Jian-ping Li

Hangzhou Normal University, Hang-hsien, Zhejiang Sheng, China

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Publications (12)3.86 Total impact

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    ABSTRACT: To investigate the serotypes distribution and ampicillin resistance of Haemophilus influenzae isolates from children with respiratory infection in Hangzhou. Haemophilus influenzae strains were identified with V factor and X factor tests. Serotypes were determined with the slide agglutination method. Nitrocefin test was used to detect beta-lactamase. The sensitivities of ampicillin to Haemophilus influenzae were determined with the Kirby-Bauer diffusion method and the E-test method. One hundred and fifty-two Haemophilus influenzae isolates, 108 from boys and 44 from girls, were identified between December 2006 and July 2007. Of the 152 isolates, 148 (97.4%) were untypable, only 4 (2.6%) were typable, including type a, type d, type e and type f (n=1 each type). Haemophilus influenzae type b and c strain was not found. Thirty-four isolates (22.4%) were beta-lactamase-positive. One hundred and thirteen isolates (74.3%) were susceptible to ampicillin, while 34 isolates (22.4%) were resistant to ampicillin. Untypable strains were the most common in Haemophilus influenzae isolates from children with respiratory infection in Hangzhou. The isolates of Haemophilus influenzae kept susceptibity to ampicillin to a certain extent.
    Zhongguo dang dai er ke za zhi = Chinese journal of contemporary pediatrics 04/2009; 11(3):217-20.
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    ABSTRACT: To evaluate the usefulness of a broad-range real-time PCR assay aimed at the 16S rRNA gene of bacteria in a clinical setting in rapid and reliable diagnosis of neonatal septicemia for improving the speed and accuracy of bacterial detection. The universal primer and TaqMan probe were designed based on the highly conserved sequences of the bacterial 16S rRNA gene. The chosen primers and probe did not show any likely cross hybridization with human, viral or fungal genome sequences. The TaqMan assay used the fluorescent signal on the probe, such as 6-carboxyfluorescin (6-FAM), and quenched by the standard 6-carboxytetramethylrhodamine (TAMRA) probes. The broad-range 16S rRNA gene real-time PCR array was established. Then, three common pathogenic microorganisms including Staphylococcus aureus, Staphylococcus epidermidis and Escherichia coli, which were prepared by a 10-fold dilution series respectively from 10(8) colony forming unit (CFU)/ml to 10(3) CFU/ml, as well as controls, were used for testing of both sensitivity and specificity of the real-time PCR assay. The blood samples from 830 cases of suspected septicemia, who were hospitalized in our neonatal ward and the neonatal intensive care unit (NICU) and developed clinical signs suggestive of infection, were tested with routine culture and bacterial 16S rRNA genes real-time PCR separately. In addition, 30 neonates without infection were enrolled as the negative control group. All the three common pathogenic bacterial species were positive on the 16S rRNA genes real-time PCR assay. There were no cross-reaction with cytomegalovirus (CMV), Epstein-Barr virus (EBV), hepatitis B virus (HBV), fungi, human DNA and blank control, and the technique showed high specificity and sensitivity. The detection limit of the TaqMan assay was tested by amplifying serial dilutions of the three common pathogenic bacterial DNA. The minimal detection limit of the TaqMan system was equivalent to 3 CFU of bacteria, the threshold cycle (CT), which is inversely proportional to the log of the amount of target DNA initially present, was 37.90 by calculation. The real-time PCR assay was evaluated on 830 blood specimens for suspected neonatal septicemia, as compared to the results obtained from the routine bacterial cultures. The positive rate by the real-time PCR assay was 5.18% (43/830) in 830 samples, and was significantly higher than that of blood culture [2.41% (20/830) (P < 0.01)]. The real-time PCR was positive in all the 20 positive blood culture samples. Thirty non-infectious blood samples were negative by both the PCR assay and blood cultures. When blood culture was used as control, the sensitivity of the real-time PCR assay was 100%, the specificity was 97.16%, and the index of accurate diagnosis was 0.972. Moreover, three of the PCR positive amplicons were confirmed by sequencing to confirm the accuracy of the real-time PCR assay in testing clinical specimens. The sequencing showed that except for one sequence, all the others were demonstrated to be Staphylococcus aureus and Escherichia coli respectively, which was in accord with the results of the blood cultures. The bacterial 16S rRNA genes real-time PCR had been established to diagnose the neonatal septicemia. The sensitivity and specificity the real-time PCR assay were higher than those of blood culture. This technique can provide a rapid way for the etiological diagnosis of neonatal septicemia, and was a convenient and accurate method in etiologic diagnosis of neonatal septicemia.
