Green-synthesized zinc oxide nanoparticles by Enterobacter sp.: unveiling characterization, antimicrobial potency, and alleviation of copper stress in Vicia faba (L.) plants.

BACKGROUND: The biosynthesis of zinc oxide nanoparticles (ZnO NPs) using Enterobacter sp. and the evaluation of their antimicrobial and copper stress (Cu+ 2)-reducing capabilities in Vicia faba (L.) plants. The green-synthesized ZnO NPs were validated using X-ray powder diffraction (XRD); Fourier transformed infrared (FTIR), Ultraviolet-Visible spectroscopy (UV-Vis), Transmission electron microscope (TEM) and scanning electron microscopy (SEM) techniques. ZnO NPs could serve as an improved bactericidal agent for various biological applications. as well as these nanoparticles used in alleviating the hazardous effects of copper stress on the morphological and physiological traits of 21-day-old Vicia faba (L.) plants. RESULTS: The results revealed that different concentrations of ZnO NPs (250, 500, or 1000 mg L-1) significantly alleviated the toxic effects of copper stress (100 mM CuSO4) and increased the growth parameters, photosynthetic efficiency (Fv/Fm), and pigments (Chlorophyll a and b) contents in Cu-stressed Vicia faba (L.) seedlings. Furthermore, applying high concentration of ZnO NPs (1000 mg L-1) was the best dose in maintaining the levels of antioxidant enzymes (CAT, SOD, and POX), total soluble carbohydrates, total soluble proteins, phenolic and flavonoid in all Cu-stressed Vicia faba (L.) seedlings. Additionally, contents of Malondialdehyde (MDA) and hydrogen peroxide (H2O2) were significantly suppressed in response to high concentrations of ZnO NPs (1000 mg L-1) in all Cu-stressed Vicia faba (L.) seedlings. Also, it demonstrates strong antibacterial action (0.9 mg/ml) against various pathogenic microorganisms. CONCLUSIONS: The ZnO NPs produced in this study demonstrated the potential to enhance plant detoxification and tolerance mechanisms, enabling plants to better cope with environmental stress. Furthermore, these nanoparticles could serve as an improved bactericidal agent for various biological applications.

Mitigating sediment cadmium contamination through combining PGPR Enterobacter ludwigii with the submerged macrophyte Vallisneria natans.

Sediment cadmium contamination poses risks to aquatic ecosystems. Phytoremediation is an environmentally sustainable method to mitigate cadmium contamination. Submerged macrophytes are affected by cadmium stress, but plant growth-promoting rhizobacteria (PGPR) can restore the health status of submerged macrophytes. Herein, we aimed to reduce sediment cadmium concentration and reveal the mechanism by which the combined application of the PGPR Enterobacter ludwigii and the submerged macrophyte Vallisneria natans mitigates cadmium contamination. Sediment cadmium concentration decreased by 21.59% after submerged macrophytes were planted with PGPR, probably because the PGPR colonized the rhizosphere and roots of the macrophytes. The PGPR induced a 5.09-fold increase in submerged macrophyte biomass and enhanced plant antioxidant response to cadmium stress, as demonstrated by decreases in oxidative product levels (reactive oxygen species and malondialdehyde), which corresponded to shift in rhizosphere metabolism, notably in antioxidant defence systems (i.e., the peroxidation of linoleic acid into 9-hydroperoxy-10E,12Z-octadecadienoic acid) and in some amino acid metabolism pathways (i.e., arginine and proline). Additionally, PGPR mineralized carbon in the sediment to promote submerged macrophyte growth. Overall, PGPR mitigated sediment cadmium accumulation via a synergistic plantmicrobe mechanism. This work revealed the mechanism by which PGPR and submerged macrophytes control cadmium concentration in contaminated sediment.

High prevalence of multidrug-resistant Enterobacterales carrying extended-spectrum beta-lactamase and AmpC genes isolated from neonatal sepsis in Ahvaz, Iran.

