Alternative splicing of the conserved drug-resistant orthologue FpNcb2 is associated with its nuclear accumulation of products and full virulence of Fusarium pseudograminearum.

BACKGROUND: Negative cofactor 2 NC2ß (Ncb2 or Dr1) is the beta subunit of a conserved heterodimeric regulator of transcription negative cofactor 2 (NC2) complex that has been identified as key regulator of drug resistance in model fungi. However, its role in plant pathogens is still unclear. RESULTS: We identified an NC2ß orthologue, FpNcb2, in Fusarium pseudograminearum, which is not only a significant regulatory function in drug resistance, but also essential for growth, conidiation and penetration. Moreover, FpNcb2 undergoes alternative splicing which creates two mRNA isoforms. As a putative CCAAT binding protein, FpNcb2 concentrates in the nuclei, contributing to the expression of two spliced mRNA of FpNcb2 in hypha, conidiophores and conidia, with exception of FpNcb2ISOA in germlings. Expression of each spliced mRNA of FpNcb2 in Δfpncb2 mutant could full complement the defects on growth, conidiation and fungicides sensitivity to that of wild type. However, FpNcb2ISOA and FpNcb2ISOB have different effects on virulence. FpNcb2 acts as a regulator for the transcription of some genes encoding drug efflux and hydrolases. CONCLUSION: Our analysis showed the existence of alternative mRNA splicing in the NC2ß orthologue, which is associated with protein subcellular localization and fungal virulence. The further elucidation of the target genes of NC2ß will provide insights into the potential regulation mechanisms in the antifungal resistance and pathogenesis of F. pseudograminearum. © 2024 Society of Chemical Industry.

Bacteriophage EPP-1, a potential antibiotic alternative for controlling edwardsiellosis caused by Edwardsiella piscicida while mitigating drug-resistant gene dissemination.

Edwardsiella piscicida causes significant economic losses to the aquaculture industry worldwide. Phage-based biocontrol methods are experiencing a renaissance because of the spread of drug-resistant genes and bacteria resulting from the heavy use of antibiotics. Here, we showed that the novel Edwardsiella phage EPP-1 could achieve comparable efficacy to florfenicol using a zebrafish model of Edwardsiella piscicida infection and could reduce the content of the floR resistance gene in zebrafish excreta. Specifically, phage EPP-1 inhibited bacterial growth in vitro and significantly improved the zebrafish survival rate in vivo (P = 0.0035), achieving an efficacy comparable to that of florfenicol (P = 0.2304). Notably, integrating the results of 16S rRNA sequencing, metagenomic sequencing, and qPCR, although the effects of phage EPP-1 converged with those of florfenicol in terms of the community composition and potential function of the zebrafish gut microbiota, it reduced the floR gene content in zebrafish excreta and aquaculture water. Overall, our study highlights the feasibility and safety of phage therapy for edwardsiellosis control, which has profound implications for the development of antibiotic alternatives to address the antibiotic crisis.

Enzyme types and drug resistance of carbapenem-resistant Enterobacteriaceae isolated from Linyi area / 中华检验医学杂志

Objective:To detect the genotype of carbapenase and investigate the drug sensibility of Ceftazidime-avibactam (CAZ/AVI) on carbapenem-resistant Enterobacteriaceae (CRE), and to provide evidence for rational use of antibacterial drugs in clinical practice. Methods:A total of 179 strains of CRE were isolated from clinical specimens of patients treated in Linyi People′s Hospital from January 2019 to December 2020. mCIM/eCIM test and GeneXpert were used to detect the genotype of carbapenemases. The drug sensibility of CAZ/AVI was detected by K-B test.Results:One hundred and seventy-four out of 179 strains of CRE were positive upon to mCIM test (97.2%), 147strains were positive upon to eCIM test (84.5%). There were 27 serine carbapenemase (15.5%) and 147 metallo-β-lactamase (84.5%). The results of Fluorescent quantitative PCR rapid detection system developed by Saipei GeneXpert were consistent with the results detected by mCIM/eCIM. In the drug sensitivity test, 58 out of 174 mCIM positive strains were sensitive to CAZ (33.3%), of which the sensitivity of 27 strains producing serine carbapenemase was 96.3% (26/27) and all 147 strains producing metallo-β-lactamase were drug-resistant to CAZ/AVI.Conclusions:The carbapenase genotype of CRE in Linyi region is mainly metal β-lactamase. The CRE producing serine carbapenemase is highly sensitive to CAZ/AVI. It is helpful to guide the rational clinical use of the CAZ/AVI according to the detection results of CRE with or without carbapenemase production capacities.

