Methicillin-resistant Staphylococcus aureus (MRSA) sequence type 45 (ST45) was rarely found in China. This study was conducted to trace the transmission and evolution of emerging MRSA ST45 strains in ...mainland China and explore its virulence. A total of 27 ST45 isolates were included for whole-genome sequencing and genetic characteristic analysis. Epidemiological results showed that MRSA ST45 isolates were often obtained from blood, primarily originated in Guangzhou, and carried diverse virulence and drug resistance genes. Staphylococcal cassette chromosome
type IV (SCC
IV) dominated in MRSA ST45 (23/27, 85.2%). ST45-SCC
V was located on a phylogenetic clade distinct from the SCC
IV cluster. We selected two representative isolates, MR370 (ST45-SCC
IV) and MR387 (ST45-SCC
V), and performed hemolysin activity, a blood killing assay, a Galleria mellonella infection model, and a mouse bacteremia model, as well as real-time fluorescence quantitative PCR. MR370 was proved to have extreme virulence in the phenotypic assays and at the mRNA level compared with ST59, ST5, and USA300 MRSA strains. MR387 was comparable to USA300-LAC on the phenotype and was verified to have higher expression of
,
,
,
,
, and
than USA300-LAC. The results emphasized the extraordinary performance of MR370 and the good potential of MR387 in virulence causing bloodstream infection. Meanwhile, we conclude that China MRSA ST45 showed two different clonotypes, which may be widespread in the future. The entire study is valuable as a timely reminder and reports virulence phenotypes of China MRSA ST45 for the first time.
Methicillin-resistant Staphylococcus aureus ST45 is epidemic worldwide. This study contributed to the awareness of the Chinese hyper-virulent MRSA ST45 strains and served as a timely reminder of its wide dissemination of clonotypes. Further, we provide novel insights for prevention from the perspective of bloodstream infections. ST45-SCC
V is a clonotype deserving special attention in China, and we performed genetic and phenotypic analyses for the first time on it.
The human fungal pathogen Candida glabrata is phylogenetically closely related to Saccharomyces cerevisiae, a model eukaryotic organism. Unlike S. cerevisiae, which has both haploid and diploid forms ...and a complete sexual cycle, C. glabrata has long been considered a haploid and asexual species. In this study, we analyzed the ploidy states of 500 clinical isolates of C. glabrata from four Chinese hospitals and found that approximately 4% of the isolates were in or able to spontaneously switch to an aneuploid (genomic DNA, 1N-2N), diploid (2N), or hyperdiploid (>2N) form under in vivo or in vitro conditions. Stable diploid cells were identified in 3% of the isolates (15/500). Of particular interest, one clinical strain existed only in the diploid form. Multilocus sequence typing (MLST) assays revealed two major genetic clusters (A and B) of C. glabrata isolates. Most of the isolates (70%) from China belonged to the A cluster, whereas most of the isolates from other countries (such as Iran, Japan, United States, and European countries) belonged to the B cluster. Further investigation indicated that C. glabrata cells of different ploidy forms differed in a number of respects, including morphologies, antifungal susceptibility, virulence, and global gene expression profiles. Additionally, C. glabrata could undergo spontaneous switching between the diploid and haploid forms under both in vitro and in vivo conditions. Given the absence of an apparent sexual phase, one would expect that the ploidy shifts could function as an alternative strategy that promotes genetic diversity and benefits the ability of the fungus to rapidly adapt to the changing environment. IMPORTANCE The human fungal pathogen Candida glabrata has long been thought to be a haploid organism. Here, we report the population structure and ploidy states of 500 clinical isolates of C. glabrata from China. To our surprise, we found that the ploidy of a subset of clinical isolates varied dramatically. Some isolates were in or able to switch to an aneuploid, diploid, or hyperdiploid form. C. glabrata cells with different ploidy differed in a number of biological respects, including morphology, antifungal susceptibility, virulence, and global gene expression profile. Given the absence of an apparent sexual phase in this fungus, we propose that ploidy switching could be a strategy for rapid adaptation to environmental changes and could function as an alternative to sexual reproduction.
