RESUMEN
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.
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Dictyostelium , Citometría de Flujo , Klebsiella pneumoniae , Dictyostelium/microbiología , Citometría de Flujo/métodos , Klebsiella pneumoniae/patogenicidad , Fagocitosis , Virulencia , Interacciones Huésped-Patógeno , Microscopía/métodosRESUMEN
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.
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Interacciones Huésped-Patógeno , Infecciones por Klebsiella , Klebsiella pneumoniae , Larva , Microinyecciones , Microscopía Fluorescente , Pez Cebra , Animales , Pez Cebra/microbiología , Klebsiella pneumoniae/inmunología , Microinyecciones/métodos , Larva/microbiología , Larva/inmunología , Microscopía Fluorescente/métodos , Interacciones Huésped-Patógeno/inmunología , Infecciones por Klebsiella/microbiología , Infecciones por Klebsiella/inmunología , Modelos Animales de EnfermedadRESUMEN
Multi-resistant bacteria such as Escherichia coli and Klebsiella pneumoniae are a growing threat worldwide. The spread of Carbapenemase-producing strains is particularly worrying. New antibiotics and combination therapies offer treatment options, but the development of resistant pathogens remains a major challenge.
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Antibacterianos , Farmacorresistencia Bacteriana Múltiple , Humanos , Antibacterianos/uso terapéutico , Klebsiella pneumoniae/efectos de los fármacos , Escherichia coli/efectos de los fármacos , Infecciones Bacterianas/tratamiento farmacológico , Infecciones Bacterianas/epidemiologíaRESUMEN
Carbapenem-resistant Klebsiella pneumoniae (CRKP) causes Gram-negative lung infections and fatal pneumonic sepsis for which limited therapeutic options are available. The lungs are densely innervated by nociceptor sensory neurons that mediate breathing, cough, and bronchoconstriction. The role of nociceptors in defense against Gram-negative lung pathogens is unknown. Here, we found that lung-innervating nociceptors promote CRKP pneumonia and pneumonic sepsis. Ablation of nociceptors in mice increased lung CRKP clearance, suppressed trans-alveolar dissemination of CRKP, and protected mice from hypothermia and death. Furthermore, ablation of nociceptors enhanced the recruitment of neutrophils and Ly6Chi monocytes and cytokine induction. Depletion of Ly6Chi monocytes, but not of neutrophils, abrogated lung and extrapulmonary CRKP clearance in ablated mice, suggesting that Ly6Chi monocytes are a critical cellular population to regulate pneumonic sepsis. Further, neuropeptide calcitonin gene-related peptide suppressed the induction of reactive oxygen species in Ly6Chi monocytes and their CRKP-killing abilities. Targeting nociceptor signaling could be a therapeutic approach for treating multidrug-resistant Gram-negative infection and pneumonic sepsis.
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Péptido Relacionado con Gen de Calcitonina , Carbapenémicos , Infecciones por Klebsiella , Klebsiella pneumoniae , Pulmón , Nociceptores , Sepsis , Animales , Klebsiella pneumoniae/fisiología , Ratones , Infecciones por Klebsiella/microbiología , Sepsis/metabolismo , Sepsis/microbiología , Pulmón/microbiología , Pulmón/metabolismo , Carbapenémicos/farmacología , Péptido Relacionado con Gen de Calcitonina/metabolismo , Nociceptores/metabolismo , Monocitos/metabolismo , Células Receptoras Sensoriales/metabolismo , Neutrófilos/metabolismo , Modelos Animales de Enfermedad , Antígenos Ly/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Neumonía Bacteriana/microbiología , Neumonía Bacteriana/metabolismo , Neumonía Bacteriana/patología , Ratones Endogámicos C57BLRESUMEN
BACKGROUND: Klebsiella pneumoniae (KP), responsible for acute lung injury (ALI) and inflammation of the gastrointestinal tract, is a zoonotic pathogen that poses a threat to livestock farming worldwide. Nevertheless, there is currently no validated vaccine to prevent KP infection. The development of mucosal vaccines against KP using Lactobacillus plantarum (L. plantarum) is an effective strategy. RESULTS: Firstly, the L. plantarum strains NC8-pSIP409-aCD11c' and NC8-pLc23-aCD11c were constructed via homologous recombination to express the aCD11c protein either inducibly or constitutively. Both NC8-pSIP409-aCD11c' and NC8-pLc23-aCD11c strains could enhance the adhesion and invasion of L. plantarum on bone marrow-derived dendritic cells (BMDCs), and stimulate the activation of BMDCs compared to the control strain NC8-pSIP409 in vitro. Following oral immunization of mice with NC8-pSIP409-aCD11c' and NC8-pLc23-aCD11c, the cellular, humoral, and mucosal immunity were significantly improved, as evidenced by the increased expression of CD4+ IL-4+ T cells in the spleen, IgG in serum, and secretory IgA (sIgA) in the intestinal lavage fluid (ILF). Furthermore, the protective effects of L. plantarum against inflammatory damage caused by KP infection were confirmed by assessing the bacterial loads in various tissues, lung wet/dry ratio (W/D), levels of inflammatory cytokines, and histological evaluation, which influenced T helper 17 (Th17) and regulatory T (Treg) cells in peripheral blood and lung. CONCLUSIONS: Both the inducible and constitutive L. plantarum strains NC8-pSIP409-aCD11c' and NC8-pLc23-aCD11c have been found to stimulate cellular and humoral immunity levels and alleviate the inflammatory response caused by KP infection. These findings have provided a basis for the development of a novel vaccine against KP.
