Flow Cytometry and Automated Microscopy Integration for Phagocytic Analysis of Hypervirulent Klebsiella pneumoniae in Dictyostelium discoideum.

This chapter introduces protocols for culturing and maintaining Dictyostelium discoideum and methods for conducting virulence assays in this organism to study bacterial pathogenicity. It outlines advanced techniques, such as automated microscopy and flow cytometry, for detailed cellular analysis and traditional microbiological approaches. These comprehensive protocols will enable researchers to probe the virulence factors of pathogens like Klebsiella pneumoniae and to elucidate the details of host-pathogen interactions within a cost-effective and adaptable laboratory framework.

Zebrafish Larvae Microinjection and Automated Fluorescence Microscopy for Studying Klebsiella pneumoniae Infection and the Host Immune Response.

Studying host-pathogen interactions is essential for understanding infectious diseases and developing possible treatments, especially for priority pathogens with increased virulence and antibiotic resistance, such as Klebsiella pneumoniae. Over time, this subject has been approached from different perspectives, often using mammal host models and invasive endpoint measurements (e.g., sacrifice and organ extraction). However, taking advantage of technological advances, it is now possible to follow the infective process by noninvasive visualization in real time, using optically amenable surrogate hosts. In this line, this chapter describes a live-cell imaging approach to monitor the interaction of K. pneumoniae and potentially other bacterial pathogens with zebrafish larvae in vivo. This methodology is based on the microinjection of fluorescent bacteria into the otic vesicle, followed by time-lapse observation by automated fluorescence microscopy with environmental control, monitoring the dynamics of immune cell recruitment, bacterial load, and larvae survival.

Computational Exploration of Limonin as a Potential Inhibitor of DapB in Klebsiella pneumoniae.

Klebsiella pneumoniae has emerged as a significant multidrug-resistant pathogen, classified as a critical priority by the World Health Organization. The rising rates of antibiotic resistance have led to increased therapeutic failures, diminishing the effectiveness of existing antibiotics. Consequently, there is an urgent need for alternative treatments to effectively inhibit the growth of K. pneumoniae and mitigate associated diseases. Phytochemicals have demonstrated potential advantages over traditional antibiotics, prompting their exploration as innovative therapeutic agents. This study aimed to identify phytochemicals that can inhibit dapB, a vital enzyme in the lysine biosynthesis pathway of K. pneumoniae, which is essential for protein synthesis and the cross-linking of the bacterial peptidoglycan cell wall. We screened 17,934 phytochemicals based on Lipinski's Rule of Five, along with their Absorption, Distribution, Metabolism, Excretion properties and toxicological parameters. Next, we conducted triplicate docking studies against dapB to evaluate the library further. The most promising molecules then underwent 100 ns Molecular Dynamics simulations in triplicate, followed by binding free energy calculations to identify potential dapB inhibitors. This in silico analysis highlighted limonin as a promising inhibitor of dapB in K. pneumoniae. Further experimental validation is crucial to enhance limonin's potential as a novel therapeutic agent against K. pneumoniae-associated diseases.

Clinical experience with ceftazidime/avibactam for the treatment of extensively drug-resistant or pandrug-resistant Klebsiella pneumoniae in neonates and children.

PURPOSE: Klebsiella pneumoniae is a significant cause of healthcare-associated infections, resulting in high morbidity and mortality rates due to limited treatment options. In this study, we aimed to evaluate the treatment outcomes and the safety of Ceftazidime-avibactam in infections caused by extensively drug-resistant or pandrug-resistant Klebsiella pneumoniae in pediatric patients. METHODS: This study included pediatric patients who received ceftazidime-avibactam treatment due to extensively drug-resistant or pandrug-resistant Klebsiella pneumoniae infections, monitored in the pediatric intensive care, neonatal intensive care, and pediatric wards of Cukurova University Faculty of Medicine between 2022 and 2023. Patients' microbiological responses, clinical responses, medication side effects, and 30-day survival rates were evaluated. RESULTS: Eleven pediatric patients were included in the study, of whom nine were male (81.8%). The median age at the initiation of ceftazidime-avibactam treatment was 15 months (range: 14 days-183 months). Sepsis was diagnosed in 9 patients (81.8%). Two premature infants (27 and 35 weeks) were admitted to the neonatal ICU. Regarding the Klebsiella pneumoniae strains, 10 (91%) were extensively drug-resistant (XDR), and 1 (9%) was pandrug-resistant (PDR). Eight strains (72.7%) were carbapenem-resistant, and 9 (81.8%) were colistin-resistant. Microbiological response was noted in 8 patients (72.7%), clinical response was evident in 6 patients (54.5%). The 30-day survival rate was 54.5%, with six patients surviving. CONCLUSION: In our study, ceftazidime-avibactam has been identified as a significant treatment option for resistant Klebsiella pneumoniae infection in critically ill children and premature infants with sepsis and organ failure, and it has been found to be well tolerated.

