Identification of toxic metal contamination in surface sediments of the Xiaoqing River under a long-term perspective (1996-2020): Risks, sources and driving factors.

The contamination of sediments by toxic metals poses a significant threat to both river ecosystems and human health. In this study, the geo-accumulation index (Igeo), biotoxicity evaluation method, and potential ecological risk index (RI) were employed to analyze the contamination level, biotoxicity risk, and potential ecological risk of toxic metals in surface sediments of the Xiaoqing River. To identify toxic metal sources, Spearman correlation and principal component analysis with multiple linear regression analysis (PCA-MLR) were employed. Additionally, redundancy analysis (RDA) was utilized to investigate potential driving factors affecting toxic metal accumulation in sediments. The results revealed that the levels of the five investigated metals (Cr, Pb, As, Hg, and Cd) showed constant fluctuations during the period 1996-2020. The midstream was found to be more polluted than the upstream and downstream. In the research area, Hg was identified as the primary contaminant with high levels of contamination, posing a biotoxicity risk and potential ecological risk. Pollution sources were identified for two periods: A (1996-2010) and B (2011-2020), with industrial, agricultural, traffic, and natural sources being the main contributors. During period A, industrial sources accounted for the highest proportion (40.8%), followed by agricultural sources (36.6%), and geological natural sources (22.6%). During period B, agricultural sources accounted for the highest proportion (42%), followed by industrial and traffic sources (32.4%), and geological natural sources (25.6%). The distribution of toxic metals in the basin was significantly influenced by water pH, sediment organic matter, population density, and per capita GDP. The study results provide fundamental data for preventing pollution and managing water resources contaminated with toxic metals in the sediments of the Xiaoqing River in Jinan. Additionally, it serves as a reference for analyzing related ecological and environmental issues in the basin.

First report of a clinical, multidrug-resistant Enterobacteriaceae isolate coharboring fosfomycin resistance gene fosA3 and carbapenemase gene blaKPC-2 on the same transposon, Tn1721.

In order to understand the genetic background and dissemination mechanism of carbapenem resistance and fosfomycin resistance in Enterobacteriaceae isolates, we studied a clinical Escherichia coli strain HS102707 isolate and an Enterobacter aerogenes strain HS112625 isolate, both of which were resistant to carbapenem and fosfomycin and positive for the bla(KPC-2) and fosA3 genes. In addition, a clinical Klebsiella pneumoniae strain HS092839 isolate which was resistant to carbapenem was also studied. A 70-kb plasmid was successfully transferred to recipient E. coli J53 by a conjugation test. PCR and Southern blot analysis showed that bla(KPC-2) was located on this plasmid. The complete sequence of pHS102707 showed that this plasmid belongs to the P11 subfamily (IncP1) and has a replication gene, several plasmid-stable genes, an intact type IV secretion system gene cluster, and a composite transposon Tn1721-Tn3 that harbored bla(KPC-2). Interestingly, a composite IS26 transposon carrying fosA3 was inserted in the Tn1721-tnpA gene in pHS102707 and pHS112625, leading to the disruption of Tn1721-tnpA and the deletion of Tn1721-tnpR. However, only IS26 with a truncated Tn21-tnpR was inserted in pHS092839 at the same position. To our knowledge, this is the first report of fosA3 and bla(KPC-2) colocated in the same Tn1721-Tn3-like composite transposon on a novel IncP group plasmid.

Contribution of ß-lactamases and porin proteins OmpK35 and OmpK36 to carbapenem resistance in clinical isolates of KPC-2-producing Klebsiella pneumoniae.

Fifty-seven carbapenem-resistant Klebsiella pneumoniae isolates belonging to ST11 (50 isolates), ST423 (5 isolates), and two other sequence types were studied. All were positive for blaKPC-2, blaTEM-1, and blaCTX-M-14. SDS-PAGE analysis of six representative isolates demonstrated varied porin expression. Nevertheless, when blaKPC-2 was deleted, carbapenem resistance was markedly reduced. Additionally, SHV-12, DHA-1, and/or VIM-1 appeared to contribute to accessory carbapenemase activity. In contrast, OmpK35 and/or OmpK36 deficiency seemed to serve only as a minor cooperative factor.

Mechanisms of linezolid resistance in staphylococci and enterococci isolated from two teaching hospitals in Shanghai, China.

