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Objective:To investigate the characteristics of hemolysis, resistance and homology of Staphylococcus cohnii urealyticum. Methods:A retrospective study was carried out on thirteen clinical strains of Staphylococcus cohnii urealyticum. They were re-identified by MALDI-TOF MS. Their colony and hemolytic characteristics on blood agar plates were observed. The co-hemolysis between Staphylococcus cohnii urealyticum and Staphylococcus aureus was demonstrated. The hemolysin genes and drug resistance genes were detected by PCR. Pulsed field gel electrophoresis and mass spectrometry were used to analyze the homology of strains. The susceptibility of strains to antimicrobial agents was detected by agar dilution. Results:The confirmed 13 strains of Staphylococcus cohnii urealyticum showed various levels of hemolysis and had enhanced synergistic hemolysis with Staphylococcus aureus. All strains were susceptible to vancomycin and tigecycline. There were 12 strains which carried mecA gene, 7 strains carried cfr gene, 7 strains carried ermC gene. The 13 strains were divided into 3 groups by MALDI-TOF MS, and 6 types by pulsed field gel electrophoresis. Conclusions:Clinical strains of Staphylococcus cohnii urealyticum demonstrated various levels of hemolysis which could be enhanced by Staphylococcus aureus. Although they carried different drug resistance genes, they were all susceptible to vancomycin and tigecycline.
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Objective@#To investigate the characteristics of hemolysis, resistance and homology of Staphylococcus cohnii urealyticum.@*Methods@#A retrospective study was carried out on thirteen clinical strains of Staphylococcus cohnii urealyticum. They were re-identified by MALDI-TOF MS. Their colony and hemolytic characteristics on blood agar plates were observed. The co-hemolysis between Staphylococcus cohnii urealyticum and Staphylococcus aureus was demonstrated. The hemolysin genes and drug resistance genes were detected by PCR. Pulsed field gel electrophoresis and mass spectrometry were used to analyze the homology of strains. The susceptibility of strains to antimicrobial agents was detected by agar dilution.@*Results@#The confirmed 13 strains of Staphylococcus cohnii urealyticum showed various levels of hemolysis and had enhanced synergistic hemolysis with Staphylococcus aureus. All strains were susceptible to vancomycin and tigecycline. There were 12 strains which carried mecA gene, 7 strains carried cfr gene, 7 strains carried ermC gene. The 13 strains were divided into 3 groups by MALDI-TOF MS, and 6 types by pulsed field gel electrophoresis.@*Conclusions@#Clinical strains of Staphylococcus cohnii urealyticum demonstrated various levels of hemolysis which could be enhanced by Staphylococcus aureus. Although they carried different drug resistance genes, they were all susceptible to vancomycin and tigecycline.
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Objective@#To understand the in vitro growth characteristics of Cryptococcus gattii VGⅠ and VGⅡ isolated in China and the diversity in their virulence to Galleria mellonella.@*Methods@#Based on the results of multilocus sequence typing for eight strains of Cryptococcus gattii isolated in China, the strains were cultured in vitro to draw growth curves, observe the melanin production and measure the capsule thickness. The median lethal time (LT50) and median lethal dose (LC50) at 48 h of Cryptococcus gattii were calculated using Galleria mellonella infection test. Fourteen strains of Cryptococcus neoformans were studied for comparison.@*Results@#The eight Cryptococcus gattii strains were six VGⅠ and two VGⅡ. The growth curves of Cryptococcus gattii VGⅠ and VGⅡ were similar to that of Cryptococcus neoformans when culture at 30℃. The total number for each of them could reach 108 CFU/ml at 96 h under 30℃. However, the total number at any time point at 37℃ was less than that at 30℃. There was no significant difference in the amount of melanin produced by Cryptococcus neoformans under 30℃ and 37℃, but both VGⅠand VGⅡ types of Cryptococcus gattii could produce more amount of melanin under 37℃ than under 30℃. The ratio of capsule/cell wall diameter of Cryptococcus gattii VGⅠwas greater at 37℃ than that at 30℃ with statistical significance (P<0.001). Cryptococcus neoformans showed the longest LT50, followed by VGⅠand VGⅡ types of Cryptococcus gattii. The LT50 of Cryptococcus gattii VGⅡ at the concentration of 1×106 CFU/ml was 72 h, and its LC50 at 48 h was 1×108 CFU/ml.@*Conclusions@#Like Cryptococcus neoformans, Cryptococcus gattii VGⅠ and VGⅡ grew faster under 30℃ than under 37℃, but more melanin was produced and thicker capsule was formed under 37℃ than under 30℃. Among Cryptococcus neoformans and VGⅠ and VGⅡ types of Cryptococcus gattii, Cryptococcus gattii VGⅡ showed the shortest LT50 and the strongest virulence to Galleria mellonella.
