Increased MCHC*RDW-SD interaction values: indicators of neurological impairment in lead-poisoned children.

Background: The neurotoxic effects of lead in children can have long-lasting and profound impacts on the developing nervous system. This study aimed to identify a reliable and easily accessible biomarker to monitor neurological impairment in lead-poisoned children. Methods: We analyzed hematological data from 356 lead-poisoned children, comparing them with age and gender-matched healthy controls. Multivariate logistic regression and receiver operating characteristic (ROC) analysis were employed to identify and evaluate potential biomarkers for neurological damage. Results: Significant changes in erythrocyte parameters were observed in lead-poisoned children. Upon further analysis, increased mean corpuscular hemoglobin concentration (MCHC) and red cell distribution width-standard deviation (RDW-SD) interaction values were found to be significantly associated with neurological impairment. The MCHC*RDW-SD interaction model demonstrated an AUC of 0.76, indicating its effectiveness in reflecting neurological damage. Additionally, the MCHC*RDW-SD Interaction value showed weak or no correlation with other erythrocyte parameters, suggesting its independence as an indicator. Conclusion: Our findings propose the increased MCHC*RDW-SD interaction value as a robust and independent biomarker for detecting neurological impairment in lead-poisoned children. This underscores the potential of utilizing specific erythrocyte parameters for screening the neurotoxic effects of lead exposure in pediatric populations.

Impact of LuxS on virulence and pathogenicity in Klebsiella pneumoniae exhibiting varied mucoid phenotypes.

How the LuxS/AI-2 quorum sensing (QS) system influences the pathogenicity of K. pneumoniae is complicated by the heterogeneity of the bacterial mucoid phenotypes. This study aims to explore the LuxS-mediated regulation of the pathogenicity of K. pneumoniae with diverse mucoid phenotypes, including hypermucoid, regular-mucoid, and nonmucoid. The wild-type, luxS knockout, and complemented strains of three K. pneumoniae clinical isolates with distinct mucoid phenotypes were constructed. The results revealed the downregulation of virulence genes of regular-mucoid, and nonmucoid but not hypermucoid strains. The deletion of luxS reduced the pathogenicity of the regular-mucoid, and nonmucoid strains in mice; while in hypermucoid strain, luxS knockout reduced virulence in late growth but enhanced virulence in the early growth phase. Furthermore, the absence of luxS led the regular-mucoid and nonmucoid strains to be more sensitive to the host cell defense, and less biofilm-productive than the wild-type at both the low and high-density growth state. Nevertheless, luxS knockout enhanced the resistances to adhesion and phagocytosis by macrophage as well as serum-killing, of hypermucoid K. pneumoniae at its early low-density growth state, while it was opposite to those in its late high-density growth phase. Collectively, our results suggested that LuxS plays a crucial role in the pathogenicity of K. pneumoniae, and it is highly relevant to the mucoid phenotypes and growth phases of the strains. LuxS probably depresses the capsule in the early low-density phase and promotes the capsule, biofilm, and pathogenicity during the late high-density phase, but inhibits lipopolysaccharide throughout the growth phase, in K. pneumoniae.IMPORTANCECharacterizing the regulation of physiological functions by the LuxS/AI-2 quorum sensing (QS) system in Klebsiella pneumoniae strains will improve our understanding of this important pathogen. The genetic heterogeneity of K. pneumoniae isolates complicates our understanding of its pathogenicity, and the association of LuxS with bacterial pathogenicity has remained poorly addressed in K. pneumoniae. Our results demonstrated strain and growth phase-dependent variation in the contributions of LuxS to the virulence and pathogenicity of K. pneumoniae. Our findings provide new insights into the important contribution of the LuxS/AI-2 QS system to the networks that regulate the pathogenicity of K. pneumoniae. Our study will facilitate our understanding of the regulatory mechanisms of LuxS/AI-2 QS on the pathogenicity of K. pneumoniae under the background of their genetic heterogeneity and help develop new strategies for diminished bacterial virulence within the clinical K. pneumoniae population.

Caspofungin at sub-inhibitory concentration promotes the formation of Candida albicans persister cells.