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Inmunidad Celular , Infecciones por Klebsiella , Klebsiella pneumoniae , Lactobacillus plantarum , Animales , Infecciones por Klebsiella/prevención & control , Infecciones por Klebsiella/veterinaria , Infecciones por Klebsiella/inmunología , Klebsiella pneumoniae/inmunología , Ratones , Administración Oral , Femenino , Ratones Endogámicos BALB C , Vacunas Bacterianas/inmunología , Vacunas Bacterianas/administración & dosificación , Células Dendríticas/inmunología , InflamaciónRESUMEN
Alcohol use is an independent risk factor for the development of bacterial pneumonia due, in part, to impaired mucus-facilitated clearance, macrophage phagocytosis, and recruitment of neutrophils. Alcohol consumption is also known to reduce peripheral natural killer (NK) cell numbers and compromise NK cell cytolytic activity, especially NK cells with a mature phenotype. However, the role of innate lymphocytes, such as NK cells during host defense against alcohol-associated bacterial pneumonia is essentially unknown. We have previously shown that indole supplementation mitigates increases in pulmonary bacterial burden and improves pulmonary NK cell recruitment in alcohol-fed mice, which were dependent on aryl hydrocarbon receptor (AhR) signaling. Employing a binge-on-chronic alcohol-feeding model we sought to define the role and interaction of indole and NK cells during pulmonary host defense against alcohol-associated pneumonia. We demonstrate that alcohol dysregulates NK cell effector function and pulmonary recruitment via alterations in two key signaling pathways. We found that alcohol increases transforming growth factor beta (TGF-ß) signaling while suppressing AhR signaling. We further demonstrated that NK cells isolated from alcohol-fed mice have a reduced ability to kill Klebsiella pneumoniae. NK cell migratory capacity to chemokines was also significantly altered by alcohol, as NK cells isolated from alcohol-fed mice exhibited preferential migration in response to CXCR3 chemokines but exhibited reduced migration in response to CCR2, CXCR4, and CX3CR1 chemokines. Together this data suggests that alcohol disrupts NK cell-specific TGF-ß and AhR signaling pathways leading to decreased pulmonary recruitment and cytolytic activity thereby increasing susceptibility to alcohol-associated bacterial pneumonia.
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Células Asesinas Naturales , Ratones Endogámicos C57BL , Neumonía Bacteriana , Receptores de Hidrocarburo de Aril , Transducción de Señal , Animales , Células Asesinas Naturales/inmunología , Neumonía Bacteriana/inmunología , Neumonía Bacteriana/microbiología , Ratones , Receptores de Hidrocarburo de Aril/metabolismo , Pulmón/inmunología , Pulmón/microbiología , Factor de Crecimiento Transformador beta/metabolismo , Etanol , Receptores CCR2/metabolismo , Receptores CCR2/genética , Modelos Animales de Enfermedad , Indoles/farmacología , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Masculino , Klebsiella pneumoniae , Receptores CXCR3/metabolismoRESUMEN
The Gram-negative bacterium Klebsiella pneumoniae is an important human pathogen. Its treatment has been complicated by the emergence of multi-drug resistant strains. The human complement system is an important part of our innate immune response that can directly kill Gram-negative bacteria by assembling membrane attack complex (MAC) pores into the bacterial outer membrane. To resist this attack, Gram-negative bacteria can modify their lipopolysaccharide (LPS). Especially the decoration of the LPS outer core with the O-antigen polysaccharide has been linked to increased bacterial survival in serum, but not studied in detail. In this study, we characterized various clinical Klebsiella pneumoniae isolates and show that expression of the LPS O1-antigen correlates with resistance to complement-mediated killing. Mechanistic data reveal that the O1-antigen does not inhibit C3b deposition and C5 conversion. In contrast, we see more efficient formation of C5a, and deposition of C6 and C9 when an O-antigen is present. Further downstream analyses revealed that the O1-antigen prevents correct insertion and polymerization of the final MAC component C9 into the bacterial membrane. Altogether, we show that the LPS O1-antigen is a key determining factor for complement resistance by K. pneumoniae and provide insights into the molecular basis of O1-mediated MAC evasion.
