Antibiotic Resistance and Virulence Gene Patterns Associated with Multi Drug Resistant Avian Pathogenic Escherichia coli (APEC) Isolated from Broiler Chickens in India.

In the present study, a total of 102 samples were collected from chickens of different flocks, died due to suspected colibacillosis. Bacteriological and PCR methods were applied to detect avian pathogenic Escherichia coli (APEC). Phenotypic antimicrobial resistance (AMR) was determined by disk diffusion method. Extended spectrum beta lactamases (ESBL) detection was carried out via PCR by targeting blaTEM, blaSHV, blaOXA, and blaCTX-M groups 1, 2, and 9. Genes of eight virulence factors and class I integrons were also detected by PCR using gene specific primers. Culture, microscopic, biochemical tests and PCR recognised 69/102 (67.64%) samples as E. coli. Phenotypic AST revealed higher resistance against fluoroquinolone antibiotics, i.e., enrofloxacin (72.46%), levofloxacin (69.56%) & ciprofloxacin (66.66%), followed by amoxyclav (63.77%) and tetracycline (59.42%). Six isolates were found as pan-drug-resistant E. coli. A total of 48 (69.56%) and 7 (10.14%) isolates were positive for the presence of blaTEM and blaCTX-M-G9 genes, respectively, whereas 2 (2.90%) isolates each were found positive for blaSHV, blaOXA, and blaCTX-M-G1 genes. Among APEC associated virulence genes, iss (79.71%) was the most predominant, followed by tsh (50.72%), ast (30.43%), cvaf (26.08%), pap (23.18%), vat (8.69%) and stx-1 (1.44%). Thirty-two isolates harboured class I integrons, either with or without ESBL genes. Conclusively, the isolates under study showed pan and multiple-drug resistance, specifically against fluoroquinolone drugs. ESBL production was mediated principally through bla TEM and blaCTX-M-G9. Multiple virulence factors, toxins, and carriage & spread factor render these as zoonotically potential pathogens for humans. Supplementary Information: The online version contains supplementary material available at 10.1007/s12088-023-01132-2.

Characterization of acquired ß-lactamases in Pseudomonas aeruginosa and quantification of their contributions to resistance.

Pseudomonas aeruginosa is a highly problematic opportunistic pathogen that causes a range of different infections. Infections are commonly treated with ß-lactam antibiotics, including cephalosporins, monobactams, penicillins, and carbapenems, with carbapenems regarded as antibiotics of last resort. Isolates of P. aeruginosa can contain horizontally acquired bla genes encoding ß-lactamase enzymes, but the extent to which these contribute to ß-lactam resistance in this species has not been systematically quantified. The overall aim of this research was to address this knowledge gap by quantifying the frequency of ß-lactamase-encoding genes in P. aeruginosa and by determining the effects of ß-lactamases on susceptibility of P. aeruginosa to ß-lactams. Genome analysis showed that ß-lactamase-encoding genes are present in 3% of P. aeruginosa but are enriched in carbapenem-resistant isolates (35%). To determine the substrate antibiotics, 10 ß-lactamases were expressed from an integrative plasmid in the chromosome of P. aeruginosa reference strain PAO1. The ß-lactamases reduced susceptibility to a variety of clinically used antibiotics, including carbapenems (meropenem, imipenem), penicillins (ticarcillin, piperacillin), cephalosporins (ceftazidime, cefepime), and a monobactam (aztreonam). Different enzymes acted on different ß-lactams. ß-lactamases encoded by the genomes of P. aeruginosa clinical isolates had similar effects to the enzymes expressed in strain PAO1. Genome engineering was used to delete ß-lactamase-encoding genes from three carbapenem-resistant clinical isolates and increased susceptibility to substrate ß-lactams. Our findings demonstrate that acquired ß-lactamases play an important role in ß-lactam resistance in P. aeruginosa, identifying substrate antibiotics for a range of enzymes and quantifying their contributions to resistance.IMPORTANCEPseudomonas aeruginosa is an extremely problematic pathogen, with isolates that are resistant to the carbapenem class of ß-lactam antibiotics being in critical need of new therapies. Genes encoding ß-lactamase enzymes that degrade ß-lactam antibiotics can be present in P. aeruginosa, including carbapenem-resistant isolates. Here, we show that ß-lactamase genes are over-represented in carbapenem-resistant isolates, indicating their key role in resistance. We also show that different ß-lactamases alter susceptibility of P. aeruginosa to different ß-lactam antibiotics and quantify the effects of selected enzymes on ß-lactam susceptibility. This research significantly advances the understanding of the contributions of acquired ß-lactamases to antibiotic resistance, including carbapenem resistance, in P. aeruginosa and by implication in other species. It has potential to expedite development of methods that use whole genome sequencing of infecting bacteria to inform antibiotic treatment, allowing more effective use of antibiotics, and facilitate the development of new antibiotics.

