Potent synergism of the combination of fluconazole and cyclosporine in Candida albicans.

Several types of drugs currently used in clinical practice were screened in vitro for their potentiation of the antifungal effect of the fungistatic agent fluconazole (FLC) on Candida albicans. These drugs included inhibitors of multidrug efflux transporters, antimicrobial agents, antifungal agents, and membrane-active compounds with no antimicrobial activity, such as antiarrhythmic agents, proton pump inhibitors, and platelet aggregation inhibitors. Among the drugs tested in an agar disk diffusion assay, cyclosporine (Cy), which had no intrinsic antifungal activity, showed a potent antifungal effect in combination with FLC. In a checkerboard microtiter plate format, however, it was observed that the MIC of FLC, as classically defined by the NCCLS recommendations, was unchanged when FLC and Cy were combined. Nevertheless, if a different reading endpoint corresponding to the minimal fungicidal concentration needed to decrease viable counts by at least 3 logs in comparison to the growth control was chosen, the combination was synergistic (fractional inhibitory concentration index of <1). This endpoint fitted to the definition of MIC-0 (optically clear wells) and reflected the absence of the trailing effect, which is the result of a residual growth at FLC concentrations greater than the MIC. The MIC-0 values of FLC and Cy tested alone in C. albicans were >32 and >10 microg/ml, respectively, and decreased to 0.5 and 0.625 microg/ml when the two drugs were combined. The combination of 0.625 microg of Cy per ml with supra-MICs of FLC resulted in a potent antifungal effect in time-kill curve experiments. This effect was fungicidal or fungistatic, depending on the C. albicans strain used. Since the Cy concentration effective in vitro is achievable in vivo, the combination of this agent with FLC represents an attractive perspective for the development of new management strategies for candidiasis.

Macrophage Migration Inhibitory Factor Deficiency Is Associated With Impaired Killing of Gram-Negative Bacteria by Macrophages and Increased Susceptibility to Klebsiella pneumoniae Sepsis.

The cytokine macrophage migration inhibitory factor (MIF) is an important component of the early proinflammatory response of the innate immune system. However, the antimicrobial defense mechanisms mediated by MIF remain fairly mysterious. In the present study, we examined whether MIF controls bacterial uptake and clearance by professional phagocytes, using wild-type and MIF-deficient macrophages. MIF deficiency did not affect bacterial phagocytosis, but it strongly impaired the killing of gram-negative bacteria by macrophages and host defenses against gram-negative bacterial infection, as shown by increased mortality in a Klebsiella pneumonia model. Consistent with MIF's regulatory role of Toll-like 4 expression in macrophages, MIF-deficient cells stimulated with lipopolysaccharide or Escherichia coli exhibited reduced nuclear factor κB activity and tumor necrosis factor (TNF) production. Addition of recombinant MIF or TNF corrected the killing defect of MIF-deficient macrophages. Together, these data show that MIF is a key mediator of host responses against gram-negative bacteria, acting in part via a modulation of bacterial killing by macrophages.

Quantifying uncertainty: physicians' estimates of infection in critically ill neonates and children.

To determine the diagnostic accuracy of physicians' prior probability estimates of serious infection in critically ill neonates and children, we conducted a prospective cohort study in 2 intensive care units. Using available clinical, laboratory, and radiographic information, 27 physicians provided 2567 probability estimates for 347 patients (follow-up rate, 92%). The median probability estimate of infection increased from 0% (i.e., no antibiotic treatment or diagnostic work-up for sepsis), to 2% on the day preceding initiation of antibiotic therapy, to 20% at initiation of antibiotic treatment (P<.001). At initiation of treatment, predictions discriminated well between episodes subsequently classified as proven infection and episodes ultimately judged unlikely to be infection (area under the curve, 0.88). Physicians also showed a good ability to predict blood culture-positive sepsis (area under the curve, 0.77). Treatment and testing thresholds were derived from the provided predictions and treatment rates. Physicians' prognoses regarding the presence of serious infection were remarkably precise. Studies investigating the value of new tests for diagnosis of sepsis should establish that they add incremental value to physicians' judgment.

Macrophage migration inhibitory factor and host innate immune defenses against bacterial sepsis.