AIMS: Low caspofungin exposure is frequently encountered in patients with invasive candidiasis caused by Candida albicans. This study aimed to investigate the effects of caspofungin on C. albicans at sub-inhibitory concentrations. METHODS AND RESULTS: First, a comparative transcriptomics analysis was performed on C. albicans receiving caspofungin at sub-minimum inhibitory concentrations (sub-MICs). The results showed that caspofungin significantly changed the mRNA expression profile in DAY185, with DE-mRNAs enriched in the functions of cell wall biosynthesis, metabolism, etc. Subsequently, cellular fitness, cell aggregation, energy metabolism activity and the proportion of persister cells of C. albicans were quantitatively and/or qualitatively assessed after sub-MIC caspofungin exposure. No significant changes in cell fitness and aggregation formation were observed during treatment of C. albicans with sub-MIC caspofungin. In C. albicans aggregation treated with sub-MIC caspofungin, we observed a decrease in respiratory metabolism and an increase in persister cells; this effect was more pronounced in als1ΔΔ than in DAY185. CONCLUSIONS: Pre-exposure to sub-MIC caspofungin suppresses C. albicans respiratory metabolism and promotes persister cell development. SIGNIFICANCE AND IMPACT OF THE STUDY: Caspofungin should be used with caution in patients with C. albicans infections, as anti-infection therapy may fail due to persister cells.

Acquisition of Tigecycline Resistance by Carbapenem-Resistant Klebsiella pneumoniae Confers Collateral Hypersensitivity to Aminoglycosides.

Tigecycline is a last-resort antibiotic for infections caused by carbapenem-resistant Klebsiella pneumoniae (CRKP). This study aimed to broaden our understanding of the acquisition of collateral hypersensitivity by CRKP, as an evolutionary trade-off of developing resistance to tigecycline. Experimental induction of tigecycline resistance was conducted with tigecycline-sensitive CRKP clinical isolates. Antimicrobial susceptibility testing, microbial fitness assessment, genotypic analysis and full-genome sequencing were carried out for these clinical isolates and their resistance-induced descendants. We found that tigecycline resistance was successfully induced after exposing CRKP clinical isolates to tigecycline at gradually increased concentrations, at a minor fitness cost of bacterial cells. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) found higher expression of the efflux pump gene acrB (5.3-64.5-fold) and its regulatory gene ramA (7.4-65.8-fold) in resistance-induced strains compared to that in the tigecycline-sensitive clinical isolates. Stable hypersensitivities to aminoglycosides and other antibiotics were noticed in resistance-induced strains, showing significantly lowered MICs (X 4 - >500 times). Full genome sequencing and plasmid analysis suggested the induced collateral hypersensitivity might be multifaceted, with the loss of an antimicrobial resistance (AMR) plasmid being a possible major player. This study rationalized the sequential combination of tigecycline with aminoglycosides for the treatment of CRKP infections.

Decreased serum bilirubin levels in children with lead poisoning.

OBJECTIVE: Lead is a toxic heavy metal, which causes irreversible damage in children. Oxidative stress is the underlying mechanism of lead toxicity, and monitoring oxidative stress of lead poisoning children in vivo is important. Our study aimed to investigate blood serum levels of biochemical parameters, including albumin, bilirubin, creatinine, and uric acid, which are regarded as non-enzymatic antioxidants, in children with lead poisoning. METHODS: We studied 355 children with lead poisoning and 355 age- and sex-matched controls. We analyzed clinical characteristics and measured serum levels of total protein, globulin, albumin, bilirubin, aspartate aminotransferase (AST), alanine aminotransferase, urea, and creatinine. RESULTS: We found that albumin, bilirubin, urea, and creatinine levels were significantly lower and AST, total protein, and globulin levels were higher in children with lead poisoning than in controls. Direct bilirubin, albumin, total protein, urea, creatinine, and AST levels were associated with lead poisoning after adjustment for other covariates. Spearman analysis showed that direct bilirubin, albumin, and urea levels were independent indicators (i.e., not related to hemoglobin or weight), while creatinine levels showed a moderate correlation with weight. CONCLUSION: Lead interferes with the non-enzymatic antioxidant system in children, and lead poisoning results in a decrease in serum bilirubin levels.

Biofilm Formation of Candida albicans Facilitates Fungal Infiltration and Persister Cell Formation in Vaginal Candidiasis.

