Biomedical subjects

Scott G Filler

Publications and source records attributed to Scott G Filler.

At least 19 recordsLinked to original sources

PubMed2006

Candida albicans protein kinase CK2 governs virulence during oropharyngeal candidiasis.

To identify Candida albicans genes whose proteins are necessary for host cell interactions and virulence, a collection of C. albicans insertion mutants was screened for strains with reduced capacity to damage endothelial cells in vitro. This screen identified CKA2. CKA2 and its homologue CKA1 encode the catalytic subunits of the protein kinase CK2. cka2delta/cka2delta strains of C. albicans were constructed and found to have significantly reduced capacity to damage both endothelial cells and an oral epithelial cell line in vitro. Although these strains invaded endothelial cells similarly to the wild-type strain, they were defective in oral epithelial cell invasion. They were also hypersusceptible to hydrogen peroxide, but not to high salt or to cell wall damaging agents. A cka1delta/cka1delta mutant caused normal damage to both endothelial cells and oral epithelial cells, and it was not hypersusceptible to hydrogen peroxide. However, overexpression of CKA1 in a cka2delta/cka2delta strain restored wild-type phenotype. Although the cka2delta/cka2delta mutant had normal virulence in the mouse model of haematogenously disseminated candidiasis, it had significantly attenuated virulence in the mouse model of oropharyngeal candidiasis. Therefore, Cka2p governs the interactions of C. albicans with endothelial and oral epithelial cells in vitro and virulence during oropharyngeal candidiasis.

Animals↗

PubMed2006

Daptomycin versus standard therapy for bacteremia and endocarditis caused by Staphylococcus aureus.

BACKGROUND: Alternative therapies for Staphylococcus aureus bacteremia and endocarditis are needed. METHODS: We randomly assigned 124 patients with S. aureus bacteremia with or without endocarditis to receive 6 mg of daptomycin intravenously per kilogram of body weight daily and 122 to receive initial low-dose gentamicin plus either an antistaphylococcal penicillin or vancomycin. The primary efficacy end point was treatment success 42 days after the end of therapy. RESULTS: Forty-two days after the end of therapy in the modified intention-to-treat analysis, a successful outcome was documented for 53 of 120 patients who received daptomycin as compared with 48 of 115 patients who received standard therapy (44.2 percent vs. 41.7 percent; absolute difference, 2.4 percent; 95 percent confidence interval, -10.2 to 15.1 percent). Our results met prespecified criteria for the noninferiority of daptomycin. The success rates were similar in subgroups of patients with complicated bacteremia, right-sided endocarditis, and methicillin-resistant S. aureus. Daptomycin therapy was associated with a higher rate of microbiologic failure than was standard therapy (19 vs. 11 patients, P=0.17). In 6 of the 19 patients with microbiologic failure in the daptomycin group, isolates with reduced susceptibility to daptomycin emerged; similarly, a reduced susceptibility to vancomycin was noted in isolates from patients treated with vancomycin. As compared with daptomycin therapy, standard therapy was associated with a nonsignificantly higher rate of adverse events that led to treatment failure due to the discontinuation of therapy (17 vs. 8, P=0.06). Clinically significant renal dysfunction occurred in 11.0 percent of patients who received daptomycin and in 26.3 percent of patients who received standard therapy (P=0.004). CONCLUSIONS: Daptomycin (6 mg per kilogram daily) is not inferior to standard therapy for S. aureus bacteremia and right-sided endocarditis. (ClinicalTrials.gov number, NCT00093067 [ClinicalTrials.gov].).

Adult↗

PubMed2006

Candida-host cell receptor-ligand interactions.

The interaction of Candida species with their cognate host receptors is a key factor in the pathogenesis of different types of candidiasis. The recognition of different forms of Candida albicans by Toll-like receptors 2 and 4 on mononuclear leukocytes has recently been discovered to determine the function and activity of regulatory T-cells, determine the balance of Type 1 and Type 2 cytokines and, thereby, influence the antifungal activity of both the innate and adaptive immune response. Different forms of C. albicans are also recognized by different lectins that are expressed on the surface macrophages. C. albicans and Candida glabrata express the ALS (agglutinin-like sequence) and EPA (epithelial adhesin) families of adhesins, respectively. A key difference between C. glabrata and C. albicans is that EPA expression in C. glabrata is governed by sub-telomeric silencing, whereas ALS expression in C. albicans is regulated by other mechanisms.

