MDR Isolates Research Articles

ASSESSING PHENOTYPIC SYNERGISTIC ACTIVITY OF CEFTAZIDIMEAVIBACTAM AND AZTREONAM AGAINST CARBAPENEMASE PRODUCING MULTIDRUG RESISTANT GRAM-NEGATIVE ORGANISMS IN A TERTIARY CARE HOSPITAL, NAVI MUMBAI.

Introduction: Multidrug resistance organisms have become a great concern globally. Ceftazidime-avibactam (CAZ/AVI) has a broad-spectrum activity against Ambler molecular Class A (narrow-spectrum & extended-spectrum beta-lactamases (ESBLs), serine carbapenemase including KPC). It also has activity against some Class C (KPC) and Class D carbapenemase including OXA-48-like enzymes in Enterobacteriaceae. The study aims to identify clinically relevant sy Aim: nergy between CAZ/AVI+Aztreonam (ATM) for multidrug resistant isolates (MDR) using E-test/disc method. A total of 60 non- Material & Methods: duplicate clinical isolates of different clinical specimens (blood, pus, sputum, urine, and sterile body uids) were collected prospectively over 3- month period (January – March 2022). Antimicrobial susceptibility determined by Kirby-Bauer disc diffusion method. CAZ/AVI+ATM synergy was determined using E- test/disc method. NMIC 500 BD phoenix automated system conrmed the presence of class of enzymes. Of Results: the 60 MDR isolates, Klebsiella (n=35; 58.3%) was the most common followed by E. coli (n=17; 28.3%). CAZ/AVI was susceptible for 25% (n= 15) of the isolates and the remaining 45(75%) were resistant. CLSI guidelines used for interpretation. Using qualitative E-test/disc diffusion method, addition of aztreonam to CAZ/AVI showed susceptibility for 66% (n=30/45) of CAZ/AVI resistant isolates demonstrating a Reverse D synergy. The remaining 15 (out of 45) isolates showed resistance to the above combination and thereby showing no synergy. The most common class of enzymes were Class D (n=25/60) followed by Class B (n=19/60). In screening for CZA Conclusion: /AVI +ATM synergy, the E-test/disc method provides a quick and practical approach for the real world screening and for the targeted use in the patients. This approach would be more relevant considering NDM and its coproduction prevalence is on the rise with limited antibiotic options for such infections.

Perchlozone Resistance in Clinical Isolates of Mycobacterium tuberculosis

The emergence of drug-resistant tuberculosis forced the development of new drugs and the screening of more effective or less toxic analogues. Mycolic acid biosynthesis is targeted by several antituberculosis drugs, isoniazid being one of the most important in tuberculosis therapy. Recently, perchlozone, acting on another step in the FAS-II cycle, was officially approved for tuberculosis treatment in the Russian Federation and was included in the Russian national clinical guidelines. Using the serial dilution method on 7H10 agar plates for perchlozone and a Sensititre MYCOTB microdilution plate, we analyzed the phenotypic properties of primary clinical isolates of M. tuberculosis and analyzed the molecular determinants of resistance to isoniazid, ethionamide, and perchlozone. We found a wide variation in the MIC of perchlozone from 2 to 64 mg/L, correlating with the overall resistance profile: the MIC was higher for MDR and pre-XDR isolates. The cross-resistance between ethionamide and perchlozone was driven by mutations in the ethA gene encoding monooxygenase responsible for the activation of both drugs. The presumably susceptible to perchlozone and wild-type strains had MICs ranging from 2 to 4 mg/L, and the breakpoint was estimated to be 4 or 8 mg/L. In conclusion, susceptibility to perchlozone is retained for a part of the MDR strains, as is susceptibility to ethionamide, providing the possibility of therapy for such cases based on phenotypic or molecular analysis.

Bacterial profile, antimicrobial resistance, and molecular detection of ESBL and quinolone resistance gene of uropathogens causing urinary tract infection in the southeastern part of Bangladesh.

