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Background. Kawasaki disease (KD) is a systemic vasculitis of unknown etiology in children. Coronary artery abnormalities are the most common complications of KD. Recent evidence showed that genetic polymorphisms may lead to susceptibility to KD. Genetic variants in platelet glycoprotein have been reported to be associated with coronary artery disease. The aim of the present study is to investigate the correlation between the role of platelet glycoprotein and coronary artery aneurysms in KD patients. Methods. We did a case-control study that enrolled 818 KD patients and 1401 healthy children with the same age and sex from January 2013 to December 2016. Analysis of single-nucleotide polymorphism (rs1126643) of the platelet glycoprotein Ia/IIa C807T was performed by multiplex polymerase chain reactions in this study. Results. A significant difference in the genotype distribution between KD cases and controls was observed for the glycoprotein Ia/IIa C807T (rs1126643) polymorphism ( p = 0.026 ). Compared with the healthy children, the rs1126643T allele carriers had odds ratio (OR) of 0.63 for developing KD (TT vs. CC: adjusted OR = 0.62, 95% confidence interval (CI) = 0.43–0.88, p = 0.0078 ; TT vs. CT/CC: adjusted OR = 0.63, 95% CI = 0.44–0.889, p = 0.0093 ). Furthermore, we also found that children less than 60 months of age and female patients with rs1126643 T allele carriers had an adjusted OR of 0.66 (95% CI = 0.46–0.95) for noncoronary artery aneurysm patients ( p = 0.0242 ). Single-nucleotide polymorphism rs1126643 TT seems to represent a protective factor against KD in coronary artery aneurysm formation in multivariate analysis. Conclusions. The platelet glycoprotein Ia/IIa T allele carriers may have a protective effect on the risk of coronary artery aneurysms of KD patients, especially in females and children aged less than 60 months. These results may provide evidence for platelet glycoprotein Ia/IIa gene polymorphisms in the pathogenesis of KD patients.

BackgroundKawasaki disease (KD) is a systemic vasculitis, and the formation of coronary artery lesions(CAL) is its most common sequela. Both genetic and environmental factors are considered to be important factors of in KD. Integrin α2 (ITGA2) is a transmembrane receptor that is associated with susceptibility to several diseases, but its relevance to KD with CAL is unclear.MethodsWe genotyped ITGA2 rs1126643 in 785 KD patients with the CAL and no-CAL(NCAL) (300 patients with CAL, and 485 age- and sex-matched patients with NCAL). OR (95% CI) and adjusted OR (95% CI) were used to evaluate the intensity of the association.ResultsWe found a significantly increased risk of KD with CAL associated with ITGA2 rs1126643 genotypes (CT vs CC: adjusted OR = 1.57, 95% CI = 1.16–2.12,P = 0.0032; CT/TT vs CC: adjusted OR = 1.49, 95% CI = 1.12–2.00,P = 0.0068; T vs C: adjusted OR = 1.66, 95% CI = 1.16–2.51,P = 0.0165). Moreover, we found that carriers of the CT/TT genotype had a significant risk of KD with coronary artery lesion susceptibility for children ≤60 months of age, and the CT/TT genotype was significantly associated with an increased risk of SCAL formation and MCAL formation when compared with the CC genotype.ConclusionITGA2 rs1126643 was associated with increased susceptibility and severity of CAL in KD.

Invasive group B Streptococcus (GBS) remains a leading cause of illness and death among infants globally. We conducted prospective and retrospective laboratory-based surveillance of GBS-positive cultures from infants

Background Invasive group B Streptococcus (GBS) disease in Chinese infants has gradually gained attention in recent years, but the molecular epidemiology of the pathogen is still not well known. Methods This multicenter study retrospectively investigated distribution of capsular serotypes, sequence types (STs), and hypervirulent GBS adhesin gene (hvgA) in clinical GBS isolates that caused invasive disease in infants aged Results From a total of 93 GBS isolates taken from 34 early-onset disease (EOD, 0–6 days after birth) and 59 late-onset disease (LOD, 7–89 days after birth) cases, four serotypes were identified: serotypes III (79.6%), Ib (12.9%), Ia (4.3%), and V (3.2%). Serotype III accounted for 73.5% of EOD and 83.1% of LOD and was responsible for 75.5% of cases involving meningitis. Fifteen STs were found, with the majority being ST17 (61.3%), ST12 (7.5%), ST19 (7.5%), and others (23.7%). 96.8% of STs belonged to only five clonal complexes (CCs): CC17 (64.5%), CC10 (12.9%), CC19 (9.7%), CC23 (6.5%), and CC1 (3.2%). The hvgA gene was detected in 66.7% of GBS isolates and 95% of CC17 isolates, all of which were serotype III except one serotype Ib/CC17 isolate. A large proportion of GBS isolates were found to be resistant to tetracycline (93.5%), clindamycin (65.5%), and erythromycin (60.2%). Genes of tetO (74.7%) and tetM (46.0%) were found in tetracycline resistant isolates, linB (24.6%) in clindamycin resistant isolates, and ermB (87.5%) and mefA (3.6%) in erythromycin resistant isolates. Conclusion Our results reveal higher prevalence of serotype III, ST17, CC17, hvgA expressing, and antibiotic resistant GBS isolates than previously reported in southern mainland China. This study provides guidance for appropriate measures of prevention and control to be taken in the future.

