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570200.9070Disinfectant Susceptibility of Third-Generation-Cephalosporin/Carbapenem-Resistant Gram-Negative Bacteria Isolated from the Oral Cavity of Residents of Long-Term-Care Facilities. We have recently reported the isolation of third-generation-cephalosporin-resistant Gram-negative bacteria from the oral cavity of residents of a long-term-care facility (LTCF). Since disinfectants are often used in the oral cavity, it is important to investigate the disinfectant susceptibility of oral bacteria. Here, we evaluated the susceptibilities of Gram-negative antimicrobial-resistant bacteria (GN-ARB), including Pseudomonas, Acinetobacter, and Enterobacteriaceae, obtained from the oral cavity of residents of LTCFs to povidone-iodine (PVPI), cetylpyridinium chloride (CPC), benzalkonium chloride (BZK), and chlorhexidine chloride (CHX). We also evaluated the susceptibilities of isolates from the rectum to the same agents to compare the susceptibility profiles of oral and rectal isolates. Next, we investigated the relationship between their susceptibility and disinfectant resistance genes delineated by whole-genome sequencing of the isolates. Additionally, we evaluated the correlation between disinfectant-resistant GN-ARB and clinical information. In oral GN-ARB, the MIC of PVPI showed almost identical values across isolates, while the MICs of CPC, BZK, and CHX showed a wide range of variation among species/strains. In particular, Pseudomonas aeruginosa exhibited high-level resistance to CPC and BZK. The disinfectant susceptibility of rectal GN-ARB showed a tendency similar to that of oral GN-ARB. The presence of qacEΔ1 was correlated with CPC/BZK resistance in P. aeruginosa, while other species exhibited no correlation between qacEΔ1 and resistance. Multiple analyses showed the correlation between the presence of CPC-resistant bacteria in the oral cavity and tube feeding. In conclusion, we found that some oral GN-ARB isolates showed resistance to not only antibiotics but also disinfectants. IMPORTANCE Antibiotic-resistant bacteria (ARB) are becoming a serious concern worldwide. We previously reported the isolation of third-generation-cephalosporin-resistant Gram-negative bacteria from the oral cavity of residents of a long-term-care facility (LTCF). To prevent infection with ARB in hospitals and eldercare facilities, we must pay more attention to the use of not only antibiotics but also disinfectants. However, the effect of disinfectants on ARB is unclear. In this study, we evaluated the susceptibility of Gram-negative ARB (GN-ARB) from the oral cavity of residents of LTCFs to some disinfectants that are often used for the oral cavity; we found that some isolates showed resistance to several disinfectants. This is the first comprehensive analysis of the disinfectant susceptibility of oral GN-ARB. These results provide some important information for infection control and suggest that disinfectants should be applied carefully.202336515531
82810.9068Screening for Resistant Bacteria, Antimicrobial Resistance Genes, Sexually Transmitted Infections and Schistosoma spp. in Tissue Samples from Predominantly Vaginally Delivered Placentae in Ivory Coast and Ghana. Medical complications during pregnancy have been frequently reported from Western Africa with a particular importance of infectious complications. Placental tissue can either become the target of infectious agents itself, such as, e.g., in the case of urogenital schistosomiasis, or be subjected to contamination with colonizing or infection-associated microorganisms of the cervix or the vagina during vaginal delivery. In the retrospective cross-sectional assessment presented here, the quantitative dimension of infection or colonization with selected resistant or pathogenic bacteria and parasites was regionally assessed. To do so, 274 collected placental tissues from Ivory Coastal and Ghanaian women were subjected to selective growth of resistant bacteria, as well as to molecular screening for beta-lactamase genes, Schistosoma spp. and selected bacterial causative agents of sexually transmitted infections (STI). Panton-Valentine-negative methicillin-resistant Staphylococcus aureus (MRSA) was grown from 1.8% of the tissue samples, comprising the spa types t008 and t688, as well as the newly detected ones, t12101 (n = 2) and t12102. While the culture-based recovery of resistant Enterobacterales and nonfermentative rod-shaped Gram-negative bacteria failed, molecular assessments confirmed beta-lactamase genes in 31.0% of the samples with multiple detections of up to four resistance genes per sample and bla(CTX-M), bla(IMP), bla(GES), bla(VIM), bla(OXA-58)-like, bla(NDM), bla(OXA-23)-like, bla(OXA-48)-like and bla(KPC) occurring in descending order of frequency. The beta-lactamase genes bla(OXA-40/24)-like, bla(NMC_A/IMI), bla(BIC), bla(SME), bla(GIM) and bla(DIM) were not detected. DNA of the urogenital schistosomiasis-associated Schistosoma haematobium complex was recorded in 18.6% of the samples, but only a single positive signal for S. mansoni with a high cycle-threshold value in real-time PCR was found. Of note, higher rates of schistosomiasis were observed in Ghana (54.9% vs. 10.3% in Ivory Coast) and Cesarean section was much more frequent in schistosomiasis patients (61.9% vs. 14.8% in women without Schistosoma spp. DNA in the placenta). Nucleic acid sequences of nonlymphogranuloma-venereum-associated Chlamydia trachomatis and of Neisseria gonorrhoeae were recorded in 1.1% and 1.9% of the samples, respectively, while molecular attempts to diagnose Treponema pallidum and Mycoplasma genitalium did not lead to positive results. Molecular detection of Schistosoma spp. or STI-associated pathogens was only exceptionally associated with multiple resistance gene detections in the same sample, suggesting epidemiological distinctness. In conclusion, the assessment confirmed considerable prevalence of urogenital schistosomiasis and resistant bacterial colonization, as well as a regionally expected abundance of STI-associated pathogens. Continuous screening offers seem advisable to minimize the risks for the pregnant women and their newborns.202337623959