    Zhonghua er ke za zhi. Chinese journal of pediatrics 06/2007; 45(6):446-9.
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    ABSTRACT: To study the pathogenic bacteria of lower respiratory tract infection (LRTI), and age and gender distribution and drug resistance of the pathogenic bacteria in children. Sputum specimens for bacterial cultures were collected in sterile tubes from all of the children with LRTI who had been admitted to the Children's Hospital of Zhejiang University between August 2001 and July 2002. Antibiotic susceptibility tests were performed using the Vitek system, the Kirby-Bauer diffuse method and the Etest method after bacteria were identified. Among the 4,238 patients with LRTI during the study period, 1,181 patients were bacteria-positive, with a positive rate of 27.9%. Streptococcus pneumoniae (S. pneumoniae) was the most common (222 strains), followed by Haemophilus influenzae (H. influenzae) (216 strains), Klebsiella pneumoniae (K. pneumoniae) (216 strains), Escherichia coil (E. coli) (169 strains) and Staphylococcus aureus (S. aureus) (89 strains). The isolation rate of S. pneumoniae in females was significantly higher than in males (6.2% vs 4.7%; P < 0.05). However, the isolation rates of K. pneumoniae and S. aureus in males were higher than in females (5.1% vs 4.1% and 2.5% vs 1.5%, respectively; P < 0.05). A higher incidence of LRTI due to S. pneumoniae and H. influenzae was found in the 1-3 years group, while the incidence of LRTI due to K. pneumoniae, E. coli, S. aureus and E. cloacae was higher in patients under 1 year of age. Antibiotic susceptibility tests showed that rates of penicillin non-susceptible S. pneumoniae, ampicillin resistant H. influenzae, oxacillin-resistant S. aureus and ESBL-positive K. pneumoniae and E. coli were 55.0%, 16.5%, 41.2%, 42.6% and 4.5%, respectively. S. pneumoniae, H. influenzae, K. pneumoniae, E. coli and S. aureus were common pathogens of LRTI in children. The infection rate varied with age and gender. Antibiotics for treating LRTI should be selected based on the drug susceptibility test.
    Zhongguo dang dai er ke za zhi = Chinese journal of contemporary pediatrics 11/2006; 8(5):365-8.
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    ABSTRACT: To study the resistance of staphylococcus aureus (S. aureus) isolated from children in Hangzhou to antibiotics and analyze the clinical value of mecA-PCR in determining oxacillin-resistant isolates. S. aureus isolates were screened by using latex agglutination test and identified with GPI card of Vitek system. Antibiotics sensitivity tests were performed using disk diffusion methods and tests for sensitivity to oxacillin and vancomycin were performed with a further E-test method. The mecA gene was detected with polymerase-chain reaction (PCR). Of all 259 S. aureus strains, 185 from clinical specimens in inpatients and 74 from pharyngeal swabs in healthy children, 247 strains (95.8%) were beta-lactamase-positive and resistant to penicillin, while 91.1% of all strains were sensitive to oxacillin. All the strains were sensitive to vacomycin and 91.9% of all the strains were susceptible to cefotaxime and ceftriaxone. Resistance to erythromycin, tetracycline, clindamycin, trimethoprim-sulfamethoxazole, chloramphenicol, ofloxacin and rifampin were 48.3%, 30.9%, 21.6%, 11.2%, 10.0%, 2.3% and 1.5%, respectively. The resistance rate to oxacillin, cefotaxime, and ceftriaxone in clinical strains were significantly higher than that in carried strains (P < 0.05), while erythromycin-resistance rate was significantly higher in carried strains than that in clinical isolates (P < 0.05). The mecA-PCR showed that the control strain ATCC25923 and all oxacillin-sensitive S. aureus were mecA-negative, while all oxacillin-resistant strains were mecA-positive instead. Only one strain was mecA-positive in 7 oxacillin-intermediate S. aureus strains. Oxacillin-resistance in S. aureus isolates was low, and mecA-PCR method is a good choice for rapid examination oxacillin-resistant strains.
    Zhonghua er ke za zhi. Chinese journal of pediatrics 06/2006; 44(5):360-3.
