Genome-wide identification and functional analysis of circRNAs in Trichophyton rubrum conidial and mycelial stages.

BACKGROUND: Circular RNAs (circRNAs) are a group of noncoding RNAs that participate in gene expression regulation in various pathways. The essential roles of circRNAs have been revealed in many species. However, knowledge of circRNAs in fungi is still not comprehensive. RESULTS: Trichophyton rubrum (T. rubrum) is considered a model organism of human pathogenic filamentous fungi and dermatophytes. In this study, we performed a genome-wide investigation of circRNAs in T. rubrum based on high-throughput sequencing and ultimately identified 4254 circRNAs. Most of these circRNAs were specific to the conidial or mycelial stage, revealing a developmental stage-specific expression pattern. In addition, 940 circRNAs were significantly differentially expressed between the conidial and mycelial stages. PCR experiments conducted on seven randomly selected differentially expressed (DE-) circRNAs confirmed the circularized structures and relative expression levels of these circRNAs. Based on their genome locations, most circRNAs originated from intergenic regions, unlike those in plants and animals. Furthermore, we constructed circRNA-miRNA-mRNA regulatory networks that included 661 DE-circRNAs targeting 140 miRNAs and further regulating 2753 mRNAs. The relative expression levels of two randomly selected circRNA-miRNA-mRNA axes were investigated by qRT-PCR, and the competing endogenous RNA (ceRNA) network theory was validated. Functional enrichment analysis of the target genes suggested that they were significantly involved in posttranscriptional processes and protein synthesis as well as some small-molecule metabolism processes. CircRNAs are relatively more conserved in closely related dermatophytes but rarely conserved in distantly related species. Tru_circ07138_001 is a highly conserved circRNA that was conserved in all ten dermatophytes analyzed in our study and three distantly related species. Its host gene TERG_07138 was also highly conserved in two of these distantly related species Gallus gallus and Caenorhabditis elegans. The specific role of this circRNA deserves further exploration. CONCLUSIONS: Our study is the first to provide a global profile of circRNAs in T. rubrum as well as dermatophytes. These results could serve as valuable resources for research on circRNA regulatory mechanisms in fungi and reveal new insights for further investigation of the physical characteristics of these significant human fungal pathogens.

Distribution of fluoroquinolone resistance determinants in Carbapenem-resistant Klebsiella pneumoniae clinical isolates associated with bloodstream infections in China.

BACKGROUND: The rate of fluoroquinolone (FQ) resistance among carbapenem-resistant Klebsiella pneumoniae (CRKP) is high. The present study aimed to investigate the distribution of fluoroquinolone resistance determinants in clinical CRKP isolates associated with bloodstream infections (BSIs). RESULTS: A total of 149 BSI-associated clinical CRKP isolates collected from 11 Chinese teaching hospitals from 2015 to 2018 were investigated for the prevalence of fluoroquinolone resistance determinants, including plasmid-mediated quinolone resistance (PMQR) genes and spontaneous mutations in the quinolone resistance-determining regions (QRDRs) of the gyrA and parC genes. Among these 149 clinical CRKP isolates, 117 (78.5%) exhibited resistance to ciprofloxacin. The GyrA substitutions (Ser83 → IIe/Phe) and (Asp87 → Gly/Ala) were found among 112 (75.2%) of 149 isolates, while the substitution (Ser80 → IIe) of ParC was found in 111 (74.5%) of the 149 isolates. In total, 70.5% (105/149) of the CRKP isolates had at least two mutations within gyrA as well as a third mutation in parC. No mutations in the QRDRs were found in 31 ciprofloxacin susceptible CRKP isolates. Eighty-nine (56.9%) of 149 were found to carry PMQR genes including qnrS1 (43.0%), aac(6')-Ib-cr (16.1%), qnrB4 (6.0%), qnrB2 (2.7%), and qnrB1 (1.3%). Nine isolates contained two or more PMQR genes, with one carrying four [aac(6')-Ib-cr, qnr-S1, qnrB2, and qnrB4]. The co-existence rate of PMQR determinants and mutations in the QRDRs of gyrA and parC reached 68.5% (61/89). Seventy-four (83.1%, 74/89) PMQR-positive isolates harbored extended-spectrum beta-lactamase (ESBL)-encoding genes. Multilocus sequence typing (MLST) analysis demonstrated that the ST11 was the most prevalent STs in our study. CONCLUSIONS: Mutations in the QRDRs of gyrA and parC were the key factors leading to the high prevalence of fluoroquinolone resistance among BSI-associated CRKP. The co-existence of PMQR genes and mutations in the QRDRs can increase the resistance level of CRKP to fluoroquinolones in clinical settings. ST11 CRKP isolates with identical QRDR substitution patterns were found throughout hospitals in China.

Comparison of therapy with ß-lactam/ß-lactamase inhibitor combinations or carbapenems for bacteraemia of nonurinary source caused by ESBL-producing Escherichia coli or Klebsiella pneumoniae.