306920.9045The hospital sink drain biofilm resistome is independent of the corresponding microbiota, the environment and disinfection measures. In hospitals, the transmission of antibiotic-resistant bacteria (ARB) may occur via biofilms present in sink drains, which can lead to infections. Despite the potential role of sink drains in the transmission of ARB in nosocomial infections, routine surveillance of these drains is lacking in most hospitals. As a result, there is currently no comprehensive understanding of the transmission of ARB and the dissemination of antimicrobial resistance genes (ARGs) and associated mobile genetic elements (MGEs) via sink drains. This study employed a multifaceted approach to monitor the total aerobic bacteria as well as the presence of carbapenemase-producing Enterobacterales (CPEs), the microbiota and the resistome of sink drain biofilms (SDBs) and hospital wastewater (WW) of two separate intensive care units (ICUs) in the same healthcare facility in France. Samples of SDB and WW were collected on a monthly basis, from January to April 2023, in the neonatal (NICU) and the adult (AICU) ICUs of Grenoble Alpes University Hospital. In the NICU, sink drain disinfection with surfactants was performed routinely. In the AICU, routine disinfection is not carried out. Culturable aerobic bacteria were quantified on non-selective media, and CPEs were screened using two selective agars. Isolates were identified by MALDI-TOF MS, and antibiotic susceptibility testing (AST) was performed on Enterobacterales and P. aeruginosa. The resistome was analyzed by high-throughput qPCR targeting >80 ARGs and MGEs. The overall bacterial microbiota was assessed via full-length 16S rRNA sequencing. No CPEs were isolated from SDBs in either ICU by bacterial culture. Culture-independent approaches revealed an overall distinct microbiota composition of the SDBs in the two ICUs. The AICU SDBs were dominated by pathogens containing Gram-negative bacterial genera including Pseudomonas, Stenotrophomona, Klebsiella, and Gram-positive Staphylococcus, while the NICU SDBs were dominated by the Gram-negative genera Achromobacter, Serratia, and Acidovorax, as well as the Gram-positive genera Weisella and Lactiplantibacillus. In contrast, the resistome of the SDBs exhibited no significant differences between the two ICUs, indicating that the abundance of ARGs and MGEs is independent of microbiota composition and disinfection practices. The AICU WW exhibited more distinct aerobic bacteria than the NICU WW. In addition, the AICU WW yielded 15 CPEs, whereas the NICU WW yielded a single CPE. All the CPEs were characterized at the species level. The microbiota of the NICU and AICU WW samples differed from their respective SDBs and exhibited distinct variations over the four-month period:the AICU WW contained a greater number of genes conferring resistance to quinolones and integron integrase genes, whereas the NICU WW exhibited a higher abundance of streptogramin resistance genes. Our study demonstrated that the resistome of the hospital SDBs in the two ICUs of the investigated healthcare institute is independent of the microbiota, the environment, and the local disinfection measures. However, the prevalence of CPEs in the WW pipes collecting the waste from the investigated drains differed. These findings offer valuable insights into the resilience of resistance genes in SDBs in ICUs, underscoring the necessity for innovative strategies to combat antimicrobial resistance in clinical environments.202540483807
85630.9033Oral colonisation by antimicrobial-resistant Gram-negative bacteria among long-term care facility residents: prevalence, risk factors, and molecular epidemiology. BACKGROUND: For residents of long-term care facilities (LTCFs), antimicrobial-resistant bacteria (ARB) are a risk factor, yet their oral colonisation, potentially leading to aspiration pneumonia, remains unclear. This study was undertaken to survey the prevalence, phenotypic characteristics, and molecular epidemiology of antimicrobial-resistant Gram-negative bacteria in the oral cavity of LTCF residents, and to analyse the risk factors for such carriers. METHODS: This study involved 98 residents of a LTCF in Hiroshima City, Japan, aged between 55 and 101 years. Oropharyngeal swabs were collected and plated on screening media for ESBL-producing and carbapenem-resistant bacteria; isolates were identified and tested for antibiotic susceptibility; biofilm formation was tested in vitro; identification of epidemic clones were pre-determined by PCR; resistance genes, sequence types, and whole-genome comparison of strains were conducted using draft genome sequences. Demographic data and clinical characterisations were collected and risk factors analysed. RESULTS: Fifty-four strains from 38% of the residents grew on screening media and comprised predominantly of Acinetobacter spp. (35%), Enterobacteriaceae spp. (22%), and Pseudomonas spp. (19%). All Escherichia coli isolates carried CTX-M-9 group and belonged to the phylogroup B2, O25:H4 ST131 fimH30 lineage. Six Acinetobacter baumannii isolates presented identical molecular characteristics and revealed more biofilm production than the others, strongly suggesting their clonal lineage. One Acinetobacter ursingii isolate displayed extensive resistance to various ß-lactams due to multiple acquired resistance genes. One Pseudomonas aeruginosa isolate showed exceptional resistance to all ß-lactams including carbapenems, aminoglycosides, and a new quinolone, showing a multidrug-resistant Pseudomonas aeruginosa (MDRP) phenotype and remarkable biofilm formation. Genome sequence analysis revealed this isolate was the bla(IMP-1)-positive clone ST235 in Japan. Strokes (cerebral infarction or cerebral haemorrhage) and percutaneous endoscopic gastrostomy tubes were recognised as risk factors for oral colonisation by ARB in the LTCF residents. CONCLUSIONS: ARB, as defined by growth on screening agar plates, which carried mobile resistance genes or elements or conferred high biofilm formation, were already prevalent in the oral cavity of LTCF residents. Health-care workers involved in oral care should be aware of antimicrobial resistance and pay special attention to transmission prevention and infection control measures to diminish ARB or mobile resistance elements dissemination in LTCFs.202032131899
