Many locks to one key: N-acetylneuraminic acid binding to proteins.

Sialic acids play crucial roles in cell surface glycans of both eukaryotic and prokaryotic organisms, mediating various biological processes, including cell-cell interactions, development, immune response, oncogenesis and host-pathogen interactions. This review focuses on the ß-anomeric form of N-acetylneuraminic acid (Neu5Ac), particularly its binding affinity towards various proteins, as elucidated by solved protein structures. Specifically, we delve into the binding mechanisms of Neu5Ac to proteins involved in sequestering and transporting Neu5Ac in Gram-negative bacteria, with implications for drug design targeting these proteins as antimicrobial agents. Unlike the initial assumptions, structural analyses revealed significant variability in the Neu5Ac binding pockets among proteins, indicating diverse evolutionary origins and binding modes. By comparing these findings with existing structures from other systems, we can effectively highlight the intricate relationship between protein structure and Neu5Ac recognition, emphasizing the need for tailored drug design strategies to inhibit Neu5Ac-binding proteins across bacterial species.

Efficacy and safety of different polymyxin-containing regimens for the treatment of pneumonia caused by multidrug-resistant gram-negative bacteria: a systematic review and network meta-analysis.

BACKGROUND: The optimal administration of polymyxins for treating multidrug-resistant gram-negative bacterial (MDR-GNB) pneumonia remains unclear. This study aimed to systematically assess the efficacy and safety of three polymyxin-containing regimens by conducting a comprehensive network meta-analysis. METHODS: We comprehensively searched nine databases. Overall mortality was the primary outcome, whereas the secondary outcomes encompassed microbial eradication rate, clinical success, acute kidney injury, and incidence of bronchospasm. Extracted study data were analyzed by pairwise and network meta-analyses. Version 2 of the Cochrane risk-of-bias tool and the Risk of Bias in Nonrandomized Studies of Interventions (ROBINS-I) assessment tool were used to assess the risk of bias in randomized trials and cohort studies, respectively. RESULTS: This study included 19 observational studies and 3 randomized controlled trials (RCTs), encompassing 3318 patients. Six studies with high risk of bias were excluded from the primary analysis. In the pairwise meta-analysis, compared to the intravenous (IV) polymyxin-containing regimen, the intravenous plus inhaled (IV + IH) polymyxin-containing regimen showed a significant decrease in overall mortality, while no statistically significant difference was found in the inhaled (IH) polymyxin-containing regimen. The network meta-analysis indicated that the IV + IH polymyxin-containing regimen had significantly lower overall mortality (OR 0.67; 95% confidence interval [CI] 0.50-0.88), higher clinical success rate (OR 1.90; 95% CI 1.20-3.00), better microbial eradication rate (OR 2.70; 95% CI 1.90-3.90) than the IV polymyxin-containing regimen, and significantly better microbial eradication rate when compared with the IH polymyxin-containing regimen (OR 2.30; 95% CI 1.30-4.20). Furthermore, compared with IV + IH and IV polymyxin-containing regimens, the IH polymyxin-containing regimen showed a significant reduction in acute kidney injury. CONCLUSIONS: Our study indicates that among the three administration regimens, the IV + IH polymyxin-containing regimen may be the most effective for treating MDR-GNB pneumonia, with a significantly lower overall mortality compared to the IV regimen and a considerably higher microbial eradication rate compared to the IH regimen. The IH regimen may be considered superior to the IV regimen due to its substantially lower incidence of acute kidney injury, even though the reduction in overall mortality was not significant.

A General Strategy for Enhanced Photodynamic Antimicrobial Therapy with Perylenequinonoid Photosensitizers Using a Macrocyclic Supramolecular Carrier.

Perylenequinonoid natural products are a class of photosensitizers (PSs) that exhibit high reactive oxygen species (ROS) generation and excellent activity for Type I/Type II dual photodynamic therapy. However, their limited activity against gram-negative bacteria and poor water solubility significantly restrict their potential in broad-spectrum photodynamic antimicrobial therapy (PDAT). Herein, a general approach to overcome the limitations of perylenequinonoid photosensitizers (PQPSs) in PDAT by utilizing a macrocyclic supramolecular carrier is presented. Specifically, AnBox·4Cl, a water-soluble cationic cyclophane, is identified as a universal macrocyclic host for PQPSs such as elsinochrome C, hypocrellin A, hypocrellin B, and hypericin, forming 1:1 host-guest complexes with high binding constants (≈107 m -1) in aqueous solutions. Each AnBox·4Cl molecule carries four positive charges that promote strong binding with the membrane of gram-negative bacteria. As a result, the AnBox·4Cl-PQPS complexes can effectively anchor on the surfaces of gram-negative bacteria, while the PQPSs alone cannot. In vitro and in vivo experiments demonstrate that these supramolecular PSs have excellent water solubility and high ROS generation, with broad-spectrum PDAT effect against both gram-negative and gram-positive bacteria. This work paves a new path to enhance PDAT by showcasing an efficient approach to improve PQPSs' water solubility and killing efficacy for gram-negative bacteria.

