Genomic and phylogenetic characterization of Elizabethkingia anophelis strains: The first two cases of life-threatening infection in Japan.

OBJECTIVE: Elizabethkingia anophelis causes meningitis, bloodstream infections, and respiratory infections in immunocompromised individuals. We examined two E. anophelis strains isolated from the first life-threatening cases caused by this species in Japan to determine the phylogenetic origin and genomic features of them. METHODS: We performed whole genome-based analysis to clarify the genetic relationship for the two strains (EK0004 and EK0079) and Elizabethkingia sp. strains isolated from worldwide and to characterize the genomic features such as the prevalence of virulence- and antimicrobial resistance (AMR)-related genes. PATIENTS: A 29-year-old man with hepatosplenic T-cell lymphoma and a 52-year-old man with systemic lupus erythematosus developed fatal bacteremia and meningitis due to E. anophelis, respectively. RESULTS: Two strains, EK0004 and EK0079, were genetically different but most closely related to the strains isolated from the largest outbreak in Wisconsin, USA from 2015 to 2016, and the strain isolated from cerebrospinal fluid of a patient in Florida, USA in 1982, respectively. The two strains contained AMR-related genes such as those encoding for an extended-spectrum ß-lactamase and multiple metallo-ß-lactamases and several virulence-related genes such as capsular polysaccharide synthesis gene clusters. CONCLUSIONS: Although further functional analyses are required to understand the virulence of these clones, these finding suggests that enough caution of E. anophelis infection in immunocompromised patients is required since the number of infections by this species is increasing outside Japan.

Initial medical management in acute type A aortic dissection patients with a thrombosed false lumen in the ascending aorta combining intramural hematoma and retrograde dissection from the descending to the ascending aorta.

BACKGROUND: The feasibility of medical management for select patients with acute type A aortic dissection has been reported from a few institutions. In this study, we retrospectively investigated the safety and feasibility of our conservative approach for patients with type A aortic dissection in daily practice. METHODS: From January 2013 to December 2017, 131 consecutive patients were admitted to our institution for acute aortic dissection, including 58 patients of type A. Initial medical management was attempted in select patients who were clinically stable and had a thrombosed false lumen of the ascending aorta without ulcer-like projections in the ascending aorta. RESULTS: Except for nine patients contraindicated for surgery, urgent surgery was performed in 26 patients (SRG group), while 23 patients (MED group) were treated with the initial medical management. The maximum diameter of the ascending aorta was significantly larger in the SRG group than in the MED group. In the MED group, the heart rate and blood pressures were well-controlled at admission to the intensive-care unit, and the systolic blood pressure was further reduced at 24 h after. The in-hospital mortality rates of the MED and SRG groups were 0% and 15%, respectively. During the follow-up period, the survival rate was significantly higher in the MED group than in the SRG group, and the aortic event-free survival at one year was 80%. CONCLUSIONS: The initial medical management for select patients with a thrombosed false lumen in the ascending aorta was a safe and feasible strategy in real-world practice.

Liver abscess due to Sterigmatomyces halophilus in a boy with acute lymphoblastic leukemia.

We report the first case of liver abscess due to Sterigmatomyces halophilus. Because this pathogen grows poorly in culture medium without added salts, it was identified by sequencing analysis targeting the rRNA gene internal transcribed spacer (ITS) region. This method could be useful for pathogens that cannot be cultured using standard methods.

Intraoperative evaluation of polymorphonuclear leukocyte during second-stage revision surgery promote overdiagnosis of persistent periprosthetic joint infection.

OBJECTIVE: The aim of this study was to evaluate whether intraoperative histopathological examination could predict the risk of relapse of infection in periprosthetic joint infections (PJI). METHODS: The study included 25 patients (14 women and 11 men, with a mean age of 67.0 years (range, 37-83 years)), who had two-staged revision surgery for a PJI. Following prosthetic removal in the first stage, all patient underwent an intraoperative histopathological examination during the second stage. The patients were divided into PMNs-positive group (≥five PMNs per high-powered field) or -negative group (

A bundle that includes active surveillance, contact precaution for carriers, and cefazolin-based antimicrobial prophylaxis prevents methicillin-resistant Staphylococcus aureus infections in clean orthopedic surgery.