Deskilled and Rapid Drug-Resistant Gene Detection by Centrifugal Force-Assisted Thermal Convection PCR Device.

Here we report the improved Cyclo olefin polymer (COP) microfluidic chip and polymerase chain reaction (PCR) amplification system for point-of-care testing (POCT) in rapid detection of Carbapenem-resistant Enterobacteriaceae (CRE). The PCR solution and thermal cycling is controlled by the relative gravitational acceleration (7G) only and is expected to pose minimal problem in operation by non-expert users. Detection is based on identifying the presence of carbapenemase encoding gene through the corresponding fluorescence signal after amplification. For preliminary tests, the device has been demonstrated to detect blaIMP-6 from patients stool samples. From the prepared samples, 96.4 fg/µL was detected with good certainty within 15 min (~106 thermocycles,) which is significantly faster than the conventional culture plate method. Moreover, the device is expected to detect other target genes in parallel as determination of the presence of blaNDM-1 and blaOXA-23 from control samples has also been demonstrated. With the rising threat of drug-resistant bacteria in global healthcare, this technology can greatly aid the health sector by enabling the appropriate use of antibiotics, accelerating the treatment of carriers, and suppressing the spread.

Drug resistance gene expression and chemotherapy sensitivity detection in Chinese women with different molecular subtypes of breast cancer.

Objective: The aim of the study was to identify specific chemosensitivity drugs for various molecular subtypes of breast tumors in Chinese women, by detecting the expression of drug resistance genes and by using the drug sensitivity test on different molecular subtypes of breast cancers. Methods: The expression of drug resistance genes including Topo II, GST-π, P-gp, LRP, and CD133 were detected with immunohistochemistry in a tissue microarray. Drug sensitivity tests included those for paclitaxel, epirubicin, carboplatin, vinorelbine, and fluorouracil and were conducted on primary cancer tissue cells and cell lines, including the T47D, BT-474, and MDA-MB-231 cells and human breast cancer xenografts in nude mice. Results: The different drug resistant genes Topo II, GST-π, P-gp, and LRP were differentially expressed among different molecular subtypes of breast cancers (P < 0.05). Positive expression of CD133 was highest in basal-like breast cancer (P < 0.05). Kaplan-Meier survival analysis showed that positive expressions of Topo II and CD133 both correlated with shorter disease-free survival (DFS) (P < 0.05) and overall survival (P < 0.05), and positive expression of LRP correlated only with shorter DFS (P < 0.05). BT-474 showed chemosensitivity to paclitaxel and epirubicin, while MDA-MB-231 showed chemosensitivities to paclitaxel, epirubicin, carboplatin, and fluorouracil (T/C ≤ 50%). The basal-like and HER2+ breast cancer primary cells showed chemosensitivities to paclitaxel and epirubicin with significant differences compared with luminal breast cancer primary cells (P < 0.05). Conclusions: The differential expression of drug resistance genes and the differential chemosensitivities of drugs in different molecular subtype of breast cancers suggested that individual treatment should be given for each type of breast cancer.

Molecular typing, antibiotic susceptibility, and biofilm production in nonencapsulated Streptococcus pneumoniae isolated from children in Japan.

The prevalence of nonencapsulated Streptococcus pneumoniae (NESp) has increased with the introduction of pneumococcal conjugate vaccines in children; however, the bacteriological characteristics of NESp have not been sufficiently clarified. In this study, NESp strains isolated from the nasopharyngeal carriage of children from four nursery schools in Japan were analyzed for molecular type, antibiotic susceptibility, and biofilm productivity. A total of 152 putative S. pneumoniae strains were identified by optochin-susceptibility analysis, of which 21 were not serotypeable by slide agglutination, quellung reaction, or multiplex PCR. Among these 21 strains, three were lytA-negative and, therefore, not S. pneumoniae. The remaining 18 strains were positive for lytA, ply, pspK, and bile solubility and were confirmed as NESp. Therefore, the isolation rate of NESp in the S. pneumoniae strains in this study was 12.0% (18/149). Molecular-typing analyses classified five strains as two existing sequence types (STs; ST7502 and ST7786), and 13 strains formed four novel STs. Horizontal spread was suspected, because strains with the same ST were often isolated from the same nursery school. The NESp isolates were generally susceptible to most antimicrobials, with the exception of macrolides; however, all isolates possessed more than one abnormal penicillin-binding protein gene. Furthermore, NESp strains were more effective than encapsulated counterparts at forming biofilms, which showed obvious differences in morphology. These data indicated that NESp strains should be continuously monitored as emerging respiratory pathogens.