BACKGROUND: Vaginal candidiasis is an important medical condition awaiting more effective treatment. How Candida albicans causes this disease and survives antifungal treatment is not yet fully understood. This study aimed to establish a comprehensive understanding of biofilm-related defensive strategies that C. albicans uses to establish vaginal candidiasis and to survive antifungal treatment. METHODS: A mouse model of vaginal candidiasis was adopted to examine the formation of biotic biofilms on the vaginal epithelium and fungal infiltration by laboratory and clinical strains of C. albicans. Histopathological changes and local inflammation in the vaginal epithelium caused by C. albicans of different biofilm phenotypes were compared. Antifungal susceptibility testing was carried out for C. albicans grown as planktonic cells, microplate-based abiotic biofilms, and epithelium-based biotic biofilms. Formation of persister cells by C. albicans in different growth modes was also quantified and compared. RESULTS: C. albicans wild-type reference strains and clinical isolates, but not the biofilm-defective mutants, formed a significant number of biotic biofilms on the vaginal epithelium of mice and infiltrated the epithelium. Biofilm formation and epithelial invasion induced local inflammatory responses and histopathological changes in the vaginal epithelium including neutrophil infiltration and subcorneal microabscesses. Biofilm growth on the vaginal epithelium also led to high resistance to antifungal treatments and promoted the formation of antifungal-tolerant persister cells. CONCLUSION: This study comprehensively assessed biofilm-related microbial strategies that C. albicans uses in vaginal candidiasis and provided experimental evidence to support the important role of biofilm formation in the histopathogenesis of vaginal candidiasis and the recalcitrance of the infection to antifungal treatment.

Characterization of the mechanism and impact of staphylokinase on the formation of Candida albicans and Staphylococcus aureus polymicrobial biofilms.

PURPOSE: Candida albicans and Staphylococcus aureus can be co-isolated in biofilm-associated infections. However, treatments have not been well established due to a lack of antibiofilm strategies. Hence, this study aims to characterize the mechanism and impact of Staphylokinase (Sak) on fungal-bacterial polymicrobial biofilms. METHODOLOGY: Sak generation levels were obtained via chromogenic analysis. C. albicans and S. aureus polymicrobial biofilm formation and integrity were analysed using a bright-field microscope and scanning electron microscopy (SEM). Metabolic mitochondrial activity, growth rate and adhesive capacity were also measured. Quantification real-time RT-PCR (qRT-PCR) was carried out to evaluate the expression levels of biofilm-related genes. Furthermore, the biofilm inhibitory potential of Sak alone or combined with antimicrobials was investigated. RESULTS: Sak production levels varied, ranging from 0.130 to 0.648. A strong decrease of biomass, metabolic activity andearly stage growth rate was demonstrated in the Sak-treated group. SEM showed S. aureus attached on hyphae of C. albicans in sporadic small microcolonies after Sak treatment. Moreover, the gene expression levels of HWP1, EFG1 and NRG1 were significantly altered, while no obvious difference was observed in ALS3. Finally, Sak had a notable impact on mature polymicrobial biofilms alone or when combined with vancomycin and fluconazole. CONCLUSION: The effect induced by Sak to C. albicans and S. aureus polymicrobial biofilms is caused by decreased biomass, biofilm integrity, metabolic activity and early stage growth rate. Alterations of gene expression levels were consistent with Sak-induced phenotypic change. Combined treatment strategies are essential for optimal activities against fungal-bacterial polymicrobial biofilms.

Description and plasmid characterization of the qnrD determinant in Proteeae in Wenzhou, Southern China.

BACKGROUND/PURPOSE: Only limited information is available about the detailed characteristics of qnrD, a plasmid-mediated quinolone resistance (PMQR) gene. This study aimed to understand the distribution of qnrD and the characterization of qnrD-carrying plasmids in Proteeae. METHODS: The distribution of qnrD genes was investigated by polymerase chain reaction (PCR) amplification in 203 consecutive nonduplicate clinical isolates of Proteeae collected from inpatients at the First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China. The minimum inhibitory concentrations (MICs) of antibiotics were measured by agar dilution method and other PMQR determinants were also determined by PCR. qnrD was positioned via Southern hybridization and the transferability of qnrD-carrying plasmids was achieved by conjugation experiment. The genetic environment of qnrD was investigated by sequencing, and chromosomal polymorphism for qnrD-positive strains was analyzed by pulsed-field gel electrophoresis (PFGE). RESULTS: Forty strains carried qnrD, showing decreased fluoroquinolone susceptibility or low-level fluoroquinolone resistance. qnrD was encoded on the plasmid of about 2.7 kb or 5.2 kb in length, which cannot be transferred by liquid conjugation or filter mating, but can be successfully transferred by transduction. The transformants showed 62.5-300-fold increases in the MICs of quinolones compared with the recipient. The plasmids carrying qnrD showed a high similarity with that of Providencia spp. and Proteus vulgaris. PFGE analysis demonstrated that these isolates were divergent and not clone related. CONCLUSION: qnrD could have originated from Proteeae or presented in these bacteria as a reservoir; furthermore, qnrD could be transferred and spread within the same or across different bacterial species if the plasmids acquired mobile elements under antimicrobial selective pressures.