Previous studies have shown that the increased prevalent ST764 clone in China, Japan, and other Asian areas. However, the knowledge of the genetic features and virulence characteristics of ...methicillin-resistant Staphylococcus aureus (MRSA) ST764 in China is still limited. In this study, we identified 52 ST764-SCCmec type II isolates collected from five cities in China between 2014 and 2021. Whole genome sequencing showed that the most common staphylococcal protein A (spa) types of ST764 in China were t002 (55.78%) and t1084 (40.38%). Virulence assays showed that ST764-t1084 isolates had high haemolytic activity and α-toxin levels. Of the critical regulatory factors affecting α-toxin production, only the SaeRS was highly expressed in ST764-t1084 isolates. Mouse abscess model indicated that the virulence of ST764-t1084 isolates was comparable to that of S. aureus USA300-LAC famous for its hypervirulence. Interestingly, ST764-t002 isolates exhibited stronger biofilm formation and cell adhesion capacities than ST764-t1084 isolates. This seems to explain why ST764-t002 subclone has become more prevalent in China in recent years. Phylogenetic analysis suggested that all ST764 isolates from China in Clade III were closely related to KUN1163 (an isolate from Japan). Notably, genomic analysis revealed that the 52 ST764 isolates did not carry arginine catabolic mobile element (ACME), which differed from ST764 isolates in Japan. Additionally, most ST764 isolates (69.23%) harboured an obvious deletion of approximately 5 kb in the SCCmec II cassette region compared to KUN1163. Our findings shed light on the potential global transmission and genotypic as well as phenotypic characteristics of ST764 lineage.
Xpert MTB/RIF (Xpert) is an automated molecular test recommended by World Health Organization (WHO) for diagnosis of tuberculosis (TB). This study evaluated the effect of Xpert implementation on the ...detection of pulmonary TB (PTB) and rifampicin-resistant TB (RR-TB) cases in Shanghai, China.
Xpert was routinely implemented in 2018 for all presumptive PTB patients. All PTB patients above 15 years-old identified within the Provincial TB Control Program during the first half of each of 2017 and 2018, were enrolled to compare the difference in proportions of bacteriological confirmation, patients with drug susceptibility test (DST) results for rifampicin (ie, DST coverage) and RR-TB detection before and after Xpert's implementation.
A total of 6047 PTB patients were included in the analysis with 1691 tested by Xpert in 2018. Percentages of bacteriological confirmation, DST coverage and RR-TB detection in 2017 and 2018 were 50% vs. 59%, 36% vs. 49% and 2% vs. 3%, respectively (all p-values < 0.05). Among 1103 PTB patients who completed sputum smear, culture and Xpert testing in 2018, Xpert detected an additional 121 (11%) PTB patients who were negative by smear and culture, but missed 248 (23%) smear and/or culture positive patients. Besides, it accounted for an increase of 9% in DST coverage and 1% in RR-TB detection. The median time from first visit to a TB hospital to RR-TB detection was 62 days (interquartile range -IQR 48-84.2) in 2017 vs. 9 days (IQR 2-45.7) in 2018 (p-value < 0.001). In the multivariate model, using Xpert was associated with decreased time to RR-TB detection (adjusted hazard ratio = 4.62, 95% confidence interval: 3.18-6.71).
Integrating Xpert with smear, culture and culture-based DST in a routine setting significantly increased bacteriological confirmation, DST coverage and RR-TB detection with a dramatic reduction in the time to RR-TB diagnosis in Shanghai, China. Our findings can be useful for other regions that attempt to integrate Xpert into routine PTB and RR-TB case-finding cascade. Further study should focus on the identification and elimination of operational level challenges to fully utilize the benefit of rapid diagnosis by Xpert.
To verify the efficacy and safety of an inexpensive standardized regimen for multidrug-resistant tuberculosis (MDR-TB) with low resistance to isoniazid (INH), a multicenter prospective study was ...conducted in eastern China.
Patients diagnosed as MDR-TB with low concentration INH resistance and rifampicin resistance, second-line/injectable agents sensitive were prospectively enrolled, given the regimen of Amikacin (Ak)-Fluoroquinolones (FQs)-Cycloserine (Cs)-Protionamide (Pto)-PasiniaZid (Pa)-Pyrazinamide (Z) for 6 months followed by 12 months of FQs-Cs-Pto-Pa-Z, and then followed up for treatment outcomes and adverse events (AEs).
