Application of metagenomic next-generation sequencing in the outbreak of healthcare-associated infection with carbapenem-resistant Acinetobacter baumannii / 中国感染控制杂志

Objective To explore the application of metagenomic next-generation sequencing(mNGS)technology in the investigation of healthcare-associated infection(HAI)outbreaks of carbapenem-resistant Acinetobacter bau-mannii(CRAB).Methods Pathogenic detection by mNGS and conventional pathogen culture were performed on 5 patients in the intensive care unit(ICU)of a hospital from June 8 to 22,2023 from whom CRAB were detected.Microbial sampling was carried out in potentially contaminated environment.Bacterial culture,identification,and antimicrobial susceptibility testing were conducted.Comprehensive control measures were taken,and the effect was evaluated.Results The time required for reporting results by mNGS was shorter than the culture time([3.92± 1.05]days vs[6.24±0.25]days,P＜0.001).CRAB was isolated from the specimens of 5 patients.mNGS de-tected OXA-23 resistance genes from all patients.After comprehensive assessment by experts,4 patients were HAI and 1 patient was due to specimen contamination.According to the definition from Guidelines for HAI outbreak control,this event was considered an outbreak of HAI.The monitoring results of environmental hygiene showed that the detection rate of CRAB in the environment during the outbreak was 51.30％(59/115),mainly from the hands of health care workers and the surface of ventilators.After implementing multidisciplinary infection control measures,clinicians'hand hygiene compliance rate and implementation rate of ventilator disinfection increased from 40.83％(49/120)and 33.33％(16/48)to 82.61％(95/115)and 83.33％(30/36),respectively.The prognosis of patients was good,and no new case emerged during subsequent monitoring.The outbreak of HAI in this hospital has been effectively controlled.Conclusion mNGS is characterized by high precision,less time consumption,and high accuracy,and can be applied to the prevention and control of HAI outbreak and the study of antimicrobial-re-sistant genomes.It is of great significance for the anti-infection treatment of patients with multidrug-resistant orga-nism infection as well as the formulation of HAI prevention and control measures.Continuous improving disinfec-tion effectiveness and hand hygiene compliance is important for preventing and controlling CRAB infection.

Clinical application and evaluation of rapid detection of carbapenem resistance genes in blood cultures by Xpert Carba-R / 中华检验医学杂志

This work aims to evaluate a rapid detection method of carbapenem resistance genes in blood cultures based on Xpert Carba-R and preliminarily evaluate its clinical application.Methods:Sixteen strains of Enterobacterales carrying different carbapenem resistance genes were selected to prepare simulated positive blood culture samples and Xpert Carba-R was used to directly detect carbapenem resistance genes in the simulated positive blood culture. From January 2022 to June, a prospective study was conducted on a total of 117 Enterobacteriaceae-positive blood culture samples in the First Affiliated Hospital of Nanjing Medical University. Xpert Carba-R, detecting five kinds of carbapenem resistance genes in these samples, was evaluated in sensitivity and specificity compared to polymerase chain reaction sequencing. Meanwhile clinical data of positive patients was collected for prognostic analysis. Results:Of the 16 simulated specimens, 14 strains had carbapenem resistance genes detected by Xpert Carba-R, including 8 bla KPC, 5 bla NDM and 1 bla IMP, showing 100% agreement with the known results. As of the 117 clinical specimens, 28 cases were determined to be Enterobacterales harboring carbapenem resistance genes, including 24 bla KPC, 2 bla NDM and 2 bla KPC+ bla NDM. In comparison to the PCR sequencing, the sensitivity and specificity of Xpert Carba-R were both 100% for blood culture samples, and furthermore, the detection time was significantly reduced. Of the 25 positive patients, 9 cases were treated with monotherapy and 5 cases were effective, other 16 cases received combined treatment and 12 cases were effective. A total of 17 cases were effective, 8 cases were ineffective and 3 of them died, the mortality rate was 12% (3/25). Conclusion:Xpert Carba-R can rapidly and accurately detect carbapenem resistance genes in blood culture, which can provide evidence for rational drug therapy in early clinical stage.

Construction,expression,purification and quality identification of kanamycin resistant strain of recombinant human interleukin-1 receptor antagonist / 中国生物制品学杂志

