Ib-cr Genes Research Articles

Bioaccumulation of antibiotics and resistance genes in lettuce following cattle manure and digestate fertilization and their effects on soil and phyllosphere microbial communities

The degradation and bioaccumulation of selected antibiotics such as the sulfonamide sulfamethoxazole (SMX) and the fluoroquinolones enrofloxacin (ENR) and ciprofloxacin (CIP) were investigated in soil microcosm experiments where Lactuca sativa was grown with manure or digestate (1%) and spiked with a mixture of the three antibiotics (7.5 mg/kg each). The soil, rhizosphere and leaf phyllosphere were sampled (at 0 and 46 days) from each microcosm to analyze the antibiotic concentrations, main resistance genes (sul1, sul2, qnrS, aac-(6’)-Ib-crand qepA), the intI1and tnpA mobile genetic elements and the microbial community structure.Overall results showed that SMX and CIP decreased (70-85% and 55-79%, respectively), and ENR was quite persistent during the 46-day experiment. In plant presence, CIP and ENR were partially up-taken from soil to plant. In fact the bioaccumulation factors were > 1, with higher values in manure than digestate amended soils. The most abundant gene in soil was sul2 in digestate- and aac-(6’)-Ib-cr in the manure-amended microcosms. In soil, neither sulfamethoxazole-resistance (sul1 and sul2), nor fluoroquinolone-resistance (aac-(6’)-Ib-cr, qepA and qnrS) gene abundances were correlated with any antibiotic concentration. On the contrary, in lettuce leaves, the aac-(6’)-Ib-cr gene was the most abundant, in accordance with the fluoroquinolone bioaccumulation. Finally, digestate stimulated a higher soil microbial biodiversity, introducing and promoting more bacterial genera associated with antibiotic degradation and involved in soil fertility and decreased fluoroquinolone bioaccumulation.

Distribution of Ciprofloxacin- and Azithromycin-Resistant Genes among Salmonella Typhi Isolated from Human Blood.

Salmonella Typhi has developed resistance to different groups of antibiotics. The purpose of the present study was to assess the distribution of ciprofloxacin- and azithromycin-resistant genes among Salmonella Typhi isolated from human blood. This cross-sectional study was conducted in the Department of Microbiology of a tertiary care hospital in Bangladesh from July 2019-June 2020. Clinically suspected enteric fever patients, irrespective of age and gender, who attended the laboratory of the Department of Microbiology and outpatient department of Medicine of tertiary care hospital. Blood culture and sensitivity tests were done. The positive growth of Salmonella Typhi was identified by Gram staining, colony morphology, and biochemical test. Then, Salmonella Typhi was identified by using Salmonella-specific antisera. Final identification was made by using 16s rRNA by polymerase chain reaction (PCR). PCR was also done to detect quinolone and azithromycin resistance genes. A total number of 83 samples yielded positive cultures, of which 50 isolated organisms were identified as Salmonella species; however, among these isolates, Salmonella Typhi was detected in 40 (48.2%) isolates. Among 12 ciprofloxacin-resistant isolates, 8 (66.67%) were positive for the gyrA gene, 1 (8.33%) was positive for the qnrB gene and qnrS gene, 2 (16.67%) were positive for aac (6´)-Ib-cr. Among 12 azithromycin-resistant isolates, 2 (16.66%) were positive for mphA and mefA genes, respectively. In conclusion, the gyrA, aac (6´)-Ib-cr, mphA, and mefA genes are found for the first time in tertiary care hospitals from the quinolones and azithromycin-resistant Salmonella Typhi.

Prevalence of Escherichia coli ST1193 Causing Intracranial Infection in Changsha, China.

ST1193 is an emerging new virulent and resistant clone among Escherichia coli with a tendency to spread rapidly across the globe. However, the prevalence of intracranial infection-causing E. coli ST1193 is rarely reported. This study aimed at determining the prevalence of E. coli ST1193 isolates, causing intracranial infections in Changsha, central China. A total of 28 E. coli isolates were collected from the cerebrospinal fluid of patients with intracranial infection over a four-year period. All isolates were differentiated using multilocus sequence typing (MLST), and phylogenetic grouping, and tested for antibiotic resistance. MLST analysis showed 11 sequence types (ST) among the 28 E. coli isolates. The most prevalent ST was B2-ST1193 (28.6%, 8/28), followed by B2-ST131 (21.4%, 6/28) and F-ST648 (10.7%, 3/28). Of the eight ST1193 isolates, three carried CTX-M-55, and one carried CTX-M-27. All eight ST1193 isolates were resistant to Ciprofloxacin, showing gyrA1AB/parC4A mutations. Two ST1193 isolates carried the aac(6′)-Ib-cr gene. All ST1193 isolates were recovered from infants with meningitis, with a fatal outcome for one three-month-old infant. ST1193 has emerged as the predominant type of E. coli strain causing intracranial infections in Changsha, China. This study highlights the importance of implementing appropriate surveillance measures to prevent the spread of this emerging public health threat.