Animals↗

PubMed2006

Efficacy of the anti-Candida rAls3p-N or rAls1p-N vaccines against disseminated and mucosal candidiasis.

We have shown that vaccination with the recombinant N terminus of Als1p (rAls1p-N) protects mice against disseminated and oropharyngeal candidiasis. We now report that vaccination of mice with a related candidate, rAls3p-N, induces a broader antibody response than rAls1p-N and a similar cell-mediated immune response. The rAls3p-N vaccine was equally as effective as rAls1p-N against disseminated candidiasis but was more effective than rAls1p-N against oropharyngeal or vaginal candidiasis. Antibody titers did not correlate with protection against disseminated candidiasis, but delayed-type hypersensitivity did. The rAls3p-N vaccine is a promising new vaccine candidate for further exploration to prevent systemic and mucosal candidal infections.

Animals↗

PubMed2006

Efficacy of ambruticin analogs in a murine model of invasive pulmonary aspergillosis.

Ambruticins are a family of polyketides. The antifungal activity of an ambruticin, KOSN-2079, was tested in the mouse model of invasive aspergillosis. KOSN-2079 significantly reduced pulmonary fungal burdens and improved survival over that with the vehicle control. These results support the continued development of ambruticins as antifungal agents.

Animals↗

PubMed2006

Standardization of an experimental murine model of invasive pulmonary aspergillosis.

Evaluating new therapeutic agents for invasive aspergillosis requires animal models that are reproducible among different laboratories. We therefore evaluated a murine model of aerosol infection in two independent laboratories and found a high level of both intra- and interlaboratory reproducibility of survival, fungal burden over time, and the efficacy of liposomal amphotericin B.

Amphotericin B↗

PubMed2006

Candida albicans Ecm33p is important for normal cell wall architecture and interactions with host cells.

Candida albicans ECM33 encodes a glycosylphosphatidylinositol-linked cell wall protein that is important for cell wall integrity. It is also critical for normal virulence in the mouse model of hematogenously disseminated candidiasis. To identify potential mechanisms through which Ecm33p contributes to virulence, we investigated the interactions of C. albicans ecm33Delta mutants with endothelial cells and the FaDu oral epithelial cell line in vitro. The growth rate of blastospores of strains containing either one or no intact copies of ECM33 was 50% slower than that of strains containing two intact copies of ECM33. However, all strains germinated at the same rate, forming similar-length hyphae on endothelial cells and oral epithelial cells. Strains containing either one or no intact copies of ECM33 had modestly reduced adherence to both types of host cells, and a markedly reduced capacity to invade and damage these cells. Saccharomyces cerevisiae expressing C. albicans ECM33 did not adhere to or invade epithelial cells, suggesting that Ecm33p by itself does not act as an adhesin or invasin. Examination of ecm33Delta mutants by transmission electron microscopy revealed that the cell wall of these strains had an abnormally electron-dense outer mannoprotein layer, which may represent a compensatory response to reduced cell wall integrity. The hyphae of these mutants also had aberrant surface localization of the adhesin Als1p. Collectively, these results suggest that Ecm33p is required for normal cell wall architecture as well as normal function and expression of cell surface proteins in C. albicans.

Candida albicans↗

PubMed2006

Critical role of Bcr1-dependent adhesins in C. albicans biofilm formation in vitro and in vivo.

The fungal pathogen Candida albicans is frequently associated with catheter-based infections because of its ability to form resilient biofilms. Prior studies have shown that the transcription factor Bcr1 governs biofilm formation in an in vitro catheter model. However, the mechanistic role of the Bcr1 pathway and its relationship to biofilm formation in vivo are unknown. Our studies of biofilm formation in vitro indicate that the surface protein Als3, a known adhesin, is a key target under Bcr1 control. We show that an als3/als3 mutant is biofilm-defective in vitro, and that ALS3 overexpression rescues the biofilm defect of the bcr1/bcr1 mutant. We extend these findings with an in vivo venous catheter model. The bcr1/bcr1 mutant is unable to populate the catheter surface, though its virulence suggests that it has no growth defect in vivo. ALS3 overexpression rescues the bcr1/bcr1 biofilm defect in vivo, thus arguing that Als3 is a pivotal Bcr1 target in this setting. Surprisingly, the als3/als3 mutant forms a biofilm in vivo, and we suggest that additional Bcr1 targets compensate for the Als3 defect in vivo. Indeed, overexpression of Bcr1 targets ALS1, ECE1, and HWP1 partially restores biofilm formation in a bcr1/bcr1 mutant background in vitro, though these genes are not required for biofilm formation in vitro. Our findings demonstrate that the Bcr1 pathway functions in vivo to promote biofilm formation, and that Als3-mediated adherence is a fundamental property under Bcr1 control. Known adhesins Als1 and Hwp1 also contribute to biofilm formation, as does the novel protein Ece1.