Humans frequently contract urinary tract infections (UTIs), which can be brought on by uropathogens (UPs) that are multi-drug resistant. Treatment for UTIs brought on by pathogenic UPs that produce extended-spectrum lactamases (ESBLs) is more costly and potentially fatal. As a result, the objective of this study was to use culture, biochemical, and 16S rRNA sequencing to identify and characterize UPs isolated from outpatients in Noakhali, Bangladesh, who had symptoms of UTIs. ESBL gene identification and quinolone resistance gene typing were then performed on the isolates using polymerase chain reaction (PCR). Throughout the trial's 8-month duration, 152 (76%) of 200 urine samples were positive for the presence of UPs. The overall number of UPs recovered was 210, with 39 individuals having multiple UPs present in their samples. Among all of the isolates, Escherichia coli (45.24%, 95/210; 95% confidence interval (CI): 35.15-57.60%), Enterobacter spp. (24.76%, 52/210; CI: 19.15-35.77%), Klebsiella spp. (20.95%; 44/210; CI: 15.15-30.20%), and Providencia spp. (9.05%; 19/210; CI: 4.95-19.25%) were the four most prevalent bacteria found in the isolates. The UPs displayed a very high level of resistance to piperacillin 96.92% (126/130), ampicillin 90% (117/130), nalidixic acid 77.69% (101/130), cefazolin 70% (91/130), amoxicillin 50% (55/130), cefazolin 42.31% (55/130), nitrofurantoin 43.08% (56/130), and ciprofloxacin 33.08% (43/130), whereas resistance to netilmicin (3.85%), amikacin (4.62%), and imipenem (9.23%) was low. Individually, every species of E. coli and Providencia spp. showed greater ampicillin, amikacin, cefazolin, cefazolin, and nalidixic acid resistance than the others. The bivariate results indicate several antibiotic pairings, and isolates had meaningful associations. All MDR isolates were subjected to PCR, which revealed that blaCTX-M-15 genes predominated among the isolates, followed by the blaTEM class (37%). Isolates also had the qnrS, aac-6´-Ib-cr, and gyrA genes. The findings provide worrying indications of a major expansion of MDR isolates in the study locations, particularly the epidemiological balCTX-M 15, with the potential for the transmission of multi-drug-resistant UP strains in the population.

Multidrug-resistant extended spectrum β-lactamase (ESBL)-producing Escherichia coli from farm produce and agricultural environments in Edo State, Nigeria.

Antimicrobial resistance (AMR) is a major public health concern, especially the extended-spectrum β-lactamase-producing (ESBL) Escherichia coli bacteria are emerging as a global human health hazard. This study characterized extended-spectrum β-lactamase Escherichia coli (ESBL-E. coli) isolates from farm sources and open markets in Edo State, Nigeria. A total of 254 samples were obtained in Edo State and included representatives from agricultural farms (soil, manure, irrigation water) and vegetables from open markets, which included ready-to-eat (RTE) salads and vegetables which could potentially be consumed uncooked. Samples were culturally tested for the ESBL phenotype using ESBL selective media, and isolates were further identified and characterized via polymerase chain reaction (PCR) for β-lactamase and other antibiotic resistance determinants. ESBL E. coli strains isolated from agricultural farms included 68% (17/25) from the soil, 84% (21/25) from manure and 28% (7/25) from irrigation water and 24.4% (19/78) from vegetables. ESBL E. coli were also isolated from RTE salads at 20% (12/60) and vegetables obtained from vendors and open markets at 36.6% (15/41). A total of 64 E. coli isolates were identified using PCR. Upon further characterization, 85.9% (55/64) of the isolates were resistant to ≥ 3 and ≤ 7 antimicrobial classes, which allows for characterizing these as being multidrug-resistant. The MDR isolates from this study harboured ≥1 and ≤5 AMR determinants. The MDR isolates also harboured ≥1 and ≤3 beta-lactamase genes. Findings from this study showed that fresh vegetables and salads could be contaminated with ESBL-E. coli, particularly fresh produce from farms that use untreated water for irrigation. Appropriate measures, including improving irrigation water quality and agricultural practices, need to be implemented, and global regulatory guiding principles are crucial to ensure public health and consumer safety.

Investigation of bedaquiline resistance and genetic mutations in multi-drug resistant Mycobacterium tuberculosis clinical isolates in Chongqing, China

BackgroundTo investigate the prevalence and molecular characterization of bedaquiline resistance among MDR-TB isolates collected from Chongqing, China.MethodsA total of 205 MDR-TB isolates were collected from Chongqing Tuberculosis Control Institute between March 2019 and June 2020. The MICs of BDQ were determined by microplate alamarblue assay. All strains were genotyped by melting curve spoligotyping, and were subjected to WGS.ResultsAmong the 205 MDR isolates, the resistance rate of BDQ was 4.4% (9/205). The 55 (26.8%) were from male patients and 50 (24.4%) were new cases. Furthermore, 81 (39.5%) of these patients exhibited lung cavitation, 13 (6.3%) patients afflicted with diabetes mellitus, and 170 (82.9%) isolates belonged to Beijing family. However, the distribution of BDQ resistant isolates showed no significant difference among these characteristics. Of the 86 OFX resistant isolates, 8 isolates were XDR (9.3%, 8/86). Six BDQ resistant isolates (66.7%, 6/9) and two BDQ susceptible isolates (1.0%, 2/196) carried mutations in Rv0678. A total of 4 mutations types were identified in BDQ resistant isolates, including mutation in A152G (50%, 3/6), T56C (16.7%, 1/6), GA492 insertion (16.7%, 1/6), and A274 insertion (16.7%, 1/6). BDQ showed excellent activity against MDR-TB in Chongqing.ConclusionsBDQ showed excellent activity against MDR-TB in Chongqing. The resistance rate of BDQ was not related to demographic and clinical characteristics. Mutations in Rv0678 gene were the major mechanism to BDQ resistance, with A152G as the most common mutation type. WGS has a good popularize value and application prospect in the rapid detection of BDQ resistance.

