Extended Spectrum Research Articles

Bactericidal activity of Ag4V2O7/β-AgVO3 heterostructures against antibiotic-resistant Klebsiella pneumoniae.

Although Ag-based materials are efficient against antibiotic-resistant bacteria, their high toxicity to living organisms represents a major challenge for obtaining useful products. In this work, we report the bactericidal activity of Ag4V2O7/β-AgVO3 heterostructures, which proved to be effective against Klebsiella pneumoniae (ATCC 1706, a standard strain; A54970, a multidrug-resistant carbapenemase (KPC)-producing strain; A34057, a multidrug-resistant strain capable of producing extended spectrum beta-lactamases (ESBL); and a community-isolated strain, A58240) at minimum inhibitory concentrations (MIC) as low as 62.5μg/mL. This activity is higher than that reported for the individual silver vanadates (Ag4V2O7 or β-AgVO3) owing to the synergistic interactions between both semiconductors. However, the most efficient heterostructure was found to be toxic to mouse 3T3 fibroblasts and to L. sativa and C. sativus seeds, as indicated by MTT ((4,5 - dimethylthiazol -2yl) 2,5 -diphenylbromide), neutral red assays and germination index measurements. The antimicrobial, phytotoxic and cytotoxic activities were all associated with an efficient generation of reactive oxygen species (ROS) in the heterostructure, especially OH and O2- radicals. The ROS production by Ag4V2O7/β-AgVO3 heterostructures was measured through photodegradation studies with Rhodamine B. While the bactericidal activity of the heterostructures is promising, especially when compared to Ag-based materials, their use in practical applications will require encapsulation either to avoid leaching or to mitigate their toxicity to humans, animals and plants.

Camellia sinensis mediated synthesis and characterization of nanoparticles and applications to control Gram-negative ESBL producing antibiotic resistant bacterial pathogens

Camellia sinensis (tea), has numerous pharmacotherapeutic benefits. The majority of its phytocompounds exhibit antibacterial properties, which are effective in treating a range of bacterial diseases, including Urinary Tract Infections (UTI). In-silico research examined the ability of the major C. sinensis phytocompounds to inhibit bacterial enzymes. Molecular docking was performed to explore the efficiency of tea phytoconstituents by evaluating their binding affinity and existence of different interactions between the bacterial proteins/enzymes and the tea phytoconstituents, to inhibit key enzymes associated with bacterial infections. Camellia sinensis silver nanoparticles (CsAgNps) were synthesized by using the biogenesis method, which comes under the bottom-up approach. The characterization results confirmed the charge, size and shape of the synthesized CsAgNps. Characterization data revealed the existence of functional groups which are responsible for the stability and effectivity of the nanoparticles. Toxicity studies were carried out to determine its safety at various dosage levels. In in vitro condition, CsAgNps synthesized from fresh tea leaves extract showed antibacterial properties even against antibiotic-resistant bacteria (ARB). In-vitro studies of this biosynthesized CsAgNps confirmed the bacteriostatic and bactericidal properties by using agar well diffusion, MIC and MBC methods at various concentrations of the synthesized CsAgNps. This proved that CsAgNps can inhibit bacterial growth against these three Gram-negative extended spectrum beta lactamase (ESBL) producers and hence it can serve as an alternative to antibiotics.

Detection of Extended spectrum β -lactamases (ESBLs) produced by Acinetobacter baumannii isolated from burn patients in Al- Nasiriyah city Iraq.

The aim of present study is to isolate and identify of Acinetobacter baumannii from burns infect as well as study their production for Extended spectrum β -lactamases (ESBLs).This study included 205 burn patients at Al-Hussien Teaching Hospital and other burn care units in Al-Nasiriyah city . The specimens were collected during period from August 2018 to February 2019. It were taken by burn swabs from pus of the burned area and then inoculated on blood agar and MacConkey agar. The isolates were diagnosed by microscopic examination, biochemical tests , API 20 E and Vitek- 2 system . It was found that 20 isolates of A.baumannii and 2 isolates of A. loffii . Where identified from this study . The study showed 18 (90%) out of 20 isolates of A. baumannii were appeared resistance to cefotaxime and ceftazidime(screening test of ESBLs) . Seven (35%) of isolates were producing ESBLs by confirmatory test(Modified double –disk synergy test (MDDST)) , and 13(65%) of isolates were negative results to production of the ESBLs.