Antibiotic Treatment of Infections Caused by AmpC-Producing Enterobacterales.

AmpC enzymes are a class of beta-lactamases produced by Gram-negative bacteria, including several Enterobacterales. When produced in sufficient amounts, AmpCs can hydrolyze third-generation cephalosporins (3GCs) and piperacillin/tazobactam, causing resistance. In Enterobacterales, the AmpC gene can be chromosomal- or plasmid-encoded. Some species, particularly Enterobacter cloacae complex, Klebsiella aerogenes, and Citrobacter freundii, harbor an inducible chromosomal AmpC gene. The expression of this gene can be derepressed during treatment with a beta-lactam, leading to AmpC overproduction and the consequent emergence of resistance to 3GCs and piperacillin/tazobactam during treatment. Because of this phenomenon, the use of carbapenems or cefepime is considered a safer option when treating these pathogens. However, many areas of uncertainty persist, including the risk of derepression related to each beta-lactam; the role of piperacillin/tazobactam compared to cefepime; the best option for severe or difficult-to-treat cases, such as high-inoculum infections (e.g., ventilator-associated pneumonia and undrainable abscesses); the role of de-escalation once clinical stability is obtained; and the best treatment for species with a lower risk of derepression during treatment (e.g., Serratia marcescens and Morganella morganii). The aim of this review is to collate the most relevant information about the microbiological properties of and therapeutic approach to AmpC-producing Enterobacterales in order to inform daily clinical practice.

Genomic Analysis of Enterobacter Species Isolated from Patients in United States Hospitals.

We analyzed the whole genome sequences (WGS) and antibiograms of 35 Enterobacter isolates, including E. hormaechei and E. asburiae, and the recently described E. bugandensis, E. kobei, E. ludwigii, and E. roggenkampii species. Isolates were obtained from human blood and urinary tract infections in patients in the United States. Our goal was to understand the genetic diversity of antimicrobial resistance genes and virulence factors among the various species. Thirty-four of 35 isolates contained an AmpC class blaACT allele; however, the E. roggenkampii isolate contained blaMIR-5. Of the six Enterobacter isolates resistant to ertapenem, imipenem, and meropenem, four harbored a carbapenemase gene, including blaKPC or blaNDM. All four isolates were mCIM-positive. The remaining two isolates had alterations in ompC genes that may have contributed to the resistance phenotype. Interpretations of cefepime test results were variable when disk diffusion and automated broth microdilution results were compared due to the Clinical Laboratory and Standards Institute use of the "susceptible dose-dependent" classification. The diversity of the blaACT alleles paralleled species identifications, as did the presence of various virulence genes. The classification of recently described Enterobacter species is consistent with their resistance gene and virulence gene profiles.

Disentangling abiotic and biotic effects of treated wastewater on stream biofilm resistomes enables the discovery of a new planctomycete beta-lactamase.

BACKGROUND: Environmental reservoirs of antibiotic resistance pose a threat to human and animal health. Aquatic biofilms impacted by wastewater effluent (WW) are known environmental reservoirs for antibiotic resistance; however, the relative importance of biotic factors and abiotic factors from WW on the abundance of antibiotic resistance genes (ARGs) within aquatic biofilms remains unclear. Additionally, experimental evidence is limited within complex aquatic microbial communities as to whether genes bearing low sequence similarity to validated reference ARGs are functional as ARGs. RESULTS: To disentangle the effects of abiotic and biotic factors on ARG abundances, natural biofilms were previously grown in flume systems with different proportions of stream water and either ultrafiltered or non-ultrafiltered WW. In this study, we conducted deep shotgun metagenomic sequencing of 75 biofilm, stream, and WW samples from these flume systems and compared the taxonomic and functional microbiome and resistome composition. Statistical analysis revealed an alignment of the resistome and microbiome composition and a significant association with experimental treatment. Several ARG classes exhibited an increase in normalized metagenomic abundances in biofilms grown with increasing percentages of non-ultrafiltered WW. In contrast, sulfonamide and extended-spectrum beta-lactamase ARGs showed greater abundances in biofilms grown in ultrafiltered WW compared to non-ultrafiltered WW. Overall, our results pointed toward the dominance of biotic factors over abiotic factors in determining ARG abundances in WW-impacted stream biofilms and suggested gene family-specific mechanisms for ARGs that exhibited divergent abundance patterns. To investigate one of these specific ARG families experimentally, we biochemically characterized a new beta-lactamase from the Planctomycetota (Phycisphaeraceae). This beta-lactamase displayed activity in the cleavage of cephalosporin analog despite sharing a low sequence identity with known ARGs. CONCLUSIONS: This discovery of a functional planctomycete beta-lactamase ARG is noteworthy, not only because it was the first beta-lactamase to be biochemically characterized from this phylum, but also because it was not detected by standard homology-based ARG tools. In summary, this study conducted a metagenomic analysis of the relative importance of biotic and abiotic factors in the context of WW discharge and their impact on both known and new ARGs in aquatic biofilms. Video Abstract.

