Phenotypic and molecular characterization of ß-lactamase-producing Klebsiella species among children discharged from hospital in Western Kenya.

BACKGROUND: The emergence and spread of ß-lactamase-producing Klebsiella spp. has been associated with a substantial healthcare burden resulting in therapeutic failures. We sought to describe the proportion of phenotypic resistance to commonly used antibiotics, characterize ß-lactamase genes among isolates with antimicrobial resistance (AMR), and assess the correlates of phenotypic AMR in Klebsiella spp. isolated from stool or rectal swab samples collected from children being discharged from hospital. METHODS: We conducted a cross-sectional study involving 245 children aged 1-59 months who were being discharged from hospitals in western Kenya between June 2016 and November 2019. Whole stool or rectal swab samples were collected and Klebsiella spp. isolated by standard microbiological culture. ß-lactamase genes were detected by PCR whilst phenotypic antimicrobial susceptibility was determined using the disc diffusion technique following standard microbiology protocols. Descriptive analyses were used to characterize phenotypic AMR and carriage of ß-lactamase-producing genes. The modified Poisson regression models were used to assess correlates of phenotypic beta-lactam resistance. RESULTS: The prevalence of ß-lactamase carriage among Klebsiella spp. isolates at hospital discharge was 62.9% (154/245). Antibiotic use during hospitalization (adjusted prevalence ratio [aPR] = 4.51; 95%CI: 1.79-11.4, p < 0.001), longer duration of hospitalization (aPR = 1.42; 95%CI: 1.14-1.77, p < 0.002), and access to treated water (aPR = 1.38; 95%CI: 1.12-1.71, p < 0.003), were significant predictors of phenotypically determined ß-lactamase. All the 154 ß-lactamase-producing Klebsiella spp. isolates had at least one genetic marker of ß-lactam/third-generation cephalosporin resistance. The most prevalent genes were blaCTX-M 142/154 (92.2%,) and blaSHV 142/154 (92.2%,) followed by blaTEM 88/154 (57.1%,) and blaOXA 48/154 (31.2%,) respectively. CONCLUSION: Carriage of ß-lactamase producing Klebsiella spp. in stool is common among children discharged from hospital in western Kenya and is associated with longer duration of hospitalization, antibiotic use, and access to treated water. The findings emphasize the need for continued monitoring of antimicrobial susceptibility patterns to inform the development and implementation of appropriate treatment guidelines. In addition, we recommend measures beyond antimicrobial stewardship and infection control within hospitals, improved sanitation, and access to safe drinking water to mitigate the spread of ß-lactamase-producing Klebsiella pathogens in these and similar settings.

Antimicrobial Resistance as Risk Factor for Recurrent Bacteremia after Staphylococcus aureus, Escherichia coli, or Klebsiella spp. Community-Onset Bacteremia.

We investigated links between antimicrobial resistance in community-onset bacteremia and 1-year bacteremia recurrence by using the clinical data warehouse of Europe's largest university hospital group in France. We included adult patients hospitalized with an incident community-onset Staphylococcus aureus, Escherichia coli, or Klebsiella spp. bacteremia during 2017-2019. We assessed risk factors of 1-year recurrence using Fine-Gray regression models. Of the 3,617 patients included, 291 (8.0%) had >1 recurrence episode. Third-generation cephalosporin (3GC)-resistance was significantly associated with increased recurrence risk after incident Klebsiella spp. (hazard ratio 3.91 [95% CI 2.32-6.59]) or E. coli (hazard ratio 2.35 [95% CI 1.50-3.68]) bacteremia. Methicillin resistance in S. aureus bacteremia had no effect on recurrence risk. Although several underlying conditions and infection sources increased recurrence risk, 3GC-resistant Klebsiella spp. was associated with the greatest increase. These results demonstrate a new facet to illness induced by 3GC-resistant Klebsiella spp. and E. coli in the community setting.

Klebsiella species: Taxonomy, hypervirulence and multidrug resistance.