Purpose: Nontyphoidal Salmonella (NTS) is a common pathogen responsible for acute gastroenteritis among all ages; however, information on the prevalence, serotypes, and antibiotic susceptibility of NTS isolates is limited. We aimed to explore the characteristics of NTS isolated from paediatric patients in Guangzhou, China. Methods: This was a retrospective study of 4586 stool culture collected at Guangzhou Women and Children’s Medical Center from 2014 to 2016. Results: We identified 220 (4.80%) NTS isolates in stool samples. Fourteen serotypes were identified among the 220 NTS isolates. Salmonella serotype Typhimurium was the most common serotype, representing 69.09%. The highest rate of resistance was recorded in relation to AMP (76.61%), followed by SXT (29.95%), CTX (29.93%), CHL (29.77%), CAZ (23.20%), CIP (7.51%), and CFS (7.18%). The resistance rates of NTS and serotype Typhimurium to CAZ in 2015 were significantly higher than those in 2014. The average hospitalisation duration of inpatients infected by NTS resistant to three or more clinically important agents was significantly longer than that of patients infected with NTS with less antibiotic resistance. Conclusion: NTS represents a major cause of paediatric gastroenteritis in Guangzhou, China, and the high level of resistance to third-generation cephalosporins coupled with increasing resistance to quinolones among isolated NTS from paediatric gastroenteritis is a serious public health concern that requires continued monitoring and rational usage of antibiotics. Keywords: Salmonella, Gastroenteritis, Paediatric, Serotyping, Drug resistance

Kankan Gao,1 Xiaoshan Guan,1 Lanlan Zeng,1 Jiabi Qian,2 Sufei Zhu,1 Qiulian Deng,1 Huamin Zhong,1 Shuying Pang,1 Fei Gao,1 Jielin Wang,1 Yan Long,1 Chien-Yi Chang,3 Haiying Liu1 1Clinical Laboratory, Guangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou, China; 2Guangzhou Institute of Pediatrics, Guangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou, China; 3School of Chemistry and Biosciences, University of Bradford, Bradford, BD7 1DP, UK Background: A multidrug-resistant (MDR) RR2 gene cluster was identified by whole-genome sequencing in several highly virulent (ST-17) Group B streptococcus (GBS) isolates, which caused neonatal invasive infections in southern China in 2016. Tracing the transmission and distribution of MDR isolates in this area is important for the effective management of future infections. The aim of this study was to obtain longitudinal data of MDR isolates to monitor epidemiological trends of general common isolates in southern China, and provide evidence for future characterization of antimicrobial resistance mechanisms.Methods: Clinical information and antimicrobial susceptibility of GBS isolates were acquired from electronic information management system databases of the hospital under study between January 2011 and December 2017. To confirm the presence of intact RR2, the tetO, ant6, lnuB, and ant9 genes located upstream, midstream, and downstream of RR2 were detected by PCR and DNA sequencing.Results: A total of 149 cases of neonatal invasive GBS infection were identified during the period 2011–2017. Among them, 119 cases (79.9%) were caused by MDR isolates, with a general increasing trend over the past 7 years. Further characterization of 11 isolates showed that six isolates causing late-onset disease (LOD) carry the tetO, ant6, and lnuB genes, which are located on RR2. Moreover, lnuB and ant9 consistently co-occurred in GBS isolates, which suggests their close proximity to one another in the RR2 gene cluster.Conclusion: The MDR GBS is responsible for a large number of neonatal invasive infections and occurs with increasing frequency over time. Particularly, the MDR GBS isolates that cause LOD are more likely to carry the RR2 gene cluster, compared with those that cause early-onset disease. The rise in number of MDR GBS isolates emphasizes the pressing need for continuous surveillance to monitor their antibiotic susceptibility and epidemiology. Keywords: invasive infection, gene cluster, neonates, multidrug-resistance

Group B Streptococcus (GBS) is an important etiological agent of maternal and neonatal infections as well as postpartum women and individuals with impaired immunity. We developed and evaluated a rapid classification method for sequence types (STs) of GBS based on statistic models with Matrix-Assisted Laser Desorption/Ionization Time-of Flight Mass Spectrometry (MALDI-TOF/MS). Whole-cell lysates MALDI-TOF/MS analysis was performed on 235 well-characterized GBS isolates from neonatal invasive infections in a multi-center study in China between 2015 and 2017. Mass spectra belonging to major STs (ST10, ST12, ST17, ST19, ST23) were selected for model generation and validation. Recognition and cross validation values were calculated by Genetic Algorithm-K Nearest Neighbor (GA-KNN), Supervised Neural Network (SNN), QuickClassifier (QC) to select models with the best performance for validation of diagnostic efficiency. Informative peaks were further screened through peak statistical analysis, ST subtyping MSP peak data and mass spectrum visualization. For major STs, the ML models generated by GA-KNN algorithms attained highest cross validation values in comparison to SNN and QC algorithms. GA-KNN models of ST10, ST17, and ST12/ST19 had good diagnostic efficiency, with high sensitivity (95–100%), specificity (91.46%–99.23%), accuracy (92.79–99.29%), positive prediction value (PPV, 80%–92.68%), negative prediction value (NPV, 94.32%–99.23%). Peak markers were firstly identified for ST10 (m/z 6250, 3125, 6891) and ST17 strains (m/z 2956, 5912, 7735, 5218). Statistical models for rapid GBS ST subtyping using MALDI-TOF/MS spectrometry contributes to easier epidemical molecular monitoring of GBS infection diseases.