PREVALENCE, PHENOTYPIC-GENOTYPIC RESISTANCE AND BIOFILM FORMATION OF Staphylococcus aureus IN CHICKEN MEAT WITH REFERENCE TO ITS PUBLIC HEALTH HAZARD

The purpose of this study was to determine the prevalence, antimicrobial resistance in S. aureus and presence of enterotoxigenic as well as biofilm-forming genes in S. aureus bacteria isolated from broiler meat from traditional shops and supermarkets in Sharkia, Egypt. For this determination, two hundred fresh raw chicken meat cotton swabs were collected from the breast and thighs (100 each). S. aureus was highlighted through a coagulase test, and then phenotypic and genotypic characterizations were studied. Uniplex PCR was used to identify the occurrence of enterotoxin genes in the selected isolates. Finally, they were subjected to a biofilm formation test using 96-well flat bottom polystyrene microtiter plates; besides, biofilm-forming genes were investigated. Nineteen isolates out of the 200 samples tested positive for S. aureus (9.5 percent), No Vancomycin resistance strains were obtained, nor did any ciprofloxacin isolates for S. aureus while, all isolates were 100 % resistant for streptomycin, amoxicillin- clavulanic acid and sulfamethoxazole-trimethoprim S. aureus isolates were found resistant to at least one antibiotic. The multiple antibiotic resistance (MAR) index of S. aureus isolates was ranged from 0.3 to 0.7 with an average of 0.4. In The mecicillin resistant S.aureus (MRSA) strains carried mecA gene with a percentage of 5%, while the blaZ gene was distributed with a percentage of 5.5%, 11/200). The obtained isolates gave 60% strong biofilm formation and 40% were non-formers, with nil results for moderate or weak production isolates. The icaA gene was 100%, in comparison to icaD which was zero. It should be noted that strong biofilm former strains were only 100% positive for sea and seb. This study pointed out the higher prevalence of MDR isolates of S. aureus isolates in chicken meat due to inadequate handling and insufficient sanitary equipment and post microbial contamination. This finding highlighted the importance of broiler meat as a reservoir for antimicrobial-resistant strains of S. aureus and biofilm-forming of Staphylococcus aureus.

Epidemiology and Drug Resistance Patterns of Mycobacterium tuberculosis in High-Burden Area in Western Siberia, Russia

Russia is a high-burden area for multidrug-resistant tuberculosis (MDR-TB). Here, we studied the epidemiological situation and drug resistance patterns of Mycobacterium tuberculosis in the Omsk region in Western Siberia. M. tuberculosis isolates (n = 851) were recovered from newly diagnosed TB patients in 2021. The isolates were tested by bacteriological and molecular methods, and long-term epidemiological data were analyzed. The TB incidence dec, this is not variablereased from 93.9 in 2012 to 48.1 in 2021, per 100,000 population, but the primary MDR-TB rate increased from 19.2% to 26.4%. The destructive forms of tuberculosis accounted for 37.8% of all cases, while 35.5% of patients were smear-positive. Of all isolates tested, 55.2% were culture-positive, of which 94.5% were further tested for phenotypic drug resistance and associated mutations. More than half (53.4%) of isolates were drug-resistant, 13.9% were monoresistant and 67.9% were MDR. Among MDR isolates, 40.4% were pre-XDR, and 19.2% were XDR. The spectrum of drug resistance included second-line drugs (new-generation fluoroquinolones, linezolid), which significantly increase the risk of an adverse outcome in patients. In conclusion, our results highlight the critical importance of monitoring drug resistance in circulating M. tuberculosis strains emerging due to ineffective treatment and active transmission.

SG-APSIC1074: Trend of ‘ESKAPE’ and their susceptibility changes for meropenem and levofloxacin during the pandemic at Sardjito Hospital Yogyakarta Indonesia