A total of 114 patients were enrolled into the study. The overall favorable treatment rate was 79.8% (91/114). Among 91 cases with favorable treatment, 75.4% (86/114) were cured and 4.4% (5/114) were completed treatment. Regarding to unfavorable outcomes, among 23 cases, 8.8% (10/114) had failures, 8.8% (10/114) losing follow up, 0.9% (1/114) had treatment terminated due to intolerance to drugs and 1.8% (2/114) died. Treatment favorable rate was significantly higher in newly treated MDR-TB (91.7%, 33/36) than that in retreated MDR-TB (74.4%, 58/78, p 0.03). The investigators recorded 42 AEs occurrences in 30 of 114 patients (26.3%). Clinicians rated most AEs as mild or moderate (95.24%, 40/42).
The regimen was proved to be effective, safe and inexpensive. It is suitable for specific drug resistant population, especially for newly-treated patients, which could be expected to be developed into a short-course regimen. Clinical trials registration China Clinical Trial Registry ChiCTR-OPC-16009380.
Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) sequence type 9 (ST9) has emerged and disseminated in Asia. It is associated with colonization or infection in both humans ...and animal hosts; however, the genetic factors underpinning its adaptation to animal and human population remain to be determined. Here, we conducted a genomic analysis of 191 ST9 S. aureus genomes collected from 12 different countries, including 174 genomes retrieved from public databases and 17 sequenced in this study. In silico spa typing, staphylococcal cassette chromosome mec (SCCmec) typing, and antimicrobial resistance and virulence gene mining were conducted, and the temporal phylogenetic signal was assessed by Bayesian inference. Our results point toward a human methicillin-susceptible S. aureus (MSSA) origin of ST9 that evolved approximately 2 centuries ago. Three major genetic events occurred during ST9 host shift from human to animals: the loss of the immune evasion cluster genes (scn, chp, and sak), which were reported to contribute to virulence in human infections, the acquisition of the SaPIbov4-like element-encoding vwb gene, which is an animal-specific virulence factor responsible for the clotting of animal plasma, and the acquisition of antibiotic resistance genes, including SCCmec, quinolone resistance-determining region (QRDR) mutations, and a multidrug resistance genetic element (MDRST9). Evidence of direct transmission of animal-adapted strains to human hosts also suggest that transmission could potentially reshape the resistance and virulence genetic pool in these isolates. The rapid clonal expansion of MDR ST9 strains in mainland China and Taiwan highlights the increasing need for effective surveillance of antibiotic consumption in animal husbandry to control antimicrobial resistance spread. IMPORTANCE Staphylococcus aureus sequence type 9 (ST9) is the main LA-MRSA clone spreading in the Asian continent. It can colonize and cause mild to severe infections both in animal and humans. Previous work described its genotypic characteristics; however, the molecular history of global spread of ST9 strains remains largely unclear. We conducted a detailed analysis of genomic evolution of global ST9 strains and identified key genetic changes associated with its adaptation to specific hosts. Our results suggest that the ST9 clone originated from human-adapted strains, which lost genes related to the evasion of the immune system. The introduction of ST9 strains in animal populations was aligned with the acquisition of animal-specific virulent factors and mobile elements harboring multiple antimicrobial resistance genes, especially in isolates from mainland China and Taiwan.
•Bacteriological diagnosis of pulmonary tuberculosis is important but difficult.•We evaluated the Xpert Mycobacterium tuberculosis/rifampicin assay (MTB/RFI).•The assay was tested on bronchoalveolar ...lavage fluid (BALF).•The Xpert MTB/RIF assay on BALF adds microbiologic evidence to clinical decisions.•The ‘very low’ results of this assay may be false-positives.
To evaluate he diagnostic performance of the Xpert Mycobacterium tuberculosis/Rifampin (MTB/RIF) assay in bronchoalveolar lavage fluid (BALF).
We retrospectively reviewed the clinical data from 671 sputum-smear negative or sputum-scarce adult patients with suspected pulmonary tuberculosis (PTB) who had an Xpert MTB/RIF assay performed on BALF. The diagnostic performance of the Xpert MTB/RIF assay, smear microscopy (SM) and MTB culture was evaluated using MTB culture or final clinical diagnosis as the reference standard.