Fluoroquinolone (FQ)-resistant Group B Streptococcus (GBS) has been reported with considerable cross-resistance, worsening the crisis of multidrug-resistant (MDR) GBS in clinical settings. However, national epidemiological data on FQ-resistant GBS in mainland China have not been well-characterized. This study aimed to determine the prevalence of FQ resistance among GBS from neonatal invasive infections and maternal colonization in northern and southern China, to investigate the serotyping, multilocus sequence typing, and antibiotic cross-resistance, and to characterize the mutations in gyrA and parC genes in quinolone resistance-determining region (QRDR). In order to provide a comprehensive view of the location and structure of resistance genes, whole-genome sequencing on III/ST19 MDR isolates were performed. Among 426 GBS, 138 (32.4%) were FQ resistant, with higher prevalence in northern China than in southern China in both neonates (57.8%, 37/64 vs. 21.7%, 39/180) and pregnant women (50.9%, 29/57 vs. 26.4%, 33/125). Serotypes were distributed as III (48.5%), Ib (39.9%), V (6.5%), and Ia (5.1%). Sequence types were mainly ST19 (53.6%) and ST10 (39.1%), followed by ST12 (1.4%), ST17 (1.4%), ST23 (1.4%), and 0.7% each of ST27, ST188, ST197, and ST597. ST19 isolates were more prevalent in southern China than in northern China in both neonates (64.1%, 25/39 vs. 27.0%, 10/37) and pregnant women (81.8%, 27/33 vs. 41.4%, 12/29), whereas ST10 isolates were more common in northern China than in southern China in both neonates (64.9%, 24/37 vs. 20.5%, 8/39) and pregnant women (58.6%, 17/29 vs. 15.2%, 5/33). Serotype III isolates were mainly ST19 (89.6%, 60/67), while Ib isolates were largely ST10 (94.5%, 52/55). Sequencing data revealed several mutations in QRDR, including Ser81Leu in gyrA (99.2%, 130/131), Ser79Phe or Tyr in parC (76.2%, 48/63), and a previously unreported Ile218Thr and Ile219Phe double mutation pattern (49.2%, 31/63) in parC. ST10 isolates were associated with Ser79Phe (84%, 21/25), while ST19 isolates were limited to Ser79Tyr (95.7%, 22/23). A new integrative and conjugative element (ICE) harboring tetM and gyrA genes was identified in a III/ST19 isolate. This study investigates the molecular characteristics of FQ-resistant GBS in northern and southern China, emphasizing the need for continuous surveillance geographically and further research to characterize the mechanisms of ICE transfer.

Fluoroquinolone (FQ)-resistant Group B Streptococcus (GBS) has been reported with considerable cross-resistance, worsening the crisis of multidrug-resistant (MDR) GBS in clinical settings. However, national epidemiological data on FQ-resistant GBS in mainland China have not been well-characterized. This study aimed to determine the prevalence of FQ resistance among GBS from neonatal invasive infections and maternal colonization in northern and southern China, to investigate the serotyping, multilocus sequence typing, and antibiotic cross-resistance, and to characterize the mutations in gyrA and parC genes in quinolone resistance-determining region (QRDR). In order to provide a comprehensive view of the location and structure of resistance genes, whole-genome sequencing on III/ST19 MDR isolates were performed. Among 426 GBS, 138 (32.4%) were FQ resistant, with higher prevalence in northern China than in southern China in both neonates (57.8%, 37/64 vs. 21.7%, 39/180) and pregnant women (50.9%, 29/57 vs. 26.4%, 33/125). Serotypes were distributed as III (48.5%), Ib (39.9%), V (6.5%), and Ia (5.1%). Sequence types were mainly ST19 (53.6%) and ST10 (39.1%), followed by ST12 (1.4%), ST17 (1.4%), ST23 (1.4%), and 0.7% each of ST27, ST188, ST197, and ST597. ST19 isolates were more prevalent in southern China than in northern China in both neonates (64.1%, 25/39 vs. 27.0%, 10/37) and pregnant women (81.8%, 27/33 vs. 41.4%, 12/29), whereas ST10 isolates were more common in northern China than in southern China in both neonates (64.9%, 24/37 vs. 20.5%, 8/39) and pregnant women (58.6%, 17/29 vs. 15.2%, 5/33). Serotype III isolates were mainly ST19 (89.6%, 60/67), while Ib isolates were largely ST10 (94.5%, 52/55). Sequencing data revealed several mutations in QRDR, including Ser81Leu in gyrA (99.2%, 130/131), Ser79Phe or Tyr in parC (76.2%, 48/63), and a previously unreported Ile218Thr and Ile219Phe double mutation pattern (49.2%, 31/63) in parC. ST10 isolates were associated with Ser79Phe (84%, 21/25), while ST19 isolates were limited to Ser79Tyr (95.7%, 22/23). A new integrative and conjugative element (ICE) harboring tetM and gyrA genes was identified in a III/ST19 isolate. This study investigates the molecular characteristics of FQ-resistant GBS in northern and southern China, emphasizing the need for continuous surveillance geographically and further research to characterize the mechanisms of ICE transfer.

