Molecular characterisation of Staphylococcus aureus in school-age children in Guangzhou: associations among agr types, virulence genes, sequence types, and antibiotic resistant phenotypes.

BACKGROUND: Staphylococcus aureus, one of the most prevalent opportunistic pathogens, mainly colonizes the nasal cavity and is a risk factor for severe infections. Virulence factors and accessory gene regulator (agr) are key to the severity and diversity of staphylococcal infection. In this study, we aimed to characterise S. aureus agr-types and virulence genes and correlated them with genetic background and antibiotic-resistant phenotypes. RESULTS: Agr types were identified in 704 isolates (98.5%), with only 11 isolates were negative for agr type. Most of our isolates were classified as agr type I, followed by types III, II and IV. The enterotoxin c gene (sec) was detected in 48.6% of isolates, showing the highest prevalence among the five enterotoxin genes detected. The positivity rates for the lukS/F-PV and tsst genes were 4% and 2.2%, respectively, while neither sed nor SasX were detected. ST45, ST59, ST338, ST188, ST6, ST7, ST22, ST25, ST398, and ST944 belonged to agr I group, while ST5 and ST15 belonged to agr II group. ST30 and ST1 were classified into agr III group, and ST121 was assigned into agr IV group. The tsst gene was found exclusively within agr I and III types belonging to ST7 and ST30 isolates, while the lukS/F-PV was predominantly carried by agr I type isolates primarily within CC59 and CC22 clones. Among the methicillin-resistant S. aureus (MRSA) isolates, 89.7% belonged to agr I group, and 97.8% of rifampicin-resistant or intermediate isolates were assigned to agr I group. MRSA isolates harboured more tested virulence genes compared to methicillin-susceptible S. aureus isolates. CONCLUSIONS: We characterized the distributions of agr types and eight major virulence genes of 715 S. aureus isolates, and our findings revealed clear associations between agr types and STs, as well as virulence genes, and drug resistant phenotypes.

Molecular Characterization and Epidemiology of Carbapenem-Resistant Enterobacteriaceae Isolated from Pediatric Patients in Guangzhou, Southern China.

Background: Carbapenem-resistant Enterobacteriaceae (CRE) are spreading worldwide, posing a serious public health concern. However, the data on CRE strains that cause infections in children in Guangzhou remain limited. Therefore, this study aimed to investigate the epidemiology of CRE, drug resistance, and resistance mechanisms in children in Guangzhou, Southern China. Methods: In total, 54 nonrepetitive CRE strains were collected in pediatric patients at three centers in Guangzhou, Southern China, from January 2016 to August 2018. CRE isolates were used for further studies on antimicrobial susceptibility, the modified Hodge test (MHT), the modified carbapenem inactivation method (mCIM), and drug resistance genes. Multilocus sequence typing (MLST) was used to elucidate the molecular epidemiology of K. pneumoniae. Results: The isolated CRE strains include 34 K. pneumoniae (63.0%), 10 E. coli (18.5%), 4 Enterobacter cloacae (7.4%), and 6 Proteus mirabilis (11.1%) strains. The strains were isolated mainly from the blood (31.5%, n = 17), sputum (31.5%, n = 17), and urine (16.7%, n = 9). All CRE isolates were highly resistant to the third- or fourth-generation cephalosporins, carbapenems, and ß-lactam + ß-lactamase inhibitors (94.4%-96.3%). In addition, the resistance rates to amikacin, ciprofloxacin, levofloxacin, tigecycline, and colistin were 5.6%, 14.8%, 16.7%, 9.3%, and 0%, respectively. Carbapenemase was detected in 35 (64.8%) of the CRE isolates. The most dominant carbapenemase gene was bla NDM (n = 17, 48.6%), followed by bla IMP (n = 13, 37.1%) and bla OXA-23 (n = 4, 11.4%). Other carbapenemase genes (bla KPC, bla sim, bla Aim, bla GES, bla Gim, bla OXA-2 , and bla OXA-48 ) and the mcr-1 gene were not detected. MLST analysis showed high diversity among the K. pneumoniae, and ST45 (11.7%, 4/34) was the dominant sequence type. Conclusion: This study revealed bla NDM was the most dominant carbapenemase gene in children in Guangzhou. Infection control measures need to be taken for the prevention and treatment of CRE infections.

