflux pumps of ABC, MFS, RND and SMR of Mtb, as well as the research progress of efflux pump inhibitors of Mtb.

Objective To report a rare case of bacteremia caused by Lactobacillus salivarius in gastric cancer patient with liver metastasis, and to discuss the risk of opportunistic infection caused by Lactobacillus salivarius. Methods The diagnosis and treatment of bloodstream infection in this patient were retrospectively analyzed. VITEK 2 compact automatic bacterial analyzer and Antu autof ms1000 automatic microbial mass spectrometry detection system were compared, and 16S rRNA sequencing was also used to identify the strains. TDR ANA-AST was used for drug sensitivity test to provide clinical medication guidance. Results Three bottles of blood culture were positive in 2-3 days after operation. Gram positive bacilli were detected by direct smear and colonoscopy. The strains was identified as Lactobacillus acidophilus by VITEK 2 compact. Antu biological Autof ms1000 automatic microbial mass spectrometry detection system and 16S rRNA sequencing were all identified the strains as Lactobacillus salivarius. After using imipenem for anti infection treatment, the blood culture was negative. Conclusion Lactobacillus salivarius was isolated from the blood culture of a gastric cancer patient with liver metastasis in Shanghai East Hospital. The drug sensitivity test provides guidance for clinical drug use.

Mouse hepatitis virus (MHV) is a model coronavirus, but the traditional etiological methods are limited to the viruses isolated from nature. The reverse genetic system can rapidly obtain the recombinant virus for research, creating a new method to study the genomic functions of MHV and other coronaviruses. In this study, whole genome of the original MHV A59 strain was divided into six fragments by reverse transcription-polymerase chain reaction (RT-PCR). All fragments were individually cloned into plasmid vectors, then the full-length genomic RNA was obtained by restriction enzyme digestion, in vitro ligation and in vitro transcription. After RNA was electroporated into cells, the recombinant virus was rescued. The results revealed that the replication characteristics of recombinant virus rA59 is similar to the original strain, providing an experimental tool for studying the biological characteristics of the virus. To detect the virus replication level, the recombinant reporter viruses rA59-ZsGreen and rA59-Nluc were obtained by inserting the green fluorescent protein ZsGreen (Zoanthus sp. green fluorescent protein) and luciferase Nluc (NanoLuc luciferase) into the open reading frame 4 (ORF4) genome, respectively. The results indicate that the reporter genes do not compromise viral replication and the reporter viruses can well reflect the viral replication property of their parental recombinant virus. These two viruses were used to verify the antiviral activity of remdesivir, by detecting the fluorescence intensity or luciferase activity after treatment. The results demonstrate that the recombinant reporter viruses rA59-ZsGreen and rA59-Nluc are suitable for high-throughput drug screening and have promising potential for application. In summary, the successful establishment of the 6-plasmid-based reverse genetic system for MHV strain A59 provides a powerful tool for studying the biological characteristics of MHV and testing antiviral drugs.

The single-cell study of hepatitis B virus (HBV) in liver tissue is of paramount significance. However, the associated research technology is still in an advanced stage and represents an unmet clinical need. Based on ddPCR (digital droplet polymerase chain reaction), this study has pioneered a single-cell ddPCR (sc-ddPCR) method for the precise quantification of HBV DNA and RNA at the single-cell level. To ascertain the linearity and lower detection limit of HBV DNA, a series of experiments were conducted by simulating a chronic HBV infection in the liver through the mixing of various proportions of HBV-positive and negative cells. The study focused on the detection of episome-derived RNA (eRNA), developing a tailored protocol for its analysis. By capitalizing on the structural disparities in HBV transcripts, the study had formulated a specific primer and probe system for eRNA, the specificity of which was rigorously confirmed in HBV liver cancer cell lines. The linearity and lower detection limit of eRNA were scrutinized through the utilization of gradient-diluted standard cell lines. The results unequivocally demonstrated the establishment of an advanced single-cell detection method for HBV DNA and RNA based on ddPCR, characterized by exceptional linearity (HBV DNA: R²＝0.998 7; eRNA: R²＝0.942 5). Moreover, the method exhibited a remarkable sensitivity, as it consistently detected positive signals in gradient-diluted standard cell lines, with a lower detection limit of 0.16% for HBV DNA and 0.2% for eRNA. The sc-ddPCR method developed in this study is versatile, enabling simultaneous detection of DNA and RNA, and it serves as a robust technical foundation for further exploration of HBV virology activities within liver tissue. It is also invaluable for optimizing and advancing HBV treatment strategies, presenting promising prospects for broad applications.