Efficacy of aspergillomarasmine A/meropenem combinations with and without avibactam against bacterial strains producing multiple ß-lactamases.

The effectiveness of ß-lactam antibiotics is increasingly threatened by resistant bacteria that harbor hydrolytic ß-lactamase enzymes. Depending on the class of ß-lactamase present, ß-lactam hydrolysis can occur through one of two general molecular mechanisms. Metallo-ß-lactamases (MBLs) require active site Zn2+ ions, whereas serine-ß-lactamases (SBLs) deploy a catalytic serine residue. The result in both cases is drug inactivation via the opening of the ß-lactam warhead of the antibiotic. MBLs confer resistance to most ß-lactams and are non-susceptible to SBL inhibitors, including recently approved diazabicyclooctanes, such as avibactam; consequently, these enzymes represent a growing threat to public health. Aspergillomarasmine A (AMA), a fungal natural product, can rescue the activity of the ß-lactam antibiotic meropenem against MBL-expressing bacterial strains. However, the effectiveness of this ß-lactam/ß-lactamase inhibitor combination against bacteria producing multiple ß-lactamases remains unknown. We systematically investigated the efficacy of AMA/meropenem combination therapy with and without avibactam against 10 Escherichia coli and 10 Klebsiella pneumoniae laboratory strains tandemly expressing single MBL and SBL enzymes. Cell-based assays demonstrated that laboratory strains producing NDM-1 and KPC-2 carbapenemases were resistant to the AMA/meropenem combination but became drug-susceptible upon adding avibactam. We also probed these combinations against 30 clinical isolates expressing multiple ß-lactamases. E. coli, Enterobacter cloacae, and K. pneumoniae clinical isolates were more susceptible to AMA, avibactam, and meropenem than Pseudomonas aeruginosa and Acinetobacter baumannii isolates. Overall, the results demonstrate that a triple combination of AMA/avibactam/meropenem has potential for empirical treatment of infections caused by multiple ß-lactamase-producing bacteria, especially Enterobacterales.

Resolved genomes of wastewater ESBL-producing Escherichia coli and metagenomic analysis of source wastewater samples.

Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli pose a serious threat to human health because of their resistance to the most commonly prescribed antibiotics: penicillins and cephalosporins. In this study, we provide a genomic and metagenomic context for the determinant beta-lactam resistance genes of ESBL-positive E. coli isolated from various wastewater treatment utilities in Oregon, USA. Class A beta-lactamase genes on chromosomes (blaCTX-M, blaTEM) were clustered with antibiotic resistance genes associated with other classes of antibiotics (sulfonamides and aminoglycosides) along with insertional elements. ESBL genes such as blaCTX-M, blaTEM, and blaSHV were also detected on conjugable plasmids of IncF and IncI incompatibility types. One novel IncF plasmid (pSHV2A_ESBLF) was identified, which carried a multidrug resistance genotype (blaSHV-2A, aadA22, aac3, aph6, tetA, and sul1) in addition to a mer (mercury resistance) operon, colicin, and aerobactin genes. Shotgun metagenomic analysis of the ESBL-producing E. coli-originating wastewater samples showed the presence of class A beta-lactamases; however, the ESBL genes identified in the E. coli genomes were below the detection limits. Other ESBL-associated genes (i.e., blaOXA.11, blaFOX.7, and blaGES.17) were identified in the wastewater samples, and their occurrences were correlated with the core microbial genera (e.g., Paraprevotella). In the E. coli genomes and wastewater samples, tetracycline, aminoglycoside, and beta-lactam resistance determinants frequently co-occurred. The combination of whole-genome and metagenomic analysis provides a holistic description of ESBL-producing organisms and genes in wastewater systems.IMPORTANCEUsing a hybrid sequencing and assembly strategy (short- and long-read sequencing), we identified the distribution of ARGs and virulence factors harbored on plasmids and chromosomes. We further characterized plasmids' incompatibility types and the co-occurrences of ARGs and virulence factors on plasmids and chromosomes. We investigated the transferability of plasmid-mediated beta-lactams via conjugation. Finally, using shotgun metagenomic analysis of the ESBL-producing Escherichia coli-originated wastewater samples, we described the microbial community, the resistome composition, and the potential associations with plasmid-mediated beta-lactam genes and other ARGs.