Healthcare Facilities as Potential Reservoirs of Antimicrobial Resistant Klebsiella pneumoniae: An Emerging Concern to Public Health in Bangladesh.

The emergence of virulent extended spectrum β-lactamase producing Klebsiella pneumoniae (ESBL-KP) including carbapenem-resistant Klebsiella pneumoniae (CRKP) in hospital-acquired infections has resulted in significant morbidity and mortality worldwide. We investigated the antibiotic resistance and virulence factors associated with ESBL-KP and CRKP in tertiary care hospitals in Bangladesh and explored their ability to form biofilm. A total of 67 ESBL-KP were isolated from 285 Klebsiella pneumoniae isolates from environmental and patient samples from January 2019 to April 2019. For ESBL-KP isolates, molecular typing was carried out using enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR), antibiotic susceptibility testing, PCR for virulence and drug-resistant genes, and biofilm assays were also performed. All 67 isolates were multidrug-resistant (MDR) to different antibiotics at high levels and 42 isolates were also carbapenem-resistant. The most common β-lactam resistance gene was blaCTX-M-1 (91%), followed by blaTEM (76.1%), blaSHV (68.7%), blaOXA-1 (29.9%), blaGES (14.9%), blaCTX-M-9 (11.9%), and blaCTX-M-2 (4.5%). The carbapenemase genes blaKPC (55.2%), blaIMP (28.4%), blaVIM (14.9%), blaNDM-1 (13.4%), and blaOXA-48 (10.4%) and virulence-associated genes such as fimH (71.6%), ugeF (58.2%), wabG (56.7%), ureA (47.8%) and kfuBC (28.4%) were also detected. About 96.2% of the environmental and 100% of the patient isolates were able to form biofilms. ERIC-PCR-based genotyping and hierarchical clustering of K. pneumoniae isolates revealed an association between environmental and patient samples, indicating clonal association with possible transmission of antimicrobial resistance genes. Our findings can help in improving patient care and infection control, and the development of public health policies related to hospital-acquired infections.

Microbiota Assessment of Pediatric Simple and Complex Acute Appendicitis.

Background and Objectives. The aim of this study is to determine the prevailing microbiota in samples from pediatric patients with acute appendicitis, as well as evaluate the antibacterial sensitivity of the isolated microorganisms, comparing the data obtained with the clinic’s antibacterial therapy guidelines. Materials and Methods. The study group consisted of 93 patients between the ages of 7 and 18. All patients underwent a laparoscopic or conventional appendectomy. The children were hospitalized with signs and symptoms suggestive of acute appendicitis. Microbiological cultures from the appendix and abdominal cavity were collected intraoperatively. Results. E. coli was identified in most cases irrespective of the clinical presentation of acute appendicitis. Most strains were susceptible to ampicillin and amoxicillin/clavulanic acid. Five strains of E. coli produced extended spectrum beta-lactamase (ESBL). Pseudomonas aeruginosa (P. aeruginosa) was the second most commonly isolated causative agent. Furthermore, it was common in cases of acute complex appendicitis. Most strains of P. aeruginosa were resistant to amoxicillin/clavulanic acid, ertapenem, ampicillin and cefotaxime, yet were susceptible to ceftazidime. Regardless of the clinical presentation, the samples yielded mixed isolates. Conclusion. E. coli is the main causative agent of acute appendicitis in the pediatric population displaying susceptibility to various antibiotics. P. aeruginosa was more prevalent in cases of acute complex appendicitis. P. aeruginosa isolates were susceptible to ceftazidime; however, they were resistant to cefotaxime, which should, therefore, be removed from guidelines for empirical antibacterial treatment of acute appendicitis due to phenotypic resistance of P. aeruginosa. We recommend antibiotics with distinct implementation to avoid antibiotic resistance.

Molecular Epidemiology and Virulence of Non-Typhoidal Salmonella in Armenia.