Modulating the biosynthesis and TLR4-interaction of lipopolysaccharide as an approach to counter gut dysbiosis and Parkinson's disease: Role of phyto-compounds.

The prevalence of the world's second leading neurodegenerative disorder Parkinson's disease (PD) is well known while its pathogenesis is still a topical issue to explore. Clinical and experimental reports suggest the prevalence of disturbed gut microflora in PD subjects, with an abundance of especially Gram-negative bacteria. The endotoxin lipopolysaccharide (LPS) released from the outer cell layer of these bacteria interacts with the toll-like receptor 4 (TLR4) present on the macrophages and it stimulates the downstream inflammatory cascade in both the gut and brain. Recent research also suggests a positive correlation between LPS, alpha-synuclein, and TLR4 levels, which indicates the contribution of a parallel LPS-alpha-synuclein-TLR4 axis in stimulating inflammation and neurodegeneration in the gut and brain, establishing a body-first type of PD. However, owing to the novelty of this paradigm, further investigation is mandatory. Modulating LPS biosynthesis and LPS-TLR4 interaction can ameliorate gut dysbiosis and PD. Several synthetic LpxC (UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase; LPS-synthesizing enzyme) inhibitors and TLR4 antagonists are reported to show beneficial effects including neuroprotection in PD models, however, are not devoid of side effects. Plant-derived compounds have been long documented for their benefits as nutraceuticals and thus to search for effective, safer, and multitarget therapeutics, the present study focused on summarizing the evidence reporting the potential of phyto-compounds as LpxC inhibitors and TLR4 antagonists. Studies demonstrating the dual potential of phyto-compounds as the modulators of LpxC and TLR4 have not yet been reported. Also, very few preliminary studies have reported LpxC inhibition by phyto-compounds. Nevertheless, remarkable neuroprotection along with TLR4 antagonism has been shown by curcumin and juglanin in PD models. The present review thus provides a wide look at the research progressed to date in discovering phyto-compounds that can serve as LpxC inhibitors and TLR4 antagonists. The study further recommends the need for expanding the search for potential candidates that can render dual protection by inhibiting both the biosynthesis and TLR4 interaction of LPS. Such multitarget therapeutic intervention is believed to bring fruitful yields in countering gut dysbiosis, neuroinflammation, and dopaminergic neuron damage in PD patients through a single treatment paradigm.

Stannous fluoride forms aggregates between outer and inner membranes leading to membrane rupture of Porphyromonas gingivalis and Prevotella pallens.

Objective: Stannous has been shown to bind to free lipopolysaccharides, thus preventing them from binding to TLR receptors. This study was undertaken to determine the histomorphological mechanism of stannous binding to anaerobic bacteria. Methods: Two bacteria associated with gingivitis and advanced periodontal disease, Porphyromonas gingivalis (P. gingivalis) and Prevotella pallens (P. pallens), were cultured in 25-1,000 µM of stannous fluoride and stannous chloride for 48 h. The growth rate was estimated using absorbance OD600. Bacterial cells were then fixed and processed for transmission electron microscopy (TEM) analysis. Results: Stannous fluoride inhibited proliferation of both P. gingivalis and P. pallens in a dose-dependent manner. There was a statistically significant suppression of the growth curve starting at 100 µM for P. pallens (P = 0.050) and 200 µM for P. gingivalis (P = 0.039). TEM analysis revealed a thick layer of polysaccharides (19.8 nm) in P. gingivalis. The outer and inner membranes were clearly visible with low electron densities in both bacteria. Stannous diffused into bacterial membranes and formed precipitates in the areas spanning outer and inner membranes and below inner membranes. Precipitates varied in size ranging from 46.4 to 84.5 nm in length, and 18.4 to 35.9 nm in width. The membranes were disintegrated in the region where stannous formed precipitates. Cytosolic contents were leaked out, and in several cases, small vesicles were formed. Stannous precipitates were more abundant in numbers and larger in size in bacteria treated with high concentrations (100-300 µM) than in low concentrations (25-50 µM) of stannous fluoride. Furthermore, most of the bacteria were disintegrated in the groups treated with 100-300 µM stannous fluoride. At low concentrations (25 µM), stannous fluoride formed complexes primarily around outer membranes, to which lipopolysaccharides are anchored. Stannous chloride results showed similar trends, but it was less potent than stannous fluoride. Conclusion: Stannous fluoride can penetrate bacteria, bind to the constituents of the membrane and form precipitates between outer and inner membranes and beneath inner membranes. These large precipitates damaged the integrity of membranes and allowed cytosolic contents to be leaked out. Stannous complexes formed at the outer membranes, even at low concentrations (25 µM).