IncC plasmid genome rearrangements influence the vertical and horizontal transmission tradeoff in Escherichia coli.

It has been shown that an evolutionary tradeoff between vertical (host growth rate) and horizontal (plasmid conjugation) transmissions contributes to global plasmid fitness. As conjugative IncC plasmids are important for the spread of multidrug resistance (MDR), in a broad range of bacterial hosts, we investigated vertical and horizontal transmissions of two multidrug-resistant IncC plasmids according to their backbones and MDR-region rearrangements, upon plasmid entry into a new host. We observed plasmid genome deletions after conjugation in three diverse natural Escherichia coli clinical strains, varying from null to high number depending on the plasmid, all occurring in the MDR region. The plasmid burden on bacterial fitness depended more on the strain background than on the structure of the MDR region, with deletions appearing to have no impact. Besides, we observed an increase in plasmid transfer rate, from ancestral host to new clinical recipient strains, when the IncC plasmid was rearranged. Finally, using a second set of conjugation experiments, we investigated the evolutionary tradeoff of the IncC plasmid during the critical period of plasmid establishment in E. coli K-12, by correlating the transfer rates of deleted or non-deleted IncC plasmids and their costs on the recipient strain. Plasmid deletions strongly improved conjugation efficiency with no negative growth effect. Our findings indicate that the flexibility of the MDR-region of the IncC plasmids can promote their dissemination, and provide diverse opportunities to capture new resistance genes. In a broader view, they suggest that the vertical-horizontal transmission tradeoff can be manipulated by the plasmid to improve its fitness.

Advancements in Immunology and Microbiology Research: A Comprehensive Exploration of Key Areas.

Immunology and microbiology research has witnessed remarkable growth and innovation globally, playing a pivotal role in advancing our understanding of immune mechanisms, disease pathogenesis, and therapeutic interventions. This manuscript presents a comprehensive exploration of the key areas in immunology research, spanning from the utilisation of bacterial proteins as antibody reagents to the intricate realms of clinical immunology and disease management. The utilisation of bacterial immunoglobulin-binding proteins (IBPs), including protein A (SpA), protein G (SpG), and protein L (SpL), has revolutionised serological diagnostics, showing promise in early disease detection and precision medicine. Microbiological studies have shed light on antimicrobial resistance patterns, particularly the emergence of extended-spectrum beta-lactamases (ESBLs), guiding antimicrobial stewardship programmes and informing therapeutic strategies. Clinical immunology research has elucidated the molecular pathways underlying immune-mediated disorders, resulting in tailored management strategies for conditions such as severe combined immunodeficiency (SCID), neuropsychiatric systemic lupus erythematosus (NPSLE), etc. Additionally, significant efforts in vaccine development against tuberculosis and HIV are highlighted, underscoring the ongoing global pursuit of effective preventive measures against these infectious diseases. In summary, immunology and microbiology research have provided significant contributions to global healthcare, fostering collaboration, innovation, and improved patient outcomes.

The burden of antibiotic resistance of the main microorganisms causing infections in humans - review of the literature.

One of the biggest threats to human well-being and public health is antibiotic resistance. If allowed to spread unchecked, it might become a major health risk and trigger another pandemic. This proves the need to develop antibiotic resistance-related global health solutions that take into consideration microdata from various global locations. Establishing positive social norms, guiding individual and group behavioral habits that support global human health, and ultimately raising public awareness of the need for such action could all have a positive impact. Antibiotic resistance is not just a growing clinical concern but also complicates therapy, making adherence to current guidelines for managing antibiotic resistance extremely difficult. Numerous genetic components have been connected to the development of resistance; some of these components have intricate paths of transfer between microorganisms. Beyond this, the subject of antibiotic resistance is becoming increasingly significant in medical microbiology as new mechanisms underpinning its development are identified. In addition to genetic factors, behaviors such as misdiagnosis, exposure to broad-spectrum antibiotics, and delayed diagnosis contribute to the development of resistance. However, advancements in bioinformatics and DNA sequencing technology have completely transformed the diagnostic sector, enabling real-time identification of the components and causes of antibiotic resistance. This information is crucial for developing effective control and prevention strategies to counter the threat.

Bloodstream infections due to multi-drug resistant bacteria in the emergency department: prevalence, risk factors and outcomes-a retrospective observational study.