OBJECTIVES: In the recent years, multidrug resistant (MDR) neonatal septicemia-causing Enterobacterales has been dramatically increased due to the extended-spectrum beta-lactamases (ESBLs) and AmpC enzymes. This study aimed to assess the antibiotic resistance pattern, prevalence of ESBLs/AmpC beta-lactamase genes, and Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) fingerprints in Enterobacterales isolated from neonatal sepsis. RESULTS: In total, 59 Enterobacterales isolates including 41 (69.5%) Enterobacter species, 15 (25.4%) Klebsiella pneumoniae and 3 (5.1%) Escherichia coli were isolated respectively. Resistance to ceftazidime and cefotaxime was seen in all of isolates. Furthermore, all of them were multidrug-resistant (resistant to three different antibiotic categories). The phenotypic tests showed that 100% of isolates were ESBL-positive. Moreover, AmpC production was observed in 84.7% (n = 50/59) of isolates. Among 59 ESBL-positive isolates, the highest percentage belonged to blaCTX-M-15 gene (66.1%) followed by blaCTX-M (45.8%), blaCTX-M-14 (30.5%), blaSHV (28.8%), and blaTEM (13.6%). The frequency of blaDHA, blaEBC, blaMOX and blaCIT genes were 24%, 24%, 4%, and 2% respectively. ERIC-PCR analysis revealed that Enterobacterales isolates were genetically diverse. The remarkable prevalence of MDR Enterobacterales isolates carrying ESBL and AmpC beta-lactamase genes emphasizes that efficient surveillance measures are essential to avoid the more expansion of drug resistance amongst isolates.

Antimicrobial Lasso Peptide Cloacaenodin Utilizes a Unique TonB-Dependent Transporter to Access Susceptible Bacteria.

The development of new antimicrobial agents effective against Gram-negative bacteria remains a major challenge in drug discovery. The lasso peptide cloacaenodin has potent antimicrobial activity against multiple strains in the Enterobacter genus, one of the ESKAPE pathogens. Here, we show that cloacaenodin uses a previously uncharacterized TonB-dependent transporter, which we name CloU, to cross the outer membrane (OM) of susceptible bacteria. Inner membrane transport is mediated by the protein SbmA. CloU is distinct from the known OM transporters (FhuA and PupB) utilized by other antimicrobial lasso peptides and thus offers important insight into the spectrum of activity of cloacaenodin. Using knowledge of the transport pathway to predict other cloacaenodin-susceptible strains, we demonstrate the activity of cloacaenodin against clinical isolates of Enterobacter and of a Kluyvera strain. Further, we use molecular dynamics simulations and mutagenesis of CloU to explain the variation in cloacaenodin susceptibility observed across different strains of Enterobacter. This work expands the currently limited understanding of lasso peptide uptake and advances the potential of cloacaenodin as an antibiotic.

Genetic characterization of KHM-1 metallo-ß-lactamase-producing Enterobacterales isolates from inpatient sources in Osaka, Japan.

OBJECTIVES: KHM-1-metallo-ß-lactamase-producing Enterobacterales strains, of which only a few have been found, were isolated from four inpatients in Osaka, Japan during 2016 to 2020. We compared whole genomes of the four KHM-1-producing isolates, including one Enterobacter hormaechei subsp. hoffmannii, one Escherichia coli, and two Citrobacter freundii. METHODS: These isolates were characterized by whole-genome sequencing, comparative analysis of blaKHM-1-encoding plasmids with earlier reported plasmids, and antimicrobial susceptibility tests. RESULTS: Multilocus sequence typing classified the E. hormaechei subsp. hoffmannii isolate to ST78, the E. coli isolate to ST354, and the two C. freundii isolates to ST95. These isolates harboured various antimicrobial resistance genes aside from blaKHM-1 on their chromosomes and plasmids. In all four isolates, blaKHM-1 was located on 137 kbp to 213 kbp plasmids of IncC replicon type. Although there were common resistance genes such as blaKHM-1-ISEc68, class I integron cassette, and fosG, the four blaKHM-1-encoding plasmids were distinguishable into two lineages based on differences of the resistance gene components and their surrounding regions. CONCLUSION: Because no epidemiological contact was observed among the inpatients, the blaKHM-1-encoding IncC plasmids might have spread horizontally to multiple bacterial species through repeated recombination and insertion.

Drug efflux and lipid A modification by 4-L-aminoarabinose are key mechanisms of polymyxin B resistance in the sepsis pathogen Enterobacter bugandensis.