In vitro evaluation of using ceftazidime/avibactam against carbapenem-resistant Acinetobacter baumannii.

OBJECTIVE: Carbapenem-resistant Acinetobacter baumannii (CRAB) is a global concern as effective treatments are very limited. We previously used a modified susceptibility testing approach to predict growth suppression in carbapenem-resistant Enterobacterales, but there are uncertainties about the generalizability of the model. The objective of this study is to verify if a similar approach can be extended to CRAB. METHOD: A clinical isolate of CRAB resistant to ceftazidime/avibactam (CAZ/AVI, MIC=32/4 mg/L) was examined. CAZ susceptibility was determined using increasing concentrations of AVI (0-64 mg/L), and MIC reduction was characterized with a sigmoid inhibitory maximum effect (Emax) model. The effectiveness of CAZ/AVI was validated in a hollow fiber infection model (HFIM) over 72 hours, using simulated unbound serum / epithelial lining fluid (ELF) exposures of 2.5 g over 2 hours every 8 hours. Baseline inocula of approximately 5.5 log CFU/mL were examined. RESULTS: An AVI concentration-dependent reduction in CAZ MIC was observed (r2=0.99). Ceftazidime MIC was dramatically reduced from 512 mg/L (no AVI) to 32 mg/L (AVI=4 mg/L), and further to 8 mg/L (AVI=16 mg/L). Pharmacokinetic simulations were satisfactory in the HFIM (r2>0.96). Bacterial suppression was observed > 24 hours with the serum exposure, but not that from the ELF. CONCLUSION: Using multiple AVI concentrations within the clinically relevant range, our susceptibility testing approach could have better insights of treatment outcome for infections caused by CRAB. This could potentially lead to effective intervention(s) overlooked by conventional susceptibility testing method. This case highlights the importance of site-specific drug exposures on determining treatment outcome.

Analysis of mcr family of colistin resistance genes in Gram-negative isolates from a tertiary care hospital in India.

AIM: Colistin serves as the drug of last resort for combating numerous multidrug-resistant (MDR) Gram-negative infections. Its efficacy is hampered by the prevalent issue of colistin resistance, which severely limits treatment options for critically ill patients. Identifying resistance genes is crucial for controlling resistance spread, with horizontal gene transfer being the primary mechanism among bacteria. This study aimed to assess the prevalence of plasmid-mediated mcr genes associated with colistin resistance in Gram-negative bacteria, utilizing both genotypic and phenotypic tests. METHODS AND RESULTS: The clinical isolates (n=913) were obtained from a tertiary care center in Chennai, India. Colistin resistance was seen among Gram-negative isolates. These strains underwent screening for mcr-1, mcr-3, mcr-4, and mcr-5 genes via conventional PCR. Additionally, mcr-positive isolates were confirmed through Sanger sequencing and phenotypic testing. The bacterial isolates predominantly comprised Klebsiella pneumoniae (62.43%), Escherichia coli (19.71%), Pseudomonas aeruginosa (10.73%), Acinetobacter baumannii (4.81%), along with other species. All isolates exhibited multidrug resistance to three or more antibiotic classes. Colistin resistance, determined via broth microdilution (BMD) using CLSI guidelines, was observed in 13.08% of the isolates studied. Notably, mcr-5 was detected in K. pneumoniae in PCR, despite their absence in Sanger sequencing and phenotypic tests (including the combined-disk test, colistin MIC in the presence of EDTA, and Zeta potential assays). This finding underscores the importance of employing multiple diagnostic approaches to accurately identify colistin resistance mechanisms. CONCLUSION AND IMPACT: The study highlights a concerning prevalence of colistin resistance among Enterobacterales, especially those producing carbapenemase, thereby impacting mortality rates. Nonetheless, further investigations are warranted to elucidate common mechanisms of colistin resistance and to evaluate the efficacy of screening techniques in detecting isolates carrying mcr genes responsible for enzyme-mediated lipopolysaccharide (LPS) modification.

