Citrobacter koseri stimulates dendritic cells to induce IL-33 expression via abundant ATP production.

Introduction. Food allergies (FAs) occur due to intestinal immune dysfunction elicited by dysbiotic conditions. It was previously determined by us that Citrobacter species propagate in the faeces of mice with FAs and worsen allergic symptoms by inducing the allergenic cytokine IL-33. Dendritic cells can play important roles in regulation of FA responses.Hypothesis. Citrobacter species propagating in intestines of mice worsen allergic symptoms by stimulating dendritic cells to induce IL-33 expression.Aim. The aim of the present study was to analyse whether C. koseri stimulates dendritic cells to induce IL-33 expression.Methodology. IL-33 expression was evaluated in a DC2.4 mouse dendritic cell line stimulated by live or heat-inactivated C. koseri JCM1658, ATP, LPS extracted from C. koseri JCM1658 or other enterobacteria by real-time PCR. The ATP concentration and number of live bacteria in the culture supernatant were measured simultaneously.Results. Live C. koseri JCM1658 induced higher levels of IL-33 expression than other enterobacteria tested, but such a response was not elicited by heat-inactivated C. koseri JCM1658. LPS extracted from C. koseri JCM1658 did not induce IL-33 expression and suppressed live C. koseri JCM1658-induced IL-33 expression via the activation of Toll-like receptor 4 signalling. Furthermore, ATP produced by C. koseri JCM1658 stimulated dendritic cells to induce IL-33 expression by stimulating the P2X7 receptor, and LPS attenuated extracellular ATP-induced IL-33 expression. C. koseri JCM1658 was observed to proliferate more vigorously and produce more ATP than other enterobacteria.Conclusion. C. koseri acts as an allergenic bacterium through ATP production, stimulating dendritic cells to induce IL-33 expression, while LPS released from inactivated C. koseri JCM1658 attenuates this allergenicity.

Myo-inositol utilization by Citrobacter koseri promotes brain infection.

Citrobacter species are opportunistic bacterial pathogens that are implicated in both nosocomial and community-acquired infections. Among the Citrobacter species, Citrobacter koseri is often isolated from clinical material, and it can cause meningitis and brain abscesses in neonates and immunocompromised individuals, thus posing a great threat to human health. However, the virulence determinants of C. koseri remain largely unknown. Myo-inositol is an abundant carbohydrate in the environment and in certain organs of the human body, especially the brain. The C. koseri genome harbors a cluster of genes, QCQ70420.1 to QCQ70429.1 (named the Ino-cluster in this study), which encode IolBCDE, MmsA, and an ATP-binding cassette transporter. The gene cluster may be involved in the utilization of myo-inositol. To investigate the functions of the Ino-cluster in C. koseri, we constructed a mutant strain by deleting the Ino-cluster and found that the mutant could not use myo-inositol as the sole carbon source, confirming that this cluster is responsible for myo-inositol utilization. Moreover, we investigated the function of the Ino-cluster and myo-inositol utilization in C. koseri pathogenicity. Deletion of the Ino-cluster significantly impaired C. koseri colonization of the brain of infected Sprague-Dawley (SD) rats and BALB/c mice, and this increased the survival rate of the infected animals, indicating that the Ino-cluster and the ability to use myo-inositol are essential for C. koseri pathogenicity. Taken together, our findings suggest that using the Ino-cluster products, C. koseri can exploit the abundant myo-inositol in the brain as a carbon source for growth, thus promoting colonization and virulence.

Citrobacter koseri causing osteomyelitis in a diabetic foot with concomitant acute gouty arthritis successfully treated with ertapenem.

We present an elderly diabetic man with left hallux pain and drainage who was initially diagnosed with acute gouty arthritis using the diagnostic rule for acute gout and monosodium urate crystals presented on synovial fluid analysis. Further investigation with surgical debridement, plain X-ray, MRI and wound culture revealed concomitant Citrobacter koseri septic arthritis with osteomyelitis. C. koseri is considered an opportunistic infection that rarely causes musculoskeletal infections. Acute gouty arthritis and septic arthritis are rarely seen occurring concomitantly in the same joint and are often difficult to differentiate due to similar findings on exam and imaging. The present case illustrates that osteomyelitis with an opportunistic organism can present concomitantly with acute gouty arthritis, and the diagnosis of one should not exclude the other.

Identical bacterial populations colonize premature infant gut, skin, and oral microbiomes and exhibit different in situ growth rates.