OBJECTIVES: A concern with the ESKAPE pathogen, Enterobacter bugandensis, and other species of the Enterobacter cloacae complex, is the frequent appearance of multidrug resistance against last-resort antibiotics, such as polymyxins. METHODS: Here, we investigated the responses to polymyxin B (PMB) in two PMB-resistant E. bugandensis clinical isolates by global transcriptomics and deletion mutagenesis. RESULTS: In both isolates, the genes of the CrrAB-regulated operon, including crrC and kexD, displayed the highest levels of upregulation in response to PMB. ∆crrC and ∆kexD mutants became highly susceptible to PMB and lost the heteroresistant phenotype. Conversely, heterologous expression of CrrC and KexD proteins increased PMB resistance in a sensitive Enterobacter ludwigii clinical isolate and in the Escherichia coli K12 strain, W3110. The efflux pump, AcrABTolC, and the two component regulators, PhoPQ and CrrAB, also contributed to PMB resistance and heteroresistance. Additionally, the lipid A modification with 4-L-aminoarabinose (L-Ara4N), mediated by the arnBCADTEF operon, was critical to determine PMB resistance. Biochemical experiments, supported by mass spectrometry and structural modelling, indicated that CrrC is an inner membrane protein that interacts with the membrane domain of the KexD pump. Similar interactions were modeled for AcrB and AcrD efflux pumps. CONCLUSION: Our results support a model where drug efflux potentiated by CrrC interaction with membrane domains of major efflux pumps combined with resistance to PMB entry by the L-Ara4N lipid A modification, under the control of PhoPQ and CrrAB, confers the bacterium high-level resistance and heteroresistance to PMB.

A novel F type plasmid encoding mcr-10 in a clinical Enterobacter ludwigii strain from a tertiary hospital in the Czech Republic.

OBJECTIVE: Here we describe a novel IncFIA plasmid harbouring mcr-10 gene in a clinical Enterobacter ludwigii strain isolated at the University Hospital in Pilsen in the Czech Republic. METHODS: The strain was subjected to antibiotic susceptibility testing. Whole genome sequencing was performed using Illumina for short-read sequencing and Oxford Nanopore Technologies for long-read sequencing followed by hybrid assembly. The resulting genome was used to detect species using average nucleotide identity, resistance genes, plasmid replicon and MLST (using centre for genomic epidemiology databases; ResFinder, PlasmidFinder and MLST, respectively) and virulence genes using VFDB. RESULTS: Τhe strain showed susceptibility against tetracycline, cefuroxime and chloramphenicol, and it was susceptible to the second and third generation of cephalosporins, carbapenems and colistin. Genome analysis identified the strain as E. ludwigii sequence type ST20 and located the mcr-10 gene on an IncFIA (HI1)/IncFII (Yp) plasmid (pI9455333_MCR10; 129 863 bp). Upon blasting the nucleotide sequence of pI9455333_MCR10 against the NCBI database, no similar plasmid sequence was detected, implying a novel plasmid structure. Nevertheless, it showed a partial similarity with pRHBSTW-00123_3 and FDAARGOS 1432, which were detected in Enterobacter cloacae complex (ECC) strains in wastewater samples in 2017 in UK and in 2021 in the United States, respectively, and pEC81-mcr, which was detected in a clinical Escherichia coli strain in 2020 in China. Moreover, I9455333cz genome carried virulence genes coding for curli fibers, fimbrial adherence determinants, siderophore aerobactin, iron uptake proteins and regulators of sigma factor. CONCLUSION: In conclusion, we identified a novel IncF plasmid harbouring mcr-10 gene in a clinical Enterobacter ludwigii strain. To our knowledge, this is the first clinical report of mcr-10 in the Czech Republic.

Dramatic increase in antimicrobial resistance in ESKAPE clinical isolates over the 2010-2020 decade in India.