Macrophages are essential effector cells of innate immunity that play a pivotal role in the recognition and elimination of invasive microorganisms. Mediators released by activated macrophages orchestrate innate and adaptive immune host responses. The cytokine macrophage migration inhibitory factor (MIF) is an integral mediator of the innate immune system. Monocytes and macrophages constitutively express large amounts of MIF, which is rapidly released after exposure to bacterial toxins and cytokines. MIF exerts potent proinflammatory activities and is an important cytokine of septic shock. Recent investigations of the mechanisms by which MIF regulates innate immune responses to endotoxin and gram-negative bacteria indicate that MIF acts by modulating the expression of Toll-like receptor 4, the signal-transducing molecule of the lipopolysaccharide receptor complex. Given its role in innate immune responses to bacterial infections, MIF is a novel target for therapeutic intervention in patients with septic shock.

Mécanismes physiopathologiques impliqués dans la dysfonction d'organes au cours du sepsis [Pathophysiological mechanisms of organ dysfunction in sepsis].

Sepsis is defined as the systemic inflammatory response to an infection. The occurrence of organ dysfunction increases the severity of sepsis. Complex interactions between multiple immunomodulating mediators and various cell populations, activated secondarily to the initial infectious insult, promote the development of organ dysfunction in sepsis. Although septic organ dysfunction has long been considered as the end result of chaotic, uncontrolled and deregulated inflammatory cascades, it might instead represent an adaptive response to avoid the occurrence of irreversible tissue damage and end-organ injury. In this article, we review the major mechanisms involved in organ dysfunction during sepsis, and also present the concept of organ dysfunction as an adaptive response to the septic process.

Candidiasis Peritoneal: prevalença i factors de risc. Estudi preliminar

Introducció: Un cultiu positiu de líquid peritoneal per a Càndida spp amb clínica associada, és diagnòstic de Candidiasis Peritoneal (CP). Objetius: L’objectiu primari és coneixer la prevalença de CP. Com objectius secundaris coneixer els possibles factors de risc. Tipus d’estudi: Prospectiu, observacional. Mètodes: S’agafa una mostra de 74 pacients amb diagnòstic de peritonitis (2007-2010). Durant la cirugía s’aspira líquid peritoneal lliure i és conrea. Resultats i conclusions: La prevalença va ser del 17.6% (46.15% C. albicans). L’ afectació del tracte gastro-intestinal-superior (OR 6.554) i l’aparició de fallada cardio-vascular durant la cirugia (OR 5.827), són factors de risc per a desenvolupar-la. És estudi preliminar.

Pseudomonas aeruginosa: study of Antibiotic Resistance and Molecular Typing in Hospital Infection Cases in a Neonatal Intensive Care Unit from Rio de Janeiro City, Brazil

This study had the objective of to analyze the demographic and bacteriologic data of 32 hospitalized newborns in an neonatal intensive care unit of a public maternity hospital in Rio de Janeiro city, Brazil, seized by Pseudomonas aeruginosa sepsis during a period ranged from July 1997 to July 1999, and to determine the antimicrobial resistance percentage, serotypes and pulsed field gel electrophoresis (PFGE) patterns of 32 strains isolated during this period. The study group presented mean age of 12.5 days, with higher prevalence of hospital infection in males (59.4%) and vaginal delivery (81.2%), than females (40.6%) and cesarean delivery (18.8%), respectively. In this group, 20 (62.5%) patients received antimicrobials before positive blood cultures presentation. A total of 87.5% of the patients were premature, 62.5% presented very low birth weight and 40.6% had asphyxia. We detected high antimicrobial resistance percentage to b-lactams, chloramphenicol, trimethoprim/sulfamethoxazole and tetracycline among the isolated strains. All isolated strains were classified as multi-drug resistant. Most strains presented serotype O11 while PFGE analysis revealed seven distinct clones with isolation predominance of a single clone (75%) isolated from July 1997 to June 1998.

