Detection of a Novel Metallo-β-Lactamase, CAM-2, in a Metagenome-Assembled Genome from China.
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ABSTRACT:
SUBMITTER: Xiang R
PROVIDER: S-EPMC9241868 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Intrinsic β-lactam resistance in Stenotrophomonas maltophilia is caused by bla L1 and/or bla L2, a kind of metallo-β-lactamase with a broad substrate spectrum including carbapenems. A rapid and sensitive molecular method for the detection of bla L1 in clinical samples is needed to guide therapeutic treatment. In present study, we first described a loop-mediated isothermal amplification (LAMP) method for the rapid detection of bla L1 in clinical samples by using two methods including a chromogenic method using calcein/Mn(2+) complex and the real-time turbidity monitoring to assess the reaction. Then dissemination of L1-producing S. maltophilia was investigated from ICU patients in three top hospital in Beijing, China. The results showed that both methods detected the target DNA within 60 min under isothermal conditions (65°C). The detection limit of LAMP was 3.79 pg/μl DNA, and its sensitivity 100-fold greater than that of conventional PCR. All 21 test strains except for S. maltophilia were negative for bla L1, indicative of the high-specificity of the primers for the bla L1. A total of 22 L1-positive isolates were identified for LAMP-based surveillance of bla L1 from 105 ICU patients with clinically suspected multi-resistant infections. The sequences of these bla L1 genes were conservative with only a few sites mutated, and the strains had highly resistant to β-lactam antibiotics. The MLST recovered that 22 strains belonged to seven different S. maltophilia sequence types (STs). Furthermore, co-occurrence of bla L1 and bla L2 genes were detected in all of isolates. Strikingly, S. maltophilia DCPS-01 was recovered to contain bla L1, bla L2, and bla NDM-1 genes, possessing an ability to hydrolyse all β-lactams antibiotics. Our data showed the diversity types of S. maltophilia carrying bla L1 and co-occurrence of many resistant genes in the clinical strains signal an ongoing and fast evolution of S. maltophilia resulting from their wide spread in the respiratory infections, and therefore will be difficult to control.
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Project description:Metallo-β-lactamases (MBLs) are present in major Gram-negative pathogens and environmental species, and pose great health risks because of their ability to hydrolyze the β-lactam rings of antibiotics such as carbapenems. PNGM-1 was the first reported case of a subclass B3 MBL protein that was identified from a metagenomic library from deep-sea sediments that predate the antibiotic era. In this study, PNGM-1 was overexpressed, purified and crystallized. Crystals of native and selenomethionine-substituted PNGM-1 diffracted to 2.10 and 2.30 Å resolution, respectively. Both the native and the selenomethionine-labelled PNGM-1 crystals belonged to the monoclinic space group P21, with unit-cell parameters a = 122, b = 83, c = 163 Å, β = 110°. Matthews coefficient (VM) calculations suggested the presence of 6-10 molecules in the asymmetric unit, corresponding to a solvent content of ∼31-58%. Structure determination is currently in progress.
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Project description:BACKGROUND:The increase in blaNDM-1 in Enterobacteriaceae has become a major concern worldwide. In previous study, we investigated clonal dissemination and mechanisms of resistance to carbapenem in China. METHODS:We carried out retrospective surveillance for blaNDM-1 among carbapenem-resistant enterobacter strains, which were isolated from patients at our hospital by bacterial strains selection, antimicrobial susceptibility testing, species identification, and molecular detection of resistance gene. RESULTS:We found three blaNDM-1 -positive isolates which were identified as Enterobacter aerogenes in clinical patients in China. The blaNDM-1 -positive Enterobacter aerogenes isolates were first found. CONCLUSION:It is important to mandate prudent usage of antibiotics and implement infection control measures to control the spread of these resistant blaNDM-1 -positive strains.