Prevalence and molecular characterization of carbapenem-resistant Enterobacterales in patients from a public referral hospital in a non-metropolitan region of Brazil during and post the SARS-CoV-2 pandemic.

Antimicrobial resistance (AMR) poses a global threat, with carbapenem-resistant Enterobacterales (CRE) representing a significant concern due to limited therapeutic options. This study investigated the prevalence of carbapenemase genes in CRE strains isolated from tracheal aspirates of patients at a Brazilian university hospital between January 2020 and August 2023. Bacterial identification was conducted using MALDI-TOF, while carbapenemase genes were detected by qPCR. Demographic and clinical data were collected, and univariate analysis was performed using the chi-square test (p < 0.05). Variables with p ≤ 0.10 were further investigated using the chi-square test for linear trend, along with stratified analysis. Out of 1,133 samples, 111 (9.79%) showed CRE growth, with 46 isolates included in the final sample, predominantly comprising Klebsiella pneumoniae (65.21%) and Serratia marcescens (19.57%). The blaKPC gene was prevalent (78.26%), while blaNDM was detected in 21.74% of cases. The identified population was predominantly male (67.39%), elderly (69.57%), white (56.52%), unmarried (63.04%), and had a low level of education (56.52%). Most patients (69.57%) were in the intensive care unit and remained hospitalized for more than 30 days (76.08%). There was a significant inverse trend between Klebsiella pneumoniae and age (p = 0.045), as well as a direct linear trend between blaNDM and the annual increase in COVID-19 cases in Brazil (p = 0.050). A high probability of finding non-Klebsiella pneumoniae bacteria was observed in patients with prolonged hospital stays, independent of COVID-19 (p = 0.006) and the type of resistance genes (p = 0.020). The persistent prevalence of CRE, especially with blaKPC, underscores the urgency of effective control measures.

High frequency of acquired virulence factors in carbapenemase-producing Klebsiella pneumoniae isolates from a large German university hospital, 2013-2021.

Carbapenemase-producing Klebsiella pneumoniae (CP-Kp) isolates are a public health concern as they can cause severe hospital-acquired infections that are difficult to treat. It has recently been shown that CP-Kp can take up virulence factors from hypervirulent K. pneumoniae lineages. In this study, 109 clinical CP-Kp isolates from the University Hospital Cologne were examined for the presence of acquired virulence factors using whole-genome sequencing and phenotypic tests, and results were linked to clinical data. The virulence factor iuc was present in 18/109 of the CP-Kp isolates. Other acquired virulence factors, such as ybt, cbt, iro, rmpA/rmpA2, peg-344, and hypervirulence-associated capsule types were detected in various combinations among these isolates. The iuc-positive isolates produced OXA-232 (n = 7), OXA-48 (n = 6), OXA-48+NDM (n = 3), NDM, and KPC (each n = 1), and 7/18 isolates were resistant to ceftazidime-avibactam, colistin, and/or cefiderocol. Four isolates carried hybrid plasmids that harbored acquired virulence factors alongside the carbapenemase genes blaNDM-1/5 or blaOXA-48. In 15/18 patients, iuc-positive CP-Kp were isolated from a clinically manifest infection site. Among these, four patients had osteomyelitis, and four patients died from pneumonia with OXA-232-producing ST231 isolates, three of them as part of an outbreak. In conclusion, acquired virulence factors are frequently detected in various combinations in carbapenemase-producing K. pneumoniae isolates in Germany, warranting continuous monitoring of infections caused by these strains.

RfaH contributes to maximal colonization and full virulence of hypervirulent Klebsiella pneumoniae.