Members of the genus Klebsiella have rapidly evolved within the past decade, generating organisms that simultaneously exhibit both multidrug resistance and hypervirulence (MDR-hv) phenotypes; such organisms are associated with severe hospital- and community-acquired infections. Carbapenem-resistant infections with unknown optimal treatment regime were of particular concern among the MDR-hv Klebsiella strains. Recent studies have revealed the molecular features and the mobile resistance elements they harbour, allowing identification of genetic loci responsible for transmission, stable inheritance, and expression of mobile resistance or virulence-encoding elements that confer the new phenotypic characteristics of MDR-hv Klebsiella spp. Here, we provide a comprehensive review on the taxonomic position, species composition and different phylotypes of Klebsiella spp., describing the diversity and worldwide distribution of the MDR-hv clones, the genetic mutation and horizontal gene transfer events that drive the evolution of such clones, and the potential impact of MDR-hv infections on human health.

Correlation between antibiotic consumption and the occurrence of multidrug-resistant organisms in a Malaysian tertiary hospital: a 3-year observational study.

Inappropriate use of antibiotics has been shown to contribute to the occurrence of multidrug-resistant organisms (MROs). A surveillance study was performed in the largest tertiary care hospital in Kuala Lumpur, Malaysia, from 2018 to 2020 to observe the trends of broad-spectrum antibiotics (beta-lactam/beta-lactamases inhibitors (BL/BLI), extended-spectrum cephalosporins (ESC), and fluoroquinolones (FQ)) and antibiotics against MRO (carbapenems, polymyxins, and glycopeptides) usage and the correlation between antibiotic consumption and MROs. The correlation between 3-year trends of antibiotic consumption (defined daily dose (DDD)/100 admissions) with MRO infection cases (per 100 admissions) was determined using a Jonckheere-Terpstra test and a Pearson's Correlation coefficient. The antimicrobial resistance trend demonstrated a positive correlation between ESC and FQ towards the development of methicillin-resistant Staphylococcus aureus (MRSA), extended-spectrum beta-lactamase (ESBL)-producing Klebsiella spp, ESBL-producing Escherichia coli (E. coli), and MRO Acinetobacter baumannii (A. baumannii). Increasing carbapenem consumption was positively correlated with the occurrence of ESBL-producing Klebsiella spp and E. coli. Polymyxin use was positively correlated with ESBL-producing Klebsiella spp, MRO A. baumannii, and carbapenem-resistant Enterobacteriaceae. The findings reinforced concerns regarding the association between MRO development, especially with a surge in ESC and FQ consumption. Stricter use of antimicrobials is thus crucial to minimise the risk of emerging resistant organisms.

Genetic Characteristics and Microbiological Profile of Hypermucoviscous Multidrug-Resistant Klebsiella variicola Coproducing IMP-4 and NDM-1 Carbapenemases.

We report here a hypermucoviscous, New Delhi metallo-ß-lactamase 1 (NDM-1) and imipenemase 4 (IMP-4) carbapenemases-coproducing Klebsiella variicola isolate obtained from a pediatric patient. This strain was resistant to carbapenems and most other ß-lactams. Although hypermucoviscous, this strain possessed attenuated virulence according to serum killing assay and Galleria mellonella infection model. Notably, two copies of blaNDM-1 were contained on two tandem ISCR1 elements and coexisted with blaIMP-4 in a novel hybrid multidrug resistance plasmid. This is the first description of the coexistence of blaNDM-1 and blaIMP-4 in a single plasmid of hypermucoviscous K. variicola. IMPORTANCE As an important member of the Klebsiella pneumoniae complex, Klebsiella variicola is poorly studied as an emerging human pathogen. We, for the first time, report a unique K. variicola isolated from a pediatric patient in China. This isolate exhibited hypermucoviscosity, a classic hypervirulence characteristic of K. pneumoniae, and contained multiple carbapenem-resistant genes, including blaIMP-1 and blaNDM-1. Interestingly, these antimicrobial resistance genes were located on a novel hybrid plasmid, and our results suggested that this plasmid might have been introduced from K. pneumoniae and undergone a series of integration and recombination evolutionary events. Overall, our study provides more insight into K. variicola and highlights its superior capability to acquire and maintain foreign resistance genes.