BACKGROUND: Extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae has become a public health concern. This study aimed to compare the clinical outcomes of patients with nonurinary source bacteraemia caused by ESBL-producing Escherichia coli (E. coli) or Klebsiella pneumoniae (ESBL-producing EK) receiving ß-lactam/ß-lactamase inhibitor combinations (BLICs) versus carbapenem treatment and assess the risk factors of mortality with these two drugs. METHODS: We conducted a retrospective single-centre study of adult hospitalised patients with ESBL-producing EK bloodstream infection (BSI) from nonurinary source at our centre over a 4-year period. One hundred and eighty patients who received BLICs or carbapenems were included in the analysis. The outcome variables were 14-day treatment failure and 30-day mortality. For more reliable results, propensity score analysis was performed to compare the efficacy of the two drugs and analyse their risk factors for 30-day mortality. RESULTS: Out of 180 patients, 114 received BLICs, and 66 received carbapenem therapy. Compared to carbapenem-treated patients, those treated with BLICs were older and had higher age-adjusted Charlson comorbidity index, but they had shorter stay in the hospital. Additionally, their Pitt bacteraemia score, SOFA score, rate of leukaemia, and immune compromise were lower. After propensity score matching (PSM), the baseline characteristics of patients in the two treatment groups were balanced. BLICs were associated with a higher 14-day treatment failure rate (20.6%, 13/63) than carbapenems (16.3%, 7/43), although the difference was not significant in either univariate analysis (P = 0.429) or multivariate analysis (P = 0.122). And the 30-day mortality rate in BTG (11.1%, 7/63) and CTG (11.6%, 5/43) did not significantly differ (univariate analysis, P = 0.926; multivariate analysis, P = 0.420). In the multivariate analysis, after PSM, leukaemia was the only independent predictor of mortality in both BTG and CTG. CONCLUSIONS: Our study showed that BLICs had higher 14-day treatment failure rate compared with carbapenems, although there were no statistically significant differences because of the small number of patients, therefore, further evaluation of the efficacy of BLICs is needed.

Resveratrol enhances the antimicrobial effect of polymyxin B on Klebsiella pneumoniae and Escherichia coli isolates with polymyxin B resistance.

BACKGROUND: Multidrug resistant (MDR) Gram-negative bacterial infections are a serious threat to human health due to the lack of effective treatments. In this study, we selected 50 Gram-negative bacterial strains, including 26 strains of Klebsiella pneumoniae and 24 strains of Escherichia coli, to explore whether resveratrol and polymyxin B have a synergistic killing effect. RESULTS: MIC values against polymyxin B were ≥ 4 µg/mL for 44 of the strains and were 2 µg/mL for the other 6 strains. MICs against polymyxin B in the isolates tested were significantly reduced by the addition of resveratrol. The degree of decline depended on the bacteria, ranging from 1/2 MIC to 1/512 MIC, and the higher the concentration of resveratrol, the greater the decrease. Checkerboard analysis indicated a synergistic effect between resveratrol and polymyxin B; the optimal drug concentration for different bacteria was different, that of resveratrol ranging from 32 µg/mL to 128 µg/mL. Subsequent time-kill experiments showed that a combination of polymyxin B and resveratrol was more effective in killing bacteria. CONCLUSIONS: Our in vitro studies have shown that resveratrol can increase the sensitivity of MDR bacterial strains to polymyxin B, suggesting a potential new approach to the treatment of MDR infections.

Comparison and evaluation of neutralization of clinically frequently used antimicrobial agents using three different culture media in simulated blood cultures.

The performance of BACT/ALERT FA/FN Plus (France) blood culture containing a novel resin, DL (China) blood culture containing common resin, and adsorbent-free REDOX (USA) blood culture relying on dilution for antimicrobial neutralization at %peak serum concentration was evaluated by measuring the recovery of organisms and time to detection (TTD) in nine simulated microorganism-antimicrobial combination blood cultures. Significant differences were observed in the recovery rates among the aerobic media: 87.5% for BACT/ALERT media, 42.9% for DL media, and 12.5% for REDOX media. In contrast, no statistical difference was found in the TTD between FA Plus media and DL aerobic media. For the anaerobic media, the recovery rates were 91.4% for BACT/ALERT media, 2.9% for DL media, and 14.3% for REDOX media, with significant differences only between BACT/ALERT FN Plus media and the others. Among the seven main antimicrobial categories, only BACT/ALERT FA/FN Plus culture media demonstrated high recovery of microorganisms, with the exception of carbapenems. The DL culture media exhibited a relatively high recovery rate of microorganisms in the presence of piperacillin/tazobactam, levofloxacin, and gentamicin, but only in aerobic conditions. Conversely, REDOX media displayed microorganism recovery solely in the presence of gentamicin. BACT/ALERT FA/FN Plus culture media with novel resin showed absolute advantages over DL and REDOX culture media and can, therefore, be selectively applied in clinical settings when antimicrobials are used prior to blood collection. DL culture media, containing common resin, outperformed adsorbent-free dilution-based REDOX culture media, making it a viable backup option. There is a need to focus on improving the neutralization of carbapenems with current inefficiency in all three medias. IMPORTANCE: We present a study on performance comparison of three different commercial culture media for neutralization of antibiotic effects in simulated blood cultures. BACT/ALERT (FA Plus and FN Plus) culture media with novel resin showed absolute advantages over DL and REDOX culture media at %PSL concentration of antimicrobials.