Lineages, Virulence Gene Associated and Integrons among Extended Spectrum β-Lactamase (ESBL) and CMY-2 Producing Enterobacteriaceae from Bovine Mastitis, in Tunisia.

Extended Spectrum Beta-Lactamase (ESBL) Enterobacteriaceae are becoming widespread enzymes in food-producing animals worldwide. Escherichia coli and Klebseilla pneumoniae are two of the most significant pathogens causing mastitis. Our study focused on the characterization of the genetic support of ESBL/pAmpC and antibiotic resistance mechanisms in cefotaxime-resistant (CTXR) and susceptible (CTXS) Enterobacteriaceae isolates, recovered from bovine mastitis in Tunisia, as well as the analyses of their clonal lineage and virulence-associated genes. The study was carried out on 17 ESBL/pAmpC E. coli and K. pneumoniae and 50 CTXS E. coli. Detection of resistance genes and clonal diversity was performed by PCR amplification and sequencing. The following β-lactamase genes were detected: blaCTX-M-15 (n = 6), blaCTX-M-15 + blaOXA-1 (2), bla CTX-M-15 + blaOXA-1 + blaTEM-1b (2), blaCTX-M-15 + blaTEM-1b (4), blaCMY-2 (3). The MLST showed the following STs: ST405 (n = 4 strains); ST58 (n = 3); ST155 (n = 3); ST471 (n = 2); and ST101 (n = 2). ST399 (n = 1) and ST617 (n = 1) were identified in p(AmpC) E. coli producer strains. The phylogroups A and B1 were the most detected ones, followed by the pathogenic phylogroup B2 that harbored the shigatoxin genes stx1/stx2, associated with the cnf, fimA, and aer virulence factors. The qnrA/qnrB, aac(6′)-Ib-cr genes and integrons class 1 with different gene cassettes were detected amongst these CTXR/S isolated strains. The presence of different genetic lineages, associated with resistance and virulence genes in pathogenic bacteria in dairy farms, may complicate antibiotic therapies and pose a potential risk to public health.

Prevalence of Plasmid-Mediated Quinolone Resistance Genes and Molecular Typing of Klebsiella pneumoniae Isolates from Blood Cultures in Milad Hospital, Tehran, Iran, Within 2018 - 2019

Background: Plasmid-mediated quinolone resistance (PMQR) determinants are commonly characterized in Klebsiella pneumoniae isolates worldwide and complicate the treatment of infections caused by this bacterium. Objectives: This study aimed to investigate the prevalence of PMQR determinants and molecular typing of blood isolates of K. pneumoniae in Milad hospital in Tehran, Iran, within 2018 - 2019. Methods: A total of 100 K. pneumoniae isolates were tested for susceptibility to quinolones using the disk diffusion method. The minimum inhibitory concentrations (MICs) of ciprofloxacin (CIP) and levofloxacin (LEV) were determined using the microdilution broth method. The PMQR determinants were detected by polymerase chain reaction (PCR) assay, and the genetic relationship between the isolates was assessed using enterobacterial repetitive intergenic consensus (ERIC)-PCR. Results: The resistance rates of the isolates to LEV, CIP, nalidixic acid, and norfloxacin were determined to be 62, 46, 29, and 23%, respectively. Eighty-one isolates were resistant to at least one tested quinolone. A high-level CIP and LEV resistance (MIC > 32 mg/L) was observed in 15 (18.51%) and 36 (44.44%) isolates, respectively. The PMQR genes were detected in 71 (87.65%) isolates. The oqxAB, qnrS, qnrD, qnrB, aac(6')-Ib-cr, qnrA, qepA, and qnrC genes were detected in 71 (87.65%), 30 (37%), 25 (30.8%), 24 (29.6%), 18 (22.2%), 17 (21%), 17 (21%), and 8 (9.9%) isolates, respectively. The ERIC-PCR revealed 64 genotypes among quinolone-resistant isolates. Conclusions: The high prevalence of PMQR genes observed in this study is a significant concern for public health since they can contribute to the spread of fluoroquinolone resistance among clinical isolates. The ERIC-PCR revealed high heterogeneity among the studied isolates, indicating that they emerged from different sources.

Fluoroquinolones resistance of Pseudomonas aeruginosa isolated from different clinical samples

A total of 156 clinical specimens were collected from patients at Al-Sadder Medical City, Al-Furat al-Awsat Hospital, and the burn center between August 2021 and February 2022. According to medical service utilities, Burn center was the highest with total number of samples with 96 samples and the highest recovery rate 17 isolates followed by Alfurat Al-Awsat Hospital with 32 samples and 4 isolates and finally tame Al-Sadder Medical City with only 28 samples and 2 isolates. The results of 23 isolates of Pseudomonas aeruginosa revealed that the most effective agent was ciprofloxacin with (60%) resistance which represents regression in the overall potential while Norfloxacin showed high resistance percentage (73%) (The least effective among them) followed by Ofaloxacin (69%) and levofloxacin with (65%).genetically, the result of aac(6´)-Ib-cr gene showed that 21/23 isolates gave positive result with percentage (91.3%).

