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Objectives: This study describes the detection and tracking of emergency neurosurgical cross-transmission infections with carbapenem-resistant Klebsiella oxytoca (CRKO)., Methods: We conducted an epidemiological investigation and a rapid screening of 66 surveillance samples using the chromogenic selective medium. Two CRKO isolates from infected patients and three from the preoperative shaving razors had similar resistance profiles identified by the clinical laboratory., Results: The whole genome sequencing (WGS) results identified all isolates as Klebsiella michiganensis (a species in the K. oxytoca complex) with sequence type 29 (ST29) and carrying resistance genes bla KPC-2 and bla OXY-5 , as well as IncF plasmids. The pairwise average nucleotide identity values of 5 isolates ranged from 99.993% to 99.999%. Moreover, these isolates displayed a maximum genetic difference of 3 among 5,229 targets in the core genome multilocus sequence typing scheme, and the razors were confirmed as the contamination source. After the implementation of controls and standardized shaving procedures, no new CRKO infections occurred., Conclusion: Contaminated razors can be sources of neurosurgical site infections with CRKO, and standard shaving procedures need to be established. Chromogenic selective medium can help rapidly identify targeted pathogens, and WGS technologies are effective mean in tracking the transmission source in an epidemic or outbreak investigation. Our findings increase the understanding of microbial transmission in surgery to improve patient care quality., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Jiang, Lyu, Liu, Li, Liu, Tai, Hu, Zhang, Chu, Zhao, Huang and Wu.)

The objective of this study was to describe the prevalence of antimicrobial resistance of Escherichia coli, Klebsiella oxytoca, Klebsiellapneumoniae, and Serratiamarcescens from quarter milk samples submitted to the udder health laboratory of the Bavarian Animal Health Services (TGD) in Southern Germany between 2014 and 2022. All samples were tested with the California Mastitis Test and analyzed with a standard microbroth dilution to determine the MIC. The antimicrobials tested were amoxicillin/clavulanate, cefazoline, kanamycin/cefalexin, cefoperazone, cefquinome, and marbofloxacin. Breakpoints were chosen in accordance with the Clinical and Laboratory Standards Institute (CLSI). Over the study period, E. coli, K. oxytoca, and K. pneumoniae showed only few resistances to all antimicrobials tested. For those pathogens MIC 50 and MIC 90 were below breakpoint for all antimicrobials except cefoperazone over the 9 years. A decrease in MIC could be seen for E. coli and K. oxytoca for all of the antimicrobials. While the MIC for K. pneumoniae stayed more stagnant, the prevalence of resistance still decreased overall. Serratiamarcescens isolates were proven intrinsically resistant to amoxicillin/clavulanate and cefazolin, and while in vitro resistances were low for all other antimicrobials tested, S. marcescens tended toward higher MIC for most of the antimicrobials over the years. Over time, there was also an overall increase in the number of isolates for all 4 pathogens per year. Starting 2018 there was a steep increase in the number of isolates particularly from clinical cases. This jump in numbers coincided with a change of the regulation for veterinary drug prescriptions in Germany in 2018 that required, among other things, antimicrobial resistance testing before a change of antibiotics in the course of treatment and the use of critically important antimicrobials. Overall, although the pathogens increased in numbers, the prevalence of their antimicrobial resistance remained low., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The Klebsiella oxytoca complex comprises diverse opportunistic bacterial pathogens associated with hospital and community-acquired infections with growing alarming antimicrobial resistance. We aimed to uncover the genomic features underlying the virulence and antimicrobial resistance of isolates from Mulago National Hospital in Uganda. We coupled whole genome sequencing with Pathogenwatch multilocus sequence typing (MLST) and downstream bioinformatic analysis to delineate sequence types (STs) capsular polysaccharide K- and O-antigen loci, along with antimicrobial resistance (AMR) profiles of eight clinical isolates from the National Referral Hospital of Uganda. Our findings revealed that only two isolates (RSM6774 and RSM7756) possess a known capsular polysaccharide K-locus (KL74). The rest carry various unknown K-loci (KL115, KL128, KLI52, KL161 and KLI63). We also found that two isolates possess unknown loci for the lipopolysaccharide O-antigen (O1/O2v1 type OL104 and unknown O1). The rest possess known O1 and O3 serotypes. From MLST, we found four novel sequence types (STs), carrying novel alleles for the housekeeping genes glyceraldehyde-6-phosphate dehydrogenase A (gapA), glucose-6-phosphate isomerase (pgi), and RNA polymerase subunit beta (rpoB). Our AMR analysis revealed that all the isolates are resistant to ampicillin and ceftriaxone, with varied resistance to other antibiotics, but all carry genes for extended-spectrum beta-lactamases (ESBLs). Notably, one strain (RSM7756) possesses outstanding chromosomal and plasmid-encoded AMR to beta-lactams, cephalosporins, fluoroquinolones and methoprims. Conclusively, clinical samples from Mulago National Referral Hospital harbor novel STs and multidrug resistant K. oxytoca strains, with significant public health importance, which could have been underrated., Competing Interests: Declaration of competing interest The authors have no conflict of interest to declare., (Copyright © 2023. Published by Elsevier B.V.)

Introduction: Multi-carbapenemase-producing Enterobacterales (M-CPE) are increasingly described. We characterized the M-CPE isolates prospectively recovered in our hospital (Madrid, Spain) over two years (2021-2022)., Methods: We collected 796 carbapenem resistant Enterobacterales (CRE) from clinical and surveillance samples. Carbapenemase production was confirmed with phenotypic (immunochromatographic, disk diffusion) and molecular (PCR, WGS) techniques. Antimicrobial susceptibility was evaluated by a standard broth microdilution method. Clinical and demographic data were collected., Results: Overall, 23 M-CPE (10 Klebsiella pneumoniae, 6 Citrobacter freundii complex, 3 Escherichia coli, 2 Klebsiella oxytoca, and 2 Enterobacter hormaechei) isolates were recovered from 17 patients (3% with CPE, 0.26-0.28 cases per 1000 admissions). OXA-48 + KPC-3 (7/23) and KPC-3 + VIM-1 (5/23) were the most frequent carbapenemase combinations. All patients had prior antibiotics exposure, including carbapenems (8/17). High resistance rates to ceftazidime/avibactam (14/23), imipenem/relebactam (16/23) and meropenem/vaborbactam (7/23) were found. Ceftazidime/avibactam + aztreonam combination was synergistic in all metallo-β-lactamase producers. Clonal and non-clonal related isolates were found, particularly in K. pneumoniae (5 ST29, 3 ST147, 3 ST307) and C. freundii (3 ST8, 2 ST125, 1 ST563). NDM-1 + OXA-48 was introduced with the ST147-K. pneumoniae high-risk clone linked to the transfer of a Ukrainian patient. We identified four possible nosocomial clonal transmission events between patients of the same clone with the same combination of carbapenemases (KPC-3 + VIM-1-ST29-K. pneumoniae, NDM-1 + OXA-48-ST147-K. pneumoniae and KPC-2 + VIM-1-ST145-K. oxytoca). Carbapenemase-encoding genes were located on different plasmids, except for VIM-1 + KPC-2-ST145-K. oxytoca. Cross-species transmission and a possible acquisition overtime was found, particularly between K. pneumoniae and E. coli producing OXA-48 + KPC-3., Conclusion: M-CPE is an emerging threat in our hospital. Co-production of different carbapenemases, including metallo-β-lactamases, limits therapeutic options and depicts the need to reinforce infection control measures., Competing Interests: Declaration of competing interest None., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: In this study, Escherichia coli, Klebsiella oxytoca, and Citrobacter amalonaticus carrying blaNDM-5 were isolated from a single patient., Methods: The antibiotic susceptibility of the isolates was evaluated by using E-test and agar dilution methods, and blaNDM-5 was identified in genomic and plasmid DNA by using polymerase chain reaction and sequencing. Whole genome sequencing and de novo assembly were used for species characterization, resistance gene identification, and plasmid analysis., Results: All three species had identical plasmids, which were similar to pEC463-NDM5, a plasmid harboring blaNDM-5. Transconjugation experiments confirmed the horizontal gene transfer of blaNDM-5, highlighting the need for a close monitoring of Enterobacteriaceae species harboring this gene., Conclusions: This study conclusively demonstrates the propensity for horizontal gene transfer of blaNDM-5 among Enterobacteriaceae species, underlining the importance of vigilant monitoring to combat antibiotic resistance.