Clinical Characteristics, Prognosis and Treatment of Bloodstream Infections with Enterobacter Cloacae Complex in a Chinese Tertiary Hospital: A Retrospective Study.

Objective: This research aimed to analyze the clinical characteristics, prognosis, and antimicrobial treatment of bloodstream infections (BSI) caused by Enterobacter cloacae complex (ECC). Methods: The clinical data of patients with bloodstream infections caused by Enterobacter cloacae complex from April 2017 to June 2023 were collected retrospectively. These data were then analyzed in subgroups based on the detection results of extended-spectrum ß-lactamase (ESBL), 30-day mortality, and the type of antimicrobial agent used (ß-lactam/ß-lactamase inhibitor combinations (BLICs) or carbapenems). Results: The proportion of ESBL-producing Enterobacter cloacae complex was 32.5% (37/114). Meanwhile, ICU admission, receiving surgical treatment within 3 months, and biliary tract infection were identified as risk factors for ESBL-producing ECC-BSI. Additionally, immunocompromised status and Sequential Organ Failure Assessment (SOFA) score ≥ 6.0 were identified as independent risk factors of 30-day mortality in patients with ECC-BSI (n = 108). Further analysis in BSI patients caused by non-ESBL-producing ECC revealed that patients treated with BLICs (n = 45) had lower SOFA scores and lower incidence of hypoproteinemia and sepsis compared with patients treated with carbapenems (n = 20). Moreover, in non-ESBL-producing ECC-BSI patients, the univariate Cox regression analysis indicated a significantly lower 30-day mortality rate in patients treated with BLICs compared to those treated with carbapenems (hazard ratios (HR) [95% CI] 0.190 [0.055-0.662], P = 0.009; adjusted HR [95% CI] 0.106 [0.013-0.863], P = 0.036). Conclusion: This study investigated the factors influencing the susceptibility to infection by ESBL-producing strains and risk factors for 30-day mortality in ECC-BSI patients. The results revealed that ESBL-negative ECC-BSI patients treated with BLICs exhibited significantly lower 30-day mortality compared to those treated with carbapenems. BLICs were found to be more effective in ECC-BSI patients with milder disease (ESBL-negative and SOFA ≤6.0).

Assessment of Effective Antimicrobial Regimens and Mortality-Related Risk Factors for Bloodstream Infections Caused by Carbapenem-Resistant Acinetobacter baumannii.

Objective: This study aimed to determine the clinical features, risk factors, and effective antimicrobial therapy for Carbapenem-resistant Acinetobacter baumannii (CRAB) bloodstream infection (BSI). Methods: This was a retrospective analysis of data from patients with CRAB bacteremia in a Chinese tertiary hospital between January 2012 and October 2021. Risk factors, predictors of 30-day mortality, and effective antimicrobial therapy for CRAB BSI were identified using logistic and cox regression analyses. Results: Data from 276 patients with Acinetobacter baumannii (AB) BSI were included, of whom 157 (56.9%) had CRAB BSI. The risk factors that were significantly associated with CRAB BSI included previous intensive care unit (ICU) stay (P < 0.001), immunocompromised status (P < 0.001), cephalosporin use (P = 0.014), and fluoroquinolone use (P = 0.007). The 30-day mortality of the CRAB BSI group was 49.7% (78/157). ICU stay after BSI (P = 0.047), sequential organ failure assessment (SOFA) score ≥10 (P < 0.001), and multiple organ failure (MOF) (P = 0.037) were independent predictors of 30-day mortality. Among antibiotic strategies for the treatment of patients with CRAB BSI, we found that definitive regimens containing cefoperazone/sulbactam were superior to those without cefoperazone/sulbactam in reducing the 30-day mortality rate (25.4% vs 53.4%, P = 0.005). After propensity score matching, we observed a significant increase in the 30-day mortality (77.8%vs 33.3%, P = 0.036) in patients receiving tigecycline monotherapy compared to those receiving cefoperazone/sulbactam monotherapy. The mortality rate of patients receiving tigecycline with cefoperazone/sulbactam was also higher than that of patients receiving cefoperazone-sulbactam monotherapy; however, the difference was not significant (28.6%vs 19.0%, P = 0.375). Conclusion: The severity of patient conditions was significantly associated with mortality in patients with CRAB BSI. Those Patients treated with cefoperazone/sulbactam had better clinical prognoses, and tigecycline should be used with caution.

Ceftazidime/Avibactam, Polymyxin or Tigecycline as a Rescue Strategy for the Treatment of Carbapenem-Resistant Klebsiella pneumoniae in Bloodstream Infection: A Retrospective Cohort Study.