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Objective To understand the in vitro growth characteristics of Cryptococcus gattii VGⅠ and VGⅡ isolated in China and the diversity in their virulence to Galleria mellonella. Methods Based on the results of multilocus sequence typing for eight strains of Cryptococcus gattii isolated in China, the strains were cultured in vitro to draw growth curves, observe the melanin production and measure the capsule thickness. The median lethal time (LT50 ) and median lethal dose (LC50 ) at 48 h of Cryptococcus gattii were calculated using Galleria mellonella infection test. Fourteen strains of Cryptococcus neoformans were studied for comparison. Results The eight Cryptococcus gattii strains were six VGⅠ and two VGⅡ. The growth curves of Cryptococcus gattii VGⅠ and VGⅡ were similar to that of Cryptococcus neoformans when culture at 30℃ . The total number for each of them could reach 108 CFU/ ml at 96 h under 30℃ . However, the total number at any time point at 37℃ was less than that at 30℃ . There was no significant difference in the amount of melanin produced by Cryptococcus neoformans under 30℃ and 37℃ , but both VGⅠand VGⅡtypes of Cryptococcus gattii could produce more amount of melanin under 37℃ than under 30℃ . The ratio of capsule/ cell wall diameter of Cryptococcus gattii VGⅠwas greater at 37℃ than that at 30℃ with statistical significance (P<0. 001). Cryptococcus neoformans showed the longest LT50 , followed by VGⅠand VGⅡtypes of Cryptococcus gattii. The LT50 of Cryptococcus gattii VGⅡ at the concentration of 1×106 CFU/ ml was 72 h, and its LC50 at 48 h was 1×108 CFU/ ml. Conclusions Like Cryptococcus neoformans, Cryptococcus gattii VGⅠ and VGⅡ grew faster under 30℃ than under 37℃ , but more melanin was produced and thicker capsule was formed under 37℃ than under 30℃ . Among Cryptococcus neoformans and VGⅠ and VGⅡtypes of Cryptococcus gattii, Cryptococcus gattii VGⅡ showed the shortest LT50 and the strongest virulence to Galleria mellonella.
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Hypervirulent Klebsiella pneumoniae ( hvKP) mainly infects healthy people and causes serious infections, such as liver abscess, meningitis, necrotizing fasciitis, endophthalmitis and severe pneu-monia. Studies have shown that hvKP is more virulent than classic Klebsiella pneumoniae characterized by ex-pressing more capsular polysaccharide and carrying the virulence factors including magA, rmpA and iron ac-quisition molecules. The greater survival and anti-phagocytosis abilities of hvKP strains contribute to the spread and metastasis of hvKP infection. This review describes the virulence factors, colonization and infec-tion of hvKP as well as the host immunity to hvKP.