Bioprocessing and integration of a high flux screening systematic platform based on isothermal amplification for the detection on 8 common pathogens.

During a large variety of common pathogens, E. coli, P. aeruginosa, MRSA, MRCNS, V. parahaemolyticus, L. monocytogenes and Salmonella are the leading pathogens responsible for large number of human infections and diseases. In this study, a high flux screening based on nucleic acid isothermal amplification technique has been developed. For the 8 common pathogens, species-specific targets had been selected and analyzed for their unique specificity. After optimization, separate LAMP reaction assays had been bioprocessed and integrated into one systematic detection platform, including 8 strips (PCR tubes) and 96-well plates. Eight standard strains verified for the accuracy. Application of the established high flux screening platform was used for detection for 48 samples in 4 different 96-well plates, with 2 groups of 2 operators using double-blind procedure. The accuracy of 100% was obtained, with the total time consumption as 66-75 min (for 12 samples detection on 8 different pathogens). As concluded, through the bioprocess of the systematic platform based on LAMP technique, it's been demonstrated to be capable of simultaneous detection of 8 pathogens, with high sensitivity, specificity, rapidity and convenience.

Evaluation of time to positivity for blood culture combined with immature granulocytes, neutrophil-to-lymphocyte ratio, and CRP in identifying bloodstream coagulase-negative Staphylococci infection in pediatric patients.

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 < .05). IG%, IG#, CRP, NE%, and NLR were all higher in the infected group as compared to the contaminated group (P < .05), while WBC was similar among groups. There was also no statistical difference in those parameters when comparing the CoNS infection and SA groups. ROC analysis showed that TTP value in identifying CoNS infection from contamination was the highest with area under the curve (AUC) of 0.913, and the sensitivity and specificity were 0.827 and 0.852, respectively, at the optimal cutoff value of 23.9 hours. This was followed by IG% (AUC = 0.712), with an optimal critical value of 0.55%, and a sensitivity of 0.519 and specificity of 0.797. All the AUC values of IG#, CRP, NE%, and NLR were <0.7. A combination of TTP with IG%, CRP, and NLR improved the AUC, sensitivity, specificity, accuracy, PPV, and NPV values to 0.977, 0.922, 0.957, 91.8%, 92.2%, and 91.3%, respectively. CONCLUSIONS: TTP within 24 hours indicates likelihood of CoNS as the pathogenic agent in pediatric patient blood culture. The combination of TTP with IG% CRP and NLR might improve the diagnostic accuracy.

Molecular characteristics of group B Streptococcus isolates from infants in southern mainland China.

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 < 3 months of age in southern mainland China between January 2013 and June 2016. Genes for antibiotic resistance to tetracycline, erythromycin, and clindamycin were also examined. 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.

Virulence and resistance on various pathogens mediated by mobile genetic integrons via high flux assays.

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.

RETRACTED: Prevalence of GBS serotype III and identification of a ST 17-like genotype from neonates with invasive diseases in Guangzhou, China.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request of the authors. There is significant discrepancy in the genomic characteristics between the currently and previously dominant GBS (2018) and previously dominant GBS (2013-2014). The dramatically rapid and unexpected evolution of GBS strains has led to the significant discrepancy from recent findings which makes all the authors strongly concerned that this will influence the accuracy and validity of GBS treatment and therapy if based on the current manuscript. For example, the genomic difference between the currently prevalent type (II and III) and previously prevalent type (III) is considerably diverse, for which the pathogenic and virulent characteristics of the strains are very different. As all authors have a strong sense of responsibility and expertise in clinical microbiology, agreed by all authors, on behalf of all authors of this manuscript, the authors consequently request for article withdrawal for this manuscript.

Epidemiology of invasive group B streptococcal disease in infants from urban area of South China, 2011-2014.