@#Objective To construct a recombinant human interleukin-1 receptor antagonist(rhIL-1Ra)strain of kanamycin resistance,express,purify and identify rhIL-1Ra protein in order to reduce the risks of β-lactam antibiotics.Methods The ampicillin-resistant rhIL-1Ra(A-rhIL-1Ra)plasmid and PET-28a plasmid were used as templates to prepare the linearized vector and kanamycin gene fragment by PCR.The kanamycin resistant rhIL-1Ra plasmid(K-rhIL-1Ra)was constructed by homologous recombination,which was transformed into E.coli BL21(DE3)to construct the recombinant engineered bacteria after correct sequencing.The engineered bacteria were induced by IPTG and then purified by CM Bestarose Fast Flow and DEAE Bestarose Fast Flow column chromatography.The purified products were collected,and then detected for the purity by 15% SDS-PAGE,size-exclusion high-performance liquid chromatography(SEC-HPLC),and reversed-phase high-performance liquid chromatography(RP-HPLC),determined for the relative molecular mass by tandem mass spectrometry,identified for the specificity by Western blot,measured for the biological activity by reporter gene assay,and compared for the related impurities by liquid chromatography-mass spectrometry(LC-MS)analysis.Results Colony PCR and sequencing results showed that K-rhIL-1Ra plasmid was constructed correctly.The expressed K-rhIL-1Ra protein had a relative molecular mass of about 17 000,mainly existing in soluble form,and the expression amount accounted for more than 30% of the total bacterial proteins.The purity of K-rhIL-1Ra protein purified by two steps was 97% and 99%,the monomer content was 99.33%and 100%,and the chromatographic purity was 91.86% and 96.96%,respectively.The mass spectral molecular mass was consistent with that of the standard(A-rhIL-1Ra protein),and the protein reacted specifically relative with mouse antihuman IL-1Ra monoclonal antibody.The biological activity of the purified K-rhIL-1Ra protein was 1.29 × 105U/mg,and the chemical modification types of related proteins were the same as those of the standard(A-rhIL-1Ra protein).Conclusion The K-rhIL-1Ra strain was successfully constructed,and the expressed and purified protein was in line with the characteristics and quality standards of rhIL-1Ra,which lays a foundation for the study of rhIL-1Ra strain change and comparability

Analysis of virulence and drug resistance characteristics of diarrheagenic Escherichia coli isolated from patients with infectious diarrhea in a hospital in Beijing / 临床检验杂志

Objective To investigate the virulence and drug resistance characteristics of diarrheagenic Escherichia coli isolated from patients with infectious diarrhea in our hospital.Methods The preliminary identification of microbes was carried out by the VITEK-MS microbial mass spectrometry detection system and virulence genes were detected by the multiplex real-time PCR.Five types of diarrhea-genic Escherichia coli(DEC)clinically isolated from patients with infectious diarrhea in our hospital were identified.The drug resist-ance characteristics of DEC strains were detected by the microbroth dilution and E-test.The drug-resistant molecular characteristics were analyzed by the next-generation sequencing and bioinformatics.The Fisher exact probability method was used for statistical analy-sis.Results The detection rate of DEC in our hospital was 11.9%,with enteroaggregative E.coli(EAEC)accounting for 37.5%,a-typical enteropathogenic Escherichia coli(EPEC)accounting for 34.38%,enterotoxigenic E.coli(ETEC)accounting for 25.0%,and enteroinvasive E.coli(EIEC)accounting for 3.12%.None of enterohemorrhagic E.coli(EHEC)strain was detected.The resistance rates of 32 DEC strains to ampicillin,tetracycline,and trimethoprim/sulfamethoxazole were 53.12%,43.75%,and 37.5%,respec-tively.ESBLs(+)strains accounted for 18.75%,and the detection rate of multidrug-resistant strains was 83.83%,significantly higher than that of ESBLs(-)strains(P=0.042).A total of 25 ST genotypes were obtained from 32 DEC strains.The dominant genotypes were ST10(4 strains,12.5%),followed by ST28(2 strains,6.25%),ST31(2 strains,6.25%),ST3153(2 strains,6.25%),and the other 21 genotypes(1 strain,3.13%).One carbapenem resistant strain carrying the blaNDM-1 gene was detected in EAEC.Conclu-sion Four virulence genes such as aggR,pic,astA,and eae,are more common in the DEC of patients with infectious diarrhea in our hospital,with EAEC and EPEC as the main subtypes.The genotypes are highly polymorphic,and multidrug-resistant strains have been detected.

ISOLATION AND SEQUENCING OF RESISTANCE GENE ANALOGS FROM THE NBS-LRR REGION OF COFFEA ARABICA GENOME

Coffee is one of the most economically important crops in India and also across the world. Apart from the abiotic stress, a USD 495.5 billion coffee industry suffers from the outbreak of various diseases caused by pathogenic fungus, bacteria and viruses. The presence of Resistance Gene Analogs (RGAs) in coffee plant is the most prominent marker of resistance against the pathogens. The current study aims to isolate resistance gene analogs from the nucleotide binding site – Leucine rich repeats (NBS-LRR) region of the Coffea arabica chromosome.

Value of drug resistance and homology of Acinetobacter baumannii in tracing the source of nosocomial infection in Jianyang, Sichuan / 中国热带医学

@#Abstract: Objective　To analyze the drug sensitivity and the carrying of carbapenem resistant gene of Acinetobacter baumannii isolated from clinical patients and clinical objects, and analyze the homology of strains to provide support for the control of nosocomial infection. Methods　A total of 38 strains of Acinetobacter baumannii isolated from patients and clinical objects surface were collected from January 2019 to August 2020. The antimicrobial susceptibility was tested by the minimum inhibitory concentration method. In addition, the resistance related genes were detected by polymerase chain reaction method, and homology analysis was performed by enterobacterial repetitive Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR). Results All 34 strains of Acinetobacter baumannii isolated from Clinical patients and 4 strains isolated from clinical objects carried blaOXA-51 and imp resistance genes, neither of them carried blaVIM gene. 32 Acinetobacter baumannii carrying blaOXA-23 gene, 28 strains carrying blaTEM gene, 7 strains carrying blaOXA-58 gene. After cluster analysis, 38 Acinetobacter baumannii isolates were classified into 7 genotypes (expressed A, B, C, D, E, F, G), and cluster E and cluster G were the main clusters, containing 12 strains (12/38, 31.6%) and 18 strains (18/38, 47.4%), respectively, as the main prevalent clonal strains. Conclusions　Acinetobacter baumannii isolated from different sources have the significant differences in drug resistance and carry different resistance genes. There is no direct correlation between patients and environmental isolates of Acinetobacter baumannii belonging to different clonal strains. Also, there aren’t significant correlation between clinical patients infected with Acinetobacter baumannii.