Predation Efficacy of Bdellovibrio bacteriovorus on Multidrug-Resistant Clinical Pathogens and Their Corresponding Biofilms.

The aim of the present study was to evaluate the predation efficacy of Bdellovibrio bacteriovorus on multidrug-resistant (MDR) or extensive drug resistant (XDR) gram-negative pathogens and their corresponding biofilms. In this study, we examined the ability of B. bacteriovorus to prey on MDR and XDR gram-negative clinical bacteria, including Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii. Results showed that B. bacteriovorus was able to prey on all planktonic cultures, among which the most efficient predation was observed for drug-resistant E. coli, with a 3.11 log10 reduction in viability. Furthermore, B. bacteriovorus demonstrated promising efficacy in preventing biofilm formation and dispersing the established biofilm. Reductions in biofilm formation of E. coli, K. pneumoniae, P. aeruginosa, and A. baumannii co-cultured with B. bacteriovorus were 65.2%, 37.1%, 44.7%, and 36.8%, respectively. Meanwhile, the established biofilms of E. coli, K. pneumoniae, P. aeruginosa, and A. baumannii were significantly reduced by 83.4%, 81.8%, 83.1%, and 79.9%, respectively. A visual analysis supported by scanning electron microscopy demonstrated the role of B. bacteriovorus in removing the established biofilms. This study highlights the potential use of B. bacteriovorus as a biological control agent with the capability to prey on MDR/XDR gram-negative pathogens and eradicate biofilms.

Antioxidant status of serum bilirubin and uric acid in patients with polymyositis and dermatomyositis.

OBJECTIVE: Oxidative stress and variations in antioxidant status are implicated in the pathogenesis of inflammatory and autoimmune diseases. Polymyositis and dermatomyositis (PM/DM) are autoimmune diseases with inflammatory cells infiltrating into skeletal muscles, and the antioxidant status is still controversial. The aim of our study was to investigate the correlation between PM/DM and the antioxidant status of serum bilirubin (Tbil, Dbil and Ibil) and uric acid (UA). MATERIALS AND METHODS: We measured serum concentrations of bilirubin (Tbil, Dbil and Ibil) and uric acid in 384 individuals, including 110 PM/DM patients and 274 healthy controls. RESULTS: We found that PM/DM patients had significantly lower serum concentrations of bilirubin (Tbil and Ibil) and uric acid than healthy controls, whether male or female. Also, after separately adjusting the covariances of age and gender, Tbil, Dbil, Ibil and UA were all relevant factors for PM/DM. Moreover, there were no significant differences in serum antioxidant molecule levels between PM and DM subgroups. CONCLUSION: Our study demonstrated the low serum levels of bilirubin and uric acid in patients with PM/DM. This suggested low antioxidant status in PM/DM patients with excessive oxidative stress.

Biofilm Formation Restrained by Subinhibitory Concentrations of Tigecyclin in Acinetobacter baumannii Is Associated with Downregulation of Efflux Pumps.

BACKGROUND: Tigecycline, one of the few therapeutic options against multidrug-resistant Acinetobacter baumannii, reaches subinhibitory serum concentrations only with cautious clinical dosing and pharmacokinetics. Subinhibitory concentrations of tigecycline might induce an A. baumannii biofilm. METHODS: Biofilm formation was assessed via the crystal violet staining method. We further analyzed the main biofilm components with NaIO4, proteinase K, and DNase. Real-time RT-PCR was applied for quantitative detection of biofilm potential-associated genes. RESULTS: In this study, A. baumannii proved to be a strong biofilm producer, and we found that proteins and extracellular DNA are crucial components of the A. baumannii biofilm. Quantitative real-time RT-PCR revealed positive correlations between biofilm formation restrained by subinhibitory concentrations of tigecycline and the expression of biofilm potential-associated genes, especially the AdeFGH efflux pump gene. CONCLUSION: Our results suggest that downregulation of efflux pumps, especially the AdeFGH efflux pump, is probably responsible for the decline in biofilm formation in A. baumannii treated with subinhibitory concentrations of tigecyclin.

Molecular Characterization and Epidemiologic Study of NDM-1-Producing Extensively Drug-Resistant Escherichia coli.