Objectives: The bacteria in the ‘ESKAPE’ group are monitored due to their ability to resist antibiotic action. During the COVID-19 pandemic at our hospital, the usage of meropenem and levofloxacin as the empirical treatment for bacterial pneumonia increased and might have contributed to the antimicrobial resistance problem. In this study, we evaluated the ESKAPE group infection rates and their susceptibility to antibiotics in Dr. Sardjito Hospital, a referral and academic hospital in Yogyakarta, Indonesia. Methods: Data for ESKAPE pathogens in 2019–2021 were taken from the microbiology laboratory of Dr. Sardjito Hospital and were evaluated. Results: The proportion of ESKAPE isolates among positive cultures during 2019–2021 slightly increased from 49.4% to 48.4% to 50.7% each year (P > .05). The dominant ESKAPE infections were pneumonia, bloodstream infection, and urinary tract infection by K. pneumoniae, and wound infection by P. aeruginosa. The susceptibility pattern of ESKAPE to meropenem decreased from 72% in 2019 to 68% in 2020 but increased to 84% in 2021. To levofloxacin, the susceptibility pattern was decreased in a fluctuating trend from 68% in 2019 to 33% in 2020 and to 39% in 2021. During the COVID-19 pandemic (2020–2021), the pattern of ESKAPE infections was similar to that of 2019. In descending order, the frequency rank was K. pneumoniae, P. aeruginosa, A. baumannii, Enterobacter spp, and S. aureus. The proportions of MDR isolates increased from the prepandemic period to the COVID-19 pandemic era for E. faecium (from 5% to 24.4%), for A. baumannii (from 9.6% to 38.5%), and for P. aeruginosa (from 7.4% to 13.5%) (P < .05). These patterns did not differ between non–COVID-19 patients and COVID-19 patients. These results highlight the general impact of overused antibiotics beyond COVID-19 patients. Usage of watched and restricted antibiotics must be more controlled because bacterial coinfection and superinfection in COVID-19 patients was relatively low. Conclusions: During the COVID-19 pandemic, ESKAPE infections increased and their susceptibility to meropenem and levofloxacin decreased. Tight control of antibiotic usage is needed.

Characterization of chicken eggs associated Escherichia coli and Staphylococcus aureus for biofilm production and antimicrobial resistance traits

The present study assessed the prevalence, virulence characteristics, antimicrobial resistance and biofilm-forming ability of E. coli and S. aureus recovered from egg samples in Ludhiana, Punjab. A total of 393 samples from hatcheries (n = 238), retail shops (n = 94), and households (n = 61) were collected. The prevalence of E. coli was observed as 11.70% and 9.16% for S. aureus. A total of 41.30% of E. coli isolates were positive for aggR gene and 52.17% were for fimA gene; while 36.11% of the S. aureus isolates were positive for coa gene. A high proportion of E. coli (76.10%) and S. aureus (69.44%) isolates were resistant toward ≥3 tested antibiotic classes. A total of 39.13% of E. coli isolates were moderate biofilm former, whereas the majority of the S. aureus (41.67%) were weak biofilm former. No significant difference regarding biofilm formation was observed between MDR and non-MDR isolates of E. coli and S. aureus. Biofilm genes viz., fimC and crl were reported in 43.47% and 80.43% of E. coli isolates, respectively; while icaA and icaD genes were reported in 58.34% and 47.22% of S. aureus isolates, respectively. A strong metabolic activity among 52.17% of E. coli and 41.66% of S. aureus isolates was observed using XTT assay. The present study highlights the need for applied food safety measures across the egg production chain of the region to prevent the development of MDR strains and biofilms.

P13 Association of biofilm production with multidrug resistance in clinical isolates of Pseudomonas aeruginosa

Abstract Background Pseudomonas aeruginosa is among the most important cause of life threatening hospital acquired infections. It is considered as top listed public health threat worldwide due to its ability to produce toxins, virulence factors, biofilms and resist multiple antibiotics. Biofilm formation is a pathogenic mechanism harboured by this pathogen which further elevates its resistance to antibiotics and host defence system. Objectives To detect the phenotypic and genotypic biofilm forming potential and its association with multi drug resistance in P. aeruginosa clinical isolates. Methods In the present study, isolation of P. aeruginosa was performed by conventional and molecular method using polymerase chain reaction (PCR) from (n=200) clinical samples. Kirby Bauer method was performed for detection of MDR isolates. The biofilm forming ability of P. aeruginosa isolates was detected by microtitre plate assay (MPA) while the genes associated with biofilm formation (pelA and pslA) were detected by PCR. Results The results of study revealed that P. aeruginosa was detected in 26% (52/200) isolates with highest frequency 38.63% in burn wound samples followed by 22.22% in sputum samples and 20.58% in surgical wound samples. Highest resistant was found to ceftriaxone (94%) followed by meropenem (92%), imipenem (90%) and cefotaxime (90%) while (n=20) isolates were detected as MDR. Biofilm formation of variable degree was detected in 49 (94%) isolates. Additionally it was recorded that all the MDR isolates exhibits biofilm formation including 55% as moderately, 40% as strongly and 5% as weakly adherent. Among the MDR P. aeruginosa, the biofilm-associated gene pelA was detected in 95% while pslA was found in 90% isolates. Conclusions It was concluded that biofilm forming P. aeruginosa are more resistant to tested antibiotics and strongly linked with presence of pelA and pslA genes. This presence of MDR strains along with biofilm formation is depicting a lethal combination of bacterial armory that poses a serious threat for public health.