Purpose: Raoultella spp. is a genus of bacteria that is known to be closely related to Klebsiella . It has been debated whether Raoultella should be reclassified as a subgroup of Klebsiella . The aim of this study is to compare clinical aspects of Raoultella and Klebsiella oxytoca , a species of Klebsiella that is known to be bacteriologically similar to Raoultella spp., Methods: Using data collected at a tertiary care hospital in the United States, we identified 43 patients with Raoultella infection and 1173 patients with Klebsiella oxytoca infection. We compared patient demographics (age and sex), hospitalization status, isolation sites and antibiotic resistance profiles between the two species., Results: There was no significant difference in patient demographics between the two bacteria species. The proportions of intensive care unit (ICU) admission were higher among patients with Raoultella infection (p=0.008). The most common site of isolation was urine for both species (39.5% of all patients with Raoultella spp. vs. 59.3% for K. oxytoca ). The second most common site of isolation was blood stream for Raoultella spp. (23.3%) and respiratory tract for K. oxytoca (10.8%). Except for the high proportion of resistant isolates of Raoultella spp. for Trimethoprim/sulfamethoxazole, the antibiotic susceptibility profiles were similar between the two bacteria species. Both were susceptible to ciprofloxacin and meropenem., Conclusion: While there are no significant differences in the patient demographics and antibiotic susceptibility profiles between Raoultella spp. and K. oxytoca , Raoultella may cause more serious infection requiring ICU admissions. Also, Raoultella may cause blood stream infection more frequently than K. oxytoca ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Mettler, Charoenngam and Colgrove.)

Probiotics produce small molecules that may serve as alternatives to conventional antibiotics by suppressing growth of antimicrobial resistant (AMR) pathogens. The objective of this study was to identify and examine antimicrobials produced and secreted by probiotics using 'omics' profiling with computer-based metabolic flux analyses. The cell-free supernatant of Gram-positive Lacticaseibacillus rhamnosus GG (LGG) and Gram-negative Escherichia coli Nissle (ECN) probiotics inhibited growth of AMR Salmonella Typhimurium, Escherichia coli , and Klebsiella oxytoca ranging between 28.85 - 41.20% (LGG) and 11.48 - 29.45% (ECN). A dose dependent analysis of probiotic supernatants showed LGG was 6.27% to 20.55% more effective at reducing AMR pathogen growth when compared to ECN. Principal component analysis showed clear separation of ECN and LGG cell free supernatant metabolomes. Among 667 metabolites in the supernatant, 304 were differentially abundant between LGG and ECN probiotics. Proteomics identified 87 proteins, whereby 67 (ECN) and 14 (LGG) showed differential expression as enzymes related to carbohydrate and energy metabolic pathways. The whole genomes and metabolomes were next used for in-silico metabolic network analysis. The model predicted the production of 166 metabolites by LGG and ECN probiotics across amino acid, carbohydrate/energy, and nucleotide metabolism with antimicrobial functions. The predictive accuracy of the metabolic flux analysis highlights the novel utility for profiling probiotic supplements as dietary-based antimicrobial alternatives in the control of AMR pathogen growth.

Background: Klebsiella spp. are opportunistic pathogens which can cause severe infections, are often multi-drug resistant and are a common cause of hospital-acquired infections. Multiple new Klebsiella species have recently been described, yet their clinical impact and antibiotic resistance profiles are largely unknown. We aimed to explore Klebsiella group- and species-specific clinical impact, antimicrobial resistance (AMR) and virulence., Methods: We analysed whole-genome sequence data of a diverse selection of Klebsiella spp. isolates and identified resistance and virulence factors. Using the genomes of 3594 Klebsiella isolates, we predicted the masses of 56 ribosomal subunit proteins and identified species-specific marker masses. We then re-analysed over 22,000 Matrix-Assisted Laser Desorption Ionization - Time Of Flight (MALDI-TOF) mass spectra routinely acquired at eight healthcare institutions in four countries looking for these species-specific markers. Analyses of clinical and microbiological endpoints from a subset of 957 patients with infections from Klebsiella species were performed using generalized linear mixed-effects models., Results: Our comparative genomic analysis shows group- and species-specific trends in accessory genome composition. With the identified species-specific marker masses, eight Klebsiella species can be distinguished using MALDI-TOF MS. We identified K. pneumoniae (71.2%; n = 12,523), K. quasipneumoniae (3.3%; n = 575), K. variicola (9.8%; n = 1717), "K. quasivariicola" (0.3%; n = 52), K. oxytoca (8.2%; n = 1445), K. michiganensis (4.8%; n = 836), K. grimontii (2.4%; n = 425) and K. huaxensis (0.1%; n = 12). Isolates belonging to the K. oxytoca group, which includes the species K. oxytoca, K. michiganensis and K. grimontii, were less often resistant to 4th-generation cephalosporins than isolates of the K. pneumoniae group, which includes the species K. pneumoniae, K. quasipneumoniae, K. variicola and "K. quasivariicola" (odds ratio = 0.17, p < 0.001, 95% confidence interval [0.09,0.28]). Within the K. pneumoniae group, isolates identified as K. pneumoniae were more often resistant to 4th-generation cephalosporins than K. variicola isolates (odds ratio = 2.61, p = 0.003, 95% confidence interval [1.38,5.06]). K. oxytoca group isolates were found to be more likely associated with invasive infection to primary sterile sites than K. pneumoniae group isolates (odds ratio = 2.39, p = 0.0044, 95% confidence interval [1.05,5.53])., Conclusions: Currently misdiagnosed Klebsiella spp. can be distinguished using a ribosomal marker-based approach for MALDI-TOF MS. Klebsiella groups and species differed in AMR profiles, and in their association with invasive infection, highlighting the importance for species identification to enable effective treatment options., (© 2021. The Author(s).)

A strain named as Pseudomonas aeruginosa 2016NX1, which could produce phenazine and cereusitin, was isolated from the root of Millettia specisoa. Phenazines were extracted, isolated and purified by chloroform, thin-layer chromatography, column chromatography and high-performance liquid chromatography. Then the purified materials were identified by analysis of nuclear magnetic resonance. The major yellow component is 1-hydroxyphenazine and the minor blue component is cereusitin A. The tests of antimicrobial activity of yellow component showed that the growth of several common plant pathogenic fungi and bacteria (such as Cochliobolus miyabeanus, Diaporthe citri, Salmonella sp., Klebsiella oxytoca) could be strongly inhibited. This study suggested that Pseudomonas aeruginosa strain 2016NX1 had a significant potential for biological control of phytopathogenic fungi. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, one bioactive substance from Pseudomonas aeruginosa 2016NX1 was identified and its antimicrobial activity was verified. This study demonstrated that one bioactive substance from P. aeruginosa can strongly inhibit the growth of plant pathogenic fungi and bacteria. This study suggested that P. aeruginosa strain 2016NX1 has a significant potential for biological control of phytopathogenic fungi., (© 2020 The Society for Applied Microbiology.)

Objective: Biofilm formation has made the therapy of bacterial infections more difficult. The objective our study was assessment of pan-drug-resistant (PDR) Klebsiella oxytoca pathogenicity and virulence factors causing AAHC in patients with colorectal cancer (CRC)., Materials and Methods: Among a total of 300 healthy and 300 patients with antibiotic-associated hemorrhagic colitis (AAHC) and CRC, 200 K. oxytoca were identified during May 2015-January 2019. The virulence properties and biofilm formation among the isolates were investigated by phenotypic, PCR, and real-time PCR (RT-qPCR) techniques., Results: The bla CTX-M1 (20%), bla SHV (11%), bla TEM1 (33%), and AmpC encoding CIT (2%) ESBL genes, carbapenemase-encoding genes blaIM (4%) and bla OXA-48 (2%), and colistin-resistant mcr-1 gene (2.5%) were detected. The virulence-encoding genes including fimA (80%), pilQ (100%), matB (100%), mrkA (80%), and npsB (100%) were amplified. Therefore, PDR K. oxytoca containing adhesins and toxin-encoding genes with ability of biofilm formation causing AAHC and CRC were isolated. There was a significant difference between healthy and patients with CRC regarding the presence of K. oxytoca (p = 00.221)., Conclusion: Bacterial enteric pathogens possibly play a role in CRC. Biofilm formation by K. oxytoca strains prevents the efficient infection elimination; therefore, rapid identification and control measure are chief requirements. Additionally, more investigations are necessary with this regard.