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Complemento C9 , Klebsiella pneumoniae , Antígenos O , Klebsiella pneumoniae/inmunología , Antígenos O/inmunología , Antígenos O/metabolismo , Humanos , Complemento C9/metabolismo , Complemento C9/inmunología , Complejo de Ataque a Membrana del Sistema Complemento/metabolismo , Complejo de Ataque a Membrana del Sistema Complemento/inmunología , Lipopolisacáridos , Polimerizacion , Infecciones por Klebsiella/inmunología , Infecciones por Klebsiella/microbiología , Complemento C3b/metabolismo , Complemento C3b/inmunologíaRESUMEN
It remains that intracranial infection has an alarming mortality and morbidity. Klebsiella pneumoniae (KP) have increasingly been isolated in ventriculitis and meningitis episodes. Intracranial infections caused by carbapenem-resistant Klebsiella pneumoniae (CRKP) account for high mortality. To understand its clinical impact and related risk factors accurately are crucial in the management of bacterial intracranial infection. The retrospective study aimed to delineate the clinical risk of death from intracranial infection and analyze the risk factors. A total of 176 Klebsiella pneumoniae intracranial infectious patients were available to divide into CRKP group and carbapenem-susceptive Klebsiella Pneumoniae (CSKP) group. We performed survival analysis and estimate the time-varying effects of CRKP and CSKP infection on 30-day mortality. Infectious patients caused by CSKP was associated with lower mortality than CRKP group. The risk factors associated with death from intracranial infection caused by Klebsiella pneumoniae included SOFA scores, ventilator therapy, CRKP, and heart failure. Longer hospital stays are independently associated with lower mortality rates. Intracranial infection caused by CRKP was associated with excess mortality. Complex comorbidities mean higher mortality. Active supportive treatment is required for complicated patients with intracranial infections caused by carbapenem-resistant Klebsiella pneumoniae.
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Carbapenémicos , Infecciones por Klebsiella , Klebsiella pneumoniae , Humanos , Masculino , Femenino , Klebsiella pneumoniae/efectos de los fármacos , Klebsiella pneumoniae/aislamiento & purificación , Infecciones por Klebsiella/mortalidad , Infecciones por Klebsiella/tratamiento farmacológico , Infecciones por Klebsiella/microbiología , Persona de Mediana Edad , Factores de Riesgo , Estudios Retrospectivos , Carbapenémicos/farmacología , Carbapenémicos/uso terapéutico , Anciano , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Adulto , Enterobacteriaceae Resistentes a los Carbapenémicos/efectos de los fármacos , Enterobacteriaceae Resistentes a los Carbapenémicos/aislamiento & purificaciónRESUMEN
While conducting genomic surveillance of carbapenemase-producing Enterobacteriaceae (CPE) from patient colonisation and clinical infections at Birmingham's Queen Elizabeth Hospital (QE), we identified an N-type plasmid lineage, pQEB1, carrying several antibiotic resistance genes, including the carbapenemase gene bla KPC-2. The pQEB1 lineage is concerning due to its conferral of multidrug resistance, its host range and apparent transmissibility, and its potential for acquiring further resistance genes. Representatives of pQEB1 were found in three sequence types (STs) of Citrobacter freundii, two STs of Enterobacter cloacae, and three species of Klebsiella. Hosts of pQEB1 were isolated from 11 different patients who stayed in various wards throughout the hospital complex over a 13 month period from January 2023 to February 2024. At present, the only representatives of the pQEB1 lineage in GenBank were carried by an Enterobacter hormaechei isolated from a blood sample at the QE in 2016 and a Klebsiella pneumoniae isolated from a urine sample at University Hospitals Coventry and Warwickshire (UHCW) in May 2023. The UHCW patient had been treated at the QE. Long-read whole-genome sequencing was performed on Oxford Nanopore R10.4.1 flow cells, facilitating comparison of complete plasmid sequences. We identified structural variants of pQEB1 and defined the molecular events responsible for them. These have included IS26-mediated inversions and acquisitions of multiple insertion sequences and transposons, including carriers of mercury or arsenic resistance genes. We found that a particular inversion variant of pQEB1 was strongly associated with the QE Liver speciality after appearing in November 2023, but was found in different specialities and wards in January/February 2024. That variant has so far been seen in five different bacterial hosts from six patients, consistent with recent and ongoing inter-host and inter-patient transmission of pQEB1 in this hospital setting.