Objective To evaluate the application value of time to positivity (TTP) for blood culture combined with inflammatory parameters that included immature granulocyte percentage (IG%), immature granulocyte count (IG#), C‐reactive protein (CRP), white blood cells (WBC) neutrophil percentage (NE%), and neutrophil‐to‐lymphocyte ratio (NLR), and to identify bloodstream infections from contamination with coagulase‐negative staphylococci (CoNS) in pediatric patients. Methods Data of 12 897 inpatients with blood culture CoNS were retrospectively collected and analyzed from January‐December 2019 at our hospital. According to pre‐defined criteria, they were divided into a CoNS infection group (132 cases) and a CoNS contamination group (124 cases). Infection with Staphylococcus aureus (SA, 27 cases) at the same period was considered a positive control group. ROC curve analysis assisted in determining the value of applying TTP combined with the above‐mentioned inflammatory parameters to distinguish CoNS infection from contamination. Results Among the 256 strains of CoNS, Staphylococcus hominis (55.1%), Staphylococcus epidermidis (32.0%), and Staphylococcus capitis (7.0%) were common. There was no significant difference in the subspecies distribution between the infection and contamination groups. The TTP of the CoNS infection group was significantly lower than the contamination group (P, ROC analysis of TTP and inflammatory parameters in identifying CoNS bloodstream infection from contamination. A: ROC curve analysis of single indicators including IG%, IG#, CRP, NE%, and NLR. The AUC of TTP was the highest, reaching a value of 0.913, the AUC of IG% was 0.712, and the AUC of IG#, CRP, NE%, and NLR were 0.679, 0.645, 0.595, and 0.605, respectively, which were all evidently lower than 0.7. B: ROC curve analysis of TTP combined with inflammatory parameters. When TTP was combined with IG%, IG#, CRP, or NLR, the AUC was 0.951, 0946, 0.945, and 0.923, respectively. When TTP was combined with IG%, CRP, and NLR, the AUC was the highest, with a value of up to 0.977.

Background Group B Streptococcus (GBS) is a leading cause of morbidity and mortality in infants in both developed and developing countries. To our knowledge, only a few studies have been reported the clinical features, treatment and outcomes of the GBS disease in China. The severity of neonatal GBS disease in China remains unclear. Population-based surveillance in China is therefore required. Methods We retrospectively collected data of

In the Viable but Non-Culturable (VBNC) state, microorganisms may survive under severe external environment. In this study, the specificity and sensitivity of PMA-LAMP assay on the detection of Vibrio Parahemolyticus (V. parahemolyticus) has been developed and evaluated, with further application on a number of food-borne V. parahemolyticus strains. Six primers were designed for recognizing 8 distinct targeting on tlh, tdh and trh gene. Through specific penetration through the damaged cell membrane of dead cells and intercalating into DNA, PMA could prevent DNA amplification of dead bacteria from LAMP, which enabled the differentiation of bacteria between VBNC state and dead state. The established PMA-LAMP showed significant advantage in rapidity, sensitivity and specificity, compared with regular PCR assay. The applicability had also been verified, demonstrating the PMA-LAMP was capable of detection on V. parahaemolyticus.

The prevalent Staphylococcus aureus clones and antibiotic susceptibility profiles are known to change dynamically and geographically; however, recent S. aureus strains causing infections in women and children in China have not been characterized. In this study, we analyzed the molecular epidemiology and antimicrobial resistance of S. aureus isolated from patients in four centers for women and children in Guangzhou, China. In total, 131 S. aureus isolates (100 from children and 31 from women) were analyzed by spa typing, multi-locus sequence typing, virulence gene and antimicrobial resistance profiling, staphylococcal chromosomal cassette mec typing, and mutation analyses of rpoB. A total of 58 spa types, 27 sequence types (STs), and 10 clonal complexes (CCs) were identified. While CC59 (ST59-IV, 48.8%; ST338-III, 35.7%) and CC45 (ST45-IV, 100%) were the major clones (84.4%) among MRSA isolates, CC5 (ST188, 24.3%; ST1, 21.6%) and CC398 (ST398, 70%) were the major ones (70.1%) among MSSA isolates. ST338-MRSA-III mostly found in pus but hardly in respiratory tract samples while ST45-MRSA-IV was on the opposite, even though they both found in blood and cerebrospinal fluid sample frequently. Staphylococcal enterotoxin genes seb-seq-sek were strongly associated with ST59 and ST338, while sec was associated with ST45, ST121, ST22, and ST30. All ST338, ST1232, and SCCmec III isolates carried lukF/S-PV genes. A total of 80% of ST338 isolates were resistant to erythromycin, clindamycin, and tetracycline. All ST45 isolates exhibited intermediate or complete resistance to rifampicin. In total, 481 HIS/ASN mutations in rpoB were found in rifampicin-resistant or intermediate-resistant isolates. ST338-III and ST45-IV emerged as two of three major clones in MRSA isolates from women and children in Guangzhou, China, though ST59-MRSA-IV remained the most prevalent MRSA clone. Clonal distribution of S. aureus varied, depending on the specimen source. Virulence genes and antibiograms were closely associated with the clonal lineage. These results clarified the molecular epidemiology of S. aureus from women and children in Guangzhou, China, and provide critical information for the control and treatment of S. aureus infections.