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    ABSTRACT: In order to investigate the neuropathological effects on the developing rat brain after intrauterine infection, identification of glail fibrillary acidic protein (GFAP), 2', 3'-cyclic nucleotide phosphodiesterase (CNPase), and neurofilament (NF) was observed. Escherichia coli (E. coli) was inoculated into uterine horn of pregnant rats when gestation was 70% complete (15 days) and the control group was inoculated with normal saline. Immunohistochemistry was used for evaluation of GFAP, CNPase, and NF expression in pup brains at postnatal day 7 (P7) and reverse transcriptase-PCR (RT-PCR) to analyze macrophage inflammatory protein-1 alpha mRNA (MIP-1 alpha mRNA), macrophage inflammatory protein-1 beta mRNA (MIP-1beta mRNA), the regulated upon activation normal T expressed and secreted chemokine mRNA (RANTES mRNA) and Eotaxin mRNA expression in pup brains at P1, P3 and P7. The numbers of GFAP-positive cells of the E. coli-treated group pups were marked increased in periventricular white matter and hippocampus at P7 compared with the control group but no significant different levels of GFAP expression in corpus callosum were found between two groups. The integrate density (ID) of CNPase-positive staining of the Escherichia coli-treated group pups were marked decreased in periventricular white matter and corpus callosum at P7 compared with the control group. The ID of NF-positive staining of the Escherichia coli-treated group pups were marked decreased in periventricular white matter at P7 compared with the control group and no significant different levels of NF expression in corpus callosum were found between two groups. The expression of MIP-1 alpha mRNA and MIP-1 beta mRNA in brain of the E. coli-treated pup rat were higher than the control at P1, but the expression of MIP-1 alpha mRNA and MIP-1 beta mRNA in brain of the pup rat at P3 and P7 had no significant difference between two groups. The alteration of expression of GFAP, CNPase, and NF in the brain of neonatal rats after intrauterine infection suggested that intrauterine infection could cause neonatal white matter damage. Moreover, the transient increase in expression of chemokine such as MIP-1 alpha, MIP-1 beta in neonatal brain after intrauterine infection indicated that MIP-1 alpha, MIP-1 beta may be a mechanism mediating between the neonatal white matter damage and the intrauterine infection.
    Journal of Perinatal Medicine 02/2005; 33(5):415-22. · 1.95 Impact Factor
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    ABSTRACT: To investigate the serotypes and antibiotics-resistance patterns of Haemophilus influenzae isolated from children in Hangzhou. Isolates were identified with api-NH card. Serotypes were determined with slide agglutination method. The sensitivities of 13 antibiotics against 247 strains of Haemophilus influenzae were determined in vitro with Kirby-Bauer diffusion methods and MICs of ampicillin were determined with E-test. Nitrocefin test was used to detect beta-lactamase. Of the 247 strains isolated from children during the period from August 2001 to July 2002, 153 strains were non-typable, while 94 strains (38.1%) were typable and 90.4% and 1.1% of them belonged to type d and type b, respectively. Higher incidence of typable Haemophilus influenzae was found in male than in female children and the difference was significant (chi(2) = 5.30, P < 0.05), while between upper and lower respiratory tract infected children the difference was not statistically significant (chi(2) = 3.60, P > 0.05). Forty-one isolates (16.6%) were beta-lactamase-positive and 14 strains could not grow on medium in antibiotics sensitivity test. Of all 233 isolates tested successfully, 85.4% were susceptible to ampicillin, and the sensitivity rate to cefaclor, ceftriaxone, cefotaxime, imipenem, rifampin, clarithromycin, and chloramphenicol were as high as 98.7%, 99.6%, 99.6%, 99.6%, 98.7%, 91.0%, and 90.6%, respectively. All strains were sensitive to amoxicillin/clavulanic acid, ampicillin/sulbactan and ofloxacin, while 107 strains (45.9%) were resistant to trimethoprim-sulfamethoxazole, followed by that of tetracycline (14.6%). Resistance to ampicillin and trimethoprim-sulfamethoxazole in typable isolates was statistically significantly higher than in non-typable strains. Twenty-six strains (10.5%) were multi-resistant isolates and the multi-resistance rate in beta-lactamase-positive strains were significantly higher than that in beta-lactamase-negative strains (chi(c)(2) = 146.8, P < 0.001). Non-typable Haemophilus influenzae was the most common type in clinical strains isolated from children in Hangzhou, while type d was the overwhelming type and type b was uncommon in typable isolates. Incidence of typable isolates was higher in male than in female children, and it was apt to intergrow with other species of pathogenic bacteria. The proportion of beta-lactamase-positive strains was not high and ampicillin or other beta-lactam actibiotics were still the treatment of choice for infections with Haemophilus influenzae.
