Pan-drug Resistance Research Articles

Expert Opinion on the Prescription Practice of Carbapenem Antibiotics for the Treatment of Drug-Resistant Bacterial Infections in Indian Settings

Objective: The current survey-based study aims to gather expert opinions on the prescription practice of carbapenem antibiotics for the treatment of drug-resistant bacterial infections in Indian clinical settings.
 Methods: The cross-sectional, multiple-response questionnaire-based survey involving professionals with expertise in treating pathogenic illnesses was carried out between 2022 and 2023. The questionnaire comprised 25 questions, the majority of which dealt with the use of antibiotics as monotherapy and in various combinations to treat pathogenic illnesses.
 Results: Out of 302 survey participants, 83% of them preferred meropenem medications more frequently than imipenem, biapenem, doripenem, and other antibiotics to treat pathogenic infections. For the management of multidrug-resistant (MDR), extensively drug-resistant (XDR), or pan-drug-resistant (PDR) infections, approximately 54% of the respondents preferred the administration of 1 gm of IV meropenem in 2 to 3 divided doses. About 61% of the responders suggested using IV meropenem together with colistin to treat infections. The combination of IV meropenem with β-lactams, aminoglycosides, tigecycline, colistin, and others was preferred by 91% of the respondents for the treatment of MDR, XDR, and PDR infections. For treating difficult intra-abdominal infections, meningitis, and urinary tract infections (UTIs), almost 43% of the respondents preferred meropenem in combination with sulbactam.
 Conclusion: The survey findings emphasize the prominent role of meropenem, either as a standalone treatment or in combination with other antibiotics in the management of challenging infections caused by MDR, XDR, or PDR pathogens. In specific scenarios involving difficult intra-abdominal infections, meningitis, and UTIs, clinicians recommended meropenem in combination with sulbactam antibiotics.

Phenotypic and Genotypic Characterization of Pan-Drug-Resistant Klebsiella pneumoniae Isolated in Qatar.

In secondary healthcare, carbapenem-resistant Enterobacterales (CREs), such as those observed in Klebsiella pneumoniae, are a global public health priority with significant clinical outcomes. In this study, we described the clinical, phenotypic, and genotypic characteristics of three pan-drug-resistant (PDR) isolates that demonstrated extended resistance to conventional and novel antimicrobials. All patients had risk factors for the acquisition of multidrug-resistant organisms, while microbiological susceptibility testing showed resistance to all conventional antimicrobials. Advanced susceptibility testing demonstrated resistance to broad agents, such as ceftazidime-avibactam, ceftolozane-tazobactam, and meropenem-vaborbactam. Nevertheless, all isolates were susceptible to cefiderocol, suggested as one of the novel antimicrobials that demonstrated potent in vitro activity against resistant Gram-negative bacteria, including CREs, pointing toward its potential therapeutic role for PDR pathogens. Expanded genomic studies revealed multiple antimicrobial-resistant genes (ARGs), including blaNMD-5 and blaOXA derivative types, as well as a mutated outer membrane porin protein (OmpK37).

Emergence of pandrug-resistant carbapenemase-producing Enterobacterales in dogs and cats: a cross-sectional study in Egypt.

One of the most important emerging health problems is the increasing role of animals in the rapid global rise in resistance to last-resort antibiotics, such as carbapenems. However, there is limited information on the role of pet animals in harboring and spreading pandrug-resistant (PDR) carbapenemase-producing Enterobacterales (CPE), especially in Egypt. This cross-sectional study was conducted to screen for CPE in healthy and diseased pets using phenotypic and molecular methods and the NG-Test CARBA 5 immunochromatographic assay. Rectal swabs were collected from 62 dogs and 48 cats, incubated overnight in tryptic soy broth containing 10 μg of meropenem disc and subsequently cultured on MacConkey agar supplemented with meropenem (1 mg/L). Sixty-six isolates (60.6%), including 56 Klebsiella pneumoniae, seven Escherichia coli, and three K. oxytoca isolates, were confirmed to be carbapenem-resistant Enterobacterales (CRE) by the disc diffusion method, broth microdilution test, CNPt-direct, and PCR assay targeting carbapenemase genes. Forty-three (65.2%) dogs and 23 (34.8%) cats carried CPE. Of these, 35 (70.0%) were healthy (including 27 dogs and 8 cats) and 31 (52.5%) were diseased (including 16 dogs and 15 cats). bla OXA-181 was the most common gene detected (42/66, 63.6%), followed by bla IMP (40/66, 60.6%), bla OXA-48-like (29/66, 43.9%), bla KPC and bla VIM (20/66, 30.3% each), and bla NDM (17/66, 25.8%). The identified genotypes were bla KPC-2, bla IMP-1, bla VIM-1, bla NDM-1, and bla NDM-5. The CARBA 5 assay showed higher sensitivity and specificity for the detection of NDM, OXA and KPC than that for VIM and IMP genes. Antimicrobial resistance profiles of CRE isolates revealed 20 PDR, 30 extensively drug-resistant (XDR), and 16 multidrug-resistant (MDR) phenotypes. This study provides evidence of colonization with PDR CPE in dogs and cats. To manage the infection or colonization of pets in veterinary clinical settings, extended surveillance systems should be considered, and the use of critical antibiotics should be strictly controlled.