523540.9033Draft genome sequences of rare Lelliottia nimipressuralis strain MEZLN61 and two Enterobacter kobei strains MEZEK193 and MEZEK194 carrying mobile colistin resistance gene mcr-9 isolated from wastewater in South Africa. OBJECTIVES: Antimicrobial-resistant bacteria of the order Enterobacterales are emerging threats to global public and animal health, leading to morbidity and mortality. The emergence of antimicrobial-resistant, livestock-associated pathogens is a great public health concern. The genera Enterobacter and Lelliottia are ubiquitous, facultatively anaerobic, motile, non-spore-forming, rod-shaped Gram-negative bacteria belonging to the Enterobacteriaceae family and include pathogens of public health importance. Here, we report the first draft genome sequences of a rare Lelliottia nimipressuralis strain MEZLN61 and two Enterobacter kobei strains MEZEK193 and MEZEK194 in Africa. METHODS: The bacteria were isolated from environmental wastewater samples. Bacteria were cultured on nutrient agar, and the pure cultures were subjected to whole-genome sequencing. Genomic DNA was sequenced using an Illumina MiSeq platform. Generated reads were trimmed and subjected to de novo assembly. The assembled contigs were analysed for virulence genes, antimicrobial resistance genes, and extra-chromosomal plasmids, and multilocus sequence typing was performed. To compare the sequenced strains with other, previously sequenced E. kobei and L. nimipressuralis strains, available raw read sequences were downloaded, and all sequence files were treated identically to generate core genome bootstrapped maximum likelihood phylogenetic trees. RESULTS: Whole-genome sequencing analyses identified strain MEZLN61 as L. nimipressuralis and strains MEZEK193 and MEZEK194 as E. kobei. MEZEK193 and MEZEK194 carried genes encoding resistance to fosfomycin (fosA), beta-lactam antibiotics (bla(ACT-9)), and colistin (mcr-9). Additionally, MEZEK193 harboured nine different virulence genes, while MEZEK194 harboured eleven different virulence genes. The phenotypic analysis showed that L. nimipressuralis strain MEZLN61 was susceptible to colistin (2 μg/mL), while E. kobei MEZEK193 (64 μg/mL) and MEZEK194 (32 μg/mL) were resistant to colistin. CONCLUSION: The genome sequences of strains L. nimipressuralis MEZLN6, E. kobei MEZEK193, and E. kobei MEZEK194 will serve as a reference point for molecular epidemiological studies of L. nimipressuralis and E. kobei in Africa. In addition, this study provides an in-depth analysis of the genomic structure and offers important information that helps clarify the pathogenesis and antimicrobial resistance of L. nimipressuralis and E. kobei. The detection of mcr-9, which is associated with very low-level colistin resistance in Enterobacter species, is alarming and may indicate the undetected dissemination of mcr genes in bacteria of the order Enterobacterales. Continuous monitoring and surveillance of the prevalence of mcr genes and their associated phenotypic changes in clinically important pathogens and environmentally associated bacteria is necessary to control and prevent the spread of colistin resistance.202336948496
147550.9032Evaluation of the FilmArray(®) Pneumonia Plus Panel for Rapid Diagnosis of Hospital-Acquired Pneumonia in Intensive Care Unit Patients. The FilmArray(®) Pneumonia plus Panel (FAPP) is a new multiplex molecular test for hospital-acquired pneumonia (HAP), which can rapidly detect 18 bacteria, 9 viruses, and 7 resistance genes. We aimed to compare the diagnosis performance of FAPP with conventional testing in 100 intensive care unit (ICU) patients who required mechanical ventilation, with clinically suspected HAP. A total of 237 samples [76 bronchoalveolar lavages (BAL(DS)) and 82 endotracheal aspirates (ETA(DS)) obtained at HAP diagnosis, and 79 ETA obtained during follow-up (ETA(TT))], were analyzed independently by routine microbiology testing and FAPP. 58 patients had paired BAL(DS) and ETA(DS). The positivity thresholds of semi-quantified bacteria were 10(3)-10(4) CFUs/mL or 10(4) copies/mL for BAL, and 10(5) CFUs/mL or copies/mL for ETA. Respiratory commensals (H. influenzae, S. aureus, E. coli, S. pneumoniae) were the most common pathogens. Discordant results for bacterial identification were observed in 33/76 (43.4%) BAL(DS) and 36/82 (43.9%) ETA(DS), and in most cases, FAPP identified one supplemental bacteria (23/33 BAL(DS) and 21/36 ETA(DS)). An absence of growth, or polybacterial cultures, explained almost equally the majority of the non-detections in culture. No linear relationship was observed between bin and CFUs/mL variables. Concordant results between paired BAL(DS) and ETA(DS) were obtained in 46/58 (79.3%) patients with FAPP. One of the 17 resistance genes detected with FAPP (mecA/C and MREJ) was not confirmed by conventional testing. Overall, FAPP enhanced the positivity rate of diagnostic testing, with increased recognition of coinfections. Implementing this strategy may allow clinicians to make more timely and informed decisions.202032983057
843960.9031Comparative genomics analysis and virulence-related factors in novel Aliarcobacter faecis and Aliarcobacter lanthieri species identified as potential opportunistic pathogens. BACKGROUND: Emerging pathogenic bacteria are an increasing threat to public health. Two recently described species of the genus Aliarcobacter, A. faecis and A. lanthieri, isolated from human or livestock feces, are closely related to Aliarcobacter zoonotic pathogens (A. cryaerophilus, A. skirrowii, and A. butzleri). In this study, comparative genomics analysis was carried out to examine the virulence-related, including virulence, antibiotic, and toxin (VAT) factors in the reference strains of A. faecis and A. lanthieri that may enable them to become potentially opportunistic zoonotic pathogens. RESULTS: Our results showed that the genomes of the reference strains of both species have flagella genes (flaA, flaB, flgG, flhA, flhB, fliI, fliP, motA and cheY1) as motility and export apparatus, as well as genes encoding the Twin-arginine translocation (Tat) (tatA, tatB and tatC), type II (pulE and pulF) and III (fliF, fliN and ylqH) secretory pathways, allowing them to secrete proteins into the periplasm and host cells. Invasion and immune evasion genes (ciaB, iamA, mviN, pldA, irgA and fur2) are found in both species, while adherence genes (cadF and cj1349) are only found in A. lanthieri. Acid (clpB), heat (clpA and clpB), osmotic (mviN), and low-iron (irgA and fur2) stress resistance genes were observed in both species, although urease genes were not found in them. In addition, arcB, gyrA and gyrB were found in both species, mutations of which may mediate the resistance to quaternary ammonium compounds (QACs). Furthermore, 11 VAT genes including six virulence (cadF, ciaB, irgA, mviN, pldA, and tlyA), two antibiotic resistance [tet(O) and tet(W)] and three cytolethal distending toxin (cdtA, cdtB, and cdtC) genes were validated with the PCR assays. A. lanthieri tested positive for all 11 VAT genes. By contrast, A. faecis showed positive for ten genes except for cdtB because no PCR assay for this gene was available for this species. CONCLUSIONS: The identification of the virulence, antibiotic-resistance, and toxin genes in the genomes of A. faecis and A. lanthieri reference strains through comparative genomics analysis and PCR assays highlighted the potential zoonotic pathogenicity of these two species. However, it is necessary to extend this study to include more clinical and environmental strains to explore inter-species and strain-level genetic variations in virulence-related genes and assess their potential to be opportunistic pathogens for animals and humans.202235761183
153570.9030Complete Genome Sequencing of Acinetobacter baumannii AC1633 and Acinetobacter nosocomialis AC1530 Unveils a Large Multidrug-Resistant Plasmid Encoding the NDM-1 and OXA-58 Carbapenemases. Carbapenem-resistant Acinetobacter spp. are considered priority drug-resistant human-pathogenic bacteria. The genomes of two carbapenem-resistant Acinetobacter spp. clinical isolates obtained from the same tertiary hospital in Terengganu, Malaysia, namely, A. baumannii AC1633 and A. nosocomialis AC1530, were sequenced. Both isolates were found to harbor the carbapenemase genes bla(NDM-1) and bla(OXA-58) in a large (ca. 170 kb) plasmid designated pAC1633-1 and pAC1530, respectively, that also encodes genes that confer resistance to aminoglycosides, sulfonamides, and macrolides. The two plasmids were almost identical except for the insertion of ISAba11 and an IS4 family element in pAC1633-1, and ISAba11 along with relBE toxin-antitoxin genes flanked by inversely orientated pdif (XerC/XerD) recombination sites in pAC1530. The bla(NDM-1) gene was encoded in a Tn125 composite transposon structure flanked by ISAba125, whereas bla(OXA-58) was flanked by ISAba11 and ISAba3 downstream and a partial ISAba3 element upstream within a pdif module. The presence of conjugative genes in plasmids pAC1633-1/pAC1530 and their discovery in two distinct species of Acinetobacter from the same hospital are suggestive of conjugative transfer, but mating experiments failed to demonstrate transmissibility under standard laboratory conditions. Comparative sequence analysis strongly inferred that pAC1633-1/pAC1530 was derived from two separate plasmids in an IS1006-mediated recombination or transposition event. A. baumannii AC1633 also harbored three other plasmids designated pAC1633-2, pAC1633-3, and pAC1633-4. Both pAC1633-3 and pAC1633-4 are cryptic plasmids, whereas pAC1633-2 is a 12,651-bp plasmid of the GR8/GR23 Rep3-superfamily group that encodes the tetA(39) tetracycline resistance determinant in a pdif module.IMPORTANCE Bacteria of the genus Acinetobacter are important hospital-acquired pathogens, with carbapenem-resistant A. baumannii listed by the World Health Organization as the one of the top priority pathogens. Whole-genome sequencing of carbapenem-resistant A. baumannii AC1633 and A. nosocomialis AC1530, which were isolated from the main tertiary hospital in Terengganu, Malaysia, led to the discovery of a large, ca. 170-kb plasmid that harbored genes encoding the New Delhi metallo-β-lactamase-1 (NDM-1) and OXA-58 carbapenemases alongside genes that conferred resistance to aminoglycosides, macrolides, and sulfonamides. The plasmid was a patchwork of multiple mobile genetic elements and comparative sequence analysis indicated that it may have been derived from two separate plasmids through an IS1006-mediated recombination or transposition event. The presence of such a potentially transmissible plasmid encoding resistance to multiple antimicrobials warrants vigilance, as its spread to susceptible strains would lead to increasing incidences of antimicrobial resistance.202133504662
140780.9022World Health Organization priority antimicrobial resistance in Enterobacterales, Acinetobacter baumannii, Pseudomonas aeruginosa, Staphylococcus aureus and Enterococcus faecium healthcare-associated bloodstream infections in Brazil (ASCENSION): a prospective, multicentre, observational study. BACKGROUND: Carbapenem-resistant Enterobacterales (CRE), Acinetobacter baumannii (CRAB), Pseudomonas aeruginosa (CRPA), methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE) are listed by World Health Organization (WHO) as priority antimicrobial-resistant bacteria. Data on WHO Priority Antimicrobial resistance Phenotype (WPAP) bacteria from low- and middle-income countries are scarce. In this study, we investigated the occurrence of WPAP in healthcare-associated bloodstream infections (BSI) in Brazil, an upper-middle-income country in South America. METHODS: ASCENSION was a prospective, multicentre, observational study conducted in 14 hospitals from four of five Brazilian regions. Enterobacterales, A. baumannii, P. aeruginosa, S. aureus and E. faecium BSIs in hospitalised patients were analysed. The primary outcome was the frequency of WPAP among all bacteria of interest. Secondary outcomes were incidence-density of bacteria isolates in hospitalised patients, WPAP proportions within bacterial species, and 28-day mortality. PCR for carbapenemase genes was performed in carbapenem-resistant Gram-negative bacteria. FINDINGS: Between August 15, 2022, and August 14, 2023, 1350 isolates (1220 BSI episodes) were included. WPAP accounted for 38.8% (n = 524; 95% Confidence Interval 32.0-46.1) of all isolates, with CRE (19.3%) as the most frequent, followed by CRAB (9.6%), MRSA (4.9%), VRE (2.7%), and CRPA (2.4%). Incidence-density of all and WPAP isolates were 1.91 and 0.77/1000 patients-day, respectively. Carbapenem-resistant Klebsiella pneumoniae (CRKP) was the most common CRE, corresponding to 14.2% of all BSIs. A. baumannii isolates presented the highest proportion of WPAP (87.8%). Mortality rates were higher in patients with BSIs by WPAP than non-WPAP isolates. KPC (64.4%) was the predominant carbapenemase in CRE, followed by NDM (28.4%) and KPC + NDM co-production (7.1%). OXA-23 was the most frequent in CRAB. INTERPRETATION: A high frequency of WPAP bacteria, particularly CRKP and CRAB, were found in healthcare-associated BSIs in Brazil, posing them as a major public health problem in this country. FUNDING: National Council for Scientific and Technological Development, Brazil.202539957800