Genomic epidemiology of Salmonella paratyphi A strains isolated in Hangzhou area / 中华微生物学和免疫学杂志

Objective To understand the genomic epidemiology of Salmonella paratyphi A strains circulating in Hangzhou area in recent years. Methods Next generation sequencing(NGS) technology was used to obtain genomes of 60 Salmonella paratyphi A strains isolated in Hangzhou area from 2002 to 2013. Genomes of 391 Salmonella paratyphi A strains were downloaded from the Sequence Read Archive (SRA) and Assembly database. After removing recombinations, the phylogenetic tree of all 451 genomes based on single nucleotide polymorphisms(SNP) was constructed using ATCC9150 as the reference. SRST2 and mul-tilocus sequence typing (MLST) were used to analyze sequence types(ST). The Salmonella In Silico Typ-ing Resource (SISTR) was used for core genome multilocus sequence typing (cgMLST). Resistant genes were screened out with SRST2 and BLASTN. Seven kinds of antibiotics were selected to conduct drug sus-ceptibility test in the 60 strains isolated in Hangzhou. Results A total of 19 258 SNP loci were found in 451 genomes. The average distance in the phylogenetic tree between strains was 0.007 0 and the distance less than 0.05 accounted for 96.73%, indicating a little difference in the 451 Salmonella paratyphi A ge-nomes. Fifty-eight Hangzhou strains of ST85 were highly similar in genomic sequences,which suggested that the clonal spread of ST85 strains caused the epidemic of paratyphoid A in Hangzhou area during 2002-2013. Salmonella paratyphi A strains isolated in Hangzhou were distantly related to five domestic strains (average distance:0.057),but close to 15 Yunnan strains(average distance:0.003 2) and closest to strains isola-ted in Cambodia (average distance: 0.001 8), suggesting the possibility of transnational spread of ST85 strains. Among two Hangzhou strains of ST129,HZ333 was closely related to two strains isolated in Jiangsu Province(average distance:0.009 7),suggesting the possibility of domestic transmission of ST129 strains. Except for two untyped strains,the other 58 strains were divided into nine cgMLST types. Except for 57 un-typed strains,the other 334 strains obtained from public databases were classified into 165 cgMLST types. Fifty-six out of the 60 Hangzhou strains carried the resistant gene aac6-Iy and the other four carried aac6-Iaa gene. Thirteen out of the 391 strains obtained from public databases didn′t carry resistant genes, while the other 378 strains carried the resistant gene aac6-Iy. Among the 60 Hangzhou strains,56 were sensitive to all of the seven kinds of antibiotics;three were resistant to cotrimoxazole and one was resistant to ampicillin and tetracycline. Conclusion The epidemic of paratyphoid A fever in Hangzhou from 2002 to 2013 was mainly caused by the clonal spread of ST85 strains that had the possibility of transnational spread. Some Hangzhou strains of ST129 had the possibility of domestic transmission. SNPs analysis had an advantage over pulse field gel electrophoresis(PFGE) technology in resolution and could be used to accurately trace the cause of paratyphoid A fever. Resistant gene aac6-Iy was generally carried. NGS technology had a promising prospect in the control and prevention of infectious diseases.