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 <3 months old infants with culture-positive GBS in sterile samples from three large urban tertiary hospitals in South China from Jan 2011 to Dec 2014. The GBS isolates and their antibiotic susceptibility were routinely identified in clinical laboratories in participating hospitals. Serotyping and multi-locus sequence typing (MLST) were also conducted for further analysis of the neonatal GBS disease. RESULTS: Total 70 cases of culture-confirmed invasive GBS infection were identified from 127,206 live births born in studying hospitals, giving an overall incidence of 0.55 per 1000 live births (95% confidence interval [CI] 0.44-0.69). They consisted of 49 with early-onset disease (EOD, 0.39 per 1000 live births (95% CI 0.29-0.51)) and 21 with late-onset disease (LOD, 0.17 per 1000 live births (95% CI 0.11-0.25)). The incidence of EOD increased significantly over the studying period. Five infants (4 EOD and 1 LOD) died before discharge giving a mortality rate of 7.1% and five infants (7.1%, 2 EOD and 3 LOD) had neurological sequelae. Within 68 GBS isolates from GBS cases who born in the studying hospitals or elsewhere, serotype III accounted for 77.9%, followed by Ib (14.7%), V (4.4%), and Ia (2.9%). MLST analysis revealed the presence of 13 different sequence types among the 68 GBS isolates and ST-17 was the most frequent sequence type (63.2%). All isolates were susceptible to penicillin, ceftriaxone, vancomycin and linezolid, while 57.4% and 51.5% were resistant to erythromycin and clindamycin, respectively. CONCLUSIONS: This study gains the insight into the spectrum of GBS infection in south China which will facilitate the development of the guidance for reasonable antibiotics usage and will provide evidence for the implementation of potential GBS vaccines in the future.

Virulence of thermolable haemolysi tlh, gastroenteritis related pathogenicity tdh and trh of the pathogens Vibrio Parahemolyticus in Viable but Non-Culturable (VBNC) state.

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.

Automated classification of group B Streptococcus into different clonal complexes using MALDI-TOF mass spectrometry.

Objectives: To evaluate the performance of Matrix-Assisted Laser Desorption/Ionization Time-of Flight Mass Spectra (MALDI-TOF MS) for automated classification of GBS (Group B Streptococcus) into five major CCs (clonal complexes) during routine GBS identification. Methods: MALDI-TOF MS of 167 GBS strains belonging to five major CCs (CC10, CC12, CC17, CC19, CC23) were grouped into a reference set (n = 67) and a validation set (n = 100) for the creation and evaluation with GBS CCs subtyping main spectrum (MSP) and MSP-M using MALDI BioTyper and ClinProTools. GBS CCs subtyping MSPs-M was generated by resetting the discriminative peaks of GBS CCs subtyping MSP according to the informative peaks from the optimal classification model of five major CCs and the contribution of each peak to the model created by ClinProTools. Results: The PPV for the GBS CCs subtyping MSP-M was greater than the subtyping MSP for CC10 (99.21% vs. 93.65%), but similar for CC12 (79.55% vs. 81.06%), CC17 (93.55% vs. 94.09%), and CC19 (92.59% vs. 95.37%), and lower for CC23 (66.67% vs. 83.33%). Conclusion: MALDI-TOF MS could be a promising tool for the automated categorization of GBS into 5 CCs by both CCs subtyping MSP and MSP-M, GBS CCs subtyping MSP-M is preferred for the accurate prediction of CCs with highly discriminative peaks.

Molecular characteristics and evaluation of the phenotypic detection of carbapenemases among Enterobacterales and Pseudomonas via whole genome sequencing.