Antimicrobial resistance characteristics of multidrug resistance and extended-spectrum beta-lactamase producing Escherichia coli from several dairy farms in Probolinggo, Indonesia

Abstract. Widodo A, Lamid M, Effendi MH, Khailrullah AR, Kurniawan SC, Silaen OSM, Riwu KHP, Yustinasari LR, Afnani DA, Dameanti FNAEP, Ramandinianto SC. 2022. Antimicrobial resistance characteristics of multidrug resistance and extended spectrum beta-lactamase producing Escherichia coli from several dairy farms in Probolinggo, Indonesia. Biodiversitas 23: 215-221. Escherichia coli bacteria initially reside in the digestive tract of humans and animals but are able to adapt to new environments that are different from their initial habitat. The pathogenicity of E. coli can occur when these bacteria grow more than normal limits, produce toxins, and are resistant to certain types of antibiotics. The purpose of this study was to investigate the antimicrobial resistance characteristics of MDR and ESBL-producing E. coli from several dairy farms in the Probolinggo district of East Java province, Indonesia. A total of 150 samples consisting of 109 milk and 41 environmental samples from 41 dairy farms were used for isolation. TSIA and IMViC biochemical tests were used to identify E. coli bacteria. Escherichia coli resistance profile was obtained through disc diffusion test on several antibiotics, namely tetracycline, streptomycin, trimethoprim, chloramphenicol and aztreonam. Escherichia coli that was resistant to 3 or more antibiotics was defined as MDR. The results of isolation and identification obtained 124 (82.6%) isolates characterizing E. coli bacteria. The antimicrobial susceptibility test of E. coli showed 9 (7.26%) MDR isolates and 2 (22.22%) ESBL isolates by double-disc synergy test (DDST). MDR E. coli was dominated by the pattern of antimicrobial drug resistance TE-S-W (tetracycline, streptomycin, trimethoprim) with a total of 8 (38.10%) isolates, followed by antimicrobial drug resistance pattern TE-S-W-ATM (tetracycline, streptomycin, trimethoprim, aztreonam) with one (4.76%) E. coli isolates. The pattern of antimicrobial drugs of ESBL E. coli showed in one (11.11%) sample of ESBL E. coli from a milk sample with the pattern of TE-S-W-ATM (tetracycline, streptomycin, trimethoprim, aztreonam) and one (11.11%) sample ESBL E. coli (AL 30) from the environmental sample with a pattern of TE-S-W pattern (tetracycline, streptomycin, trimethoprim). The discovery of MDR E. coli isolates and ESBL E. coli from milk and environmental samples at several dairy farms in Probolinggo district, East Java, Indonesia is a matter of concern and requires real action to reduce antibiotic resistance.

Role of MexAB-OprM efflux pump in the emergence of multidrug-resistant clinical isolates of Pseudomonas aeruginosa in Mazandaran province of Iran.

Multidrug-resistant clinical isolates can cause many therapeutic problems. The MexAB-OprM efflux pump plays a significant role in expelling toxins and drugs from the bacterial cells resulting in multidrug-resistant Pseudomonas aeruginosa isolates. This study aimed to investigate the effect of the MexAB-OprM efflux pump in the emergence of multidrug-resistant clinical isolates of P. aeruginosa. For the present study, 100 clinical isolates of P. aeruginosa were collected from different wards of teaching hospitals (2018-2019). After confirmation and detection of bacteria by standard methods, the antibiotic resistance pattern of the isolates was determined by the disk agar diffusion method. Also, the minimum inhibitory concentration (MIC) of ciprofloxacin was measured in the presence and absence of phenylalanine arginine beta-naphthylamide by the broth microdilution method. Then, the real-time PCR was used to investigate the expression level of the mexB gene compared to the standard PAO1 strain. Forty-one/100 isolates exhibited multidrug-resistant phenotype (MDR), while piperacillin-tazobactam and levofloxacin were the most and least effective antibiotics tested, respectively. Also, 54/100 isolates showed no increased expression of mexB gene compared to the standard PAO1 strain. However, among the 41 MDR isolates, 12 (29.26%) showed a more than three-fold increase in the expression level of the mexB gene. In this study, a significant relationship was observed between the resistance to tested antibiotics in MDR strains and the increased expression of the mexB gene. We found that increasing the expression of the mexB gene can cause the emergence of multidrug-resistant strains by increasing the minimum inhibitory concentration of the antibiotics. Then, we need to evaluate the resistance mechanisms separately in different area of a country to improve the antibiotic stewardship.

Susceptibility of Salmonella serotypes isolated from meat and meat contact surfaces to thermal, acidic, and alkaline treatments and disinfectants.