    Zhonghua er ke za zhi. Chinese journal of pediatrics 11/2004; 42(11):854-8.
  • Zhonghua liu xing bing xue za zhi = Zhonghua liuxingbingxue zazhi 11/2004; 25(10):915-6.
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    ABSTRACT: In order to investigate the neuropathological effects on the developing rat brain after intrauterine infection, identification of GFAP was observed. Escherichia coli (E. coli) was inoculated into uterine horn of pregnant rats when gestation was 70% complete (15 days) and the control group was inoculated with normal saline. Immunohistochemistry was used for evaluation of GFAP expression in pup brains at postnatal day 1 (P1), P3, P7, P14 and P21, and RT-PCR was used to analyze GFAP mRNA, interleukin-1beta, mRNA (IL-1beta mRNA) and tumor necrosis factor-alpha mRNA (TNF-alpha mRNA) expression in pup brains at P1, P3 and P7. At P1 and P3, GFAP was expressed very scarcely in periventricular white matter but not in other brain regions between the two groups. Compared with the control group, at P7 GFAP expression of the E. coli-treated pups was remarkably increased in periventricular white matter and hippocampus. The E. coli-treated pups at P14 showed a marked increase of GFAP expression in periventricular white matter, corpus callosum and cortex. However, no significant difference in levels of GFAP expression in any brain regions were found at P21 between the two groups. GFAP mRNA expression of the E. coli-treated pups was higher than the control at P1 and P3, but there was no significant difference between the two groups at P7. IL-1beta mRNA and TNF-alpha mRNA expressions of the E. coli-treated pups were higher than the control at P1 but there was no significant difference between the two groups at P3 and P7. These present results suggest that intrauterine infection could increase GFAP expression in the pup brain and indicate that intrauterine infection might damage the developing white matter and IL-1beta, TNF-alpha might be a mechanism mediating between the two events.
    Neuropathology 07/2004; 24(2):136-43. · 1.91 Impact Factor
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    ABSTRACT: To investigate the antibiotics-resistance type and molecular epidemiology of Streptococcus pneumoniae isolated from children in Hangzhou. The sensitivities of 323 strains of Streptococcus pneumoniae to 9 antibiotics were determined in vitro by Kirby-Bauer diffuse methods, and MICs of penicillin and cefotaxime were determined by E-test methods. Among all 323 strains isolated from children during the period from August 2001 to July 2002, 136 strains (42.1%) were sensitive to penicillin, while 57 strains (17.7%) were penicillin-resistant. Penicillin MICs ranged from 0.012 microg/ml to 4.0 microg/ml. All the strains were sensitive to cefotaxime and its MICs ranged from 0.012 microg/ml to 4.0 microg/ml. The most resistant antibiotic was erythromycin and it's resistant-rate was as high as 90.7%, followed by tetracycline (87.6%), trimethoprim-sulfamethoxazole (48.6%) and chloromycetin (14.9%). Totally 197 strains (61.0%) were multi-drug-resistant pneumococci and most of them were resistant to trimethoprim-sulfamethoxazole, erythromycin and tetracycline at the same time. Two strains (0.6%) were resistant to rifampin and none was resistant to vancomycin and ofloxacin. BOX PCR typing was carried out and no overwhelming fingerprinting pattern was found among penicillin resistant Streptococcus pneumoniae strains which were isolated from patients, while the banding patterns were always similar or identical among the strains isolated from the same specimen or from the same patient at different time, respectively. The antibiotics-resistant rate of pneumococci was high in Hangzhou, but the third-generation cephalosporins were still the best antibiotics against Streptococcus pneumoniae. One child could be infected or colonized by more than one pneumococci clone at the same or different time.
    Zhonghua er ke za zhi. Chinese journal of pediatrics 02/2004; 42(1):16-9.