Cytotoxic rhamnolipid micelles drive acute virulence in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic human pathogen that has developed multi- or even pan-drug resistance toward most frontline and last resort antibiotics, leading to increasing frequency of infections and deaths among hospitalized patients, especially those with compromised immune systems. Further complicating treatment, P. aeruginosa produces numerous virulence factors that contribute to host tissue damage and immune evasion, promoting bacterial colonization and pathogenesis. In this study, we demonstrate the importance of rhamnolipid production in host-pathogen interactions. Secreted rhamnolipids form micelles that exhibited highly acute toxicity toward murine macrophages, rupturing the plasma membrane and causing organellar membrane damage within minutes of exposure. While rhamnolipid micelles (RMs) were particularly toxic to macrophages, they also caused membrane damage in human lung epithelial cells, red blood cells, Gram-positive bacteria, and even noncellular models like giant plasma membrane vesicles. Most importantly, rhamnolipid production strongly correlated with P. aeruginosa virulence against murine macrophages in various panels of clinical isolates. Altogether, our findings suggest that rhamnolipid micelles are highly cytotoxic virulence factors that drive acute cellular damage and immune evasion during P. aeruginosa infections.

Molecular Characteristics of Colistin Resistance in Acinetobacter baumannii and the Activity of Antimicrobial Combination Therapy in a Tertiary Care Medical Center in Lebanon.

(1) Background: Infections with pan-drug-resistant (PDR) bacteria, such as A. baumannii, are becoming increasingly common, especially in healthcare facilities. In this study, we selected 15 colistin-resistant clinical A. baumannii isolates from a hospital in Beirut, Lebanon, to test combination therapies and determine their sequence types (STs) and the mechanism of colistin resistance using whole-genome sequencing (WGS). (2) Methods: Antimicrobial susceptibility testing via broth microdilution against 12 antimicrobials from different classes and growth rate assays were performed. A checkerboard assay was conducted on PDR isolates using six different antimicrobials, each in combination with colistin. Genomic DNA was extracted from all isolates and subjected to WGS. (3) Results: All isolates were resistant to all tested antimicrobials with the one exception that was susceptible to gentamicin. Combining colistin with either meropenem, ceftolozane-tazobactam, or teicoplanin showed synergistic activity. Sequencing data revealed that 67% of the isolates belonged to Pasteur ST2 and 33% to ST187. Furthermore, these isolates harbored a number of resistance genes, including blaOXA-23. Mutations in the pmrC gene were behind colistin resistance. (4) Conclusions: With the rise in antimicrobial resistance and the absence of novel antimicrobial production, alternative treatments must be found. The combination therapy results from this study suggest treatment options for PDR ST2 A. baumannii-infected patients.

A comprehensive investigation of the medicinal efficacy of antimicrobial fusion peptides expressed in probiotic bacteria for the treatment of pan drug-resistant (PDR) infections.

The present work aimed to examine the intracellular antibacterial efficacy of Recombinant Lactobacillus acidophilus/antimicrobial peptides (AMPs) Melittin and Alyteserin-1a, specifically targeting Gram-negative bacteria. The first assessment was to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of Recombinant L. acidophilus/AMPs versus Gram-negative and Gram-positive bacteria. In addition, the researchers examined the in vitro viability and safety of AMPs generated by L. acidophilus. The experiments included exposing the AMPs to elevated temperatures, proteases, cationic salts at physiological levels, and specific pH settings. The safety aspect was evaluated using hemolytic analysis utilizing sheep erythrocytes; cytotoxicity assays employing cell lines, and experiments on beneficial gut lactobacilli. An experiment was done using a time-kill method to assess the intracellular antibacterial efficacy of Recombinant L. acidophilus/AMPs compared to pathogenic varieties in HEp-2 cells. Previous investigations have shown that the MBC levels of recombinant L. acidophilus/AMPs were consistently two to four times higher than the equivalent MIC values when evaluated versus Gram-negative bacteria. Furthermore, the stability of the Recombinant L. acidophilus/AMPs showed variability when exposed to elevated temperatures (70 and 90℃), treated with protease enzymes (proteinase K, lysozyme), exposed to higher concentrations of physiological salts (150mM NaCl and 2mM MgCl2), and varying pH levels (ranging from 4.0 to 9.0). The recombinant L. acidophilus/AMPs are non-hemolytic towards sheep erythrocytes, exhibit little cytotoxicity in RAW 264.7 and HEp-2 cells, and are considered safe when compared to beneficial gut lactobacilli. The research examined the intracellular bacteriostatic effects of recombinant L. acidophilus/AMPs on Gram-negative bacteria inside HEp-2 cells. Nevertheless, no notable bactericidal impact was seen on Gram-positive bacteria (P > 0.05). The research shows that recombinant L. acidophilus/AMPs, namely (L. acidophilus/melittin/Alyteserin-1a) as the focus of the investigation, effectively eliminate Gram-negative bacteria. Therefore, more investigation is necessary to elaborate on these discoveries.

