Endemic fungal infections in solid organ and hematopoietic cell transplant recipients enrolled in the Transplant-Associated Infection Surveillance Network (TRANSNET).

BACKGROUND: Invasive fungal infections are a major cause of morbidity and mortality among solid organ transplant (SOT) and hematopoietic cell transplant (HCT) recipients, but few data have been reported on the epidemiology of endemic fungal infections in these populations. METHODS: Fifteen institutions belonging to the Transplant-Associated Infection Surveillance Network prospectively enrolled SOT and HCT recipients with histoplasmosis, blastomycosis, or coccidioidomycosis occurring between March 2001 and March 2006. RESULTS: A total of 70 patients (64 SOT recipients and 6 HCT recipients) had infection with an endemic mycosis, including 52 with histoplasmosis, 9 with blastomycosis, and 9 with coccidioidomycosis. The 12-month cumulative incidence rate among SOT recipients for histoplasmosis was 0.102%. Occurrence of infection was bimodal; 28 (40%) infections occurred in the first 6 months post transplantation, and 24 (34%) occurred between 2 and 11 years post transplantation. Three patients were documented to have acquired infection from the donor organ. Seven SOT recipients with histoplasmosis and 3 with coccidioidomycosis died (16%); no HCT recipient died. CONCLUSIONS: This 5-year multicenter prospective surveillance study found that endemic mycoses occur uncommonly in SOT and HCT recipients, and that the period at risk extends for years after transplantation.

TLR9 KO mice, haplotypes and CPG indices in Chlamydia trachomatis infection.

Previous studies have investigated the role of Toll-like receptor (TLR)2 and TLR4 in susceptibility to and severity of Chlamydia trachomatis infections. In this study we employ a unique integrated approach to study the role of the intracellular CpG DNA receptor: we use a murine knockout (KO) model to assess TLR9 relevance, study human TLR9 genotypes and haplotypes in sexually transmitted disease (STD) patients and subfertile women with or without tubal pathology and use in silico TLR9 CpG index calculations to assess potential immunostimulatory properties of the Chlamydia bacterium. Although no significant differences in the course of initial infections were observed between KO mice and wild-type mice the TLR9 KO mice showed a significant level of protection upon reinfection (P = 0.02). We did not observe significant differences in genotype frequencies between C. trachomatis-positive and C. trachomatisnegative women (STD patients). However, haplotype analyses revealed a trend between C. trachomatis-positive and C. trachomatis-negative women in the carriage of haplotype IV (P = 0.061; OR: 2.6; 95% CI: 1.0-6.8). In women with subfertility, odds ratios between 2 and 3 were found for tubal pathology risk, but they did not reach significance due to cohort size limitations. Finally, CpG sequence analysis showed mildly immunostimulatory properties for the genomic sequences of Chlamydia serovars B and D. Based on the murine model, human immunogenetic studies and in silico CpG index analyses, TLR9 seems to play a modest role in C. trachomatis infections. Extension of the human cohorts is necessary to significantly prove the effect in humans.

TLR4 in Chlamydia trachomatis infections: knockout mice, STD patients and women with tubal factor subfertility.

Chlamydia trachomatis is the most prevalent sexually transmitted bacterium in the world with almost 100 million new cases each year, some of which will develop tubal pathology. Clear differences in its clinical course of infections have been observed, and recently it has been shown that 40% is based on host genetic factors. We used an integrated approach based on infection of Toll-like receptor 4 (TLR4) knockout mice and immunogenetic analysis of female sexually transmitted disease (STD) patients (susceptibility) and women with C. trachomatis-associated tubal factor subfertility (severity). The results in TLR4 knockout mice suggest that the protection against reinfection is more solid in normal as compared to the TLR4-deficient mice. In humans the functional TLR4 single nucleotide polymorphism studied was not involved in the susceptibility to infection. However, C. trachomatis immunoglobulin (Ig) G-positive subfertile women with tubal pathology were more than twice as likely to be carriers of the mutant TLR4 +896 G allele as compared to those without tubal pathology; however this observation did not reach statistical significance. In conclusion, both the murine model and the human immunogenetics studies show a slight effect upon TLR4 deficiency in the severity of infection but not in the susceptibility to infection.

Description of the ICTI consortium: an integrated approach to the study of Chlamydia trachomatis infection.