Molecular features and drug-resistance genes of 70 erythromycin-resistant group B Streptococcus strains in Zhengzhou / 中华微生物学和免疫学杂志

Objective:To investigate the phenotypes, molecular types and drug-resistance genes of erythromycin (ERY)-resistant group B Streptococcus (GBS) in pregnant women in late pregnancy in Zhengzhou and provide basic data for the prevention, control and treatment of GBS infection. Methods:This study retrospectively collected 86 GBS strains isolated from the vaginal secretions of pregnant women in late pregnancy at Maternal and Child Health Hospital of Henan Province from 2021 to 2022. ERY-resistant GBS strains were selected using the ERY disk diffusion method, and their susceptibility to 10 different antibiotics was tested. Whole-genome sequencing was performed to analyze their molecular features including molecular types, clonal complex groups and drug-resistance genes. Drug-resistance genes carried by GBS strains belonging to different clonal complex groups were compared.Results:There were 70 ERY-resistant GBS strains. Among them, 7.14%(5/70) exhibited an inducible resistance phenotype to macrolide-lincosamide-streptogramin B (MLSB) antibiotics; 84.29%(59/70) showed constitutive resistance to MLSB antibiotics; 8.57%(6/70) were resistant to macrolides but susceptible to lincosamides. The resistance rates of these strains to clindamycin (CLI), tetracycline (TE) and levofloxacin (LEV) were 91.43%(64/70), 54.29%(38/70) and 60.00%(42/70), respectively. These ERY-resistant strains exhibited multidrug resistance patterns with 40.00%(28/70) showing ERY-CLI-LEV resistance phenotype and 30.00%(21/70) showing ERY-CLI-TE resistance phenotype. The major drug-resistance genes carried by the 70 GBS strains were macrolide/lincosamide resistance genes mreA (100.00%) and ermB (53/70, 75.71%), aminoglycoside resistance genes ant (6)-Ⅰ a (22/70, 31.43%) and aph(3′)-Ⅲ (18/70, 25.71%), and tetracycline resistance genes tetM (22/70, 31.43%) and tetO (13/70, 18.57%). These strains belonged to 12 sequence types derived from seven clonal complexes (CCs) and 48.57%(34/70) of them were clustered into CC12. All CC12 strains harbored ermB, but none carried ermA. The positive rates of lsaE, lunB, and aac (6′)- aph(2" ) in CC19 and CC651 strains, ant (6)-Ⅰ a in CC651 and CC452 strains, and mefA and msrD in CC19 and CC23 strains were significantly higher than those in CC12 strains ( P<0.001). Conclusions:ERY-resistant GBS in Zhengzhou exhibited diverse drug resistance phenotypes and molecular types. CC12 was the most prevalent clonal complex in this region. The constitutive MLSB resistance phenotype and ermB gene were the most common ERY resistance phenotype and genotype, respectively, and tetM gene was related to tetracycline resistance. Furthermore, the drug-resistance genes varied in GBS strains of different clonal complexes. This study suggested that close attention should be paid to the epidemiological situation of GBS in this region and the effectiveness of antibiotics used for clinical prevention and treatment of GBS infection should be carefully evaluated.

Positive antibiotic and resistance genes in source water of three regions and correlation analysis / 环境与职业医学

Background China is a big country in the production and use of antibiotics. The abuse of antibiotics enables bacteria in water environment to acquire resistance, and promotes the generation and spread of antibiotics resistance genes (ARGs). The problem of antibiotic-resistant bacteria is increasingly serious and has become a public security issue of global concern. Water environment is a huge reservoir of antibiotics and ARGs. It is of great significance to study the pollution of antibiotics and ARGs in water to protect water sources and optimize the biosecurity of drinking water. Objective To evaluate the detection of antibiotics and ARGs in typical water sources, and to explore the relationship between antibiotics and ARGs. Methods Water samples were collected in Heilongjiang, Liaoning, and Hubei provinces during the wet season (from August to October) in 2020. Ten water samples were collected from each of the three places, and a total of 30 water samples were collected in this study. Five kinds of antibiotics, including macrolides, quinolones, sulfonamides, tetracycline, and β-lactam, were detected by ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The integron (Intl1), 16S rRNA, and 6 kinds of ARGs were detected by quantitative real-time PCR (qPCR). The ARGs include one macrolide ARGs (ermB), one β-lactam ARGs (blaTEM), two tetracycline ARGs (tetC, tetQ), and two sulfonamide ARGs (sul1, sul2). Results The types of detected antibiotics varied by the three regions, and the concentration ranges of the same antibiotics varied by the three regions (P<0.05). The concentration ranges of selected five kinds of antibiotics were 0.11-418.80 ng·L−1 in region A, 0.12-23.23 ng·L−1 in region B, and 4.69-285.75 ng·L−1 in region C, respectively. The detection rates of all six ARGs were 100%. The absolute abundance of ARGs in region A ranged from 22.56 to 94355.91 copies·mL−1, that in region B ranged from 27.99 to 80584.32 copies·mL−1, and that in region C ranged from 41.99 to 111068.19 copies·mL−1. The absolute abundance of blaTEM was higher among the ARGs, followed by sul1 and sul2. In addition, the absolute abundance of Intl1 was also at a high level. The results of correlation analysis showed that the abundance of ARGs was positively correlated with each other. There was no correlation between specific antibiotics and corresponding ARGs. There was a positive correlation between Intl1 and sul1 or sul2 (P<0.05). Conclusion The types and concentrations of antibiotics and the abundance of ARGs in source water vary greatly in the study areas. The association between antibiotics and ARGs is uncertain. Intl1 may play an important role in the horizontal transfer of sulfonamide resistance genes.