142590.9022Distribution and Antimicrobial Resistance of Complicated Intraabdominal Infection Pathogens in Two Tertiary Hospitals in Egypt. Background: Management of complicated intraabdominal infections (cIAIs) requires containment of the source and appropriate initial antimicrobial therapy. Identifying the local data is important to guide the empirical selection of antimicrobial therapy. In this study, we aimed to describe the pathogen distribution and antimicrobial resistance of cIAI. Methods: In two major tertiary care hospitals in Egypt, we enrolled patients who met the case definition of cIAI from October 2022 to September 2023. Blood cultures were performed using the BACTAlert system (BioMerieux, Marcy l'Etoile, France). A culture of aspirated fluid, resected material, or debridement of the infection site was performed. Identification of pathogens and antimicrobial susceptibility testing were conducted by the VITEK-2 system (BioMerieux, Marcy l'Etoile, France). Gram-negative resistance genes were identified by PCR and confirmed by whole bacterial genome sequencing using the Nextera XT DNA Library Preparation Kit and sequencing with the MiSeq Reagent Kit 600 v3 (Illumina, USA) on the Illumina MiSeq. Results: We enrolled 423 patients, 275 (65.01%) males. The median age was 61.35 (range 25-72 years). We studied 452 recovered bacterial isolates. Gram-negative bacteria were the vast majority, dominated by E. coli, followed by Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, and Proteus mirabilis (33.6%, 30.5%, 13.7%, 13%, and 5.4%, respectively). High rates of resistance were detected to third- and fourth-generation cephalosporins and fluoroquinolones. No resistance was detected to colistin. Resistance to amikacin and tigecycline was low among all isolates. Resistance to meropenem and ceftazidime/avibactam was moderate. ESBL genes were common in E. coli and K. pneumoniae. CTX-M15 gene was the most frequent. Among Enterobacterales, bla(OXA-48) and bla(NDM) were the most prevalent carbapenemase genes. Pseudomonas aeruginosa isolates harbored a wide variety of carbapenemase genes (OXA, NDM, VIM, SIM, GIM, SPM, IMP, AIM), dominated by metallo-beta-lactamases. In 20.6% of isolates, we identified two or more resistance genes. Conclusion: High resistance rates were detected to third- and fourth-generation cephalosporins and fluoroquinolones. Amikacin and tigecyclines were the most active antimicrobials. Our data call for urgent implementation of antimicrobial stewardship programs and reinforcement of infection control.202439172656
3064100.9022High Diversity but Monodominance of Multidrug-Resistant Bacteria in Immunocompromised Pediatric Patients with Acute Lymphoblastic Leukemia Developing GVHD Are Not Associated with Changes in Gut Mycobiome. Graft-versus-host disease (GvHD) is a severe complication after hematopoietic stem cell transplantation (HSCT). Our study focused on identifying multidrug-resistant (MDR) gut bacteria associated with GvHD-prone guts and association with gut microbiota (GM) diversity, bacteriome, and mycobiome composition in post-HSCT patients. We examined 11 pediatric patients with acute lymphoblastic leukemia (ALL), including six with GvHD, within three time points: seven days pre-HSCT, seven days post-, and 28 days post-HSCT. The gut microbiome and its resistome were investigated using metagenomic sequencing, taxonomically classified with Kraken2, and statistically evaluated for significance using appropriate tests. We observed an increase in the abundance of MDR bacteria, mainly Enterococcus faecium strains carrying msr(C), erm(T), aac(6')-li, dfrG, and ant(6)-la genes, in GvHD patients one week post-HSCT. Conversely, non-GvHD patients had more MDR beneficial bacteria pre-HSCT, promoting immunosurveillance, with resistance genes increasing one-month post-HSCT. MDR beneficial bacteria included the anti-inflammatory Bacteroides fragilis, Ruminococcus gnavus, and Turicibacter, while most MDR bacteria represented the dominant species of GM. Changes in the gut mycobiome were not associated with MDR bacterial monodominance or GvHD. Significant α-diversity decline (Shannon index) one week and one month post-HSCT in GvHD patients (p < 0.05) was accompanied by increased Pseudomonadota and decreased Bacteroidota post-HSCT. Our findings suggest that MDR commensal gut bacteria may preserve diversity and enhance immunosurveillance, potentially preventing GvHD in pediatric ALL patients undergoing HSCT. This observation has therapeutic implications.202338136701
5161110.9021Genomic analysis of contaminant Stenotrophomonas maltophilia, from placental swab culture, carrying antibiotic resistance: a potential hospital laboratory contaminant. Acute chorioamnionitis has been considered as reflective of amniotic fluid infection. Standard microbiological work ups for causative microorganism of intra-amniotic infection is based on microbial identification. However, frequency of positive placental culture is varied depending on placental sampling techniques, contaminations, methods of microbiologic work ups or comprehensive microbiologic work ups. In this report, we performed a hybrid whole genome sequencing of a proven bacterial contaminant obtained from placental culture in a patient with preterm labor and acute chorioamnionitis. This is to unveil genetic characterization of contaminant Stenotrophomonas maltophilia habouring antibiotic resistance genes. Stenotrophomonas maltiphilia was proven to be bacterial contaminant since Ureaplasma urealyticum was subsequently demonstrated in amniotic fluid by 16 S rRNA gene Sanger sequencing. Cultivation results from other sources were no growth. We identified Stenotrophomonas maltiphilia strain RAOG732 which carried several antibiotic resistance genes, including aminoglycoside, fluoroquiolone and beta-lactam. Biofilm production genes were also identified in this genome. We firstly utilized a hybrid sequencing approach to investigate the genome of S. maltiphilia in the patient with preterm and acute chorioamnionitis, a proven bacterial laboratory contaminant. The analysis provided several antibiotic resistance-associated and genes biofilm-associated genes. The detection of S. maltiphilia raised the awareness of the colonization of biofilm-producing bacteria in hospitals, where surveillance for decontamination is necessary.202540594762