Characterization of plasmid-mediated quinolone resistance genes and extended-spectrum beta-lactamases in non-typhoidal Salmonella enterica isolated from broiler chickens.

Background and Aim:Antibiotic-resistant Salmonella is a public health concern. Fluoroquinolones and extended-spectrum beta-lactams are widely used for the treatment of Salmonella infections. This study focused on the detection of plasmid-mediated quinolone resistance (PMQR) and extended-spectrum beta-lactamase (ESBL) genes among multidrug-resistant (MDR) Salmonella enterica isolated from broilers.Materials and Methods:A total of 40 non-typhoidal S. enterica isolates were collected from 28 broiler chicken farms in four Iraqi Governorates. These isolates were examined for their susceptibility to 10 antimicrobial agents by disk-diffusion method followed by polymerase chain reaction assay for the detection of PMQR determinants and ESBLs genes.Results:Salmonella strains revealed high levels of resistance to the following antibiotics: Nalidixic acid 100%, levofloxacin (LEV) 97.5%, amoxicillin-clavulanic acid 95.0%, tetracycline 92.5%, and nitrofurantoin 80.0%. Otherwise, all isolates were susceptible to cefotaxime and ceftriaxone. All isolates were MDR, with 15 different profiles observed. Among 38 amoxicillin/clavulanic acid-resistant Salmonella isolates, 20 (52.6%) had the blaTEM gene, while blaSHV, blaCTX-M, and blaOXA genes were not detected. Only 5 (12.8%) out of 39 LEV-resistant isolates were positive for qnrB, three of which had blaTEM. No qnrC or qnrD, qnrS, aac(6`)-Ib-cr, qunA, and oqxAB genes were found in any of the tested isolates.Conclusion:This study demonstrates that broiler chickens may be considered a potential source for spreading MDR non-typhoidal Salmonella and ESBL traits in poultry production. Therefore, it is important to continuously monitor ESBL and PMQR genes to avoid the spread of resistant strains in the food chain and impact public health.

Characteristics of Stenotrophomonas maltophilia isolates from cystic fibrosis patients in Russia.

Stenotrophomonas maltophilia is a common opportunistic microorganism and an important respiratory pathogen in cystic fibrosis (CF). The aim of this study was to determine antimicrobial resistance phenotypes, sequence-types (ST) and genetic determinants of antibiotic resistance in S. maltophilia strains recovered from CF patients in Russia. S. maltophilia isolates recovered from 170 CF patients were analyzed. Minimum inhibitory concentrations of antibacterial agents were determined using Sensititre Gram Negative GNX2F plates and the results were interpreted according to Clinical and Laboratory Standards Institute (CLSI) criteria. Whole-genome sequencing (WGS) was performed on MGISEQ-2000 platform. SPAdes software, Galaxy, ResFinder, Integrall and PubMLST were used for analysis of WGS data. S. maltophilia strains were identified from 24/170 (14%) CF patients. In total, 25 isolates were detected, two strains were isolated from the same patient. The isolates belonged to 17 different STs, including 5 new STs; ST4 was the most prevalent ST. Resistance to ceftazidime was observed in 60% of strains, to ticarcillin-clavulanate - in 32%, to levofloxacin - in 24%, to trimethoprim/sulfamethoxazole - in 12% of strains. All isolates were susceptible to minocycline. All ST4 isolates were resistant or intermediate to ceftazidime and ticarcillin-clavulanate. In two isolates, the sul1 gene was detected. In one isolate, sul1 was part of a class 1 integron. The detected integron also contained the blaGES-7 and aac(6')-Ib-cr genes. The ST4 sequence-type was the most prevalent ST among S. maltophilia strains recovered from CF patients in Russia. Antibiotic resistance genes, including sul1, blaGES-7, aac(6')-Ib-cr, were detected in single strains.

Antimicrobial Resistance in Escherichia coli from the Broiler Farm Environment, with Detection of SHV-12-Producing Isolates.