Multidrug-resistant organisms (MDROs) are prevalent in patients admitted to the Emergency Department (ED) and increase the risk of inappropriate empirical antibiotic therapy. Risk stratification for MDRO infection is essential to early identify patients requiring empirical broad-spectrum antibiotic therapy, but it remains challenging for emergency physicians. This study aimed to evaluate prevalence, risk factors, and outcomes of patients admitted to the ED with a bloodstream infection (BSI) caused by MDROs. A retrospective observational study enrolling all consecutive adult patients admitted with a BSI to the ED of Niguarda Hospital, Italy, from January 2019 to December 2021 was performed. 757 patients were enrolled, 14.1% with septic shock. 156 (20%) patients had a BSI caused by MDRO: extended-spectrum beta-lactamase (ESBL) producing Enterobacterales were the most prevalent followed by methicillin-resistant Staphylococcus aureus (MRSA). Risk factors for BSI due to MDRO and specifically for ESBL were chronic renal failure (OR 2.2; 95%CI 1.4-3.6), nursing home residency (OR 4.4; 95%CI 1.9-10.2) and antibiotic therapy in the last 90-days (OR 2.6; 95%CI 1.7-4), whereas for MRSA were dialysis (OR 12.3; 95%CI 1.8-83), antibiotic therapy and/or hospital admission in the past 90-days (OR 3.6; 95%CI 1.2-10.6) and ureteral stent or nephrostomy (OR 7.8; 95%CI 1.5-40.9). Patients with BSI due to MDRO had a higher rate of inappropriate empirical antibiotic therapy (50%) and longer length of stay, but no higher in-hospital mortality. Among patients admitted to the ED with a BSI, MDROs are frequent and often associated with inappropriate empirical antibiotic therapy. Specific updated risk factors for MDRO may help clinicians to better identify patients requiring a broader antibiotic therapy in the ED, while awaiting microbiological results.

Complete genome sequence of a Pasteurella multocida isolate from a pig in Australia.

Here, we announce the complete genome sequence of a Pasteurella multocida isolate (PM1463) obtained from a diseased pig as a part of routine diagnostic investigations by the field veterinarian in Queensland, Australia. The assembly consists of a 2,321,605-bp chromosome and a 4,769-bp plasmid. The assembly has an average GC content of 40.33%.

Cefiderocol heteroresistance associated with mutations in TonB-dependent receptor genes in Pseudomonas aeruginosa of clinical origin.

The siderophore-cephalosporin cefiderocol (FDC) presents a promising treatment option for carbapenem-resistant (CR) P. aeruginosa (PA). FDC circumvents traditional porin and efflux-mediated resistance by utilizing TonB-dependent receptors (TBDRs) to access the periplasmic space. Emerging FDC resistance has been associated with loss of function mutations within TBDR genes or the regulatory genes controlling TBDR expression. Further, difficulties with antimicrobial susceptibility testing (AST) and unexpected negative clinical treatment outcomes have prompted concerns for heteroresistance, where a single lineage isolate contains resistant subpopulations not detectable by standard AST. This study aimed to evaluate the prevalence of TBDR mutations among clinical isolates of P. aeruginosa and the phenotypic effect on FDC susceptibility and heteroresistance. We evaluated the sequence of pirR, pirS, pirA, piuA, or piuD from 498 unique isolates collected before the introduction of FDC from four clinical sites in Portland, OR (1), Houston, TX (2), and Santiago, Chile (1). At some clinical sites, TBDR mutations were seen in up to 25% of isolates, and insertion, deletion, or frameshift mutations were predicted to impair protein function were seen in 3% of all isolates (n = 15). Using population analysis profile testing, we found that P. aeruginosa with major TBDR mutations were enriched for a heteroresistant phenotype and undergo a shift in the susceptibility distribution of the population as compared to susceptible strains with wild-type TBDR genes. Our results indicate that mutations in TBDR genes predate the clinical introduction of FDC, and these mutations may predispose to the emergence of FDC resistance.

Blue Light Compromises Bacterial ß-Lactamases Activity to Overcome ß-Lactam Resistance.