This paper presents the results of a pilot study of difficult-to-treat patients (exhibiting several previous treatment failures or detection of extended-spectrum beta-lactamase [ESBL] strains) with chronic bacterial prostatitis (CBP) who underwent treatment with fosfomycin trometamol (FT) and N-acetyl-L-cysteine (NAC). Twenty-eight patients with clinically- and microbiologically-confirmed CBP who attended a single urological institution between January 2018 and March 2019 were treated with oral administration of 3 g FT once a day for 2 days, followed by a dose of 3 g every 48 h for 2 weeks, in combination with oral administration of NAC 600 mg once a day for 2 weeks. Clinical and microbiological analyses were carried out at the time of admission (T0) and during follow-up at 1 month (T1) and 6 months (T2) after the end of treatment. Symptoms were assessed by the National Institutes of Health Chronic Prostatitis Symptom Index (NIH-CPSI) and International Prostatic Symptom Score (IPSS), and quality of life was assessed by Quality of Well-Being (QoL) questionnaires. Isolated strains were Escherichia coli (23 patients), Enterococcus spp. (3 patients), and Klebsiella oxytoca (2 patients). ESBL strain was found in 19 (67.8%) patients. Microbiological eradication was documented in 21 (75%) patients at the second follow-up visit and clinical cure was achieved in 20 (71.4%) patients. Significant changes on questionnaires were recorded between baseline and follow-up visits. Fifteen of 19 patients (78.9%) with ESBL strains were cured. No significant side effects were reported. FT in combination with NAC is a promising alternative therapy in difficult-to-treat CBP patients., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The expression of the bla KPC gene plays a key role in carbapenem resistance in Enterobacteriaceae However, the genetic regulators of the bla KPC gene have not been completely elucidated, especially the genes in Tn 3 -Tn 4401 chimeras. Two novel Tn 3 -Tn 4401 chimera isoforms were characterized in our hospital, isoform A (CTA), which harbors a 121-bp deletion containing the PX promoter and was present in 22.6% (54/239) of isolates, and isoform C (CTC), which harbors a 624-bp insertion and a P1 promoter deletion and was present in only 1 isolate. The carbapenem MICs of both isoforms were 2-fold or more higher than those of the wild type (Tn 3 -Tn 4401 chimera, CTB), and bla KPC was most highly expressed in CTA. Bioinformatics and 5' rapid amplification of cDNA ends (5' RACE) experiments indicated a novel strong putative promoter, PY, at the 3' end of the IS Kpn8 gene. PY mutation nearly abrogated bla KPC expression ( P  < 0.01) and restored carbapenem susceptibility in all 3 isoforms. Although the mutation of PX or P1 halved bla KPC expression in CTB ( P  < 0.05), PX deletion caused a 68% increase in bla KPC expression ( P  = 0.037) in CTA. The level of bla KPC mRNA in CTC was 8-fold higher than that in InCTC, which harbors P1 ( P  = 0.011). These results suggest that PY is a core promoter of the bla KPC gene in the chimeras and that the deletion of the PX and P1 promoters enhanced gene expression in CTA and CTC, respectively., (Copyright © 2020 American Society for Microbiology.)

Klebsiella sp. are responsible for a multitude of infectious diseases in both humans and animals. In this study, phylogenetic relationships, virulence and antimicrobial resistance gene properties of 16 Klebsiella sp. isolated from 49 pet turtles were investigated. The isolates including Klebsiella oxytoca (n = 13) and Klebsiella pneumoniae (n = 3) were identified using 16S rRNA gene sequencing and each species formed distinct clusters in the neighbour-joining phylogenetic tree. The prevalence of virulence genes including ureC (100%) and kfu (68·75%) was observed among the isolates using Polymerase chain reaction (PCR) assay. The fimH, mrkD and rmpA genes were detected in all K. pneumoniae while these were absent in every K. oxytoca isolate. In antimicrobial susceptibility testing, high resistance rates were observed against ampicillin (100%) and cephalothin (62·50%). The resistance rates against imipenem, tetracycline, trimethoprim/sulfamethoxazole, nalidixic acid and ciprofloxacin were 12·50, 12·50, 12·50, 6·25 and 6·25% respectively. The presence of antimicrobial resistance genes such as plasmid-mediated quinolone resistance (PMQR) [qnrB (37·50%), qnrA (31·25%), qnrS (12·50%) and aac(6')-Ib-cr (12·50%)], extended-spectrum β-lactamase (ESBL) [bla CTX-M (18·75%)], β-lactamase [bla SHV-1 (18·75%)] and tetracycline resistance [tetE (12·50%)] was observed. The results revealed that pet turtle-borne Klebsiella sp. may carry different types of virulence and antimicrobial resistance genes which represents a potential threat to public health. SIGNIFICANCE AND IMPACT OF THE STUDY: Klebsiella sp. are nonmotile Gram-negative bacteria that are found in different environments. The virulence and antimicrobial resistance properties of pet turtle-borne Klebsiella sp. have not been studied before. Phylogenetic relationships, virulence traits and antimicrobial resistance profiles of pet turtle-borne Klebsiella sp. were characterized for the first time in Korea. Multiple virulence and antimicrobial resistance genes were observed among the isolates. The occurrence of virulence and antimicrobial resistance determinants in Klebsiella sp. may represent a potential threat to public health., (© 2019 The Society for Applied Microbiology.)

Klebsiella oxytoca causes antibiotic-associated hemorrhagic colitis and diarrhea. This was attributed largely to its secreted cytotoxins tilivalline and tilimycin, inductors of epithelial apoptosis. To study whether Klebsiella oxytoca exerts further barrier effects, T84 monolayers were challenged with bacterial supernatants derived from tilivalline/tilimycin-producing AHC6 or its isogeneic tilivalline/tilimycin-deficient strain Mut-89. Both preparations decreased transepithelial resistance, enhanced fluorescein and FITC-dextran-4kDa permeabilities, and reduced expression of barrier-forming tight junction proteins claudin-5 and -8. Laser scanning microscopy indicated redistribution of both claudins off the tight junction region in T84 monolayers as well as in colon crypts of mice infected with AHC6 or Mut-89, indicating that these effects are tilivalline/tilimycin-independent. Furthermore, claudin-1 was affected, but only in a tilivalline/tilimycin-dependent manner. In conclusion, Klebsiella oxytoca induced intestinal barrier impairment by two mechanisms: the tilivalline/tilimycin-dependent one, acting by increasing cellular apoptosis and a tilivalline/tilimycin-independent one, acting by weakening the paracellular pathway through the tight junction proteins claudin-5 and -8.

During the period from April 2012 to May 2013, 13 newborns (1 to 4 weeks of age) and 1 child in a pediatric hospital ward in Germany were colonized with Klebsiella oxytoca producing an extended-spectrum beta-lactamase (ESBL) (CTX-M-15). A microbiological source-tracking analysis with human and environmental samples was carried out to identify the source and transmission pathways of the K. oxytoca clone. In addition, different hygienic intervention methods were evaluated. K. oxytoca isolates were detected in the detergent drawer and on the rubber door seal of a domestic washer-extractor machine that was used in the same ward to wash laundry for the newborns, as well as in two sinks. These strains were typed using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. The environmental findings were compared with those for the human strains and the isolates detected on clothing. The results from both techniques showed that the strains were identical (sequence type 201 and PFGE type 00531, a clone specific to this hospital and not previously isolated in Germany), emphasizing the washing machine as a reservoir and fomite for the transmission of these multidrug-resistant bacteria. After the washing machine was taken out of use, no further colonizations were detected during the subsequent 4-year period. IMPORTANCE Washing machines should be further investigated as possible sites for horizontal gene transfer (ESBL genes) and cross-contamination with clinically important Gram-negative strains. Particularly in the health care sector, the knowledge of possible (re-)contamination of laundry (patients' clothes and staff uniforms) with multidrug-resistant Gram-negative bacteria could help to prevent and to control nosocomial infections. This report describes an outbreak with a single strain of a multidrug-resistant bacterium ( Klebsiella oxytoca sequence type 201) in a neonatal intensive care unit that was terminated only when the washing machine was removed. In addition, the study implies that changes in washing machine design and processing are required to prevent accumulation of residual water where microbial growth can occur and contaminate clothes., (Copyright © 2019 Schmithausen et al.)