BACKGROUND: Linezolid is one of the most effective treatments against Gram-positive pathogens. However, linezolid-resistant/intermediate strains have recently emerged in worldwide. The purpose of this study was to analyse the prevalence and resistance mechanisms of linezolid-resistant/intermediate staphylococci and enterococci in Shanghai, China. RESULTS: Thirty-two linezolid-resistant/intermediate strains, including 14 Staphylococcus capitis, three Staphylococcus aureus, 14 Enterococcus faecalis and one Enterococcus faecium clinical isolates, were collected in this study which displayed linezolid MICs of 8 to 512 µg/ml, 8-32 µg/ml, 4-8 µg/ml and 4 µg/ml, respectively. All linezolid-resistant S. capitis isolates had a novel C2131T mutation and a G2603T mutation in the 23S rRNA region, and some had a C316T (Arg106Cys) substitution in protein L4 and/or harboured cfr. Linezolid-resistant S. aureus isolates carried a C389G (Ala130Gly) substitution in protein L3, and/or harboured cfr. The cfr gene was flanked by two copies of the IS256-like element, with a downstream orf1 gene. Linezolid-resistant/intermediate enterococci lacked major resistance mechanisms. The semi-quantitative biofilm assay showed that 14 linezolid-resistant E. faecalis isolates produced a larger biofilm than linezolid-susceptible E. faecalis strains. Transmission electron microscopy showed the cell walls of linezolid-resistant/intermediate strains were thicker than linezolid-susceptible strains. CONCLUSION: Our data indicated that major resistance mechanisms, such as mutations in 23S rRNA and ribosomal proteins L3 and L4, along with cfr acquisition, played an important role in linezolid resistance. Secondary resistance mechanisms, such as biofilm formation and cell wall thickness, should also be taken into account.

Distribution, sources, and risk analysis of heavy metals in sediments of Xiaoqing River basin, Shandong province, China.

The accumulation of heavy metals in river sediment poses a major threat to ecological safety. The Xiaoqing River originates in western Jinan, with higher population density and per capita gross domestic product (GDP) in its basin compared to the Shandong province average. This study analyzed the spatial characteristics, ecological risk, human health risk, and contamination sources of heavy metals by collecting sediment samples from Xiaoqing River. We use the methods such as geo-accumulation index (Igeo), ecological risk assessment based on the interval number sorting method, and health risk assessment to evaluate the risk of heavy metals in sediments. The research finding suggests heavy metals including Pb, As, Ni, and Cr are low ecological risks, while Hg and Cd have reached high and extreme ecological risks. Correlation analysis and principal component analysis were used to analyze the correlation and sources of different heavy metals. The six heavy metals were categorized into three groups. Factor 1, comprising Hg, Cr, and Pb, was identified as a mixed source with a contribution rate of 37.76%. Factor 2 is an agricultural source and comprises Ni, Cd, and As with a contribution rate of 27.05%. Factor 3 includes Pb and Ni contributing to 15.30% as a natural source. This study offers valuable insights for the prevention of heavy metal pollution, as well as promoting sustainable urban development.

Clinical and Microbiological Characteristics of Endogenous Klebsiella pneumoniae Endophthalmitis in a Tertiary Hospital in Shanghai, China.

PURPOSE: To investigate the clinical features of endogenous Klebsiella pneumoniae endophthalmitis (EKPE) and the molecular characteristics of pathogenic K. pneumoniae. METHODS: A retrospective study was conducted between January 2014 and January 2021. Clinical data were extracted. K. pneumoniae isolates cultured from EKPE patients' specimens were characterized by antimicrobial susceptibility testing, multilocus sequence typing, capsular serotyping, and virulence gene profiles. Virulence phenotypes were confirmed by mouse lethality assay. RESULTS: Thirty-nine patients (47 eyes) were diagnosed with EKPE. Administration of systemic antibiotics and eye treatment within 24 hours of ocular symptom onset was statistically associated with better visual outcomes (p = .007). Sixteen K. pneumoniae isolates were collected, three of which were carbapenem-resistant and avirulent. Only salmochelin encoding gene was harbored by all isolates. CONCLUSIONS: Systemic and eye treatment administered within 24 hours from the onset improves visual prognosis. Besides hypervirulent strains, avirulent carbapenem-resistant K. pneumoniae can cause EKPE.

Characteristics and prognostic risk factors of patients with sequence type 5 lineage-associated cryptococcosis in China.