Evaluation of Rapid Immunochromatographic Tests for the Direct Detection of Extended Spectrum Beta-Lactamases and Carbapenemases in Enterobacterales Isolated from Positive Blood Cultures.

NG-Test CTX-M MULTI and NG-Test Carba 5 (NG Biotech) are two rapid in vitro immunochromatographic assays that are widely used for the detection of the most common extended spectrum beta-lactamases (ESBL) and carbapenemases in Enterobacterales. ESBL and carbapenemases are leading causes of morbidity and mortality worldwide and their rapid detection from positive blood cultures is crucial for early initiation of effective antimicrobial therapy in bloodstream infections (BSI) involving antibiotic-resistant organisms. In this study, we developed a rapid workflow for positive blood cultures for direct identification of Enterobacterales by MALDI-TOF mass-spectrometry, followed by detection of ESBL and carbapenemases using NG-Test CTX-M MULTI and NG-Test Carba 5 (NG Biotech). The workflow was evaluated using Enterobacterales isolates (n = 114), primarily Klebsiella species (n = 50) and Escherichia coli (n = 40). Compared to the standard testing approach in our institution using BD Phoenix, our new testing approach demonstrates 100% sensitivity and specificity for organism identification and detection of ESBL and carbapenemases. Implementation of a rapid workflow in diagnostic microbiology laboratories will enable more effective antimicrobial management of patients with BSI due to ESBL- and carbapenemase-producing Enterobacterales. IMPORTANCE The incidence of bloodstream infections (BSI) with extended spectrum beta-lactamase (ESBL) producing and carbapenemase producing Enterobacterales (CPE) is increasing at an alarming rate, for which only limited therapeutic options remain available. Rapid identification of these bacteria along with their antibiotic resistance mechanisms in positive blood cultures with Gram-negative bacteria will allow for early initiation of effective therapy and limit the overuse of broad-spectrum antibiotics in BSI (1). In this study we evaluated a combined approach of testing positive blood cultures directly, using MALDI-TOF MS followed by rapid immunochromatographic tests, for the detection of ESBLs and CPEs. Our approach demonstrates 100% sensitivity and specificity for the identification of Enterobacterales and detection of ESBLs and CPEs in positive blood culture with a turnaround time (TAT) of ≤60 min compared to a TAT of 48 h required by conventional culture and susceptibility testing methods.

Polymerization of C9 enhances bacterial cell envelope damage and killing by membrane attack complex pores.

Complement proteins can form membrane attack complex (MAC) pores that directly kill Gram-negative bacteria. MAC pores assemble by stepwise binding of C5b, C6, C7, C8 and finally C9, which can polymerize into a transmembrane ring of up to 18 C9 monomers. It is still unclear if the assembly of a polymeric-C9 ring is necessary to sufficiently damage the bacterial cell envelope to kill bacteria. In this paper, polymerization of C9 was prevented without affecting binding of C9 to C5b-8, by locking the first transmembrane helix domain of C9. Using this system, we show that polymerization of C9 strongly enhanced damage to both the bacterial outer and inner membrane, resulting in more rapid killing of several Escherichia coli and Klebsiella strains in serum. By comparing binding of wildtype and 'locked' C9 by flow cytometry, we also show that polymerization of C9 is impaired when the amount of available C9 per C5b-8 is limited. This suggests that an excess of C9 is required to efficiently form polymeric-C9. Finally, we show that polymerization of C9 was impaired on complement-resistant E. coli strains that survive killing by MAC pores. This suggests that these bacteria can specifically block polymerization of C9. All tested complement-resistant E. coli expressed LPS O-antigen (O-Ag), compared to only one out of four complement-sensitive E. coli. By restoring O-Ag expression in an O-Ag negative strain, we show that the O-Ag impairs polymerization of C9 and results in complement-resistance. Altogether, these insights are important to understand how MAC pores kill bacteria and how bacterial pathogens can resist MAC-dependent killing.