Background: Bacterial adhesins play an important role in the bacterial attachment and colonization. The aim of this study was comparison of adhesin genes expression in the planktonic and biofilm mode of growth among ESBL-non-producers isolates of K. oxytoca and effect of imipenem., Materials and Methods: A total of eight extended-spectrum beta-lactamase (ESBL) non-producer K. oxytoca isolates were included from patients with hemorrhagic colitis. The adhesin genes including fimA (type 1 fimbria), mrkA (type 3 fimbria), pilQ and the capsular matB genes were adopted. Phenotypic biofilm production was assessed by microtiter tissue plate assay. Expression of adhesin genes in the planktonic and biofilm growth conditions was calculated using quantitative Real-time PCR (RT-qPCR) technique and sub-MIC (0.25 μg/ml) levels of imipenem were also added to broth culture of isolates to evaluate the gene expression., Results: The isolates produced biofilm in moderate level. The expression of pilQ, mrkA and matB but not fimA genes was significantly higher in biofilm conditions compared to the planktonic mode of growth (p = 0.002, p = 0.011 and p = 001, respectively). In addition, imipenem sub-MIC treatment led to a significant overexpression of matB (p = 0.002) and mrkA (p = 0.003) genes compared to the control group., Conclusion: Although none of isolates produced strong biofilm, biofilm conditions led to the increase in the expression of adhesin encoding genes in non-ESBL-producing K. oxytoca. Furthermore, β-lactams; and especially carbapenems possibly increase the colonization of K. oxytoca and increase the biofilm formation. Hence, accurate consumption of antibiotics must be considered., (Copyright © 2019. Published by Elsevier Ltd.)

Advances in neonatal care have led to the increasing survival of smaller and sicker infants, but nosocomial infections continue to be a serious problem, associated with increased mortality rates, immediate and long-term morbidity, prolonged hospital stay, and increased cost of care. We report a case of hospital-acquired sepsis in a preterm baby secondary to Klebsiella oxytoca, resulting from contaminated intravenous fluid., (Copyright © 2019 Association for Professionals in Infection Control and Epidemiology, Inc. All rights reserved.)

Purpose: Biofilm formation and resistance to last-line antibiotics have restricted chemotherapy options toward infection eradication., Methodology: Fifty K. oxytoca isolates were collected from patients with antibiotic-associated haemorrhagic colitis (AAHC). Antibiotic susceptibility tests were conducted and phenotypic biofilm formation was assessed using microtitre tissue plate (MTP) assay. PCR was employed to amplify the adhesins, extended-spectrum β-lactamases (ESBLs), carbapenemase and colistin resistance genes. The expression of adhesin genes was evaluated using quantitative real-time PCR (RT-qPCR).Results/Key findings. The previous antibiotic consumption and hospitalization (P<0.05) and older ages (P=0.0033) were significantly associated with AAHC. None of the isolates produced biofilm strongly, but 70% of them produced moderate-level biofilm. The blaCTX-M (12/14), the blaIMP (8/14 MICIMI =4 µg ml -1 ) and blaOXA-48-like (5/14) and mcr-1 (4/14) genes were predominant, three of which harbouring all the genes. The expression of matB (0.023) and mrkA (0.011) was significantly different between multidrug-resistant and susceptible isolates. Furthermore, moderately biofilm producer isolates significantly exhibited higher expression of fimA (P=.0117), pilQ (P=0.002) and mrkA (P=0.020) genes compared to biofilm non-producers. No significant difference regarding gene expression was observed among ESBL alleles., Conclusion: Bacterial attachment by adhesins and biofilm formation among extensive drug-resistant K. oxytoca isolates hinder the efficient infection eradication. Hence, control and surveillance studies should be performed and other therapeutic auspicious approaches must be taken into account against AAHC, biofilm formation and drug resistance spread. Furthermore, previous antibiotic consumption and long-term hospitalization should be controlled.

A new series of copper (II), cobalt (II), zinc (II), nickel (II), manganese (II), iron (II) complexes with a novel Schiff base were synthesized by the condensation of sulphadizine and thiophene-2-carbaldehyde.The ligand and its complexes were characterized by using diverse instrumental procedures like microanalysis, thermo gravimetric examination and spectroscopy. The integrated ligand and its metal complexes were subjected to antibacterial studies. These studies demonstrated the enhanced activity of metal complexes against reported microbes with respect to the Schiff base.

The antibiotic susceptibility profile and antimicrobial resistance determinants were characterized on Gram-negative bacilli (GNB) isolated from Algerian hospital effluents. Among the 94 isolates, Enterobacteriaceae was the predominant family, with Escherichia coli and Klebsiella pneumoniae being the most isolated species. In non-Enterobacteriaceae, Acinetobacter and Aeromonas were the predominant species followed by Pseudomonas, Comamonas, Pasteurella, and Shewanella spp. The majority of the isolates were multidrug-resistant (MDR) and carried different antimicrobial resistance genes including bla CTX-M , bla TEM , bla SHV , bla OXA-48 -like, bla OXA-23 , bla OXA-51 , qnrB, qnrS, tet(A), tet(B), tet(C), dfrA1, aac(3)-IIc (aacC2), aac(6')-1b, sul1, and sul2. The qacEΔ1-sul1 and intI2 signatures of class 1 and class 2 integrons, respectively, were also detected. Microarray hybridization on MDR E. coli revealed additional resistance genes (aadA1 and aph3strA, tet30, mphA, dfrA12, bla cmy2 , bla ROB1 , and cmlA1) and classified the tested strains as commensals, thus highlighting the potential role of humans in antibiotic resistance dissemination. This study is the first report of bla OXA-48 -like in Klebsiella oxytoca in Algeria and bla OXA-23 in A. baumannii in Algerian hospital effluents. The presence of these bacteria and resistance genes in hospital effluents represents a serious public health concern since they can be disseminated in the environment and can colonize other hosts.

The worldwide emergence of antibiotic resistance poses a serious threat to human health. A molecular understanding of resistance strategies employed by bacteria is obligatory to generate less-susceptible antibiotics. Albicidin is a highly potent antibacterial compound synthesized by the plant-pathogenic bacterium Xanthomonas albilineans. The drug-binding protein AlbA confers albicidin resistance to Klebsiella oxytoca. Here we show that AlbA binds albicidin with low nanomolar affinity resulting in full inhibition of its antibacterial activity. We report on the crystal structure of the drug-binding domain of AlbA (AlbAS) in complex with albicidin. Both α-helical repeat domains of AlbAS are required to cooperatively clamp albicidin, which is unusual for drug-binding proteins of the MerR family. Structure-guided NMR binding studies employing synthetic albicidin derivatives give valuable information about ligand promiscuity of AlbAS. Our findings thus expand the general understanding of antibiotic resistance mechanisms and support current drug-design efforts directed at more effective albicidin analogs.