Drug resistant gene detection and homology analysis of Acinetobacter baumannii in patients with bloodstream infection / 国际检验医学杂志

Objective To investigate the drug resistant mechanism and the genetic homology of Acineto-bacter baumannii(A.ba)isolated from blood in the patients with bloodstream infection so as to provide a basis for the prevention and treatment of multi-drug resistant A.ba infection.Methods The different sites of sam-ples from the inpatients with bloodstream infection caused by A.ba were collected from January 2015 to June 2016.A total of 38 strains of A.Ba were isolated.The drug susceptibility test of all bacterial strains was per-formed according to the uniform operating specification.The carbapenemase gene was amplified by PCR and its products were analyzed by agarose gel electrophoresis.The homology study was performed by using multi-locus sequence typing(MLST)and eBURST analysis software.Results All 38 strains of A.ba were multi-drug-resistant.And in all of them,OXA-23-like and OXA-51-like gene,and ISAbal sequence were detected. OXA-23-like gene was associated with ISAbal sequence;the MLST typing showed that 38 A.ba strains mainly included the type ST195(55.3%),STn-2(15.8%)and STn-3(10.5%).The eBURST analysis indicated that the main prevalent clones were CC92,accounting for 86.8%.Conclusion Multi-drug resistant A.ba carries the same drug-resistant gene and its strain has higher genetic homology,indicating that the clone spread ex-ists,w hich provides a molecular biological basis for reducing the risks of A.ba related bloodstream infection in hospital.

Clinical application of Carba NP test in detecting CRE strains / 西安交通大学学报(医学版)

Objective To investigate the clinical application of Carba NP test in detecting carbapenem-resistant enterobacteriaceae.Methods Routine method was adopted to identify the 51 strains from clinical isolates and K-B test was conducted to screen out the carbapenem-resistant enterobacteriaceae and detect antimicrobial susceptibility.E-test was used to detect the minimum inhibitory concentration (MIC)of carbapenems.Modified Hodge test (MHT)and Carba NPⅠ test were used to screen out the phenotypes of carbapenemase,respectively. Then Carba NPⅡ test was used to classify carbapenemase which were positive in Carba NP I test.E-test was used for the detection of metalloenzymes.Polymerase chain reaction (PCR)was used for the detection of resistance genes. Results Totally 41 strains were positive in Carba NP 1 test and classified as B-class by Carba NPⅡ test.They were positive in metalloenzyme by E-test and carried NDM-1 gene by PCR.Conclusion Carba NP test can be used conveniently,has high consistency with PCR method,and the result is easy to determine.Therefore,it can be used as a routine laboratory method.

Research on rapid detection of Acinetobacter baumannii produced carbapenemase by CarbaNP method / 国际检验医学杂志

Objective To understand the phenotype and enzyme genotype of pan‐drug resistant carbapenemase‐producing Acine‐tobacter baumannii to provide the evidence for clinical rational use of antibiotics and monitoring hospital infection .Methods A total of 117 clinically isolated strains of Acinetobacter baumannii were collected and performed the routine microbiological detection . Multi‐drug resistant Acinetobacter baumannii was screened by K‐B disk diffusion method .The phenotype of carbapenemase‐produ‐cing strains was detected by using the Carba NP colorimetry and modified Hodge test .The drug resistant genotype of multi‐drug re‐sistant Acinetobacter baumannii was verified by PCR .Results Among clinically isolated 117 strains of Acinetobacter baumannii ,64 strains were multi‐drug resistant Acinetobacter baumannii ,in which 33 strains were carbapenemase positive .OXA‐23 drug‐resistant genotype of carbapenemase was detected by PCR ,while IMP ,VIM and NDM‐1 drug resistant genes were not detected .Conclusion The CarbaNP method can rapidly detect carbapenemase‐producing strains with the advantages of strong sensitivity and simple oper‐ation ,which conduces to improve the detection rate of carbapenemase‐producing strains and monitor the nosocomial infection .

ClassⅠ integron and its correlation with genes coding forESBLs in ESBLs-producing Escherichia coli from chil-dren / 临床儿科杂志