An efficient method for synthesis, characterization and molecular docking study of new sulfamethoxazole derivatives as antibacterial agents.

A new series of sulfamethoxazole derivatives bearing some biologically active heterocycles such as pyrazole (2), 3,4-dihydropyrimidin (3-7, 11, 12), pyrrole (9) and 1,3-dihydropyrimidin (10) rings were successfully synthesized. Identification of designed compounds was done by physicochemical properties and spectral measurements (1H-NMR, 13C-NMR and FT-IR). New prepared derivatives were assay for their (in vitro) antibacterial efficacy against four types of pathogenic bacterial isolates. Significant of the newly prepared compounds appeared promising activity comparison to the cephalexin standard drug. Most of the active compounds are docked into the effective site of tested bacterial enzymes obtained by crystal structure; results reveal the binding template to enzymes of bacteria, which closely related to the laboratory results.

A new mutation in mgrb mediating polymyxin resistance in Klebsiella variicola.

Polymyxin resistance is a public health concern - present in humans, animals and the environment - caused by chromosomal-encoding or plasmid-encoding mechanisms. Chromosomal alterations in MgrB are frequently detected in Klebsiella spp., but not yet reported and characterised in Klebsiella variicola (K. variicola). This study performed microbiological and genomic characterisation of three polymyxin-resistant K. variicola isolates (M14, M15 and M50) recovered from the microbiota of migratory birds in Brazil. The isolates were submitted to SpeI-PFGE, broth microdilution and whole genome sequencing using Illumina MiSeq for analysis of genetic relatedness, sequence typing and detection of antimicrobial-resistance genes. K. variicola isolates belonged to two clones, and susceptibility tests showed resistance only for polymyxins. Sequences of chromosomal two-component systems (PmrAB, PhoPQ, RstAB, CrrAB) and MgrB were evaluated by blastN and blastP against a polymyxin-susceptible K. variicola (A58243), and mutations with biological effect were checked by the PROVEAN tool. K. variicola isolates belonged to two clones, and susceptibility tests showed resistance for polymyxins. In M14 and M15, phoQ deleterious mutations (D90N, I122S and G385S) were identified, while an mgrB variant containing a single deletion (C deletion on position 93) leading to the production of a non-functional protein was detected in M50. mgrB complementation studies showed restoration of polymyxin susceptibility (64 to ≤ 0.25 mg/L) as a wild-type mgrB was inserted into the mgrB-deficient M50. This study confirmed the role of a non-functional mgrB variant in conferring polymyxin resistance, stressing the role of this regulator in K. variicola and drawing attention to novel polymyxin resistance mechanisms emerging in wildlife.

Rapid detection of fosfomycin resistance in Escherichia coli and Klebsiella spp. strains isolated from urinary tract infections.