As the research progresses on intestinal flora, more and more evidences show that the imbalance of intestinal flora is closely related to the occurrence and progression of many chronic diseases. Probiotic treatment has become a worldwide hotspot, and Akkermansia muciniphlia (A. muciniphila), a common colonizer in the mucous layer of human intestines, has gradually been regarded as a promising candidate for the next-generation probiotics. The present paper summarizes the ameliorating effects of A. muciniphila on chronic diseases and the possible mechanisms, providing a new idea for the treatment of diseases.

Fecal microbiota transplantation (FMT) is an innovative method to reconstruct the intestinal microecological balance by transplanting the fecal microbiota of healthy donors into patients, which provides help for the treatment of diseases. It has a good effect on the treatment of recurrent Clostridioides difficile infection (rCDI), and its therapeutic potential is not only limited to gastrointestinal diseases, but also can be continuously explored in other microbial-related diseases. In the exploration of factors for the success of FMT in the treatment of diseases, in addition to the recovery of intestinal bacteria, the regulation of intestinal phages is also involved. Intestinal phages, an important part of the gut microbiota, play an important role in the complex dynamics of bacteria; their transfer may be related to the curative effect of FMT. This review analyzed the biological characteristics of intestinal phage and the main bioinformatics analysis strategies, summarized the clinical research on the role of intestinal phage in FMT, explored the changes in the intestinal phage community in FMT and the possible mechanisms of action, and discussed the relationship between intestinal phage and the efficacy of FMT, as well as the existing safety issues. It can not only improve the understanding of the role of intestinal phages in FMT, but also promote the clinical application of FMT.

In order to explore the ability of routine laboratory identification methods to identify clinical isolates of Herbaspirillum huttiense (H. huttiense), the source of infection of H. huttiense in the blood of this patient was analyzed. Automated biochemical identification system, matrix-assisted laser desorption ionization-time of flight mass spectrometer (MALDI-TOF MS), 16S rRNA gene sequencing and rpoB gene sequencing were used to identify H. huttiense, and the specimens from multiple parts of the patient were detected. The results showed that the automatic biochemical identification system misidentified H. huttiense as Burkholderia cepacia. MALDI-TOF MS and 16S rRNA gene sequencing were not able to distinguish H. huttiense from Herbaspirillum aquaticum and Herbaspirillum camelliae, while rpoB gene sequencing could accurately identify H. huttiense. In addition to the blood, only a small amount of H. huttiense was detected in the patient’s feces. The results of enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) showed the same DNA band patterns of H. huttiense from blood and feces. MALDI-TOF MS main spectra projection(MSP)cluster analysis showed that they were located at the same node, and the distance level was under 50, with the same resistance phenotype. The blood and fecal origins of H. huttiense were highly homologous. This study demonstrated the performance of automated biochemical identification system, MALDI-TOF MS, 16S rRNA gene sequencing and rpoB gene sequencing for identification of H. huttiense, which confirmed that H. huttiense could infect gastrointestinal tract through contaminated food and cause sepsis in patients with postoperative chemotherapy for rectal adenocarcinoma.

Mycobacterium tuberculosis (M. tuberculosis) is the pathogen of tuberculosis, a global epidemic and chronic infectious disease. Its pathogenic mechanism and drug resistance are closely related to its specific cell wall components, especially lipids, such as lipoarabinomannans, trehalose-6, 6’-dimycolates, phenolic glycolipids and other glycolipids. It is of great value to study the lipids of M. tuberculosis cell wall for the diagnosis and prevention of tuberculosis. In order to provide a reference for the related basic research of M. tuberculosis, this review summarized the recent progress on the bioactivity of various lipids of M. tuberculosis cell wall.

