Objective To investigate the infection status and drug resistance of imipenem‐resistant Pseudomonas aeruginosa (PAE) isolated from the submitted specimens from children patients in our hospital between October 2012 to September 2013 for mastering the antimicrobial resistance status of Pseudomonas aeruginosa infection among children in Guangzhou and the occurrence situation of imipenem resistant strains .Methods The detection situation of Pseudomonas aeruginosa from infected children during this period was analyzed and the VITEK 2 Compact analyzer produced by the France bioMerieux company was used to identify the bacteria .The minimal inhibitory concentration(MIC) of imipenem resistant Pseudomonas aeruginosa to 11 kinds of antibiotics was detected .Results 161 strains of Pseudomonas aeruginosa were detected from 36 600 specimens ,including 24 strains(14 .9% ) of imipenem resistant Pseudomonas aeruginosa ,the positive strains were mainly originated from phlegmy (50 .3% ) ,the isolation was highest in ICU (27 .4% ) and NICU(21 .8% ) .The drug resistance rate of imipenem‐resistant P .aeruginosa was higher to ceftriax‐one sodium(91 .7% ) ,cefoperazone(29 .0% ) ,ceftazidime(29 .0% ) ,cefoperazone /shubatan(29 .0% ) and aztreonam (25 .0% ) .Con‐clusion Imipenem‐resistant Pseudomonas aeruginosa has the higher detection rate among children in this area and usually has re‐sistance to multiple antibiotics ,which should be paid more attention to and antibiotics should be rationally used .

Objective To explore the main pathogenic bacteria and antibiotic resistance patterns in children with bacterial diar‐rhea from Guangzhou region .Methods Regular bacterial culture of stool samples from children with suspicious bacterial diarrhea was performed to isolate the pathogen during 2011 to 2012 ,followed by the analysis of its composition and serum type ,ward distri‐bution characteristics and drug resistance to 12 antimicrobacterial drugs .Results 416 strains of pathogenic bacteria were isolated from diarrhea children during 2011-2012 ,in which salmonella ,enteropathogenic E .coli ,Campylobacter jejuni and Candida albicans isolates accounted for 53 .61% ,37 .98% ,5 .29% and 1 .68% respectively .Drug resistance rate of the main strains to 12 antimicrobi‐al agents was 85 .25% to ampicillin ,54 .28% to compound sulfamethoxazole ,44 .70% to cefotaxime ,42 .53% to ceftriaxone , 40 .66% to chloramphenicol ,23 .55% to ceftazidime ,23 .36% to aztreonam ,14 .88% to ciprofloxacin ,8 .07% to cefepime ,7 .99% to cefperazone/sulbactam ,7 .42% to piperacillin/tazobactam respectively ,and no resistance to imipenem was detected .Conclusion The pathogenic bacteria causing diarrhea mainly includes salmonella ,pathogenic e .coli ,campylobacter jejuni in children from guang‐zhou region ,the top five sensitive antimicrobial reagents for the main strains includes imipenem ,piperacillin/tazobactam ,cefpera‐zone/sulbactam ,cefepime and ciprofloxacin .

OBJECTIVE This study was conducted to analyze the underlying bacterial pathogens of the tonsils and adenoids in children with sleep-disordered breathing(SDB).METHODS The core tissue from the tonsils and/or adenoids of 163 SDB children was cultured aerobically. Of the 163 cases, 120 children underwent adenoidectomy and tonsillectomy simultaneously(A+T), 39 children underwent adenoidectomy(A) and 4 tonsillectom(T) only. 124 children who underwent tonsillectomy were subdivided into two groups based on history(with or without a history of recurrent tonsillitis). 71 children with the history were enrolled in the 'recurrent tonsillitis group' and 53 children without the history were enrolled in 'non- recurrent tonsillitis group'.RESULTS Of the total 120 cases who underwent A+T, 114(95.00%) cases had same distribution of bacteria detected in both sides in the same patient. Besides this, 17 cases in whom mixed organisms were identified in both sites shared common pathogen. No significant difference in the detection rates of staphylococcus aureus and haemophilus influenzae were found when we compared seasons(Tonsil:χ2=8.538,P=0.201; Adenoid:χ2=5.427, P=0.490). No significant difference in the type and detection rate of essential bacteria was found when we compared between recurrent tonsillitis group and 'non-recurrent tonsillitis group' (χ2=3.028,P=0.387).CONCLUSION The bacterial isolates from the tonsils and adenoids are virtually identical in type and detection rate in the same SDB patient. The bacterial distribution of the tonsillar and adenoidal core is unaffected by the seasonal variation and history of recurrent tonsillitis.