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Project description:Metallo β-Lactamases (MBLs) degrade most clinical β-lactam antibiotics, especially Carbapenem, posing a huge threat to global health. Studies on environmental MBLs are important for risk assessment of the MBLs transmission among connected habitats, and between environment and human. Here, we described a novel metallo β-Lactamases, named SZM-1 (Shenzhen metallo-β-lactamase), from an Arenimonas metagenome-assembled genome recovered from the river sediment in the Shenzhen Bay area, south China. Phylogenetic analysis, primary sequence comparison, structural modeling suggested that the SZM-1 belongs to B1 MBL family, likely harboring a typical di-zinc catalytic center. Furthermore, the gene encoding the MBLs was cloned into Escherichia coli TOP10 for Carba NP test and antimicrobial susceptibility test. The results indicated that the SZM-1 had carbapenemase activity, and conferred the carrier to increased resistance toward carbapenems. Taken together, our results raise alarms about the emergence and spread of the SZM-1, and suggest further surveillance, especially in hospital settings and clinical isolates, to determine whether bla SZM-1 is a mobilizable antibiotic resistance.
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Project description:A multidrug-resistant Vibrio alginolyticus isolate recovered from a shrimp sample with reduced carbapenem susceptibility produced a novel metallo-β-lactamase (MBL), VAM-1. That carbapenemase shared 67% to 70% amino acid identity with several VMB family subclass B1 MBLs, which were recently reported among some marine bacteria including Vibrio, Glaciecola, and Thalassomonas. The blaVAM-1 gene was located in a novel conjugative plasmid, namely, pC1579, and multiple copies of blaVAM-1 via an unusual mechanism of gene amplification were detected in pC1579. These findings underline the emergence of marine organisms acting as natural reservoirs for MBL genes and the importance of continuous bacterial antibiotic resistance surveillance.
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Project description:Being the second leading cause of death and the leading cause of disability-adjusted life years worldwide, infectious diseases remain-contrary to earlier predictions-a major consideration for the public health of the 21st century. Resistance development of microbes to antimicrobial drugs constitutes a large part of this devastating problem. The most widely spread mechanism of bacterial resistance operates through the degradation of existing β-lactam antibiotics. Inhibition of metallo-β-lactamases is expected to allow the continued use of existing antibiotics, whose applicability is becoming ever more limited. Herein, we describe the synthesis, the metallo-β-lactamase inhibition activity, the cytotoxicity studies, and the NMR spectroscopic determination of the protein binding site of phosphonamidate monoesters. The expression of single- and double-labeled NDM-1 and its backbone NMR assignment are also disclosed, providing helpful information for future development of NDM-1 inhibitors. We show phosphonamidates to have the potential to become a new generation of antibiotic therapeutics to combat metallo-β-lactamase-resistant bacteria.
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Project description:Wautersiella falsenii is a rarely non-fermenting Gram-negative bacterium and belongs to the Flavobacteriaceae family. This nosocomial pathogen can cause several human infections, especially among immunocompromised patients. Here, we describe the whole genome sequence of a clinical W. falsenii strain isolated from a urine sample of a 35-year-old woman with a urinary tract infection in Tunisia. We investigated its phenotype and genotype. After bacterial identification by the MALDI-TOF method, the whole-genome sequencing of this strain was performed. This isolate was not susceptible to various antibiotics, including β-lactams, aminoglycosides, and quinolones. However, it remains susceptible to imipenem (MIC = 0.25 mg/l), ertapenem (MIC = 0.75 mg/l), and meropenem (MIC = 0.19 mg/l). Interestingly, the E-TEST® (MP/MPI) showed a reduced MIC of meropenem +/- EDTA (0.064 μg/ml). Besides, the color change from yellow to red in the β CARBA test only after 24 hours of incubation can be interpreted in two ways. On the one hand, as a likely low expression of the gene encoding metallo-β-lactamase. On the other hand, and more likely, it may be a false-positive result because, according to the test manufacturer's recommendations, the test should be read after 30 minutes. Perhaps, therefore, this gene is not expressed in the tested strain. Moreover, the whole-genome sequence analysis demonstrated the presence of a novel chromosomally located subclass B1 metallo-β-lactamase EBR-like enzyme, sharing 94.92% amino acid identity with a previously described carbapenemase produced by Empedobacter brevis, EBR-1. The results also showed the detection of other antibiotic resistance genes and the absence of plasmids. So far, this study is the first report on the detection of W. falsenii in Tunisia. These findings prove that W. falsenii could be a potential reservoir of antibiotic resistance genes, e.g., β-lactamases. Collaborative efforts and effective hygiene measures should be established to prevent the emergence of this species in our health care settings.
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