In this work, we analysed human isolates of nontyphoidal Salmonella enterica subsp. enterica (NTS), which were collected from salmonellosis cases in Armenia from 1996 to 2019. This disease became a leading food-borne bacterial infection in the region, with the younger age groups especially affected. The isolates were characterised by serotyping, Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR) typing, and whole genome sequencing (WGS). The main serotypes were S. Typhimurium, S. Enteritidis, and S. Arizonae. ERIC-PCR indicated a high degree of clonality among S. Typhimurium strains, which were also multidrug-resistant and produced extended spectrum beta-lactamases. During the study period, the frequency of S. Typhimurium and S. Arizonae isolations decreased, but with the increase in S. Enteritidis and other NTS. A total of 42 NTS isolates were subjected to WGS and explored for virulence-related traits and the corresponding genetic elements. Some virulence and genetic factors were shared by all NTS serotypes, while the main differences were attributed to the serotype-specific diversity of virulence genes, SPIs, virulence plasmids, and phages. The results indicated the variability and dynamics in the epidemiology of salmonellosis and a high virulence potential of human NTS isolates circulating in the region.

Environmental Contamination with Multi-drug Resistant Organisms in a Nigerian Neonatal Intensive Care Unit. Impactful Infection Source

Background: Neonates admitted into neonatal intensive care unit (NICU) are at high risk of acquiring infections due to the immaturity of their immune systems and the barrage of intensive medical care. Multiple drug resistant organisms are mostly implicated in causing these infections, usually acquired through contaminated environment. This research was designed to assess the environmental contamination with multi-drug resistant organisms in neonatal intensive care unit.
 Methods: Environmental samples were collected from all the equipment in each of the 3 rooms in neonatal intensive care unit and cultured on 5% sheep blood agar, chocolate agar and MacConkey agar plates. The isolates were identified with standard biochemical tests. Antibiotic susceptibility profile was done using modified Kirby Bauer disc diffusion method and interpreted according to the guidelines by Clinical and Laboratory Standard Institute. Methicillin resistant Staphylococcus aureus was detected using cefoxitin disc while extended spectrum beta-lactamase (ESBL) producers were confirmed with ESBL chromogenic agar.
 Results: Out of 36 samples collected from different sites in the 3 rooms, 19(53%) yielded bacterial growth. A total of 28 (78%) bacterial isolates were detected, with majority 25(89%) isolated from room 1. Staphylococcus aureus 15(42%) was the predominant isolate followed by Klebsiella pneumoniae 5(14%) and non-Candida albicans spp 3(8%). Most of Staphylococcus aureus isolates were from samples collected from incubators. Out of 15 Staphylococcus aureus isolated, 11 (73%) were MRSA. Among the Gram negatives, all 5(100%) Klebsiella pneumonioae and 3(60) Acinetobacter species were positive for extended spectrum beta-lactamase production. All the isolates were resistant to 3 or more classes of antibiotics. Imipenem and linezolid were the most sensitive antibiotics to the isolated organisms.
 Conclusion: High degree of contamination of the environment in NICU with multi drug resistant bacteria is worrisome because of the serious threat it poses. This calls for immediate holistic interventional measures. Strict Compliance with infection prevention and control measures. Additionally, there should be regular antimicrobial resistance surveillance in NICU.

Molecular Detection of Extended Spectrum Beta-Lactamases in Clinical Isolates of Pseudomonas aeruginosa

Pseudomonas aeruginosa producing extended spectrum beta lactamases (ESBL) is a major concern in the hospital settings. It is usually reported in Enterobacteriaceae and is less frequently observed in P. aeruginosa. There is no recommended test for ESBL detection in P.aeruginosa. Therefore, we determined the occurrence of ESBL in clinical isolates of P.aeruginosa by both phenotypic and genotypic methods. Antimicrobial susceptibility tests were done on two hundred and thirteen isolates of P. aeruginosa. Phenotypic detection of ESBL was performed using combined disk method and ESBL encoding genes such as blaVEB, blaPER, blaPSE, blaGES, blaTEM, blaSHV, blaCTX-M, blaBEL, blaOXA1, blaOXA10, blaOXA2 were studied by simplex PCR. Of the 213 isolates, 85 were identified as resistant to ceftazidime and 27/85 isolates were confirmed to be ESBL producers by phenotypic method. The presence of genes encoding ESBLs comprising of blaTEM (n=44), blaOXA-10 (n=19) isolates, blaOXA-1 (n=5), blaOXA-2 (n=3) were found. All OXA gene positive isolates exhibited the ESBL phenotype. The blaGES gene were identified in 4/85 (5%) isolates. This study shows the prevalence of ESBL among clinical isolates of P.aeruginosa and in particular, the presence of GES β lactamases.