OBJECTIVE: In this study, we evaluated the effectiveness of antimicrobial blue light (aBL; 410 nm wavelength) against ß-lactamase-carrying bacteria and the effect of aBL on the activity of ß-lactamases. METHODS: Pseudomonas aeruginosa, Escherichia coli, and Klebsiella pneumoniae strains carrying ß-lactamases as well as a purified ß-lactamase enzymes were studied. ß-lactamase activity was assessed using a chromogenic cephalosporin hydrolysis assay. Additionally, we evaluated the role of porphyrins in the photoreaction, as well as protein degradation by sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Finally, we investigated the bactericidal effect of combined aBL-ceftazidime exposure against a metallo-ß-lactamase expressing P. aeruginosa strain. RESULTS: Our study demonstrated that aBL effectively killed ß-lactamase-producing bacteria and reduced ß-lactamase activity. After an aBL exposure of 1.52 J/cm2, a 50% reduction in enzymatic activity was observed in P. aeruginosa. Additionally, we found a 40% decrease in the photoreaction activity of porphyrins following an aBL exposure of 64.8 J/cm2. We also revealed that aBL reduced ß-lactamase activity via protein degradation (after 136.4 J/cm2). Additionally, aBL markedly improved the bactericidal effect of ceftazidime (by >4-log10) in the metallo-ß-lactamase P. aeruginosa strain. CONCLUSION: Our results provide evidence that aBL compromises bacterial ß-lactamase activity, offering a potential approach to overcome ß-lactam resistance in bacteria.

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.

Synergistic Effect of Ceftazidime-Avibactam with Aztreonam on Carbapenemase-Positive Klebsiella pneumoniae MBL+, NDM.

Background: The difficulties in attaining effective antibiotic therapy arising from the multidrug resistance of Gram-negative bacilli compel the exploration of new possibilities for synergistic interactions among existing antibiotics. Research Design and Methods: An analysis was conducted to assess the efficacy of two antibiotic therapy regimens in the treatment of infections caused by Klebsiella pneumoniae strains producing carbapenemases (MBL). Two patient groups were considered: Group A - individuals in whom the treatment of infection involved the application of ceftazidime-avibactam in combination with aztreonam. Group B comprised patients subjected to an alternative antibiotic therapy regimen. Results: In the group subjected to the treatment regimen involving ceftazidime-avibactam and aztreonam, as compared to alternative antibiotic combinations, a statistically lower mortality rate during the course of treatment and a faster clinical response to the administered therapy were evident. Conclusion: The results obtained may be applicable to routine in vitro assays performed and serve as valuable guidance for the potential utilization of the positive effect of antibiotic therapy through the synergy between ceftazidime-avibactam and aztreonam. The selection of antibiotics employed in the therapy of invasive infections caused by K. pneumoniae influences the ultimate treatment outcome.

Prevalence and risk factors of extended-spectrum beta-lactamase producing E. coli causing urinary tract infections in Iceland during 2012-2021.

PURPOSE: To investigate the association of potential risk factors for urinary tract infections (UTI) caused by E. coli producing ESBL vs. not producing ESBL in Iceland. METHODS: Observational, case-control study including a cohort of 27,747 patients (22,800 females, 4,947 males; 1207 cases, 26,540 controls) of all ages with UTI caused by E. coli in 2012 to 2021 at the clinical microbiology laboratory covering about 2/3 of the Icelandic population. Clinical patient data was obtained from three national databases. Logistic regression was used to calculate odds ratios (ORs) and 95% confidence intervals (CIs) as a measure of association between ESBL and exposure variables. RESULTS: The proportion of samples with ESBL-producing E. coli increased during the study period, from 2.6% in 2012 to 7.6% in 2021 (p < 0.001). ESBL-positive strains were detected in 1207 individuals (4.4%), 905 females (4.0%) and 302 males (6.1%). The following risk factors were identified: Male sex, higher age, institution type (hospital, nursing home), hospital-associated UTI, Charlson comorbidity index score ≥ 3, history of cystitis or hospitalization in the past year, and prescriptions for certain antibiotics or proton pump inhibitors (PPIs: OR 1.51) in the past half year. The antibiotic associated with the highest risk was ciprofloxacin (OR 2.45). CONCLUSION: The prevalence of UTIs caused by ESBL-producing E. coli has been increasing in Iceland. The strongest risk factors for ESBL production were previous antibiotic use, especially ciprofloxacin, and previous PPI use, both considered to be overprescribed. It is important to promote the prudent use of these drugs.

A potential space-making role in cell wall biogenesis for SltB1and DacB revealed by a beta-lactamase induction phenotype in Pseudomonas aeruginosa.