This study evaluates whether the rapid fosfomycin resistance (fosfomycin NP) method can be used for detecting fosfomycin resistance in routine laboratory work. Results from the disk diffusion and rapid fosfomycin NP methods were compared with the reference agar dilution method for Escherichia coli and Klebsiella spp. strains isolated from urinary tract infections. The study included 57 E. coli and 48 Klebsiella spp. isolates from urinary tract infections. The reference agar dilution and disk diffusion methods were performed in accordance with EUCAST recommendations, and the results were evaluated according to EUCAST V.10.0. The method developed by Nordmann et al. was used for rapid detection of fosfomycin resistance (Nordmann, P., Poirel, L., Mueller, L., 2019. Rapid Detection of Fosfomycin Resistance in Escherichia coli. J Clin Microbiol. 57(1), e01531-18. doi:https://doi.org/10.1128/JCM.01531-18). The acceptable categorical agreement (CA ≥ 90%) and the rates of major error (ME <3%) and very major error (VME < 3%) of the two methods were compared with the reference method according to the criteria of ISO 20776-1. Fosfomycin resistance was detected in 15.8% of E. coli and 75% of Klebsiella spp. isolates using the reference method. Disk diffusion method showed CA 89.5%, ME 12.5% in E. coli isolates, and CA 75%, ME 100% in Klebsiella spp. isolates. No VME was detected in both methods. The rapid fosfomycin NP method resulted in CA 96.4%, ME 0.0%, VME 22.2% in E. coli isolates, and CA 77.3%, ME 81.8%, and VME 3% in Klebsiella spp. isolates. We believe the results from both of disk diffusion assay and rapid fosfomycin NP for the E. coli and Klebsiella spp. isolates are incompatible with the reference method and should not be used as an alternative to the agar dilution method.

Klebsiella spp. cause severe and fatal disease in Mozambican children: antimicrobial resistance profile and molecular characterization.

BACKGROUND: Klebsiella spp. are important pathogens associated with bacteremia among admitted children and is among the leading cause of death in children < 5 years in postmortem studies, supporting a larger role than previously considered in childhood mortality. Herein, we compared the antimicrobial susceptibility, mechanisms of resistance, and the virulence profile of Klebsiella spp. from admitted and postmortem children. METHODS: Antimicrobial susceptibility and virulence factors of Klebsiella spp. recovered from blood samples collected upon admission to the hospital (n = 88) and postmortem blood (n = 23) from children < 5 years were assessed by disk diffusion and multiplex PCR. RESULTS: Klebsiella isolates from postmortem blood were likely to be ceftriaxone resistant (69.6%, 16/23 vs. 48.9%, 43/88, p = 0.045) or extended-spectrum ß-lactamase (ESBL) producers (60.9%, 14/23 vs. 25%, 22/88, p = 0.001) compared to those from admitted children. blaCTX-M-15 was the most frequent ESBL gene: 65.3%, 9/14 in postmortem isolates and 22.7% (5/22) from admitted children. We found higher frequency of genes associated with hypermucoviscosity phenotype and invasin in postmortem isolates than those from admitted children: rmpA (30.4%; 7/23 vs. 9.1%, 8/88, p = 0.011), wzi-K1 (34.7%; 8/23 vs. 8%; 7/88, p = 0.002) and traT (60.8%; 14/23 vs. 10.2%; 9/88, p < 0.0001), respectively. Additionally, serine protease auto-transporters of Enterobacteriaceae were detected from 1.8% (pic) to 12.6% (pet) among all isolates. Klebsiella case fatality rate was 30.7% (23/75). CONCLUSION: Multidrug resistant Klebsiella spp. harboring genes associated with hypermucoviscosity phenotype has emerged in Mozambique causing invasive fatal disease in children; highlighting the urgent need for prompt diagnosis, appropriate treatment and effective preventive measures for infection control.

Antimicrobial resistance of Escherichia coli and Klebsiella spp. conventionally sampled from factory-farmed chickens - clinical submissions.

INTRODUCTION: The article discusses the antimicrobial resistance of poultry-isolated bacteria in the Wielkopolska region of Poland. MATERIAL AND METHODS: From August 2014 - June 2016, antibiotic resistance screening tests were performed involving 4,496 samples of Escherichia coli and 84 samples of Klebsiella spp., and the following antibiotics: amoxicillin, amoxicillin with clavulanic acid, colistin, doxycycline, enrofloxacin, florfenicol, neomycin, norfloxacin, spectinomycin, and trimethoprim with sulfamethoxazole. The research used broth the microdilution method and CLSI standards. RESULTS: During the investigation period of 22 months a growing percentage of E. coli isolates showed antibiotic resistance to amoxicillin, amoxicillin with clavulanic acid, colistin, enrofloxacin, neomycin, norfloxacin, spectinomycin, and trimethoprim with sulfamethoxazole. Resistance to doxycycline and florfenicol decreased. The most efficient antibiotics against E. coli were colistin (84.64 %), neomycin (80.62 %), and amoxicillin with clavulanic acid (73.05 %). Klebsiella samples were the most susceptible to neomycin (85.71 %), colistin (84.52 %), and trimethoprim with sulfamethoxazole (73.81 %). CONCLUSIONS: Antibiotic resistance of pathogenic micro-organisms, such as Escherichia coli and Klebsiella spp., is a serious problem both for poultry producers and for public health protection. Low efficiency of numerous antibiotic groups forces reflection on limiting the use of medicines in food-producing animals.