Distribution characteristics of antibiotic resistance genes in the domestic water of residents in Haimen， Jiangsu Province / 上海预防医学

ObjectiveAntibiotic resistance genes （ARGs） have received wide attention all over the world. The purpose of this study was to explore the bacterial community structure， the types and levels of antibiotic resistance genes in a water body in east China， and to compare and analyze the characteristics of microbial species distribution and antibiotic resistance gene distribution in various water environments. MethodsA total of 10 households in Haimen City， Jiangsu Province were selected and their surrounding water environment samples were collected. 21 water samples including river water （4）， Mingou water （9） and well water （8） were collected for metagenomics sequencing， assembled with MetaWRAP， annotated with CARD database， and analyzed with R software. ResultsIn various water bodies， the dominant bacteria phyla was Proteobacteria， the dominant bacteria genera were Deuterostomia， Pseudomonas， Flavobacteriales and Streptomycetaceae. The ARGs annotated were mainly composed of quinolones， aminoglycosides， macrolides and beta-lactams antibiotic resistance genes. The top four relative abundance of resistance genes were macB， RanA， evgS and TxR， The average absolute abundance and expression of resistance genes in well water and Mingou water were higher than those in river water. ConclusionMultiple ARGs are detected to varying degrees in well water， river water， and Mingou water bodies， and the expression of resistance genes in well water and Mingou water bodies is higher than that in river water bodies， possibly due to human production and living activities.

Diagnostic efficacy of joint detection of Xpert MTBRIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid for smear-negative pulmonary tuberculosis / 中国热带医学

@#Abstract: Objective　To investigate the diagnostic value of joint detection of Mycobacterium tuberculosis rifampicin resistance gene (Xpert MTB/RIF), Mycobacterium tuberculosis ribonucleic acid (TB-RNA) and Mycobacterium tuberculosis deoxyribonucleic acid (TB-DNA) in bronchoalveolar lavage fluid for smear-negative pulmonary tuberculosis. Methods A total of 806 patients with suspected smear-negative pulmonary tuberculosis admitted to our hospital from May 2020 to July 2022 were selected, 506 patients diagnosed as bacterial negative pulmonary tuberculosis by clinical, X-ray and sputum samples were classified as bacterial negative pulmonary tuberculosis group, and the other 300 patients with non-tuberculous pulmonary disease were classified as non-tuberculous pulmonary disease group. XpertMTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid of all patients were detected. With clinical, X-ray and sputum specimen examination of mycobacterium tuberculosis as the gold standard, the diagnostic efficacy of alveolar lavage solution Xpert MTB/RIF, TB-RNA and TB-DNA alone and in combination was analyzed. Results The positive detection rates of Xpert MTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid of the smear-negative pulmonary tuberculosis group and the non-tuberculosis pulmonary disease group were 69.96% (354/506) and 2.67% (8/300), 61.46% (311/506) and 5.00% (15/300), and 63.64% (322/506) and 8.00% (24/300), respectively. The rates in the smear-negative pulmonary tuberculosis group were higher than those in the non-tuberculosis lung disease group, and the differences were statistically significant (χ2=342.005, 246.930, 235.687, P<0.01). Compared with the gold standard, the sensitivity, specificity, accuracy, positive predictive value and negative predictive value of Xpert MTB/RIF in the diagnosis of smear-negative pulmonary tuberculosis were 69.96%, 97.33%, 80.15%, 97.79% and 65.77%, respectively; those values of TB-RNA were 61.46%, 95.00%, 73.95%, 95.40% and 59.38%, respectively; those values of TB-DNA were 63.64%, 92.00%, 74.19%, 93.06% and 60.00%, respectively; those values of combined diagnosis with Xpert MTB/RIF, TB-RNA and TB-DNA were 61.26%, 100.00%, 75.68%, 100.00% and 60.48%, respectively; the specificity and positive predictive value of combined detection were higher than those of single detection (P<0.05). Conclusions The joint detection of Xpert MTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid can improve the diagnostic efficacy of smear-negative pulmonary tuberculosis and is worthy of clinical promotion and application.