Background/purposes: The continuously increasing carbapenem resistance within Enterobacterales and Pseudomonas poses a threat to public health, nevertheless, the molecular characteristics of which in southern China still remain limited. And carbapenemase identification is a key factor in effective early therapy of carbapenem-resistant bacteria infections. We aimed to determine the molecular characteristics of these pathogens and compare commercial combined disc tests (CDTs) with the modified carbapenem inactivation method (mCIM) and EDTA-CIM (eCIM) in detecting and distinguishing carbapenemases using whole genome sequencing (WGS). Methods: A total of 78 Enterobacterales, 30 Pseudomonas were obtained from two tertiary hospitals in southern China. Susceptibility tests were conducted using an automated VITEK2 compact system with confirmation via the Kirby-Bauer method. The WGS was conducted on all clinical isolates and the molecular characteristics were analyzed by screening the whole genome sequences. CDTs with or without cloxacillin, mCIM, and eCIM, were performed and compared by taking WGS results as the benchmark. Results: A total of 103 carbapenem non-susceptible and 5 carbapenem susceptible bacteria were determined, with Klebsiella pneumoniae (42.7%), Pseudomonas aeruginosa (23.3%) and Escherichia coli (18.4%) being most prevalent. Carbapenemase genes were detected in 58 (56.3%) of the 103 carbapenem-non-susceptible clinical isolates, including 46 NDM, 6 KPC, 3 IMP, 1 IPM+VIM,1NDM+KPC, and 1 OXA-181. Carbapenemase-producing isolates were detected more frequently in Enterobacterales (76.3%). Among K. pneumoniae, the major sequence types were st307 and st11, while among E. coli and P. aeruginosa, the most prevalent ones were st410 and st242 respectively. For carbapenemase detection in Enterobacterales, the mCIM method achieved 100.00% (95% CI, 92.13-100.00%) sensitivity and 94.44% (70.63-99.71%) specificity (kappa, 0.96); for Pseudomonas, detection sensitivity was 100% (5.46-100.00%), and 100% (84.50-100.00%) specificity (kappa, 0.65). Commercial CDT carbapenemase detection sensitivity for Enterobacterales was 96.49% (86.84-99.39%), and 95.24% (74.13-99.75%) specificity (kappa, 0.90); for Pseudomonas, carbapenemase detection sensitivity was 100.00% (5.46-100.00%) and 37.93% (21.30-57.64%) specificity (kappa, 0.04). When cloxacillin testing was added, CDT specificity reached 84.61% (64.27-94.95%). Conclusion: The molecular epidemiology of carbapenem-non-susceptible isolates from pediatric patients in Southern China exhibited distinctive characteristics. Both the mCIM-eCIM combination and CDT methods effectively detected and differentiated carbapenemases among Enterobacterales isolates, and the former performed better than CDT among Pseudomonas.

Prevalence, serotype, and antimicrobial resistance profiles of children infected with Salmonella in Guangzhou, southern China, 2016-2021.

Purpose: Salmonella infection is a key global public health concern and has lead to an increased economic burden on society. We investigated the epidemiological characteristics and antimicrobial resistance profiles of clinically isolated Salmonella strains in Guangzhou Women and Children's Medical Center. Patients and methods: This was a retrospective study of 1,338 Salmonella strains collected from children in Guangzhou Women and Children's Medical Center during 2016 to 2021. Results: The results revealed that 1,338 cases of Salmonella were mainly isolated from feces and blood samples. The age distribution was dominated by infants under 3 years old. The seasonal distribution was high in summer and autumn. 48 serotypes were detected, and S. typhimurium (78.7%) was the predominant serogroup. The results of antimicrobial susceptibility showed that the highest resistance was observed in ampicillin (84.5%), while lower resistance was observed in piperacillin/tazobactam, cefoperazone/sulbactam and ciprofloxacin. The antimicrobial resistance rate of fecal isolates was higher than that of blood isolates. The five-year average detection rate of multi-drug resistant Salmonella was 8.5% (114/1338) and the MDR rate of S. typhimurium was the lowest (6.9%; 73/1053). Conclusion: We concluded that antibacterial treatment should be carefully selected according to serotype and antimicrobial sensitivity results in children. Antimicrobial resistance monitoring for multi-drug resistant Salmonella is still required.

Genomic Basis of Occurrence of Cryptic Resistance among Oxacillin- and Cefoxitin-Susceptible mecA-Positive Staphylococcus aureus.