Comparative studies of chitosan and carboxymethyl chitosan doped with nickel and copper: Characterization and antibacterial potential.

The study focused on the preparation and antibacterial evaluation of chitosan (CHT), carboxymethyl chitosan (CMC) and their respective metal composites. All the samples were characterized using Fourier Transform Infrared (FTIR), scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDS) and X-ray diffraction (XRD). The antibacterial potentials of the samples were tested against Escherichia coli, Klebsiella sp., Pseudomonas aeruginosa A, and Pseudomonas aeruginosa B. SEM results revealed different changes in samples surfaces as a result of chemical modification. EDS revealed the presence of Ni and Cu in the composites. XRD spectra of CMC showed that the crystalline region of CHT was reduced by the modification. The antibacterial results indicated that the samples had inhibitory and bactericidal effects against Escherichia coli and Klebsiella sp. at 1000, 500, and 250 mg mL-1. The study showed that CMC and CMC-metal composites performed better at inhibiting the growth of microorganisms than CHT and CHT-metal composites.

Performance of a MALDI-TOF mass spectrometry-based method for rapid detection of third-generation oxymino-cephalosporin-resistant Escherichia coli and Klebsiella spp. from blood cultures.

We optimized and prospectively evaluated a simple MALDI-TOF MS-based method for direct detection of third-generation oxymino-cephalosporin resistance (3rd CephR) in Escherichia coli and Klebsiella spp. from blood cultures (BC). In addition, we assessed the performance of a lateral flow immunochromatographic assay (LFIC) for detecting extended-spectrum ß-lactamases (ESBL) (NG-Test CTX-M MULTI assay) using bacterial pellets from BC. A total of 168 BCs from unique patients were included. A pre-established volume of BC flagged as positive was transferred in brain heart infusion with or without ceftriaxone (2 mg/ml). After 2-h incubation, intact bacterial pellets were used for MALDI-TOF MS testing. Identification of bacterial species (index score > 2) in the presence of CRO was considered marker of 3rd CephR. The LFIC assay was evaluated in 141 BC. Bacteremia episodes were caused by E. coli (n = 115) or Klebsiella spp. (n = 53). A total of 49 strains were 3rd CephR by broth microdilution, of which 41 were ESBL producers, seven expressed ESBL and OXA-48 type D carbapenemase, and one harbored a plasmid-mediated AmpC. The MALDI-TOF MS method yielded four very major errors (false susceptibility) and two major errors (false resistance). The overall sensitivity of the assay was 91.8% and the specificity 98.3%. Concordance between the LFIC assay and the MALDI-TOF MS method for detection of ESBL-mediated 3rd CephR was 100%. Both evaluated methods may prove useful for early adjustment of empirical therapy in patients with E. coli and Klebsiella spp. bloodstream infections. Whether their use has a beneficial impact on patient outcomes is currently under investigation.

Presence of Colistin- and Tigecycline-Resistant Klebsiella pneumoniae ST29 in Municipal Wastewater Influents in Japan.