ObjectiveTo study the correlation with Genes Coding forESBLs and ClassⅠIntegron in ESBLs-producing Escherichia coli from children.MethodsPCR was used for gene typing detection of 100 strains of ESBLs-producingEsche-richia colistrains. While detection of class I integrase gene in the 100 strains ESBLs-producingEscherichia coli and 100 strains of non-ESBLs producingEscherichia coli were separately performed by PCR. It provides the solid base not only to reveal the relationship between class I integron and drug resistance, but also the relationship between class I integron and ESBLs-producing. ResultsThe most frequently genotyping from ESBLs-producingEscherichia coli in children isCTX-M (84%), followed by TEM-1(63%). The predominant distribution of genotype in ESBL- producing strains isTEM-1 +CTX-M (45%), followed by CTX-M (34%). Class I integrase gene detected in ESBLs- producing and non- ESBLs producing strain were 100 cases (100%) and 25 cases (25%) separately, the difference was statistically signiifcant (P<0.05); drug resistance in class I integron positive strains were signiifcantly higher than in class I integron negative strains, especially in Ciprolfoxacin, Levolfoxacin, and Amino-glycoside (86.4%, 88%, and 80%).ConclusionsThe distribution of Class I integron in ESBLs-producingEscherichia coli is signiifcantly higher than that in non-ESBLs-producing strains, It is rational that Class I integron highly correlate with strong drug resistance in ESBLs-producing strains.

Study on drug resistance genes forβ-lactams in multidrug-resistant Acine-tobacter baumannii / 中国感染控制杂志

Objective To investigate prevalence of resistance genes for β-lactams in clinically isolated multidrug-resistant Acinetobacter baumannii (MDRAB)in a hospital.Methods 22 clinically isolated MDRAB strains were performed antimicrobial susceptibility testing,resistance genes forβ-lactams (TEM,SHV,CTX-M,PER,DHA, IMP ,VIM ,SIM,OXA-23,OXA-24,and OXA-58)in these strains were detected with polymerase chain reaction. Results The resistant rates of 22 isolates of MDRAB to ceftazidime,cefotaxime,cefepime,gentamycin,amikacin, ciprofloxacin,and compound sulfamethoxazole were all 100.00%;to imipenem,meropenem,and cefoperazone/sul-bactam were 50.00%,40.91 %,and 31 .82% respectively,intermediated rates were 31 .82%,36.36%,and 31 .82% respectively.The carriage rates of OXA-23,TEM,IMP ,and PER were 100.00% (n =22),72.73 %(n=16),54.55% (n = 12),and 18.18% (n =4)respectively.Detection results of SHV,CTX-M,DHA,VIM , SIM,OXA-24,and OXA-58 were all negative.Conclusion Carriage of IMP ,TEM,PER,and OXA-23 resistance genes are the major resistance mechanisms of MDRAB to β-lactamase antimicrobial agents in this hospital.

Analysis of the genetic homology and resistant genes in clinical isolates of extensively drug-resistant Acinetobacter baumannii / 中国感染与化疗杂志

Objective To investigate the genetic homology in clinical isolates of extensively drug-resistant Acinetobacter baumannii (XDRAB),so as to provide evidence for better controlling hospital infections.Methods The genotypes of 28 clinical XDRAB isolates were determined by Enterobacterial Repetitive Intergenic Consensus PCR (ERIC-PCR).PCR was conducted to analyze 9 types of β-lactamase genes (blaKPC,blaIMP,blaVIM,blaNDM-1,blaOXA-23,blaOXA-24,blaOXA-48, blaOXA-51 and blaOXA-58),the outer membrane porin gene (CarO)and insertion sequence (IS)ISAba1.We also carried out linkage analysis for ISAba1-OXA23 and ISAba1-OXA58.Results Most of the above 28 XDRAB strains were isolated from respiratory tract specimens from intensive care patients.The result of ERIC-PCR showed that there was high homology between all the strains,suggesting that they might derive from the same clone.The genes blaOXA-23,blaOXA-51,CarO and the IS ISAba1 except Class Bβ-lactamase genes,blaOXA-24,blaOXA-48,blaOXA-58,ISAba1-OXA23 and ISAba1-OXA58 were detected in all the clinical strains by PCR.Conclusions All the XDRAB isolates belong to the same clone and carry the same drug-resistant genes,indicating that there was clone spread among XDRAB isolates.