Next generation antibiotic combinations to combat pan-drug resistant Klebsiella pneumoniae

Antimicrobial resistance has emerged as one of the leading public health threats of the twenty-first century. Gram-negative pathogens have been a major contributor to the declining efficacy of antibiotics through both acquired resistance and tolerance. In this study, a pan-drug resistant (PDR), NDM-1 and CTX-M-15 co-producing isolate of K. pneumoniae, CDC Nevada, (Kp Nevada) was exposed to the clinical combination of aztreonam + ceftazidime/avibactam (ATM/CAZ/AVI) to overcome metallo-β-lactamases. Unexpectedly, the β-lactam combination resulted in long filamentous cell formation induced by PBP3 inhibition over 168 h in the hollow fiber infection model experiments with eventual reversion of the total population upon drug removal. However, the addition of imipenem to the two drug β-lactam combination was highly synergistic with suppression of all drug resistant subpopulations over 5 days. Scanning electron microscopy and fluorescence microscopy for all imipenem combinations in time kill studies suggested a role for imipenem in suppression of long filamentous persisters, via the formation of metabolically active spheroplasts. To complement the imaging studies, salient transcriptomic changes were quantified using RT-PCR and novel cassette assay evaluated β-lactam permeability. This showed significant upregulation of both spheroplast protein Y (SPY), a periplasmic chaperone protein that has been shown to be related to spheroplast formation, and penicillin binding proteins (PBP1, PBP2, PBP3) for all combinations involving imipenem. However, with aztreonam alone, pbp1, pbp3 and spy remained unchanged while pbp2 levels were downregulated by > 25%. Imipenem displayed 207-fold higher permeability as compared with aztreonam (mean permeability coefficient of 17,200 nm/s). Although the clinical combination of aztreonam/avibactam and ceftazidime has been proposed as an important treatment of MBL Gram-negatives, we report the first occurrence of long filamentous persister formation. To our knowledge, this is the first study that defines novel β-lactam combinations involving imipenem via maximal suppression of filamentous persisters to combat PDR CDC Nevada K. pneumoniae.

Phage therapy as a glimmer of hope in the fight against the recurrence or emergence of surgical site bacterial infections.

Over the last decade, surgery rates have risen alarmingly, and surgical-site infections are expanding these concerns. In spite of advances in infection control practices, surgical infections continue to be a significant cause of death, prolonged hospitalization, and morbidity. As well as the presence of bacterial infections and their antibiotic resistance, biofilm formation is one of the challenges in the treatment of surgical wounds. This review article was based on published studies on inpatients and laboratory animals receiving phage therapy for surgical wounds, phage therapy for tissue and bone infections treated with surgery to prevent recurrence, antibiotic-resistant wound infections treated with phage therapy, and biofilm-involved surgical wounds treated with phage therapy which were searched without date restrictions. It has been shown in this review article that phage therapy can be used to treat surgical-site infections in patients and animals, eliminate biofilms at the surgical site, prevent infection recurrence in wounds that have been operated on, and eradicate antibiotic-resistant infections in surgical wounds, including multi-drug resistance (MDR), extensively drug resistance (XDR), and pan-drug resistance (PDR). A cocktail of phages and antibiotics can also reduce surgical-site infections more effectively than phages alone. In light of these encouraging results, clinical trials and research with phages will continue in the near future to treat surgical-site infections, biofilm removal, and antibiotic-resistant wounds, all of which could be used to prescribe phages as an alternative to antibiotics.

