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Objective:To study the changes of β-lactam resistance of Haemophilus influenzae (Hi) strain isolated from neonatal lower respiratory tract and the molecular mechanism of β-lactam resistance.Methods:Nineteen Hi strains isolated from neonatal lower respiratory tract infection in the previous multicenter prospective epidemiological study were re-identified, and the P6, fucK and Cap genes were detected by PCR.The minimum inhibitory concentration(MIC) of ampicillin, amoxicillin clavulanic acid and cefuroxime were detected by microdilution method, and tem-1 gene, rob-1 gene and ftsI gene were sequenced and analyzed.Results:(1) Nineteen strains of Hi were confirmed to be capsule-free type by P6 gene, fucK gene and cap gene, which was non-typeable Haemophilus influenzae(NTHi). (2)Compared with 2003-2004, the MIC values of ampicillin, amoxicillin clavulanic acid and cefuroxime of NTHi isolated from the lower respiratory tract of the newborn from 2013-2014 were significantly higher( P<0.05). (3)The rates of β-lactamase producing strains during 2003-2004 and 2013-2014 were 33.33% (3/9) and 30.00% (3/10), respectively.There was no significant difference between them during 10 years ( P>0.05). The detection of the β-lactamase gene showed that the β-lactamase of the all six strains were of the tem-1 type, and the rob-1 type was not detected.(4)Only one gBLNAR strain ( n=9) was found during 2003~2004, and gBLNAR 1, gBLNAI 3, gBLPAR 3, gBLPACR 1 ( n=10)appeared during 2013~2014.(5)There were 11 amino acid substitution patterns in ftsI gene during 2013~2014, but only five amino acid substitution patterns in 2003~2004.The mutation rate of the S357N, S385T, N526K and T532S of ftsI gene significantly increased during the past ten years ( P<0.05). One strain of gBLNAR/gBLNACR resistant to ampicillin, amoxicillin clavulanic acid and cefuroxime isolated in 2014 showed D350N, S357N, M377I, S385T, L389F, A502T and N526K variation at the same time. Conclusion:Neonatal patients with lower respiratory tract NTHi infection may rapidly face the severe challenge of multiple drug resistance of β-lactam antibiotics.
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Objective Broth dilution method was used as a reference method to observe the capability of Kirby-Bauer disc diffusion assay(K-B)for correcting automated ampicillin susceptibility detection of He-mophilus influenzae(HI).Methods A total of 228 HI strains isolated were collected,broth dilution assay,K-B and automated microdilution broth test(ATB)were used to determine the susceptibility of HI to ampicillin. Analyze the essential agreements of the three methods and the correction of K-B to the errors of A TB. Results The essential agreement of K-B or ATB with broth dilution method were 77.19%,70.18% respec-tively,combination of K-B and ATB could make the essential agreement increase up to 86.0%,which was sig-nificantly higher than ATB(χ2=16.600,P=0.000).Major error of ATB(42.0%)was higher than that of K-B(10.0%)(χ2=13.306,P=0.001),but very major error and minor error showed no significant difference be-tween the two methods(χ2=1.208,P=0.272;χ2=1.182,P=0.227),meanwhile,76.19% of major error of ATB could be corrected by K-B.For the very major error of ATB,53.57% could be corrected by K-B.Howev-er,the corrective capability of K-B to minor error of ATB was relative low.Conclusion K-B test could correct some errors generated by ATB.For the β-lactamase negative strains which were judged as ampicillin resistance by A TB,K-B test should be used to correct the errors by ATB.Moreover,it is necessary to apply K-B to confirm am-picillin sensitivity of the β-lactamase positive strains which were judged as ampicillin susceptible by ATB.
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Objective To investigate the epidemic strains of biological type, drug resistance, and the basic clinical characteristics of haemophilus influenzae(Hi)isolated from hospitalized adults with lower respiratory tract infection in Chengdu area.Methods A prospective cross-sectional study was conducted to analyze the biological typing,capsular genes detected by PCR technique,and drug resistance tested by drug sensitive test of Hi epidemic strains isolated from the sputum of adults aged above 18 years who were hospitalized in two tertiary hospitals of west Sichuan in China.Results The positive rate of pathogenic bacteria in adults aged above 18 years who were hospitalized in the two hospitals was 46.71%(15 447/33 069)between November 2013 and October 2014.The positive rate of Hi isolated from the sputum of 100 adults with lower respiratory tract infection was 0.31%(101/33 069).The constituent ratio of Hi in lower respiratory tract infection pathogens was 0.65%(101/15 447).The Hi were all undifferentiated type detected by PCR,and the biological typing of Hi were typeⅠ(42.57%),Ⅳ(29.7%),Ⅱ(15.84%),Ⅲ(9.9%),Ⅶ(1.98%), and Ⅵ(0.9%).The diseases of Hi positive were acute phase of chronic obstructive pulmonary disease(59%), pneumonia(35%), and bronchitis(6%), in which community acquired infection was 55%.The rate of β-lactamase enzyme production was 38.61%.The frequencice of β-lactamase -nonproducing-ampicillin-resistant(BLNAR)strains was 2.97%,and of intermediary strains was 4.95%(5/101).The drug resistance rate of amoxicillin and clavulafiate was 2.97%.The drug resistance rate of cefuroxime was 12.87%, and intermediary rate was 12.87%.The drug resistance rate of cefaclor was 29.7%,and intermediary rate was 8.91%.The drug resistance rate of cefotaxime and ofloxacin was 6.93%and 1.99%.There were no obvious statistical differences between the drug resistance rates of the two hospitals.Conclusions The Hi epidemic strains isolated from the sputum of adults with lower respiratory tract infection were all undifferentiated type,and the common biological types were Ⅰ,Ⅳ,Ⅱ, andⅢ in west Sichuan in China.It should pay attention to the BLNAR strains and ofloxacin-resistant strains.