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Brotes de Enfermedades , Plásmidos , beta-Lactamasas , Humanos , Plásmidos/genética , beta-Lactamasas/genética , Infecciones por Enterobacteriaceae/microbiología , Infecciones por Enterobacteriaceae/epidemiología , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/aislamiento & purificación , Klebsiella pneumoniae/efectos de los fármacos , Proteínas Bacterianas/genética , Enterobacter cloacae/genética , Enterobacter cloacae/aislamiento & purificación , Enterobacter cloacae/efectos de los fármacos , Farmacorresistencia Bacteriana Múltiple/genética , Infección Hospitalaria/microbiología , Antibacterianos/farmacología , Citrobacter freundii/genética , Citrobacter freundii/aislamiento & purificación , Enterobacteriaceae Resistentes a los Carbapenémicos/genética , Enterobacteriaceae Resistentes a los Carbapenémicos/aislamiento & purificación , Hospitales , EnterobacterRESUMEN
Background: Klebsiella pneumoniae is a major cause of hospital-acquired infections (HAIs), primarily spread through environmental contamination in hospitals. The effectiveness of current chemical disinfectants is waning due to emerging resistance, which poses environmental hazards and fosters new resistance in pathogens. Developing environmentally friendly and effective disinfectants against multidrug-resistant organisms is increasingly important. Methods: This study developed a bacteriophage cocktail targeting two common carbapenem-resistant Klebsiella pneumoniae (CRKP) strains, ST11 KL47 and ST11 KL64. The cocktail was used as an adjunctive disinfectant in a hospital's respiratory intensive care unit (RICU) via ultrasonic nebulization. Digital PCR was used to quantify CRKP levels post-intervention. The microbial community composition was analyzed via 16S rRNA sequencing to assess the intervention's impact on overall diversity. Results: The phage cocktail significantly reduced CRKP levels within the first 24 hours post-treatment. While a slight increase in pathogen levels was observed after 24 hours, they remained significantly lower than those treated with conventional disinfectants. 16S rRNA sequencing showed a decrease in the target pathogens' relative abundance, while overall species diversity remained stable, confirming that phages selectively target CRKP without disrupting ecological balance. Discussion: The findings highlight the efficacy and safety of phage-based biocleaners as a sustainable alternative to conventional disinfectants. Phages selectively reduce multidrug-resistant pathogens while preserving microbial diversity, making them a promising tool for infection control.
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Bacteriófagos , Descontaminación , Unidades de Cuidados Intensivos , Klebsiella pneumoniae , ARN Ribosómico 16S , ARN Ribosómico 16S/genética , Klebsiella pneumoniae/virología , Klebsiella pneumoniae/genética , Descontaminación/métodos , Bacteriófagos/genética , Humanos , Reacción en Cadena de la Polimerasa/métodos , Infección Hospitalaria/prevención & control , Infección Hospitalaria/microbiología , Desinfectantes/farmacología , Infecciones por Klebsiella/prevención & control , Infecciones por Klebsiella/microbiología , Análisis de Secuencia de ADNRESUMEN
Antibody-dependent complement activation plays a key role in the natural human immune response to infections. Currently, the understanding of which antibody-antigen combinations drive a potent complement response on bacteria is limited. Here, we develop an antigen-agnostic approach to stain and single-cell sort human IgG memory B cells recognizing intact bacterial cells, keeping surface antigens in their natural context. With this method we successfully identified 29 antibodies against K. pneumoniae, a dominant cause of hospital-acquired infections with increasing antibiotic resistance. Combining genetic tools and functional analyses, we reveal that the capacity of antibodies to activate complement on K. pneumoniae critically depends on their antigenic target. Furthermore, we find that antibody combinations can synergistically activate complement on K. pneumoniae by strengthening each other's binding in an Fc-independent manner. Understanding the molecular basis of effective complement activation by antibody combinations to mimic a polyclonal response could accelerate the development of antibody-based therapies against problematic infections.