Clinical and microbiological features of ceftolozane/tazobactam-resistant Pseudomonas aeruginosa isolates in a university hospital in central Italy

Ceftolozane/tazobactam (C/T) is a novel cephalosporin and β-lactamase inhibitor combination with great activity against Pseudomonas aeruginosa. To assess P. aeruginosa susceptibility to C/T, a surveillance study was conducted from October 2018 to March 2019 at the University Hospital 'Ospedali Riuniti' in Ancona, Italy. Minimum inhibitory concentrations (MICs) to C/T were determined by Etest strip. Resistant isolates were characterized by phenotypic (broth microdilution antimicrobial susceptibility testing and modified Carbapenem Inactivation Method [mCIM]) and genotypic (Polymerase Chain Reaction [PCR], Pulsed Field Gel Electrophoresis [PFGE], and whole-genome sequencing [WGS]) methods. Clinical variables of patients infected by C/T-resistant P. aeruginosa were collected from medical records. Fifteen of 317 P. aeruginosa collected showed resistance to C/T (4.7%). Ten strains demonstrated carbapenemase activity by mCIM method, and PCR confirmed that eight strains harbored a blaVIM gene while the other two were positive for blaIMP. Additionally, three isolates carried acquired extended spectrum β-lactamase genes (two isolates carried blaPER and one carried blaGES). Eight strains were strictly related by PFGE and WGS analysis confirmed that they belonged to sequence type (ST)111. The other STs found were ST175 (two isolates), ST235 (two isolates), ST70 (one isolate), ST621 (one isolate), and the new ST3354 (one isolate). Most patients had received previous antibiotic therapies, carried invasive devices, and experienced prolonged hospitalization. This study demonstrated the presence of C/T-resistant P. aeruginosa isolates in a regional hospital carrying a number of resistance mechanisms acquired by different high-risk clones.

Sheep and goat milk in Jordan is a reservoir of multidrug resistant extended spectrum and AmpC beta-lactamases Escherichia coli

Limited data is available on the prevalence and antimicrobial resistance of extended spectrum (ESBL) and AmpC β-lactamases Escherichia coli in sheep and goats in Jordan. This study determined the molecular prevalence and antimicrobial-resistance of ESBL and AmpC β-lactamases E. coli in 155 sheep and goat flocks across Jordan by testing 948 milk samples. The samples were enriched in MacConkey broth, and then plated on MacConkey agar with cefotaxime. The presence of ESBL and AmpC genes in the E. coli isolates was determined by PCR and the resistance toward critically important antimicrobials was tested by disc diffusion. In total, 1016 E. coli isolates were isolated from the cefotaxime supplemented MacConkey, 382 isolates harbored ESBL genes and 54 harbored blaCMY. The prevalence of blaCTX-M, blaTEM, blaCMY and blaSHVE. coli in the milk samples were 33.5 %, 31.7 %, 5.7 %. and 1.1 %, respectively. At the flock level, 30.3 % flocks had β-lactamase E. coli, specifically 25.2 %, 20.7 %, 5.2 % and 2.6 % had blaCTX-M, blaTEM, blaCMY and blaSHV E. coli, respectively. About 52.2 % of the isolates harbored both blaCTX-M and blaTEM. A high percentage (>59 %) of ESBL and AmpC β-lactamases E. coli resisted sulfamethoxazole/trimethoprim, tetracycline and nalidixic acid. Overall, 93.5 % and 96.3 % of ESBL and AmpC E. coli were resistant to ≥1 another antimicrobial class and 44.5 % and 44.4 % were resistant to ≥3 another antimicrobial class, respectively. This study shows that sheep and goat milk is a reservoir of multidrug resistant ESBL and AmpC β-lactamases E. coli, thus good hygienic practices and judicious antimicrobials use are important in small ruminants' farming.

Reduced penetrance of an eastern French mutation in ATL1 autosomal-dominant inheritance (SPG3A): extended phenotypic spectrum coupled with brain 18F-FDG PET.