Objective: To analyze the clinical characteristics, outcomes, and risk factors of patients treated with ceftazidime/avibactam, polymyxin, or tigecycline (CPT) compared with those receiving a conventional therapy (CT) (ie, imipenem, levofloxacin, or gentamicin). Methods: A single-center retrospective cohort study included patients with carbapenem-resistant Klebsiella pneumoniae bloodstream infection (CRKP-BSI) treated at one Chinese tertiary hospital between March 2012 and November 2022 was performed. Clinical characteristics, outcomes, and risk factors of patients treated with CPT or CT were compared. Predictors of 30-day mortality of patients with CRKP-BSI were also analysed in our study. Results: Among 184 recruited patients with CRKP-BSI, 39.7% (73/184) were treated with CPT, while 60.3% (111/184) were treated with CT. Compared to patients treated with CT, patients treated with CPT had worse conditions, as evidenced by a higher rate of underlying diseases and invasive procedures; however, they also had a better prognosis and lower rates of 14-day treatment failure (p = 0.024). In addition, univariate analysis and multivariate analysis showed that SOFA score [odds ratio (OR) = 1.310, 95% confidence interval (CI) 1.157-1.483; p < 0.001] and cold weather (OR = 3.658, 95% CI 1.474-9.081; p = 0.005) were independent risk factors for 30-day mortality. Conclusion: Compared to CRKP-BSI patients treated with CT, patients treated with CPT had worse conditions but better prognoses. CRKP-BSI occurred more frequently in hot weather; however, higher 30-day mortality was associated with cold weather. A randomized trial is needed to confirm these observational results.

Comparison of Molecular Characteristics Between Methicillin-Resistant and -Susceptible Staphylococcus aureus Clinical Isolates by Whole-Genome Sequencing.

Introduction: The transmission of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible Staphylococcus aureus (MSSA) are great public health concern worldwide. To better understand S. aureus evolution and dissemination, we compared the molecular features of MSSA and MRSA isolates. Methods: In this study, 74 MSSA and 102 MRSA non-duplicate isolates were recovered from clinical samples between 2016 and 2020. Molecular epidemiology, antimicrobial resistance determinants, and virulence gene profiles were carried out by whole-genome sequencing (WGS). Results: Twenty distinct sequence types were identified in MRSA isolates, with the most common being ST59, ST630, and ST338. The major genotypes of MSSA were ST188 and ST7. The toxin genes clfA, sek, and seq were significantly associated with MRSA, while splA/B, clfB, map, sdrC/D, and sem-sen-seo-seu were detected more frequently in MSSA isolates than MRSA (P < 0.05). The tst positive isolates were more commonly identified in CC1 and CC72, whereas lukE/D was mainly found in the CC7, CC15, CC88, and completely absent in CC59 clones. Conclusion: Our results compared the genetic diversity between MRSA and MSSA strains, suggesting efforts to fight infections caused by MSSA need to be intensified due to MSSA isolates carrying wide range of virulence factors. Comparative epidemiological studies of large populations of MSSA and MRSA will be necessary in the future to understand how MSSA and MRSA populations may co-evolve and interact in the future.

Emergence of the Coexistence of mcr-1, bla NDM-5, and bla CTX-M-55 in Klebsiella pneumoniae ST485 Clinical Isolates in China.

INTRODUCTION: Polymyxin resistance caused by the plasmid-mediated mcr-1 gene in gram-negative bacilli poses a huge threat to our health. In recent years, many regions have reported that mcr-1 and ß-lactamase genes can coexist in a single strain. METHODS: In this study, 107 nonduplicate Klebsiella pneumoniae (K. pneumoniae) isolates were collected from a tertiary hospital in Jiangxi, China. Antimicrobial susceptibility testing of isolates was performed using gram-negative susceptibility cards on the VITEK system. The minimum inhibitory concentrations (MICs) of polymyxin B was detected using the microdilution broth method. The presence of resistance genes was assessed using polymerase chain reaction (PCR). We subjected isolates to genotyping using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) and analyzed the transferability of plasmids with filter mating and electroporation. Subsequently, whole-genome sequencing was performed for plasmids. RESULTS: Of the 107 K. pneumoniae isolates, 15 (14.0%) were resistant to polymyxin B. All polymyxin B-resistant isolates harbored at least one of the extended-spectrum ß-lactamase genes tested. Only one isolate simultaneously harbored mcr-1, blaNDM-5, blaCTX-M-55 , and blaSHV-27 genes. MLST results showed that 15 carbapenem-resistant K. pneumoniae isolates belonged to five sequence types (STs). PFGE results displayed nine different PFGE clusters. Conjugation and transformation experiments and sequencing analysis showed that the strain had three plasmids, and mcr-1, blaNDM-5 , and blaCTX-M-55 were located on different plasmids. CONCLUSION: The present study demonstrated for the first time the coexistence of mcr-1, blaNDM-5 , and blaCTX-M-55 in a K. pneumoniae ST485 isolate. The three plasmids carrying the mcr-1, blaNDM-5 , and blaCTX-M-55 genes can be transmitted in Enterobacteriaceae strains, which may lead to more severe bacterial resistance.