Animals↗

PubMed2006

Fungal invasion of normally non-phagocytic host cells.

Many fungi that cause invasive disease invade host epithelial cells during mucosal and respiratory infection, and subsequently invade endothelial cells during hematogenous infection. Most fungi invade these normally non-phagocytic host cells by inducing their own uptake. Candida albicans hyphae interact with endothelial cells in vitro by binding to N-cadherin on the endothelial cell surface. This binding induces rearrangement of endothelial cell microfilaments, which results in the endocytosis of the organism. The capsule of Cryptococcus neoformans is composed of glucuronoxylomannan, which binds specifically to brain endothelial cells, and appears to mediate both adherence and induction of endocytosis. The mechanisms by which other fungal pathogens induce their own uptake are largely unknown. Some angioinvasive fungi, such as Aspergillus species and the Zygomycetes, invade endothelial cells from the abluminal surface during the initiation of invasive disease, and subsequently invade the luminal surface of endothelial cells during hematogenous dissemination. Invasion of normally non-phagocytic host cells has different consequences, depending on the type of invading fungus. Aspergillus fumigatus blocks apoptosis of pulmonary epithelial cells, whereas Paracoccidioides brasiliensis induces apoptosis of epithelial cells. This review summarizes the mechanisms by which diverse fungal pathogens invade normally non-phagocytic host cells and discusses gaps in our knowledge that provide opportunities for future research.

Apoptosis↗

PubMed2005

Current treatment strategies for disseminated candidiasis.

The incidence of disseminated candidiasis has increased dramatically over the past several decades. Fortunately, in recent years, a variety of new antifungal agents have become available to treat these infections. On the basis of efficacy, safety, and cost considerations, fluconazole is the agent of choice for the empirical treatment of disseminated candidiasis in nonneutropenic, hemodynamically stable patients, unless a patient is suspected to be infected with an azole-resistant species (i.e., Candida glabrata or Candida krusei). For hemodynamically unstable or neutropenic patients, agents with broader species coverage, such as polyenes, echinocandins, or, possibly, voriconazole, are preferred for empirical treatment of candidemia. Modification of the initial, empirical regimen depends on the response to therapy and the subsequent identification of the species of the offending pathogen. Echinocandins or high-dose polyenes are preferred for the treatment of infections with C. glabrata or C. krusei. Central venous catheters should be removed from all patients who have disseminated candidiasis, if feasible, and antifungal therapy should be administered to all patients who have candidemia or proven candidiasis.

Antifungal Agents↗

PubMed2005

A randomized study of the use of fluconazole in continuous versus episodic therapy in patients with advanced HIV infection and a history of oropharyngeal candidiasis: AIDS Clinical Trials Group Study 323/Mycoses Study Group Study 40.

BACKGROUND: In human immunodeficiency virus (HIV)-infected patients, fluconazole prophylaxis is associated with reductions in the rate of fungal infection. However, concerns exist with regard to the use of fluconazole prophylaxis and the risk of development of fluconazole treatment-refractory infections. METHODS: We performed a randomized, open-label trial that compared oral fluconazole given continuously (200 mg 3 times weekly; the "continuous fluconazole arm") with fluconazole that was provided only for episodes of orophayngeal candidiasis (OPC) or esophageal candidiasis (EC) (the "episodic fluconazole arm") in HIV-infected persons with CD4+ T cell counts of <150 cells/mm3 and a history of OPC. The primary study end point was the time to development of fluconazole-refractory OPC or EC, which was defined as lack of response to 200 mg fluconazole given daily for 14 or 21 days, respectively. RESULTS: A total of 413 subjects were randomized to receive continuous fluconazole, and 416 were randomized to receive episodic fluconazole. After 42 months, 17 subjects (4.1%) in the continuous fluconazole arm developed fluconazole-refractory OPC or EC infections, compared with 18 subjects (4.3%) in the episodic fluconazole arm, with no difference between treatment arms with regard to the time to development of a fluconazole-refractory infection within 24 months (P=.88, by log-rank test) or before the end of the study (P=.97, by the log-rank test). Continuous fluconazole therapy was associated with fewer cases of OPC or EC (0.29 vs. 1.08 episodes per patient-year; P<.0001) and fewer invasive fungal infections (15 vs. 28 episodes; P=.04, by chi2 test), but not with improved survival, compared with episodic fluconazole therapy. CONCLUSION: Continuous fluconazole therapy is not associated with significant risk of fluconazole-refractory OPC or EC, compared with episodic fluconazole therapy, in HIV-infected patients with access to active antiretroviral therapy.