The oxacillin- and cefoxitin-susceptible mecA-positive Staphylococcus aureus is a novel "stealth" methicillin-resistant S. aureus (MRSA) type. Here, we sequenced the whole genome of two oxacillin- and cefoxitin-susceptible mecA-positive MRSA isolates from breast abscesses in a lactating woman and a nasal swab of a healthy student in Guangzhou for investigating the mechanism underlying its occurrence. The reversion of these isolates was selected by exposure to sub-MICs of cefoxitin with or without mupirocin. The mecA expression of both parental strains and their revertants was determined, and the whole genome of the revertants was sequenced. Comparative whole-genome analyses performed for both strains revealed that mecA of the clinical strain was mutated by a single-bp insertion at the 262nd position in the tandem repeat region of the gene, and this mutation that led to the formation of a premature stop codon. The colonizing strain was mutated by a novel G-to-A base substitution in the second promoter region (-35 bp) of mecA. The mecA expression level of strain 697 revertant was 37 times higher than that of the parental strain. Although the mecA expression level was even higher for parental strain 199 compared with that for its revertant, its cDNA sequence contained a single-bp insertion. Collectively, both the missense and single substitution mutations of the second promoter of mecA could render MRSA isolates as "stealth" MRSA, thereby emphasizing the importance of combining phenotype tests with mecA or penicillin-binding protein 2a detection for the identification of MRSA. IMPORTANCE The oxacillin- and cefoxitin-susceptible mecA-positive Staphylococcus aureus is a novel type of "stealth" methicillin-resistant S. aureus (MRSA), which is difficult to be detected using conventional methods. To investigate the genomic basis of their occurrence, we sequenced the whole genome of two previously recovered oxacillin- and cefoxitin-susceptible mecA-positive MRSA isolates from breast abscesses in a lactating woman and a nasal swab of a healthy student in Guangzhou. Complete SCCmec structure was absent except for mecA in clinical isolate 199. Additionally, a novel single-base pair insertion was observed in the clinical strain, which resulted in premature termination and a frameshift mutation. The colonizing isolate 697 had a Scc-mec-type IVa, and the second promoter region (-35 bp) of mecA was mutated by a novel G-to-A base substitution. The reversion of oxacillin- and cefoxitin-susceptible mecA-positive S. aureus to resistant MRSA isolates was selected by exposure to subminimum inhibitory cefoxitin with or without mupirocin.

Distribution of Biocide Resistance Genes and Association with Clonal Complex Genotypes in Staphylococcus aureus Isolated from School-Age Children in Guangzhou.

Purpose: Chlorhexidine and mupirocin are often prescribed to children in affected communities to prevent colonization and transmission of Staphylococcus aureus, but this has led to an increasing rate of biocide resistance. In this study, we aimed to determine the distribution of biocide resistance genes among S. aureus isolates from school-age children in Guangzhou, investigate chlorhexidine gluconate and mupirocin susceptibility and clonal complex (CC) genotypes in strains carrying biocide-resistance genes, and further explore the role of biofilms in this resistance. Patients and Methods: Antibiotic resistance and multilocus sequence genotyping were performed on 722 S. aureus isolates from previous study. The distribution of nine biocide genes (qacA/B, mupA, mepA, sepA, norA, lmrS, smr, mupB, qacG) was determined by PCR. Isolates carrying qacA/B or mupA genes were further tested for susceptibility to chlorhexidine gluconate (CHG) and mupirocin and biofilm formation abilities. Results: The most prevalent of the nine biocide resistance genes were mepA (95.57%), followed by norA (78.81%), lmrS (77.01%), and sepA (58.17%). The qacG gene was not detected. Distribution of sepA was significantly decreased in CC30 and CC45 genotypes, and presence of sepA was associated with resistance to antibiotics such as CLI, ERY, TCY, SXT, CIP, and LVX. In addition, 64 (94.1%, n=68) qacA/B+ isolates showed CHG resistance, 12 (100.0%, n=12) mupA+ isolates were mupirocin resistant, and 4 (80%, n=5) and 5 (100%, n=5) qacA/B+mupA+ isolates were CHG and mupirocin resistant, respectively. Of these 85 isolates, 98.8% (n=84) had different degrees of biofilm-forming abilities, which were positively associated with CHG and mupirocin resistance. Conclusion: The distribution of biocide resistance genes was associated with special CCs. The qacA/B and mupA genes are highly associated with resistance to CHG and mupirocin, and biofilm formation was found to contribute to this biocide resistance.