This study characterized the β-lactamase content of baseline pathogens recovered from patients with complicated urinary tract infections (cUTI), including acute pyelonephritis, who were enrolled in two phase 3 clinical trials of ceftazidime-avibactam (RECAPTURE 1 and 2), and correlated the clinical efficacy of ceftazidime-avibactam and the comparator doripenem according to resistance mechanisms. A total of 26.2% (93/355) ceftazidime-avibactam and 26.8% (101/377) doripenem patients had baseline isolates that met the MIC screening criteria. The majority of Enterobacteriaceae (87.5%; 154/176) carried bla CTX-M . This pattern was mainly observed in Escherichia coli (96.8%; 92/95) and Klebsiella pneumoniae (96.0%; 48/50), whereas most Proteus mirabilis (80.0%; 8/10) carried plasmid AmpC genes. Two K. pneumoniae and 1 Klebsiella oxytoca carried bla OXA-48 and 1 K. pneumoniae carried bla NDM-1 . Five (13/35; 37.1%) Pseudomonas aeruginosa isolates were screened, and 2 carbapenemase producers (IMP-18 and VIM-2) were detected. Among patients enrolled in the ceftazidime-avibactam arm who were infected by MIC screen-positive Enterobacteriaceae, clinical cure occurred in 85.7-95.5%, regardless of β-lactamase content; the respective rate in the doripenem arm was 82.1-92.5%. A total of 75.0% in the ceftazidime-avibactam arm and 100.0% in the doripenem arm of patients infected by P. aeruginosa with MIC screen-positive criteria were clinically cured. Ceftazidime-avibactam efficacy was comparable to doripenem efficacy for treating cUTI caused by uropathogens producing extended-spectrum and/or AmpC β-lactamases., (Copyright © 2018. Published by Elsevier B.V.)

The emergence of extended-spectrum β-lactamase-producing Enterobacteriaceae (ESBL-E) infections requires re-assessment of therapeutic choices. Here we report the efficacy of cefoxitin-based antibiotic therapy for ESBL-E prostatitis. A prospective study including patients with ESBL-E prostatitis resistant to trimethoprim/sulfamethoxazole and fluoroquinolones from January 2014 to March 2016 was conducted. Cefoxitin was administered by continuous infusion for 3 weeks in the case of acute bacterial prostatitis or 6 weeks in the case of chronic bacterial prostatitis (CBP), with intravenous fosfomycin for the first 5 days. Urological investigations were performed to diagnose underlying urinary tract pathology. Clinical and microbiological efficacy were evaluated 3 months (M3) and 6 months (M6) after the end of therapy. A total of 23 patients were included in the study. The median patient age was 74 years (range 48-88 years). Of the 23 infections, 14 (61%) were CBP and 12 (52%) were healthcare-associated infections. The bacteria involved were Escherichia coli in 11 cases, Klebsiella pneumoniae in 10 cases and Klebsiella oxytoca in 2 cases. Clinical cure was observed in 19/23 patients (83%) at M3 and in 17/22 patients (77%) at M6. Urocultures were sterile in 13/23 patients (57%) at M3 and in 9/19 patients (47%) and M6. Urinary colonisation was observed in 6/19 patients (32%) with clinical cure at M3 and 5/14 patients (36%) with clinical cure at M6. No resistance to cefoxitin was detected. Surgical treatment was required for 7/23 patients (30%). In conclusion, cefoxitin-based antibiotic therapy is suitable for difficult-to-treat ESBL-E infections such as prostatitis., (Copyright © 2018 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.)

Bacterial biofilms formed on equipment surfaces are potential sources of cross-contamination and can be responsible for the spread of bacteria involved in food spoilage, such as some Enterobacteriaceae family members. In this study, the effect of chlorite-based disinfectants, including sodium hypochlorite (SH), chlorine dioxide (CD), strongly acidic electrolyzed water (StAEW), and neutral electrolyzed water (NEW), on inactivation of mono-biofilms of Enterobacter cloacae, Klebsiella oxytoca, and Citrobacter freundii was evaluated separately. All the strains were enumerated by the viable plate-count method after disinfection for 30 min. A comparison of the surviving cells after disinfection indicated that E. cloacae biofilms were more resistant to disinfectants than the biofilms of the other two strains, and treatment with all the disinfectants improved sanitizing. SH (200 mg/L) was the most effective in the reduction of cell number in the biofilms of all strains. Considering the safety of use and environmental protection, electrolyzed oxidizing water, especially StAEW, was a good suggestion for the inactivation of cells in K. oxytoca or C. freundii biofilms. These results suggest that the cells in biofilm of E. cloacae, K. oxytoca, and C. freundii were highly sensitive to chlorite-based disinfectants and provide insights into the efficacy of disinfectants in killing bacteria., Practical Application: The Enterobacteriaceae biofilms formed on equipment surfaces, which can cause cross-contamination and food spoilage, are greatly challenging bacterial contaminants of food products. Electrolyzed oxidizing water is a novel, environmentally friendly disinfectant that can effectively treat Enterobacteriaceae biofilms. The results of this study may be used to design effective measures to disinfect biofilms on equipment contact surfaces., (© 2018 Institute of Food Technologists®.)

Background: The increasing incidence of carbapenem-resistant Enterobacteriaceae (CRE), has resulted in a difficult problem in the current clinical anti-infective treatment. We performed a retrospective analysis of prevalence and treatment for CRE infections patients., Methods: This study was conducted in three tertiary care hospitals from January 1, 2010 to December 30, 2016. Baseline data, treatment, and outcomes were collected in patients with ventilator-associated bacterial pneumonia (VABP), bacteremia, complicated urinary tract infection (cUTI)/acute pyelonephritis (AP), hospital-acquired bacterial pneumonia (HABP), superficial wound infection (SWI), biliary tract infection (BTI), deep wound infection (DWI) and sterile body fluids infection (SBFI) due to CRE., Results: One hundred twenty-four cases of CRE infection were identified: 31 VABP, 22 bacteremia, 18 cUTI/AP, 16 HABP, 16 SWI, 9 BTI, 7 DWI and 5 SBFI. The patient population had significant immunocompromised (33 of 124, 26.6%) and severe sepsis (43 of 124, 34.7%). The most common CRE pathogens were Klebsiella pneumoniae (84 of 124, 67.7%) and Enterobacter cloacae (24 of 124, 19.4%). And the production of IMP-type carbapenemase was the main antibiotic resistance mechanism. The majority of patients to take monotherapy for empiric therapy and dual therapy for direct treatment. Outcomes were universally poor (28-day mortality was 22.6%, 28 of 124) across all sites of infection., Conclusions: We identified a large number of cases of CRE infection in 7 years from different parts, most of these pathogens have been confirmed to produce IMP-type carbapenemases. The retrospective analysis of cases of such bacterial infections will help to control future infections of these pathogens. Despite the high mortality rate, we still found that the selection of quinolone antibiotics can be effective in the treatment of CRE producing IMP type enzymes.

This study describes highly aminoglycoside-resistant Klebsiella pneumoniae and Klebsiella oxytoca clinical isolates obtained from an inpatient in Okinawa, Japan, with no known record of traveling overseas. The minimum inhibitory concentrations of amikacin and arbekacin against these strains were >1024 μg/ml. Whole-genome sequencing analysis revealed that these isolates harbored armA, which encodes a 16S rRNA methylase, ArmA, that confers pan-aminoglycoside resistance. This is the second report of K. pneumoniae harboring armA and the first report of K. oxytoca harboring a 16S rRNA methylase encoding gene in Japan., (Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

The Rapid Polymyxin NP test has been recently developed to rapidly detect polymyxin resistance in Enterobacteriaceae. Here we evaluated this test for detecting MCR-1/MCR-2-producing Enterobacteriaceae using a collection of 70 non-redundant strains either recovered from the environment, animals, or humans. Sensitivity and specificity were found to be 100%., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Background: Bacterial biofilms exhibit reduced sensitivity to conventional antibiotics and host defence systems and contribute to the persistence of chronic bacterial infections., Hypothesis: The antibiofilm approach using plant alkaloids provides an alternative to antibiotic strategies., Study Design: In this study, the antibiofilm activities of various plant alkaloids were investigated against enterohemorrhagic Escherichia coli O157:H7 and Pseudomonas aeruginosa. In the subsequent investigation, the effects of five norharmane derivatives were investigated., Result: Harmaline significantly inhibited biofilm formation by E. coli O157:H7, P. aeruginosa PAO1, P. aeruginosa PA14, and Klebsiella oxytoca, and norharmane (β-carboline) was found to have antibiofilm activity. It was also found that functional groups at the C-1 and C-7 positions of norharmane could play important roles in its antibiofilm activity. Confocal and electron microscopic observations confirmed biofilm inhibition by harmaline and norharmane, and both reduced fimbriae production and swarming and swimming motilities. Furthermore, harmaline and norharmane attenuated the virulence of E. coli O157:H7 in a Caenorhabditis elegans nematode model., Conclusion: These findings strongly suggest that harmaline and norharmane could have potential use in antibiofilm strategy against persistent bacterial infections., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