Lei Pi,1,* Di Che,1,* Haifeng Long,2,3,* Zhenzhen Fang,4 Jiawen Li,1 Shuyi Lin,2 Yunfeng Liu,2 Meiai Li,2 Lijuan Bao,2 Wenli Li,2 Yuan Zhang,2 Qiulian Deng,2 Techang Liu,5 Li Zhang,5 Xiaoqiong Gu1,2 1Department of Clinical Biological Resource Bank, Guangzhou Institute of Pediatrics, Guangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou, China; 2Department of Clinical Laboratory, Guangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou, China; 3Department of Clinical Laboratory, The First Clinical Medical College, Jinan University, Guangzhou, China; 4Program of Molecular Medicine, Guangzhou Women and Children’s Hospital, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China; 5Department of Cardiology, Guangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou, China *These authors contributed equally to this work Introduction: Kawasaki disease is a kind of systemic vasculitis that mainly damages moderate and small-sized blood vessels, and is a leading cause of coronary artery lesions (CAL). Antiplatelet therapy is a routine component of Kawasaki disease treatment strategies. So it is important to evaluate the antiplatelet effect of aspirin because of the individual biological variability of antiplatelet effect of aspirin. The immature platelet fraction (IPF) has attracted particular attention as it may influence the antiplatelet effect of aspirin. This study investigated the prognostic factors for evaluating the degree of vasculitis and the effect of antiplatelet therapy in children with Kawasaki disease. Materials and methods: Blood samples were collected from 44 patients with Kawasaki disease before aspirin treatment and 7 to 10 days after treatment. The IPF counts, percentage of the IPF, and highly fluorescent IPF were detected by a Sysmex XE-5000 instrument. The levels of 11-dehydrothromboxane B2 (11-DH-TXB2), soluble CD40 ligand (sCD40L), and soluble P-selectin (sP-selectin) were measured by ELISA. The correlation between the measured factors and the degree of coronary artery damage in Kawasaki disease was analyzed. Results: We found that 11-DH-TXB2, sP-selectin, and sCD40L levels were much more elevated in the CAL group than in the non-coronary artery lesions (NCAL) group before aspirin treatment. The concentrations of 11-DH-TXB2, sCD40L, sP-selectin, and IPF were reduced after aspirin treatment in the NCAL group but not the CAL group. This is related to the degree of coronary artery damage in Kawasaki disease patients. Additionally, 11-DH-TXB2, sCD40L, sP-selectin, and IPF were positively correlated with the degree of coronary artery damage in Kawasaki disease patients. Conclusion: The current study suggests that the presence of high plasma concentrations of 11-DH-TXB2, sCD40L, sP-selectin, and IPF can be considered a risk factor and experimental biomarker for CAL in Kawasaki disease patients. Keywords: Kawasaki disease, coronary artery lesions, aspirin, immature platelet fraction

Kawasaki disease (KD) is a type of disease that includes the development of a fever that lasts at least 5 days and involves the clinical manifestation of multicellular vasculitis. KD has become one of the most common pediatric cardiovascular diseases. Previous studies have reported that miR-218 rs11134527 A>G is associated with susceptibility to various cancer risks. However, there is a lack of evidence regarding the relationship between this polymorphism and KD risk. The present study explored the correlation between the miR-218 rs11134527 A>G polymorphism and the risk of KD. We recruited 532 patients with KD and 623 controls to genotype the miR-218 rs11134527 A>G polymorphism with a TaqMan allelic discrimination assay. Our results illustrated that the miR-218 rs11134527 A>G polymorphism was not associated with KD risk. In an analysis stratified by age, sex, and coronary artery lesions, we found only that the risk of KD was significantly decreased for children older than 5 years (GG vs. AA/AG: adjusted OR = 0.26, 95% CI = 0.07–0.94, P=0.041). The present study demonstrated that the miR-218 rs1113452 A>G polymorphism may have an age-related relationship with KD susceptibility that has not previously been revealed.