The initial microbiome impacts the health and future development of premature infants. Methodological limitations have led to gaps in our understanding of the habitat range and subpopulation complexity of founding strains, as well as how different body sites support microbial growth. Here, we used metagenomics to reconstruct genomes of strains that colonized the skin, mouth, and gut of two hospitalized premature infants during the first month of life. Seven bacterial populations, considered to be identical given whole-genome average nucleotide identity of >99.9%, colonized multiple body sites, yet none were shared between infants. Gut-associated Citrobacter koseri genomes harbored 47 polymorphic sites that we used to define 10 subpopulations, one of which appeared in the gut after 1 wk but did not spread to other body sites. Differential genome coverage was used to measure bacterial population replication rates in situ. In all cases where the same bacterial population was detected in multiple body sites, replication rates were faster in mouth and skin compared to the gut. The ability of identical strains to colonize multiple body sites underscores the habit flexibility of initial colonists, whereas differences in microbial replication rates between body sites suggest differences in host control and/or resource availability. Population genomic analyses revealed microdiversity within bacterial populations, implying initial inoculation by multiple individual cells with distinct genotypes. Overall, however, the overlap of strains across body sites implies that the premature infant microbiome can exhibit very low microbial diversity.

Detection of Citrobacter koseri carrying beta-lactamase KPC-2 in a hospitalised patient, Greece, July 2011.

This report describes the detection of Citrobacter koseri carrying K. pneumoniae carbapenemase (KPC-2) isolated in July 2011 from a Greek patient, who was also colonised by a Klebsiella pneumoniae strain coproducing KPC-2 and Verona integron-encoded metallo-beta-lactamase (VIM)-1.

Abscess caused by Citrobacter koseri infection: three case reports and a literature review.

In this report, we present 3 cases of abscess caused by Citrobacter koseri. All infected patients recovered after initial empirical antibiotic treatment and percutaneous drainage of the abscess. We reviewed the literature and found 9 adult cases of C. koseri abscess. Most of these patients recovered after timely antibiotic treatment and drainage.

MyD88 is pivotal for immune recognition of Citrobacter koseri and astrocyte activation during CNS infection.

Citrobacter koseri (C. koseri) is a Gram-negative bacterium that can cause a highly aggressive form of neonatal meningitis, which often progresses to establish multi-focal brain abscesses. The roles of Toll-like receptor 4 (TLR4) and its signaling adaptor MyD88 during CNS C. koseri infection have not yet been examined, which is important since recent evidence indicates that innate immune responses are tailored towards specific pathogen classes. Here TLR4 WT (C3H/FeJ) and TLR4 mutant (C3H/HeJ) mice as well as MyD88 KO animals were infected intracerebrally with live C. koseri, resulting in meningitis and ventriculitis with accompanying brain abscess formation. MyD88 KO mice were exquisitely sensitive to C. koseri, demonstrating enhanced mortality rates and significantly elevated bacterial burdens compared to WT animals. Interestingly, although early proinflammatory mediator release (i.e. 12 h) was MyD88-dependent, a role for MyD88-independent signaling was evident at 24 h, revealing a compensatory response to CNS C. koseri infection. In contrast, TLR4 did not significantly impact bacterial burdens or proinflammatory mediator production in response to C. koseri. Similar findings were obtained with primary astrocytes, where MyD88-dependent pathways were essential for chemokine release in response to intact C. koseri, whereas TLR4 was dispensable; implicating the involvement of alternative TLRs since highly enriched astrocytes did not produce IL-1 upon bacterial exposure, which also signals via MyD88. Collectively, these findings demonstrate the importance of MyD88-dependent mechanisms in eliciting maximal proinflammatory responses, astrocyte activation, and bacterial containment during CNS C. koseri infection, as well as a late-phase MyD88-independent signaling pathway for cytokine/chemokine production.

Citrobacter koseri as a cause of early periprosthetic infection after primary total hip arthroplasty.

Periprosthetic joint infection in the acute setting is usually caused by gram-positive species and remains a major problem facing total joint surgeons. We report a case of a 53-year-old male who presented with drainage 3 weeks after primary total hip arthroplasty. Citrobacter koseri was cultured from an infected hematoma in his deep tissues. Surgical treatment included irrigation and debridement with femoral head and liner exchange. He received a 6-week course of ertapenem and is currently asymptomatic. We present C. koseri as a rare cause of acute periprosthetic infection and offer an effective treatment protocol.

Citrobacter koseri meningitis: a neurosurgical condition?

A 2-month-old girl developed meningitis, ventriculitis and brain abscess in the course of Citrobacter koseri infection. She was successfully treated with the combined use of antibiotics, intra-cavitary urokinase and surgery, thus avoiding the development of hydrocephalus and of ventricular loculation. C. koseri is a Gram-negative pathogen with a strong predilection for the neonatal brain. Brain abscesses develop in roughly 77% of cases, causing severe neurological sequels in one-half and death in one-third of patients. The authors aim to report the role of neurosurgical treatment for managing the severe complications that may arise in the course of C. koseri brain infection and the use of urokinase for preventing the development of loculated hydrocephalus.

Endogenous endophthalmitis caused by Citrobacter koseri.