The bioassay-guided fractionation of the ethyl acetate extract of the fungus Cochliobolus sp. highlighted leishmanicidal activity and allowed for anhydrocochlioquinone A (ANDC-A) isolation. MS, 1D and 2D NMR spectra of this compound were in agreement with those published in the literature. ANDC-A exhibited leishmanicidal activity with EC 50 value of 22.4 microgram/mL (44 mu M) and, when submitted to the microdilution assay against Gram-ositive and Gram-negative bacteria, showed a minimal inhibitory concentration against Staphylococcus aureus ATCC 25295 of 128 microgram/mL (248.7 mu M). It was also active against five human cancer cell lines, showing IC 50 values from 5.4 to 20.3 mu M. ANDC-A demonstrated a differential selectivity for HL-60 (SI 5.5) and THP-1 (SI 4.3) cell lines in comparison with Vero cells and was more selective than cisplatin and doxorubicin against MCF-7 cell line in comparison with human peripheral blood mononuclear cells. ANDC-A was able to eradicate clonogenic tumour cells at concentrations of 20 and 50 mu M and induced apoptosis in all tumour cell lines at 20 mu M. These results suggest that ANDC-A might be used as a biochemical tool in the study of tumour cells biochemistry as well as an anticancer agent with durable effects on tumours.

Cytotoxin-producing Klebsiella oxytoca is the causative agent of antibiotic-associated hemorrhagic colitis (AAHC). Recently, the cytotoxin associated with AAHC was identified as tilivalline, a known pentacyclic pyrrolobenzodiazepine (PBD) metabolite produced by K. oxytoca Although this assertion of tilivalline's role in AAHC is supported by evidence from animal experiments, some key aspects of this finding appear to be incompatible with toxicity mechanisms of known PBD toxins. We therefore hypothesized that K. oxytoca may produce some other uncharacterized cytotoxins. To address this question, we investigated whether tilivalline alone is indeed necessary and sufficient to induce cytotoxicity or whether K. oxytoca also produces other cytotoxins. LC-MS- and NMR-based metabolomic analyses revealed the presence of an abundant tricyclic PBD, provisionally designated kleboxymycin, in the supernatant of toxigenic K. oxytoca strains. Moreover, by generating multiple mutants with gene deletions affecting tilivalline biosynthesis, we show that a tryptophanase-deficient, tilivalline-negative K. oxytoca mutant induced cytotoxicity in vitro similar to tilivalline-positive K. oxytoca strains. Furthermore, synthetic kleboxymycin exhibited greater than 9-fold higher cytotoxicity than tilivalline in TC 50 cell culture assays. We also found that the biosynthetic pathways for kleboxymycin and tilivalline appear to overlap, as tilivalline is an indole derivative of kleboxymycin. In summary, our results indicate that tilivalline is not essential for inducing cytotoxicity observed in K. oxytoca -associated AAHC and that kleboxymycin is a tilivalline-related bacterial metabolite with even higher cytotoxicity., (© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.)

Objectives: Plazomicin, a novel aminoglycoside with in vitro activity against MDR Gram-negative organisms, is under development to treat patients with serious enterobacterial infections. We evaluated the activity of plazomicin and comparators against colistin-resistant enterobacterial isolates., Methods: Susceptibility to plazomicin and comparators was tested by broth microdilution for a collection of 95 colistin-resistant enterobacterial isolates collected from 29 hospitals in eight countries. Forty-two isolates (Klebsiella pneumoniae and Klebsiella oxytoca) possessed chromosomally encoded resistance mechanisms to colistin, 21 isolates (Escherichia coli and Salmonella enterica) expressed the mcr-1 gene, 8 isolates (Serratia, Proteus, Morganella and Hafnia) were intrinsically resistant to colistin and 24 isolates (K. pneumoniae, E. coli and Enterobacter spp.) had undefined, non-mcr-1 mechanisms. Susceptibility profiles were defined according to CLSI for aminoglycosides and to EUCAST for colistin and tigecycline., Results: Plazomicin inhibited 89.5% and 93.7% of the colistin-resistant enterobacterial isolates at ≤ 2 and ≤4 mg/L, respectively. MICs of plazomicin were ≤2 mg/L for all of the mcr-1 positive isolates and ≤4 mg/L for all the intrinsic colistin-resistant Enterobacteriaceae. Non-susceptibility to currently marketed aminoglycosides was common: amikacin, 16.8%; gentamicin, 47.4%; and tobramycin, 63.2%. Plazomicin was the most potent aminoglycoside tested with an MIC90 of 4 mg/L, compared with 32, >64 and 64 mg/L for amikacin, gentamicin and tobramycin, respectively., Conclusions: Plazomicin displayed potent activity against colistin-resistant clinical enterobacterial isolates, including those expressing the mcr-1 gene. Plazomicin was more active than other aminoglycosides against this collection of isolates. The further development of plazomicin for the treatment of infections due to MDR Enterobacteriaceae is warranted., (© The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

We investigated the efficacy of liposomal gentamicin formulations of different surface charges against Pseudomonas aeruginosa and Klebsiella oxytoca . The liposomal gentamicin formulations were prepared by the dehydration-rehydration method, and their sizes and zeta potential were measured. Gentamicin encapsulation efficiency inside the liposomal formulations was determined by microbiologic assay, and stability of the formulations in biologic fluid was evaluated for a period of 48 h. The minimum inhibitory concentration and the minimum bactericidal concentration were determined, and the in vitro time kill studies of the free form of gentamicin and liposomal gentamicin formulations were performed. The activities of liposomal gentamicin in preventing and reducing biofilm-forming P. aeruginosa and K. oxytoca were compared to those of free antibiotic. The sizes of the liposomal formulations ranged from 625 to 806.6 nm in diameter, with the zeta potential ranging from -0.22 to -31.7 mV. Gentamicin encapsulation efficiency inside the liposomal formulation ranged from 1.8% to 43.6%. The liposomes retained >60% of their gentamicin content during the 48 h time period. The minimum inhibitory concentration of neutral formulation was lower than that of free gentamicin (0.25 versus 1 mg/L for P. aeruginosa and 0.5 versus 1 mg/L for K. oxytoca ). The negatively charged formulation exhibited the same bacteriostatic concentration as that of free gentamicin. The minimum bactericidal concentration of neutral liposomes on planktonic bacterial culture was twofold lower than that of free gentamicin, whereas the negatively charged formulations were comparable to free gentamicin. The killing time curve values for the neutral negatively charged formulation against planktonic P. aeruginosa and K. oxytoca were better than those of free gentamicin. Furthermore, liposomal formulations prevent the biofilm-formation ability of these strains better than free gentamicin. In summary, liposomal formulations could be an effective lipid nanoparticle to combat acute infections where planktonic bacteria are predominant., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Competing Interests: Conflict of Interest: The authors declare no conflict of interest.

The nucleotide sequences of five plasmids from one Klebsiella oxytoca isolate were determined using the PacBio RS II system. Plasmid analysis revealed that bla NDM-1 was carried on an IncX3 plasmid. The bla IMP-4 and bla KPC-2 genes were located on IncN and IncP-6 plasmids, respectively. Comparative sequence analysis highlighted the successful spread of carbapenemase-harboring plasmids among different enterobacterial species. We report for the first time, to our knowledge, coproducing NDM-1, KPC-2, and IMP-4 carbapenemases on a K. oxytoca isolate., (Copyright © 2017 American Society for Microbiology.)

The R1 antibiotic resistance plasmid, originally discovered in a clinical Salmonella isolate in London, 1963, has served for decades as a key model for understanding conjugative plasmids. Despite its scientific importance, a complete sequence of this plasmid has never been reported. We present the complete genome sequence of R1 along with a brief review of the current knowledge concerning its various genetic systems and a comparison to the F and R100 plasmids. R1 is 97,566 nucleotides long and contains 120 genes. The plasmid consists of a backbone largely similar to that of F and R100, a Tn21-like transposon that is nearly identical to that of R100, and a unique 9-kb sequence that bears some resemblance to sequences found in certain Klebsiella oxytoca strains. These three regions of R1 are separated by copies of the insertion sequence IS1. Overall, the structure of R1 and comparison to F and R100 suggest a fairly stable shared conjugative plasmid backbone into which a variety of mobile elements have inserted to form an "accessory" genome, containing multiple antibiotic resistance genes, transposons, remnants of phage genes, and genes whose functions remain unknown., (Copyright © 2017 Elsevier Inc. All rights reserved.)