Pseudomonas aeruginosa encodes the beta-lactamase AmpC, which promotes resistance to beta-lactam antibiotics. Expression of ampC is induced by anhydro-muropeptides (AMPs) released from the peptidoglycan (PG) cell wall upon beta-lactam treatment. AmpC can also be induced via genetic inactivation of PG biogenesis factors such as the endopeptidase DacB that cleaves PG crosslinks. Mutants in dacB occur in beta-lactam-resistant clinical isolates of P. aeruginosa, but it has remained unclear why DacB inactivation promotes ampC induction. Similarly, the inactivation of lytic transglycosylase (LT) enzymes such as SltB1 that cut PG glycans has also been associated with ampC induction and beta-lactam resistance. Given that LT enzymes are capable of producing AMP products that serve as ampC inducers, this latter observation has been especially difficult to explain. Here, we show that ampC induction in sltB1 or dacB mutants requires another LT enzyme called MltG. In Escherichia coli, MltG has been implicated in the degradation of nascent PG strands produced upon beta-lactam treatment. Accordingly, in P. aeruginosa sltB1 and dacB mutants, we detected the MltG-dependent production of pentapeptide-containing AMP products that are signatures of nascent PG degradation. Our results therefore support a model in which SltB1 and DacB use their PG-cleaving activity to open space in the PG matrix for the insertion of new material. Thus, their inactivation mimics low-level beta-lactam treatment by reducing the efficiency of new PG insertion into the wall, causing the degradation of some nascent PG material by MltG to produce the ampC-inducing signal. IMPORTANCE: Inducible beta-lactamases like the ampC system of Pseudomonas aeruginosa are a common determinant of beta-lactam resistance among gram-negative bacteria. The regulation of ampC is elegantly tuned to detect defects in cell wall synthesis caused by beta-lactam drugs. Studies of mutations causing ampC induction in the absence of drug therefore promise to reveal new insights into the process of cell wall biogenesis in addition to aiding our understanding of how resistance to beta-lactam antibiotics arises in the clinic. In this study, the ampC induction phenotype for mutants lacking a glycan-cleaving enzyme or an enzyme that cuts cell wall crosslinks was used to uncover a potential role for these enzymes in making space in the wall matrix for the insertion of new material during cell growth.

Cefiderocol for treating severe aerobic Gram-negative bacterial infections: technology evaluation to inform a novel subscription-style payment model.

Background: To limit the use of antimicrobials without disincentivising the development of novel antimicrobials, there is interest in establishing innovative models that fund antimicrobials based on an evaluation of their value as opposed to the volumes used. The aim of this project was to evaluate the population-level health benefit of cefiderocol in the NHS in England, for the treatment of severe aerobic Gram-negative bacterial infections when used within its licensed indications. The results were used to inform the National Institute for Health and Care Excellence guidance in support of commercial discussions regarding contract value between the manufacturer and NHS England. Methods: The health benefit of cefiderocol was first derived for a series of high-value clinical scenarios. These represented uses that were expected to have a significant impact on patients' mortality risks and health-related quality of life. The clinical effectiveness of cefiderocol relative to its comparators was estimated by synthesising evidence on susceptibility of the pathogens of interest to the antimicrobials in a network meta-analysis. Patient-level costs and health outcomes of cefiderocol under various usage scenarios compared with alternative management strategies were quantified using decision modelling. Results were reported as incremental net health effects expressed in quality-adjusted life-years, which were scaled to 20-year population values using infection number forecasts based on data from Public Health England. The outcomes estimated for the high-value clinical scenarios were extrapolated to other expected uses for cefiderocol. Results: Among Enterobacterales isolates with the metallo-beta-lactamase resistance mechanism, the base-case network meta-analysis found that cefiderocol was associated with a lower susceptibility relative to colistin (odds ratio 0.32, 95% credible intervals 0.04 to 2.47), but the result was not statistically significant. The other treatments were also associated with lower susceptibility than colistin, but the results were not statistically significant. In the metallo-beta-lactamase Pseudomonas aeruginosa base-case network meta-analysis, cefiderocol was associated with a lower susceptibility relative to colistin (odds ratio 0.44, 95% credible intervals 0.03 to 3.94), but the result was not statistically significant. The other treatments were associated with no susceptibility. In the base case, patient-level benefit of cefiderocol was between 0.02 and 0.15 quality-adjusted life-years, depending on the site of infection, the pathogen and the usage scenario. There was a high degree of uncertainty surrounding the benefits of cefiderocol across all subgroups. There was substantial uncertainty in the number of infections that are suitable for treatment with cefiderocol, so population-level results are presented for a range of scenarios for the current infection numbers, the expected increases in infections over time and rates of emergence of resistance. The population-level benefits varied substantially across the base-case scenarios, from 896 to 3559 quality-adjusted life-years over 20 years. Conclusion: This work has provided quantitative estimates of the value of cefiderocol within its areas of expected usage within the NHS. Limitations: Given existing evidence, the estimates of the value of cefiderocol are highly uncertain. Future work: Future evaluations of antimicrobials would benefit from improvements to NHS data linkages; research to support appropriate synthesis of susceptibility studies; and application of routine data and decision modelling to assess enablement value. Study registration: No registration of this study was undertaken. Funding: This award was funded by the National Institute for Health and Care Research (NIHR) Health Technology Assessment Policy Research Programme (NIHR award ref: NIHR135591), conducted through the Policy Research Unit in Economic Methods of Evaluation in Health and Social Care Interventions, PR-PRU-1217-20401, and is published in full in Health Technology Assessment; Vol. 28, No. 28. See the NIHR Funding and Awards website for further award information.