ATL1-related spastic paraplegia SPG3A is a pure form of hereditary spastic paraplegia. Rare complex phenotypes have been described, but few data concerning cognitive evaluation or molecular imaging of these patients are available. We relate a retrospective collection of patients with SPG3A from the Neurology Department of Nancy University Hospital, France. For each patient were carried out a 18F-FDG PET (positron emission tomography), a electromyography (EMG), a sudoscan®, a cerebral and spinal cord MRI (magnetic resonance imaging) with measurement of cervical and thoracic surfaces, a neuropsychological assessment. The present report outlines standardised clinical and paraclinical data of five patients from two east-France families carrying the same missense pathogenic variation, NM_015915.4(ATL1): c.1483C > T p.(Arg495Trp) in ATL1. Mean age at onset was 14 ± 15.01years. Semi-quantitatively and in comparison to healthy age-matched subjects, PET scans showed a significant cerebellar and upper or mild temporal hypometabolism in all four adult patients and hypometabolism of the prefrontal cortex or precuneus in three of them. Sudoscan® showed signs of small fibre neuropathy in three patients. Cervical and thoracic patients' spinal cords were significantly thinner than matched-control, respectively 71 ± 6.59mm2 (p = 0.01) and 35.64 ± 4.35mm2 (p = 0.015). Two patients presented with a dysexecutive syndrome. While adding new clinical and paraclinical signs associated with ATL1 pathogenic variations, we insist here on the variable penetrance and expressivity. We report small fibre neuropathy, cerebellar hypometabolism and dysexecutive syndromes associated with SPG3A. These cognitive impairments and PET findings may be related to a cortico-cerebellar bundle axonopathy described in the cerebellar cognitive affective syndrome (CCAS).

Incidence and antibiotic susceptibility profile of uropathogenic Escherichia coli positive for extended spectrum β-lactamase among HIV/AIDS patients in Awka metropolis, Nigeria.

This study investigated the incidence and antibiotic susceptibility profile of extended spectrum β-lactamase (ESBL) producing uropathogenic Escherichia coli recovered from HIV/AIDS patients in Awka metropolis, Nigeria. A total of 363 urine samples were bacteriologically analyzed for the isolation of E. coli isolates which were further characterized using standard microbiology techniques. The isolated uropathogenic E. coli was tested for susceptibility to a range of clinically important antibiotics using the modified disk diffusion technique. All E. coli isolates were phenotypically screened for ESBL production using the combined disk technique, and strains which were positive were further confirmed for the presence of ESBL genes using PCR technique. A total 160 (44.1%) non-duplicate isolates were bacteriologically confirmed to be uropathogenic E. coli (UPEC). The E. coli isolates showed reduced susceptibility to important antibiotics including ceftazidime (76.88%), cefuroxime (77.5%), cefixime (61.88%), amoxicillin-clavulanic (32.5%) and ciprofloxacin (34.38%). Twenty-seven of the UPEC isolates were phenotypically confirmed to be ESBL producers. PCR test confirmed some important genes mediating ESBL production in Gram negative bacteria including bla TEM (5.0%) and bla CTX-M-15 (6.9%) genes. We report a high prevalence of ESBL producers among HIV/AIDS patients in Awka, Nigeria. This result is important as antibiotic resistance (ABR) particularly those mediated by multidrug resistant bacteria as reported in this current study could complicate treatment outcome, worsen the individual's health, and even increase cost of treatment and hospitalization. It is therefore important to lookout for ESBL positive UPEC amongst HIV/AIDS patients in Nigeria.

Bloodstream Infections in the Intensive Care Unit: a Single-Center Retrospective Bacteriological Analysis Between 2007 and 2019.

Hospital-acquired bloodstream infections are a severe worldwide problem associated with significant morbidity and mortality. This retrospective, single-center study aimed to analyze bloodstream infections in patients hospitalized in the intensive care unit of the Military Institute of Medicine, Poland. Data from the years 2007–2019 were analyzed. When the infection was suspected, blood samples were drawn and analyzed microbiologically. When bacterial growth was observed, an antimicrobial susceptibility/resistance analysis was performed. Among 12,619 analyzed samples, 1,509 were positive, and 1,557 pathogens were isolated. In 278/1,509 of the positive cases, a central line catheter infection was confirmed. Gram-negative bacteria were the most frequently (770/1,557) isolated, including Acinetobacter baumannii (312/770), Klebsiella pneumoniae (165/770; 67/165 were the isolates that expressed extended spectrum beta-lactamases (ESBL), 5/165 isolates produced the New Delhi metallo-β-lactamases (NDM), 4/165 isolates expressed Klebsiella pneumoniae carbapenemase (KPC), and 1/165 isolate produced OXA48 carbapenemase), Pseudomonas aeruginosa (111/770; 2/111 isolates produced metallo-β-lactamase (MBL), and Escherichia coli (69/770; 11/69 – ESBL). Most Gram-positive pathogens were staphylococci (545/733), mainly coagulase-negative (368/545). Among 545 isolates of the staphylococci, 58 represented methicillin-resistant Staphylococcus aureus (MRSA). Fungi were isolated from 3.5% of samples. All isolated MRSA and methicillin-resistant coagulase-negative Staphylococcus (MRCNS) strains were susceptible to vancomycin, methicillin-sensitive Staphylococcus aureus (MSSA) isolates – to isoxazolyl penicillins, and vancomycin-resistant Enterococcus (VRE) – to linezolid and tigecycline. However, colistin was the only therapeutic option in some infections caused by A. baumannii and KPC-producing K. pneumoniae. P. aeruginosa was still susceptible to cefepime and ceftazidime. Echinocandins were effective therapeutics in the treatment of fungal infections.