The use of an integrated approach to the study of Chlamydia trachomatis infection of the female genital tract, presented at the mini-symposium "Chlamydia trachomatis infections" and described in the thesis of Joseph M. Lyons, has resulted in the creation of the ICTI consortium. The ICTI consortium is based on strong interaction and collaboration between basic scientists, clinicians, epidemiologists, and health care policy makers. This translational approach will help to further the valuable insight into the immunopathogenesis of this sexually transmitted infection (STI) and the development of new intervention strategies, including the vaccines and screening programs necessary to effectively diagnose, treat and prevent C. trachomatis infection. A background of the need for this integrated approach is presented and the goals and participants of the consortium are described.

Effectiveness of amphotericin B lipid complex (ABLC) treatment in allogeneic hematopoietic cell transplant (HCT) recipients with invasive aspergillosis (IA).

A total of 85 allogeneic hematopoietic cell transplant (HCT) recipients with invasive aspergillosis treated with amphotericin B lipid complex (ABLC) were identified from the Collaborative Exchange of Antifungal Research (CLEAR) database. Of these patients, 78% (66/85) presented with pulmonary aspergillosis. Graft-versus-host disease (GVHD) was present in 24 of 85 patients. The response rate to ABLC was 31% (26/85) overall and 21% (5/24) in patients with GVHD. The overall response rate to first-line ABLC treatment was 41% (11/27). Four of nine (44%) patients with GVHD responded to first-line treatment with ABLC, while only one of 13 (8%) responded to ABLC as second-line therapy. Five of 18 (28%) and four of 14 (29%) patients, respectively, responded to sequential or concurrent treatment with ABLC and itraconazole. None of seven patients responded who continued receiving itraconazole after the start of ABLC therapy. At the end of ABLC therapy, serum creatinine had doubled in 12% of patients (10/85), and 2% (2/85) had developed a requirement for dialysis. These data suggest that ABLC, especially when administered as first-line therapy, can result in clinical response even in the most immunocompromised patients, that is, HCT recipients with GVHD, with minimal effects on renal function.

Differences in growth characteristics and elementary body associated cytotoxicity between Chlamydia trachomatis oculogenital serovars D and H and Chlamydia muridarum.

AIM: In vitro growth and elementary body (EB) associated cytotoxicity of two Chlamydia trachomatis strains belonging to serovars D and H and C muridarum were compared to identify difference(s) that correlate with virulence variations between these strains in the mouse model of human female genital tract infection, and phenotypic characteristics that could explain human epidemiological data on serovar prevalence and levels of shedding during serovar D and H infection. METHODS: Replication cycle kinetics, inclusion characteristics, and EB associated cytotoxicity were assessed in McCoy cell monolayers using culture, light microscopy, and lactate dehydrogenase release. RESULTS: Over 72 hours, more rapid production and release of inclusion forming units (ifu) allowed C muridarum to initiate two replication rounds, resulting in 4-8 times more ifu/input unit of infection than with serovars D and H. Although C muridarum EBs were significantly more cytotoxic to McCoy cell monolayers than serovar D at moderate and high multiplicity of infection ratios (MOI), serovar H EBs were significantly more cytotoxic than C muridarum, even at the lowest MOI tested. CONCLUSIONS: These phenotypic differences are consistent with the more invasive course and severe pathological outcome of infection in mice infected with C muridarum, providing an objective basis for questioning the appropriateness of C muridarum as a surrogate for the human biovar of C trachomatis in the murine model of female genital tract infection. The differences seen between the human strains could help explain human epidemiological data relating to differences in prevalence and level of shedding that occurs during infection with oculogenital serovars D and H.

Chlamydia trachomatis serovar E isolates from patients with different clinical manifestations have similar courses of infection in a murine model: host factors as major determinants of C trachomatis mediated pathogenesis.

BACKGROUND: Some investigators have proposed an association between certain Chlamydia trachomatis serovars and the clinical course of infection in humans. A recent study of over 1100 patients with culture confirmed and serotyped C trachomatis urogenital infection detected no such association. AIMS: To corroborate these results using a murine model of female genital tract infection. METHODS: Various parameters of infection were assessed in mice intravaginally infected with human genital isolates of C trachomatis serovar E from four cases with either a clear symptomatic or asymptomatic clinical course in both the patient and their partner. RESULTS: No differences were seen among the strains in the incidence or duration of infection, polymorphonuclear granulocyte responses, or upper genital tract progression. CONCLUSIONS: An investigation to determine the correlation between the clinical manifestations of different isolates of C trachomatis serovar E in humans and certain parameters of microbial pathogenesis in a mouse model failed to reveal an association between the measured parameters and the tendency of serovar E to produce symptomatic versus asymptomatic infections in humans. These findings suggest that differences in the clinical course of infection in humans seen with these strains may be more related to host factors than to genetic variation among strains.