The present study was conducted to evaluate the response of 29 Salmonella isolates to exposure to thermal (60°C for 2min), acidic (pH2.9 for 30 min), and alkaline (pH 11 for 60 min) treatments and investigate the susceptibility of the isolates and their biofilms to disinfectants. The reductions of Salmonella isolates populations subjected to each treatment were analyzed according to their isolation source, serotype, antibiotic resistance pattern, and biofilm formation ability. Median reductions for all of Salmonella isolates populations after thermal, acidic, and alkaline treatments were 1.8, 2.1, and 0.7 log CFU/ml, respectively. The isolates behavior under stress conditions were not related to their isolation source, serotype, or biofilm formation ability. The median reduction after alkaline treatment in non-MDR (multidrug- resistant) isolates populations was significantly (p < .05) higher than MDR isolates. The median reduction in biofilms of moderate biofilm producers by disinfectants was significantly (p < .05) higher than that of strong biofilm producers. In conclusion, Salmonella isolates showed the highest susceptibility to acidic treatment and MDR isolates were more resistant to alkaline treatment than non-MDR ones. The current study also revealed that the strong biofilm producer isolates were more resistant to disinfectants than moderate biofilm producers. This study facilitated the understanding of the relationship between Salmonella characteristics (isolation source, serotype, antibiotic resistance pattern, and biofilm formation ability) and its susceptibility to thermal, acidic, and alkaline treatments and disinfectants. The findings are helpful for the prevention and control of Salmonella.

Detection of Genes Related to Linezolid Resistance (poxtA, cfr, and optrA) in Clinical Isolates of Enterococcus spp. from Humans: A First Report from Iran.

Enterococci may develop resistance to linezolid through chromosomal mutations that involve specific linezolid resistance genes, such as cfr, optrA, and poxtA. The objective of this study was to evaluate the antibiotic susceptibility of enterococcal isolates and identify cfr, optrA, and poxtA genes in MDR isolates. Enterococcal isolates were collected from various clinical specimens at Al-Zahra, Amin, and Khorshid Hospitals in Isfahan. The Enterococcus isolates were identified as belonging to the E. faecalis and E. faecium species by using specific gene (D alanine D alanine ligase ddl) sets in PCR. To detect cfr, optrA, and poxtA genes among the species, a multiplex-PCR assay was performed. Out of 175 isolates, E. faecalis predominated 129/175 (73.7%). Furthermore, the prevalence of vancomycin-resistant Enterococci (VRE) and linezolid-resistant Enterococci (LRE) was 29.7% and 4%, respectively. The overall prevalence of MDR was 91.1%, 68.9%, and 66.6% of E. faecium, E. faecalis, and other Enterococcus spp., respectively. Interestingly, the frequency of optrA (71.4%) in E. faecium and poxtA and crf (42.8%) in E. faecalis were detected among LRE species. A statistically significant relationship (P < 0.05) was found between the presence of the three genes and the occurrence of LRE. This is the first study to report the detection of linezolid resistance genes (cfr, optrA, and poxtA) in clinical Enterococcus spp. isolates from Iran, conducted at Isfahan University of Medical Sciences hospitals. The emergence of enterococcal strains that resist linezolid is concerning as it can lead to the spread of resistant strains among patients, resulting in treatment failure.

Molecular Characterization of Resistance to Second-Line Anti-Mycobacterial Drugs among Clinical Isolates of Multidrug-Resistant Mycobacterium tuberculosis

The emergence of multidrug resistance and extensively drug-resistant tuberculosis is a serious public health crisis. Using rapid and inexpensive molecular methods such as HRM assay in the detection of second-line drugs resistance in M. tuberculosis would be helpful in the treatment and control of XDR tuberculosis cases. MDR-TB isolates were collected from Iranian tuberculosis laboratories. Drug susceptibility test performed via the indirect proportion method utilizing LJ Medium. Susceptibility to ciprofloxacin, ofloxacin, amikacin, kanamycin, and capreomycin, as second-line anti-tuberculosis agents were assessed. Single point mutations in gyrA, rrs and eis genes were detected via HRM assay and DNA sequencing. A DST test was performed for 56 MDR isolates and at least 27 (48.2%) isolates were resistant to CIP or OFL. Also, 14 (25%), 12 (21.4%), and 15 (26.7%) isolates were resistant to capreomycin, amikacin, and kanamycin, respectively. D94G, A90V, and G88C mutations were the most frequent mutations in gyrA gene. Also, A1401G mutation was detected more than the other mutations in rrs gene. The frequency of CIP/OFL and AMK/CAP/KAN-resistant TB is considerable among Iranian tuberculosis cases. HRM assay is a rapid and inexpensive test and can detect important mutation-based drug resistance in MDR-TB and XDR-TB isolates.

Multi-drug Resistance, β-Lactamases Production, and Coexistence of bla NDM-1 and mcr-1 in Escherichia coli Clinical Isolates From a Referral Hospital in Kathmandu, Nepal.