OBJECTIVES: Cryptococcus neoformans sequence type 5 (ST5) lineage could infect immunocompetent hosts and cause a significant medical burden. We sought to identify characteristics and prognostic risk factors of ST5 lineage-associated cryptococcosis. METHODS: Multilocus sequence typing and antifungal susceptibility testing were conducted for Cryptococcus isolates. The clinical and laboratory characteristics of cryptococcosis patients were investigated. The multivariable logistic regression identified variables independently associated with 30-day mortality in patients with ST5 lineage-associated cryptococcosis without HIV. RESULTS: The infection rate of the ST5 isolates was 89.4% (370/414) in China. The proportion of ST5 isolates with nonwild-type minimum inhibitory concentrations to amphotericin B, 5-flucytosine, voriconazole, posaconazole, itraconazole, and fluconazole were 0%, 5.4%, 0.3%, 1.4%, 0.3%, and 8.1%, respectively. The ST5 lineage-infected group exhibited significantly higher blood platelet count, lower blood cryptococcal antigen (CrAg) titer, lower cerebrospinal fluid (CSF) CrAg titer than the non-ST5 lineage-infected group, and lower hemoglobin and lower CSF CrAg titer than the Cryptococcus gattii isolates-infected group. Seven baseline parameters, including underlying disease, dyskinesia, anemia, high peripheral blood neutrophils, low platelet count, high CSF fungal burden, and high CSF opening pressure, were associated independently with the 30-day mortality of patients with ST5 lineage-associated cryptococcosis without HIV. CONCLUSION: Our study has provided an understanding of the ST5 lineage associated with cryptococcosis.

A LAMP-based microfluidic module for rapid detection of pathogen in cryptococcal meningitis.

Cryptococcal meningitis (CM) is a global threat with significant attributable morbidity and mortality. Information on microfluidic detection for CM diagnosis is still limited. We developed a multifunctional microfluidic module that integrated the pathogen enrichment and on-chip nucleic acid extraction. The module adopted a simple filtration membrane to effectively capture Cryptococcus cells and simplify the process, and combined lyticase digestion, alkaline lysis and heating methods to optimize the strategy to achieve nucleic acid extraction. The entire process was operated in the module, which reduced the exposure risk of directly processing cryptococcal samples. A portable one-pot lyophilized LAMP reagent bead with no temperature limit was developed, which improved the flexibility of operation. This module did not require any additional instrument, and is promising to develop a simple, rapid, and efficient approach to realize the "sample in and answer out" detection of real CSF samples. This microfluidic module had practical prospects and is expected to replace LFA for efficacy evaluation and follow-up in the middle and late stages of CM treatment, and could be used as an auxiliary method to confirm cases with questionable LFA results in the early diagnosis of CM.

KlcAHS genes are ubiquitous in clinical, blaKPC-2-positive, Klebsiella pneumoniae isolates.

Carbapenemase-producing Klebsiella pneumoniae has emerged and spread widely throughout the world. The mechanisms involved remain unclear. To provide insight, five plasmids were obtained from carbapenemase-producing K. pneumoniae clinical isolates. The five sequences were acquired, aligned and analyzed. In addition to the blaKPC-2 gene, which encodes beta lactamase, essentially all the plasmids contained a putative anti-restriction protein-encoding gene, KlcAHS. The KlcAHS gene was found in 98.2% of the blaKPC-2-positive, imipenem-resistant K. pneumoniae clinical isolates and in <1% of the blaKPC-2-negative control group. A searched of the GenBank database indicated that KlcAHS was mainly submitted by Chinese investigators beginning in 2010. Seventeen different KlcA amino acid sequences were found in the database using the restricting words: KlcA and Klebsiella pneumoniae. These sequences were used to generate a phylogenetic tree via MEGA6 software, revealing a distant evolutionary relationship between KlcAHS and other KlcAs. The secondary structure of KlcAHS, predicted with PROMALS3D software, exhibited highly conserved α-helices and ß-strands. KlcAHS expressed anti-restriction activity in vivo. In summary, KlcAHS genes are ubiquitous in blaKPC-2-positive Klebsiella pneumoniae clinical isolates collected at Huashan Hospital, China. The KlcAHS protein possesses a secondary structure similar to that exhibited by anti-restriction proteins and displays anti-restriction activity. As such, KlcAHS is a probable factor in the accelerated spread of blaKPC-2 and carbapenem-resistance among clinical, K. pneumoniae isolates.