To analyze the distribution and drug sensitivity of pathogenic bacteria of bloodstream infections in patients hospitalized in Xinchang County Hospital of Traditional Chinese Medicine to provide a basis for the diagnosis and treatment of clinical bloodstream infections. All blood culture samples from Xinchang County Hospital of Traditional Chinese Medicine were collected from January 2019 to December 2021, and The pathogen species and drug sensitivity in vitro were analyzed statistically. 10,879 blood culture samples were collected, of which 787 were culture-positive (7.23%), mainly from the intensive care unit. The pathogens detected included 760 aerobic (96.57%), 5 anaerobic (0.64%) and 22 fungal (2.80%) strains; the gram-negative bacteria (40.03%) were mainly Escherichia coli, Klebsiella pneumoniae and Aspergillus chimaerae, and the gram-positive bacteria (55.78%) were mainly Staphylococcus epidermidis, Staphylococcus aureus and Staphylococcus hominis. Drug sensitivity results showed that Escherichia coli and Klebsiella pneumoniae were more than 80% resistant to ampicillin and more than 80% sensitive to cephalosporins; Staphylococcus epidermidis, Staphylococcus aureus and Staphylococcus hominis were more than 70% resistant to penicillin and more sensitive to antimicrobial drugs such as linezolid and tigecycline (greater than 90%). Blood culture positive samples are mainly from immunocompromised patients' wards, and the pathogenic bacteria are mainly Escherichia coli, Klebsiella pneumoniae, Staphylococcus epidermidis and Staphylococcus aureus which are sensitive to some antibacterial drugs, but with the application of antibiotics in treatment, drug resistance will change, and the clinic should strengthen the detection of pathogenic bacteria and drug susceptibility of bloodstream infections to provide a basis for the diagnosis and treatment of bloodstream infections.

To study the effect of SopE, a type III secretory system effector encoded by Salmonella enterica serovar Typhi, on SCV induced by S. Typhi after invasion of macrophages. S.Typhi GIFU1007was used as the wild type (WT) strain to construct the sopE mutant (ΔsopE) by homologous recombination using the suicide plasmid pGMB151. The ΔsopE complementary strain (C-ΔsopE) was constructed with the plasmid complementary method by using pBAD33.The recombinant plasmid pET28-SFGFP was introduced into the WT, ΔsopE and C-ΔsopE , respectively, to yield WT/pBAD33::pET28-SFGFP, ΔsopE/pBAD33::pET28-SFGFP and C-ΔsopE/pET28-SFGFP. The effect of SopE on the intracellular viability of S. Typhi was studied by THP-1 intracellular viability assay of macrophages. The effect of SopE on the stability of late SCV was investigated by immunofluorescence and real-time quantitative PCR (qRT-PCR). The relationship between SopE and autophagy was further investigated by qRT-PCR and Western blot. The intracellular viability of THP-1 macrophages with ΔsopE/pBAD33::PET28-SFGFP was significantly lower than that of WT/pBAD33::PET28-SFGFP and C-ΔsopE/PET28-SFGFP. The immunofluorescence assay and qRT-PCR results showed that deletion of sopE decreased the stability of late SCV. The qRT-PCR and Western blot results showed that SopE inhibited the expression of autophagy related genes. The data presented here showed that SopE regulated the stability of late SCV by affecting autophagy.

The human vaginal microbiota (VMB), which plays a vital role in maintaining health and homeostasis, exhibits low diversity compared to the microbiomes of other organs. Cervical cancer (CC) is a common malignancy in women and has been shown to be highly associated with persistent infection with high-risk human papillomavirus (HPV). VMB is associated with human papillomavirus infection and cervical lesions, and may play a positive role in the progression of HPV infection and cervical intraepithelial neoplasia (CIN). On the one hand, lactobacillus can reduce the permeability of cervical cells, and reduce inflammatory response, inhibit the growth of cervical cancer cells, but the bacterias enhancing the diversity of VMB can express virulence and attachment genes, which cause damage to cervix and epithelial cells, causing HPV infection and high-grade disease states; On the other hand, HPV-associated E7 oncoprotein can reduce the secretion of defense peptides which are conducive to the growth of lactobacillus through NF-β-κB and Wnt/β-catenin signaling pathways, resulting in an increase in vaginal pH, which further facilitates the growth of vaginal pathogenic bacteria, and ultimately leads to structural changes of VMB. This review focuses on the relationship between the vaginal microbiome, persistent HPV infection and cervical dysplasia and the factors that mediate these relationships, which will help to find new targets for VMB-related diseases.