Characterization of β-Lactamases and Multidrug Resistance Mechanisms in Enterobacterales from Hospital Effluents and Wastewater Treatment Plant.

Antimicrobials in wastewater promote the emergence of antibiotic resistance, facilitated by selective pressure and transfer of resistant genes. Enteric bacteria belonging to Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter cloacae, and Citrobacter species (n = 126) from hospital effluents and proximate wastewater treatment plant were assayed for susceptibility to four antimicrobial classes. The β-lactamase encoding genes harbored in plasmids were genotyped and the plasmids were sequenced. A multidrug resistance phenotype was found in 72% (n = 58) of E. coli isolates, 70% (n = 43) of Klebsiella species isolates, and 40% (n = 25) of Enterobacter and Citrobacter species. Moreover, 86% (n = 50) of E. coli, 77% (n = 33) of Klebsiella species, and 25% (n = 4) of Citrobacter species isolates phenotypically expressed extended spectrum β-lactamase. Regarding ESBL genes, blaCTX-M-27 and blaTEM-1 were found in E. coli, while Klebsiella species harbored blaCTX-M-15, blaCTX-M-30, or blaSHV-12. Genes coding for aminoglycoside modifying enzymes, adenylyltransferases (aadA1, aadA5), phosphotransferases (aph(6)-1d, aph(3″)-Ib), acetyltransferases (aac(3)-IIa), (aac(6)-Ib), sulfonamide/trimethoprim resistant dihydropteroate synthase (sul), dihydrofolate reductase (dfrA), and quinolone resistance protein (qnrB1) were also identified. Monitoring wastewater from human sources for acquired resistance in clinically important bacteria may provide a cheaper alternative in regions facing challenges that limit clinical surveillance.

Biotyping, virulotyping and biofilm formation ability of ESBL-Klebsiella pneumoniae isolates from nosocomial infections.

The aim of this study was to investigate the frequency, molecular characterization, virulence genes, resistance genes and antimicrobial profile of nosocomial extended spectrum beta lactamase producing Klebsiella species. A total of 22 (12.2%) K. pneumoniae strains were isolated from 180 clinical samples collected from hospitalized patients in Egypt. K. pneumoniae biotypes were B1 (72.8%), B3 (13.6%) and B4 (13.6%). The isolates were classified for the capsular serotypes, 86.4% (20/22) were of K1 serotype, while only two isolates (13.64%) were of K2 serotype. Hypermucoviscous K. pneumoniae isolates accounted for 68.2%. Biofilm formation ability of K. pneumoniae was determined by microtitre plate method. The majority of the isolates (40.9%) were moderate biofilm producers, while 27.3% were strong biofilm producers. All K. pneumoniae strains were positive for fimH and traT genes, while magA was identified in only 63.6% of the isolates. The antibiotic susceptibility profile of the isolates (n=22) was determined by the disc diffusion technique using 23 different antibiotics. Streptomycin and imipenem are the most effective antibiotics against 22 tested K. pneumoniae isolates with sensitivity rates of 63.64% and 54.54% respectively. All tested K. pneumoniae isolates showed high resistance to amoxicillin∕clavulanate (100%), cefuroxime (100%) and ceftazidime (95.45%). Extended spectrum beta lactamases (ESBL) production and the presence of ESBL-related genes were tested in the isolates. All the isolates tested positive for blaVIM, NDM1 and blaTEM, while only 81.8%tested positive for the blaSHV gene. Increasing antimicrobial resistance in K. pneumoniae causing nosocomial infections limits the use of antimicrobial agents for treatment. Furthermore, the spread of biofilm, multiple drug resistant and ESBL-producing K. pneumoniae isolates is a public threat for hospitalized patients.