7130120.9020Microbial community structure and resistome dynamics on elevator buttons in response to surface disinfection practices. BACKGROUND: Disinfectants have been extensively used in public environments since the COVID-19 outbreak to help control the spread of the virus. This study aims to investigate whether disinfectant use influences the structure of bacterial communities and contributes to bacterial resistance to disinfectants and antibiotics. METHODS: Using molecular biology techniques-including metagenomic sequencing and quantitative PCR (qPCR)-we analyzed the bacterial communities on elevator button surfaces from two tertiary hospitals, one infectious disease hospital, two quarantine hotels (designated for COVID-19 control), and five general hotels in Nanjing, Jiangsu Province, during the COVID-19 pandemic. We focused on detecting disinfectant resistance genes (DRGs), antibiotic resistance genes (ARGs), and mobile genetic elements (MGEs). RESULTS: Significant differences were observed in the bacterial community structures on elevator button surfaces across the four types of environments. Quarantine hotels, which implemented the most frequent disinfection protocols, exhibited distinct bacterial profiles at the phylum, genus, and species levels. Both α-diversity (within-sample diversity) and β-diversity (between-sample diversity) were lower and more distinct in quarantine hotels compared to the other environments. The abundance of DRGs, ARGs, and MGEs was also significantly higher on elevator button surfaces in quarantine hotels. Notably, antibiotic-resistant bacteria (ARBs), including Escherichia coli, Acinetobacter baumannii, and Pseudomonas aeruginosa, were detected in all four settings. CONCLUSION: The structure of bacterial communities on elevator button surfaces varies across different environments, likely influenced by the frequency of disinfectant use. Increased resistance gene abundance in quarantine hotels suggests that disinfection practices may contribute to the selection and spread of resistant bacteria. Enhanced monitoring of disinfection effectiveness and refinement of protocols in high-risk environments such as hospitals and hotels are essential to limit the spread of resistant pathogens.202540520307
1538130.9019KPC-2 allelic variants in Klebsiella pneumoniae isolates resistant to ceftazidime-avibactam from Argentina: bla(KPC-80), bla(KPC-81), bla(KPC-96) and bla(KPC-97). Ceftazidime-avibactam (CZA) therapy has significantly improved survival rates for patients infected by carbapenem-resistant bacteria, including KPC producers. However, resistance to CZA is a growing concern, attributed to multiple mechanisms. In this study, we characterized four clinical CZA-resistant Klebsiella pneumoniae isolates obtained between July 2019 and December 2020. These isolates expressed novel allelic variants of bla(KPC-2) resulting from changes in hotspots of the mature protein, particularly in loops surrounding the active site of KPC. Notably, KPC-80 had an K269_D270insPNK mutation near the Lys270-loop, KPC-81 had a del_I173 mutation within the Ω-loop, KPC-96 showed a Y241N substitution within the Val240-loop and KPC-97 had an V277_I278insNSEAV mutation within the Lys270-loop. Three of the four isolates exhibited low-level resistance to imipenem (4 µg/mL), while all remained susceptible to meropenem. Avibactam and relebactam effectively restored carbapenem susceptibility in resistant isolates. Cloning mutant bla(KPC) genes into pMBLe increased imipenem MICs in recipient Escherichia coli TOP10 for bla(KPC-80), bla(KPC-96), and bla(KPC-97) by two dilutions; again, these MICs were restored by avibactam and relebactam. Frameshift mutations disrupted ompK35 in three isolates. Additional resistance genes, including bla(TEM-1), bla(OXA-18) and bla(OXA-1), were also identified. Interestingly, three isolates belonged to clonal complex 11 (ST258 and ST11) and one to ST629. This study highlights the emergence of CZA resistance including unique allelic variants of bla(KPC-2) and impermeability. Comprehensive epidemiological surveillance and in-depth molecular studies are imperative for understanding and monitoring these complex resistance mechanisms, crucial for effective antimicrobial treatment strategies. IMPORTANCE: The emergence of ceftazidime-avibactam (CZA) resistance poses a significant threat to the efficacy of this life-saving therapy against carbapenem-resistant bacteria, particularly Klebsiella pneumoniae-producing KPC enzymes. This study investigates four clinical isolates exhibiting resistance to CZA, revealing novel allelic variants of the key resistance gene, bla(KPC-2). The mutations identified in hotspots surrounding the active site of KPC, such as K269_D270insPNK, del_I173, Y241N and V277_I278insNSEAV, prove the adaptability of these pathogens. Intriguingly, low-level resistance to imipenem and disruptions in porin genes were observed, emphasizing the complexity of the resistance mechanisms. Interestingly, three of four isolates belonged to clonal complex 11. This research not only sheds light on the clinical significance of CZA resistance but also shows the urgency for comprehensive surveillance and molecular studies to inform effective antimicrobial treatment strategies in the face of evolving bacterial resistance.202438319084