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) is a frequent cause of orthopedic surgical site infections (SSIs). The aim of this study was to evaluate the effect of a bundle approach in the prevention of orthopedic MRSA SSIs. MATERIAL AND METHODS: MRSA active surveillance and decolonization were performed preoperatively at our institution from July 2004 until 2007. In January 2008, a bundle approach comprising contact precautions for MRSA-positive patients and cefazolin-based antimicrobial prophylaxis (AMP) stewardship was implemented. Data on the prevalence of MRSA SSIs, antimicrobial use density, duration of AMP, and the use of an alcohol antiseptic agent (L/1,000 patient-days) were evaluated during 2 periods: July 2004-December 2007 (period A) and January 2008-December 2012 (period B). RESULTS AND DISCUSSION: The MRSA SSI rate during period B (0.97%; 19 out of 1,966) was significantly lower than that during period A (2.17%; 29 out of 1,333; P = .003). The infection rate correlated negatively with both the cefazolin antimicrobial use density (r = -0.76; P = .0002) and the use of an alcohol antiseptic agent (r = -0.68; P = .002). CONCLUSIONS: An infection-prevention bundle consisting of contact precautions for carriers and AMP stewardship in addition to active surveillance was associated with a significant decrease in the incidence of orthopedic MRSA SSIs.

Factors affecting treatment and recurrence of Clostridium difficile infections.

The antimicrobial agents vancomycin and metronidazole have been used to treat Clostridium difficile infections (CDIs). However, it remains unclear why patients are at risk of treatment failure and recurrence. Therefore, this study retrospectively examined 98 patients with CDIs who were diagnosed based on the detection of toxin-positive C. difficile to determine the risk factors affecting drug treatment responses and the recurrence of CDI. No significant difference was observed in the cure rate or dosage between the vancomycin and metronidazole groups. The 90-d mortality rate and total number of drugs associated with CDIs, including antiinfective agents used within 2 months before the detection of toxin-positive C. difficile, were significantly lower in the treatment success group than in the failure group. The total number of antiinfective agents and gastric acid-suppressive agents used during CDI therapy was also significantly lower in the success group than in the failure group. The period from the completion of CDI therapy to restarting the administration of anticancer agents and steroids was significantly longer in patients without than in patients with recurrence. These results indicate that the total number of drugs associated with CDIs should be minimized to reduce the risk of CDIs, that not only antibiotics but also gastric acid-suppressive agents should be discontinued during CDI therapy to increase therapeutic efficacy, and that the use of anticancer agents and steroids should be delayed as long as possible after patients are cured by CDI therapy to prevent recurrence.

Ramsay Hunt syndrome in a girl with acute lymphoblastic leukemia during maintenance therapy.

A 5-year-old girl with precursor B-cell acute lymphoblastic leukemia developed peripheral-type right facial palsy and very faint erythema on her right pinna during maintenance therapy. Acyclovir was started for possible zoster infection. The following day, vesicles appeared and a diagnosis of Ramsay Hunt syndrome was made. Prednisolone was started on day 5 after onset. Her facial palsy recovered within 6 months. Ramsay Hunt syndrome is a rare cause of facial palsy in patients with acute lymphoblastic leukemia, and this is the first case report. Preemptive therapy with acyclovir before the development of vesicles should help the patient recover from facial palsy.

Environmental mutagens may be implicated in the emergence of drug-resistant microorganisms.

The emergence of drug-resistant microorganisms is an important medical and social problem. Drug-resistant microorganisms are thought to grow selectively in the presence of antibiotics. Most clinically isolated drug-resistant microorganisms have mutations in the target genes for the drugs. While any of the many mutagens in the environment may cause such genetic mutations, no reports have yet described whether these mutagens can confer drug resistance to clinically important microorganisms. We investigated how environmental mutagens might be implicated in acquired resistance to antibiotics in clinically important microorganisms, which causes human diseases. We selected mutagens found in the environment, in cigarette smoke, or in drugs, and then exposed Pseudomonas aeruginosa to them. After exposure, the incidence of rifampicin- and ciprofloxacin-resistant P. aeruginosa strains markedly increased, and we found mutations in genes for the antibiotic-target molecule. These mutations were similar to those found in drug-resistant microorganisms isolated from clinical samples. Our findings show that environmental mutagens, and an anticancer drug, are capable of inducing drug-resistant P. aeruginosa similar to strains found in clinical settings.

IGFBP-4 is an inhibitor of canonical Wnt signalling required for cardiogenesis.