Partial sequence diagram for seven house-keeping genes. adhP gene(A), pheS gene(B), atr gene(C), glnA gene(D), sdhA gene(E), glcK gene(F), tkt gene(G). (DOCX 849 kb)

Background Recognized as a resistance mechanism responsible for the emergence and prevalence of antimicrobial resistance, integron is widely distributed and spread among clinical microorganisms and play a key role in the dissemination of such antimicrobial resistance, which may eventually contribute to the unleashing of “Super Bugs” In this study, detection assays based on loop-mediated isothermal amplification (LAMP) methodologies targeting on class 1 to class 3 integrase genes was developed and evaluated. Methods LAMP methodology was employed to develop novel detection assays on class 1, 2 and 3 integrons. Firstly, this protocol was specifically designed to detect such integrons by targeting integrase genes intI1, intI2 and intI3. Development, evaluation and optimization of such LAMP assays was studied, including the reaction temperature, volumn, time, sensitivity and specificity of both primers and targets. A total of 1082 strains, including 397 integron positive and 685 integron negative microorganisms, were included for the application verification of the established LAMP assays. Results The indispensability of each primer was confirmed, and the optimal amplification was obtained under 63 °C for 45 min, with 25 μl reaction found to be the most cost-efficient volume. As application was concerned, all of the 397 integron-positive isolates yielded positive amplicons and other 685 integron-negative bacteria were negative for the integron-LAMP assays, revealing totaling 100% detection rate and specificity. Conclusions The established integron-LAMP assays was demonstrated to be a valid and rapid detection method for integrons screening, which may aid in both the laboratory and clinical integron screening for microorganisms.

Three kinds of PMQR determinants (qnr genes, aac(6')-Ib-cr, and qepA) have been discovered and shown to be widely distributed among clinical isolates. To characterize the prevalence of PMQR determinants in ESBL or AmpC-producing E. coli clinical isolates in Chinese children, a total of 292 ESBL or AmpC-producing E. coli clinical isolates collected from five children's hospitals in China from 2005 to 2006 were screened for PMQR determinants by PCR. Twenty (6.8%) of the 292 isolates were positive for PMQR determinants. A total of 12 (4.1%) isolates were positive for qnr genes, comprising three positive for qnrA (1.0%), three for qnrB (1.0%), and six for qnrS (2.1%). Twenty-four (8.2%) isolates were positive for aac(6')-Ib, of which 10 (3.4% of 292) had the -cr variant. There was no qepA gene detected in the isolates. Conjugation revealed that qnr, aac(6')-Ib-cr, and ESBL-encoding genes were transferred together.

On the basis of the single-input single-output (SISO) RBF-ARX model proposed in previous works [Peng, H., et al. (2003b). Stability analysis of the RBF-ARX model based nonlinear predictive control. In Proceedings of the ECC2003; Peng, H., et al. (2003c). Modeling and control of nonlinear nitrogen oxide decomposition process. In Proceedings of the CDC’03; Peng, H., et al. (2004). RBF-ARX model based nonlinear system modeling and predictive control with application to a NOx decomposition process. Control Engineering Practice, 12, 191–203; Peng, H., et al. (2007). Nonlinear predictive control using neural nets-based local linearization ARX model—Stability and industrial application. IEEE Transactions on Control Systems Technology, 15, 130–143] the multi-input multi-output (MIMO) RBF-ARX model and its state-space representation are derived to describe the dynamics of a class of multivariable nonlinear systems whose working-point varies with time and which may be linearized around the working-point. The proposed MIMO RBF-ARX model has a basic regression-model structure that is analogous to the linear ARX model structure, and the elements of its regression matrices are composed of Gaussian radial basis function (RBF) neural networks that are dependent on the working-point state of the current system. An off-line estimation approach to parameters and orders of the MIMO RBF-ARX model is presented, and, on the basis of the estimated MIMO RBF-ARX model, a predictive control strategy is designed to control the underlying nonlinear system. A case study on a simulator of a thermal power plant is also given to illustrate the effectiveness of the nonlinear modeling and control method proposed in this paper.

The purpose of this study was to investigate the prevalence of plasmid-mediated AmpC beta-lactamases in Escherichia coli and Klebsiella pneumoniae from five children's hospitals in China. A total of 494 E. coli and 637 K. pneumoniae isolates were collected from five children's hospitals in China from 2005 to 2006. The isolates with decreased susceptibility to cefoxitin were subjected to confirmation test with 3-aminophenyl boronic acid. Polymerase chain reaction (PCR) amplification of the blaAmpC, blaTEM, blaCTXM, and blaSHV genes and their gene sequencing were performed. Transconjugants were achieved by conjugation experiments. Plasmid-mediated AmpC beta-lactamases were found in 10.1% of K. pneumoniae (64/637) and in 2.0% of E. coli (10/494) strains. The proportion of plasmid-mediated AmpC-producing strains significantly increased from 2005 (2.6%) to 2006 (9.3%) (p0.001). The DHA-1-producing isolates were the most prevalent type (93.2%, 69/74). The sequences of blaDHA-1 genes were all identical to those from the GenBank. Strains of blaCMY-2 were isolated from five isolates (6.8%), which were all from E. coli. One sequence of blaCMY-2 differs from blaCMY-2 in the GenBank. Eighteen of the 74 (24.3%) AmpC-producing K. pneumoniae and E. coli isolates coproduced an extended-spectrum beta-lactamase (ESBL). Cefoxitin resistance was transferred to 15 of the 74 positive strains (20.3%). Our study has demonstrated the occurrence of plasmid-mediated AmpC beta-lactamases in E. coli and K. pneumoniae in Chinese pediatric patients and DHA-1 type AmpC enzymes had the highest prevalent rate. The CMY-2 AmpC beta-lactamases from the children's hospitals in China in this study are the first reported. Hence, continuous surveillance of the prevalence and evolution of AmpC beta-lactamase is important.