The present paper aims to construct the eukaryotic overexpression vector and knockdown vector of LncRNA-GAS5 (long non-coding RNA-growth arrest-specific transcript 5), and to study its role in the expression of SNAP29 in HeLa cells infected by Acinetobacter baumannii (A. baumannii). LncRNA-GAS5 gene was amplified by polymerase chain reaction (PCR) and cloned into pcDNA3.1 (＋) eukaryotic expression vector to form pcDNA3.1-GAS5 eukaryotic expression vector. Through RNA interference technology, the oligonucleotide sequences for knockdown of GAS5 were designed and constructed into pRNAT-U6.1/Neo vector to form pRNAT-U6.1-GAS5 knockdown vector. The expression of GAS5 was detected by real-time PCR to determine whether the overexpression and knockdown were successful or not. Western blotting was used to detect the changes of SNAP29 expression in HeLa cells after overexpression or knockdown of GAS5. In addition, the expression of SNAP29 in A. baumannii-infected HeLa cells at different times and the expression of SNAP29 in A. baumannii-reinfected HeLa cells after knockdown or overexpression of GAS5 were detected by Western blotting. The results showed that pcDNA3.1-GAS5 and pRNAT-U6.1-GAS5 were successfully constructed. pcDNA3.1-GAS5 could induce high expression of LncRNA-GAS5 in HeLa cells. pRNAT-U6.1-GAS5 significantly reduced the expression of GAS5 in HeLa cells. The overexpression of LncRNA-GAS5 decreased the expression of SNAP29 in HeLa cells, while the knockdown of LncRNA-GAS5 increased the expression of SNAP29 in HeLa cells. The expression of SNAP29 decreased progressively with A. baumannii infection time. The knockdown of LncRNA-GAS5 could restore the expression of SNAP29 degraded by A. baumannii, while the overexpression of LncRNA-GAS5 could aggravate the degradation of SNAP29 by A. baumannii. The results suggest that LncRNA-GAS5 has a negatively regulatory effect on SNAP29 expression, and the degradation of SNAP29 by A. baumannii depends on LncRNA-GAS5. It lays a theoretical foundation for further discovering the deep mechanism of autophagy degradation disorder caused by A. baumannii.

This study aims to investigate the virulence, antibiotic resistance gene profiles, and molecular types of Staphylococcus aureus small colony variants (SASCVs) in periprosthetic joint infections (PJIs) to provide fundamental data for understanding their pathogenic mechanisms. A total of 19 SASCV strains isolated from synovial fluid samples of PJI patients were collected, and whole genome sequencing, bioinformatics analysis, and database comparison were performed to determine the presence of virulence and antibiotic resistance genes. Multilocus sequence typing (MLST) was employed to classify strain types and a phylogenetic tree was constructed by MEGA software. The results demonstrated that among the 19 SASCV strains, eight were found to coexist with Staphylococcus aureus with a normal phenotype. All 27 strains underwent whole genome sequencing, with genome lengths ranging from 2 696 437 bp to 3 011 803 bp and G＋C content of (32.8±0.05)%. A total of 23 virulence-related genes were detected: 14 adhesin-related genes had detection rates between 21.1% and 100.0%; four immune evasion-related genes had detection rates ranging from 36.8% to 68.4%; and five hemolytic and leukocidin genes were also identified. Twelve resistance genes were detected: blaZ was detected in 78.9% (15/19) of the samples; mecA was detected in 31.6% (6/19) of the samples; resistance genes for macrolide and aminoglycoside antibiotics were detected with varying rates; and methicillin-resistant Staphylococcus aureus (MRSA) had a higher number of detected resistance genes compared to methicillin-sensitive Staphylococcus aureus (MSSA). MLST analysis revealed 10 different sequence types (STs) among the 19 SASCV strains. There were eight groups of phenotypically mixed strains. In two groups SCVs and their normal phenotype (NP) counterparts belonged to different geno-types, with significant differences in virulence and antibiotic resistance genes, while in the remaining six groups SCVs and NP strains belonged to the same genotype. This study revealed that in PJIs, the detection rate of biofilm-related adhesion genes in SASCVs is high, and the prevalence of mecA is relatively elevated. Among the MLST genotypes, ST59 is the most common, followed by ST398 and ST25. When SCVs coexist with normal phenotype strains, they may belong to different genotypes.