RATIONALE AND OBJECTIVES: ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) constitute a threat to humans worldwide. India is now the most populous country. The goal was to investigate the evolution of the rates of antimicrobial resistance in ESKAPE pathogens across India over the 2010-20 decade. METHODS: The data (89 studies) were retrieved from the Medline PubMed repository using specific keywords. RESULTS: The study of 20 177 ESKAPE isolates showed that A. baumannii isolates were the most represented (35.9%, n = 7238), followed by P. aeruginosa (25.3%, n = 5113), K. pneumoniae (19.5%, n = 3934), S. aureus (16.3%, n = 3286), E. faecium (2.6%, n = 517) and Enterobacter spp. (0.4%, n = 89). A notable increase in the resistance rates to antimicrobial agents occurred over the 2010-20 decade. The most important levels of resistance were observed in 2016-20 for A. baumannii (90% of resistance to the amoxicillin-clavulanate combination) and K. pneumoniae (81.6% of resistance to gentamycin). The rise in ß-lactamase activities was correlated with an increase in the positivity of Gram-negative isolates for ß-lactamase genes. CONCLUSIONS: This review highlighted that, in contrast to developed countries that kept resistance levels under control, a considerable increase in resistance to various classes of antibiotics occurred in ESKAPE pathogens in India over the 2010-2020 decade.

Antimicrobial Resistance Genes, Virulence Genes, and Associated Mobile Genetic Elements of Eight Multidrug-Resistant Enterobacterales Isolated from Hospital-Acquired Urinary Tract Infections in Sri Lanka.

The study describes the first isolation of multidrug-resistant (MDR) Klebsiella pneumoniae ST16, Escherichia coli ST131 (Esc), and Enterobacter hormaechei subsp. steigerwaltii ST93 (Enterobacter cloacae complex [ECC]) in Sri Lanka. Eight MDR strains of uropathogenic Enterobacterales isolated from hospital acquired urinary tract infections (UTIs) were analyzed using genomic sequencing and comparative genomics. Isolates carried multiple carbapenemase, AmpC, and ESBL (extended-spectrum ß-lactamase) genes. ECC manifested both blaNDM-4 and blaOXA-181. The K. pneumoniae strains harbored fimbrial genes that facilitate pathogenesis of UTI. Several extraintestinal pathogenic E. coli associated virulence genes were identified in Esc. The efflux pump gene, acrA, and the T6SS gene cluster were detected in ECC. Many antimicrobial resistance (AMR) and virulence genes were identified associated with mobile genetic elements. ISEcp1 flanked upstream of blaCTX-M-15. The carbapenemase genes were carried on ColKP3 plasmids and were associated with ISEcp1. In Esc, the AMR gene blaTEM-1B and virulence gene traT were found on an IncF plasmid replicon. In K. pneumoniae the AMR genes sul1 and tetB present on IncR plasmid replicons and were associated with the insertion sequence IS6100. In Kp5, blaLAP-2 and qnrS1 coexisted and were flanked by ISEcl. AMR gene clusters, conferring resistance to multiple antimicrobial classes, flanked by mobile elements were identified in seven isolates.

A Novel Lipid-Based MALDI-TOF Assay for the Rapid Detection of Colistin-Resistant Enterobacter Species.

Enterobacter species are classified as high-priority pathogens due to high prevalence of multidrug resistance from persistent antibiotic use. For Enterobacter infections caused by multidrug-resistant isolates, colistin (polymyxin E), a last-resort antibiotic, is a potential treatment option. Treatment with colistin has been shown to lead to emergence of polymyxin resistance. The primary mechanism for colistin resistance is modification of terminal phosphate moieties of lipid A, leading to decreased membrane electronegativity and reducing colistin binding affinity. Detection of these modifications, including the addition of phosphoethanolamine and 4-amino-4-deoxy-l-arabinose (Ara4N), can be used for prediction of colistin resistance using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). The objective of this study was to identify lipid A markers for colistin resistance in Enterobacter species and Klebsiella aerogenes (formerly Enterobacter aerogenes). Using a collection of Enterobacter and Klebsiella aerogenes clinical isolates, broth MICs for colistin were determined initially. Subsequently, killing assays were carried out to determine how the concentration of colistin at which there is approximately 50% survival (kill50) equates to their MICs. Finally, lipid A analysis was conducted via MALDI-TOF MS using the novel rapid extraction method, termed fast lipid analysis technique (FLAT), to correlate MIC and killing efficacy with predictive lipid A modifications. Sensitivity and specificity of the MS assay compared to MIC interpretation were 100% and 53.4%, respectively. A receiver operator characteristic (ROC) demonstrated that MS was highly correlated with killing, with area under the curve of 0.97. This analysis demonstrated the potential utility of MALDI-TOF MS as a rapid diagnostic platform of colistin resistance in Enterobacter species. IMPORTANCE In this study, we develop a novel method for identifying colistin resistance in Enterobacter species and Klebsiella aerogenes without performing antimicrobial susceptibility testing. Typically, susceptibility testing requires an additional 24 to 48 h, while the MS assay described in this study allows for resistant identifications in under 1 h after initial culture. Identification using MALDI-TOF MS would save time and prevent inappropriate use of colistin. MALDI-TOF MS is an easy-to-use, readily available, robust diagnostic tool in clinical laboratories. Furthermore, this study highlights limitations of polymyxin susceptibility testing. Use of a killing assay best captures how colistin treats infection and is shown to be highly correlated with our MS assay; thus, the MS assay in this study effectively predicts how colistin would treat a patient's infection. Use of MALDI-TOF MS for accurate and early identification of antimicrobial resistance can improve antimicrobial stewardship and patient outcomes.