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Objective To detect the hypermucoviscosity phenotype , capsular serotype and virulence gene of Klebsiella pneumonia (K.pneumonia) from various kinds of clinical specimens and understand the characteristics of different K.pneumonia causing infections.Methods A retrospective study was conducted through collection of 178 K.pneumonia isolates from blood, sputum, bronchoalveolar lavage fluid , urine, normally sterilized fluid , puncture fluid from liver abscess and other abscesses between January 2010 and December 2012 in General Hospital of Chinese PLA.String test was carried out for detection of hypermucoviscosity phenotype.Capsular serotype and virulence gene ( rmpA) were checked by polymerase chain reaction.Analysis was made according to the hypermucoviscosity , capsular serotype , rmpA gene , as well as the sources of K.pneumoniae.Statistic data was analyzed by contingency table analysis and χ2 test.Results Eighty-three out of 178 ( 46.6%) strains of K.pneumonia were hypermucoviscous with positive string test, the positive rate of virulence gene rmpA was 92.8%(77/83).K1/K2/K57 capsular serotypes were the predominant serotypes in the group of puncture fluid from liver abscess and other abscesses (75.0%,27/36)than the group of blood (32.4%,12/37), urine(21.7%,5/23) and normally sterilized fluid(25.0%,5/20), and also more than in the group of sputum , bronchoalveolar lavage fluid (50.0%,22/44),χ2 =21.19,P<0.01.The positive rate of string test in the group of puncture fluid from liver abscess and other abscesses (77.8%, 28/36)was significantly higher than the group of blood (29.7%, 11/37), urine (30.4%, 7/23), or normally sterilized fluid (25.0%, 5/20),χ2 =27.90,P<0.01.The positive rate of rmpA gene in the isolates from puncture fluid of liver abscess and other abscesses was higher than other groups.Conclusions As the pathogens of various kinds of infections , mainly abscess and respiratory infection, hypermucoviscous strains of K.pneumonia were of great clinical significance.Capsular serotype K57, as well as K1 and K2, possessed hypermucoviscosity and hypervirulence in China.
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Objective To study the education of medical mycology for undergraduates of medical laboratory specialty and provide a basis for teaching reformation.Method Setting of mycology related courses of medical mycology for undergraduates in 5 medical schools and 85 inspection and technical personnel's detection of fungi in 81 hospitals were investigated through consultation and questionnaire survey.Results More than 140 class hours for medical mycology were arranged in 5 schools,but as to medical mycology,22 class hours in 1 school and less than 10 class hours in 4 schools,the minimum class hours were 5.Although various numbers of Candida and filamentous fungi could be isolated in hospitals investigated,more than half laboratory workers could not identify penicillium,thermally dimorphic fungi,Zygomycetes and Dematiaceous fungi.Conclusion Education on medical mycology for medical laboratory specialty undergraduates is insufficient and the corresponding teaching lacks such content as medically important pathogenic fungi detection methods and identification characteristics.The hospital technical personnel's fungal identification ability cannot meet the situation of increasing fungal infection involved in clinical medicine,so it is necessary to carry out teaching reformation of medical mycology for undergraduates in laboratory medicine,including adding class hours,increasing course contents and so on.
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Objective To detect the main virulence genes and biofilm formation of Enterococci strains isolated from blood samples .Methods Twenty-eight strains of Enterococcus faecalis ( E.faecalis) and 54 strains of Enterococcus faecium ( E.faecium) were collected from blood samples .Five main virulence genes (asa1, esp, hyl, cylA and gelE) were detected by multiplex PCR.Biofilm formation was investigated by using microtiter dish biofilm formation assay .Results All E.faecalis strains were positive for at least one kind of virulence genes , of which 14 strains were concurrently positive for asa1, esp, cylA and gelE.asa1, cylA and gelE were only detected in E.faecalis strains, while hyl gene only existed in E.faecium strains. Twenty-seven strains of E.faecium were esp positive, of which 12 strains were both hyl and esp positive. None of the 5 virulence genes were identified in 10 strains of E.faecium.85.7% of E.faecalis strains and 63.0%of E.faecium strains could form biofilm.Conclusion Compared with E.faecium strains, more types of virulence genes were detected in E.faecalis strains with higher positive rates .Moreover , E.faecalis strains were more likely to form biofilms than E.faecium strains.
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ObjectiveTo evaluate the application value of the quantitative procalcitonin (PCT) test in bloodstream infection.Methods Of 1066 patients with blood culture and PCT detection were collected in our hospital,retrospectively,1010 were effective cases.The relationship between blood culture results and serum PCT levels was investigated.PCT levels in gram-negative bacterial infection,gram-positive bacterial infection and candidiasis were compared.The prognosis of 33 blood culture positive patients with repeated PCT detection results were analyzed.Mann-Whitney U test was used to compare the PCT value among the three groups,and Fisher' s test was used to compare the death rate among the three groups.ResultsIn the patients with negative blood culture results,the median of PCT was 0.37 (0.11 - 1.67) μg/L.But in the patients with positive blood culture results,the median of PCT were 2.24(0.57 -11.59) μg/L The positive rate of PCT in gram-negative bacteria infection,gram-positive bacterial infection and candidiasis were 86.6％,72.0％ and 75.7％,respectively.In the 33 patients subjected to repeated PCT detections,the mortality of the patients with decreasing PCT was lower than the others.The patients whose PCT levels were greater than 5 μg/L had poor prognosis.ConclusionsQuantitative PCT is proved to be an effective method for rapid diagnosis of bloodstream infection.The changing trends of PCT test results has certain reference value for the patients' prognosis.