A carbapenemase-producing colistin-resistant Klebsiella oxytoca isolate was recovered from a blood culture of a female patient without previous report of risk factors to obtain multidrug-resistant Gram-negative bacilli. A combination of biochemical and molecular methods was used to identify the resistance mechanism of this isolate. Carbapenemase production was mediated by Verona integron-encoded metallo-β-lactamase (VIM)-2. Colistin resistance was not due to plasmid- borne mcr-1 gene, but we found an integration of IS5-like sequence in the mgrB gene of K. oxytoca. This gene is known to be an important regulator of the PhoPQ two-component system, and the disruption of this gene is most likely the cause of lipid A modification resulting in colistin resistance of our isolate. To the best of our knowledge this constitutes the first report of a carbapenemase-producing K. oxytoca with colistin resistance, a case that demonstrates the limited treatment options for infections with multidrug-resistant organisms., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

Ceftazidime-avibactam (CAZ-AVI) is a recently approved β-lactam-β-lactamase inhibitor combination with the potential to treat serious infections caused by carbapenem-resistant organisms. Few patients with such infections were included in the CAZ-AVI clinical trials, and clinical experience is lacking. We present a case series of patients with infections caused by carbapenem-resistant Enterobacteriaceae (CRE) or Pseudomonas aeruginosa (CRPa) who were treated with CAZ-AVI salvage therapy on a compassionate-use basis. Physicians who had prescribed CAZ-AVI completed a case report form. We used descriptive statistics to summarize patient characteristics and treatment outcomes. We used the Wilcoxon rank sum test and Fisher's exact test to compare patients by treatment outcome. The sample included 36 patients infected with CRE and two with CRPa. The most common infections were intra-abdominal. Physicians categorized 60.5% of patients as having life-threatening infections. All but two patients received other antibiotics before CAZ-AVI, for a median of 13 days. The median duration of CAZ-AVI treatment was 16 days. Twenty-five patients (65.8%) concurrently received other antibiotics to which their pathogen was nonresistant in vitro Twenty-eight patients (73.7%, 95% confidence interval [CI], 56.9 to 86.6%) experienced clinical and/or microbiological cure. Five patients (20.8%) with documented microbiological cure died, whereas 10 patients (71.4%) with no documented microbiological cure died (P = 0.01). In three-quarters of cases, CAZ-AVI (alone or combined with other antibiotics) cured infections caused by carbapenem-resistant organisms, 95% of which had failed previous therapy. Microbiological cure was associated with improved survival. CAZ-AVI shows promising clinical results for infections for which treatment options are limited., (Copyright © 2017 American Society for Microbiology.)

We describe a method for investigating wet foci in the hospital environment of a nosocomial outbreak due to carbapenem-resistant Klebsiella oxytoca. By searching in areas that are usually inaccessible, by immersing sampling swabs in thioglycolate broth, and by concentrating the sampled water, reservoirs of infection that would have gone unnoticed can be detected., (Copyright © 2016 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.)

The mammalian gastrointestinal tract is colonized by a high-density polymicrobial community where bacteria compete for niches and resources. One key competition strategy includes cell contact-dependent mechanisms of interbacterial antagonism, such as the type VI secretion system (T6SS), a multiprotein needle-like apparatus that injects effector proteins into prokaryotic and/or eukaryotic target cells. However, the contribution of T6SS antibacterial activity during pathogen invasion of the gut has not been demonstrated. We report that successful establishment in the gut by the enteropathogenic bacterium Salmonella enterica serovar Typhimurium requires a T6SS encoded within Salmonella pathogenicity island-6 (SPI-6). In an in vitro setting, we demonstrate that bile salts increase SPI-6 antibacterial activity and that S Typhimurium kills commensal bacteria in a T6SS-dependent manner. Furthermore, we provide evidence that one of the two T6SS nanotube subunits, Hcp1, is required for killing Klebsiella oxytoca in vitro and that this activity is mediated by the specific interaction of Hcp1 with the antibacterial amidase Tae4. Finally, we show that K. oxytoca is killed in the host gut in an Hcp1-dependent manner and that the T6SS antibacterial activity is essential for Salmonella to establish infection within the host gut. Our findings provide an example of pathogen T6SS-dependent killing of commensal bacteria as a mechanism to successfully colonize the host gut., Competing Interests: The authors declare no conflict of interest.

Antibiotic-associated diarrhoea (AAD) is a common and unintended consequence of antibiotic use. Clostridium difficile is the most common infectious aetiology of AAD; however, only approximately 25% of all AAD cases are associated with C. difficile infection, with the aetiology in the majority of cases remaining undetermined. Numerous other bacterial infectious agents have been implicated in AAD, including Clostridium perfringens, Staphylococcus aureus, and Klebsiella oxytoca. AAD is a complex disease that is influenced by the host, the infectious agent involved, and numerous clinical factors, including antibiotic treatment regimes. This review re-examines AAD and presents current perspectives on this disease, with a particular focus on the current understanding of bacterial causes other than C. difficile and the virulence factors involved in pathogenesis. VIDEO ABSTRACT., (Crown Copyright © 2016. Published by Elsevier Ltd. All rights reserved.)

Background and Study Aim: Colitis is a common complication after treatment with antibiotics such as β-lactams, quinolones, and aminoglycosides. Recently, Klebsiella oxytoca has been implicated in this type of diarrhoea. The prevalence and characterisations of K. oxytoca isolated from patients with antibiotic-associated diarrhoea were investigated. The K. oxytoca isolates were also tested for cytotoxin production., Patients and Methods: This study was conducted from May 2011 to Dec 2013. Faecal samples were collected from hospitalised patients receiving antibiotic treatment. Initial cultivation was performed on specific media. The clinical isolates were confirmed by polymerase chain reaction (PCR) using the specific K. oxytoca polygalacturonase (pehX) gene. The double-disc diffusion test was used to detect extended-spectrum beta-lactamase (ESBL)-producing strains. Tracking of ESBL-encoding genes was performed via PCR. The organism was cultured on Hep-2 cell lines for cytotoxin production., Results: Out of 331 samples collected from patients, 40 were confirmed molecularly to be clinical isolates of K. oxytoca. Fourteen (35%) ESBL-producing strains were isolated using the double-disc diffusion method. Among the molecularly confirmed K. oxytoca isolates, seven (17.5%) tested positive for the blaSHV gene, 12 (30%) for blaTEM, 10 (25%) for blaCTX-M, three (7.5%) for blaOXA, nine (22.5%) for blaCTX-M-15, and seven (17.5%) for blaTEM-1. Five (12%) isolates showed cytotoxin activity below 30%, 12 (30%) strains showed moderate cytotoxin activity between 30% and 60%, and 23 (58%) strains showed cytotoxin activity ⩾60%., Conclusions: The cytotoxin-producing K. oxytoca is found to be one of the causes of antibiotic-induced colitis. Discontinuing treatment and allowing normal intestinal flora to be established or prescribing appropriate medication after antibiogram can help patients with antibiotic-induced haemorrhagic colitis in a timely manner., (Copyright © 2016 Arab Journal of Gastroenterology. Published by Elsevier B.V. All rights reserved.)