The plasmid-mediated quinolone resistance qnr genes in clinical isolates in adults have been described in different countries; however, the frequency of their occurrence has not been detected in pediatric patients. A total of 410 clinical isolates of Klebsiella pneumoniae, identified as producers of an extended-spectrum beta-lactamase (ESBL), or AmpC beta-lactamase, were collected from five children's hospitals in China during 2005-2006. The isolates were screened for the presence of the qnrA, qnrB, and qnrS genes, and then the harboring qnr gene isolates were detected for a bla gene coding for the TEM, SHV, CTX-M, and plasmid-mediated ampC gene by a PCR experiment. Ninety-two isolates (22.7%) were positive for the qnr gene, including 10 of qnrA (2.4%), 25 of qnrB (6.1%), and 62 of qnrS (15.1%). Eighty-one of the 92 (88.0%) qnr-positive isolates carried at least one bla gene for TEM, SHV, CTX-M, or DHA-1. The ciprofloxacin resistance increased 16-256-fold and oflaxacin resistance increased 2-32-fold in transconjugants, respectively. These results indicated that the plasmid-mediated qnr quinolone resistance gene was qnrS, followed by qnrB and qnrA. Most of the isolates also carried a bla gene coding ESBL or ampC gene coding DHA-1 among Klebsiella pneumoniae isolated from Chinese pediatric patients.

In order to demonstrate antibiotic usage in Chinese Pediatric Intensive Care Units (PICUs), and provide some data to further study on relation of antimicrobial use and resistance. We reviewed the use of antibiotics in Chinese PICUs from 2002 to 2006. All data, including general data and antibiotic use data, were obtained from five PICUs of pediatric teaching hospitals in China. The results of antibiotic use were expressed as defined daily doses (DDDs) per 100 patient-days by WHO in this study. All 12 743 patients were included from the five PICUs in this study. Length of stay in PICUs was essentially unchanged, but the percentage of antimicrobial costs vs. drug costs dramatically decreased in this study period (p < 0.01). The percentage of empiric treatment decreased by year (from 82.2% to 70.2%). while the percentage of therapeutic treatment increased by year (from 11.2% to 24.2%) from 2002 to 2006. Total antibiotic usage decreased from 72.1 DDDs per 100 patient-days to 35.5 DDDs per 100 patient-days from 2002 to 2006 (p < 0.05). The significant increase was found in the DDDs per 100 patient-days of second generation cephalosporins in this study (p < 0.05). While usages of the some antibiotics decreased, for example penicillins, third-generation cephalosporins, and macrolides in this study period. The data of antimicrobial use were obtained from five PICUs of biggest pediatric teaching hospital in this 5-year period, which could serve as a basis of antibiotic treatment and a benchmark in future study of antibiotic use.

To investigate the pathogenic bacteria distribution and drug susceptibility in children with acute otitis media (AOM) in different age and different season in the Pearl River Delta region.Four hundred and forty-two children diagnosed as AOM were divided into three groups by age factor and four groups by season factor. Midge ear pus collecting and culturing were used for bacteria and antimicrobial susceptibility test.(1) Strains of bacteria were isolated from 356 children with the positive rate of 80.5%. Streptococcus pneumoniae (39.2%), staphylococcus aureus (25.9%) and haemophilus influenzae (7.4%) were the most frequently isolated pathogens. (2) Streptococcus pneumoniae was the main pathogenic bacteria in 0-1 year group and1-3 years group (P0.05), staphylococcus aureus was the most frequently isolated pathogen in3 years group (P0.05); (3) In the season groups, the number of children with AOM decreased significantly in July-September group. There was no significant difference of streptococcus pneumoniae distribution among the four groups (P0.05). Staphylococcus aureus was the main pathogen in January-March group (P0.05); (4) Drug sensitivity shown that linezolid and ofloxacin were most sensitive to streptococcus pneumoniae and staphylococcus aureus, and macrolides had a good therapy effect to haemophilus influenzae.The pathogens distribution and drug susceptibility in children with AOM were varies in different age and different season. As a result, a treatment should be done based on the climate, environment, and pathogens distribution of a region.

SUMMARYThe characteristics of 359 group A streptococcal (GAS) isolates collected from Chinese paediatric patients in two periods (1993–1994, 2005–2006) were studied. Isolates were assigned toemmtypes and assayed for eight superantigen (SAg) genes (speA,speC,speH,speI,speG,speJ,ssa,SMEZ). Typesemm1 andemm12 were consistently the most prevalent during the two periods, while others varied in frequency. GAS isolates carrying six or more SAg genes increased from 46·53% (1993–1994) to 78·39% (2005–2006);ssa,speHandspeJgenes (PspeAdeclined (Pemmtype, but strains of the sameemmtype sometimes carried different SAg genes in the two periods. No significant difference inemm-type distribution and SAg gene profile was noted between isolates from different diseases. These data may contribute towards the development of a GAS vaccine in China.