Objective To perform the amplification ,sequencing and prokaryotic expression of APH (3′′)‐Ⅰ and AAC (2′)‐Ⅰgenes from the clinically isolated gzch810 strain(SM gzch810)of Stenotrophomonas maltophilia to provide the basic materials for the next step functional test .Methods The SM gzch810 genome chromosome was extracted ,the APH (3′′)‐Ⅰ ,AAC (2′)‐Ⅰ whole genes were amplified by PCR and sequenced after being cloned into pMD18‐T vector .The recombination were subcloned into pGEX‐4T‐1 vector and the expression of the recombinant APH (3′′)‐Ⅰ and AAC (2′)‐Ⅰ were analyzed by SDS‐PAGE .Results The 800bp and 550bp DNA fragments of APH(3′′)‐Ⅰ ,AAC(2′)‐Ⅰ gene were amplified from SM gzch810 by PCR and sequenced ;the sequence comparison analysis showed that DNA and amino acid sequence identities of APH (3′′)‐Ⅰand AAC (2′)‐Ⅰ genes with other strains were 91% and 95% respectively .The sequence of APH (3′′)‐Ⅰand AAC(2′)‐Ⅰ of SM gzch810 were submitted to GenBank(accession number :HQ315852 and HQ315853);two major protein bands corresponding to the expected recombinant GST‐TP fusion proteins (56 × 103 and 46 × 103 respectively) were identified by SDS‐PAGE .Conclusion APH(3′′)‐Ⅰand AAC(2′)‐Ⅰgene of SM gzch810 are successfully cloned and expressed ,which lays a good foundation for further detecting corresponding antibi‐otic resistance and functional evaluation of above two kinds of recombinant E .coli .

Objective To analyse distribution and antibacterial resistance status of pathogenic bacteria isolated from blood cul‐tures of hospitalized infants ,in order to provide references for rational use of antimicrobial agents in the treatment of bloodstream infection .Methods A total of 299 strains of pathogenic bacteria isolated from positive blood culture specimens from infants(3 or less than 3 months of age) suspected with bloodstream infections in this hospital from January 2011 to May 2015 were collected ,the bacteria identification and drug sensitivity test were carried out by using the VITEK 2 Compact automatic microorganism analyzer . The composition and antibacterial resistance of these isolates were analyzed .Results Among the 299 strains of pathogenic bacteria , there were 169 strains of gram‐positive cocci(accounted for 56 .5% ) ,including 95 strains of coagulase negative Staphylococcus (ac‐counted for 31 .8% ) which was the main isolates ,and followed by 28 strains of Staphylococcus aureus(accounted for 9 .4% );there were 120 strains of gram‐negative bacilli (accounted for 40 .1% ) and mainly were Escherichia coli (53 strains ,accounted for 17 .7% );otherwise ,there were 8 strains of fungi (accounted for 2 .7% ) and 2 strains of gram‐positive bacillus (accounted for 0 .7% ) .The results of drug susceptibility test indicated that the gram‐positive cocci had multiple drug resistance to antibacterial a‐gents except for vancomycin and linezolid;the gram‐negative bacilli shown multiple drug resistance except for amikacin ,imipenem and meropenem .The fungus ,however ,displayed high sensitivty to all antifungal drugs .Conclusion Gram‐positive and gram‐nega‐tive bacteria are the main pathogens of hospitalized infants with bloodstream infection ,and are severely resistant to antibacterial a‐gents .Rational use of antimicrobial agents should be recommend for improving clinical efficacy and prohibiting the emergence of drug‐resistant strains .

Objective To understand the molecular epidemiology of penicillin resistance Streptococcus pneumonia (PNSP) isolated from children in Guangzhou area to provide the experimental basis for clinical prevention and control of Streptococcus pneumonia infectious diseases.Methods Specific primers were designed according to Genebank,penicillin binding protein(PBP) genes PBP1A,PBP1B,PBP2A,PBP2B,PBP2X,PBP3 were amplified by PCR.The sequencing analysis was performed.The PCR products were digested by Hinf I,and the restriction fragment length polymorphism (RFLP) was analyzed.Results DNA of PNSP was successfully extracted,the PCR results showed that in 50 strains of PNSP,the positive rates of bacterial strains containing PBP1A,PBP1B,PBP2A,PBP2B,PBP2X and PBP3 were 48.9%,64.4%,71.1%,31.1%,40.0% and 31.1% respectively.The sequencing showed that their homologies with known sequences in GenBank were 99%,98%,100%,97%,95% and 100% respectively.Using RFLP in Hinf I showed that PBP1A,PBP1B,PBP2A and PBP3 only had one kind of genotype,PBP2B and PBP2X had two kinds of genotypes,the positive rates were 71.4%,28.6%,66.7% and 33.3% respectively.Conclusion The gene distribution of PNSP strains among children in Guangzhou is dominated by PBP2A,PBP1B and PBP1A,there are two subtypes in PBP2B,PBP2X when digested by Hinf I,in which the predominant subtype >65%.