Human Amnion-Derived Mesenchymal Stromal Cells: A New Potential Treatment for Carbapenem-Resistant Enterobacterales in Decompensated Cirrhosis.

BACKGROUND: Spontaneous bacterial peritonitis (SBP) is a severe and often fatal infection in patients with decompensated cirrhosis and ascites. The only cure for SBP is antibiotic therapy, but the emerging problem of bacterial resistance requires novel therapeutic strategies. Human amniotic mesenchymal stromal cells (hA-MSCs) possess immunomodulatory and anti-inflammatory properties that can be harnessed as a therapy in such a context. METHODS: An in vitro applications of hA-MSCs in ascitic fluid (AF) of cirrhotic patients, subsequently infected with carbapenem-resistant Enterobacterales, was performed. We evaluated the effects of hA-MSCs on bacterial load, innate immunity factors, and macrophage phenotypic expression. RESULTS: hA-MSCs added to AF significantly reduce the proliferation of both bacterial strains at 24 h and diversely affect M1 and M2 polarization, C3a complement protein, and ficolin 3 concentrations during the course of infection, in a bacterial strain-dependent fashion. CONCLUSION: This study shows the potential usefulness of hA-MSC in treating ascites infected with carbapenem-resistant bacteria and lays the foundation to further investigate antibacterial and anti-inflammatory roles of hA-MSC in in vivo models.

Analysis of an IncR Plasmid Carrying blaNDM-1 Linked to an Azithromycin Resistance Region in Enterobacter hormaechei Involved in an Outbreak in Quebec.

In the context of a recent rise in prevalence of NDM-encoding carbapenemase-producing Enterobacterales (CPE) in the province of QC, Canada, the genetic environment of blaNDM-1 was investigated. Three NDM-producing clinical isolates of Enterobacter hormaechei recovered from hospitalized patients involved in a putative outbreak were further characterized by whole-genome sequencing (WGS). Two isolates were confirmed by pulsed-field gel electrophoresis and WGS to be closely related. In addition to a ∼128 kb IncFII conjugative multidrug-resistance (MDR) plasmid, these isolates possessed a ∼45 kb mobilizable IncR MDR plasmid containing 2 MDR regions: a complex class 1 integron harboring blaNDM-1 and 7 other AMR genes, and the IS26-mph(A)-mrx-mphR(A)-IS6100 azithromycin resistance unit. The predicted antimicrobial resistance (AMR) genes correlated with the antimicrobial susceptibility testing results. The multidrug-resistant phenotype in addition to the presence of two important mobile genetic elements, suggest a potent role as a reservoir of antibiotic resistance for such a small IncR plasmid. IMPORTANCE Analyzing the genetic environment of clinically relevant MDR genes can provide information on the way in which such genes are maintained and disseminated. Understanding this phenomenon is of interest for clinicians as it can also provide insight on where these genes might have been sourced, possibly supporting outbreak investigations.

A hospital-wide outbreak of IMI-17-producing Enterobacter ludwigii in an Israeli hospital.