Hypervirulent K. pneumoniae (hvKp) have emerged as clinically important pathogens, posing a serious threat to human health. RfaH, a transcriptional elongation factor, has been regarded as implicated in facilitating the transcription of long virulence operons in certain bacterial species. In K. pneumoniae, RfaH plays a vital role in promoting CPS synthesis and hypermucoviscosity, as well as mediating bacterial fitness during lung infection. In this study, we aim to conduct a systematic investigation of the roles of rfaH in the survival, dissemination, and colonization of hvKp through in vitro and in vivo assays. We found that bacterial cells and colonies displayed capsule -deficient phenotypes subsequent to the deletion of rfaH in K. pneumoniae NTUH-K2044. We confirmed that rfaH is required for the synthesis of capsule and lipopolysaccharide (LPS) by positively regulating the expression of CPS and LPS gene clusters. We found that the ΔrfaH mutant led to a significantly decreased mortality of K. pneumoniae in a mouse intraperitoneal infection model. We further demonstrated that the absence of rfaH was associated with slower bacterial growth under conditions of low nutrition or iron limitation. ΔrfaH displayed reduced survival rates in the presence of human serum. Besides, the engulfment of the ΔrfaH mutant was significantly higher than that of NTUH-K2044 by macrophages in vivo, indicating an indispensable role of RfaH in the phagocytosis resistance of hvKp in mice. Both mouse intranasal and intraperitoneal infection models revealed a higher bacterial clearance rate of ΔrfaH in lungs, livers, and spleens of mice compared to its wild type, suggesting an important role of RfaH in the bacterial survival, dissemination, and colonization of hvKp in vivo. Histopathological results supported that RfaH contributes to the pathogenicity of hvKp in mice. In conclusion, our study demonstrates crucial roles of RfaH in the survival, colonization and full virulence of hvKp, which provides several implications for the development of RfaH as an antibacterial target.

Characterization of novel phage pK3-24 targeting multidrug-resistant Klebsiella pneumoniae and its therapeutic efficacy in Galleria mellonella larvae.

Klebsiella pneumoniae is a common, conditionally pathogenic bacterium that often has a multidrug-resistant phenotype, leading to failure of antibiotic therapies. It can therefore induce serious diseases, including community-acquired pneumonia and bloodstream infections. As an emerging alternative to antibiotics, phages are considered key to solving the problem of drug-resistant bacterial infections. Here, we report a novel phage, pK3-24, that mainly targets ST447 K. pneumoniae. Phage pK3-24 is a T7-like short-tailed phage with a fast adsorption capacity that forms translucent plaques with halos on bacterial lawns. The optimal multiplicity of infection (MOI) is 0.01, and the average burst size is 50 PFU/mL. Phage pK3-24 shows environmental stability, surviving at below 50°C and at pH values of 6-10. It has a double-stranded DNA genome of 40,327 bp and carries no antibiotic-resistance, virulence, or lysogeny genes. Phylogenetic analysis assigned phage pK3-24 to the genus Przondovirus as a new species. Phage pK3-24 inhibited the production of biofilm. Moreover, treatment with pK3-24 at doses with an MOI > 1 effectively reduced the mortality of Galleria mellonella larvae infected with ST447 K. pneumoniae.

Pharmacodynamic Target Attainment at Infection Site during Treatment of Post-Neurosurgical Ventriculitis Caused by Carbapenem-Resistant Klebsiella pneumoniae with Ceftazidime-Avibactam-Based Regimens: a case report.

A patient developed a post-neurosurgical ventriculitis with carbapenem-resistant Klebsiella pneumoniae and mold, initially treated with ceftazidime/avibactam and voriconazole. A Klebsiella pneumoniae carbapenemase mutation led to therapy adjustment to ceftazidime/avibactam and polymyxin B, achieving cure. Pharmacokinetic/pharmacodynamic analysis highlights effective ceftazidime/avibactam's brain penetration and bacterial clearance efficacy.

In vivo development of resistance to novel ß-lactam/ß-lactamase inhibitor combinations in KPC-producing Klebsiella pneumoniae infections: a case series.