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Anticuerpos Antibacterianos , Activación de Complemento , Inmunoglobulina G , Klebsiella pneumoniae , Humanos , Activación de Complemento/inmunología , Anticuerpos Antibacterianos/inmunología , Klebsiella pneumoniae/inmunología , Inmunoglobulina G/inmunología , Linfocitos B/inmunología , Células B de Memoria/inmunologíaRESUMEN
BACKGROUND: Sophorolipids are glycolipid biosurfactants with potential antibacterial, antifungal, and anticancer applications, rendering them promising for research. Therefore, this study hypothesizes that sophorolipids may have a notable impact on disrupting membrane integrity and triggering the production of reactive oxygen species, ultimately resulting in the eradication of pathogenic microbes. RESULTS: The current study resulted in the isolation of two Metschnikowia novel yeast strains. Sophorolipids production from these strains reached maximum yields of 23.24 g/l and 21.75 g/l, respectively, at the bioreactors level. Biosurfactants sophorolipids were characterized using FTIR and LC-MS techniques and found to be a mixture of acidic and lactonic forms with molecular weights of m/z 678 and 700. Our research elucidated sophorolipids' mechanism in disrupting bacterial and fungal membranes through ROS generation, confirmed by transmission electron microscopy and FACS analysis. The results showed that these compounds disrupted the membrane integrity and induced ROS production, leading to cell death in Klebsiella pneumoniae and Fusarium solani. In addition, the anticancer properties of sophorolipids were investigated on the A549 lung cancer cell line and found that sophorolipid-11D (SL-11D) and sophorolipid-11X (SL-11X) disrupted the actin cytoskeleton, as evidenced by immunofluorescence microscopy. The A549 cells were stained with Acridine orange/Ethidium bromide, which showed that they underwent necrosis. This was confirmed by flow cytometric analysis using Annexin/PI staining. The SL-11D and SL-11X molecules exhibited low levels of haemolytic activity and in-vitro cytotoxicity in HEK293, Caco-2, and L929 cell lines. CONCLUSION: In this work, novel yeast species CIG-11DT and CIG-11XT, isolated from the bee's gut, produce significant yields of sophorolipids without needing secondary oil sources, indicating a more economical production method. Our research shows that sophorolipids disrupt bacterial and fungal membranes via ROS production. They suggest they may act as chemo-preventive agents by inducing apoptosis in lung cancer cells, offering the potential for enhancing anticancer therapies.
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Antifúngicos , Antineoplásicos , Metschnikowia , Estrés Oxidativo , Especies Reactivas de Oxígeno , Tensoactivos , Antifúngicos/farmacología , Antifúngicos/química , Antifúngicos/metabolismo , Humanos , Tensoactivos/farmacología , Tensoactivos/metabolismo , Tensoactivos/química , Estrés Oxidativo/efectos de los fármacos , Antineoplásicos/farmacología , Especies Reactivas de Oxígeno/metabolismo , Células A549 , Metschnikowia/metabolismo , Metschnikowia/efectos de los fármacos , Fusarium/efectos de los fármacos , Fusarium/metabolismo , Klebsiella pneumoniae/efectos de los fármacos , Glucolípidos/farmacología , Glucolípidos/metabolismo , Pruebas de Sensibilidad Microbiana , Ácidos OléicosRESUMEN
The increase of antibiotic-resistant bacteria has become a global health emergency and the need to explore alternative therapeutic options arises. Phage therapy uses bacteriophages to target specific bacterial strains. Phages are highly specific and can target resistant bacteria. Currently, research in this regard is focused on ensuring reliability and safety to bring this tool into clinical practice. The first step is to conduct comprehensive preclinical research. In this work, we present two novel bacteriophages vB_Kpn_F13 and vB_Kpn_F14 isolated against clinical carbapenem-resistant Klebsiella pneumoniae strains obtained from hospital sewage. Multiple studies in vitro were conducted, such as sequencing, electron microscopy, stability, host range infectivity, planktonic effect and biofilm inhibition in order to discover their ability to be used against carbapenem-resistant K. pneumoniae pathogens causing difficult-to-treat infections.
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Bacteriófagos , Biopelículas , Enterobacteriaceae Resistentes a los Carbapenémicos , Carbapenémicos , Infecciones por Klebsiella , Klebsiella pneumoniae , Terapia de Fagos , Klebsiella pneumoniae/virología , Klebsiella pneumoniae/efectos de los fármacos , Bacteriófagos/aislamiento & purificación , Bacteriófagos/fisiología , Bacteriófagos/genética , Enterobacteriaceae Resistentes a los Carbapenémicos/aislamiento & purificación , Enterobacteriaceae Resistentes a los Carbapenémicos/virología , Infecciones por Klebsiella/microbiología , Infecciones por Klebsiella/terapia , Carbapenémicos/farmacología , Biopelículas/crecimiento & desarrollo , Biopelículas/efectos de los fármacos , Humanos , Especificidad del Huésped , Aguas del Alcantarillado/virología , Aguas del Alcantarillado/microbiología , Antibacterianos/farmacología , Genoma Viral , Pruebas de Sensibilidad MicrobianaRESUMEN