The ability of pathogenic Escherichia coli to produce carbapenemase enzymes is a characteristic that allows them to resist various antibiotics, including last-resort antibiotics like colistin and carbapenem. Our objectives were to identify rapidly developing antibiotic resistance (AR), assess β-lactamases production, and detect mcr-1 and bla NDM-1 genes in the isolates. A prospective cross-sectional study was carried out in a referral hospital located in Kathmandu from November 2019 to December 2020 using standard laboratory and molecular protocols. Among 77 total E. coli isolates, 64 (83.1%) of them were categorized as MDR. Phenotypically 13 (20.3%) colistin-resistant, 30 (46.9%) ESBL and 8 (12.5%) AmpC producers, and 5 (7.8%) ESBL/AmpC co-producers were distributed among MDR-E. coli. Minimum inhibitory concentrations (MIC) against the majority of MDR isolates were exhibited at 1 g/L. Of these 77 E. coli isolates, 24 (31.2%) were carbapenem-resistant. Among these carbapenem-resistant bacteria, 11 (45.9%) isolates were reported to be colistin-resistant, while 15 (62.5%) and 2 (8.3%) were MBL and KPC producers, respectively. Out of 15 MBL producers, 6 (40%) harbored bla NDM-1, and 8 (61.5%) out of 13 colistin-resistant pathogens possessed mcr-1. The resistance by colistin- and carbapenem were statistically associated (P < .001). However, only 2 (18.2%) of the co-resistant bacteria were found to have both genes. Our study revealed the highly prevalent MDR and the carbapenem-resistant E. coli and emphasized that the pathogens possess a wide range of capabilities to synthesize β-lactamases. These findings could assist to expand the understanding of AR in terms of enzyme production.

Urinary tract Infection in Female Patients Visiting Tertiary Care Hospital and Determination of ESBL Producing MDR Isolates

Objectives: The main purpose of this study was to assess the urinary tract infection (UTI) in female patients and determine the extended spectrum of beta-lactamase producers among MDR isolates. Methods: A three-month hospital-based cross-sectional study was conducted from Marchto April 2022 at the Microbiology laboratory of Alka hospital, Lalitpur. Following the standard protocol, urine samples were taken for culture and identification. Urine samples were cultured using a semi-quantitative approach and antimicrobial susceptibility test was performed by disc diffusion method. Results: Out of 493 female urine samples examined, significant growth was found in 106 samples. The positive isolates belonged to 5 different species. Out of them E.coli (77.36%) was the major pathogens isolated followed by Klebsiella pneumoniae (6.61%), Klebsiella oxytoca and Pseudomonas aeruginosa (1.88%) as Gram negative bacteria. And the most common Gram positive bacteria was found to be Staphylococcus aureus (12.26%). The most effective drug for Gram negative isolates was Amikacin (93.396%) followed by Nitrofurantoin (91.505%). The least effective drug against Gram negative bacteria was found to be Cefexime (64.16%) and Amoxicillin (58.49%). And for Gram positive isolates Ceftazidime (100%) and Amikacin (100%) were the choice of drug showing high susceptibility. The least effective drug used against Gram positive isolates tends to be Cloxacillin (15.38%). Out of 106 isolates 63 were found to be multidrug resistant among which 31 of them were ESBL producers confirmed by cephalosporin/clavulanate double disc synergy method. The urinary tract infection was most common in the age group 20-30 (20.75%) followed by age group 30-40 (16.98%). Conclusion: UTI was least common among age groups 0-10 and 80-90 years. Bacterial isolates causing UTI had variation in the antibiotic susceptibility pattern with resistance against commonly used antibiotics which is a serious concern.

In Vitro Inhibitory Potential of Lawsonia inermis Extracts against Multidrug Resistant Clinically-Relevant Bacteria: a Phytochemical, Quantitative Antimicrobial and Toxicological Assessment

Objective: Majority of the current antibiotics have become less effective due to widespread of multidrug-resistant microorganisms. Medicinal plants are promising candidates that could be used to manage this menace. Therefore, phytochemical, toxicological and antimicrobial potentiality of Lawsonia inermis extracts against MDR clinical bacteria were carried out. &#x0D; Material-Method: Henna leaf and seed were extracted by cold maceration technique using methanol and water and screened phytochemically. Eight MDR isolates, four of which are ESβL-producers were used for this study. In vitro antimicrobial efficacy and quantitative antimicrobial potency of extracts were estimated. MIC and MBC were determined using broth macrodilution technique. Cytotoxicity test was conducted using brine shrimp lethality assay and LC50 was determined. &#x0D; Results: The findings of this study revealed that aqueous leaf extract possesses maximum percentage yield of 25.58%. Tannins and phenolic compounds were detected in all extracts, while steroid was absent. Methanol seed extract showed the highest antimicrobial efficacy against all bacteria with 100 percent activity. The highest and lowest zones of inhibition were recorded at 30.0±0.00 and 10.0±0.00 mm, respectively. The zones of inhibition of extracts differed significantly. All extracts displayed highest activity index against the ESβL-producing Enterobacter aerogenes 196 that was isolated from wound with highest value at 4.28. Pseudomonas aeruginosa U109 showed maximum susceptibility index (93.75%); majority of MIC values recorded were within the range of 1.95-62.5 mg/mL. Cytotoxicity test of methanol and aqueous extracts displayed 10001000, respectively. &#x0D; Conclusion: Findings from this study elucidate the efficacy of Lawsonia inermis as a potential remedy to manage MDR-related infectious bacteria.