Endogenous endophthalmitis occurs when organisms are hematogenously disseminated in to the eye from a distant focus of infection. The most common isolated organisms that cause endogenous endophthalmitis are Klebsiella pneumoniae and Escherichia coli. Previous reports on endophthalmitis caused by Citrobacter species are limited. We present the first case of endogenous endophthalmitis caused by Citrobacter koseri bacteremia and renal abscesses.

fliP influences Citrobacter koseri macrophage uptake, cytokine expression and brain abscess formation in the neonatal rat.

Citrobacter koseri causes neonatal meningitis frequently complicated with multiple brain abscesses. During C. koseri central nervous system infection in the neonatal rat model, previous studies have documented many bacteria-filled macrophages within the neonatal rat brain and abscesses. Previous studies have also shown that C. koseri is taken up by, survives phagolysosomal fusion and replicates in macrophages in vitro and in vivo. In this study, in order to elucidate genetic and cellular factors contributing to C. koseri persistence, a combinatory technique of differential fluorescence induction and transposon mutagenesis was employed to isolate C. koseri genes induced while inside macrophages. Several banks of mutants were subjected to a series of enrichments to select for gfp : : transposon fusion into genes that are turned off in vitro but expressed when intracellular within macrophages. Further screening identified several mutants attenuated in their recovery from macrophages compared with the wild-type. A mutation within an Escherichia coli fliP homologue caused significant attenuation in uptake and hypervirulence in vivo, resulting in death within 24 h. Furthermore, analysis of the immunoregulatory interleukin (IL)-10/IL-12 cytokine response during infection suggested that C. koseri fliP expression may alter this response. A better understanding of the bacteria-macrophage interaction at the molecular level and its contribution to brain abscess formation will assist in developing preventative and therapeutic strategies.

Citrobacter koseri brain abscess in the neonatal rat: survival and replication within human and rat macrophages.

A unique feature of Citrobacter koseri is the extremely high propensity to initiate brain abscesses during neonatal meningitis. Previous clinical reports and studies on infant rats have documented many Citrobacter-filled macrophages within the ventricles and brain abscesses. It has been hypothesized that intracellular survival and replication within macrophages may be a mechanism by which C. koseri subverts the host response and elicits chronic infection, resulting in brain abscess formation. In this study, we showed that C. koseri causes meningitis and brain abscesses in the neonatal rat model, and we utilized histology and magnetic resonance imaging technology to visualize brain abscess formation. Histology and electron microscopy (EM) revealed that macrophages (and not fibroblasts, astrocytes, oligodendrocytes, or neurons) were the primary target for long-term C. koseri infection. To better understand C. koseri pathogenesis, we have characterized the interactions of C. koseri with human macrophages. We found that C. koseri survives and replicates within macrophages in vitro and that uptake of C. koseri increases in the presence of human pooled serum in a dose-dependent manner. EM studies lend support to the hypothesis that C. koseri uses morphologically different methods of uptake to enter macrophages. FcgammaRI blocking experiments show that this receptor primarily facilitates the entry of opsonized C. koseri into macrophages. Further, confocal fluorescence microscopy demonstrates that C. koseri survives phagolysosomal fusion and that more than 90% of intracellular C. koseri organisms are colocalized within phagolysosomes. The ability of C. koseri to survive phagolysosome fusion and replicate within macrophages may contribute to the establishment of chronic central nervous system infection including brain abscesses.

Anaerobic cocci populating the deep tissues of chronic wounds impair cellular wound healing responses in vitro.

BACKGROUND: Anaerobic cocci are estimated to be present in the deep tissues of over 50% of chronic skin wounds. While the part they play in the chronicity of these wounds is uninvestigated, anaerobic cocci have previously been shown to be involved in other chronic inflammatory human conditions. METHODS: In this study the anaerobic microflora of the deep tissues of 18 patients with refractory chronic venous leg ulcers (mean age 80.3 years; mean duration > 24 months) was characterized using strict anaerobic culture conditions. The effect of the anaerobic organisms isolated from these tissues on extracellular matrix (ECM) proteolysis and cellular wound healing responses was studied using in vitro models. RESULTS: Anaerobic organisms were present in the deep tissues of 14 of 18 wounds and were principally Peptostreptococcus spp. The effects of three Peptostreptococcus spp. isolated from these wounds (P. magnus, P. vaginalis and P. asaccharolyticus) on cellular wound healing responses were compared with those of two pathogenic organisms also isolated from these wounds (Pseudomonas aeruginosa and Citrobacter diversus). While the direct ECM proteolytic activity exhibited by the Peptostreptococcus spp. was limited, they did significantly inhibit both fibroblast and keratinocyte proliferation, but only at high concentrations. However, at lower concentrations peptostreptococcal supernatants profoundly inhibited keratinocyte wound repopulation and endothelial tubule formation. The magnitude of these effects varied between strains and they were distinct from those demonstrated by Pseudomonas aeruginosa and Citrobacter diversus. CONCLUSIONS: These studies confirm the importance of anaerobic organisms in chronic wounds and demonstrate an indirect, strain-specific mechanism by which these microorganisms may play a part in mediating the chronicity of these wounds.