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ObjectiveTo establish the rapid molecular diagnosis of 16 common coagulase negative Staphylococcus(CNS).MethodsDNA sequencing of 16 CNS would be obtained with gap gene.After the alignment gap gene sequences which were available in the GenBank,the bacteria were identified with homological alignment and phylogenetic tree,and compared with the 16S rRNA gene.ResultsThe sequence similarity of the gap sequences ranged from 39% to 98% in 16 CNS.There were the highest similarity (98%) between S.hominis and S.hominis subsp,and the lowest(39% ) between S.saprophyticus and S.xylosus.The sequence similarity of the 16S rRNA sequences ranged from 96 to 98%,at least two species of bacteria similar rate of 99% and the most four species similar rate of 99%.Phylogenetic homology analysis showed that it was a high confidence(99% ) in the detection ofS.xylosus and S.lentus,S.chromogenes and S.intermedius,S.hominis and S.hominis subsp,but for 10 other species of bacteria,gap homology analysis has less unreliable confidence(49%,56% ) and 16S rRNA has more unreliable confidence(43%,43%,50%,56%,63%,65%,76% ).ConclusionAnalysis of gap sequence could identify 16 CNS timely and accurately,with higher confidence than 16S rRNA.
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OBJECTIVE To study the prevalance and resistance characteristics of clinical isolates of Enterobacter cloacae.METHODS Clinical isolates of E.cloacae were studied by K-B methods,and the data of MIC were(analyzed ) by WHONET 5.3 software.RESULTS Totally 416 clinical isolates of E.cloacae isolated from 2000 to 2004 were studied and they were mainly isolated from department of respiration,surgical ICU and department of(neurology) and their rate was 25.2%,9.6%,and 8.6%,respectively.The E.cloacae strain was(resistant) to ciprofloxacin,amikacin and most of penicillins,cephalosporins and ?-lactams combined with the(?-lactamase) inhibitors.It was susceptible to cefepime,imipenem and etrapenem.CONCLUSIONS The E.cloacae is resistant to many commonly used antibiotics in clinics and it is important to control antibiotic(resistance) by using antibiotics reasonably.
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OBJECTIVE To establish the implanted biofilm model of rabbit implant infection.METHODS SEP was cultured,purified,identified,and tested for the drug sensitivity and the biofilm production.Healthy adult rabbits were randomly divided into 4 groups,8 in each group.The stainless steel screws and washer UHMWPEs were implanted into the femoral condyle of the non-articular surface of the rabbit right stifle knee.The knee joints were inoculated with 1ml sterile saline,102,103 and 104 CFU SEP,respectively.Wounds were observed postoperatively.After the 14th days postoperation,the knee synovium of 32 rabbits sacrificed were taken out by aseptic technique and were cultured to determine whether the knees were infected and which concentration of SEP was the appropriate for causing knee joint infection.UHMWPEs from the appropriate ID group were observed to find whether there were biofilms on the surface by SEM and LCSM.RESULTS The bacterial strain was identified as SEP and could produce biofilm.Among the knee joints inoculated with 1ml saline,102,103 and 104 CFU SEP,the infective rate was 0,37.5%,100.0% and 100.0% and poor wound healing was in 0,1,2 and 4 rabbits,respectively.It showed 103 CFU of SEP was the appropriate ID.Biofilms were found on all UHMWPE surfaces from 103 CFU SEP group by SEM and LCSM.SEM showed SEP in the biofilms on the surface of UHMWPE was agglomerated and wrapped in the matrix.The structure of biofilms in which SEP radiated red fluorescence was inlaid in mucopolysaccharide stained green fluorescence was observed by LCSM.CONCLUSIONS The model provides an effective method to investigate the biofilm.