Objective To establish and optimize a loop-mediated isothermal amplification (LAMP) method for the rapid detection of Escherichia coli and its microbial toxin.Methods The LAMP reaction system and reaction conditions were determined by optimizing LAMP reaction,and the optimized LAMP system was used for the detection.Results Primers targeting shiga toxin (stx) gene and O157 antigen gene rfbe were designed.The established and optimized LAMP amplification system contained 1.2 mmol/L dNTPs,10 mmol/L MgSO4,0.4 mol/L betaine,1 μl 10 × Bst DNA polymerase Buffer,8 U Bst DNA polymerase fragment,2 μl DNA template,and the ratio of inner-primer (FIP and BIP) and outerprimer (F3 and B3) were 8∶ 1.Time and temperature for LAMP was 60 min,60 ℃.The sensitivity was 103 times higher than polymerase chain reaction (PCR),reached 5 × 101 CFU/ml.When LAMP was applied to 19 reference strains,102 EHEC strains,the specification was 100％ while identification rate of rfbe,stx1 and stx2 gene reached 100％,95.2％,92.9％.Conclusions The LAMP method showed a promising prospect for the rapid detection of common nosocomial pathogens microbial toxin.

Group B streptococcus ( GBS) is the most common cause of neonatal invasive infections in western countries , leading to severe pneumonia , sepsis and meningitis with high mortality and morbidity . Neonatal GBS infections are mainly transmitted from mother to baby .Since the end of 1990 s, prevention strategies of antenatal GBS screening and intrapartum antibiotic prophylaxis ( IAP) have been implemented by European and American area , decreasing the incidence of neonatal GBS disease significantly .The harm of GBS to neonates has been recognized and valued in recent years in China , but the authorized prevention measures are still lacking.Efficacy of IAP-based prevention depends on the accuracy of GBS screening results during pregnancy .Here, the GBS prevention guideline or consensus recommended by European and American CDC, including the screening strategies , detection methods, sample collection, storage and transportation were interpreted .

Objective:To establish a classification model for rapid identification of hypervirulent subtype ST17 clones of Group B Streptococcus (GBS) using matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS).Methods:In a retrospective study, 235 strains of GBS strains were selected from multiple centers in China during 2015-2018. For model generation,45 strains of ST17 and 50 strains of non-ST17 (20 ST19, 15 ST12 and 15 ST10 strains) were enrolled as the modeling group. The remaining 90 main ST strains (40 ST17, 16 ST10, 17 ST12 and 17 ST19) were served as validation group. 50 GBS strains classified as other minor ST subtypes were regarded as taxonomic groups. MS spectra were collected by Bruker mass spectrometry, and then loaded for model generation and verification, and screening of differential peptide peaks by genetic algorithm (GA) and model verification on ClinProTools 3.0 software.Results:The recognition rate for ST17-GA model were 99.4% with cross validation value of 96.9%. Among the ten differential peptide peaks for the classification model, the weights of both two main peptide peaks m/z 2 956 and m/z 5 912 were greater than 1, while the weights of the all other eight peptide peaks were less than 0.5. Model validation showed only one of the ST17 was misjudged as non-ST17 strain, resulting in diagnostic accuracy of 98.9%, sensitivity of 97.5% and specificity of 100%, positive predictive value of 100% and negative predictive value of 98.0%, respectively. For other sporadic STs, 42.0% (21/50) of them were misdiagnosed as ST17 subtype.Conclusion:A MALDI-TOF MS classification model for hypervirulent subtype of ST17 GBS strains has been successfully established with good diagnostic efficacy.

Objective To investigate the clinical characteristics,antibiotic susceptibilities and serotypes of Group B Streptococcus(GBS)isolated from neonatal meningitis to provide references for the prevention and treatment of neonatal GBS meningitis. Methods From June 2013 to June 2016,we surveyed the GBS strains iso-lated from purulent meningitis of < 90 days infants from Guangzhou Women and Children′s Medical Center. The GBS isolates were identified and the minimum inhibitory concentration of the antibiotics was determined by Vitek 2 Compact automatic bacterial identification system.GBS serotyping was performed using Strep-B-Latex?rapid latex agglutination test kit. Results A total of 46 cases of neonatal GBS meningitis,15 cases of early-onset infection and 31 cases of late-onset infection were diagnosed. 78.3% of GBS meningitis with varying degrees of complica-tions.Among 41 survivors with 3~24 months follow-up,50% of the early-onset and 44.8% of the late-onset GBS meningitis with varying degrees of neurological sequelae.Four capsular types were identified among the 46 isolates, serotype-Ⅲwas the most prevalent(73.9%),followed by Ib(19.6%),V(4.3%)and Ia(2.2%).All the isolates were susceptible to penicillins,cephalosporins,linezolid and vancomycin. Conclusion The highly pathogenic serotype-Ⅲ was the predominant serotype among neonatal GBS meningitis in Guangzhou,Therefore,it is neces-sary to strengthen the epidemiological surveillance of GBS invasive infection and the effective implementation of pre-ventive measures.