Impact of inappropriate empirical antibiotic treatment on clinical outcomes of urinary tract infections caused by Escherichia coli: a retrospective cohort study.

OBJECTIVES: We aimed to determine the clinical impact of inappropriate empirical antibiotic treatment (IEAT) compared with appropriate empirical antibiotic treatment (AEAT) in hospitalised patients with urinary tract infections (UTIs) caused by Escherichia coli (E. coli). METHODS: This retrospective cohort study included adult patients with a primary diagnosis of UTI who were treated with empirical antibiotics at a tertiary hospital in southern China over a 2-year period. Clinical data of patients who received IEAT were compared with those of patients receiving AEAT. We used multivariable logistic regression to identify the predictors for receiving IEAT and the risk factors affecting clinical outcomes. RESULTS: A total of 213 patients were enrolled (median age, 61 years), of whom 103 (48.4%) received IEAT. IEAT was associated with empirical use of fluoroquinolones, male sex and age-adjusted Charlson comorbidity index (aCCI) score >6. Hospital length of stay (LOS) was longer for patients who received IEAT than for those who received AEAT (13.6 ± 8.6 days vs. 10.8 ± 7.9 days; P = 0.008). IEAT was an independent risk factor for longer LOS along with aCCI score ≥2, lung disease and cardiac disease. CONCLUSION: Empirical use of fluoroquinolones for UTIs should be avoided, especially in male patients with aCCI score >6. Improved empirical antimicrobial therapy may have a beneficial impact in reducing bacterial resistance and healthcare costs by decreasing the LOS. Therefore, interventions to promote in-depth antibiotic stewardship programmes in China are needed.

Rifampin-resistance-associated mutations in the rifampin-resistance-determining region of the rpoB gene of Mycobacterium tuberculosis clinical isolates in Shanghai, PR China.

Introduction. Resistance to rifampin (RIF) in Mycobacterium tuberculosis infection is associated with mutations in the rpoB gene coding for the ß-subunit of RNA polymerase. The contribution of various rpoB mutations to the development and level of RIF resistance remains elusive.Hypothesis/Gap Statement. Various rpoB mutations may be associated with differential levels of RIF resistance.Aim. This study aimed to investigate the relationship between specific rpoB mutations and the MICs of RIF and rifabutin (RFB) against M. tuberculosis.Methodology. Of the 195 clinical isolates, 105 and 90 isolates were randomly selected from isolates resistant to RIF and sensitive to RIF, respectively. The MICs of 12 agents for M. tuberculosis isolates were determined using commercial Sensititre M. tuberculosis MIC plates and the broth microdilution method. Strains were screened for rpoB mutations by DNA extraction, rpoB gene amplification and DNA sequence analysis.Results. One hundred isolates (95.24 %) were found to have mutations in the RIF-resistance-determining region (RRDR) of the rpoB gene. Three rpoB mutations were identified in 90 RIF-susceptible isolates. Out of 105 isolates, 86 (81.90 %) were cross-resistant to both RIF and RFB. The most frequent mutation occurred at codons 450 and 445. We also found a novel nine-nucleotide (ATCATGCAT) deletion (between positions 1543 and 1551) in the rpoB gene in two strains (1.90 %) with resistance to RIF, but susceptibility to RFB. In addition, the mutation frequency at codon 450 was significantly higher in RIF-resistant/RFB-resistant (RIFR/RFBR) strains than in RIFR/RFBS strains (75.58 % versus 21.05 %, P<0.01), whereas the mutation frequency at codon 435 was significantly lower in RIFR/RFBR strains than in RIFR/RFBS strains (1.16 % versus 26.32 %, P<0.01).Conclusion. Our data support previous findings, which reported that various rpoB mutations are associated with differential levels of RIF resistance. The specific mutations in the rpoB gene in RIFR/RFBR isolates differed from those in the RIFR/RFBS isolates. A novel deletion mutation in the RRDR might be associated with resistance to RIF, but not to RFB. Further clinical studies are required to investigate the efficacy of RFB in the treatment of infections caused by M. tuberculosis strains harbouring these mutations.

Study on acrylate peelable nuclear detergent for film formation at low temperature.

The human handling of radioactive contamination on the surface of nuclear facilities has become an important issue in the development of nuclear industry. In order to solve the poor film-forming of nuclear detergent in low temperature, a new type low temperature film-forming peelable nuclear decontaminant was proposed. The acrylate copolymers were developed by solution polymerization of acrylic monomers in the mass ratio of BA (butyl acrylate): MMA (methyl methacrylate): AA (acrylic acid): EA (ethyl acetate) is equal to 50:45:5:100 considering different proportioning methods and reaction temperature. Then the orthogonal test was carried out to study the effects of crosslinker, plasticizer, thickener, release agent on the decontamination performance based on the orthogonal test method, and the optimal peelable formula of peelable nuclear detergent was obtained. Next the infrared properties, glass transition temperature, molecular weight of polymer and decontamination efficiency of peelable nuclear decontaminant were studied by experiments. The results show that the proposed low-temperature film-forming peelable radioactive decontamination agent can form film on concrete, sand, terrazzo, stainless steel and other surfaces, and is easy to peel off. The decontamination rate of this material is more than 82%.