Ignatzschineria indica Bacteremia Initially Misdiagnosed in a Patient With a Maggot-Infested Wound.

Gram-negative bacteremia in hospitalized patients often leads to prolonged hospital stays, increased healthcare costs, and mortality rates. Simultaneously, the presence of comorbidities like chronic wounds increases the risk of severe infection and complicated hospital courses involving amputation, broad-spectrum antibiotic use, and repeat hospital admissions, after discharge. This case presents a 72-year-old male with a past medical history significant for chronic lower extremity cellulitis with multiple prior hospitalizations. On admission, the patient had a chief complaint of progressively worsening left lower extremity pain along with nausea, vomiting, and diarrhea. CT imaging of the left lower extremity suggested severe cellulitis without signs of osteomyelitis. Blood cultures initially suggested Corynebacterium jeikeium, but were sent to an outside facility due to ambiguity of results. The outside facility identified the pathogen as Ignatzschineria indica. After confirming the results, antibiotics were appropriately de-escalated to oral levofloxacin. The patient continued to show clinical improvement and was discharged with follow-up appointments scheduled for infectious disease and bi-weekly visits to wound care. Considering the increasing prevalence of chronic wounds in the United States, awareness and recognition of emerging pathogens are crucial for the timely diagnosis, treatment, and management of these complex patients. Our case adds to the growing body of reports on the management of I. indica bacteremia resulting from maggot-infested wounds.

Analysis of Risk Factors for Urinary Tract Infections in Pregnant Women: A Retrospective Study.

OBJECTIVE: Urinary tract infections (UTIs) are the most common bacterial infection during pregnancy. This study aimed to investigate the risk factors of UTI during pregnancy. METHODS: In this study, pregnant women who underwent prenatal examination in our hospital from October 2019 to October 2023 were divided into UTI group and non-UTI group in accordance with whether or not they had a UTI. The general data, clinical data and laboratory indicators of the participants were collected. Multivariate logistic regression was used to analyse the influencing factors of UTI in pregnant women, and the results were shown with odds ratio (OR) and 95% confidence interval (95% CI). RESULTS: A total of 600 pregnant women were included in the study. The results found that 56 women (9.33%) had a combined UTI. The results of midstream urinary bacterial culture in the UTI group showed that Gram-negative bacteria accounted for 60.71% of all detected pathogenic bacteria, and Escherichia coli and Staphylococcus aureus were common strains, accounting for 46.43% and 23.21%, respectively. The proportions of patients in the UTI group who were ≥35 years old, had a high school education or below, had a history of abortion, had gestational diabetes, had ≥three vaginal and anal examinations, had a history of UTI and had urinary tract stones were significantly higher than the non-UTI group (p < 0.05). Multivariate logistic regression analysis showed that age ≥35 years (OR = 9.127; 95% CI: 4.668-17.810; p < 0.001), educational level of high school or lower (OR = 4.184; 95% CI: 2.448-7.160; p < 0.001), gestational diabetes (OR = 3.494; 95% CI: 1.789-6.803; p < 0.001), UTI history (OR = 2.074; 95% CI: 1.114-3.834; p < 0.001) and haemoglobin (Hb) <100 g/L (OR = 8.022; 95% CI: 4.532-14.325; p < 0.001) are risk factors for UTI in pregnant women. CONCLUSIONS: The common pathogenic bacteria of pregnant women with UTI are mainly Gram-negative bacteria. Older pregnant women, low educational level, gestational diabetes mellitus, history of UTI and anaemia may be risk factors for UTI in pregnant women.

Molecular characteristics and evaluation of the phenotypic detection of carbapenemases among Enterobacterales and Pseudomonas via whole genome sequencing.