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Objective To develop a novel multiplex polymerase chain reaction (PCR) to detect multidrug-resistant Acinetobacter baumannii.Methods One hundred and five strains of multidrug-resistance A. baumannii were isolated from January 2006 to April 2007. The bacterial DNA was obtained by boiling the pure growth of A. baumannii. All isolates were subjected to the multiplex PCR to detect genes of blaOXA-23-like,blaOXA-24-like,blaOXA-51-like,blaOXA-58-like,intI 1 and intI 2.Results Among 105 isolates,76 were positive for blaOXA-51-like,blaOXA-23-like,and intI 1,18 were positive for blaOXA-51-like and intI 1,10 were positive for blaOXA-51-like and blaOXA-23-like,1 was positive for blaOXA-51-like and blaOXA-23-like,1 was positive for blaOXA-51-like,blaOXA-23-like,and blaOXA-58-like,and all were negative for blaOXA-24-like and intI 2.Conclusion The presence of OXA carbapenemase and integrase genes was correlated with multidrug resistance in A.baumannii.
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OBJECTIVE To evaluate the antibiotic effects of polymyxin B combined with meropenem against 110 strains of multidrug-resistant Acinetobacter baumannii.METHODS The protocol was designed by checkerboard method and the MICs of polymyxin B combined with meropenem against the 110 strains of A.baumannii were determined by broth dilution method,the FIC index was calculated according to MIC results.RESULTS The percentage of the FIC indexes less than 0.5,from 0.5 to 1,from 1 to 2 and more than 2 were 98%,2%,0% and 0%,respectively.CONCLUSIONS When polymyxin B combined with meropenem against 110 strains of A.baumannii the synergism and additivity are the main,there are no autonomy and antagonism.
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OBJECTIVE To investigate the in vitro effect of 4 kinds of implant materials on the biofilm formation of Staphylococcus epidermidis.METHODS S.epidermidis was cultured,purified and identified.Susceptibility test was done for S.epidermidis and the ability to produce biofilm was proven.The test samples were made into wafer shape for titanium alloy,Co-Cr-Mo alloy,ultrahigh molecular weight polyethylene(UHMWPE)and home-made polymethyl methacrylate(PMMA).S.epidermidis was cultured with 4 kinds of test samples for 5 days respectively.Bacteria adhering on surfaces of 4 kinds of test samples were dissolved with trypsin,and then diluted into bacterial suspensions.Each bacterial suspension was inoculated quantitatively and CFU were counted.Biofilm on surfaces of 4 kinds of test samples prepared by vacuum drying method was observed with SEM.RESULTS The strain proven to be S.epidermidis,was resistant to semisynthetic penicillins,and could produce biofilm.CFU count showed that CFU were the most on the UHMWPE surface and the number of CFU were(24.96?1.459)?105.CFU were(17.44?1.883)?105 on the PMMP surface.(0.424?0.065)?105 CFU were discovered on the surfaces of titanium alloy and(0.382?0.075)?105 CFU on Co-Cr-Mo alloy.When each group compared with UHMWPE and domestic PMMA respectively,all P value was
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Objective To study the function of aminophenylboronic acid(APB)and clavulanate(CA)for detecting ESBLs in Enterobacter cloacae.Methods The phenotype of ESBLs of 61 Enterobacter cloacae isolates was detected with adding single beta-lactamase inhibitor CA to ceftazidime(CAZ)and cefotaxime(CTX),and double beta-lactamase inhibitors CA/APB to ceftazidime(CAZ)and cefotaxime(CTX)respectively.PCR was used to detect ESBLs genes of 61 Enterobacter cloacae isolates.The results of the enzymatic inhibitor potentiation test and PCR were compared and analyzed.Results With adding single enzymatic inhibitor CA to CAZ,28 isolates of Enterobacter cloacae producing ESBLs were detected,while 14 isolates were detected with adding CA to CTX.With adding double enzymatic inhibitors CA/APB to CAZ,28 isolates of Enterobacter cloacae producing ESBLs were detected,while 44 isolates were detected with adding CA/APB to CTX.By PCR positive ESBLs genes were detected in 47 isolates of Enterobacter cloacaes.Conclusions The potentiation test with double beta-lactamase inhibtion can be used to detect ESBLs in Enterobacter cloacae.