Ultrasound monitoring of structural urinary tract disease in Schistosoma haematobium infection

A major advance in our understanding of the natural history of Schistosoma haematobium-related morbidity has come through the introduction of the portable ultrasound machines for non-invasive examination of the kidneys and bladder. With the use of generators or battery packs to supply power in non-clinical field settings, and with the use of instant photography or miniaturized thermal printers to record permanent images, it is possible to examine scores of individuals in endemic communities every day. Broad-based ultrasound screening has allowed better definition of age-specific disease risks in urinary schistosomiasis. Results indicate that urinary tract abnormalities are common (18% overall prevalence) in S. haematobium transmission areas, with a 2-4% risk of either severe bladder abnormality or advanced ureteral obstruction. In longitudinal surveys, ultrasound studies have shown that praziquantel and metrifonate therapy are rapidly effective in reversing urinary tract abnormalities among children. The benefits of treating adults are less well known, but research in progress should help to define this issue. Similarly, the prognosis of specific ultrasound findings needs to be clarified, and the ease of sonographic examination will make such long-term follow-up studies feasible. In summary, the painless, quick, and reproducible ultrasound examination has become an essential tool in the study of urinary schistosomiasis.

Systemic antibody responses to gut commensal bacteria during chronic HIV-1 infection.

BACKGROUND: Human systemic antibody responses to commensal microbiota are not well characterised during health and disease. Of particular interest is the analysis of their potential modulation caused by chronic HIV-1 infection which is associated with sustained enteropathy and systemic B cell disturbances reflected by impaired B cell responses and chronic B cell hyperactivity. The mechanisms underlying B cell hyperactivation and the specificities of the resulting hypergammaglobulinaemia are only poorly understood. METHODS: By a technique referred to as live bacterial FACS (fluorescence-activated cell sorting), the present study investigated systemic antibody responses to several gut and skin commensal bacteria as well as Candida albicans in longitudinal plasma and serum samples from healthy donors, chronic HIV-1-infected individuals with or without diarrhoea and patients with inflammatory bowel disease (IBD). RESULTS: The data show that systemic antibody responses to the commensal microbiota were abundantly present in humans and remained remarkably stable over years. Overall systemic antibody responses to gut commensal bacteria were not affected during chronic HIV-1 infection, with titres decreasing when normalised to elevated plasma immunoglobulin G (IgG) levels found in patients with HIV. In contrast, increases in the titres of high affinity antimicrobiota antibodies were detected in patients with IBD, demonstrating that conditions with known increased intestinal permeability and aberrant mutualism can induce changes in antibody titres observed in these assays. CONCLUSION: Neither HIV-associated enteropathy nor B cell dysfunction impact on the high-affinity systemic antibody responses to gut commensal bacteria. HIV-associated hypergammaglobulinaemia is therefore unlikely to be driven by induction of antimicrobiota antibodies.

Species distribution and antifungal susceptibility profile of Candida spp. bloodstream isolates from Latin American hospitals

From March 1999 to March 2000, we conducted a prospective multicenter study of candidemia involving five tertiary care hospitals from four countries in Latin America. Yeast isolates were identified by classical methods and the antifungal susceptibility profile was determined according to the National Committee for Clinical Laboratory Standards microbroth assay method. During a 12 month-period we were able to collect a total of 103 bloodstream isolates of Candida spp. C. albicans was the most frequently isolated species accounting for 42% of all isolates. Non-albicans Candida species strains accounted for 58% of all episodes of candidemia and were mostly represented by C. tropicalis (24.2%) and C. parapsilosis (21.3%). It is noteworthy that we were able to identify two cases of C. lusitaniae from different institutions. In our casuistic, non-albicans Candida species isolates related to candidemic episodes were susceptible to fluconazole. Continuously surveillance programs are needed in order to identify possible changes in the species distribution and antifungal susceptibility patterns of yeasts that may occurs after increasing the use of azoles in Latin American hospitals.