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    ABSTRACT: To investigate the expression of glial fibrillary acidic protein (GFAP), GFAP mRNA and interleukin-1beta mRNA (IL-1beta mRNA), tumor necrosis factor-alpha mRNA (TNF-alpha mRNA) in neonatal rat brain after intrauterine infection. Escherichia coli (E. coli) was inoculated into both uterine horns of pregnant rats when gestation was 70% complete (15 days). The control group was treated with normal saline. The pups were killed on the postnatal day 1 (P1), P3 and P7, respectively. The cerebral white matter damage of the neonatal rats was determined by HE staining. Immunohistochemistry was used for evaluation of GFAP expression in neonatal rat brains and RT-PCR to analyze GFAP mRNA, IL-1beta mRNA and TNF-alpha mRNA expression at P1, P3 and P7. The major histopathological changes in neonatal cerebral white matter at P7 after intrauterine infections were: weak staining of cerebral white matter and focal rarefaction. GFAP-positive cells were observed in both the control and the E. coli-treated groups. The numbers of GFAP-positive cells of the E. coli-treated group pups were markedly increased in periventricular white matter and hippocampus at P7 compared with those of the control group (periventricular white matter: 9.73 +/- 3.55 vs 5.67 +/- 1.90, P < 0.05 and hippocampus: 7.81 +/- 3.61 vs 2.16 +/- 1.11, P < 0.05, respectively). No significantly different levels of GFAP expression in corpus callosum were found between two groups (P > 0.05). The expression of GFAP mRNA in brain of the E. coli-treated neonatal rat was higher than the control at P1, P3 (P1: 0.25 +/- 0.07 vs 0.15 +/- 0.08, P < 0.05 and P3: 0.50 +/- 0.09 vs 0.39 +/- 0.08, P < 0.05, respectively), but the expression of GFAP mRNA in brain of the neonatal rat at P7 had no significant difference between two groups (P > 0.05). The expression of IL-1beta mRNA and TNF-alpha mRNA in brain of the E. coli-treated neonatal rat were higher than of the control at P1 (IL-1beta mRNA: 0.83 +/- 0.19 vs 0.50 +/- 0.30, P < 0.05 and TNF-alpha mRNA: 0.74 +/- 0.30 vs 0.30 +/- 0.20, P < 0.05, respectively), but the expression of IL-1beta mRNA and TNF-alpha mRNA in brain of the neonatal rat at P3 and P7 had no significant difference between two groups (P > 0.05). The intrauterine infection could cause neonatal white matter damage and IL-1beta, TNF-alpha may be a mechanism mediating between the two events.
    Zhonghua er ke za zhi. Chinese journal of pediatrics 12/2003; 41(12):893-6.
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    ABSTRACT: OBJECTIVE: To examine the use of PCR utilizing 16S-23S rRNA gene spacer regions in the identification of bacteria. METHODS Primers used in PCR were designed by using the target sequences from the genes encoding 16S-23S rRNA spacer regions. PCR was used for the detection of different standard and clinical bacterial strains. RESULTS Characteristic DNA maps were present after using the PCR to identify 27 standard strains from 27 species. The maps could be directly used for classification of the tested bacterial strains or further differentiated by RFLP. The sensitivity of the PCR may be as high as 2.5 CFU/ml. No non-specific amplification products were observed when using DNA from human PBMC funguses or viruses as templates. Thirty-two strains of bacteria isolated from clinical strains showed DNA maps similar to the DNA maps amplified from standard strains. CONCLUSION The PCR detection of bacteria using 16S-23S rRNA gene spacer regions is sensitive, rapid, specific and accurate for identification of bacteria and provides a new rapid method for determining the clinical diagnosis and the etiology of sepsis.
    Zhejiang da xue xue bao. Yi xue ban = Journal of Zhejiang University. Medical sciences 09/2002; 31(6):448-452.
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    ABSTRACT: OBJECTIVE: To study the role of Haemophilus influenzae(Hi) in pneumonia and that of PCR in the diagnosis of Hi pneumonia. METHODS: Hi genus-specific PCR, Hib type-specific PCR and selective Hi culture media were used to detect 83 samples of deep nasal pharyngeal aspiration (NPA), 51 sera from 83 children younger than 3 years with pneumonia and 37 samples of pharyngeal swabs from healthy children. RESULTS: Of 83 NPA samples, 20(24.1 %) were positive by culture, 36 (43.4 %) positive by Hi-PCR and 19 (22.9 %) positive by Hib-PCR.Six out of 51 sera were positive by Hi-PCR and Hib-PCR, but none positive by culture. Of 37 pharyngeal swabs from healthy children, 3 ( 8.1 %) were positive by culture, 6 ( 16.2 %) positive by Hi-PCR and none positive by Hib-PCR. CONCLUSION: Hib-PCR is more appropriate for detecting NPA samples from children with pneumonia because of the high rate of non-typeable Hi carriers in healthy children.
    Zhejiang da xue xue bao. Yi xue ban = Journal of Zhejiang University. Medical sciences 03/2002; 31(1):47-50.