Emergence of High-Level Antibiotic Resistance in Klebsiella pneumoniae: A Narrative Review

In an era marked by remarkable advancements in medicine, the persistent emergence of high-level antibiotic resistance in Klebsiella pneumoniae poses a critical threat to public health globally. As the worldwide spread of extensively drug-resistant (XDR) and pan-drug-resistant (PDR) K. pneumoniae strains continues to grow, a significant shift in how we approach treatment is on the horizon. The complex interaction of genetic factors, which encompasses a wide range of beta-lactamases, aminoglycoside-modifying enzymes, and chromosomal mutations, creates a dynamic resistance mechanism that counters the effects of antibiotics. These intricate adaptations, arising from both gene transfers facilitated by plasmids and changes in the genome itself, present a challenging obstacle to our efforts in managing antimicrobial effectiveness. Klebsiella infections come back stronger armed with molecular tactics that challenge healthcare systems, prolong hospital stays, and increase mortality. Beyond healthcare settings, the economic and social dimensions grow as resources are redirected, intensifying the impact on vulnerable groups. This review delves into the intricate mechanisms behind the high-level antibiotic resistance in K. pneumoniae, examining its epidemiological, molecular, and clinical facets. Highlighting the necessity for coordinated research, medical protocols, and policies, the review underscores the importance of judicious antibiotic utilization, drug innovation, and rigorous infection management.

Meropenem plus Ertapenem and Ceftazidime-Avibactam plus Aztreonam for the Treatment of Ventilator Associated Pneumonia Caused by Pan-Drug Resistant Klebsiella pneumonia.

Gram-negative bacteria (GNB) account for about 70% of infections in the intensive care unit (ICU) setting and are associated with significant morbidity and mortality. In recent years, pan-drug resistant (PDR) strains, strains that are not susceptible to any antibiotic, have been emerged and new treatment strategies are required. Fifty eligible patients were recruited in the three groups. A statistically significant reduction in the Sequential Organ Failure Assessment (SOFA) score was observed in the control group on day 4 in comparison to day 0 of VAP (p = 0.005). The Clinical Pulmonary Infection Score (CPIS) was also reduced on day 4 (p = 0.0016) and day 7 in comparison to day 0 (p = 0.001). Patients that received combination therapy, CAZ-AVI + ATM and DCT, presented with a lower SOFA score and CPIS on day 7 in comparison to day 0 (p = 0.0288 and p = 0.037, respectively). No differences in the ΔSOFA score and ΔCPIS were found between the groups. The control group presented with a significantly lower ICU stay and duration of mechanical ventilation (p = 0.03 and p = 0.02, respectively). There was no difference in mortality. This is a retrospective analysis. This study was conducted in a mixed ICU in the University Hospital of Larissa, Thessaly, Greece during a three-year period (2020-2022). Patients suffering from ventilator associated pneumonia (VAP) due to carbapenem-resistant K. pneumonia (CR-KP) were divided in three different groups: the first one was treated using ceftazidime-avibactam plus aztreonam (CAZ-AVI + ATM group), the second was treated using double carbapenems (DCT group), and the last one (control group) received appropriate therapy since the strain was susceptible in vitro to at least to one antibiotic. Treatment with CAZ-AVI +ATM or DCT may offer a clinical benefit in patients suffering with infections due to PDR K. pneumoniae. Larger studies are required to confirm our findings.

MICROBIAL LOAD AND ANTIBIOTICS RESISTANCE PATTERNS OF BACTERIAL ISOLATES FROM NIGERIA NAIRA NOTES

In our society today, exchange of goods and services usually involves exchange of currency notes by hand. This can serve as a means of transferring microorganisms and antibiotics resistant bacterial isolates. This study was aimed at assessing the contamination of Naira notes in Ungwan Rimi area of Kaduna State and to determine the antibiotics resistance patterns of the isolated bacteria. A total of 160 samples of naira notes, were collected randomly from staff and students, marketers, hospital and pharmacies, food vendors and transporters at strategic spots in Ungwan Rimi. Bacteria isolation and identification of samples were carried out using spread plate technique on medium and standard biochemical characterization, respectively. Antibiotics susceptibility testing were carried out using agar disc diffusion method. Isolates that were non-susceptible to at least one antimicrobial agent in three or more antimicrobial classes were classified as multidrug resistant (MDR), those that remain susceptible to agent(s) in only one or two classes as extended drug resistant (XDR) while pandrug resistant (PDR) isolates were classified as those isolates that were resistant to all antibiotics used. Highest bacterial load was observed with N100 notes with marketers (5.33 x 105) while the least bacterial load was 1.2 x 104 in N50 notes from hospitals and pharmacies. The following bacteria were isolated Staphylococcus aureus 70 (26.8%), Bacillus spp 60 (23.0%), Clostridium spp 31 (11.9%), Escherichia coli 30 (11.5%), Streptococcus species 25 (9.6%), Klebsiella spp 24 (9.2%), Salmonella 15 (5.8%), Shigella species 4 (1.5%) and Pseudomonas species 2 (0.8%)...