ABSRACT: BACKGROUND: To describe the course and intervention of an hospital-wide IMI-Producing Enterobacter ludwigii outbreak. METHODS: This was an outbreak interventional study, done at a tertiary care center in Tel-Aviv, Israel. Data was collected on the course of the outbreak and the demographic and clinical characteristics of all patients involved in the outbreak. The intervention measures included patients' cohorting, contact isolation precautions, environmental cleaning and screening of contacts. The molecular features and phylogeny of outbreak-related isolates were studied by whole-genome based analysis. RESULTS: The outbreak included 34 patients that were colonized by IMI-Producing E. ludwigii and were identified in 24 wards throughout the hospital. Colonization was identified in the first 72 h of admission in 13/34 patients (38.2%). Most patients (91.2%) were admitted from home and had relatively low level of comorbidities. The majority of them (88%) had no recent use of invasive catheters and none had previous carriage of other multi-drug resistant bacteria. All available isolates harbored the blaIMI-17 allele and belonged to Sequence-Type 385. With the exception of two isolates, all isolates were closely related with less than a 20-SNP difference between them. CONCLUSIONS: This outbreak had most likely originated in the community and subsequently disseminated inside our institution. More studies are required in order to elucidate the epidemiology of IMI-Producing E. ludwigii and the possible role of environmental sources in its dissemination.

Has resistance to chlorhexidine increased among clinically-relevant bacteria? A systematic review of time course and subpopulation data.

Chlorhexidine (CHX) was introduced for use as an antimicrobial more than 70 years ago. CHX has been and continues to be used broadly for disinfecting surfaces in medical and food service facilities as well as directly on skin of humans and animals. Considering its widespread use over many decades, questions of resistance to CHX have been raised. Additionally, questions of possible coincident resistance to the biocide and resistance to clinically relevant antibiotics have also been raised. A number of important questions remain, including is there consistent evidence of resistance, what is the degree of resistance, especially among clinically isolated microbial strains, and what is the degree of resistance compared to the typical concentrations of the biocide used? Data for microbial species isolated over the last 70+ years were compiled to construct as complete a picture as practical regarding possible resistance, especially among species in which resistance to commonly used antibiotics has been noted to be increasing. This is a compilation and analysis of individual MIC values for CHX reported in the literature, not a compilation of the conclusions individual authors reached. The data were analyzed using straight-forward and robust statistical procedures to detect changes in susceptibility to CHX over time, i.e. linear regression. Linear regression was supplemented with the use of nonlinear least squares regression analysis to detect the presence of population parameters associated with subpopulations of microbial strains which exhibit increased resistance to CHX. Pseudomonas aeruginosa, Klebsiella pneumoniae, and Acinetobacter baumannii were all found to have an increased resistance to CHX over time with the most profound change detected in A. baumannii. Additionally, subpopulations with log-normal distributions were found consistent with the presence of a baseline subpopulation of susceptible strains and a subpopulation with increased resistance to CHX. However, the CHX-resistant subpopulations did not correlate exactly with antibiotic resistance, so details of the relationship remain to be addressed. Increased resistance over time was not detected for Escherichia coli, Enterobacter faecalis, Staphylococcus aureus, or Candida albicans, although a subpopulation with greater than baseline resistance to CHX was detected among strains of E. faecalis and C. albicans. A difference in susceptibility to CHX was also detected between methicillin-resistant (MRSA) and methicillin-sensitive (MSSA) S. aureus strains. The levels of resistance to CHX detected were all markedly lower than concentrations routinely used in medical and food service applications. Reaching conclusions regarding the relationship between antibiotic and CHX resistance was complicated by the limited overlap between tests of CHX and antibiotic resistance for several species. The results compiled here may serve as a foundation for monitoring changes in resistance to CHX and possible relationships between the use of CHX and resistance to antibiotics commonly used in clinical medicine.

Evaluation of the MICRONAUT MIC-strip colistin assay for colistin susceptibility testing of carbapenem-resistant Acinetobacter baumannii and Enterobacterales.

We sought to evaluate the MICRONAUT MIC-Strip colistin (MMS) assay, a commercial broth microdilution (BMD) panel, using 273 carbapenem-resistant Acinetobacter baumannii and Enterobacterales isolates. BMD was used as gold standard. MMS had 98.5% sensitivity and 99.5% specificity. All 37 isolates with MICs close to the breakpoint exhibited complete agreement.

Presence of ß-Lactamase-producing Enterobacterales and Salmonella Isolates in Marine Mammals.