AIDS-Related Opportunistic Infections↗

PubMed2005

The Aspergillus fumigatus StuA protein governs the up-regulation of a discrete transcriptional program during the acquisition of developmental competence.

Members of the Asm1p, Phd1p, Sok2p, Efg1p, and StuAp (APSES) family of fungal proteins regulate morphogenesis and virulence in ascomycetes. We cloned the Aspergillus fumigatus APSES gene encoding StuAp and demonstrated that stuA transcription is markedly up-regulated after the acquisition of developmental competence. A. fumigatus DeltastuA mutants were impaired in their ability to undergo asexual reproduction. Conidiophore morphology was markedly abnormal, and only small numbers of dysmorphic conidia were produced, which exhibited precocious germination. Whole genome transcriptional analysis during the onset of developmental competence was performed and identified a subset of developmentally regulated genes that were stuA dependent, including a cluster of putative secondary metabolite biosynthesis genes, genes encoding proteins implicated in the regulation of morphogenesis, and genes encoding allergens and other antigenic proteins. Additionally, hyphae of the DeltastuA mutant displayed reduced expression of the catalase gene CAT1 and were hypersusceptible to hydrogen peroxide.

Amino Acid Sequence↗

PubMed2005

Tumor necrosis factor inhibition and invasive fungal infections.

Candida albicans and Aspergillus fumigatus occur ubiquitously in nature; C. albicans is part of the natural flora of most healthy individuals, and A. fumigatus is commonly found in soil, plant debris, and indoor air. Neither fungus poses a threat to healthy individuals, but each can cause fatal infections in immunocompromised patients. The use of tumor necrosis factor (TNF) antagonists for the treatment of rheumatoid arthritis and other autoimmune diseases has been associated with an increased incidence of opportunistic infections, including infections with both of these fungi. Because the use of TNF antagonists is expected to increase in the future, understanding the role that TNF plays and the effect of its antagonism on host defense against infections with these fungi is critical for reducing the associated morbidity and mortality.

Humans↗

PubMed2005

Mice with disseminated candidiasis die of progressive sepsis.

BACKGROUND: Candida species are among the most common etiologies of nosocomial bloodstream infections, causing a mortality of >40%. The murine model of hematogenously disseminated candidiasis is the standard for investigating both the activity of antifungal agents and the pathogenesis of this disease. However, despite decades of use, little is known about the physiological characteristics of the host in this model, and the cause of death remains unclear. METHODS: Using i-STAT technology, we measured blood chemistry and hemodynamic parameters to define host physiological characteristics during murine disseminated candidiasis. RESULTS: Mice with hematogenously disseminated candidiasis died of progressive sepsis, as manifested by worsening hypotension, tachycardia, and hypothermia. The mice developed metabolic acidosis, as well as profound acidemia and hypoglycemia. They also developed renal insufficiency, which became severe only shortly before death. Kidney fungal burden was correlated with severity of renal failure and systemic acidosis. The presence of significant weight loss, hypotension, or hypothermia was predictive of imminent death. CONCLUSIONS: These findings indicate that the murine model of hematogenously disseminated candidiasis accurately recapitulates the progressive sepsis seen during severe clinical cases. The results underscore the validity of the model for study of the pathophysiological aspects of this disease, as well as for the evaluation of antifungal drug efficacy.

Animals↗

PubMed2005

N-cadherin mediates endocytosis of Candida albicans by endothelial cells.

Candida albicans is the most common cause of fungal bloodstream infections. To invade the deep tissues, blood-borne organisms must cross the endothelial cell lining of the vasculature. We have found previously that C. albicans hyphae, but not blastospores, invade endothelial cells in vitro by inducing their own endocytosis. Therefore, we set out to identify the endothelial cell receptor that mediates the endocytosis of C. albicans. We determined that endocytosis of C. albicans was not mediated by bridging molecules in the serum and that it was partially dependent on the presence of extracellular calcium. Using an affinity purification procedure, we discovered that endothelial cell N-cadherin bound to C. albicans hyphae but not blastospores. N-cadherin also co-localized with C. albicans hyphae that were being endocytosed by endothelial cells. Chinese hamster ovary (CHO) cells expressing human N-cadherin endocytosed significantly more C. albicans hyphae than did CHO cells expressing either human VE-cadherin or no human cadherins. The expression of N-cadherin by the CHO cells resulted in enhanced endocytosis of hyphae, but not blastospores, indicating the selectivity of the N-cadherin-mediated endocytosis. Down-regulation of endothelial cell N-cadherin expression with small interfering RNA significantly inhibited the endocytosis of C. albicans hyphae. Therefore, a novel function of N-cadherin is that it serves as an endothelial cell receptor, which mediates the endocytosis of C. albicans.