Antimicrobial resistance is an important One Health challenge that encompasses the human, animal, and environmental fields. A total of 111 Escherichia coli isolates previously recovered from manure (n = 57) and indoor air (n = 54) samples from a broiler farm were analyzed to determine their phenotypes and genotypes of antimicrobial resistance and integron characterization; in addition, plasmid replicon analysis and molecular typing were performed in extended-spectrum-beta-lactamase (ESBL) producer isolates. A multidrug-resistance phenotype was detected in 46.8% of the isolates, and the highest rates of resistance were found for ampicillin, trimethoprim–sulfamethoxazole, and tetracycline (>40%); moreover, 15 isolates (13.5%) showed susceptibility to all tested antibiotics. None of the isolates showed imipenem and/or cefoxitin resistance. Twenty-three of the one hundred and eleven E. coli isolates (20.7%) were ESBL producers and carried the blaSHV-12 gene; one of these isolates was recovered from the air, and the remaining 22 were from manure samples. Most of ESBL-positive isolates carried the cmlA (n = 23), tet(A) (n = 19), and aac(6′)-Ib-cr (n = 11) genes. The following genetic lineages were identified among the ESBL-producing isolates (sequence type-phylogroup-clonotype): ST770-E-CH116–552 (n = 12), ST117-B2-CH45–97 (n = 4), ST68-E-CH26–382/49 (n = 3), ST68-E-CH26–49 (n = 1), and ST10992-A/B1-CH11–23/41/580 (n = 4); the latter two were detected for the first time in the poultry sector. At least two plasmid replicon types were detected in the ESBL-producing E. coli isolates, with IncF, IncF1B, IncK, and IncHI1 being the most frequently found. The following antimicrobial resistance genes were identified among the non-ESBL-producing isolates (number of isolates): blaTEM (58), aac(6′)-Ib-cr (6), qnrS (2), aac(3)-II (2), cmlA (6), tet(A)/tet(B) (22), and sul1/2/3 (51). Four different gene-cassette arrays were detected in the variable region of class 1 (dfrA1-aadA1, dfrA12-aadA2, and dfrA12-orf-aadA2-cmlA) and class 2 integrons (sat2-aadA1-orfX). This work reveals the worrying presence of antimicrobial-resistant E. coli in the broiler farm environment, with ESBL-producing isolates of SHV-12 type being extensively disseminated.

Occurrence of plasmid-mediated quinolone resistance genes in Pseudomonas aeruginosa strains isolated from clinical specimens in southwest Iran: a multicentral study

This study aimed to assess the presence of qnrA, qnrB, qnrC, qnrD, qnrS, qepA, and aac(6′)-Ib-cr determinants as well as quinolone resistance pattern of clinical isolates of P. aeruginosa in Ahvaz, southwest Iran. A total of 185 clinical isolates of P. aeruginosa were collected from 5 university-affiliated hospitals in Ahvaz, southwest Iran. The disk diffusion method was applied to assess the quinolone resistance pattern. The presence of qnrA, qnrB, qnrC, qnrD, qnrS, qepA, and aac(6′)-Ib-cr genes was investigated by the polymerase chain reaction (PCR) method. Overall, 120 (64.9%) isolates were non-susceptible to quinolones. The most and the less quinolone resistance rates were observed against ciprofloxacin (59.4%) and ofloxacin (45.9%), respectively. The prevalence rates of qnr genes were as follows: qnrA (25.8%), qnrB (29.2%), and qnrS (20.8%). The qnrB gene was the most common type of qnr genes. The qnr genes were occurred in 37.5% (n = 45/120) of quinolne-resistant isolates, simultaneously. The qnrC, qnrD, qepA, and aac(6′)-Ib-cr genes were not recognized in any isolates. In conclusion, the ofloxacin was the most effective quinolone. This study was the first to shed light on the prevalence of PMQR genes among P. aeruginosa isolates in southwest Iran.

Plasmid-Mediated Fluoroquinolone Resistance in Pseudomonas aeruginosa and Acinetobacter baumannii.

Introduction Pseudomonas aeruginosa and Acinetobacter baumannii are important pathogens in health care–associated infections. Fluoroquinolone resistance has emerged in these pathogens. In this study, we aimed to determine the occurrence of plasmid-mediated quinolone resistance (PMQR) determinants ( qnrA , qnrB , qnrS , aac(6′)-Ib-cr , oqxAB , and qepA ) by polymerase chain reaction (PCR) and the transmissibility of plasmid-borne resistance determinants in clinical isolates of P. aeruginosa and A. baumannii . Materials and Methods The study included P. aeruginosa (85) and A. baumannii (45) which were nonduplicate, clinically significant, and ciprofloxacin resistant. Antibiotic susceptibility testing was done by disk diffusion method for other antimicrobial agents, namely amikacin, ceftazidime, piperacillin/tazobactam, ofloxacin, levofloxacin, and imipenem. Minimum inhibitory concentration of ciprofloxacin was determined. Efflux pump activity was evaluated using carbonyl-cyanide m-chlorophenylhydrazone (CCCP). The presence of PMQR genes was screened by PCR amplification. Transferability of PMQR genes was determined by conjugation experiment, and plasmid-based replicon typing was performed. Results Resistance to other classes of antimicrobial agents was as follows: ceftazidime (86.9%), piperacillin/tazobactam (73.8%), imipenem (69.2%), and amikacin (63.8%). The minimal inhibitory concentration (MIC)50 and MIC90 for ciprofloxacin were 64 and greater than or equal to 256 µg/mL, respectively. There was a reduction in MIC for 37 (28.4%) isolates with CCCP. In P. aeruginosa , 12 (14.1%) isolates harbored qnrB , 12 (14.1%) qnrS , 9 (10.5%) both qnrB and qnrS , 66 (77.6%) aac(6′)-Ib-cr , and 3 (3.5%) oqxAB gene. In A. baumannii , qnrB was detected in 2 (4.4%), 1 (2.2%) harbored both the qnrA and qnrS , 1 isolate harbored qnrB and qnrS , 21 (46.6%) aac(6′)-Ib-cr , and 1 (2.2%) isolate harbored oqxAB gene. Notably, qepA gene was not detected in any of the study isolates. Conjugation experiments revealed that 12 (9.2%) were transferable. Of the transconjugants, seven (58.3%) belonged to IncFII type plasmid replicon, followed by four (33.3%) IncA/C and one (8.3%) IncFIC type. Conclusion The plasmid-mediated resistance aac(6′)-Ib-cr gene is primarily responsible for mediating fluoroquinolone resistance in clinical isolates of P . aeruginosa and A. baumannii . The predominant plasmid type is IncFII.