There have been increasing reports of Klebsiella pneumoniae resistant to ß-lactam antibiotics. This study aimed to determine the prevalence of some selected carbapenemase genes among clinical isolates of Klebsiella pneumoniae recovered from patients attending a private tertiary hospital in Southwestern Nigeria. The study was conducted over two months (February-March 2024). A total of 50 clinical isolates of Klebsiella pneumoniae from different clinical specimens were obtained from the Medical Microbiology Department, Babcock University Teaching Hospital (BUTH). The clinical isolates were then characterized using standard microbiological procedures and were tested for susceptibility to meropenem and other classes of antibiotics according to Clinical and Laboratory Standards Institute (CLSI) guidelines. Polymerase Chain Reaction (PCR) detection for OXA-48 and NDM-1 carbapenemase genes was performed on the 50 clinical isolates. PCR analysis showed that 9 (18%) clinical isolates were positive for the OXA-48 gene, 22 (44%) were positive for the NDM-1 gene, 4 (8%) possessed both the OXA-48 and NDM-1 genes, and 23 (46%) possessed neither the OXA-48 nor NDM-1 genes. Antibiotic Susceptibility Testing (AST) revealed that all the clinical isolates were resistant to meropenem. In conclusion, this study demonstrates the presence of OXA-48 and NDM-1 genes in clinical isolates of Klebsiella pneumoniae recovered from patients attending a private tertiary hospital in Southwestern Nigeria, highlighting the role of ESBL (extended-spectrum beta-lactamase) as a major resistance mechanism alongside other mechanisms. Population-based surveillance programs should be implemented to monitor the prevalence and epidemiology of Klebsiella pneumoniae infections at the community level, facilitating early detection of outbreaks and identification of emerging antimicrobial resistance patterns. CORE TIP: This study highlights the significant prevalence of NDM-1 and OXA-48 carbapenemase genes among Klebsiella pneumoniae clinical isolates in a private tertiary hospital in Southwestern Nigeria, with 44% and 18% of isolates harboring these genes, respectively. Notably, 46% of isolates were resistant to carbapenems despite lacking these genes, suggesting alternative resistance mechanisms. The findings underscore the urgent need for enhanced surveillance, infection control measures, and antibiotic stewardship programs to combat the spread of multidrug-resistant Klebsiella pneumoniae in healthcare settings.
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Antibacterianos , Proteínas Bacterianas , Infecciones por Klebsiella , Klebsiella pneumoniae , Pruebas de Sensibilidad Microbiana , Centros de Atención Terciaria , beta-Lactamasas , beta-Lactamasas/genética , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/aislamiento & purificación , Klebsiella pneumoniae/efectos de los fármacos , Klebsiella pneumoniae/enzimología , Humanos , Centros de Atención Terciaria/estadística & datos numéricos , Infecciones por Klebsiella/microbiología , Infecciones por Klebsiella/epidemiología , Nigeria/epidemiología , Antibacterianos/farmacología , Proteínas Bacterianas/genética , Masculino , Femenino , Persona de Mediana Edad , Adulto , Anciano , Adulto JovenRESUMEN
BACKGROUND: Extended-spectrum ß-lactamase -producing Enterobacterales (ESBL-E) are important zoonotic pathogens that can cause serious clinical infections, also in horses. Preventing the spread of ESBL-E, especially in the equine hospital environment, is key to reducing the number of difficult-to-treat infections. Estimating the local prevalence of ESBL-E in horses is crucial to establish targeted infection control programs at equine hospitals. We conducted a prevalence and risk factor study in equine patients on admission to an equine teaching hospital in Finland through a rectal ESBL-E screening specimen of the horse and a questionnaire. RESULTS: The prevalence of ESBL-E in admitted horses was 3% (5/161, 95% CI 1-7%); none of the tested factors remained statistically significant in multivariate analysis, although antimicrobial treatment within three months was borderline significant (p = 0.052). Extended-spectrum ß-lactamase -producing Klebsiella pneumoniae ST6179:CTX-M-15 was detected in three horses using whole-genome sequencing, which in combination with patient records suggested nosocomial transmission. Escherichia coli isolates were ST1250:CTX-M-1 (n = 1), ST1079:CTX-M-1 (n = 1), and ST1245:CTX-M-14 (n = 1). Multiple virulence genes were detected in the ESBL-E isolates. In the ESBL-E positive horses enrolled in a one-year follow-up study, ESBL-E were unlikely to be isolated in rectal screening specimens after the initial positive specimen. CONCLUSIONS: The prevalence of ESBL-E in horses visiting a veterinary teaching hospital in Finland is low, indicating an overall low prevalence estimate in the country's equine population. No statistically significant risk factors were identified, likely due to the low number of cases. The duration of ESBL-E carriage is likely to be very short in horses.