INTRODUCTION: Understanding the dynamics that may characterize the emergence of KPC variants with resistance to novel ß-lactam/ß-lactamase inhibitor combinations (ßL/ßLICs) represents a challenge to be overcome in the appropriate use of recently introduced antibiotics. METHODS: Retrospective case series describing development of multiple resistance to novel ßL/ßLICs in patients with KPC-producing Klebsiella pneumoniae (KPC-Kp) infections treated with these drugs. Clinical-microbiological investigation and characterization of longitudinal strains by Whole-Genome Sequencing were performed. RESULTS: Four patients with KPC-Kp bloodstream infections were included. Most frequent clinical features were kidney disease, obesity, cardiac surgery as reason for admission, ICU stay, treatment with ceftazidime/avibactam, and pneumonia and/or acute kidney injury needing renal replacement therapy as KPC-Kp sepsis-associated complications. The development of resistance to ceftazidime/avibactam was observed in four longitudinal strains (three of which were co-resistant to aztreonam/avibactam and cefiderocol) following treatments with ceftazidime/avibactam (n = 3) or cefiderocol (n = 1). Resistance to meropenem/vaborbactam and imipenem/cilastatin/relebactam was observed in one case after exposure to ceftazidime/avibactam and imipenem/cilastatin/relebactam. Resistome analysis showed that resistance to novel ßL/ßLICs was related to specific mutations within blaKPC carbapenemase gene (D179Y mutation [KPC-33]; deletion Δ242-GT-243 [KPC-14]) in three longitudinal strains, while porin loss (truncated OmpK35 and OmpK36 porins) was observed in one case. CONCLUSION: Therapy with novel ßL/ßLICs or cefiderocol may lead to the selection of resistant mutants in the presence of factors influencing the achievement of PK/PD targets. KPC variants are mainly associated with resistance to ceftazidime/avibactam, and some of them (e.g. KPC-14) may also be associated with reduced susceptibility to aztreonam/avibactam and/or cefiderocol. Loss of function of the OmpK35 and OmpK36 porins appears to play a role in the development of resistance to meropenem/vaborbactam and/or imipenem/relebactam, but other mechanisms may also be involved.

Lemierre's syndrome associated-diabetic ketoacidosis in an elderly female: a case report.

BACKGROUND: The co-occurrence of Lemierre's syndrome, primarily triggered by Fusobacterium necrophorum following oropharyngeal infection, with diabetic ketoacidosis (DKA) in diabetes mellitus (DM) patients, underscores a rare but life-threatening clinical scenario. Lemierre's syndrome induced DKA is extremely rare, with only one case report in adult and no case yet reported in elderly. CASE PRESENTATION: We reported a case of a 69-year-old female who presented with DKA triggered by deep neck space infection (DNSI), leading to rapid clinical deterioration within 6 h that necessitated high flow nasal cannula (HFNC) and antibiotic administration. Laboratory findings included leukocytosis, elevated serum C-reactive protein, hyperglycemia, ketonemia, and severe metabolic acidosis. Culture of the fluid from a neck mass puncture drainage and blood were positive for Klebsiella pneumoniae. The patient was further complicated by thrombosis of the left internal jugular vein with extension to the sigmoid and a neck abscess surrounding the carotid artery sheath, consistent with Lemierre's syndrome. This condition was managed aggressively with fluid resuscitation, insulin therapy, surgical drainage, antibiotics, and anticoagulation led to a significant improvement in her condition. Following a 13-day hospitalization, there was significant clinical improvement, culminating in the patient's discharge. CONCLUSIONS: The case highlights the need for greater awareness and understanding of the interrelated and mutually promoting conditions of DKA and Lemierre's syndrome among clinicians. Early recognition and treatment are crucial to prevent mortality in such complex cases.

Development and validation of a clinical-radiomics nomogram for the early prediction of Klebsiella pneumoniae liver abscess.