Isolation and identification of multidrug-resistant Escherichia coli from cattle, sheep, poultry and human in Cumilla, Bangladesh

Aims@#Antimicrobial resistance (AMR) is a significant public health concern of modern civilization. The potential risk of AMR is significant in terms of both human and animal health. This study aims to assess the antimicrobial resistance pattern of selected antimicrobials against Escherichia coli of animal, poultry and human origin in the Cumilla district of Bangladesh.@*Methodology and results@#A total of 200 samples were collected from different sources. Isolation and identification of commensal E. coli were performed following standard bacteriological and molecular techniques. Antimicrobial susceptibility testing was performed following the Kirby-Bauer disc diffusion technique. Ampicillin, tetracycline and sulfamethoxazole-trimethoprim resistance genes were detected by polymerase chain reactions (PCR). A total of 152 (76%; 95% confidence interval (CI) 70-81%) E. coli were isolated from cattle, sheep, chicken and human, where 37.5% of isolates were found to be multidrug-resistant (MDR). In the cultural sensitivity test, E. coli showed the highest resistance to sulfamethoxazole-trimethoprim (71%), tetracycline (63%), ampicillin (62%), where gentamicin (23%) showed the lowest resistance, followed by ceftriaxone (26%). The prevalence of resistance genes like blaTEM, tetA, tetB, tetC, sul1 and sul2 were 100%, 95%, 11%, 8%, 58% and 52%, respectively.@*Conclusion, significance and impact of study@#The emergence of multidrug-resistant commensal E. coli and resistance genes circulating in animals, poultry and humans limit the treatment options for serious infections.

Diversity of antibiotic resistance genes in raw water from drinking water source in Shanghai / 上海预防医学

ObjectiveTo determine the distribution of various antibiotic resistance genes （ARGs） in raw water from drinking water source， and to explore the correlation between the ARGs and common carbapenem-resistant and multidrug-resistant bacteria isolated from drinking water source， so as to provide scientific evidence for improving the safety of urban drinking water. MethodsA total of 30 raw water samples were collected from a major drinking water source in Shanghai in 2020. Bacterial strains were selectively cultured on Columbia blood agar medium containing 1 μg·μL-1 meropenem， and then identified by MALDI-TOF-MS mass spectrometry system. Minimum inhibitory concentration （MIC） of the strains was detected by broth microdilution method. The water samples were filtered through a 0.45 μm filter membrane and diversity of ARGs was determined by using high-throughput metagenomic sequencing. ResultsA total of 64 strains of carbapenem-resistant bacteria were isolated from the water samples， including Stenotrophomonas maltophilia， Enterococcus faecium， Pseudomonas aeruginosa， Acinetobacter baumannii and Klebsiella pneumoniae， which were resistant to a variety of common antibiotics. Using metagenomic sequencing，1 244 ARGs were identified. The relative average abundance of the top 100 ARGs accounted for 96.1%， and that of the multidrug-resistant ARGs accounted for 63.41%. Furthermore， the multidrug-resistant ARGs were mainly adeJ， mexT， adeC， oprM， mexF， mdfA， mexB， mdtK， adeK， etc. Using Spearman's correlation， five multidrug-resistant bacteria isolated from the drinking water source were significantly associated with the ARGs. ConclusionRelative abundance of multidrug-resistant ARGs is high in raw water from main drinking water source. The five isolated carbapenem-resistant and multidrug-resistant bacteria are significantly correlated with the ARGs. It warrants strengthening the rational and standardized application of antibiotics to protect water resources and ensure the safety of drinking water.

Clinical and bronchoscopy characteristics of Mycoplasma pneumoniae pneumonia in children with 23S rRNA resistance gene positive in bronchoalveolar lavage fluid / 中华实用儿科临床杂志