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The incidence of invasive fungal infections has increased year by year. However, the current antifungal drugs are scarce and the development of new drugs is relatively slow. Therefore, it is urgent to develop new antifungal drugs. Natural products have the characteristics of wide sources and low side effects. They are one of the sources of antifungal drugs. A large number of studies have proved that many compounds isolated from plants and traditional Chinese medicine have antifungal activities, and the antifungal mechanisms are different. This review discusses the latest progress in development of natural products antifungal drugs, investigates the natural products with potential antifungal activity, and puts forward the prospect of the development of new antifungal candidate drugs.

The outbreak of Omicron variant infectious disease not only affects Shanghai’s overall medical operation mode, but also generates enormous challenges and roadblocks of the implementation of treatment work, the accomplishment of the core medical systems and regimes, and the protection of patient safety to designated hospitals in Shanghai. As one of designated hospitals of Shanghai, the Baoshan Campus of Huashan Hospital has formed various multi-level management systems under the current circumstance, including the “hospital-level medical core management work system”， “specialized treatment guarantee-related work system” and “new crown-related special work system”. With the limited medical resources, Huashan Hospital regards flexible and intelligent implementation of the core system as an important task, and has built a series of management models. This paper has illustrated practical experience, and has provided reference for the purpose of maximizing the protection of patient’s safety and properly reacting during the outbreak of epidemic.

The present paper reported a case of corneal ulcer caused by Lasiodiplodia theobromae. The male patient was 54 years old. His right eye became red and the pain increased for 3 days. Ophthalmic examination showed mixed hyperemia in the bulbar conjunctiva of the right eye. The whole cornea was gray and cloudy, representing an uneven surface like a map. Light white deposition could be seen below the anterior chamber, and the remaining conditions in the eye were not clear. Corneal curettage specimens were taken for fungal fluorescence staining and culture. The cultured strains were identified by morphology and molecular biology. The drug sensitivity of filamentous fungi was tested by E-test. Septahypha can be seen under fluorescent staining of fungi. After incubation for 14 days, conidia could be seen in the conidiophores. The pathogen was identified as Lasiodiplodia theobromae by sequencing. Drug susceptibility result showed that amphotericin B and voriconazole were relatively sensitive.The corneal ulcer was confirmed to be caused by Lasiodiplodia theobromae. The right eye was treated with fluconazole eye drops q2h, and the fluconazole sodium chloride injection was used with 0.4 g qd via intervenous drop infusion. The patient did not get better after 5 days of treatment and asked to be discharged.

Aspergillus is an opportunistic pathogen, which can cause invasive aspergillosis, chronic pulmonary aspergillosis, allergic bronchopulmonary aspergillosis, and can also cause skin, bone and other parts of the infection and aspergillus poisoning. Peritonitis caused by Aspergillus is relatively rare, but its severity and mortality are relatively high, and it is associated with high mortality and morbidity in peritoneal dialysis patients. Peritonitis caused by Aspergillus flavus has not been reported in China. We report a case of Aspergillus flavus infection secondary to a bacterial peritonitis in a patient with peritoneal penetration, and finally received effective treatment through accurate identification of isolates and accurate antimicrobial susceptibility results. The aim of this study is to deepen the understanding of Aspergillus flavus and provide important guidance for early diagnosis, timely detection, and effective treatment in clinical practice.