Phenotypic and molecular characterization of blaCTX-M-3 and blaCTX-M-55-producing monophasic Salmonella enterica serovar Typhimurium in Greece

To characterize three Salmonella enterica serovar Typhimurium strains using whole genome sequencing (WGS) and conventional methods. The isolates were recovered from three pediatric patients in Greece as part of the hospital's epidemiological surveillance system during 2016 to 2018. Bacterial identification and antimicrobial susceptibility testing was performed using the VITEK 2 automated system, disc diffusion test, and MIC gradient test while serotyping by the slide agglutination method. Detection of resistance genes, eBurst groups (eBG), assignment to sequence types, single nucleotide polymorphism (SNP) typing, location and characterization of drug resistance regions, and in silico plasmid detection were carried out using WGS. All strains were identified as S. Typhimurium-monophasic, ST34, eBG1 with antigenic formula 1,4, [5], 12:i:-. They were phenotypically resistant to most antibiotics tested except piperacillin/tazobactam, imipenem, and co-trimoxazole. WGS revealed the chromosomally located genes encoding the ASSuT (ampicillin, streptomycin, sulfonamides, and tetracycline) resistant pattern in all three strains. WGS revealed extended spectrum ß-lactamase (ESBL) production in all three strains, the presence of blaCTX-M-3 on an IncI1 plasmid in two strains isolated in 2018, and the chromosomally encoded blaCTX-M-55 plus qnrS1 (resistance to ciprofloxacin) in the strain isolated in 2016. The two strains from 2018 were isolated from the same hospital ward and were genetically related. The emergence of ESBL among S. 1,4,[5], 12:i:- is worrisome due to its increasing antimicrobial resistance phenotype, making clinical treatment difficult. WGS provides an alternative to traditional methods of identification and genomic characterisation of strains, and serves to better understand their epidemiological dynamics and bacterial pathogenesis.

Molecular characterization of blaNDM, blaOXA-48, mcr-1 and blaTEM-52 positive and concurrently carbapenem and colistin resistant and extended spectrum beta-lactamase producing Escherichia coli in chicken in Malaysia

BackgroundAntimicrobial resistance (AMR) is a global public health threat and the use of antibiotics growth promoters in food animals has been implicated as a potential contributing factor in the emergence and spread of AMR. This study was conducted to investigate colistin and carbapenem resistance and extended spectrum beta-lactamase producing E. coli from live broiler chicken and chicken meat in Kelantan, Malaysia.ResultsAmong the E. coli isolates, 37.5% (27/72 were positive for at least one of the resistance genes and one isolate was positive for mcr-1, blaTEM-52, blaNDM and blaOXA-48 whereas 4.17% (3/72) and 2.78% (2/72) were positive for mcr-1, blaTEM-52 and blaOXA-48, and mcr-1, blaTEM-52 and blaIMP. Multilocus sequence typing (MLST) results revealed the presence of widespread E. coli strains belonging to the sequence types ST410 and ST155 and other extra-intestinal E. coli (ExPEC) strains. Phylogroup A made up the majority 51.85% (14/27) followed by phylogroup B1 22.22% (6/27).ConclusionsThe findings imply the potential threats of colistin, extended-spectrum beta-lactamase producing and carbapenem resistant E. coli in food animals to the public health and underscores the need for judicious use of antibiotics in animal production and good hygiene practices to curb the rising risks of AMR.

Antisense Peptide Nucleic Acid-Diaminobutanoic Acid Dendron Conjugates with SbmA-Independent Antimicrobial Activity against Gram-Negative Bacteria.

Precision antisense antibacterial agents may be developed into novel antibiotics in the fight against multidrug-resistant Gram-negative bacteria. In this study, a series of diaminobutanoic acid (DAB) dendrons are presented as novel carriers for the delivery of antisense antibacterial peptide nucleic acids (PNAs). The dendron–PNA conjugates targeting the essential acpP gene exhibit specific antisense antimicrobial bactericidal activity against Escherichia coli and Klebsiella pneumoniae at one-digit micromolar concentrations, while showing low toxicity to human cells. One compound selected from a structure–activity relationship series showed high stability in mouse and human serum (t1/2 ≫ 24 h) as well as in vivo activity against a multidrug-resistant, extended spectrum beta-lactamase-producing E. coli in a murine peritonitis model. The compound was also well tolerated in mice upon i.v. administration up to a dose of 20 mg/kg, and in vivo fluorescence imaging indicated clearance via renal excretion with slight accumulation in the kidneys and liver. Thus, DAB-based dendrons constitute a promising new chemistry platform for development of effective delivery agents for antibacterial drugs with possible in vivo use.