The in vitro activity of ceftazidime-avibactam was evaluated against 34,062 isolates of Enterobacteriaceae from patients with intra-abdominal, urinary tract, skin and soft-tissue, lower respiratory tract, and blood infections collected in the INFORM (International Network For Optimal Resistance Monitoring) global surveillance study (176 medical center laboratories in 39 countries) in 2012 to 2014. Overall, 99.5% of Enterobacteriaceae isolates were susceptible to ceftazidime-avibactam using FDA approved breakpoints (susceptible MIC of ≤8 μg/ml; resistant MIC of ≥16 μg/ml). For individual species of the Enterobacteriaceae, the ceftazidime-avibactam MIC inhibiting ≥90% of isolates (MIC90) ranged from 0.06 μg/ml for Proteus species to 1 μg/ml for Enterobacter spp. and Klebsiella pneumoniae Carbapenem-susceptible isolates of Escherichia coli, K. pneumoniae, Klebsiella oxytoca, and Proteus mirabilis with a confirmed extended-spectrum β-lactamase (ESBL) phenotype, or a ceftazidime MIC of ≥16 μg/ml if the ESBL phenotype was not confirmed by clavulanic acid inhibition, were characterized further to identify the presence of specific ESBL- and plasmid-mediated AmpC β-lactamase genes using a microarray-based assay and additional PCR assays. Ceftazidime-avibactam demonstrated potent activity against molecularly confirmed ESBL-producing (n = 5,354; MIC90, 0.5 μg/ml; 99.9% susceptible), plasmid-mediated AmpC-producing (n = 246; MIC90, 0.5 μg/ml; 100% susceptible), and ESBL- and AmpC-producing (n = 152; MIC90, 1 μg/ml; 100% susceptible) isolates of E. coli, K. pneumoniae, K. oxytoca, and P. mirabilis We conclude that ceftazidime-avibactam demonstrates potent in vitro activity against globally collected clinical isolates of Enterobacteriaceae, including isolates producing ESBLs and AmpC β-lactamases., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

We determined the complete nucleotide sequence of a self-transmissible IncL/M plasmid, pKOI-34, from a Klebsiella oxytoca isolate. pKOI-34 possessed the core structure of an IncL/M plasmid found in Erwinia amylovora, pEL60, with two mobile elements inserted, a transposon carrying the arsenic resistance operon and a Tn21-like core module (tnp and mer modules) piggybacking blaIMP-34 as a class 1 integron, In808, where blaIMP-34 confers a resistance to carbapenems in K. oxytoca and Klebsiella pneumoniae., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Objectives: The objective of this study was to examine Klebsiella oxytoca clonal and phylogenetic diversity, based on an international collection of carriage isolates non-susceptible to expanded-spectrum cephalosporins (ESCs)., Methods: The study material comprised 68 rectal carriage K. oxytoca isolates non-susceptible to ESCs recovered in 2008-11 from patients in 14 hospitals across Europe and Israel. ESC resistance was tested phenotypically; genes encoding ESBLs, AmpC cephalosporinases and carbapenemases were amplified and sequenced. The isolates were typed by PFGE and MLST, followed by sequencing of blaOXY genes., Results: MLST and PFGE distinguished 34 STs and 47 pulsotypes among the isolates, respectively. Six STs were split into several pulsotypes each. Five STs were more prevalent (n = 2-9) and occurred in several countries each, including ST2, ST9 and ST141, which belong to a growing international clonal complex (CC), CC2. Four phylogenetic lineages were distinguished, each with another type of chromosomal OXY-type β-lactamase. Three of these, with OXY-1/-5, OXY-2 types and OXY-4, corresponded to previously described phylogroups KoI, KoII and KoIV, respectively. A single isolate from Israel represented a distinct lineage with a newly defined OXY-7 type. The phylogroups showed interesting differences in mechanisms of ESC resistance; KoI strains rarely overexpressed the OXY enzymes but commonly produced ESBLs, whereas KoII strains often were OXY hyperproducers and carried ESBLs much less frequently. AmpCs (DHA-1) and carbapenemases (VIM-1) occurred sporadically., Conclusions: The study confirmed the high genetic diversity of the collection of K. oxytoca ESC-non-susceptible isolates, composed of phylogroups with distinct types of OXY-type β-lactamases, and revealed some STs of broad geographical distribution., (© The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Carbapenemase-producing Enterobacteriaceae (CPE) cause serious infections and are associated with high mortality in part due to limited treatment options. The in vitro activities of the new aminoglycoside plazomicin and comparators were evaluated against a collection of 164 CPE (VIM-1, n=125; KPC-2, n=34; OXA-48, n=4; and IMP-22, n=1). MIC90 values of gentamicin, tobramycin and amikacin were 256, 64 and 16 mg/L, respectively. Plazomicin exhibited an MIC range of 0.12-4 mg/L with MIC50 and MIC90 values of 0.25 and 1 mg/L. The MICs of plazomicin did not correlate with the other aminoglycoside MICs, with the resistance phenotype or with the carbapenemase harboured. Chequerboard experiments against 10 carbapenemase-producing Klebsiella pneumoniae isolates showed that combinations of plazomicin with colistin yielded synergy against 60% of the strains. Synergy of plazomicin with meropenem or fosfomycin was detected against 20% and 25% of the isolates, respectively. Using time-kill methodology, the interactions of plazomicin at 2×, 1× and 0.5× MIC with meropenem, colistin, fosfomycin or tigecycline at steady-state concentrations against two K. pneumoniae carrying the VIM-1 enzyme were investigated. Bactericidal activity was evident for both isolates at 2× MIC of plazomicin. Synergy was observed when plazomicin was combined with meropenem, colistin or fosfomycin against both isolates, whilst the combination with tigecycline resulted in indifference. Antagonism was not observed for any of the combinations tested. The results of this study suggest that plazomicin may address the need for new therapeutic options for the treatment of infections due to CPE., (Copyright © 2015 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)

We characterized 12 clinical isolates of Klebsiella oxytoca with the extended-spectrum β-lactamase (ESBL) phenotype (high minimum inhibitory concentration [MIC] values of ceftriaxone) recovered over 9 months at a university hospital in Japan. To determine the clonality of the isolates, we used pulsed-field gel electrophoresis (PFGE), multi-locus sequence typing (MLST), and PCR analyses to detect blaRBI, which encodes the β-lactamase RbiA, OXY-2-4 with overproduce-type promoter. Moreover, we performed the isoelectric focusing (IEF) of β-lactamases, and the determination of the MICs of β-lactams including piperacillin/tazobactam for 12 clinical isolates and E. coli HB101 with pKOB23, which contains blaRBI, by the agar dilution method. Finally, we performed the initial screening and phenotypic confirmatory tests for ESBLs. Each of the 12 clinical isolates had an identical PFGE pulsotype and MLST sequence type (ST9). All 12 clinical isolates harbored identical blaRBI. The IEF revealed that the clinical isolate produced only one β-lactamase. E. coli HB101 (pKOB23) and all 12 isolates demonstrated equally resistance to piperacillin/tazobactam (MICs, >128 μg/ml). The phenotypic confirmatory test after the initial screening test for ESBLs can discriminate β-lactamase RbiA-producing K. oxytoca from β-lactamase CTX-M-producing K. oxytoca. Twelve clinical isolates of K. oxytoca, which were recovered from an outbreak at one university hospital, had identical genotypes and produced β-lactamase RbiA that conferred resistance to piperacillin/tazobactam. In order to detect K. oxytoca isolates that produce RbiA to promote research concerning β-lactamase RbiA-producing K. oxytoca, the phenotypic confirmatory test after the initial screening test for ESBLs would be useful.

Microbial fermentation produces a racemic mixture of 2,3-butanediol ((R,R)-BD, (S,S)-BD, and meso-BD), and the compositions and physiochemical properties vary from microorganism to microorganism. Although the meso form is much more difficult to transport and store because of its higher freezing point than those of the optically active forms, most microorganisms capable of producing 2,3-BD mainly yield meso-2,3-BD. Thus, we developed a metabolically engineered (R,R)-2,3-BD overproducing strain using a Klebsiella oxytoca ΔldhA ΔpflB strain, which shows an outstanding 2,3-BD production performance with more than 90 % of the meso form. A budC gene encoding 2,3-BD dehydrogenase in the K. oxytoca ΔldhA ΔpflB strain was replaced with an exogenous gene encoding (R,R)-2,3-BD dehydrogenase from Paenibacillus polymyxa (K. oxytoca ΔldhA ΔpflB ΔbudC::PBDH strain), and then its expression level was further amplified with using a pBBR1MCS plasmid. The fed-batch fermentation of the K. oxytoca ΔldhA ΔpflB ΔbudC::PBDH (pBBR-PBDH) strain with intermittent glucose feeding allowed the production of 106.7 g/L of (R,R)-2,3-BD [meso-2,3-BD, 9.3 g/L], with a yield of 0.40 g/g and a productivity of 3.1 g/L/h, which should be useful for the industrial application of 2,3-BD.