Background/purposes: The continuously increasing carbapenem resistance within Enterobacterales and Pseudomonas poses a threat to public health, nevertheless, the molecular characteristics of which in southern China still remain limited. And carbapenemase identification is a key factor in effective early therapy of carbapenem-resistant bacteria infections. We aimed to determine the molecular characteristics of these pathogens and compare commercial combined disc tests (CDTs) with the modified carbapenem inactivation method (mCIM) and EDTA-CIM (eCIM) in detecting and distinguishing carbapenemases using whole genome sequencing (WGS). Methods: A total of 78 Enterobacterales, 30 Pseudomonas were obtained from two tertiary hospitals in southern China. Susceptibility tests were conducted using an automated VITEK2 compact system with confirmation via the Kirby-Bauer method. The WGS was conducted on all clinical isolates and the molecular characteristics were analyzed by screening the whole genome sequences. CDTs with or without cloxacillin, mCIM, and eCIM, were performed and compared by taking WGS results as the benchmark. Results: A total of 103 carbapenem non-susceptible and 5 carbapenem susceptible bacteria were determined, with Klebsiella pneumoniae (42.7%), Pseudomonas aeruginosa (23.3%) and Escherichia coli (18.4%) being most prevalent. Carbapenemase genes were detected in 58 (56.3%) of the 103 carbapenem-non-susceptible clinical isolates, including 46 NDM, 6 KPC, 3 IMP, 1 IPM+VIM,1NDM+KPC, and 1 OXA-181. Carbapenemase-producing isolates were detected more frequently in Enterobacterales (76.3%). Among K. pneumoniae, the major sequence types were st307 and st11, while among E. coli and P. aeruginosa, the most prevalent ones were st410 and st242 respectively. For carbapenemase detection in Enterobacterales, the mCIM method achieved 100.00% (95% CI, 92.13-100.00%) sensitivity and 94.44% (70.63-99.71%) specificity (kappa, 0.96); for Pseudomonas, detection sensitivity was 100% (5.46-100.00%), and 100% (84.50-100.00%) specificity (kappa, 0.65). Commercial CDT carbapenemase detection sensitivity for Enterobacterales was 96.49% (86.84-99.39%), and 95.24% (74.13-99.75%) specificity (kappa, 0.90); for Pseudomonas, carbapenemase detection sensitivity was 100.00% (5.46-100.00%) and 37.93% (21.30-57.64%) specificity (kappa, 0.04). When cloxacillin testing was added, CDT specificity reached 84.61% (64.27-94.95%). Conclusion: The molecular epidemiology of carbapenem-non-susceptible isolates from pediatric patients in Southern China exhibited distinctive characteristics. Both the mCIM-eCIM combination and CDT methods effectively detected and differentiated carbapenemases among Enterobacterales isolates, and the former performed better than CDT among Pseudomonas.

Risk factors and mortality rates of carbapenem-resistant Gram-negative bacterial infections in intensive care units.

Background: The prevalence of hospital-acquired infections caused by carbapenem-resistant gram-negative bacteria (CRGNB) is increasing worldwide. Several risk factors have been associated with such infections. The present study aimed to identify risk factors and determine the mortality rates associated with CRGNB infections in intensive care units. Methods: This retrospective case-control study was conducted at Erciyes University Hospital (Kayseri, Turkey) between January 2017 and December 2021. Demographic and laboratory data were obtained from the Infection Control Committee data and record system. Patients who had CRGNB infection 48-72 h after hospitalization were assigned to the case group, while those who were not infected with CRGNB during hospitalization formed the control group. Risk factors, comorbidity, demographic data, and mortality rates were compared between the two groups. Results: Approximately 1449 patients (8.97%) were monitored during the active follow-up period; of those, 1171 patients were included in this analysis. CRGNB infection developed in 14 patients (70.00%) who had CRGNB colonization at admission; in 162 (78.26%) were colonized during hospitalization, whereas 515 (54.56%) were not colonized. There was no significant difference in age, sex (male/female) or comorbidities. The total length of hospital stay was statistically significantly longer (P=0.001) in the case group (median: 24 [interquartile range: 3-378] days) than the control group (median: 16 [interquartile range: 3-135] days). The rates of colonization at admission (25.5%; vs. 10.6%, P=0.001) and mortality (64.4% vs. 45.8%, P=0.001) were also significantly higher in the cases than in the control group, respectively. In the univariate analysis, prolonged hospitalization, the time from intensive care unit admission to the development of infection, presence of CRGNB colonization at admission, transfer from other hospitals, previous antibiotic use, enteral nutrition, transfusion, hemodialysis, mechanical ventilation, tracheostomy, reintubation, central venous catheter, arterial catheterization, chest tube, total parenteral nutrition, nasogastric tube use, and bronchoscopy procedures were significantly associated with CRGNB infections (P <0.05). Multivariate analysis identified the total length of stay in the hospital (odds ratio [OR]=1.02; 95% confidence interval [CI]: 1.01 to 1.03; P=0.001), colonization (OR=2.19; 95% CI: 1.53 to 3.13; P=0.001), previous antibiotic use (OR=2.36; 95% CI: 1.53 to 3.62; P=0.001), intubation (OR=1.59; 95% CI: 1.14 to 2.20; P=0.006), tracheostomy (OR=1.42; 95% CI: 1.01 to 1.99; P=0.047), and central venous catheter use (OR=1.62; 95% CI: 1.20 to 2.19; P=0.002) as the most important risk factors for CRGNB infection. Conclusions: Colonization, previous use of antibiotics, and invasive interventions were recognized as the most important risk factors for infections. Future research should focus on measures for the control of these parameters.