Prevalence and molecular characterization of foodborne and human-derived Salmonella strains for resistance to critically important antibiotics.

The primary goals of this cross-sectional study were to screen various food/water, and human samples for the presence of Salmonella species, and to assess the phenotypic and genetic relationship between resistances found in food and human Salmonella isolates to critically important antibiotics. Between November 2019 and May 2021, 501 samples were randomly collected for Salmonella isolation and identification using standard culturing methods, biochemical, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and PCR techniques. Antimicrobial susceptibility testing was performed on confirmed Salmonella species, and PCR was used to investigate the genetic components that confer these resistance traits. Salmonella enterica subspecies enterica was confirmed in 35 (6.99%) of the samples (raw food=23, ready-to-eat food/drink [REF/D]=5, human=7). Seventeen of them were antibiotic-resistant to at least one class, and eight were multidrug-resistant (MDR) isolates (raw food=7, human=1). All Salmonella isolates were susceptible to carbapenems, third- and fourth-generation cephalosporins and monobactam antibiotics. Resistance phenotypes to aminoglycosides (48.57%), β-lactams (20%) and tetracycline (17.14%), as well as associated genes such as aadA, blaTEM , blaZ and tetA, as well as dfrA and sul1, were prevalent in Salmonella isolates. Colistin resistance genotype (mcr1) was detected in three (8.57%) isolates recovered from egg, cattle mince and rabbit meat, and the total incidence was 14.29% when two isolates exhibited resistance phenotypes were considered. Furthermore, four (11.43%) MDR isolates shared the blaTEM and blaZ genes, and one (2.86%) isolate contained three extended spectrum β-lactams producing genes (ESBL), namely blaCTX , blaTEM and blaZ . The gyrA gene was expressed by one of three foodborne Salmonella isolates (8.57%) with ciprofloxacin resistance phenotypes. To the best of our knowledge, this is the first report from Egypt identifying colistin resistance in Salmonella enterica recovered from cattle minced meat and rabbit meat. Overall, thehighest incidence rate of Salmonella enterica was found in cattle-derived products, and it was slightly more prevalent in RTE/D foods than in raw foods. Resistance to critical and clinically important antibiotics, particularly in Salmonella from RTE/D food, suggests that these antibiotics are being abused in the investigated area's veterinary field, and raises the potential of these isolates being transmitted to high-risk humans, which would be a serious problem. Future research using whole-genome sequencing is needed to clarify Salmonella resistance mechanisms to critically important antimicrobial agents or those exhibiting multidrug resistance.

Prevalence of Extended Spectrum Beta Lactamase producing Escherichia coli and Klebsiella spp. from urinary specimen in a tertiary care hospital

Escherichia coli and Klebsiella spp. are two important uropathogens showing resistance to beta-lactams by producing extended spectrum beta lactamase (ESBL), leading to difficulty in treating infections with these bacteria. Majority of the ESBLs are of TEM, CTXM and SHV types reported from Nepal. This study was conducted in Clinical Microbiology Department of Nepal Medical College Teaching Hospital from November 2021 to mid-January 2022 to determine prevalence of ESBL and antimicrobial susceptibility pattern among two common uropathogens. A total of 402 E. coli and Klebsiella spp. isolated from urine samples were included in the study. ESBL producing strains were identified by combination disc method and antibiotic susceptibility testing was done by Kirby Bauer’s disc diffusion method. PCR was done to detect the TEM and CTXM genes. Prevalence of ESBL among E. coli and Klebsiella spp. was found to be 14.9% (60/402). Among the E. coli, 16.1% (58/360) and only 4.8% (2/42) of Klebsiella spp. were ESBL producers. Antibiogram showed, 56.7% ESBL producers were susceptible to ciprofloxacin, 46.7% to co-trimoxazole, 96.7% to nitrofurantoin. However, all isolates were susceptible to amikacin, piperacillin-tazobactam, and carbapenems (both meropenem and imipenem). Among total 30 ESBL isolates selected by systematic random sampling, 3 isolates (all three E. coli) showed presence of TEM and 3 other isolates (two E. coli and one Klebsiella spp.) showed CTXM genes.