Diagnostic Performance of Various Methodologies for Group B Streptococcus Screening in Pregnant Woman in China.

Maternal vaginal/rectal colonization of group B streptococcus (GBS) is a main risk for neonatal invasive infection. Efficient determination of GBS colonization in pregnant women is crucial. This study aimed to investigate the prevalence of GBS carriage and evaluate the diagnostic performance of six methodologies for GBS screening conducted in China, including blood agar plate, liquid chromogenic medium, and loop-mediated isothermal amplification (LAMP) without pre-enrichment, chromogenic agar plate with pre-enrichment, and GBS antigen detection without and with pre-enrichment in comparison with the standard reference method (Lim broth-enriched subculture with plating on 5% sheep blood agar). Vaginal/rectal swabs were collected from 1,281 pregnant women at 35-37 weeks of gestation. Of them, 309 were taken in triplicate, one for Lim broth-enriched subculture, one for blood agar plate, and the third for GBS antigen detection (Reagent W); 177 were acquired in duplicate, one for Lim broth-enriched subculture and the other for GBS antigen detection (Reagent H); 502 were obtained in duplicate, one for Lim broth-enriched subculture and the other for liquid chromogenic medium; 158 were collected in duplicate, one for Lim broth-enriched subculture and the other for LAMP; and 135 were inoculated in Lim broth-enriched for GBS antigen detection (Reagent W) and subculture with chromogenic agar plate and 5% blood agar plate. The overall prevalence of GBS carriage was 10.1% (130/1,281, 95% CI: 8.5-12.1%) according to the standard reference method. Compared with the standard reference method, the LAMP had excellent performance of sensitivity (100%, 95%CI: 83.4-100%), specificity (94%, 95%CI: 88.1-97.1%), and Yoden index (0.940); as well as the blood agar plate with sensitivity (81.5%, 95%CI: 61.3-93.0%), specificity (100%, 95%CI: 98.3-100.0%), and Yoden index (0.815). The other four methods were not sufficient to reach the threshold in terms of sensitivity or specificity compared to the standard reference method. Furthermore, for LAMP, results can be obtained within 0.5-1 h, while for blood agar plate, which needed 24-48 h, and further identification was required. Our data suggested that the performance of LAMP was highly comparable to the standard Lim broth-enriched subculture and LAMP is considered as an alternative for fast and accurate GBS screening.

Predominance of III/ST19 and Ib/ST10 Lineages With High Multidrug Resistance in Fluoroquinolone-Resistant Group B Streptococci Isolates in Which a New Integrative and Conjugative Element Was Identified.

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.

Rapid Classification of Multilocus Sequence Subtype for Group B Streptococcus Based on MALDI-TOF Mass Spectrometry and Statistical Models.

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.

Rapid and visual detection of Group B streptococcus using recombinase polymerase amplification combined with lateral flow strips.

Conventional culture method for detecting Group B streptococcus (GBS), a common pathogen of neonatal meningitis and sepsis, is time-consuming and unsensitive. Even though real-time fluorescence PCR-based molecular method is more accurate, it need special instrument and elaborate protocol. Here, we established a novel molecular method combining recombinase polymerase amplification with lateral flow strips for detecting GBS. The cAMP factor (cfb) gene is a highly specific and sensitive biomarker to identify GBS and is detectable by using 100 genomic copies as the amplification template. Clinical performance of this assay was evaluated by testing 130 samples, in comparison with culture method and real-time fluorescence PCR, and the results achieved 100% accuracy, which were the same with those of real-time fluorescence PCR, and were better than those of culture method with false-negative detection. This study provides a rapid and visual method, with clinical potential, for the detection of GBS infection of patients.

Prevalence, serotypes, and drug resistance of nontyphoidal Salmonella among paediatric patients in a tertiary hospital in Guangzhou, China, 2014-2016.

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.