Objective:To investigate clinical and bronchoscopy characteristics of Mycoplasma pneumoniae pneumonia in children with 23S rRNA resistance gene positive in bronchoalveolar lavage fluid(BALF), and find clinical indicators that can identify Mycoplasma pneumoniae drug resistance early.Methods:The clinical data of 61 hospita-lized children diagnosed with Mycoplasma pneumoniae pneumonia as subjects from October 2017 to June 2018 in the Department of Pediatric Respiratory Medicine of Shengjing Hospital Affiliated to China Medical University were analyzed retrospectively.Bronchoscopy was performed on each subject and the BALF was taken and used to detect the 2063 site mutation of the 23S rRNA V region gene in BALF, and they were divided into drug-resistant gene positive group and drug-resistant gene negative group.The clinical manifestations, relevant laboratory data, imaging data, and bronchoscopy findings in different load groups were compared.Statistical methods such as t test, rank sum test, χ2 test, Fisher′ s exact probability method, and multivariate Logistic regression analysis were used to analyze the data. Results:Among the 61 children, 38 cases (62.30%) were in the drug resistance gene positive group, and 23 cases (37.70%) were in the drug resistance gene negative group.There were no significant differences in gender and age between the two groups (all P>0.05). The days of hospitalization and fever in the children with positive drug resistance genes were longer than those in the negative drug resistance gene groups, and they were more likely to have refractory mycoplasma pneumoniae pneumonia(RMPP) and extra pulmonary complications, with statistically significant differences ( P<0.05), but there were no significant differences in hypoxemia ( P>0.05). There were significant differences in white blood cell(WBC), C-reactive protein(CRP), procalcitonin(PCT), D-Dimer (D-D) and interleukin-6 (IL-6) between the two groups (all P<0.05). Except that the WBC level in the drug-resistant gene-positive group was lower than that in the drug-resistant gene-negative group, the rest of the test results indicated that the drug-resistant gene-positive group was higher than the drug-resistant gene-negative group.There were no significant differences in the concentrations of serum lactate dehydrogenlase(LDH)and IL-6 in BALF ( P>0.05). In this study, Logistic regression analysis was performed on several statistically significant laboratory indicators.It was found that WBC was more sensitive to identify drug resistance genes, and the optimal critical value was 8.55×10 9/L.The specificity of D-D in identifying drug resistance genes was higher, and the optimal cut-off value was 523 μg/L.In the drug resistance gene positive group, 35 cases (92.11%) showed extensive lung consolidation/atelectasis on imaging, and the drug resistance gene negative group was 13 cases (56.52%), with statistically significant differences ( P<0.05). The drug resistance gene-positive group mainly showed mucosal erosion, necrosis, phlegm plug/plastic phlegm plug and bronchitis stenosis, with a total of 19 cases (50.00%). In the drug resistance gene negative group, the main manifestations of mucosal longitudinal wrinkle, flocculent and viscous secretions were 14 cases (60.88%), with statistically significant differences ( P<0.05). Conclusions:The point mutation of 23S rRNA V region gene is closely related to the clinical characteristics of children with Mycoplasma pneumoniae pneumonia.Children with A2063G mutation are more prone to have RMPP and extrapulmonary complications, and their imaging manifestation and bronchoscopy are more severe.The levels of leukocytes and D-D in the blood have significance for the early identification of drug resistance.The systemic excessive immune inflammatory response caused by Mycoplasma pneumoniae pneumonia with drug-resistant gene positive needs to be valued.

Comparison of drug susceptibility of and drug resistance mutations in fluconazole-resistant Candida albicans strains from superficial and deep infections / 中华皮肤科杂志

Objective:To compare the in vitro susceptibility of fluconazole-resistant Candida albicans strains from superficial and deep infections to 8 antifungal drugs, and to compare drug resistance mutations in these strains. Methods:According to the Clinical and Laboratory Standards Institute (CLSI) protocol M27-A4, 26 deep infection-derived and 33 superficial infection-derived drug-resistant Candida albicans strains were tested for in vitro susceptibility to 8 antifungal drugs (fluconazole, voriconazole, itraconazole, posaconazole, amphotericin B, fluorocytosine, terbinafine, and micafungin) alone or in combination. DNA was extracted from all drug-resistant strains, and mutations in 3 drug resistance genes, including ERG3, ERG11 and FUR1, were detected by PCR. Normally distributed measurement data with homogeneous variance were compared between two groups by using two-independent-sample t test, non-normally distributed measurement data with non-homogeneous variance were compared using Mann-Whitney U test, and enumeration data were compared using chi-square test. Results:The minimum inhibitory concentrations (MICs) of fluconazole, itraconazole, voriconazole, posaconazole and fluorocytosine all significantly differed between the superficial infection group and deep infection group (all P < 0.05) , while there was no significant difference in the MIC of amphotericin B or micafungin between the two groups (both P > 0.05) . The MIC of terbinafine was >64 μg/ml in 96.6% of the above strains, so could not be compared between groups. As combination drug susceptibility testing revealed, the combination of terbinafine with azoles (fluconazole, voriconazole, itraconazole or posaconazole) showed synergistic inhibitory effects against 15 Candida albicans strains (7 strains from deep infections, 8 strains from superficial infections) , with fractional inhibitory concentration (FIC) indices being 0.033 to 0.187; no marked synergistic effect was observed in the combinations between fluorocytosine and azoles, between fluorocytosine and amphotericin B, or between amphotericin B and fluconazole, with the FIC indices being 0.56 to 1.125. The missense mutation V351A in the ERG3 gene was identified in all the 33 (100%) superficial infection-derived strains, as well as in 13 (50%) deep infection-derived strains, and the mutation A353T in the ERG3 gene was identified in 4 (15%) deep infection-derived strains; as for the ERG11 gene, missense mutations identified in the superficial infection-derived strains included I437V (32 strains, 97%) , Y132H (23 strains, 70%) , T123I (16 strains, 48%) , K128T (6 strains, 18%) , D116E (5 strains, 15%) , A114S (4 strains, 12%) , E266D (2 strains, 6%) , G448E (2 strains, 6%) , and G465S (2 strains, 6%) , while missense mutations identified in the deep infection-derived strains included I437V (23 strains, 88%) , E266D (13 strains, 50%) , E260G (5 strains, 19%) , and V488I (4 strains, 15%) ; the missense mutation R101C in the FUR1 gene was identified in 11 (33%) superficial infection-derived strains, but not identified in deep infection-derived strains. Conclusion:The drug susceptibility and drug resistance mutations differed to some extent between superficial infection- and deep infection-derived fluconazole-resistant Candida albicans strains.