The aim of this study was to investigate the presence of colistin- and/or tigecycline-resistant Klebsiella spp. in influents from four wastewater treatment plants (WWTPs), which partly reflect the gut microbiome of human populations. Colistin- and tigecycline-resistant Klebsiella pneumoniae isolates (K30/ST29) were detected four times from the WWTP A during a period of 3 months. Disruptions of the mgrB and ramR genes by ISEc68 and ISKpn21, respectively, were identified in those four isolates. They also shared the IncL/M 86,197-bp plasmids carrying a blaCTX-M-3 and Tn1548-associated armA [IS26-IntI1-dfrA12-gucF-aadA2-qacEΔ1-sul1-ISCR1-ISEc28-armA-ISEc29-msr(E)-mph(E)-IS26]. Those isolates formed a distinct cluster within wgMLST clusters of ST29 K30 public reference strains of human origin and were unique due to harboring of Tn21-like mercury resistance operon transposons in addition to silver, copper, and arsenic resistance determinants. Five K. pneumoniae strains with different STs and 1 Klebsiella quasipneumoniae strain, exhibiting colistin resistance, were detected in WWTPs B, C, and D. For these isolates, disruptions of mgrB by ISEc68 (three isolates) or ISEcl1 (one isolate), insertion of IS2 in the mgrB promoter region (one isolate), and inactivation of MgrB by a nonsense mutation (one isolate) were identified. Close monitoring of these mcr-negative colistin- and/or tigecycline-resistant bacteria in wastewater influents is imperative to avoid further limiting of treatment options.

Screening a Strain of Klebsiella sp. O852 and the Optimization of Fermentation Conditions for Trans-Dihydrocarvone Production.

Flavors and fragrances have high commercial value in the food, cosmetic, chemical and pharmaceutical industries. It is interesting to investigate the isolation and characterization of new microorganisms with the ability to produce flavor compounds. In this study, a new strain of Klebsiella sp. O852 (accession number CCTCC M2020509) was isolated from decayed navel orange (Citrus sinensis (L.) Osbeck), which was proved to be capable of converting limonene to trans-dihydrocarvone. Besides, the optimization of various reaction parameters to enhance the trans-dihydrocarvone production in shake flask was performed for Klebsiella sp. O852. The results showed that the yield of trans-dihydrocarvone reached up to 1 058 mg/L when Klebsiella sp. O852 was incubated using LB-M medium for 4 h at 36 °C and 150 rpm, and the biotransformation process was monitored for 36 h after adding 1680 mg/L limonene/ethanol (final ethanol concentration of 0.8% (v/v)). The content of trans-dihydrocarvone increased 16 times after optimization. This study provided a basis and reference for producing trans-dihydrocarvone by biotransformation.

Prevalence and antimicrobial resistance of Klebsiella species isolated from clinically ill companion animals.

BACKGROUND: Klebsiella spp. is an important conditional pathogen in humans and animals. However, due to the indiscriminate use of antibiotics, the incidence of antimicrobial resistance has increased. OBJECTIVES: The purpose of this study was to investigate antimicrobial resistance in strains of Klebsiella strains and the phylogenetic relatedness of extended-spectrum cephalosporin (ESC)-resistance among Klebsiella strains isolated from clinically ill companion animals. METHODS: A total of 336 clinical specimens were collected from animal hospitals. Identification of Klebsiella species, determination of minimum inhibitory concentrations, detection of ESC resistance genes, polymerase chain reaction-based replicon typing of plasmids by conjugation, and multilocus sequence typing were performed. RESULTS: Forty-three Klebsiella strains were isolated and, subsequently, 28 were identified as K. pneumoniae, 11 as K. oxytoca, and 4 as K. aerogenes. Eleven strains were isolated from feces, followed by 10 from ear, 7 from the nasal cavity, 6 from urine, 5 from genitals, and 4 from skin. Klebsiella isolates showed more than 40% resistance to penicillin, cephalosporin, fluoroquinolone, and aminoglycoside. ESCresistance genes, CTX-M groups (CTX-M-3, CTX-M-15, and CTX-M-65), and AmpC (CMY-2 and DHA-1) were most common in the K. pneumoniae strains. Some K. pneumoniae carrying CTX-M or AmpC were transferred via IncFII plasmids. Two sequence types, ST709 and ST307, from K. pneumoniae were most common. CONCLUSIONS: In conclusion, this is the first report on the prevalence, ESCresistance genotypes, and sequence types of Klebsiella strains isolated from clinically ill companion animals. The combination of infectious diseases and antimicrobial resistance by Klebsiella in companion animals suggest that, in clinical veterinary, antibiotic selection should be made carefully and in conjunction with the disease diagnosis.