Marine mammals have been described as sentinels of the health of marine ecosystems. Therefore, the aim of this study was to investigate (i) the presence of extended-spectrum ß-lactamase (ESBL)- and AmpC-producing Enterobacterales, which comprise several bacterial families important to the healthcare sector, as well as (ii) the presence of Salmonella in these coastal animals. The antimicrobial resistance pheno- and genotypes, as well as biocide susceptibility of Enterobacterales isolated from stranded marine mammals, were determined prior to their rehabilitation. All E. coli isolates (n = 27) were screened for virulence genes via DNA-based microarray, and twelve selected E. coli isolates were analyzed by whole-genome sequencing. Seventy-one percent of the Enterobacterales isolates exhibited a multidrug-resistant (MDR) pheno- and genotype. The gene blaCMY (n = 51) was the predominant ß-lactamase gene. In addition, blaTEM-1 (n = 38), blaSHV-33 (n = 8), blaCTX-M-15 (n = 7), blaOXA-1 (n = 7), blaSHV-11 (n = 3), and blaDHA-1 (n = 2) were detected. The most prevalent non-ß-lactamase genes were sul2 (n = 38), strA (n = 34), strB (n = 34), and tet(A) (n = 34). Escherichia coli isolates belonging to the pandemic sequence types (STs) ST38, ST167, and ST648 were identified. Among Salmonella isolates (n = 18), S. Havana was the most prevalent serotype. The present study revealed a high prevalence of MDR bacteria and the presence of pandemic high-risk clones, both of which are indicators of anthropogenic antimicrobial pollution, in marine mammals.

Effects of vancomycin-induced gut microbiome alteration on the pharmacodynamics of metformin in healthy male subjects.

Metformin is a major treatment for type 2 diabetes. This study was conducted to investigate the impact of gut microbiome dysbiosis on the pharmacokinetics and antihyperglycemic effects of metformin. Healthy adult males aged 19-45 years with no defecation abnormalities were recruited for this 4-period clinical study: baseline; post-metformin (i.e., multiple oral doses of 1000 mg metformin on days 1-4); post-vancomycin (i.e., multiple oral doses of 500 mg vancomycin on days 11-17 inducing gut microbiome changes); and post-metformin + vancomycin (i.e., multiple oral doses of 1000 mg metformin on days 16-19). In each period, serum glucose and insulin concentrations following an oral glucose tolerance test, fecal samples for gut microbiome composition, and safety data were obtained. Following metformin dosing, plasma and urine samples for pharmacokinetics were collected. Nine subjects completed the study. The pharmacokinetics of metformin remained unchanged, and the antihyperglycemic effect was significantly decreased after vancomycin administration (p value = 0.039), demonstrating the weak relationship between the pharmacokinetics and pharmacodynamics of metformin. Relative abundances of some genus were changed after vancomycin administration, and tended to correlate with the antihyperglycemic effects of metformin (p value = 0.062 for Erysipelatoclostridium; p value = 0.039 for Enterobacter; and p value = 0.086 for Faecalibacterium). Adverse events occurred in all subjects and were resolved without sequelae. In conclusion, a decrease in the antihyperglycemic effect of metformin was observed after concomitant administration with vancomycin, without changes in metformin pharmacokinetics. The antihyperglycemic effect was tended to correlate with the relative abundance of several genus, suggesting that the effect of metformin is partly attributable to the gut microbiome (ClinicalTrials.gov, NCT03809260).

Institutional outbreak involving multiple clades of IMP-producing Enterobacter cloacae complex sequence type 78 at a cancer center in Tokyo, Japan.

BACKGROUND: Information about the clinical and microbiological characteristics of IMP-producing Enterobacterales has been limited. Here, we describe an institutional outbreak of IMP-producing Enterobacter cloacae complex (ECC) involving multiple clades of ECC sequence type (ST) 78 strains. METHODS: Antimicrobial susceptibility testing, whole-genome sequencing, and conjugation experiments of 18 IMP-producing ECC strains isolated during four-year study period were performed. Species and subspecies were determined by average nucleotide identity analysis and clonal relatedness of the isolates was analyzed with multilocus sequence typing and core-genome single nucleotide polymorphism (SNP) analysis. Relevant clinical information was extracted from medical records. RESULTS: Fourteen of 18 IMP-producing ECC isolates were determined as Enterobacter hormaechei ST78. Sixteen isolates, including 13 isolates belonging to ST78, carried blaIMP-1 in In316-like class 1 integron and also carried IncHI2 plasmids. Conjugation experiments were successful for 12 isolates carrying blaIMP-1 on IncHI2 plasmids and for an isolate carrying blaIMP-11 on an IncL/M plasmid. Although isolation of ST78 strains was clustered in a 14-months period suggesting nosocomial transmission, these strains were subdivided into three clades by SNP analysis: clade A (n = 10), clade B (n = 1), clade C (n = 3). A part of clonal relatedness was unexpected by the epidemiological information at the time of isolation of the strains. Most of the IMP-producing ECC strains were susceptible to non-ß-lactam antibiotics and had relatively low minimum inhibitory concentrations to carbapenems (≤4 µg/mL). Five of six infections caused by IMP-producing ECC were treated successfully. CONCLUSIONS: Whole-genome sequencing analysis revealed the outbreak was caused by three different clades of ST78 strains, where patients had favorable treatment outcome of the infections compared with that caused by Enterobacterales producing other carbapenemases, possibly due to their non-multidrug-resistant phenotype.