Compared with MTB culture, the sensitivity and specificity were 87.8% and 72.7% for the Xpert MTB/RIF assay and 11.0% and 99.2% for SM, respectively. Compared with final diagnosis, diagnostic performance was 58.9% and 83.9% for the Xpert MTB/RIF assay, 5.0% and 98.3% for SM, and 43.3% and 100% for culture, for sensitivity and specificity respectively. The Xpert MTB/RIF assay had low specificity and high sensitivity. When very low results were re-evaluated and considered MTB-negative, the specificity increased significantly. The sensitivity remained higher than SM and was similar to that of culture.
The Xpert MTB/RIF assay adds microbiologic evidence to clinical decisions; however, close attention should be paid to very low semi-quantitative positive results.
Methicillin-resistant Staphylococcus aureus (MRSA) ST8 strains have spread worldwide, causing outbreaks in various regions. However, this clone has only been sporadically reported in China. ...Consequently, detailed information regarding the phylogeny and potential virulence of S. aureus ST8 strains in China remains unknown. In this study, we characterized six ST8 strains collected from three tertiary hospitals in China, including three MRSA (MR50, MR526, and MR254) and three MSSA (H78, H849 and H863). Whole genome sequencing and phylogenetic analysis showed that the six strains formed two separate clusters, including two (MR50 and MR526) and four (MR254, H78, H849 and H863) isolates, respectively. Among them, MR50 and MR526 harboured spa t008, SCCmec IVa, arginine catabolic mobile element, and were phylogenetically close to the epidemic USA300 strains, while other four strains belonged to spa t9101 and formed a unique branch. MR254 carried a novel hybrid SCCmec element (namely SCCmec254). Same as the USA300 prototype strain LAC, the China S. aureus ST8 strains produced weak biofilms except MR254. Among them, MR254 had significantly stronger haemolysis ability and higher α-toxin levels than others, while MR526 showed comparable haemolysis and α-toxin production levels as USA300-LAC. In mouse skin abscess model, MR254 showed particularly strong invasions, accompanied by necrosis, while MR526 exhibited similar infection levels as USA300-LAC. These data suggested that the China MRSA ST8 isolates (e.g. MR254 and MR526) were highly virulent, displaying higher or similar virulence potential as the epidemic USA300 strain. Active surveillance should be enacted to closely monitor the further spread of these hyper-virulent MRSA strains in China.
Fluoroquinolones (FQ) are essential for the treatment of multidrug-resistant tuberculosis (MDR-TB). The FQ resistance (FQ-R) rate in MDR-TB in China and its risk factors remain poorly understood. We ...conducted a retrospective, population-based genomic epidemiology study of MDR-TB patients in Shanghai, China, from 2009 to 2018. A genomic cluster was defined as strains with genetic distances ≤ 12 single nucleotide polymorphisms. The transmitted FQ-R was defined as the same FQ resistance-conferring mutations shared by ≥ 2 strains in a genomic cluster. We used multivariable logistic regression analysis to identify the risk factors for drug resistance. Among the total 850 MDR-TB patients included in the study, 72.8% (619/850) were male, the median age was 39 (interquartile range 28, 55) years, 52.7% (448/850) were migrants, and 34.5% (293/850) were previously treated patients. Most of the MDR-TB strains belong to the Beijing lineage (91.7%, 779/850). Overall, the genotypic resistance rate of FQ was 34.7% (295/850), and 47.1% (139/295) FQ-R patients were in genomic clusters, of which 98 (33.2%, 98/295) were presumed as transmitted FQ-R. Patients with treatment-naïve (aOR = 1.84; 95% CI: 1.09, 3.16), diagnosed in a district-level hospital (aOR = 2.69; 95% CI: 1.56, 4.75), and streptomycin resistance (aOR = 3.69; 95% CI: 1.65, 9.42) were significantly associated with the transmission of FQ-R. In summary, the prevalence of FQ-R among MDR-TB patients was high in Shanghai, and at least one-third were transmitted. Enforced interventions including surveillance of FQ drug susceptibility testing and screening among MDR-TB before initiation of treatment were urgently needed.
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