The aim of this study was to investigate the effects of Fusobacterium nucleatum (F. nucleatum) on proliferation and apoptosis of human colorectal cancer (CRC) cells as well as its mechanism. Human CRC cells HCT116 at logarithmic growth stage were divided into control group, F. nucleatum group (infected with F. nucleatum), si-NEAT1 group (transfected with si-NEAT1) and F. nucleatum+si-NEAT1 group (infected with F. nucleatum after transfection with si-NEAT1). qRT-PCR was used to measure the level of RNA in cells. CCK-8 assay was performed to detect the proliferation of cells, while flow cytometry was used to calculate the apoptosis rate of cells. The expression levels of apoptosis related proteins caspase-3, Bcl-2 and Bax were determined by western blot. The proliferation of cells was enhanced, and the apoptosis rate of cells was downregulated. The expressions of caspase-3 and Bax were decreased, while the level of Bcl-2 was lifted. In addition, the expression of NEAT1 in CRC cells increased after infection with F. nucleatum. After transfection with si-NEAT1, the proliferation ability of CRC cells was decreased, the apoptosis rate was increased, and the expressions of caspase-3 and Bax protein were added, while the expression of Bcl-2 was reduced. Knockdown NEAT1 expression reversed the effect of F. nucleatum on proliferation and apoptosis of CRC cells. These results suggested that F. nucleatum could promote the proliferation and inhibit the apoptosis of CRC cells by up-regulating of NEAT1.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has a serious impact on people’s health and lives. At present, SARS-CoV-2 is still widely spread, but effective treatments for SARS-CoV-2 are still limited. Therefore, it is of great significance to find broad-spectrum neutralizing antibodies to block SARS-CoV-2 infection. In this study, we successfully expressed and purified three human-mouse chimeric antibodies targeting receptor-binding domain (RBD) of SARS-CoV-2, and then the neutralizing activities of these antibodies against a variety of SARS-CoV-2 pseudoviruses and authentic virus were tested. The results showed that these three antibodies could specifically neutralize SARS-CoV-2 wild-type pseudovirus, and the IC50 values were 0.03 μg/mL, 0.06 μg/mL and 0.03 μg/mL, respectively. Furthermore, these antibodies exhibited broad-spectrum neutralizing activity against Alpha, Delta and Lambda pseudoviruses, and could also inhibit the replication of SARS-CoV-2 Delta authentic virus. Mechanism studies showed that these antibodies could block cell-cell fusion mediated by SARS-CoV-2 S protein, and K417, E484 and N501 on receptor-binding domain (RBD) may be the key sites for binding with these three antibodies.

Chronic hepatitis B (CHB) is a major risk factor for liver fibrosis, cirrhosis, and hepatocellular carcinoma (HCC). Despite the use of interferons and nucleoside analogues as treatment options, a complete cure remains elusive, making the exploration of HBV infection's immunological mechanisms, especially the role of host genetic background, crucial for developing new therapeutic strategies. This study utilized the AAV8-rcccDNA mouse model to investigate the differences in immune response to HBV infection between mice of two genetic backgrounds, FVB/N×C57BL/6 and DBA/2×C57BL/6. The results showed that FVB/N×C57BL/6 mice exhibited an immune-tolerant state, characterized by persistent HBsAg expression and low levels of HBsAb. In contrast, DBA/2×C57BL/6 mice simulated the recovery phase of acute infection, specifically showing HBsAg seroclearance, HBsAb positivity, and mid-to-low levels of viral replication. This study underscores the necessity of considering the host genetic background in HBV infection treatment and research and highlights the potential of adaptive immune system regulation in achieving HBV cure. The improvement of the AAV8-rcccDNA mouse model provides an effective tool for simulating the human immune state during HBV infection, facilitating the optimization and development of HBV treatment strategies.

To analyze the clinical characteristics, pathogens and drug sensitivity of children urinary tract infection (UTI), so as to help pediatricians better manage UTI in children, clinical data of 591 hospitalized children with UTI from January, 2014 to February, 2021 were collected and analyzed. The results showed that the incidence of male and female, clinical manifestations and co-exist diseases were statistically significant in different age groups (P<0.05). The common pathogens were mainly E. coli and Enterococcus faecium. The proportion of pathogens in 2014-2017 and 2018-2021 was statistically significant (P<0.05). The enzyme-producing rate of E. coli decreased during 2018 and 2021, and the enzyme-producing strains showed higher drug resistance rate than the non-enzyme-producing strains, and the resistance rate of some antimicrobials decreased in recent years. The drug resistance rate of Enterococcus faecium to gentamycin and levofloxacin was higher than 50%, and the sensitivity rate to daptomycin and macrodantin was higher than 60%. It was concluded that the incidence, clinical manifestations and co-exist diseases in children with UTI varied with ages. In recent years, the proportion of gram-negative bacteria show an upward trend, while the proportion of gram-positive bacteria show a downward trend, and β lactamase-producing E．coli and Enterococcus faecium are resistant to many commonly used antibiotics.