BACKGROUND AND AIM: Pyogenic liver abscess (PLA) is a devastating and potentially life-threatening disease globally, with Klebsiella pneumoniae liver abscess (KPLA) being the most prevalent in Asia. This study aims to develop an effective and comprehensive nomogram combining clinical and radiomics features for early prediction of KPLA. METHODS: 255 patients with PLA from 2013 to 2023 were enrolled and randomly divided into the training and validation cohorts at a 7:3 ratio. The differences between the two cohorts of patients were assessed via univariate analysis. The radiomics features were extracted from imaging data from enhanced CT of liver abscesses. The optimal radiomics features were filtered using the independent sample t-test and least absolute shrinkage and selection operator, and a radiomics score (Rad-score) was calculated by weighting their respective coefficients. Clinically independent predictors were identified from the clinical data and combined with the Rad-score to develop a nomogram by multivariate logistic regression. The predictive performance was evaluated using the area under the receiver operating characteristic curve (AUC), calibration curve, and clinical decision curve. RESULTS: The nomogram incorporated four clinical features of diabetes mellitus, cryptogenic liver abscess, C-reactive protein level, and splenomegaly, and the Rad-score that was constructed based on seven optimal radiomics features. It had an AUC of 0.929 (95% CI, 0.894-0.964) and 0.923 (95% CI, 0.864-0.981) in the training and validation cohorts, respectively. The calibration and decision curves showed that the nomogram had good agreement and clinical applicability. CONCLUSIONS: The clinical-radiomics nomogram performed well in predicting KPLA, hopefully serving as a reference for early diagnosis of KPLA.

Ackermannviridae bacteriophage against carbapenem-resistant Klebsiella pneumoniae of capsular type 64.

Lytic bacteriophages (phages) are promising clinically viable therapeutic options against carbapenem-resistant Klebsiella pneumoniae (CRKP). In China, the predominant strains are those assigned to sequence type 11 and capsular type 64 (ST11-KL64). The emergence of phage resistance is a major bottleneck hindering effective phage therapy, requiring more new phages to provide the flexibility for creating different phage cocktails. However, the majority of phages against ST11-KL64 CRKP belong to the genus Przondovirus of the family Autographiviridae, which limits the options for constructing cocktails. We recovered a novel lytic phage of the genus Taipeivirus within the family Ackermannviridae against ST11-KL64 CRKP from a river in China. We phenotypically characterized this phage and obtained its genome sequence for analysis. This phage can inhibit the growth of ST11-KL64 CRKP for 6.5 h at a 0.1 multiplicity of infection and exhibits a narrow host range, being unable to attack CRKP strains of the other 30 capsular types. This phage carries no genes encoding antimicrobial resistance, virulence, or lysogeny. It is stable across a wide range of temperatures and pH values, making it suitable for phage therapy. Unlike other Taipeivirus phages, P01 has two tail spike proteins and a unique tail fiber protein. The distinct tail composition of this phage contributes to its activity against ST11-KL64 CRKP and its narrow host range. Taken together, we recovered a phage of a novel viral species with the potential for therapy, which expands the phage biobank against CRKP.

Isolation and characterization of pathogenic Klebsiella pneumoniae strains from lettuce: a potential source of antibiotic resistance and development of a mathematical model for ANOVA results.

Introduction: This study aimed to evaluate the prevalence of Klebsiella pneumoniae contamination in raw lettuce from Risalpur, Pakistan, and to analyze the antibiotic susceptibility profiles of the isolated strains. The presence of foodborne pathogens such as K. pneumoniae poses significant public health risks, particularly in regions with suboptimal hygiene practices and improper food handling. Methods: Lettuce samples were collected from various sources in Risalpur and screened for K. pneumoniae. Antimicrobial susceptibility testing was performed to evaluate the effectiveness of various antibiotics against the isolated strains. Statistical analyses, including ANOVA and linear regression, were conducted to assess differences in inhibition zones and to predict antibiotic effectiveness based on concentration. Results: The results revealed a significant prevalence of K. pneumoniae in the lettuce samples, highlighting the risks associated with poor hygiene, transportation, storage, and contaminated irrigation water. The isolated strains exhibited high susceptibility to gentamicin but demonstrated notable resistance to doxycycline, vancomycin, and ticarcillin. Multidrug-resistant (MDR) strains were identified. ANOVA showed significant differences in inhibition zones, and the linear regression model predicted a Zone of Inhibition based on antibiotic concentration (ß0 = 10.6667, ß1 = 0.4556). Discussion: The identification of MDR strains of K. pneumoniae underscores the urgent need for enhanced antibiotic stewardship and food safety protocols to manage foodborne pathogens. Improved hygiene practices throughout the food production and supply chain are critical to mitigate health risks and address the challenge of growing antibiotic resistance.