7646140.9018Assessment of Bacterial Community and Other Microorganism Along the Lam Takhong Watercourse, Nakhon Ratchasima, Thailand. Lam Takhong, a vital watercourse in Nakhon Ratchasima province, Thailand, supports agricultural, recreational, and urban activities. Originating in a national park, it flows through urban areas before discharging into a dam and running off via the sluice gate. While water quality monitoring is routine, microbial community data have never been reported. This study assesses the microorganism diversity and functional genes in Lam Takhong watercourse using a shotgun sequencing metagenomics approach. Water samples were collected from the upstream, midstream, and downstream sections. The midstream area exhibited the highest abundance of fecal coliform bacteria, plankton, and benthos, suggesting elevated pollution levels. Genes related to metabolism, particularly carbohydrate and amino acid pathways, were predominant. Proteobacteria was the most abundant phylum found in the water, with Limnohabitans as the dominant planktonic bacteria. Bacteria such as Staphylococcus, Mycobacterium, Escherichia, Pseudomonas, Enterococcus, Neisseria, Streptomyces, and Salmonella were detected, along with antibiotic resistance genes, raising public health concerns. These findings emphasize the need for microbial monitoring in the Lam Takhong to determine the potential water quality bioindicator and prevent potential disease spread through the water system.202540244481
5193150.9018Antibiotic resistance genes prediction via whole genome sequence analysis of Stenotrophomonas maltophilia. BACKGROUND: Stenotrophomonas maltophilia (S. maltophilia) is the first dominant ubiquitous bacterial species identified from the genus Stenotrophomonas in 1943 from a human source. S. maltophilia clinical strains are resistance to several therapies, this study is designed to investigate the whole genome sequence and antimicrobial resistance genes prediction in Stenotrophomonas maltophilia (S. maltophilia) SARC-5 and SARC-6 strains, isolated from the nasopharyngeal samples of an immunocompromised patient. METHODS: These bacterial strains were obtained from Pakistan Institute of Medical Sciences (PIMS) Hospital, Pakistan. The bacterial genome was sequenced using a whole-genome shotgun via a commercial service that used an NGS (Next Generation Sequencing) technology called as Illumina Hiseq 2000 system for genomic sequencing. Moreover, detailed in-silico analyses were done to predict the presence of antibiotic resistance genes in S. maltophilia. RESULTS: Results showed that S. maltophilia is a rare gram negative, rod-shaped, non sporulating bacteria. The genome assembly results in 24 contigs (>500 bp) having a size of 4668,850 bp with 65.8% GC contents. Phylogenetic analysis showed that SARC-5 and SARC-6 were closely related to S. maltophilia B111, S. maltophilia BAB-5317, S. maltophilia AHL, S. maltophilia BAB-5307, S. maltophilia RD-AZPVI_04, S. maltophilia JFZ2, S. maltophilia RD_MAAMIB_06 and lastly with S. maltophilia sp ROi7. Moreover, the whole genome sequence analysis of both SARC-5 and SARC-6 revealed the presence of four resistance genes adeF, qacG, adeF, and smeR. CONCLUSION: Our study confirmed that S. maltophilia SARC-5 and SARC-6 are one of the leading causes of nosocomial infection which carry multiple antibiotic resistance genes.202438128408
1991160.9017A strain defined as a novel species in the Acinetobacter genus co-harboring chromosomal associated tet(X3) and plasmid associated bla (NDM-1) from a beef cattle farm in Hebei, China. INTRODUCTION: The co-existence phenomenon of antibiotic resistance genes (ARGs), particularly of last-resort antibiotics in multi-drug resistant (MDR) bacteria, is of particular concern in the least studied bacterial species. METHODS: In 2023, strain M2 was isolated from the sludge sample at a commercial bovine farm in Hebei province, China, using a MacConkey plate containing meropenem. PCR amplification and Sanger sequencing verified it co-carrying bla (NDM) and tet(X) genes. It was classified within the Acinetobacter genus by MALDI-TOF-MS and 16S rDNA analyses. Whole-genome sequencing (WGS) was performed on the Oxford Nanopore platform, with species-level identification via ANI and dDDH. Antimicrobial susceptibility testing was performed against 20 antibiotics. Conjugation assays employed the filter-mating method using E. coli J53 and Salmonella LGJ2 as recipients. RESULTS: This strain was confirmed as a novel species of Acinetobacter genus, showing resistance to meropenem, ampicillin, ceftazidime, cefepime, gentamicin, kanamycin, fosfomycin, imipenem, ertapenem, and tetracycline. Despite carrying tet(X3), it remained susceptible to tigecycline, omadacycline, and doxycycline. The genome carried 11 ARG types, multiple metal resistance genes (MRGs), and virulence factor (VF) genes. The bla (NDM-1) was located in a skeleton, ISAba125-bla (NDM-1)-ble (MBL)-trpF, which was carried by an ISAba14-mediated rolling-circle-like structure in pM2-2-NDM-1 (rep_cluster_481). Integrative and conjugative element (ICE) and multiple pdif modules (driven by the XerCD site-specific recombination (XerCD SSR) system), which were associated with the mobilization of resistance determinants, were identified in this plasmid. Chromosomal tet(X3) was mediated by ISVsa3, forming a skeleton, ISVsa3-XerD-tet (X3)-res-ISVsa3. DISCUSSION: The co-occurrence of bla (NDM) and tet(X) in a novel species of the Acinetobacter genus hints that substantial undiscovered bacteria co-carrying high-risk ARGs are concealing in the agroecological system, which should cause particular concern.202540673007