Evaluation of role of Tigecycline among clinically significant multidrug resistant pathogens from a tertiary care hospital

Background Tigecycline, a glycylcycline antibiotic is a promising option for the treatment of single or multidrug resistant pathogens. The aim of the study was to evaluate the in-vitro Tigecycline susceptibility of various pathogens from clinical samples received at the tertiary care hospitals in South India. Methods The analysis of specimens from patients admitted were carried out in this prospective cross sectional study. The identification and antimicrobial susceptibility testing was performed by semi-automated Vitek 2 systems and Kirby Bauer method. Pattern of data analysis was done by descriptive statistics. Results Among 2574 isolates, 812 isolates were gram positive pathogens and 1762 isolates were gram negative pathogens. Resistance to Tigecycline was more common among gram negative pathogens (18.62%) in comparison to the gram positive pathogens (0.49%). Among 740 Extended Spectrum Beta Lactamases (ESBL) producers such as Klebsiella species & E coli, 629 isolates were susceptible, and 93 isolates were resistant to the tigecycline. All the methicillin resistant Staphylococcus aureus (MRSA) isolates were susceptible to tigecycline. Conclusion Multidrug resistant (MDR) pathogens like Acinetobacter species, and Klebsiella species were found to be highly effective in vitro to tigecycline for elimination of infections caused by both gram positive and gram negative pathogens. The use of combination therapy becomes crucial to prevent the development of Pan Drug resistance.

P08 An usual case of pandrug-resistant Serratia marcescens in a 63 year-old-male: a case report

Abstract Objectives To highlight the growing burden of antimicrobial resistance in developing countries where there are no officially agreed local antibiotics guidelines. We are characterizing a case of growth of pandrug-resistant (PDR) Serratia marcescens in the urine culture of a 63-year-old-male. Patient and methods Our patient, who had a history of recurrent renal stones, underwent multiple urological procedures (including ureteroscopy and DJ stenting) in various regional hospitals in northern Pakistan. It is postulated that he became infected during these repeated procedures which were done in various hospitals without adequate sterile equipment. He presented with recurrent UTIs to GPs and was subsequently treated with oral fosfomycin for over 1 year in total. His baseline renal function remains unknown due to the absence of online records, but he was noted to have CKD IV. Subsequently, during his later visits to the hospital, he developed sepsis and his urine cultures confirmed carbapenem-resistant Pseudomonas aeruginosa, which was treated with IV antibiotics as per the susceptibility report. He was prescribed IV piperacillin/tazobactam (Tazocin) for a total period of 3 months on various occasions due to repeated presentations with urosepsis. He improved clinically with IV piperacillin/tazobactam. Following this, the patient was lost to follow-up. The patient once again presented acutely unwell with urosepsis. This time urine cultures confirmed the presence of S. marcescens (May 2023). The patient had pandrug resistance to all antibiotics including IV piperacillin/tazobactam. However it was decided to treat him with IV piperacillin/tazobactam due to previous good results. The patient improved clinically but his renal functions declined to CKD stage V. He was once again lost to follow up. He presented again to renal consultants with uraemic symptoms and at the moment he is being counselled for dialysis due to his end-stage renal disease. At present the patient is having asymptomatic bacteriuria; his urine cultures remain positive without any symptoms. A more recent urine culture (September 2023) showed growth of Escherichia coli (ESBL). Due to lack of symptoms, renal consultants agreed to manage the patient without further antibiotics. Results In May 2023, an antibiotic susceptibility profile was done under supervision of microbiologists. Unfortunately for the patient, the urine culture reported growth of S. marcescens and that the patient had pandrug-resistance. The profile included testing for the following antibiotics: amikacin, co-amoxiclav, aztreonam, cefepime, cefoxitin, ceftazidime/avibactam, cefalexin, co-trimoxazole, fosfomycin, meropenem, amoxicillin, ampicillin, cefaclor, cefixime, cefpodoxime, cefuroxime, ciprofloxacin, colistin, imipenem, nitrofurantoin, norfloxacin, polymyxin, piperacillin/tazobactam and sulbactam/cefoperazone. In September 2023, the second urine culture report with the growth of E. coli showed susceptibility to amikacin, cefoperazone/sulbactam, fosfomycin, imipenem and tigecycline. This report also confirmed resistance co-amoxiclav, ampicillin, cefixime, cefotaxime, ceftazidime, ceftriaxone, cefuroxime, cefradine, levofloxacin, nalidixic acid, nitrofurantoin, piperacillin/tazobactam and trimethoprim/sulfamethoxazole. Conclusions Unfortunately, the urine culture and susceptibility report signify the threat this patient faces in case he develops further urine infections. It is imperative to encourage development of local guidelines in developing countries too. We would recommend a multidisciplinary approach in treating difficult and long-term infections with constant support of microbiologists.