Molecular epidemiology of extended-spectrum beta-lactamase-producing-Klebsiella species in East Tennessee dairy cattle farms.

Introduction: The rising prevalence of Extended-Spectrum Beta-Lactamase (ESBL)-producing Klebsiella species (spp.) poses a significant threat to human and animal health and environmental safety. To address this pressing issue, a comprehensive study was undertaken to elucidate the burden and dissemination mechanisms of ESBL-Klebsiella spp. in dairy cattle farms. Methods: Fifty-seven Klebsiella species were isolated on CHROMagar™ ESBL plates and confirmed with MADLI-TOF MS and whole genome sequenced from 14 dairy farms. Results and discussion: Six families of beta-lactamase (bla) (bla CTX-M, bla SHV, bla TEM, bla OXY, bla OXA, and bla SED) were detected in ESBL-Klebsiella spp. genomes. Most (73%) of isolates had the first three types of beta-lactamase genes, with bla SHV being the most frequent, followed by bla CTX-M. Most (93%) isolates harbored two or more bla genes. The isolates were genotypically MDR, with 26 distinct types of antibiotic resistance genes (ARGs) and point mutations in gyrA, gyrB, and parC genes. The genomes also harbored 22 different plasmid replicon types, including three novel IncFII. The IncFII and Col440I plasmids were the most frequent and were associated with bla CTXM-27 and qnrB19 genes, respectively. Eighteen distinct sequence types (STs), including eight isolates with novel STs of K. pneumoniae, were detected. The most frequently occurring STs were ST353 (n = 8), ST469 (n = 6), and the novel ST7501 (n = 6). Clusters of ESBL-Klebsiella strains with identical STs, plasmids, and ARGs were detected in multiple farms, suggesting possible clonal expansion. The same ESBL variant was linked to identical plasmids in different Klebsiella STs in some farms, suggesting horizontal spread of the resistance gene. The high burden and dual spread mechanism of ESBL genes in Klebsiella species, combined with the emergence of novel sequence types, could swiftly increase the prevalence of ESBL-Klebsiella spp., posing significant risks to human, animal, and environmental health. Immediate action is needed to implement rigorous surveillance and control measures to mitigate this risk.

Association between the presence of CRISPR-Cas system genes and antibiotic resistance in Klebsiella pneumoniae isolated from patients admitted in Ahvaz teaching hospitals.

BACKGROUND: This study aims to investigate the frequency of cas1 and cas3 and CRISPR1,2,3 genes in Klebsiella pneumoniae isolates, as well as their connection with antibiotic resistance. MATERIALS AND METHODS: 106 K. pneumoniae isolates were identified by biochemical assays and PCR. The susceptibility to antibiotics was determined by Kirby-Bauer disk diffusion method. Screening of ESBLs was undertaken by using double disk diffusion and standard disk diffusion methods. The E-test and mCIM techniques was used to confirm the disc diffusion-based carbapenem resistance profiles. CRISPR-Cas system genes were identified using PCR. RESULTS: ESBL production was found in 19% of isolates. Carbapenemase production was found in 46% of the isolates. Furthermore, the bacteria were classified as multidrug (76%), extensively drug-resistant (4%), or pan-drug-resistant (2%). When CRISPR/Cas systems were present, antibiotic resistance was lower; conversely, when they were absent, resistance was higher. CONCLUSIONS: If the CRISPR/Cas modules aren't present, the bacteria can still acquire foreign DNA, including antibiotic resistance genes. K. pneumoniae isolates with a CRISPR-Cas system were less likely to carry antibiotic-resistance genes than those lacking this defense system.

Klebsiella pneumoniae infections and phage therapy.

OBJECTIVE: Carbapenem-colistin-resistant Klebsiella pneumoniae has emerged as a serious global problem. Klebsiella pneumoniae is a major culprit in healthcare settings and is responsible for septicemia, urinary tract infections, pneumonia, meningitis, burn wound and surgical site infections, and liver abscesses even in younger and healthier population worldwide. The formation of biofilm prevents antibiotics from reaching the bacteria and exerting their effector mechanism. The non-availability of therapeutic alternatives (antibiotic therapy) further complicates the scenario. However, in the era of antibiotic resistance, bacteriophage therapy emerges as a ray of hope against antibiotic-resistant bacteria. METHOD: The present review focuses on the therapeutic potential of bacteriophages as an antimicrobial agent with special reference to safety, specificity, efficacy, dosage, and dosage frequency against Pan-Drug Resistant (PDR) K. pneumoniae, both in-vitro and in-vivo (animals and human) studies. RESULT: This review highlights the perspectives therapeutic potential of bacteriophages, their impact on the host immune system, combination therapy, and bacteriophage-encoded gene product endolysin, artificial lysins (Artilysins), polysaccharide depolymerase, and peptidoglycan hydrolases. CONCLUSION: This review briefly describes the application of bacteriophage and its encoded gene products in clinical trials.