Astragalus polysaccharides affects multidrug resistance gene 1 and P-glycoprotein 170 in adriamycin nephropathy rats via regulating microRNA-16/NF-κB axis / 中南大学学报(医学版)

OBJECTIVES@#Nephrotic syndrome is a common disease of the urinary system. The aim of this study is to explore the effect of astragalus polysaccharides (APS) on multidrug resistance gene 1 (MDR1) and P-glycoprotein 170 (P-gp170) in adriamycin nephropathy rats and the underlying mechanisms.@*METHODS@#A total of 72 male Wistar rats were divided into a control group, a model group, an APS low-dose group, an APS high-dose group, an APS+micro RNA (miR)-16 antagomir group and an APS+miR-16 antagomir control group, with 12 rats in each group. Urine protein (UP) was detected by urine analyzer, and serum cholesterol (CHOL), albumin (ALB), blood urea nitrogen (BUN), and creatinine (SCr) were detected by automatic biochemical analyzer; serum interleukin-6 (IL-6), IL-1β, tumor necrosis factor α (TNF-α) levels were detected by ELISA kit; the morphological changes of kidney tissues were observed by HE staining; the levels of miR-16 and MDR1 mRNA in kidney tissues were detected by real-time RT-PCR; the expression levels of NF-κB p65, p-NF-κB p65, and P-gp170 protein in kidney tissues were detected by Western blotting; and dual luciferase was used to verify the relationship between miR-16 and NF-κB.@*RESULTS@#The renal tissue structure of rats in the control group was normal without inflammatory cell infiltration. The renal glomeruli of rats in the model group were mildly congested, capillary stenosis or occlusion, and inflammatory cell infiltration was obvious. The rats in the low-dose and high-dose APS groups had no obvious glomerular congestion, the proliferation of mesangial cells was significantly reduced, and the inflammatory cells were reduced. Compared with the high-dose APS group and the APS+miR-16 antagomir control group, there were more severe renal tissue structure damages in the APS + miR-16 antagomir group. Compared with the control group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 in the model group were significantly increased (all P<0.05); the levels of ALB and miR-16 were significantly decreased (both P<0.05). Compared with the model group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of pNF-κB p65 and P-gp170 in the low-dose and high-dose APS groups were significant decreased (all P<0.05); and the levels of ALB and miR-16 were significantly increased (both P<0.05). Compared with APS+miR-16 antagomir control group, the UP, CHOL, BUN, SCr, IL-6, IL-1β, and TNF-α levels, MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 were significantly increased (all P<0.05). The levels of ALB and miR-16 were significantly decreased in the APS+miR-16 antagomir group compared with the APS+miR-16 antagomir control group (both P<0.05).@*CONCLUSIONS@#APS can regulate the miR-16/NF-κB signaling pathway, thereby affecting the levels of MDR1 and P-gp170, and reducing the inflammation in the kidney tissues in the adriamycin nephropathy rats.

Analysis of serotype distribution and antibiotic resistance of Haemophilus influenzae in hospitalized children / 中华实用儿科临床杂志

Objective:To explore the serotype, antibiotic resistance and β-lactamase gene of Haemophilus influenzae strains isolated from hospitalized children, thus providing reference for clinical diagnosis and treatment. Methods:A total of 148 Haemophilus influenzae strains collected from January 2016 to December 2018 in hospitalized children of Children′s Hospital, Capital Institute of Pediatrics were retrospectively analyzed.The serotype and genotype of Haemophilus influenzae strains were examined by slide agglutination test and PCR, respectively.The sensitivity of isolates to Ampicillin and other antimicrobials was detected by the E-test and disk diffusion methods.The β-lactamase phenotype was tested by nitrocefin disk method.The carrying of β-lactamase gene TEM-1 and ROB-1 were detected by PCR.The drug resistance rate was compared by χ2 test. Results:All the 148 strains were nontypeable Haemophilus influenzae (NTHi), and capsular gene was not amplified.The rate of resistance to Ampicillin, Ampicillin/Sulbactam, Cefuroxime, and Azithromycin were 68.9%(102/148 strains), 40.5%(60/148 strains), 53.4%(79/148 strains) and 56.1%(83/148 strains), respectively.The Haemophilus influenzae isolates showed the highest resistance rate to Trimethoprim-sulfamethoxazole, which was up to 91.9%(136/148 strains). The sensitive rate of isolates to Ceftriaxone, Meropenem and Levofloxacin were all 100.0%(148/148 strains). The prevalence of β-lactamase was 64.8%(96/148 strains) in Haemophilus influenzae and the genotype was TEM-1.The drug resistance rates of β-lactamase positive strains to Ampicillin, Ampicillin/Sulbactam and Azithromycin were significantly higher than those of other strains( χ2=123.222, 27.973, 70.273, all P<0.01). Conclusions:The most prevalent serotype of Haemophilus influenzae is NTHi in children. Haemophilus influenzae carried TEM-1 gene had a high positive rate of β-lactamase production, which was the main mechanism of drug resistance to Ampicillin.Ceftriaxone and Meropenem were the most active agents against Haemophilus influenzae.