Antimicrobial resistance characteristics and mechanisms of imipenem-re-sistant Pseudomonas aeruginosa / 中国感染控制杂志

Objective To investigate antimicrobial resistance characteristics and mechanisms of imipenem-resistant Pseudomonas aeruginosa (P .aeruginosa)isolated from clinical specimens.Methods Bacterial strains were identi-fied by BD Phoenix 100 automatic microbial analysis system,antimicrobial susceptibility testing was performed by Kirby-Bauer method;Carbapenemase genes (IMP ,VIM ,OXA,GES )and outer membrane protein gene oprD2 were detected by polymerase chain reaction.Results Resistant rates of imipenem-resistant P .aeruginosa strains to ami-kacin was the lowest (8.33%);resistant rates to gentamicin and tobramycin were60%,and all strains were resistant to ampicillin/sulbactam.The positive rate of OXA-17 gene was 2.78%(n=1 ),deletion rate of oprD2 was 38.89%,the other drug-resistant genes were not detected.Conclusion Except aminoglycosides,resistance of imipenem-resistant P .aeruginosa to other antimicrobial agents is serious;resistance of P .aeruginosa to imipenem may be due to lacking of OprD2 and production of carbapenemases.

Drug-resistant Gene and Disinfectant-resistant Gene for MRSA / 中华医院感染学杂志

OBJECTIVE To study the drug resistance,drug-resistant genes and(disinfectant)-resistant genes of(MRS)A.METHODS The drug resistance and mecA gene of the ?-lactamase and aac(6′)/aph(2″),aph(3′)-Ⅲ,ant(4′,4″) genes of aminoglycoside and qac(A/B) disinfectant-resistant genes were(detected) in 47 strains of MRSA.(RESULTS) In all 47 strains of MRSA,46 MRSA isolates were mecA positive,39 MRSA isolates were aac(6′)/aph(2″) positive,30 MRSA isolates were aph(3′)-Ⅲ positive,6 MRSA isolates were ant(4′,4″) positive,and 19 MRSA isolates were qac(A/B) positive.CONCLUSIONS MRSA is multiple-drug resistant.The main resistant mechanisms of MRSA to aminoglycosides and disinfectant are related to the drug-resistant genes of aminoglycoside and disinfectant-resistant genes.Clinic physician must pay attention to the diagnosis and(therapy) of MRSA,and control the hospital infection.

Drug Resistant Genes of CARB and aac(6′)-Ⅱ Prevailing in Pseudomonas aeruginosa Isolated from Burned Patients / 中华医院感染学杂志

OBJECTIVE To investigate drug resistant genes associated with beta-lactamases,aminoglycosides and sulfonamides in Pseudomonas aeruginosa isolated from burned patients.METHODS Common drug resistant genes associated with beta-lactamases,aminoglycosides and sulfonamides of 20 strains of P.aeruginosa isolated from burned patients were detected by PCR.RESULTS The positive rates of genes of TEM,aac(6′)-Ⅰb,sul1 and ant(3″)-Ⅰwere 85%,85%,90% and 10%,respectively and genes of CARB and aac(6′)-Ⅱ were both positive(100%).Genes of oprD2 were all absent and genes of SHV,GES,IMP,VIM,DHA and armA were all negative.Furthermore,drug resistant genes of strains isolated from different sites of one patient on different time could be different.CONCLUSIONS Drug resistant genes of CARB and aac(6′)-Ⅱprevail in P.aeruginosa isolated from burned patients.Detections of drug resistant genes in P.aeruginosa isolated,especially continuously isolated from burned patients are essential.

Relationship of Acinetobacter baumannii by Genetic Mark of Drug-resistant Gene,Integron and Transposon / 中华医院感染学杂志

OBJECTIVE To analyze the existence state of drug-resistant gene,integron and Tn21/Tn501 transposon and the relationship of Acinetobacter baumannii in Ningbo No.2 Hospital by genetic mark.METHODS Twenty seven strains of A.baumannii were clinically isolated.Drug-resistant gene,integron,and Tn21/Tn501 transposon were analyzed using polymerase chain reaction(PCR).The relationship was analyzed with cluster analysis methods.RESULTS The positive rates of blaTEM,blaOXA-23 cluster,blaADC,aac(3)-Ⅰ,aac(6′)-Ⅰ,ant(3″)-Ⅰ,qacE△1-sulⅠ,and intⅠ1 were 81.5%,44.4%,85.2%,85.2%,66.7%,81.5%,85.2%,and 85.2%,respectively.The others were negative.The result of multi-gene cluster analysis showed that the clone transmission existed.CONCLUSIONS The clone transmission of A.baumannii exists in Ningbo No.2 Hospital.The outbreak of A.baumannii may be possible because there are more than 3 clone transmission strains.