Overexpression of ß-lactamase genes (blaKPC, blaSHV) and novel CirA deficiencies contribute to decreased cefiderocol susceptibility in carbapenem-resistant Klebsiella pneumoniae before its approval in China.

Cefiderocol (FDC) is an effective antibiotic that is used to treat severe infections caused by carbapenem-resistant Klebsiella pneumoniae (CRKP). The mechanisms underlying FDC resistance and molecular epidemiology in China remain unclear. We collected 477 non-duplicate CRKP clinical isolates in central China and characterized their susceptibility to FDC, virulence genes, and sequence typing. The overall FDC susceptibility rate of CRKP was 99.2% in central China, which was higher than that in North America and Europe (96.1%), with MIC50/90 values of 1/2 mg/L. The decrease in FDC susceptibility in central China was concentrated in the ST11 CRKP-carrying virulence plasmids. Whole-genome sequencing (WGS) and quantitative reverse transcription PCR (qRT-PCR) experiments showed that serine ß-lactamases, especially highly expressed KPC and SHV, substantially decreased FDC susceptibility in four FDC non-susceptible isolates (two resistant and two intermediate isolates). Notably, different CirA deficiencies, p.E450GfsTer16 and p.E133Ter, were found in both of the resistant isolates. In contrast, global WGS data indicate that the resistance mechanisms in North America and Europe were primarily associated with NDM and KPC variants, predominantly found in ST307 and ST147. Overall, FDC exhibits excellent activity against CRKP in central China, with resistance mechanisms primarily related to high KPC and SHV expression, along with deficiencies in CirA, frequently observed in ST11. This is remarkably different from the situation in North America and Europe and will directly impact the choice of clinical interventions. Additionally, the surveillance of FDC resistance in China is imperative.

Interactions of biofilm polysaccharides produced by human infective bacteria with molecules of the quorum sensing system. A microscopy and NMR study.

Biofilms are the most common lifestyle adopted by bacterial communities where cells live embedded in a self-produced hydrated matrix. Although polysaccharides are considered essential for matrix architecture, their possible functional roles are still rather unexplored. The primary structure of polysaccharides produced by Klebsiella pneumoniae and species of the Burkholderia cepacia Complex revealed a composition rich in rhamnose. The methyl group on carbon 6 of rhamnose units lowers the polymer hydrophilicity and can form low polarity regions on the polysaccharide chains. These regions promote chain-chain interactions that contribute to the biofilm matrix stability, but may also act as binding sites for low-polarity molecules, aiding their mobility through the hydrated matrix. In particular, quorum sensing system components crucial for the biofilm life cycle often display poor solubility in water. Therefore, cis-11-methyl-2-dodecenoic acid and L-homoserine-lactones were investigated by NMR spectroscopy for their possible interaction with polysaccharides. In addition, the macromolecular morphology of the polysaccharides was assessed using atomic force and electron microscopies to define the role of Rha residues on the three-dimensional conformation of the polymer. NMR data revealed that quorum sensing components interact with Rhamnose-rich polysaccharides, and the extent of interaction depends on the specific primary structure of each polysaccharide.

Variable Presentations of Emphysematous Liver Abscesses: Experiences From Four Cases at a Tertiary Care Center.

Emphysematous liver abscesses usually present with fever and abdominal pain with radiological investigations showing air inside the abscess cavity and biochemical parameters suggestive of sepsis. This is a condition that needs urgent intervention, but it can present with variable presentations and the gas under the right dome of the diaphragm makes the diagnosis in dilemma, confusing it with hollow viscous perforation. Hereby we present a case series of variable presentations of emphysematous liver abscesses, successfully managed by timely intervention.