Antibiogram and Molecular Characterization of AmpC and ESBL-Producing Gram-Negative Bacteria from Poultry and Abattoir Samples.

BACKGROUND AND OBJECTIVE: The global antibiotic resistance threat posed by ESBL and AmpC-producing Gram-Negative Bacteria (GNB) is a public health menace that rolls back the gains of 'One Health'. This study investigated the antibiogram and prevalence of AmpC and ESBL genes in Escherichia coli, Klebsiella spp. and Pseudomonas spp. from poultry and abattoir milieus in Enugu and Ebonyi States, Nigeria. MATERIALS AND METHODS: Isolation, identification and characterization of GNB from samples (150 abattoirs and 300 poultry) were done using standard microbiological techniques. Antimicrobial Susceptibility Testing (AST), as well as phenotypic screening for ESBL and AmpC enzymes, was performed using the Kirby-Bauer disc diffusion technique. PCR technique was used to screen isolated GNB for AmpC and ESBL genes. RESULTS: Exactly 42 E. coli and 8 Klebsiella spp. isolate from poultry samples and another 5 P. aeruginosa isolates from abattoir samples were phenotypically confirmed to be ESBL-producers. AmpC enzymes were phenotypically detected in 8 E. coli and 13 P. aeruginosa isolates from poultry samples. All ESBL and AmpC-positive bacteria exhibited high resistance frequencies to tested antibiotics, especially to the carbapenems and cephalosporins. ESBL genes (CTX-M, SHV-1, TEM) and AmpC genes (ACC-M, MOX-M, DHA-M) were harbored by the isolated GNB in this study. Overall, the DHA-M and CTX-M genes, mediating AmpC and ESBL production respectively were the most prevalent genes harbored by the tested GNB. CONCLUSION: This study reported that AmpC and ESBL genes are harbored by Gram-negative bacteria (E. coli, Klebsiella species and P. aeruginosa) that emanated from poultry and abattoir milieus.

Anchoring Drugs to a Zinc(II) Coordination Polymer Network: Exploiting Structural Rationale toward the Design of Metallogels for Drug-Delivery Applications.

A new series of coordination polymers (CPs) were synthesized and crystallographically characterized by single-crystal X-ray diffraction with the aim of developing drug-delivery systems via metallogel formation. Structural rationale was employed to design such coordination-polymer-based metallogels. As many as nine CPs were obtained by reacting two bis(pyridyl)urea ligands, namely, 1,3-dipyridin-3-ylurea (3U) and 1,3-dipyridin-4-ylurea (4U), and the sodium salt of various nonsteroidal antiinflammatory drugs, namely, ibuprofen (IBU), naproxen (NAP), fenoprofen (FEN), diclofenac (DIC), meclofenamic acid (MEC), mefenamic acid (MEF), and Zn(NO3)2. All of the CPs displayed 1D polymeric chains that were self-assembled through various hydrogen-bonding interactions involving the urea N-H and carboxylate O atoms and, in a few cases, lattice-occluded water molecules. The reacting components of the CPs produced five metallogels in dimethyl sulfoxide/water. The gels were characterized by rheology and transmission electron microscopy. Three selected metallogelators, namely, 3UMEFg, 3UNAPg, and 3UMECg, showed in vitro anticancer, cell imaging, and multidrug delivery for antibacterial applications, respectively. The shear-thinning properties of 3UMECg (rheoreversibility and injectability) make it a potential candidate for plausible topical application.