Burden of Fluoroquinolone Resistance in Clinical Isolates of Escherichia coli at the Ho Teaching Hospital, Ghana.

The growing burden of antibiotic resistance is a threat to the management of infections. Infections by Escherichia coli are routinely treated with fluoroquinolone antimicrobial agents. Due to their frequent use, there has been increasing resistance to these drugs. We set out to determine the burden of fluoroquinolone resistance among clinical E. coli isolates at the Ho Teaching Hospital, Ghana. This was a cross-sectional study conducted from July 2018 to June 2019. One hundred and thirty-five E. coli isolates were cultured from various clinical samples. Antimicrobial susceptibility testing was performed using the Kirby-Bauer disk diffusion method with discs of nalidixic acid (NAL), ciprofloxacin (CIP), norfloxacin (NOR) and levofloxacin (LEV). Deoxyribonucleic acid (DNA) was extracted from the resistant isolates for the detection of fluoroquinolone resistant genes by polymerase chain reaction. Ninety of the 135 isolates (66.7%) were resistant to at least one of the four fluoroquinolone drugs investigated. Resistance to NAL, CIP, NOR, and LEV was 51.0%, 51.1%, 38.8% and 35.7% respectively. Out of the fluoroquinolone resistant isolates, 69 carried one or more fluoroquinolone resistant genes. The predominant resistant genes were aac(6')-Ib-cr (48.9%) and qnrD (25.6%). Seven of the isolates carried both qnrS and aac(6')-Ib-cr genes. Two isolates carried 5 different fluoroquinolone resistant genes. High prevalence of resistance to 4 fluoroquinolone drugs was recorded with associated resistant genes. This is a threat to current efforts to control the spread of antibiotic resistance and calls for concerted efforts to curb the spread of these resistant organisms.

Plasmid-mediated quinolone resistance determinants in clinical bacterial pathogens isolated from the Western Region of Ghana: a cross-sectional study.

quinolones are critically important antibiotics that are reserved for treating very severe infections caused by multidrug-resistant bacterial pathogens. However, their indiscriminate uses have resulted in an increased number of resistant strains in many parts of the world including Ghana. We determined the quinolone resistance profile of Gram-negative bacterial pathogens and characterized the underlying molecular determinants of resistance. Gram-negative pathogens obtained from clinical specimens at three hospital laboratories were tested for resistance to quinolones and other commonly used antibiotics. ESBL production among the Enterobacterial isolates was confirmed using the combined disc diffusion method. We then used PCR to determine seven types of plasmid-mediated quinolone resistance genes present in the isolates resistant to nalidixic acid and ciprofloxacin. in this study, 29.5% of the isolates were resistant to ciprofloxacin, with the highest of 50% among E. coli resistance to the other quinolones was levofloxacin (24.4%), norfloxacin (24.9%), and nalidixic acid (38.9%). Significant proportions of the quinolone-resistant isolates were ESBL producers (P-values < 0.001). The aac(6´)-Ib-cr, qnrS, oqxA, and qepA genes were present in 43 (89.6%), 27 (56.3%), 23 (47.9%), and one (2.1%) of the isolates, respectively. None of the isolates tested positive to qnrA, qnrB, and oqxB genes. The presence of the aac(6´)-Ib-cr gene positively correlated with resistance to ceftriaxone, cefotaxime, and gentamicin (P-values < 0.05). high proportions of Gram-negative bacterial isolates were resistant to quinolones and most of these isolates possessed multiple PMQR genes. There is a need to implement measures to limit the spread of these organisms.

Pan-Resistome Characterization of Uropathogenic Escherichia coli and Klebsiella pneumoniae Strains Circulating in Uganda and Kenya, Isolated from 2017-2018.