Subinhibitory Concentrations of Fusidic Acid May Reduce the Virulence of S. aureus by Down-Regulating sarA and saeRS to Reduce Biofilm Formation and α-Toxin Expression.

Staphylococcus aureus is an important pathogen in hospital and community infections. Fusidic acid is particularly effective in treating skin and wound infections caused by staphylococci. The purpose of our study was to clarify the effect of fusidic acid on the biofilm formation and α-toxin expression of S. aureus at subinhibitory concentrations [1/64, 1/32, and 1/16 × minimum inhibitory concentration (MIC)]. A total of 504 genes greater than a twofold or less than twofold change in expression of S. aureus effected by subinhibitory concentrations of fusidic acid were found, including 232 up-regulated genes and 272 down-regulated genes, which were determined by transcriptome sequencing. Our results showed subinhibitory concentrations of fusidic acid significantly inhibited the expression of hla, spa, icaA, and cidA at the mRNA level in clinical S. aureus strains tested. And subinhibitory concentrations of fusidic acid can significantly reduce the hemolysis activity and α-toxin production of S. aureus. In addition, the subinhibitory concentrations of fusidic acid significantly inhibited biofilm formation, autolysis, cell aggregation, and polysaccharide intercellular adhesin (PIA) production of S. aureus. Moreover, fusidic acid effectively reduces the damage of mouse skin lesion area. Furthermore, fusidic acid reduced the expression of the two-component regulatory system saeRS and staphylococcal accessory gene regulator (sarA). In conclusion, our results suggested that the subinhibitory concentrations of fusidic acid may reduce the virulence of S. aureus by down-regulating sarA and saeRS to reduce biofilm formation and α-toxin expression, which will provide a theoretical basis for the clinical treatment of S. aureus infection. This is the first report that fusidic acid has an inhibitory effect on the virulence of S. aureus, and this broadens the clinical application of fusidic acid.

Antimicrobial Susceptibility of Mycobacterium abscessus Complex Clinical Isolates from a Chinese Tertiary Hospital.

INTRODUCTION: Mycobacterium abscessus complex (MABC) is a group of important infectious agents that are highly associated with drug resistance, and antibiotic treatment is usually ineffective. This study investigated the characteristics of antimicrobial susceptibility of MABC isolates and the synergy between certain ß-lactam combinations against MABC infection. METHODS: We collected 129 MABC isolates from patients with lower respiratory tract infections and categorized them into three subspecies. The minimum inhibitory concentrations (MICs) of 15 antimicrobials for the MABC isolates were determined using commercial Sensititre RAPMYCOI MIC plates and the broth microdilution method, as recommended in the CLSI (M24-A2). In addition, the MICs of imipenem, alone and with ceftazidime and/or avibactam, were assessed in vitro for all isolates. The erm(41) and rrl genes were also sequenced. RESULTS: The MABC isolates exhibited >80% resistance to 11 of the 15 antimicrobials. Regarding the remaining four antimicrobials, the isolates were least resistant to tigecycline (12.4%) and amikacin (3.9%), and only partially resistant to two cefoxitin (39.5%) and imipenem (40.3%). Compared with M. massiliense isolates, M. abscessus and M. bolletii isolates were more resistant to amikacin and imipenem, whereas M. abscessus was significantly less resistant to tigecycline relative to M. massiliense and M. bolletii isolates. The clarithromycin inducible resistance rate was 68.4% and 74.3% among M. bolletii and M. abscessus isolates. Furthermore, 88.7% of the M. abscessus isolates carried a T at position 28 of erm(41), which is associated with inducible clarithromycin resistance. In addition, compared to imipenem with avibactam only, the MIC50 and MIC90values of imipenem after adding ceftazidime plus avibactam were decreased fourfold. CONCLUSION: The antimicrobial resistance rates and the characteristics of the erm(41) gene associated with inducible clarithromycin resistance were different among the three MABC subspecies. There was also synergy between imipenem and 100µg/mL ceftazidime against MABC isolates.

High Prevalence of 16S rRNA Methyltransferase Genes in Carbapenem-Resistant Klebsiella pneumoniae Clinical Isolates Associated with Bloodstream Infections in 11 Chinese Teaching Hospitals.