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Infecciones por Enterobacteriaceae , Enfermedades de los Caballos , Hospitales Veterinarios , beta-Lactamasas , Animales , Caballos , Enfermedades de los Caballos/microbiología , Enfermedades de los Caballos/epidemiología , beta-Lactamasas/metabolismo , beta-Lactamasas/genética , Prevalencia , Factores de Riesgo , Finlandia/epidemiología , Infecciones por Enterobacteriaceae/veterinaria , Infecciones por Enterobacteriaceae/epidemiología , Infecciones por Enterobacteriaceae/microbiología , Masculino , Femenino , Klebsiella pneumoniae/enzimología , Klebsiella pneumoniae/efectos de los fármacos , Klebsiella pneumoniae/aislamiento & purificación , Infección Hospitalaria/epidemiología , Infección Hospitalaria/veterinaria , Infección Hospitalaria/microbiología , Escherichia coli/enzimología , Escherichia coli/aislamiento & purificación , Enterobacteriaceae/enzimología , Enterobacteriaceae/aislamiento & purificación , Enterobacteriaceae/efectos de los fármacos , Antibacterianos/farmacologíaRESUMEN
Virulence factor genes (VFGs) play pivotal roles in bacterial infections and have been identified within the human gut microbiota. However, their involvement in chronic diseases remains poorly understood. Here, we establish an expanded VFG database (VFDB 2.0) consisting of 62,332 nonredundant orthologues and alleles of VFGs using species-specific average nucleotide identity ( https://github.com/Wanting-Dong/MetaVF_toolkit/tree/main/databases ). We further develop the MetaVF toolkit, facilitating the precise identification of pathobiont-carried VFGs at the species level. A thorough characterization of VFGs for 5452 commensal isolates from healthy individuals reveals that only 11 of 301 species harbour these factors. Further analyses of VFGs within the gut microbiomes of nine chronic diseases reveal both common and disease-specific VFG features. Notably, in type 2 diabetes patients, long HiFi sequencing confirms that shared VF features are carried by pathobiont strains of Escherichia coli and Klebsiella pneumoniae. These findings underscore the critical importance of identifying and understanding VFGs in microbiome-associated diseases.
Asunto(s)
Microbioma Gastrointestinal , Factores de Virulencia , Humanos , Factores de Virulencia/genética , Enfermedad Crónica , Microbioma Gastrointestinal/genética , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/patogenicidad , Klebsiella pneumoniae/aislamiento & purificación , Diabetes Mellitus Tipo 2/microbiología , Diabetes Mellitus Tipo 2/genética , Escherichia coli/genética , Escherichia coli/patogenicidad , Escherichia coli/aislamiento & purificación , Bacterias/genética , Bacterias/clasificación , Bacterias/aislamiento & purificación , Bacterias/patogenicidad , Bases de Datos Genéticas , Infecciones Bacterianas/microbiologíaRESUMEN
Current data on fosfomycin usage in children are limited. We present data on the clinical use of intravenous (IV) fosfomycin in children. Hospitalized patients who received ≥3 days of IV fosfomycin between April 2021 and March 2023 were analyzed retrospectively. Forty-three episodes of infection in 39 patients were evaluated. The mean age of the patients was 5.35 (10 days to 17.5 years) years, and 54% were male. Infections were hospital-acquired in 79% of the episodes. Indications for fosfomycin were urinary tract infection (35%), bacteremia (32.6%), catheter-related bloodstream infection (16.3%), soft tissue infection (4.7%), sepsis (4.7%), surgical site infection (2.3%), burn infection (2.3%), and pneumonia (2.3%). Klebsiella pneumoniae was identified in 46.5% of the episodes, and a pan-drug or extensive drug resistance was detected in 75% of them. Carbapenem was used before fosfomycin at significantly higher rates in K. pneumoniae episodes (P = .006). Most (88.5%) patients received fosfomycin as a combination therapy. Culture negativity was achieved in 80% of episodes within a median treatment period of 3 (2-22) days, which was significantly shorter in K. pneumoniae episodes (P < .001). Treatment-related side effects were seen in 9.3% of the episodes. Side effects were significant after 3 weeks of treatment (P = .013). The unresponsivity rate to fosfomycin was 23.3%. Nine (21%) of the patients who were followed up in the intensive care units mainly died because of sepsis (56%). IV fosfomycin is an effective agent in treating severe pediatric infections caused by resistant microorganisms. Fosfomycin can be used in various indications and is generally safe for children.