Urinary tract infection (UTI) develops after a pathogen adheres to the inner lining of the urinary tract. Cases of UTIs are predominantly caused by several Gram-negative bacteria and account for high morbidity in the clinical and community settings. Of greater concern are the strains carrying antimicrobial resistance (AMR)-conferring genes. The gravity of a UTI is also determined by a spectrum of other virulence factors. This study represents a pilot project to investigate the burden of AMR among uropathogens in East Africa. We examined bacterial samples isolated in 2017–2018 from in- and out-patients in Kenya (KY) and Uganda (UG) that presented with clinical symptoms of UTI. We reconstructed the evolutionary history of the strains, investigated their population structure, and performed comparative analysis their pangenome contents. We found 55 Escherichia coli and 19 Klebsiella pneumoniae strains confirmed uropathogenic following screening for the prevalence of UTI virulence genes including fimH, iutA, feoA/B/C, mrkD, and foc. We identified 18 different sequence types in E. coli population while all K. pneumoniae strains belong to ST11. The most prevalent E. coli sequence types were ST131 (26%), ST335/1193 (10%), and ST10 (6%). Diverse plasmid types were observed in both collections such as Incompatibility (IncF/IncH/IncQ1/IncX4) and Col groups. Pangenome analysis of each set revealed a total of 2862 and 3464 genes comprised the core genome of E. coli and K. pneumoniae population, respectively. Among these are acquired AMR determinants including fluoroquinolone resistance-conferring genes aac(3)-Ib-cr and other significant genes: aad, tet, sul1, sul2, and cat, which are associated with aminoglycoside, tetracycline, sulfonamide, and chloramphenicol resistance, respectively. Accessory genomes of both species collections were detected several β-lactamase genes, blaCTX-M, blaTEM and blaOXA, or blaNDM. Overall, 93% are multi-drug resistant in the E. coli collection while 100% of the K. pneumoniae strains contained genes that are associated with resistance to three or more antibiotic classes. Our findings illustrate the abundant acquired resistome and virulome repertoire in uropathogenic E. coli and K. pneumoniae, which are mainly disseminated via clonal and horizontal transfer, circulating in the East African region. We further demonstrate here that routine genomic surveillance is necessary for high-resolution bacterial epidemiology of these important AMR pathogens.

118. Machine Learning Approaches to Predicting Treatment Outcomes for Carbapenem-Resistant Enterobacterales in a Region with High Prevalence of Non-Carbapenemase Producers

BackgroundCarbapenem-resistant Enterobacterales are a growing threat globally. Early detection of CRE is necessary for appropriate treatment and infection control measures. Many hospital labs can test for carbapenemase production; however, in some regions, including South Texas, the majority of CRE are non-carbapenemase producing (NCPE). This study had two interrelated aims to develop decision rules tailored to a region with high prevalence of NCPE to predict 1) antimicrobial resistance (AMR) from whole genome sequencing (WGS) data and 2) CRE treatment outcomes.MethodsTo better understand links between resistome, phenotypic AMR, and prediction of outcomes for CRE, we developed decision rules to build machine learning prediction models. We conducted WGS and antibiotic susceptibility testing (21 antibiotics) on CRE isolates from unique patients across 5 hospitals in the South Texas region between 2013 and 2020. Day 30 outcomes were based on desirability of outcome ranking (DOOR). The overall classification accuracies of the models are reported.ResultsOverall 146 CRE isolates were included, 97 were used to train each model, and 49 were used for validation. Among the K. pneumoniae and E. coli CRE isolates that were available with susceptibility data, the majority (62%) were NCPE. For the clinical recovery model (DOOR), the combination of admission ICU status, piperacillin-tazobactam (PT) MIC > 16, presence of sul gene, and polymyxin-sparring regimens associated with an overall accuracy of 95% for having a worse DOOR. Majority (60%) of patients were empirically treated with piperacillin-tazobactam; notably, less than 33% isolates had PT MIC ≤ 16. Interestingly, combined effects of isolates that did not harbor carbapenemases, blaOXA-1, blaCTX-M-15, blaCMY, or aac(6’)ib-cr genes resulted in a decision rule with a 95.7% overall accuracy for susceptibility to PT (MIC < 16 ug/mL).ConclusionHerein, we used machine learning approaches to predict AMR and treatment-based outcomes linked with WGS data in a region with predominantly NCPE infections. Machine learning can obtain a model that can make repeatable predictions, further data can improve the accuracy and guide tailored clinical decision-making.Disclosures Grace Lee, PharmD, PhD, BCPS, Merck Co. (Grant/Research Support)NIA/NIH (Research Grant or Support)

Antibiotics and antibiotic resistance genes in anaerobic digesters and predicted concentrations in agroecosystems

In recent decades, the innovative practice of management and valorization of agrozootechnical waste as energy through anaerobic digestion (AD) has been rapidly growing. However, whether applying digestate to soil as biofertilizer can be a source of antibiotics (ABs) and antibiotic resistance genes (ARGs) has not been fully investigated so far. In this work the ARGs responsible for sulfamethoxazole (SMX) resistance (sul1, sul2), ciprofloxacin (CIP) resistance (qnrS, qepA, aac-(6′)-Ib-cr) and the mobile genetic element intl1, together with the concentrations of the antibiotics SMX and CIP, were measured in several anaerobic digesters located in Central Italy. Based on these results, the concentrations of antibiotics and ARGs which can potentially reach soil through amendment with digestate were also estimated. The highest CIP and SMX concentrations were found during winter and spring in anaerobic digesters. The highest ARG abundances were found for the aac-(6’)-Ib-cr and sul2 genes. The overall results showed that application of digestate to soil does not exclude AB contamination and spread of ARGs in agroecosystems, especially in the case of ciprofloxacin, owing to its high intrinsic persistence.