OBJECTIVE: The 16S rRNA methylase-mediated high-level resistance to aminoglycosides has become a great concern. The purpose of the study was to investigate the occurrence of 16S rRNA methyltransferase (RMTase) genes in carbapenem-resistant Klebsiella pneumoniae (CRKP) clinical isolates associated with bloodstream infections (BSIs) in China. METHODS: From July 2015 to December 2018, a total of 137 unique CRKP clinical isolates associated with BSIs were collected from 11 Chinese teaching hospitals. PCR and DNA sequencing were used to identify 16S RMTase genes. Whole-genome sequencing (WGS) was performed on all CRKP clinical isolates. Relevant information was extracted from WGS data (antibiotic resistance determinants, K-type and wzi allelic types). All 16S RMTase-producing CRKP clinical isolates were characterized by antimicrobial susceptibility testing, multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). RESULTS: In this study, 137 CRKPs were found to harbor at least one carbapenemase gene. Among 137 CRKPs, 78 (56.9%, 78/137) were positive for 16S RMTase genes (5 for armA, 70 for rmtB, 3 for both armA and rmtB) and highly resistant to gentamicin and amikacin (MICs ≥256 mg/L). Seventy-five isolates harboring 16S RMTase genes also produced ESBLs. In this study, 5 sequence types (STs) and 6 capsule serotypes were found among 78 isolates positive for 16S RMTases genes, while 14 STs and 6 capsule serotypes were found among 59 isolates negative for 16S RMTases genes. Compared with the isolates negative for 16S RMTases genes, the STs and capsular serotypes of 16S RMTases-positive strains are more concentrated. Among 78 16S RMTases-positive strains, the most prevalent clone type is ST11-PFGE-B-KL64-wzi64 (62.8%, 49/78), which mainly carries the rmtB and blaKPC genes and is distributed in 7 provinces in China. CONCLUSION: A high prevalence of 16S RMTase genes was found among CRKP clinical isolates associated with BSIs from Chinese teaching hospitals, which was attributed to the dissemination of the ST11-PFGE-B-KL64-wzi64 clone.

A Retrospective Analysis of Risk Factors and Patient Outcomes of Bloodstream Infection with Extended-Spectrum ß-Lactamase-Producing Escherichia coli in a Chinese Tertiary Hospital.

OBJECTIVE: The present study assessed risk factors and patient outcomes of bloodstream infection (BSI) caused by extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli (E. coli). METHODS: A retrospective study was performed to analyze risk factors and patient outcomes of BSI caused by extended-spectrum ß-lactamase-producing Escherichia coli (ESBL-EC) in one Chinese tertiary hospital over a 7.5-year period. The clinical characteristics of patients infected with ESBL-producing and non-ESBL-producing E. coli were compared. Predictors of 30-day mortality in patients with E. coli BSI were also identified in our study. RESULTS: The results of drug sensitivity showed that quinolones, aminoglycosides, ß-lactam/ß-lactamase inhibitor combinations (BLICs) and trimethoprim/sulfamethoxazole exhibited significant differences between the ESBL and non-ESBL groups. Of the 963 patients with E. coli BSI, 57.6% developed ESBL-EC. Multivariate analysis showed that biliary tract infection (BTI) [P<0.001,OR (95% CI):1.798 (1.334-2.425)], urinary tract obstructive disease [P=0.001,OR (95% CI):2.106 (1.366-3.248)], surgery within 3 months [P=0.002,OR (95% CI):1.591 (1.178-2.147)], hospitalization within 3 months [P<0.001,OR (95% CI):2.075 (1.579-2.725)], ICU admission [P=0.011,OR (95% CI):1.684 (1.124-2.522)] and history of cephalosporin use [P=0.006,OR (95% CI):3.097 (1.392-6.891)] were statistically significant. In mortality analysis, aCCI>2 [P=0.016,OR (95% CI): 2.453 (1.179-5.103)], gastrointestinal catheterization [P=0.004, OR (95% CI): 2.525 (1.333-4.782)] were significantly associated with 30-day mortality. According to Kaplan-Meier survival analysis, we found that in SOFA<2 group and SOFA≥2 group, the mortality rate of patients treated with BLICs were lower than that of carbapenems(P<0.05). CONCLUSION: This study showed that BTI, urinary tract obstructive disease, surgery within 3 months, hospitalization within 3 months, ICU admission and cephalosporin exposure were independent risk factors for the emergence of ESBL-EC BSI. Analysis of risk factors for 30-day mortality revealed that the factors independently associated with a higher risk of mortality were aCCI>2, gastrointestinal catheterization. Compared to carbapenems, the BLICs had preferable effect to treat patients with ESBL-EC BSI. Notably, patients with severe illness were inlcined to use carbapenems, which affected the analysis results. Therefore, we suggest that BLICs could be recommended to treat mild patients with ESBL-EC bacteremia.

Characterization of Pneumocystis jirovecii pneumonia at three tertiary comprehensive hospitals in southern China.