Involvement of caveolin-1 in cholesterol enrichment of high density lipoprotein during its assembly by apolipoprotein and THP-1 cells.

High density lipoprotein (HDL) is assembled by interaction of apolipoprotein A-I with human monocytic leukemia cell line THP-1 by removing cellular cholesterol and phospholipid. Although the HDL formed with undifferentiated THP-1 cells contained only phosphatidylcholine and almost no cholesterol, the cells differentiated with phorbol 12-myristate 13-acetate (PMA) generated HDL enriched in cholesterol. The extent of cholesterol enrichment related to the cellular cholesterol level in the differentiated cells, but only weakly in the undifferentiated cells. In contrast, the differentiation had no influence on the diffusion-mediated cellular cholesterol efflux. The undifferentiated cells expressed the messages of ATP-binding cassette transporter 1 and caveolin-1, at low levels, and the PMA-induced differentiation resulted in substantial expression of both messages. Caveolin-1 protein expression was also highly induced by the PMA treatment of THP-1 cells. When the cells were treated with the antisense DNA of caveolin-1 and differentiated, both caveolin-1 synthesis and cholesterol incorporation into the HDL were reduced in parallel to generate the cholesterol-poor HDL. We concluded that caveolin-1 is involved in enrichment with cholesterol of the HDL generated by the apolipoprotein-cell interaction. This function is independent of the assembly of HDL particles with cellular phospholipid and of nonspecific, diffusion-mediated efflux of cellular cholesterol.

Cholesterol homeostasis in rat astrocytoma cells GA-1.

Astrocytes play a key role in cholesterol metabolism in central nervous system. We have shown that fetal rat astrocytes in primary culture secrete cholesterol-rich HDL with the endogenous apolipoprotein (apo) E and generate cholesterol-poor HDL with exogenous apoE and apoA-I [Ito et al. (1999) J. Neurochem. 72, 2362]. In order to study these reactions in relation to the stage of cell differentiation, we examined generation of HDL by rat astrocytoma cells. Lack of apoE secretion was found in three astrocytoma cell lines, human T98G, rat C6, and GA-1 [Kano-Tanaka et al. (1986) Proc. Jpn. Acad. Ser. B 62, 109]. GA-1 produced apoE at very low level and therefore generated much less HDL by itself than the astrocytes in primary culture. In contrast, GA-1 interacted with exogenous apoE and apoA-I to produce cholesterol-rich HDL while the astrocytes produced cholesterol-poor HDL with these apolipoproteins. Cholesterol biosynthesis rate measured from mevalonate was higher and down-regulated more by LDL in the astrocytes than GA-1. On the other hand, the cellular cholesterol level, uptake of LDL, and cyclodextrin-mediated non-specific diffusion of cholesterol from cell surface were same between these two cells. Treatment of GA-1 with acidic fibroblast growth factor influenced neither the production of apoE nor the baseline lipid secretion, but increased the cholesterol synthesis from mevalonate and the magnitude of its down-regulation by LDL, and decreased cholesterol content in the HDL produced by exogenous apoA-I. In conclusion, suppression of apoE biosynthesis in the undifferentiated astrocytes GA-1 resulted in poor secretion of cholesterol-rich HDL and in turn more production of HDL with exogenous apolipoprotein. Cellular cholesterol homeostasis was altered accordingly.

Sialosylcholesterol induces reorganization of astrocyte filament network.