MicroRNA-200a inhibits cell growth and metastasis by targeting Foxa2 in hepatocellular carcinoma.

Background: MicroRNAs (miRNAs) are a class of endogenous, small non-coding RNAs which function as essential posttranscriptional modulators of gene expression tightly involved in a wide range of diseases, including the hepatocellular carcinoma (HCC). Here, the present study was designed to investigate the expression levels and cellular roles of miR-200a in HCC. Methods: Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was used to detect the expression levels of miR-200a in serums and cell lines. Bioinformation analysis, the luciferase reporter assay, qRT-PCR and western blotting were employed to validate Foxa2 as a direct target gene of miR-200a. Cell proliferation, migration and invasion were assessed to identify whether miR-200a could regulate the biological behaviors of HCC cells by targeting Foxa2. Results: In this study, a low level of miR-200a was observed in patients' serums and HCC cell lines. Overexpression of miR-200a in HCC cell lines reduced cell proliferation, migration and invasion. In addition, transcription factor forkhead box A2 (Foxa2) was identified as a novel target of miR-200a and downregulated at mRNA and protein levels in miR-200a overexpressed cells. Meanwhile, restoration of Foxa2 significantly reversed the tumor suppressive effects of miR-200a. Conclusions: These findings indicate that miR-200a regulates the proliferation, migration and invasion of HCC cells by targeting Foxa2, suggesting that miR-200a may function as a potential therapeutic molecular for the diagnosis and treatment of the liver cancer.

Rapid Identification and Multiple Susceptibility Testing of Pathogens from Positive-Culture Sterile Body Fluids by a Combined MALDI-TOF Mass Spectrometry and Vitek Susceptibility System.

Infections of the bloodstream, central nervous system, peritoneum, joints, and other sterile areas are associated with high morbidity and sequelae risk. Timely initiation of effective antimicrobial therapy is crucial to improving patient prognosis. However, standard final identification and antimicrobial susceptibility tests (ASTs) are reported 16-48 h after a positive alert. For a rapid, effective and low-cost diagnosis, we combined matrix-assisted laser desorption/ionization time of flight mass spectrometry with a Vitek AST system, and performed rapid microbial identification (RMI) and rapid multiple AST (RMAST) on non-duplicated positive body fluid cultures collected from a hospital in Shanghai, China. Sterile body fluid positive culture and blood positive culture caused by Gram negative (GN) or polymicrobial were applied to the MALDI-TOF measurement directly. When positive blood culture caused by Gram positive (GP) bacteria or yeasts, they were resuspended in 1 ml brain heart infusion for 2 or 4 h enrichment, respectively. Regardless of enrichment, the RMI (completed in 40 min per sample) accurately identified GN and GP bacteria (98.9 and 87.2%, respectively), fungi (75.7%), and anaerobes (94.7%). Dominant species in multiple cultures and bacteria that failed to grow on the routing plates were correctly identified in 81.2 and 100% of cases, respectively. The category agreements of RMAST results, determined in the presence of various antibiotics, were similarly to previous studies. The RMI and RMAST results not only reduce the turnaround time of the patient report by 18-36 h, but also indicate whether a patient's antibiotic treatment should be accelerated, ceased or de-escalated, and adjusted the essential drugs modification for an optimized therapy.

MRSA epidemic linked to a quickly spreading colonization and virulence determinant.

The molecular processes underlying epidemic waves of methicillin-resistant Staphylococcus aureus (MRSA) infection are poorly understood(1). Although a major role has been attributed to the acquisition of virulence determinants by horizontal gene transfer(2), there are insufficient epidemiological and functional data supporting that concept. We here report the spread of clones containing a previously extremely rare(3,4) mobile genetic element­encoded gene, sasX. We demonstrate that sasX has a key role in MRSA colonization and pathogenesis, substantially enhancing nasal colonization, lung disease and abscess formation and promoting mechanisms of immune evasion. Moreover, we observed the recent spread of sasX from sequence type 239 (ST239) to invasive clones belonging to other sequence types. Our study identifies sasX as a quickly spreading crucial determinant of MRSA pathogenic success and a promising target for therapeutic interference. Our results provide proof of principle that horizontal gene transfer of key virulence determinants drives MRSA epidemic waves.