P28 Evaluation of one in-house and three commercial phenotypic tests for detection of KPC, NDM and OXA-48-like carbapenemase in Klebsiella pneumoniae: is routine detection possible?

Abstract Background Phenotypic characterization of the OXA-48-like carbapenemase XDR and pandrug-resistant (PDR) Klebsiella pneumoniae in clinical isolates is essential for making informed empirical decisions and critical for strengthening antimicrobial stewardship programmes. This study focused on assessing one in-house and three commercial phenotypic tests for detection of KPC, NDM and OXA-48-like carbapenemases in an attempt to study the feasibility of making detection of OXA-48-like carbapenemases a routine activity of a diagnostic microbiology laboratory. Methods A total of 40 representative non-duplicate XDR and PDR clinical Klebsiella pneumoniae isolates were screened on the basis of susceptibility profile, resistance to piperacillin/tazobactam; susceptible, intermediate, or resistant to third generation cephalosporins; elevated MIC for meropenem. These isolates were then subjected to one in-house test (OXA-48 disc test) and three commercial tests: Xpert® Carba-R (Cepheid), MASTDISCS® Combi Carba Plus (D73C), Immunochromatographic assay (ICT); KPC/IMP/NDM/OXA-48 Combo test (LFIA), Medomics. Results Xpert ®Carba-R identified OXA-48 in 62.5% (n=25) isolates, NDM in 17.5% (n=7), KPC+NDM co-carriage in 15% (n=6) and KPC in one isolate. Both Xpert® Carba-R and ICT were considered as possible gold standards and the outcomes of the other tests were compared with these two. With Xpert® Carba-R as the gold standard, high sensitivity of 96.15% was detected in OXA-48-like isolates, followed by NDM (92.85%) and NDM+KPC (85.7%). The lowest sensitivity was seen with the OXA-48 disc test (48% in OXA-48, 53.8% in NDM) followed by D73C test (64% in OXA-48, 38.46% in NDM). These two tests could not detect co- carriage of NDM and KPC in six isolates, although in two cases one of the two was detected. Specificity was high among all the tests (96.29%–93.33%). With ICT as the gold standard, 100% sensitivity was reported for NDM and KPC isolates, followed by OXA-48 and KPC+NDM (96% and 83.33%, respectively). The specificity was higher with ICT (99%) than Xpert® Carba-R (96.29%) as in detection of co- carriage of NDM and KPC. In terms of cost ICT ($16 per test) was much cheaper than Xpert® Carba-R($52 per test). Conclusions We recommend immunochromatographic assay (ICT) as an excellent tool for detection of carbapenemases (KPC/IMP/NDM/OXA-48). OXA-48 disc test and MASTDISCS® Combi Carba plus (D73C) cannot be considered as good standard since they could not detect co-carriage of NDM and KPC and had lower sensitivity.

Molecular mechanisms of colistin resistance mediated by pmrCAB genes in Acinetobacter baumannii isolated from patients hospitalized in Isfahan medical centers

Abstract Introduction In recent years, colistin-resistant Acinetobacter baumannii (A. baumannii) has been found all over the world. In this current study, the main purpose was to examine the occurrence of extensively drug resistant (XDR), resistance to colistin and characterization and mutations in pmrCAB genes among A. baumannii obtained from inpatients. Materials and Methods A total of 108 clinical isolates of A. baumannii were collected from several hospitals located in Isfahan, Iran. The Kirby-Bauer assay was performed to assess the antimicrobial resistance. The Phoenix automated system was utilized to determine the minimum inhibitory concentration (MIC) of colistin for each of the bacterial isolates. Polymerase chain reaction was used to screen for pmrCAB genes that mediate colistin resistance, and sequencing was used to determine the amplicon’s nucleotide sequence. Results The results revealed that all A. baumannii isolates (100%) were resistant to piperacillin tazobactam, meropenem and ciprofloxacin. All isolates were classified as XDR, with seven isolates being pan-drug resistant (PDR). Colistin resistance (CoR) was found in 6.48% (7/108) of studied isolates, all of which were positive for pmrCAB genes. The sequencing results showed a substitution in pmrB and two isolates showed a substitution in pmrC. Conclusions In conclusion, this study is the initial report of the existence and mutations of pmrB and C genes in the clinical isolate of A. baumannii in our region. This outcome highlights the necessity to explore additional mutations in the PMR operon of A. baumannii in forthcoming studies. Moreover, our results highlight the high occurrence of XDR-A. baumannii strain in Isfahan, Iran.