Membrane Disruption-Enhanced Photodynamic Therapy against Gram-Negative Bacteria by a Peptide-Photosensitizer Conjugate.

Photodynamic therapy (PDT) emerges as a promising strategy for combating bacteria with minimal drug resistance. However, a significant hurdle lies in the ineffectiveness of most photosensitizers against Gram-negative bacteria, primarily attributable to their characteristic impermeable outer membrane (OM) barrier. To tackle this obstacle, we herein report an amphipathic peptide-photosensitizer conjugate (PPC) with intrinsic outer membrane disruption capability to enhance PDT efficiency against Gram-negative bacteria. PPC is constructed by conjugating a hydrophilic ultrashort cationic peptide to a hydrophobic photosensitizer. PPC could efficiently bind to the OM of Gram-negative bacteria through electrostatic adsorption, and subsequently disrupt the structural integrity of the OM. Mechanistic investigations revealed that PPC triggers membrane disruption by binding to both lipopolysaccharide (LPS) and phospholipid leaflet in the OM, enabling effective penetration of PPC into the Gram-negative bacteria interior. Upon light irradiation, PPC inside bacteria generates singlet oxygen not only to effectively decrease the survival of Gram-negative bacteria P. aeruginosa and E. coli to nearly zero in vitro, but also successfully cure the full-thickness skin infection and bacterial keratitis (BK) induced by P. aeruginosa in animal models. Thus, this study provides a broad-spectrum antibacterial phototherapeutic design strategy by the synergistic action of membrane disruption and PDT to combat Gram-negative bacteria.

Burden and Management of Multi-Drug Resistant Organism Infections in Solid Organ Transplant Recipients Across the World: A Narrative Review.

Solid organ transplant (SOT) recipients are particularly susceptible to infections caused by multidrug-resistant organisms (MDRO) and are often the first to be affected by an emerging resistant pathogen. Unfortunately, their prevalence and impact on morbidity and mortality according to the type of graft is not systematically reported from high-as well as from low and middle-income countries (HIC and LMIC). Thus, epidemiology on MDRO in SOT recipients could be subjected to reporting bias. In addition, screening practices and diagnostic resources may vary between countries, as well as the availability of new drugs. In this review, we aimed to depict the burden of main Gram-negative MDRO in SOT patients across HIC and LMIC and to provide an overview of current diagnostic and therapeutic resources.

Phenotypic identification of different ß-Lactamases in intrinsic and acquired colistin resistant uropathogenic gram negative bacteria.

Objective: Identification of MBL, AmpC and ESBLs in colistin intrinsic and acquired resistant uropathogenic gram negative bacteria. Method: Urine samples were collected from Hayatabad Medical Complex, Peshawar during 17 January to 30 June 2019. Collected urine samples were aseptically transported microbiology lab of Health Research Institution (HRI), National Institute of Health (NIH), Khyber Medical College, Peshawar and streaked on different media. Positive growth was identified by API-10s. Antibiotic sensitivity profile was done by Modified Kirby Bauer disc diffusion method. Detection of metallo ßlactamases (MBL) production by Imipenem EDTA synergy test, Double Disc Synergy Test (DDST) for detection of ESBLs and D-test for the detection of inducible AmpC beta lactamases test was used. Colistin resistance was identified via broth micro dilution according to CLSI manual. Colistin resistant bacteria was divided in two categories; acquired and intrinsic resistant bacteria according to CLSI manual. Results: Out of 2000 urine samples, 281(14%) gram-negative bacteria were isolated. Among positive samples, acquired colistin resistant bacteria were 241 and intrinsic resistant bacteria were 40 isolates. MBL was produce by twenty one (11.7%) E.coli and seventeen (40.5%) Pseudomonas aeruginosa. E. coli, Pseudomonas aeruginosa, Klebsiella Pneumoniae, Serratia Oderifora and Proteus Marblis were ESBLs producing bacteria. AmpC production was prevalent in fourteen (7.8%) E. coli and twelve (28.6%) Pseudomonas aeruginosa. Fifty-five samples showed resistance to colistin out of 241 samples. In colistin resistant bacteria, two E.coli were MBL, ESBLs, while one E.coli was ESBLs, AmpC co-producing bacteria. The most prevalent extended drug resistant bacteria were Pseudomonas aeruginosa (28.6%) and Escherichia coli (6.1%), While 155(86.6%) Escherichia coli, 25 (59.5%) Pseudomonas aeruginosa and 22 (95.7%) Serratia Oderifora was multi drug resistant bacteria. Conclusion: Current study concluded that ESBL, MBL AmpC enzymes and their co-expression was observed with colistin resistance in E.coli and Pseudomonas aeruginosa.