1678. Activity of mecillinam against urinary tract clinical isolates from the United States during 2017-2020 including isolates resistant to comparator antibiotics and multi-drug resistant isolates

Abstract Background Mecillinam is a first-in-class amidinopenicillin antibiotic, being the only β-lactam to exert its antibacterial activity through exclusive binding to penicillin-binding protein 2. Pivmecillinam is the oral prodrug of mecillinam recommended as a first-line therapy by the Infectious Disease Society of America guidelines for uncomplicated urinary tract infections (uUTIs) and is approved for the treatment of uUTI in Europe and Canada though not yet in the USA. To support the clinical development of mecillinam and pivmecillinam in the USA for the treatment of cUTI and uUTI, this study investigated the activity of mecillinam compared with other antibiotics against Enterobacterales isolates from patients with UTI in the US during 2017-2020 Methods A total of 3303 Enterobacterales clinical isolates from UTIs from hospitals throughout the USA during 2017-2020 were tested. MIC tests were performed by agar dilution in line with CLSI susceptibility testing standards. Results Overall, susceptibility was highest for fosfomycin (97.1% susceptible) and mecillinam (94.9% susceptible). Against isolates resistant to ceftriaxone, ciprofloxacin, fosfomycin, nitrofurantoin and trimethoprim/sulfamethoxazole susceptibility to mecillinam was high with 89.6 %, 90.9 %, 82.4 %, 86.7 % and 93.6 %, respectively. Only fosfomycin exhibited similar activity to mecillinam. The most active antibiotics against MDR Enterobacterales were fosfomycin (86% susceptible) and mecillinam (79.2 % susceptible). Of the 288 MDR Enterobacterales, the majority were E. coli (n = 128) and K. pneumoniae (n = 103). The most active agents against MDR E. coli and MDR K. pneumoniae were fosfomycin and mecillinam that had susceptibilities of 93.8 % and 92.2 % and 79.6 % and 72.8 %, respectively. Conclusion Mecillinam exhibited promising activity against the majority of Enterobacterales tested. The drug was notably active against isolates resistant to antibiotics in routine clinical use and notably was similarly active against many MDR isolates. The data from our study strongly support the clinical utility of mecillinam and pivmecillinam in the USA for the treatment of cUTI and uUTI. Disclosures Anne Santerre Henriksen, PhD, UTILITY therapeutics Ltd: Advisor/Consultant.

616. Pharmacodynamics (PD) of the lipopeptide QPX9003 Against Acinetobacter baumannii and Pseudomonas aeruginosa in the Neutropenic Mouse Thigh Infection Model

Abstract Background Acinetobacter baumannii and Pseudomonas aeruginosa are important pathogens in hospital-acquired infections and can be resistant to multiple classes of antibiotics. QPX9003 is a novel synthetic lipopeptide that is highly active in vitro against MDR Acinetobacter sp. and P. aeruginosa. The purpose of these studies was to determine the pharmacodynamic parameter that best described the bacterial killing of QPX9003 in vivo. PK-PD of QPX9003 against A. baumannii PK-PD of QPX9003 against P. aeruginosa Methods Seven A. baumannii and six P. aeruginosa isolates were used in these studies. Swiss-Webster mice were rendered neutropenic by 150 mg/kg IP injections of cyclophosphamide 4 and 1 days prior to infection. The bacterial inoculum was prepared from an overnight culture adjusted to 108 CFU/mL then further diluted for inoculation into the thighs. Various dose and dose regimens of QPX9003 were administered by the IP route starting 2 hours post infection and continuing for 24 hours. Untreated control groups were sacrificed at the start of treatment. Treated and untreated control groups were sacrificed 24 hours after the start of treatment, thighs harvested, homogenized and plated to determine colony counts. PK was conducted in neutropenic, infected mice at doses ranging from 3 to 50 mg/kg administered by the IP route. The relationship between PK-PD parameters and the reduction in the log number of CFU per thigh between time zero and 24 h after the start of treatment were analyzed by using the sigmoid maximum reduction (Emax) PD model. Results The activity of QPX9003 was best described by the 24h free AUC/MIC ratio, with stasis and 1 log kill targets of 47.1 and 67.1 for A. baumannii and 96.1 and 117 for P. aeruginosa, respectively. Conclusion The PD of QPX9003 was best described by the 24h free AUC/MIC ratio. Based on the QPX9003 MIC90 for P. aeruginosa (0.25 mg/L) and A. baumannii (1 mg/L), 24h free AUCs between 30 and 67 mg*h/L will produce 1-log of bacterial killing for ∼90% of P. aeruginosa and A. baumannii strains, respectively. QPX9003 is a potentially useful treatment for infections due to P. aeruginosa and A. baumannii, including MDR isolates, that is advancing in clinical studies. Disclosures Ziad Tarazi, Qpex Biopharma: Employee Niki Roos, n/a, Qpex Biopharma: Employee Ted Page, n/a, Qpex Biopharma: Employee David Griffith, n/a, Qpex Biopharma: Employee.