Sialosylcholesterol induces the differentiation of astrocytes with respect to their morphological appearance (Kato et al., Brain Res. 438 (1988) 277-285; Ito et al., 481 (1989) 335-343), while in a cell-free condition it depolymerizes the astrocyte cellular filaments, the glia filaments and microfilaments (Ito et al., J. Neurochem. 61 (1993) 80-84). To solve this paradox, we examined hetero-interaction between the glia filaments and microfilaments in the presence of sialosylcholesterol. Each filament was prepared in a depolymerized form in low ionic strength, and was adjusted to physiological ionic strength to prevent from repolymerization by sialosylcholesterol. When the two filament preparations in this form were mixed, repolymerization took place in spite of the presence of sialosylcholesterol. The filament formed in the mixture was found almost exclusively composed of vimentin and actin, the major component of the glia filaments and microfilaments preparation, respectively. An excess amount of vimentin over actin in the precipitate implicated that the main mechanism for the hetero-polymerization was the enhancement of vimentin polymerization by actin. To support this view, pre-polymerization of the microfilaments before mixing with the depolymerized glia filaments resulted in a marked decrease in polymerization of the glia filaments. A similar hetero-interaction was found between the purified vimentin and actin. When polymerized vimentin and actin were directly depolymerized by sialosylcholesterol and mixed, polymer formation was demonstrated between these two proteins. Electronmicroscopy indicated direct interaction of the actin filament with the vimentin filament. The results indicate that sialosylcholesterol induces reorganization of the cellular filament network, such as disorganization of vimentin and actin filaments, and provokes their hetero-interaction to form the hetero-filament. Hence, this may be one of the key mechanisms for the induction of cellular differentiation by sialocylcholesterol.

Role of gamma interferon in controlling murine chlamydial genital tract infection.

Earlier investigations have not shown an important role for gamma interferon (IFN-gamma) in the early clearance of chlamydial infection from the murine female genital tract. In a model using a human genital isolate of Chlamydia trachomatis in IFN-gamma and IFN-gamma receptor knockout mice, we were able to demonstrate a major role for IFN-gamma in mediating control of infection throughout the course of infection.

Comparison of amphotropic and pseudotyped VSV-G retroviral transduction in human CD34+ peripheral blood progenitor cells from adult donors with HIV-1 infection or cancer.

In this study we compared the transduction efficiency of conventional amphotropic MoMLV (LPONL[A]) with the MoMLV pseudotyped with that of VSV-G (LPONL[G]) in peripheral blood progenitor cells (PBPCs) from cancer patients and human immunodeficiency virus (HIV)-infected donors. The results showed that LPONL(A) and LPONL(G) infected the progenitor cells from these sources with equal efficiencies. The transgene neoR was detectable by polymerase chain reaction assay in colonies from 14-day colony-forming unit (CFU) assays and in those derived from long-term culture-initiating cell (LTC-ICs) assays. Although the overall levels of transduction efficiency were similar in cord blood and PBPCs from noninfected cancer donors (25-22%) when either LPONL(G) or LPONL(A) was used, they were significantly lower in HIV-1-infected donors compared with noninfected cancer donors when LPONL(G) was used (13 vs. 25%; p = 0.027), and when LPONL(A) was used (12 vs. 22%; p = 0.087). The clonogenic potentials of infected and noninfected CD34+ cells were similar; thus no toxicity could be attributed to the virus preparation. We conclude that PBPCs from HIV-1-infected individuals are transduced less efficiently than those from non-HIV-infected cancer donors. Nonetheless, PBPCs from HIV-infected persons serve as potential targets in gene therapy for acquired immune deficiency syndrome.

Hormonal regulation of aldolase B gene expression in rat primary cultured hepatocytes.

Gene expression of aldolase B, an important enzyme for glucose and fructose metabolism, is regulated by hormones. We examined direct effects of major hormones on aldolase B gene expression in rat primary cultured hepatocytes, in comparison with those on the gene expression of phospho(enol)pyruvate carboxykinase (PEPCK), a key enzyme for gluconeogenesis. Insulin, dexamethasone, and high concentration of glucose increased aldolase B mRNA abundance in the hepatocytes. Glucagon strongly suppressed aldolase B gene expression, and this hormone canceled the stimulative effects of insulin, dexamethasone, and high concentration of glucose. Epinephrine and thyroxine slightly reduced aldolase B mRNA abundance, but these hormones did not cancel the stimulative effects of insulin and dexamethasone. To the contrary, expression of PEPCK gene was suppressed by insulin, dexamethasone, and high concentration of glucose, and remarkably induced by glucagon. Glucagon rapidly suppressed aldolase B gene expression at the transcriptional level. Forskolin and dibutyryl cAMP mimicked the suppressive effect of glucagon on aldolase B gene expression. These results suggest that glucagon may be a key regulator of aldolase B gene transcription through a cAMP/protein kinase A-signaling pathway.