Adult acute leukemia patients with gram-negative bacteria bloodstream infection: Risk factors and outcomes of antibiotic-resistant bacteria.

This study aims to analyze the risk factors for the development of multidrug-resistant (MDR) and carbapenem-resistant (CR) bacteria bloodstream infection (BSI) in a patient with acute leukemia (AL) and the mortality in gram-negative bacteria (GNB) BSI. This is a retrospective study conducted at West China Hospital of Sichuan University, which included patients diagnosed with AL and concomitant GNB BSI from 2016 to 2021. A total of 206 patients with GNB BSI in AL were included. The 30-day mortality rate for all patients was 26.2%, with rates of 25.8% for those with MDR GNB BSI and 59.1% for those with CR GNB BSI. Univariate and multivariate analyses revealed that exposure to quinolones (Odds ratio (OR) = 3.111, 95% confidence interval (95%CI): 1.623-5.964, p = 0.001) within the preceding 30 days was an independent risk factor for MDR GNB BSI, while placement of urinary catheter (OR = 6.311, 95%CI: 2.478-16.073, p < 0.001) and exposure to cephalosporins (OR = 2.340, 95%CI: 1.090-5.025, p = 0.029) and carbapenems (OR = 2.558, 95%CI: 1.190-5.497, p = 0.016) within the preceding 30 days were independently associated with CR GNB BSI. Additionally, CR GNB BSI (OR = 2.960, 95% CI: 1.016-8.624, p = 0.047), relapsed/refractory AL (OR = 3.035, 95% CI: 1.265-7.354, p = 0.013), septic shock (OR = 5.108, 95% CI: 1.794-14.547, p = 0.002), platelets < 30 × 109/L before BSI (OR = 7.785, 95% CI: 2.055-29.492, p = 0.003), and inappropriate empiric antibiotic therapy (OR = 3.140, 95% CI: 1.171-8.417, p = 0.023) were independent risk factors for 30-day mortality in AL patients with GNB BSI. Prior antibiotic exposure was a significant factor in the occurrence of MDR GNB BSI and CR GNB BSI. CR GNB BSI increased the risk of mortality in AL patients with GNB BSI.

Potent synergy and sustained bactericidal activity of polymyxins combined with Gram-positive only class of antibiotics versus four Gram-negative bacteria.

BACKGROUND: Gram-negative bacteria (GNB) are becoming increasingly resistant to a wide variety of antibiotics. There are currently limited treatments for GNB, and the combination of antibiotics with complementary mechanisms has been reported to be a feasible strategy for treating GNB infection. The inability to cross the GNB outer membrane (OM) is an important reason that a broad spectrum of Gram-positive only class of antibiotics (GPOAs) is lacking. Polymyxins may help GPOAs to permeate by disrupting OM of GNB. OBJECTIVE: To identify what kind of GPOAs can be aided to broaden their anti-GNB spectrum by polymyxins, we systematically investigated the synergy of eight GPOAs in combination with colistin (COL) and polymyxin B (PMB) against GNB in vitro. METHODS: The synergistic effect of COL or PMB and GPOAs combinations against GNB reference strains and clinical isolates were determined by checkerboard tests. The killing kinetics of the combinations were assessed using time-kill assays. RESULTS: In the checkerboard tests, polymyxins-GPOAs combinations exert synergistic effects characterized by species and strain specificity. The synergistic interactions on P. aeruginosa strains are significantly lower than those on strains of A. baumannii, K. pneumoniae and E. coli. Among all the combinations, COL has shown the best synergistic effect in combination with dalbavancin (DAL) or oritavancin (ORI) versus almost all of the strains tested, with FICIs from 0.16 to 0.50 and 0.13 to < 0.28, respectively. In addition, the time-kill assays demonstrated that COL/DAL and COL/ORI had sustained bactericidal activity. CONCLUSIONS: Our results indicated that polymyxins could help GPOAs to permeate the OM of specific GNB, thus showed synergistic effects and bactericidal effects in the in vitro assays. In vivo combination studies should be further conducted to validate the results of this study.

Assessment of Posttransplant Bacteremia Caused by Extended-Spectrum Beta-Lactamase-Producing Gram-Negative Bacteria Among Kidney Transplant Recipients.