The plasmid-mediated quinolone resistance qnr genes in clinical isolates in adults have been described in different countries; however, the frequency of their occurrence has not been detected in pediatric patients. A total of 410 clinical isolates of Klebsiella pneumoniae, identified as producers of an extended-spectrum beta-lactamase (ESBL), or AmpC beta-lactamase, were collected from five children's hospitals in China during 2005-2006. The isolates were screened for the presence of the qnrA, qnrB, and qnrS genes, and then the harboring qnr gene isolates were detected for a bla gene coding for the TEM, SHV, CTX-M, and plasmid-mediated ampC gene by a PCR experiment. Ninety-two isolates (22.7%) were positive for the qnr gene, including 10 of qnrA (2.4%), 25 of qnrB (6.1%), and 62 of qnrS (15.1%). Eighty-one of the 92 (88.0%) qnr-positive isolates carried at least one bla gene for TEM, SHV, CTX-M, or DHA-1. The ciprofloxacin resistance increased 16-256-fold and oflaxacin resistance increased 2-32-fold in transconjugants, respectively. These results indicated that the plasmid-mediated qnr quinolone resistance gene was qnrS, followed by qnrB and qnrA. Most of the isolates also carried a bla gene coding ESBL or ampC gene coding DHA-1 among Klebsiella pneumoniae isolated from Chinese pediatric patients.

Background Quinolone resistance in Enterobacteriaceae results mainly from mutations in type II DNA topoisomerase genes and/or changes in the expression of outer membrane and efflux pumps. Several recent studies have indicated that plasmid-mediated resistance mechanisms also play a significant role in fluoroquinolone resistance, and its prevalence is increasing worldwide. In China, the presence of the qnr gene in the clinical isolates of Enterobacteriaceae has been reported, but this transmissible quinolone resistance gene has not been detected in strains isolated singly from pediatric patients. Because quinolones associated with a variety of adverse side effects on children, they are not authorized for pediatric use. This study therefore aimed to investigate the presence of the qnr gene in clinical isolates of E. coli and K. pneumoniae from pediatric patients in China. Methods A total 213 of non-repetitive clinical isolates resistant to ciprofloxacin from E. coli and K. pneumoniae were collected from hospitalized patients at five children's hospital in Beijing, Shanghai, Guangzhou, and Chongqing. The isolates were screened for the plasmid-mediated quinolone resistance genes of qnrA, qnrB, and qnrS by PCR. Transferability was examined by conjugation with the sodium azide-resistant E. coli J53. All qnr-positive were analyzed for clonality by enterobacterial repetitive intergenic consensus (ERIC)-PCR. Results The study found that 19 ciprofloxacin-resistant clinical isolates of E. coli and K. pneumoniae were positive for the qnr gene, and most of the qnr positive strains were ESBL producers. Conjugation experiments showed that quinolone resitance could be transferred to recipients. Apart from this, different DNA banding patterns were obtained by ERIC-PCR from positive strains, which means that most of them were not clonally related. Conclusion This report on transferable fluoroquinolone resistance due to the qnr gene among E. coli and K. pneumoniae strains indicated that plasmid-mediated quinolone resistance has emerged in pediatric patients in China.

To investigate the nasal carriage of antibiotic-resistant pneumococci in children of5 years old in the following four cities, Beijing, Shanghai, Guangzhou and Xi'an.A total of 647 pneumococci strains were isolated and detected. Minimal inhibition concentrations (MICs) of antibiotics were determined by E-test. Disk diffusion test was used for the measurement of antimicrobial susceptibility.Prevalence of penicillin non-susceptible Streptococcus pneumoniae in the four cities was 41%, with Guangzhou (60.8%) ranking first, followed by Xi'an (45%), Shanghai (37%) and Beijing (25.9%). The majority of penicillin non-susceptibility isolates (23.9% - 53.8%) had a low level of resistance (MIC 0.64 - 1.5 microg/ml). The most sensitive antimicrobials in terms of percentage of susceptible organisms were amoxicillin-clavulanic acid (99.4%), followed by ceftriaxone (92.1%); cefurxime and cefaclor were slightly more sensitive than penicillin with susceptibility of 74.8% and 77.9%. Erythromycin, tetracycline and TMP-SMZ were highly resistant (83.6%, 82.1% and 76.2% respectively). Among erythromycin resistant isolates, 100% were resistant to azithromycin, 98.6% to clarithromycin, 97.2% to roxithromycin and spiramycin, and 96.6% to clindamycin. 97.2% (141/145) were typical of the macrolides-lincosamides-streptogramons B (MLSB) resistance phenotype, and 2.8% (4/145) were M phenotype. The group of PRSP was with significantly higher rates of non-susceptibility for ceftriaxone (18.4%), cefurxime (58.6%), cefaclor (53.4%), compared with the group of PEN-S (0.5%, 1.8% and 0.2%, respectively) and the rate of multi-drug resistance in the isolates of PRSP group (92.9%) was significantly higher than that of PEN-S group (59.2%).The rates of penicillin and multi-drug resistance among isolates of pneumococci carried nasally in are high children and the high prevalence of multi-drug resistance in the Chinese population may be becoming one of the most serious problems in this century.