3072170.9017Faecal microbiota and antibiotic resistance genes in migratory waterbirds with contrasting habitat use. Migratory birds may have a vital role in the spread of antimicrobial resistance across habitats and regions, but empirical data remain scarce. We investigated differences in the gut microbiome composition and the abundance of antibiotic resistance genes (ARGs) in faeces from four migratory waterbirds wintering in South-West Spain that differ in their habitat use. The white stork Ciconia ciconia and lesser black-backed gull Larus fuscus are omnivorous and opportunistic birds that use highly anthropogenic habitats such as landfills and urban areas. The greylag goose Anser anser and common crane Grus grus are herbivores and use more natural habitats. Fresh faeces from 15 individuals of each species were analysed to assess the composition of bacterial communities using 16S rRNA amplicon-targeted sequencing, and to quantify the abundance of the Class I integron integrase gene (intI1) as well as genes encoding resistance to sulfonamides (sul1), beta-lactams (bla(TEM), bla(KPC) and bla(NDM)), tetracyclines (tetW), fluoroquinolones (qnrS), and colistin (mcr-1) using qPCR. Bacterial communities in gull faeces were the richest and most diverse. Beta diversity analysis showed segregation in faecal communities between bird species, but those from storks and gulls were the most similar, these being the species that regularly feed in landfills. Potential bacterial pathogens identified in faeces differed significantly between bird species, with higher relative abundance in gulls. Faeces from birds that feed in landfills (stork and gull) contained a significantly higher abundance of ARGs (sul1, bla(TEM), and tetW). Genes conferring resistance to last resort antibiotics such as carbapenems (bla(KPC)) and colistin (mcr-1) were only observed in faeces from gulls. These results show that these bird species are reservoirs of antimicrobial resistant bacteria and suggest that waterbirds may disseminate antibiotic resistance across environments (e.g., from landfills to ricefields or water supplies), and thus constitute a risk for their further spread to wildlife and humans.202133872913
1821180.9016Emergence and dissemination of bla(KPC-31) and bla(PAC-2) among different species of Enterobacterales in Colombia: a new challenge for the microbiological laboratories. Ceftazidime/avibactam (CZA) is a promising treatment option for infections caused by carbapenem-resistant Enterobacterales (CRE). However, CZA resistance is increasingly reported worldwide, largely due to the emergence of KPC variants and increase of metallo-β-lactamases (MBL). This study describes the mechanisms associated with CZA resistance in circulating Enterobacterales isolates from Colombia, highlighting the challenge this represents for microbiological identification. Between 2021 and 2024, 68 CZA-resistant Enterobacterales isolates were identified by automated methods in seven Colombian cities. Resistance to CZA was subsequently confirmed by broth microdilution and E-test. Carbapenemase production was evaluated using phenotypic tests, such as the mCIM test, Carba NP, lateral flow assay, and qPCR (bla(KPC), bla(NDM), bla(VIM), bla(IMP), and bla(OXA-48)). Whole-genome sequencing was performed on 15 isolates that tested negative for MBL genes. Whole-genome sequencing of these 15 isolates revealed a variety of resistance determinants: six isolates harbored bla(KPC-31), one bla(KPC-33), one bla(KPC-8), five harbored bla(PAC-2), and two co-harbored bla(PAC-2) and bla(KPC-2). Notably, bla(PAC-2) was located on an IncQ plasmid. However, some of these variants were not detected by phenotypic assays, likely due to their low or undetectable carbapenemase activity. CZA resistance in non-MBL producing Enterobacterales in Colombia is primarily mediated by the presence of bla(KPC-31) and emergence of bla(PAC-2). These resistance mechanisms pose significant diagnostic, therapeutic, and epidemiological challenges, as they frequently go undetected by conventional microbiological methods. In this context, enhanced molecular surveillance and improved diagnostic strategies are urgently needed to enable early detection, guide antimicrobial therapy, and support infection control and stewardship efforts.IMPORTANCEAntibiotic resistance is a serious global health threat. Ceftazidime/avibactam (CZA) is a key treatment option for multidrug-resistant (MDR) Enterobacterales often used when other antibiotics fail. However, bacteria are now developing resistance to this drug as well, making infections increasingly difficult to treat. In this study, we examined CZA-resistant bacteria from multiple cities in Colombia and found uncommon resistance genes across several bacterial species. These genes are frequently missed, as they often do not test positive due to the limitations of most routinely used laboratory tests. Importantly, some of these genes can be transferred between bacteria, increasing the likelihood of indiscriminate dissemination in the hospital setting. Therefore, our findings highlight the urgent need for improved diagnostic tools and molecular surveillance. Early detection will help physicians select effective treatments quickly and prevent the wider dissemination of these MDR-resistant bacteria.202541070989
5160190.9015Multiomics analysis reveals the presence of a microbiome in the gut of fetal lambs. OBJECTIVE: Microbial exposure is critical to neonatal and infant development, growth and immunity. However, whether a microbiome is present in the fetal gut prior to birth remains debated. In this study, lambs delivered by aseptic hysterectomy at full term were used as an animal model to investigate the presence of a microbiome in the prenatal gut using a multiomics approach. DESIGN: Lambs were euthanised immediately after aseptic caesarean section and their cecal content and umbilical cord blood samples were aseptically acquired. Cecal content samples were assessed using metagenomic and metatranscriptomic sequencing to characterise any existing microbiome. Both sample types were analysed using metabolomics in order to detect microbial metabolites. RESULTS: We detected a low-diversity and low-biomass microbiome in the prenatal fetal gut, which was mainly composed of bacteria belonging to the phyla Proteobacteria, Actinobacteria and Firmicutes. Escherichia coli was the most abundant species in the prenatal fetal gut. We also detected multiple microbial metabolites including short chain fatty acids, deoxynojirimycin, mitomycin and tobramycin, further indicating the presence of metabolically active microbiota. Additionally, bacteriophage phiX174 and Orf virus, as well as antibiotic resistance genes, were detected in the fetal gut, suggesting that bacteriophage, viruses and bacteria carrying antibiotic resistance genes can be transmitted from the mother to the fetus during the gestation period. CONCLUSIONS: This study provides strong evidence that the prenatal gut harbours a microbiome and that microbial colonisation of the fetal gut commences in utero.202133589511