Due to the increasing use of immunosuppressant therapy, Pneumocystis jirovecii pneumonia (PJP) has become an emerging concern in human immunodeficiency virus (HIV)-negative patients. In this study, we conducted a retrospective study of 96 hospitalized patients with PJP from January 2015 to June 2019 at three tertiary comprehensive hospitals in Southern China. Information was collected regarding patient demographics, clinical manifestations, risk factors, laboratory analyses, radiological images, and treatment outcomes. PJP infection was most commonly found in middle-aged men. Kidney diseases (35.5%) and connective tissue diseases (38.7%) were the predominant risk factors for PJP. About half of the patients (48.4%) received glucocorticoid, immunosuppressant, and/or chemotherapy in a low dose or in a short-term (< 3 months). None of the patients had previously received trimethoprim-sulfamethoxazole (TMP-SMX) for PJP prophylaxis. All patients had two or more clinical manifestations (cough, dyspnea, fever, and chest pain). Biochemical investigations of CRP, ESR, PaO2, LDH, and KL-6 showed that over 90% of the patients exceeded the reference range of indicators. Our analyses revealed the dominant risk factors (HIV, kidney diseases, and connective tissue diseases) and the most consistent biochemical indicators (LDH, BG, and KL-6) for PJP. Moreover, early prophylaxis, diagnosis, and treatment should contribute to improve the survival of these PJP patients.

Proteome-Wide Identification of Lysine Propionylation in the Conidial and Mycelial Stages of Trichophyton rubrum.

Posttranslational modifications (PTMs) exist in a wide variety of organisms and play key roles in regulating various essential biological processes. Lysine propionylation is a newly discovered PTM that has rarely been identified in fungi. Trichophyton rubrum (T. rubrum) is one of the most common fungal pathogens in the world and has been studied as an important model organism of anthropic pathogenic filamentous fungi. In this study, we performed a proteome-wide propionylation analysis in the conidial and mycelial stages of T. rubrum. A total of 157 propionylated sites on 115 proteins were identified, and the high confidence of propionylation identification was validated by parallel reaction monitoring (PRM) assay. The results show that the propionylated proteins were mostly involved in various metabolic pathways. Histones and 15 pathogenicity-related proteins were also targets for propionylation modification, suggesting their roles in epigenetic regulation and pathogenicity. A comparison of the conidial and mycelial stages revealed that most propionylated proteins and sites were growth-stage specific and independent of protein abundance. Based on the function classifications, the propionylated proteins had a similar distribution in both stages; however, some differences were also identified. Furthermore, our results show that the concentration of propionyl-CoA had a significant influence on the propionylation level. In addition to the acetylation, succinylation and propionylation identified in T. rubrum, 26 other PTMs were also found to exist in this fungus. Overall, our study provides the first global propionylation profile of a pathogenic fungus. These results would be a foundation for further research on the regulation mechanism of propionylation in T. rubrum, which will enhance our understanding of the physiological features of T. rubrum and provide some clues for the exploration of improved therapies to treat this medically important fungus.

Coexistence of bla NDM-1 and rmtC on a Transferrable Plasmid of a Novel ST192 Klebsiella aerogenes Clinical Isolate.

INTRODUCTION: The occurrence and development of antibiotic resistance are mainly caused by the spread of large plasmids carrying multiple antibiotic resistance genes. Recently, the association between 16S rRNA methyltransferase genes and ß-lactamase genes carried by the same plasmid is of concern. METHODS: The Klebsiella aerogenes 1564 was isolated from the catheter tip of a patient in a tertiary hospital, Shanghai, China. The presence of the bla NDM-1 and rmtC genes were assessed by PCR. Complete sequence of plasmid p1564 was determined. The K. aerogenes 1564 was characterized by antimicrobial susceptibility testing, Carbapenemase phenotype confirmation testing, conjugation experiment, S1-PFGE and multilocus sequence typing (MLST). RESULTS: Herein, we found that a New Delhi Metallo-ß-lactamase-1 gene (bla NDM-1) and a 16S rRNA methyltransferase gene (rmtC) coexisted on a transferrable plasmid of a carbapenem-resistant K. aerogenes clinical isolate. The K. aerogenes clinical isolate was found to belong to a novel sequence type 192 (ST192) determined by MLST. The sequencing results of the plasmid p1564 carrying bla NDM-1 gene and rmtC gene showed that the size and guanine-cytosine content of the plasmid were 136, 902 bp and 51.8%, with 164 putative ORFs and two multidrug resistance gene islands. In addition to bla NDM-1and rmtC, the plasmid contained bleomycin resistance gene (ble MBL), CMY-6ß-lactamase gene (bla CMY-6), quaternary ammonium compound resistance gene (sugE), truncated quaternary ammonium compound resistance gene (qacEΔ1), aminoglycoside resistance gene (aacA4) and sulfonamide resistance gene (sul1). By comparison, p1564 has high homology with pHS36-NDM from Salmonella enterica subsp. enterica serovar Stanley reported in China, with similar size and both belonging to plasmid incompatibility group A/C. CONCLUSION: The present study demonstrated for the first time the co-existence of rmtC and bla NDM-1 in a novel ST192 K. aerogenes. The spread of plasmids harboring both bla NDM-1 and rmtC may occur among Enterobacteriaceae in China.