Multi-drug resistant (MDR) Gram-negative pathogenic bacteria isolated from poultry in the Noakhali region of Bangladesh.

Rapidly increasing antibiotic-resistant bacterial strains in Bangladesh's food and farm animals stem from the excessive and inappropriate use of antibiotics. To assess the prevalence of multi-drug resistant (MDR) Gram-negative bacteria in poultry chicks, we sought to isolate and identify strains carrying antimicrobial resistance genes. Isolation and identification involved biochemical tests, 16S rRNA sequencing, and PCR screening of species-specific genes. MDR patterns were evaluated using CLSI guidelines with seventeen antibiotics across twelve classes. Targeted gene sequences were amplified for the detection of Extended-spectrum β-Lactamase (ESBL), carbapenem, tetracycline, sulfonamide, and colistin resistance genes. Common isolates, such as Escherichia coli, Klebsiella pneumoniae, Proteus penneri, and Enterobacter hormaechei, exhibited average Multiple Antimicrobial Resistance (MAR) indices of 0.66, 0.76, 0.8, 0.84, and 0.81, 0.76, 0.84, 0.41 for broiler and layer chicken, respectively. Providencia stuartii and Salmonella enterica, exclusive to broiler samples, had MAR indices of 0.82 and 0.84, respectively. Additional isolates Morganella morganii, Aeromonas spp., and Wohlfahrtiimonas chitiniclastica were found in layers (Average MAR indices: 0.73, 0.71, and 0.91). Notably, M. morganii, E. hormaechei and W. chitiniclastica were identified for the first time in Bangladeshi poultry chicken, although their evolution is yet to be understood. In this study, Pan-drug resistance was observed in one P. stuartii (broiler) and one Aeromonas spp. (layer) with a MAR index 1, while all isolates exhibited MAR indices >0.2, indicating MDR. Antimicrobial resistance (AMR) gene screening identified blaTEM, blaSHV, tetA, and sul1 in a majority of the MDR strains. Interestingly, E. coli (lactose positive and negative) and E. hormaechei were exclusively found to possess the tetB gene. In addition, E. coli (lactose negative), Klebsiella pneumoniae, Enterobacter hormaechei, M. morganii, and P. stuartii were observed to carry the colistin-resistant mcr-1 gene, whereas sul2 was detected in E. coli (lactose positive and negative), E. hormaechei, P. stuartii, and P. penneri. These findings emphasize the health risk of our consumers of both broiler and layer chickens as they have turned into a potent reservoir of various AMR gene carrying MDR and Pan-drug resistant bacteria.

Identification and Functional Annotation of Hypothetical Proteins of Pan-Drug-Resistant Providencia rettgeri Strain MRSN845308 Toward Designing Antimicrobial Drug Targets.

Providencia rettgeri has increasingly been responsible for several infections, including urinary tract, post-burn wounds, neonatal sepsis, and others. The emergence of drug-resistant isolates of P rettgeri, accompanied by intrinsic and acquired antibiotic resistance, has exacerbated the challenge of treating such infections, necessitating the development of novel therapeutics. Hypothetical proteins (HPs) form a major portion of cellular proteins and can be targeted by these novel therapeutics. In this study, 410 HPs from a pan-drug-resistant (PDR) P rettgeri strain (MRSN845308) were functionally annotated and characterized by physicochemical properties, localization, virulence, essentiality, druggability, and functionality. Among 410 HPs, the VirulentPred 2.0 tool and VICMpred combinedly predicted 33 HPs as virulent, whereas 48 HPs were highly interacting proteins based on the STRING v12 database. BlastKOALA and eggNOG-mapper v2.1.12 predicted 13 HPs involved in several metabolic pathways like Riboflavin metabolism and Lipopolysaccharide biosynthesis. Overall, 83 HPs were selected as primary drug targets; however, only 80 remained after nonhomology searching and essentiality analysis. In addition, all were detected as novel drug targets according to DrugBank 5.1.12. Considering the potential of membrane and extracellular proteins, 29 HPs (extracellular, outer, and inner membrane) were selected based on the combined prediction from PSORTb v3.0.3, CELLO v.2.5, BUSCA, SOSUIGramN, and PSLpred. According to the prevalence of those HPs in different strains of P rettgeri sequences in National Center for Biotechnology Information Identical Protein Groups (NCBI-IPG), 5 HPs were selected as final drug targets. In addition, 5 other HPs annotated as transporter proteins were also added to the list. As no crystal structures of our targets are present, 3-dimensional structures of selected HPs were predicted by the AlphaFold Server powered by AlphaFold 3. Our findings might facilitate a better understanding of the mechanism of virulence and pathogenesis, and up-to-date annotations can make uncharacterized HPs easy to identify as targets for novel therapeutics.