Risk factor analysis for piperacillin-tazobactam-resistant Enterobacter spp. bacteremia at a tertiary hospital.

This study aimed to analyze the risk factors for piperacillin-tazobactam (TZP1)-resistant Enterobacter spp. bacteremia. The medical records of 111 patients with Enterobacter spp. bacteremia divided into a TZP-susceptible group (minimum inhibitory concentrations [MICs2] ≤16 µg/mL) and TZP-resistant group (MICs >16 µg/mL) were retrospectively reviewed. The male-to-female ratio, age, underlying disease, and infection site did not differ between the 2 groups. Multivariate analysis revealed that the independent predictor associated with TZP-resistant Enterobacter spp. bacteremia was the previous usage of third-generation cephalosporins (P = 0.036). In conclusion, TZP administration in cases of suspected Enterobacter spp. bacteremia previously treated with third-generation cephalosporin should be cautiously considered.

Co-infections of two carbapenemase-producing Enterobacter hormaechei clinical strains isolated from the same diabetes individual in China.

Introduction. Since mcr-1 was first reported in China, there have been ten variants of MCR appearing nationwide so far. Multidrug-resistant Enterobacteriaceae bacteria carrying both NDM and MCR have become a serious threat to global public health.Hypothesis/Gap Statement. The genetic structure of mcr-9 needs to be better understood in order to better prevent and control the transmission of drug-resistant genes.Aims. The aim of this study was to characterize the presence of two Enterobacter hormaechei isolates, which carries bla NDM-5 CME2 and the coexistence of mcr-9 and bla NDM-1 strain CMD2, which were isolated from a patient with diabetes in Sichuan, China.Methodology. The microbroth dilution method was used for antibiotic susceptibility. Conjugation experiment was used to investigate the transferability of bla NDM-1, bla NDM-5 and mcr-9. Whole-genome sequencing was performed on Illumina HiSeq platform. The ability of biofilm formation was detected by crystal-violet staining, the virulence of the bacteria was measured by Galleria mellonella killing assay.Results. bla NDM-5 carrier CME2 and CMD2 with bla NDM-1 and mcr-9 were resistant to carbapenems, ß-lactam, aminoglycoside, quinolone and tetracycline, while CMD2 was also resistant to colistin. Conjugation assay and plasmid replicon typing further demonstrated that both bla NDM-1 and bla NDM-5 were respectively present on the self-transferrable IncX3 plasmid, mcr-9 was located on the self-transferrable IncHI2 plasmid. Through the analysis of mcr-9 gene context, the structure was DUF4942-rcnR-rcnA-copS-IS903-mcr-9-wbuC-qseC-qseB-IS1R-ΔsilR-IS903, bla NDM-1 context was IS3000-ΔISAba125-IS5-bla NDM-1-ble-trpF-groS-groL-insE-ΔIS26 structure, bla NDM-5 structure was IS3000-bla NDM-5-ble-trpF-dsbC-ΔIS26-umuD-ISKox3-tnpR-parA. Biofilm formation of CME2 was stronger than CMD2. There was no significant difference in virulence between the two strains.Conclusion. This study reveals two multiple drug-resistant E. hormaechei isolates from diabetes patient samples. E. hormaechei carrying two NDM-resistant genes is already a serious threat, where MCR is an important cause of treatment failure in bacterial infections. This study is a reminder not only to prevent infection in patients with diabetes, but also to constantly monitor the epidemic and spread of the drug-resistant gene.