Comparative Study of the Genetic Diversity, Antimicrobial Resistance, and Pathogenicity of / 生物医学与环境科学（英文）

Objective@#This study was performed to compare the genetic diversity, virulence, and antimicrobial resistance of @*Methods@#A total of 38 clinical strains and 19 strains from healthy individuals were isolated from the samples collected in Ma'anshan City, Anhui Province. Their taxonomy was investigated using concatenated @*Results@#The 57 @*Conclusions@#The taxonomy, virulence properties, and antibiotic resistance of

Pre-treatment drug resistance of newly reported HIV-1 infected individuals in Wenzhou / 预防医学

Objective@#To know the pre-treatment drug resistance ( PDR ) status of newly reported human immunodeficiency virus type 1 ( HIV-1 ) infected individuals in Wenzhou, so as to provide guidance for antiretroviral therapy ( ART ). @*Methods@# Totally 232 plasma samples of newly reported HIV-1 infected individuals who had not received ART were collected in Wenzhou in 2019. Virus ( HIV-1 ) RNA was extracted, followed by reverse transcription PCR and nested PCR to amplify the pol region and sequence. Resistance mutations and resistance to non-nucleoside reverse transcriptase inhibitors ( NNRTIs ), nucleoside reverse transcriptase inhibitors ( NRTIs ) and protease inhibitors ( PIs ) was analyzed.@*Results@#The pol region sequences from 199 infected patients were obtained and the incidence of PDR was 8.04% ( 16/199 ). Eight genotypes were detected, including circulating recombinant forms ( CRFs ) CRF07_BC ( 47.24%, 94/199 ) and CRF01_AE ( 29.15%, 58/199 ) which were the dominant types. Two unique recombinant forms ( URFs ) were detected, namely URF( CRF01_AE/BC ) and URF( B/C ) . Thirty-one cases ( 15.58% 31/199 ) had drug-resistant mutations. For NNRTIs, NRTIs and PIs, 20 cases ( 64.52% ) , 2 cases ( 6.45% ) and 9 cases ( 29.03% ) with drug resistance mutations were detected, respectively. The resistance mutations to NNRTIs included K101E, K103N/R, V106I, E138K, V179D/E/T, Y181C, G190A and H221Y. Four cases each had two resistance mutations to NNRTIs. The resistance mutations to NRTIs were V75M and M184V. The resistance mutations to PIs were M46I, L33F and Q58E. For the newly released NNRTI drug Doravirine ( DOR ), two cases were found to have mutations of resistance. @*Conclusions@#The incidence of PDR among newly reported HIV-1 patients in Wenzhou is 8.04%, mainly caused by NNRTIs drug-resistant mutation. Resistance to the new drug DOR has emerged. The surveillance of drug resistance should continue to be strengthened.

Analysis of serum and drug resistance levels between food source and human salmonella / 公共卫生与预防医学

Objective To analyze serotype distribution, drug resistance, quinolone resistance gene carrying status and genetic relationship of foodborne Salmonella and human Salmonella isolates in Changzhou from 2012 to 2018, to provide scientific basis for the prevention and control of Salmonella. Methods The serum type was identified by serum agglutination and liquid chip. The antibiotic sensitivity was determined by micro broth dilution method. The quinolone antibiotic resistance gene was determined by gene sequencing method. The multilocus sequence typing ( MLST ) typing was performed on quinolone-resistant Salmonella, and the genetic relationship was analyzed by BioNumerics 8.0. Results A total of 10 and 36 serotypes were detected in 46 foodborne Salmonella strains and 152 human Salmonella strains, respectively. The dominant serotypes were Indiana Salmonella and Salmonella typhimurium. Erythromycin resistance rate was the highest in both Salmonella strains, and the proportion of multidrug-resistant bacteria was 93.47 % ( 43 / 46 ) and 80.92 % ( 123 / 152 ), respectively. 38 strains of quinolone-resistant foodborne Salmonella GyrA subunit mainly occurred double mutations Asp87Asn, Ser83Phe, ParC subunit mainly occurred single mutation Ser80Arg, 119 strains of quinolone-resistant human Salmonella qnrS gene detection rate was higher, reached 68.1 % ( 81 / 119 ) ; The dominant ST types of quinolone-resistant Salmonella from two sources were ST17 and ST19, respectively. Conclusions The antibiotic sensitivity of the two Salmonella resistant strains from Changzhou was the same ; Synergistic drug resistance, but both quinolone resistance genemutations and carry inconsistent ; The ST type distribution of quinolone resistant strains isalso inconsistent, and the genetic relationship is far. It is suggested that the probability of Salmonella resistant bacteria infection caused by food transmission in our region is small, and the treatment of the two should be differentiated.

Research progress on antibiotic resistance of Escherichia coli O157:H7 / 预防医学

@#Contamination of foodborne pathogens is the main cause of related diseases. Escherichia coli O157:H7 (E.coli O157:H7), as a representative of pathogenic E.coli, is one of the most severe and commonly reported E.coli in the world, but there is still no effective clinical treatment against the infection. Antibiotics show effective in the early infection of E.coli O157:H7. However, their extensive use has led to drug-resistant bacteria and genes in recent years, which becomes a great threat to public health. This article reviews the research progress of E.coli O157:H7 from the prevalence of antibiotic-resistant bacteria, the resistance mechanism, and the prevention and control methods, in order to provide a reference for its prevention, early clinical treatment and related research.