The emergence and dissemination of NDM-1 (New Delhi metallo-ß-lactamase-1)-producing Enterobacteriaceae have resulted in a worldwide public health risk. This study described a high incidence and endemic spread of NDM-1-producing extensively drug-resistant Escherichia coli in a teaching hospital in Zhejiang province, China. We recovered six nonduplicated NDM-1-producing E. coli isolates from May 2014 to August 2014 with positive modified Hodge test and EDTA synergistic test. These isolates were highly resistant to ß-lactam antimicrobials, aminoglycosides, and quinolones. PCR and DNA sequences analysis showed that all isolates carried the blaNDM-1, blaSHV-11, aac(6')-ib-cr, and qnrB. Several isolates also harbored blaCTX-M-1, blaCTX-M-9, rmtB, and qnrA. Southern blot confirmed that blaNDM-1 was located on the same ∼55 kb plasmid and conjugation experiments further proved the contransferable characteristic of blaNDM-1. The ompC sequences showed various mutations, which was related to multidrug resistance in E. coli. Pulsed-field gel electrophoresis identified four of six isolates that belonged to the same genotype. Multilocus sequence typing assigned them to ST2, except one strain that belonged to ST594. Our study demonstrated that the resistance-associated genes and the loss of the outer membrane proteins could account for high resistance of NDM-1-producing E. coli to multiple antimicrobial drugs. Both horizontal transfer of IncN and transmission of ST2 were responsible for the spread of drug resistance. These findings highlighted an urgent need to limit the further dissemination of NDM-1-producing E. coli in this region.

Prevalence and molecular characterization of carbapenemase-producing gram-negative bacteria from a university hospital in China.

BACKGROUND: The increasing emergence of carbapenem resistance in gram-negative bacteria associated with carbapenemase prompted the initiation of this study. METHODS: A total of 3139 gram-negative bacteria were recovered from a 3380-bed university hospital in Wenzhou during 2008 and 2012. Antimicrobial susceptibility was determined using the VITEK2 Compact System and agar dilution method. The phenotype and genotype of carbapenemase were demonstrated using the modified Hodge test, PCR and sequencing. A conjugation experiment was performed to reveal the transferability of resistant genes. The location of the carbapenemase gene was studied by plasmid analysis and southern blot hybridization. Clonal relatedness of the isolates was investigated by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). RESULTS: Overall, 751 of 3139 isolates (71/2055 Enterobacteriaceae, 510/620 Acinetobacter baumannii and 170/464 Pseudomonas aeruginosa) exhibited resistance to carbapenem. Carbapenemase-encoding genes were detected in 70.4% (50/71) of carbapenem-resistant Enterobacteriaceae, including blaKPC (80%) and blaIMP (20%). All A. baumannii subjected to genotype analysis were positive for blaOXA-51-like and co-harboured blaOXA-23-like (80.4%) and blaIMP (7.8%). ISAba1 was found upstream of blaOXA-23-like and blaOXA-51-like. Eight and seven strains of 170 P. aeruginosa carried blaIMP and blaVIM, respectively. PFGE analysis identified at least one dominant genotype in certain species. Four KPC-2-producing Klebsiella pneumoniae belonged to the same sequence type ST11. The plasmids carrying blaKPC were successfully transferred into recipient strains. CONCLUSION: This study highlights the challenge of increasing prevalence of carbapenem resistance associated with carbapenemase genes and dissemination of epidemic clones in Wenzhou, China.

[Spermatozoal immobilization ability and virulence genes of Staphylococcus aureus isolated from the semen of infertile men].

OBJECTIVE: To investigate the impact of Staphylococcus aureus from infertile men on sperm motility and the relationship between virulence genes and the activity of spermatozoal immobilization. METHODS: We collected 60 strains of non-repeated Staphylococcus aureus from the semen of 589 infertile males and analyzed the influence of Staphylococcus aureus on sperm motility using the computer-aided sperm analysis system. We selected the strains that apparently decreased sperm motility and detected their virulence genes by PCR. RESULTS: Sperm motility was significantly decreased in 17 of the 60 strains of Staphylococcus aureus (P < 0.05). The main virulence genes in these strains were hlg (33.3%), scn (23.3%), cna (20%), hlb (20%), and clfA (18.3%), others including icaA, fnbA, tst, seb, hld, eta and sea. The scn gene carriers accounted for 47.1% in the spermatozal immobilization positive group, significantly higher than 14% in the negative group (P < 0.05). No statistically significant differences were found in the percentages of the carriers of the other virulence genes between the two groups (P > 0.05). CONCLUSION: Infections of Staphylococcus aureus in male reproductive system can lead to the decrease of sperm motility, which may be associated with the Staphylococcus complement inhibitor encoding gene scn.