BACKGROUND: Extended-spectrum beta-lactamase-producing gram-negative rods (ESBL-GNR) are a rising cause of bacteremia in kidney transplant recipients (KT). The study purpose was to examine patient mortality, allograft survival, estimated glomerular filtration rate (eGFR) at the end of 1 year, and readmission rates while looking at treatment strategies among KTs with ESBL-GNR and non-ESBL-GNR bacteremia at our institution. METHODS: This study was a retrospective, cohort analysis of KTs with gram-negative bacteremia from January 1, 2020, to December 31, 2021. The primary outcome of the study was mortality. Patient outcomes were assessed for 365 days after positive blood cultures. RESULTS: The study included 63 patients. Of these, 18 (29%) patients had bacteremia caused by an ESBL-GNR and 45 (71%) patients had bacteremia caused by a non-ESBL-GNR. Patient survival at 90 days was 94% in the ESBL-GNR group and 96% in the non-ESBL-GNR group. Ciprofloxacin was the most common antimicrobial therapy at discharge (68.9%) in the non-ESBL-GNR group whereas ertapenem was the most common in the ESBL-GNR group (44.5%). Median eGFR at discharge was 41 mL/min/1.73 m2 in the ESBL-GNR group and 48 mL/min/1.73 m2 in the non-ESBL-GNR group. Ninety-day readmission occurred in 9 (50%) ESBL-GNR patients and 14 (32%) non-ESBL-GNR patients. None of the above comparisons are statistically significant (p > 0.05). Eleven (61%) ESBL-GNR and 2 (4%) non-ESBL-GNR patients used outpatient parenteral antimicrobial therapy (p < 0.001). CONCLUSIONS: Among KTs with ESBL-GNR bacteremia, no significant difference was detected in mortality or allograft function compared to non-ESBL-GNR bacteremia.

What's old is new: leveraging existing antimicrobial susceptibility test methods for rapid results in patients with bloodstream infections.

The use of rapid disk diffusion or modified automated antimicrobial susceptibility testing (AST) system approaches demonstrates excellent performance for gram-negative organisms directly from blood cultures. In a recent study, S. Khan, A. Das, A. Mishra, A. Vidyarthi, et al. (Microbiol Spectr 12:e03081-23, 2024, https://doi.org/10.1128/spectrum.03081-23) compared the performance of three direct-from-blood AST methods against standard of care disk diffusion and automated AST. The results demonstrated high categorical agreements and low error rates across three protocols. The study suggests that locally validated direct-from-blood AST protocols offer reliable and fast results, particularly for resource-limited settings. However, local context and workflows should be considered prior to implementing rapid AST protocols, and more research is needed on the performance of rapid AST protocols for gram-positive organisms.

Antibiotic adjuvants against multidrug-resistant Gram-negative bacteria: important component of future antimicrobial therapy.

The swift emergence and propagation of multidrug-resistant (MDR) bacterial pathogens constitute a tremendous global health crisis. Among these pathogens, the challenge of antibiotic resistance in Gram-negative bacteria is particularly pressing due to their distinctive structure, such as highly impermeable outer membrane, overexpressed efflux pumps, and mutations. Several strategies have been documented to combat MDR Gram-negative bacteria, including the structural modification of existing antibiotics, the development of antimicrobial adjuvants, and research on novel targets that MDR bacteria are sensitive to. Drugs functioning as adjuvants to mitigate resistance to existing antibiotics may play a pivotal role in future antibacterial therapy strategies. In this review, we provide a brief overview of potential antibacterial adjuvants against Gram-negative bacteria and their mechanisms of action, and discuss the application prospects and potential for bacterial resistance to these adjuvants, along with strategies to reduce this risk.

Bacterial efflux pump OMPs as vaccine candidates against Multidrug-Resistant Gram-negative bacteria.

The emergence and propagation of bacteria resistant to antimicrobial drugs is a serious public health threat worldwide. The current antibacterial arsenal is becoming obsolete and the pace of drug development is decreasing, highlighting the importance of investment in alternative approaches to treat or prevent infections caused by antimicrobial-resistant bacteria. A significant mechanism of antimicrobial resistance employed by Gram-negative bacteria is the overexpression of efflux pumps that can extrude several compounds from the bacteria, including antimicrobials. The overexpression of efflux pump proteins has been detected in several multidrug resistant (MDR) Gram-negative bacteria, drawing attention to these proteins as potential targets against these pathogens. This review will focus on the role of outer membrane proteins (OMPs) from efflux pumps as potential vaccine candidates against clinically relevant MDR Gram-negative bacteria, discussing advantages and pitfalls. Additionally, we will explore the relevance of efflux pump OMP diversity and the possible impact of vaccination on microbiota.