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OBJECTIVE To investigate the phenotype and genotype of plasmid-encoded AmpC and extended spectrum beta-lactamases in Klebsiella pneumoniae.METHODS 3-Aminophenylboronic acid(APB) test and ESBLs confirmatory test were used for phenotypic detection of AmpC and ESBLs.Conjugation was conducted in order to understand the spread of plasmid in bacteria.The size and genotype of ampC and ESBL genes were studied by extraction and purification of plasmid,PCR and sequencing analysis.RESULTS A plasmid of about 15kb was extracted from K.pneumoniae.This plasmid carrying resistance genes to antibiotics could be spread from K.pneumoniae to recipient Escherichia coli NK5449 through conjugation.DHA-type ampC gene and SHV-type ESBLs gene could be amplified from plasmids extracted from both K.pneumoniae and its conjugant in E.coli,they were DHA-1 ampC gene and SHV-12 ESBLs gene confirmed by sequencing analysis.CONCLUSIONS DHA-1 ampC gene and SHV-12 ESBLs gene are detected from the plasmid of K.pneumoniae.
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OBJECTIVE To investigate the prevalence of DHA AmpC ?-lactamases mediated by plasmid in Klebsiella pneumoniae in China.METHODS Antimicrobial susceptibility test was conducted by the methods of double agar dilution and ESBLs confirmatory in K-B method according to the criteria of guidelines of CLSI.AmpC ?-lactamases were detected on the basis that AmpC ?-lactamases could be inhibited by 3-aminophenylboronic acid(APB).Gene chip technology and PCR were used to detect ESBLs and AmpC gene.RESULTS Among total 34 isolates of K.pneumoniae 32(94.1%) produced AmpC ?-lactamases and ESBLs.The most common(38.3%) were types DHA and TEM and SHV.MIC50 and MIC90 of all strains to all tested antimicrobial agents were lower than 34 strains tested 0.25?g/ml and 0.5?g/ml.Fourteen strains AmpC and ESBLs were conjugated successfully.CONCLUSIONS DHA AmpC ?-lactamases mediated by plasmid are the most common in K.pneumoniae in General Hospital of PLA of China.The most common(38.3%) are types DHA and TEM and SHV.Fourteen(41.2%) strains can be spreaded by plasmid.
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OBJECTIVE To study the phenotypic existence,genetic type and gene transfer of extended spectrum beta-lactamases(ESBLs) and AmpC beta-lactamase from Klebsiella pneumoniae and K.oxytoca. METHODS Disk confirmation test and 3-aminophenylboronic acid(APB) disk potentiation test were used to detect ESBLs and AmpC beta-lactamase.The genetic types of these two kinds of beta-lactamases were examined by gene chip technology and sequence analysis.The transfer of resistance genes was conducted by conjugation. RESULTS From 72 strains of K.pneumoniae and 20 strains of K.oxytoca which were not susceptible to cefoxitin,coexistence of AmpC(beta-lactamase) with ESBLs together was very common,accounted for 54.2% and 75.0%,single ESBLs accounted for 22.2% and 25.0%,respectively.There were 12.5% single AmpC in(K.pneumoniae).DHA type ampC gene and SHV type ESBLs gene were the main molecular types.These genes could be transferred from clinical isolates to recipient E.coli J53. CONCLUSIONS ESBLs as well as AmpC(beta-lactamase) are the most important resistance mechanism in K.pneumoniae and K.oxytoca.The resistance could be transferred through the bacterial conjugation.
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OBJECTIVE To evaluate the antibacterial effect of cefoperazone-sulbactam combined with minocycline or levofloxacin against 110 strains of multidrug-resistant Acinetobacter baumannii.METHODS Checkerboard method was designed for the minimal inhibitory concentration(MIC) testing of cefoperazone-sulbactam combined with minocycline or levofloxacin against 110 strains of multidrug-resistant A.baumannii by agar dilution method,fractional inhibitory concentration(FIC) index was calculated according to MIC value.RESULTS The MIC50 of cefoperazone-sulbactam was reduced significantly and the antimicrobial activities were reinforced remarkably when combined with minocycline or levofloxacin.The FIC results suggested that the main action be synergistic and additivie(53%,59% and 39%,37%),there was less autonomy(8%,4%) and no antagonism.CONCLUSIONS Combined with minocycline or levofloxacin respectively,cefoperazone-sulbactam expresses synergism and additivity against multidrug-resistant A.baumannii and there is no autonomy and antagonism.