Impact of target site mutations and plasmid associated resistance genes acquisition on resistance of Acinetobacter baumannii to fluoroquinolones

Among bacterial species implicated in hospital-acquired infections are the emerging Pan-Drug Resistant (PDR) and Extensively Drug-Resistant (XDR) Acinetobacter (A.) baumannii strains as they are difficult to eradicate. From 1600 clinical specimens, only 100 A. baumannii isolates could be recovered. A high prevalence of ≥ 78% resistant isolates was recorded for the recovered isolates against a total of 19 tested antimicrobial agents. These isolates could be divided into 12 profiles according to the number of antimicrobial agents to which they were resistant. The isolates were assorted as XDR (68; 68%), Multi-Drug Resistant (MDR: 30; 30%), and PDR (2; 2%). Genotypically, the isolates showed three major clusters with similarities ranging from 10.5 to 97.8% as revealed by ERIC-PCR technique. As a resistance mechanism to fluoroquinolones (FQs), target site mutation analyses in gyrA and parC genes amplified from twelve selected A. baumannii isolates and subjected to sequencing showed 12 profiles. The selected isolates included two CIP-susceptible ones, these showed the wild-type profile of being have no mutations. For the ten selected CIP-resistant isolates, 9 of them (9/10; 90%) had 1 gyrA/1 parC mutations (Ser 81 → Leu mutation for gyrA gene and Ser 84 → Leu mutation for parC gene). The remaining CIP-resistant isolate (1/10; 10%) had 0 gyrA/1 parC mutation (Ser 84 → Leu mutation for parC gene). Detection of plasmid-associated resistance genes revealed that the 86 ciprofloxacin-resistant isolates carry qnrA (66.27%; 57/86), qnrS (70.93%; 61/86), aac (6')-Ib-cr (52.32%; 45/86), oqxA (73.25%; 63/86) and oqxB (39.53%; 34/86), while qepA and qnrB were undetected in these isolates. Different isolates were selected from profiles 1, 2, and 3 and qnrS, acc(6,)-ib-cr, oqxA, and oqxB genes harbored by these isolates were amplified and sequenced. The BLAST results revealed that the oqxA and oqxB sequences were not identified previously in A. baumannii but they were identified in Klebsiella aerogenes strain NCTC9793 and Klebsiella pneumoniae, respectively. On the other hand, the sequence of qnrS, and acc(6,)-ib-cr showed homology to those of A. baumannii. MDR, XDR, and PDR A. baumannii isolates are becoming prevalent in certain hospitals. Chromosomal mutations in the sequences of GyrA and ParC encoding genes and acquisition of PAFQR encoding genes (up to five genes per isolate) are demonstrated to be resistance mechanisms exhibited by fluoroquinolones resistant A. baumannii isolates. It is advisable to monitor the antimicrobial resistance profiles of pathogens causing nosocomial infections and properly apply and update antibiotic stewardship in hospitals and outpatients to control infectious diseases and prevent development of the microbial resistance to antimicrobial agents.

The existence and expression of aminoglycoside resistance genes among multidrug-resistant Escherichia coli isolates in intensive care unit centers