This project tested new methods for estimating the value to the NHS of an antimicrobial, cefiderocol, so its manufacturer could be paid fairly even if very little drug is used in order to reduce the risk of bacteria becoming resistant to the product. Clinicians said that the greatest benefit of cefiderocol is when used for complicated urinary tract infections and pneumonia acquired within hospitals caused by two types of bacteria (called Enterobacterales and Pseudomonas aeruginosa), with a resistance mechanism called metallo-beta-lactamase. Because there were no relevant clinical trial data, we estimated how effective cefiderocol and alternative treatments were by doing a systematic literature review of studies that grew bacteria from infections in the laboratory and tested the drugs on them. We linked this to data estimating the long-term health and survival of patients. Some evidence was obtained by asking clinicians detailed questions about what they thought the effects would be based on their experience and the available evidence. We included the side effects of the alternative treatments, some of which can cause kidney damage. We estimated how many infections there would be in the UK, whether they would increase over time and how resistance to treatments may change over time. Clinicians told us that they would also use cefiderocol to treat intra-abdominal and bloodstream infections, and some infections caused by another bacteria called Stenotrophomonas. We estimated how many of these infections there would be, and assumed the same health benefits as for other types of infections. The total value to the NHS was calculated using these estimates. We also considered whether we had missed any additional elements of value. We estimated that the value to the NHS was £18­71 million over 20 years. This reflects the maximum the NHS could pay for use of cefiderocol if the health lost as a result of making these payments rather than funding other NHS services is not to exceed the health benefits of using this antimicrobial. However, these estimates are uncertain due to limitations with the evidence used to produce them and assumptions that had to be made.

In vitro and in vivo activities of KSP-1007, a broad-spectrum inhibitor of serine- and metallo-ß-lactamases, in combination with meropenem against carbapenem-resistant Gram-negative bacteria.

KSP-1007 is a novel bicyclic boronate-based broad-spectrum ß-lactamase inhibitor and is being developed in combination with meropenem (MEM) for the treatment of infections caused by carbapenem-resistant Gram-negative bacteria, a global health concern, and here, we describe its characteristics. KSP-1007 exhibited low apparent inhibition constant (Ki app) values against all classes of ß-lactamase, including imipenemase types and oxacillinase types from Acinetobacter baumannii. Against 207 Enterobacterales and 55 A. baumannii, including carbapenemase producers, KSP-1007 at fixed concentrations of 4, 8, and 16 µg/mL dose-dependently potentiated the in vitro activity of MEM in broth microdilution MIC testing. The MIC90 of MEM/KSP-1007 at 8 µg/mL against Enterobacterales was lower than those of MEM/vaborbactam, ceftazidime/avibactam, imipenem/relebactam, and colistin and similar to those of aztreonam/avibactam, cefiderocol, and tigecycline. The in vitro activity of MEM/KSP-1007 at ≥4 µg/mL against Enterobacterales harboring metallo-ß-lactamase was superior to that of cefepime/taniborbactam. MEM/KSP-1007 showed excellent activity against Escherichia coli with PBP3 mutations and New Delhi metallo-ß-lactamase compared to aztreonam/avibactam, cefepime/taniborbactam, and cefiderocol. MEM/KSP-1007 at 8 µg/mL showed greater efficacy against A. baumannii than these comparators except for cefiderocol, tigecycline, and colistin. A 2-fold reduction in MEM MIC against 96 Pseudomonas aeruginosa was observed in combination with KSP-1007. MEM/KSP-1007 demonstrated bactericidal activity against carbapenemase-producing Enterobacterales, A. baumannii, and P. aeruginosa based on minimum bactericidal concentration/MIC ratios of ≤4. KSP-1007 enhanced the in vivo activity of MEM against carbapenemase-producing Enterobacterales, A. baumannii, and P. aeruginosa in murine systemic, complicated urinary tract, and thigh infection models. Collectively, MEM/KSP-1007 has a good profile for treating carbapenem-resistant Gram-negative bacterial infections.