Sever fever with thrombocytopenia syndrome virus (SFTSV) is a novel virus discovered in recent years. Sever fever with thrombocytopenia syndrome (SFTS) is caused by SFTSV infection, characterized by high fever, thrombocytopenia, leukopenia and multiple organ function damage, with a mortality rate from 5% to 30%. There is no proven effective antiviral drugs and vaccine available now. SFTSV infection can cause immune dysfunction in patients, and B lymphocytes are one of the important target cells for SFTSV infection, but the impact of SFTSV infection on B lymphocyte function is still unclear. This article reviews research on the abnormalities and underlying mechanism of B lymphocyte functions caused by SFTSV infection, so as to promote the research progress of the immunopathological mechanism of SFTSV infection, providing a basis for a deeper understanding of the impact of SFTSV infection on B lymphocytes and the development of related diagnostic and therapeutic technologies.

Viral encephalitis/meningitis is a severe inflammatory disease of the central nervous system that poses a significant threat to public health. Despite the crucial role played by medical history, symptoms, routine tests and imaging in its diagnosis, and continuous improvements in molecular diagnostic techniques, limitations including technical limitations, assay variability, sample types, and detection time windows still constrain its diagnosis. Moreover, the selection of appropriate diagnostic techniques depends on different clinical features, leaving 40% to 60% of patients with an unclear etiology. Timely and accurate diagnosis is fundamental for initiating targeted and appropriate treatment. Sixty to eighty percent of patients with viral encephalitis initially seek medical attention at primary healthcare institutions. However, due to technical constraints, the diagnostic rate in these institutions is below 20%. Therefore, establishing a standardized diagnostic pathway based on etiology and implementing a tiered diagnostic strategy are particularly necessary. In situations where resources are limited at primary healthcare institutions, the preliminary diagnosis can be established by detailed inquiry into medical history, comprehensive physical examination, and basic laboratory tests. Based on the suspected etiology, routine molecular biology techniques such as polymerase chain reaction (PCR) can be employed for initial pathogen screening. In cases whose PCR results are negative or not feasible, non-targeted broad-coverage techniques like metagenomic next-generation sequencing (mNGS) should be considered. The establishment of a multidisciplinary team for diagnosis and treatment, along with the creation of clinical diagnostic and treatment networks, are essential to enhance the diagnostic rate and improve clinical outcomes. With advancements in diagnostic technologies and the application of artificial intelligence, the diagnosis of viral encephalitis is expected to become more precise.

Corynebacterium kroppenstedtii (C. kroppenstedtii) is a member of the genus Corynebacterium. Genome sequence analysis of the C. kroppenstedtii type strain has revealed a lipophilic (lipid-requiring) lifestyle. The clinical isolates of C. kroppenstedtii have been obtained almost exclusively from female patients, and mainly from the diseased breast tissue samples of the patients with breast abscess and granulomatous mastitis (GM). However, the role of C. kroppenstedtii in breast pathologies remains unclear. This paper reviews the research progress on the correlation between C. kroppenstedtii and granulomatous mastitis.

The purpose of this study is to evaluate the effect of online teaching of medical microbiology course in Dalian Medical University, find and summarize the problems in online teaching in time, and provide a basis for teaching curriculum reform. The examination paper of 641 students who participated in the online teaching and examination of medical microbiology course in 2021 was analyzed by using the network question bank and examination evaluation system of Dalian Medical University, including the accuracy rate of each chapter of the paper, difficulty coefficient, reliability and discrimination. The results showed that the score of this exam was normal distribution, the accuracy of each chapter was between 70% and 90%, the average score of the exam was 72.28, the pass rate was 85%, and the excellent rate was 4%. The overall quality of the examination paper was good.The difficulty of the examination paper was slightly easier, and the difficulty coefficient was 0.72. The discrimination was 0.29, the reliability was 0.9. There was no significant difference in the final exam scores between online teaching students in 2021 and offline teaching students in 2019 (P>0.05). This study confirms that the overall quality of the online examination paper is good, which can objectively reflect the teaching quality and students’ academic level. The online teaching effect is basically satisfied. This study also compares the difficulty coefficient and differentiation of each question type in the test paper to provide the basis and reference for the development of scientific and reasonable online examination papers in the future.