Insulin-like growth-factor-binding proteins (IGFBPs) bind to and modulate the actions of insulin-like growth factors (IGFs). Although some of the actions of IGFBPs have been reported to be independent of IGFs, the precise mechanisms of IGF-independent actions of IGFBPs are largely unknown. Here we report a previously unknown function for IGFBP-4 as a cardiogenic growth factor. IGFBP-4 enhanced cardiomyocyte differentiation in vitro, and knockdown of Igfbp4 attenuated cardiomyogenesis both in vitro and in vivo. The cardiogenic effect of IGFBP-4 was independent of its IGF-binding activity but was mediated by the inhibitory effect on canonical Wnt signalling. IGFBP-4 physically interacted with a Wnt receptor, Frizzled 8 (Frz8), and a Wnt co-receptor, low-density lipoprotein receptor-related protein 6 (LRP6), and inhibited the binding of Wnt3A to Frz8 and LRP6. Although IGF-independent, the cardiogenic effect of IGFBP-4 was attenuated by IGFs through IGFBP-4 sequestration. IGFBP-4 is therefore an inhibitor of the canonical Wnt signalling required for cardiogenesis and provides a molecular link between IGF signalling and Wnt signalling.

Vascular endothelial growth factor receptor-1 regulates postnatal angiogenesis through inhibition of the excessive activation of Akt.

Vascular endothelial growth factor (VEGF) binds both VEGF receptor-1 (VEGFR-1) and VEGF receptor-2 (VEGFR-2). Activation of VEGFR-2 is thought to play a major role in the regulation of endothelial function by VEGF. Recently, specific ligands for VEGFR-1 have been reported to have beneficial effects when used to treat ischemic diseases. However, the role of VEGFR-1 in angiogenesis is not fully understood. In this study, we showed that VEGFR-1 performs "fine tuning" of VEGF signaling to induce neovascularization. We examined the effects of retroviral vectors expressing a small interference RNA that targeted either the VEGFR-1 gene or the VEGFR-2 gene. Deletion of either VEGFR-1 or VEGFR-2 reduced the ability of endothelial cells to form capillaries. Deletion of VEGFR-1 markedly reduced endothelial cell proliferation and induced premature senescence of endothelial cells. In contrast, deletion of VEGFR-2 significantly impaired endothelial cell survival. When VEGFR-1 expression was blocked, VEGF constitutively activated Akt signals and thus induced endothelial cell senescence via a p53-dependent pathway. VEGFR-1(+/-) mice exhibited an increase of endothelial Akt activity and showed an impaired neovascularization in response to ischemia, and this impairment was ameliorated in VEGFR-1(+/-) Akt1(+/-) mice. These results suggest that VEGFR-1 plays a critical role in the maintenance of endothelial integrity by modulating the VEGF/Akt signaling pathway.

Proinflammatory cytokines in cerebrospinal fluid from patients with nontyphoidal Salmonella encephalopathy.

Nontyphoidal Salmonella (NTS) encephalopathy is characterized by rapidly progressive brain dysfunction that develops after NTS enteritis. The mechanism of central nervous system involvement remains unclear. We examined cerebrospinal fluids from 7 patients for cytokines and found elevated interleukin-6, interleukin-8, and monocyte chemotactic protein-1 concentrations in all the patients, suggesting that the proinflammatory cytokines are involved in the pathogenesis of NTS encephalopathy.

Critical roles of muscle-secreted angiogenic factors in therapeutic neovascularization.

The discovery of bone marrow-derived endothelial progenitors in the peripheral blood has promoted intensive studies on the potential of cell therapy for various human diseases. Accumulating evidence has suggested that implantation of bone marrow mononuclear cells effectively promotes neovascularization in ischemic tissues. It has also been reported that the implanted cells are incorporated not only into the newly formed vessels but also secrete angiogenic factors. However, the mechanism by which cell therapy improves tissue ischemia remains obscure. We enrolled 29 "no-option" patients with critical limb ischemia and treated ischemic limbs by implantation of peripheral mononuclear cells. Cell therapy using peripheral mononuclear cells was very effective for the treatment of limb ischemia, and its efficacy was associated with increases in the plasma levels of angiogenic factors, in particular interleukin-1beta (IL-1beta). We then examined an experimental model of limb ischemia using IL-1beta-deficient mice. Implantation of IL-1beta-deficient mononuclear cells improved tissue ischemia as efficiently as that of wild-type cells. Both wild-type and IL-1beta-deficient mononuclear cells increased expression of IL-1beta and thus induced angiogenic factors in muscle cells of ischemic limbs to a similar extent. In contrast, inability of muscle cells to secrete IL-1beta markedly reduces induction of angiogenic factors and impairs neovascularization by cell implantation. Implanted cells do not secret angiogenic factors sufficient for neovascularization but, instead, stimulate muscle cells to produce angiogenic factors, thereby promoting neovascularization in ischemic tissues. Further studies will allow us to develop more effective treatments for ischemic vascular disease.