BackgroundE. coli is among the substantial agents of nosocomial infections. The aim of this study was detection of the aminoglycoside resistance genes and their expression among multidrug-resistant (MDR) E. coli isolates in intensive care units (ICUs). MethodsTotally, 500 isolates of E. coli were collected from ICU centers in Baghdad, Iraq. The antibiotic susceptibility profile of them was determined using disk diffusion test in which seven antibiotics were employed. The gentamicin minimum inhibitory concentration (MIC) was determined using agar dilution method with an antibiotic dilution range of 0.5–256 μg/mL. The polymerase chain reaction (PCR) was implemented to amplify the aac(3)-IIa, ant(2″)-Ia, aac(6′)-lb, aac(3)-Ia, ant(4′)-IIb and aac(6′)-Ib-genes responsible for aminoglycosides resistance. The expression of genes was compared between MDR and non-MDR strains. ResultsThe patients' mean age included 57.33 ± 2 years (66% females vs 34% males, p = 0.0233). Four-hundred and fifty (90%) of isolates were from urinary tract infections (UTIs) and 24 from trachea (48%). Three-hundred and sixty (70%) of isolates were MDR, 260 (52%) of which being resistant to gentamicin. Prior hospitalization (p = 0.0261), prior β-lactams (p < 0.001) and aminoglycoside (p = 0.0113) consumption and diabetes (0.0117) were significant risk factors of MDR-E. coli isolation. The gentamicin MIC of MDR-E. coli included 6–256 μg/mL, 234 (46.8%) of which being in resistant range (64–256 μg/mL). The predominant gene included the aac(3)-IIa (92.6%, n = 463, MIC range: 32–256 μg/mL), followed by aac(6′)-Ib and aac(6′)-Ib-cr, aac(3)-Ia genes being in 224 (44.8%), 37 (7.4%) and 16 (2.2%) and 0% of isolates, respectively. The AMEs gene were detected among isolates with MICs>16 μg/mL and aac(3)-IIa, aac(6′)-Ib and aac(6′)-Ib-cr genes were significantly associated with MIC range of 128–256 μg/mL (p < 0.001). Most of genes expression was significantly higher in MDR than non-MDR strains. ConclusionThe majority of E. coli from ICU was MDR. Prior hospitalization, β-lactams and aminoglycoside consumption and diabetes were significant risk factors of MDR-E. coli isolation. Moreover, the aac(3)-IIa and aac(6′)-lb genes were predominant AMEs which were associated with higher MIC levels. Mechanisms affecting drug resistance genes expression in MDR strains maybe different from those in non-MDR strains.

Molecular Characterization of Cephalosporin and Fluoroquinolone Resistant Salmonella Choleraesuis Isolated from Patients with Systemic Salmonellosis in Thailand.

The antimicrobial resistance of nontyphoidal Salmonella has become a major clinical and public health problem. Southeast Asia has a high level of multidrug-resistant Salmonella and isolates resistant to both fluoroquinolone and third-generation cephalosporins. The incidence of co-resistance to both drug classes is a serious therapeutic problem in Thailand. The aim of this study was to determine the antimicrobial resistance patterns, antimicrobial resistance genes and genotypic relatedness of third-generation cephalosporins and/or fluoroquinolone-resistant Salmonella Choleraesuis isolated from patients with systemic salmonellosis in Thailand. Antimicrobial susceptibility testing was performed using the agar disk diffusion method, and ESBL production was detected by the combination disc method. A molecular evaluation of S. Choleraesuis isolates was performed using PCR and DNA sequencing. Then, a genotypic relatedness study of S. Choleraesuis was performed by pulse field gel electrophoresis. All 62 cefotaxime-resistant S. Choleraesuis isolates obtained from 61 clinical specimens were multidrug resistant. Forty-four isolates (44/62, 71.0%) were positive for ESBL phenotypes. Based on the PCR sequencing, 21, 1, 13, 23, 20 and 6 ESBL-producing isolates harboured the ESBL genes blaCTX-M-14, blaCTX-M-15, blaCTX-M-55, blaCMY-2, blaACC-1 and blaTEM-1, respectively. This study also found that nine (9/62, 14.5%) isolates exhibited co-resistance to ciprofloxacin and cefotaxime. All of the co-resistant isolates harboured at least one PMQR gene. The qnr genes and the aac(6′)-Ib-cr gene were the most prevalent genes detected. The QRDR mutation, including the gyrA (D87Y and D87G) and parC (T57S) genes, was also detected. PFGE patterns revealed a high degree of clonal diversity among the ESBL-producing isolates.

Antibiotic Resistance and Antibiotic Resistance Genes Among Edwardsiella Tarda Isolated from Fish

Edwardsiella tarda is a common fish pathogen, causing significant septicemic diseases in fish. This study was carried out to investigate prevalence of E. tarda among Tilapia zilli at EL manzla lake, Dakahlia governorate and characterize the isolates phenotypically and genotypically in addition to detection of multi-drug resistance genes (B-lactames genes and plasmid-mediated quinolone resistant genes by PCR assay. Therefore, (100) samples of Tilapia zilli collected from different localities. Fish samples were subjected to clinical and post-mortem examination then bacteriological examination from liver, kidney and spleen. The suspected isolates were characterized by cultural and morphological characters, some conventional biochemical tests and API 20E system then by PCR assay. Eighteen % isolates were characterized as E. tarda. Furthermore, detection of antimicrobial resistance genes by PCR, the recovered isolates harbored blaSHV and blaOXA-1 with a prevalence of 50% and 25%, respectively with no detection of aac (6′)-Ib-cr and qepA genes in examined isolates. The results of antibiotics sensitivity showed resistance to Nalidixic acid, Lincomycin, Amoxicillin and Norfloxacin and most of isolate were found to be sensitive to Florfenicol and Neomycin.55.6% of recovered isolates showed resistance to three antibiotic classes and 27.7% of recovered isolates showed resistance to four antibiotic classes and considered as MDR. The emergence of MDR-strains represents a threat-alarm and PCR is very rapid method for identification of E. tarda isolates which may be helpful in control of Edwardsiellosis.