Asunto(s)
Administración Intravenosa , Antibacterianos , Bacteriemia , Fosfomicina , Humanos , Fosfomicina/administración & dosificación , Fosfomicina/uso terapéutico , Masculino , Femenino , Antibacterianos/administración & dosificación , Antibacterianos/uso terapéutico , Niño , Estudios Retrospectivos , Turquía , Lactante , Adolescente , Preescolar , Resultado del Tratamiento , Bacteriemia/tratamiento farmacológico , Recién Nacido , Klebsiella pneumoniae/efectos de los fármacos , Klebsiella pneumoniae/aislamiento & purificación , Infección Hospitalaria/tratamiento farmacológico , Sepsis/tratamiento farmacológico , Infecciones Urinarias/tratamiento farmacológico , Infecciones por Klebsiella/tratamiento farmacológicoRESUMEN
BACKGROUND: Pyogenic liver abscess (PLA) caused by Klebsiella pneumoniae can vary in severity, and several risk factors for the development of organ dysfunction in PLA have been implicated. However, few studies to date have explored the most common risk factors for clinical severity. METHODS: We conducted a study on patients with PLA caused by Klebsiella pneumoniae between February 2013 and December 2022.Using logistic regression analysis, we sought to identify factors associated with positive blood culture, septic shock, and intensive care unit (ICU) admission. RESULTS: After included 200 patients, we found that an elevated procalcitonin (PCT) level (p = 0.03), higher glucose level (p = 0.03), and lower total cholesterol (TC) level (p = 0.01) were associated with a higher likelihood of positive blood bacteriological culture. Additionally, an increased PCT level (p = 0.02) and lower TC level (p < 0.01) were associated with an elevated risk of septic shock. Furthermore, a higher PCT level (p < 0.01) was associated with a higher probability of ICU admission. CONCLUSION: In patients with PLA caused by Klebsiella pneumoniae, the PCT, glucose, and TC levels were found to be associated with positive blood culture, septic shock, and ICU admission.
Asunto(s)
Colesterol , Unidades de Cuidados Intensivos , Infecciones por Klebsiella , Klebsiella pneumoniae , Absceso Piógeno Hepático , Choque Séptico , Humanos , Absceso Piógeno Hepático/microbiología , Absceso Piógeno Hepático/sangre , Klebsiella pneumoniae/aislamiento & purificación , Masculino , Factores de Riesgo , Femenino , Infecciones por Klebsiella/complicaciones , Infecciones por Klebsiella/diagnóstico , Infecciones por Klebsiella/epidemiología , Persona de Mediana Edad , Choque Séptico/microbiología , Choque Séptico/sangre , Pronóstico , Anciano , Unidades de Cuidados Intensivos/estadística & datos numéricos , Colesterol/sangre , Polipéptido alfa Relacionado con Calcitonina/sangre , Glucemia/metabolismo , Glucemia/análisis , Estudios Retrospectivos , Modelos Logísticos , Índice de Severidad de la EnfermedadRESUMEN
Despite extensive knowledge on phage resistance at bacterium level, the resistance of bacterial communities is still not well-understood. Given its ubiquity, it is essential to understand resistance at the community level. We performed quantitative investigations on the dynamics of phage infection in Klebsiella pneumoniae biofilms. We found that the biofilms quickly developed resistance and resumed growth. Instead of mutations, the resistance was caused by unassembled phage tail fibers released by the phage-lysed bacteria. The tail fibers degraded the bacterial capsule essential for infection and induced spreading of capsule loss in the biofilm, and tuning tail fiber and capsule levels altered the resistance. Latent infections sustained in the biofilm despite resistance, allowing stable phage-bacteria coexistence. Last, we showed that the resistance exposed vulnerabilities in the biofilm. Our findings indicate that phage lysate plays important roles in shaping phage-biofilm interactions and open more dimensions for the rational design of strategies to counter bacteria with phage.
Asunto(s)
Bacteriófagos , Biopelículas , Klebsiella pneumoniae , Biopelículas/crecimiento & desarrollo , Bacteriófagos/fisiología , Klebsiella pneumoniae/virología , Klebsiella pneumoniae/fisiología , Cápsulas Bacterianas/metabolismo , MutaciónRESUMEN
The murine model is invaluable for studying intricate interactions among gut microbes; hosts; and diseases. However; the impact of genetic variations in the murine microbiome; especially in disease contexts such as Klebsiella pneumoniae (Kp) infection; still needs to be explored. Kp; an opportunistic global pathogen; is becoming increasingly prevalent in regions like Asia; especially China. This study explored the role of the gut microbiota during Kp infection using mouse model; including wild-type and rpoS mutants of Kp138; KpC4; and KpE4 from human; maize; and ditch water; respectively. Under stress conditions; RpoS reconfigures global gene expression in bacteria; shifting the cells from active growth to survival mode. Our study examined notable differences in microbiome composition; finding that Lactobacillus and Klebsiella (particularly in WKp138) were the most abundant genera in mice guts at the genus level in all wild-type treated mice. In contrast; Firmicutes were predominant in the healthy control mice. Furthermore; Clostridium was the dominant genus in all mutants; mainly in ∆KpC4; and was absent in wild-type treated mice. Differential abundance analysis identified that these candidate taxa potentially influence disease progression and pathogen virulence. Functional prediction analysis showed that most bacterial groups were functionally involved in biosynthesis; precursor metabolites; degradation; energy generation; and metabolic cluster formation. These findings challenge the conventional understanding and highlight the need for nuanced interpretations in murine studies. Additionally; this study sheds light on microbiome-immune interactions in K. pneumoniae infection and proposes new potential therapeutic strategies.