Cellular senescence impairs circadian expression of clock genes in vitro and in vivo.

Circadian rhythms are regulated by a set of clock genes that form transcriptional feedback loops and generate circadian oscillation with a 24-hour cycle. Aging alters a broad spectrum of physiological, endocrine, and behavioral rhythms. Although recent evidence suggests that cellular aging contributes to various age-associated diseases, its effects on the circadian rhythms have not been examined. We report here that cellular senescence impairs circadian rhythmicity both in vitro and in vivo. Circadian expression of clock genes in serum-stimulated senescent cells was significantly weaker compared with that in young cells. Introduction of telomerase completely prevented this reduction of clock gene expression associated with senescence. Stimulation by serum activated the cAMP response element-binding protein, but the activation of this signaling pathway was significantly weaker in senescent cells. Treatment with activators of this pathway effectively restored the impaired clock gene expression of senescent cells. When young cells were implanted into young mice or old mice, the implanted cells were effectively entrained by the circadian rhythm of the recipients. In contrast, the entrainment of implanted senescent cells was markedly impaired. These results suggest that senescence decreases the ability of cells to transmit circadian signals to their clocks and that regulation of clock gene expression may be a novel strategy for the treatment of age-associated impairment of circadian rhythmicity.

G-CSF prevents cardiac remodeling after myocardial infarction by activating the Jak-Stat pathway in cardiomyocytes.

Granulocyte colony-stimulating factor (G-CSF) was reported to induce myocardial regeneration by promoting mobilization of bone marrow stem cells to the injured heart after myocardial infarction, but the precise mechanisms of the beneficial effects of G-CSF are not fully understood. Here we show that G-CSF acts directly on cardiomyocytes and promotes their survival after myocardial infarction. G-CSF receptor was expressed on cardiomyocytes and G-CSF activated the Jak/Stat pathway in cardiomyocytes. The G-CSF treatment did not affect initial infarct size at 3 d but improved cardiac function as early as 1 week after myocardial infarction. Moreover, the beneficial effects of G-CSF on cardiac function were reduced by delayed start of the treatment. G-CSF induced antiapoptotic proteins and inhibited apoptotic death of cardiomyocytes in the infarcted hearts. G-CSF also reduced apoptosis of endothelial cells and increased vascularization in the infarcted hearts, further protecting against ischemic injury. All these effects of G-CSF on infarcted hearts were abolished by overexpression of a dominant-negative mutant Stat3 protein in cardiomyocytes. These results suggest that G-CSF promotes survival of cardiac myocytes and prevents left ventricular remodeling after myocardial infarction through the functional communication between cardiomyocytes and noncardiomyocytes.

The export of coat protein from enteroaggregative Escherichia coli by a specific ATP-binding cassette transporter system.

Enteroaggregative Escherichia coli (EAEC) is an emerging enteric pathogen characterized by aggregative adherence (AA) to cultured human mucosal epithelium cells. We have recently characterized a 10.2-kDa protein, called dispersin, which is exported from the bacteria and which promotes dispersal of EAEC across the intestinal mucosa. Here, we present evidence that dispersin is exported by a putative ABC transporter complex, which is encoded by a genetic locus of the EAEC virulence plasmid pAA2. We demonstrate that the locus comprises a cluster of five genes (designated aat-PABCD), including homologs of an inner-membrane permease (AatP), an ATP-binding cassette protein (AatC) and the outer membrane protein TolC (AatA). We show that, like TolC, AatA localizes to the outer membrane independently of its ABC partner. Dispersin appears to require the Aat complex for outer membrane translocation but not for secretion across the inner membrane. We also show that, like the dispersin gene, transcription of the aat cluster is dependent on AggR, a regulator of virulence genes in EAEC. We propose that the aat cluster encodes a specialized ABC transporter, which plays a role in the pathogenesis of EAEC by transporting dispersin out of the bacterial cell.