Fosfomycin in the treatment of New Delhi Metallo-β-Lactamase-5 (blaNDM-5)-producing Escherichia coli infection

The worldwide spread of Gram-negative bacteria showing pan-drug resistance raises significant concerns. The World Health Organization (WHO) designated carbapenem-resistant Enterobacteriaceae (CRE) as a critical priority on the global pathogen list in 2017. This issue has captured increased attention to research in the field of antimicrobial resistance, specifically concentrating on the discovery of novel antibiotics. The primary mechanism of carbapenem resistance revolves around the production of acquired carbapenemase, including class A Klebsiella pneumoniae carbapenem-resistant (KPC), class B New Delhi Metallo-β-Lactamase (NDM), or class D, such as OXA-48 β-lactamases. These carbapenemases are especially prevalent in Enterobacterales. Given that these various resistance mechanisms are frequently widespread, the available therapeutic options can be severely restricted. The high susceptibility rates to fosfomycin in strains with acquired resistance to carbapenems indicate the potential effectiveness of fosfomycin against such strains. The present study aimed to determine the in-vitro activity of aztreonam, aztreonam-avibactam, and fosfomycin against 64 E. coli isolates exhibiting diverse blaNDM genes. From the data obtained, it can be inferred that resistance to aztreonam is 70% and drops with the combined use of avibactam. However, this combination cannot be used in the treatment of patients with diseases triggered by E. coli that produce blaNDM-5 . Meanwhile, all strains tested were susceptible to fosfomycin. Therefore, a remedy for elevated minimal inhibitor concentration of aztreonam, aztreonam-avibactam among blaNDM-5 -producing E. coli may be fosfomycin.

Antibiogram of Pseudomonas aeruginosa Clinical Isolates Tested for Pan, Extensive and Multidrug Resistances

Background: Pseudomonas aeruginosa usually cause nosocomial infections with concurrent morbidity and mortality and is generally resistant to many antibiotics.
 Aim: This study was aimed to determine the proportion of pan-drug-resistant (PDR), extensively drug-resistant (XDR), and multidrug-resistant (MDR) P. aeruginosa strains recovered from human samples.
 Methodology: The retrospective study was conducted in the University of Nigeria Teaching Hospital Enugu in 2023. Clinical samples obtained from patients between October 2022 and April 2023 were analysed. A total of 100 Pseudomonas aeruginosa isolates recovered from 780 clinical samples were used. Standard microbiological techniques were used to identify and categorize the isolates. Antibiotic susceptibility pattern was determined using the Kirby Bauer disc diffusion method.
 Results: Isolates recovered from wound was 38%, voided urine, catheter tip urine, ear swab, and high vaginal swab samples recorded 29%, 11%, 10% and 4%, respectively. Ceftazidime recorded the highest level of resistance (70.0%) and the least was Colistin (20%). Resistance patterns showed that 32(32.0%) bacterial strains were MDR, 68(68.0%) were XDR and no PDR was recorded.
 Conclusions: For the best selection of empirical therapy, P. aeruginosa susceptibility monitoring is essential due to the high prevalence of antibiotic resistance. The resistance pattern raises the possibility of misuse of broad-spectrum antibiotics. Treatment for bacterial infections should be directed by the results of antimicrobial susceptibility tests.

Virulence of Pseudomonas aeruginosa in Cystic Fibrosis: Relationships between Normoxia and Anoxia Lifestyle.

The airways of cystic fibrosis (CF) patients are colonized by many pathogens and the most common is Pseudomonas aeruginosa, an environmental pathogen that is able to infect immunocompromised patients thanks to its ability to develop resistance to conventional antibiotics. Over 12% of all patients colonized by P. aeruginosa harbour multi-drug resistant species. During airway infection in CF, P. aeruginosa adopts various mechanisms to survive in a hostile ecological niche characterized by low oxygen concentration, nutrient limitation and high osmotic pressure. To this end, P. aeruginosa uses a variety of virulence factors including pigment production, biofilm formation, motility and the secretion of toxins and proteases. This study represents the first report that systematically analyzes the differences in virulence features, in normoxia and anoxia, of clinical P. aeruginosa isolated from CF patients, characterized by multi- or pan-drug antibiotic resistance compared to antibiotic sensitive strains. The virulence features, such as biofilm formation, protease secretion and motility, are highly diversified in anaerobiosis, which reflects the condition of chronic CF infection. These findings may contribute to the understanding of the real-world lifestyle of pathogens isolated during disease progression in each particular patient and to assist in the design of therapeutic protocols for personalized medicine.