Suppression by gangliosides of polymerization of glial cytoskeletons prepared from rat astrocytes: a role of sialic acid moiety.

This study investigated in vitro the effects of gangliosides on polymerization of either the depolymerized microfilament preparation (MF) or depolymerized glia filament preparation (GF) extracted separately from the crude cytoskeletal fraction of rat astrocytes. Gangliosides GM1, GM2 and GM3 markedly suppressed polymerization of both MF and GF. The concentration of GM1, GM2 or GM3 required to induce 50% inhibition of the polymerization of 7.5 micrograms MF protein/200 microliters (IC50 of GM1, GM2, or GM3) was 3.2, 2.8 or 5.6 micrograms/200 microliters, respectively. The IC50 of each ganglioside for the polymerization of 7.5 micrograms/200 microliters of GF, furthermore, was 3.3, 3.5 or 7.4 micrograms/200 microliters, respectively, suggesting that the inhibitory activities of GM1 and GM2 on polymerization of both MF and GF were greater than those of GM3. GM1, GM2 and GM3 also suppressed dose-dependently the polymerization of both actin and vimentin. The inhibitory activities of GM1 and GM2 on the polymerization of actin or vimentin were greater than GM3, as in the case of polymerization of MF or GF. The IC50S of GD1a and GT1b for MF polymerization at the same concentration were 2.2 and 1.2 micrograms/200 microliters, respectively, and those for GF polymerization were 2.7 and 1.7 micrograms/200 microliters, respectively. The IC50 of GD3 for MF polymerization was 3.9 micrograms/200 microliters, and that for GF polymerization 4.0 micrograms/200 microliters, implying that the inhibitory activities of GD3 on polymerization of both MF and GF were greater than those of GM3. The findings suggested that the inhibitory activities of gangliosides on MF or GF polymerization became greater with increasing number of sialic acid residues. AsialoGM1 suppressed neither MF nor GF polymerization, and inhibited dose-dependently the ability of GM1 to suppress MF polymerization.

Reducing the risk of Chlamydia trachomatis genital tract infection by evaluating the prophylactic potential of vaginally applied chemicals.

The need to provide women with a partner-independent method of prophylaxis against sexually transmitted disease (STD) against, including Chlamydia trachomatis, has led to a direct effort to develop a vaginally applied, broad-spectrum antimicrobial preparation that is both safe and effective. Using a murine model of C. trachomatis lower genital tract infection, we tested the ability of various vaginally applied chemicals to alter the course of infection in female mice challenged with infectious doses ranging from 10(3) to 10(6) inclusion forming units of an oculogenital serovar of C. trachomatis. When administered in a methylcellulose/propylene glycol-base gel 15 minutes prior to challenge, four widely used topical antimicrobials (benzalkonium chloride, chlorhexidine, nonoxynol-9, and polymyxin B) were shown to protect against infection. These results support the possibility that this type of interventional method may be a clinically relevant means of prophylaxis against sexually transmitted infection with C. trachomatis, the most common bacterial STD agent.

Idiopathic CD4+ T-lymphocytopenia and recurrent vulvar intraepithelial neoplasia.

BACKGROUND: CD4+ T-lymphocytopenia immunodeficiency without human immunodeficiency virus (HIV) infection has been reported recently. The association between immunodeficiency and anogenital neoplasia secondary to human papillomavirus infections is well documented. CASE: A woman with recurrent vulvar intraepithelial neoplasia (VIN) had idiopathic CD4+ T-lymphocytopenia without HIV infection. CONCLUSION: Human papillomavirus-related VIN may be associated with idiopathic CD4+ T-lymphocytopenia.

Variation in virulence among oculogenital serovars of Chlamydia trachomatis in experimental genital tract infection.

Seven different oculogenital serovars (D, E, F, G, H, I, and K) of Chlamydia trachomatis were inoculated intravaginally into CF-1 mice, and subsequent infection was monitored. The duration of infection was longest with serovars D and E. This may help to explain clinical surveys which demonstrate a high (50%) prevalence of these serovars. Furthermore, a comparison of the invasiveness of strains D and H demonstrated a much higher frequency of uterine horn infection with serovar D.