Animals↗

PubMed2005

Role of the fungal Ras-protein kinase A pathway in governing epithelial cell interactions during oropharyngeal candidiasis.

Tpk1p, Tpk2p and Efg1p are members of the Ras-protein kinase A pathway that governs the yeast-to-hyphal transition in Candida albicans. We used tpk1Delta/tpk1Delta, tpk2Delta/tpk2Delta and efg1Delta/efg1Delta mutants to investigate the role of these proteins in regulating the interactions of C. albicans with oral epithelial cell lines in vitro and virulence in murine models of oropharyngeal candidiasis (OPC) and haematogenously disseminated candidiasis (HDC). The tpk1Delta/tpk1Delta strain adhered to, invaded and damaged oral epithelial cells in vitro similarly to the wild-type strain. In contrast, both the tpk2Delta/tpk2Delta and efg1Delta/efg1Delta strains had reduced capacity to invade and damage oral epithelial cells, and the efg1Delta/efg1Delta strain also exhibited decreased adherence to these cells. Consistent with these in vitro findings, the tpk2Delta/tpk2Delta and efg1Delta/efg1Delta strains also had significantly attenuated virulence during OPC. Therefore, Tpk2p and Efg1p both govern factors that enable C. albicans to invade and damage oral epithelial cells in vitro and cause OPC. These results also suggest that hyphal formation mediated by the Ras-protein kinase A pathway is a key virulence mechanism during OPC. Interestingly, the efg1Delta/efg1Delta strain, but not the tpk2Delta/tpk2Delta had reduced virulence during HDC. Thus, Tpk2p may be more important for governing virulence during OPC than HDC.

Animals↗

PubMed2005

Vaccination with recombinant N-terminal domain of Als1p improves survival during murine disseminated candidiasis by enhancing cell-mediated, not humoral, immunity.

Candida spp. are opportunistic fungal pathogens that are among the most common causes of nosocomial bloodstream infections. The mortality attributable to disseminated candidiasis is 40 to 50% despite antifungal therapy. Clearly, new strategies are needed to prevent this life-threatening infection. Because risk factors for disseminated candidiasis are well defined and frequently of limited duration, vaccination is an appealing prophylactic strategy. We have identified a cell surface protein, Als1p, that mediates adherence of Candida albicans to a variety of human substrates and plastic. Here we report that immunizing BALB/c mice with the recombinant N-terminal domain of Als1p (rAls1p-N) improved survival during a subsequent challenge with a lethal inoculum of C. albicans. The protective 20-mug dose of rAls1p-N significantly increased Candida stimulation of Th1 splenocytes and increased in vivo delayed-type hypersensitivity. In contrast, antibody titers did not correlate with protection. Finally, the vaccine was not protective in T-cell-deficient mice but was protective in B-cell-deficient mice. These data indicate that the mechanism of action of the rAls1p-N vaccine is stimulation of cell-mediated, rather than humoral, immunity against C. albicans. The majority of efforts to date have focused on the development of passive immunization strategies to prevent or treat disseminated candidiasis. In contrast, our results provide proof of principle for vaccination with an adhesin of C. albicans and emphasize the potential for cell-mediated immune modulation as a prophylactic or therapeutic strategy against disseminated candidiasis.

Animals↗

PubMed2005

The anti-Candida albicans vaccine composed of the recombinant N terminus of Als1p reduces fungal burden and improves survival in both immunocompetent and immunocompromised mice.

We have previously shown that intraperitoneal vaccination with the recombinant N terminus of Als1p (rAls1p-N) modestly improves survival during murine disseminated candidiasis. We now report marked efficacy with subcutaneous rAls1p-N vaccination. Efficacy is retained in neutropenic and corticosteroid-treated mice. The rAls1p-N vaccine is a promising candidate for the prevention of invasive candidiasis.

Animals↗