Candida dubliniensis identification in Brazilian yeast stock collection

We investigated the presence of Candida dubliniensis among isolates previously identified as Candida albicans and maintained in a yeast stock collection from 1994 to 2000. All isolates were serotyped and further evaluated for antifungal susceptibility profile. After doing a screening test for C. dubliniensis isolates based on the capability of colonies to grow at 42°C, its final identification was obtained by randomly amplified polymorphic DNA (RAPD) analysis using three different primers. A total of 46 out of 548 screened isolates did not exhibit growth at 42°C and were further genotyped by RAPD. Eleven isolates were identified as C. dubliniensis with RAPD analysis. Regarding serotypes, 81.5% of C. albicans and all C. dubliniensis isolates belonged to serotype A. Of note, 9 out of 11 C. dubliniensis isolates were obtained from patients with acquired immunodeficiency syndrome (Aids) and all of them were susceptible to azoles and amphotericin B. We found 17 (3%) C. albicans isolates that were dose-dependent susceptibility or resistant to azoles. In conclusion, we found a low rate of C. dubliniensis isolates among stock cultures of yeasts previously identified as C. albicans. Most of these isolates were recovered from oral samples of Aids patients and exhibited high susceptibility to amphotericin B and azoles. C. albicans serotype A susceptible to all antifungal drugs is the major phenotype found in our stock culture.

Factors Supporting Cysteine Tolerance and Sulfite Production in Candida albicans.

The amino acid cysteine has long been known to be toxic at elevated levels for bacteria, fungi, and humans. However, mechanisms of cysteine tolerance in microbes remain largely obscure. Here we show that the human pathogenic yeast Candida albicans excretes sulfite when confronted with increasing cysteine concentrations. Mutant construction and phenotypic analysis revealed that sulfite formation from cysteine in C. albicans relies on cysteine dioxygenase Cdg1, an enzyme with similar functions in humans. Environmental cysteine induced not only the expression of the CDG1 gene in C. albicans, but also the expression of SSU1, encoding a putative sulfite efflux pump. Accordingly, the deletion of SSU1 resulted in enhanced sensitivity of the fungal cells to both cysteine and sulfite. To study the regulation of sulfite/cysteine tolerance in more detail, we screened a C. albicans library of transcription factor mutants in the presence of sulfite. This approach and subsequent independent mutant analysis identified the zinc cluster transcription factor Zcf2 to govern sulfite/cysteine tolerance, as well as cysteine-inducible SSU1 and CDG1 gene expression. cdg1Δ and ssu1Δ mutants displayed reduced hypha formation in the presence of cysteine, indicating a possible role of the newly proposed mechanisms of cysteine tolerance and sulfite secretion in the pathogenicity of C. albicans. Moreover, cdg1Δ mutants induced delayed mortality in a mouse model of disseminated infection. Since sulfite is toxic and a potent reducing agent, its production by C. albicans suggests diverse roles during host adaptation and pathogenicity.

Genetic variability analysis among clinical Candida spp. isolates using random amplified polymorphic DNA

The patterns of genetic variation of samples of Candida spp. isolated from patients infected with human immunodeficiency virus in Vitória, state of Espírito Santo, Brazil, were examined. Thirty-seven strains were isolated from different anatomical sites obtained from different infection episodes of 11 patients infected with the human immunodeficiency virus (HIV). These samples were subjected to randomly amplified polymorphic DNA (RAPD) analysis using 9 different primers. Reproducible and complex DNA banding patterns were obtained. The experiments indicated evidence of dynamic process of yeast colonization in HIV-infected patients, and also that certain primers are efficient in the identification of species of the Candida genus. Thus, we conclude that RAPD analysis may be useful in providing genotypic characters for Candida species typing in epidemiological investigations, and also for the rapid identification of pathogenic fungi.

Experimental kinetics of infection induced by Yersinia pseudotuberculosis isolated from stock animals

The course of in vivo infection of five isolates of Yersinia pseudotuberculosis was followed for three weeks in Swiss mice. The strains were isolated from diarrheic and normal feces and mesenteric lymph nodes of healthy and sick stock animals. Four strains of serogroup O:3 and one of serogroup O:1a, with and without the virulence plasmid, were inoculated intragastrically and intravenously in the mice. Groups of five animals were sacrificed at 6 h and 3, 6, 10, 15, and 21 days after inoculation, and organs and tissues were checked for possible macroscopic alterations. Development of infection was monitored at these times by performing viable bacterial counts in homogenates of selected tissues. The animals were cheked daily for clinical alterations. The results of the study showed that strains with the virulence plasmid infected organs and tissues at various times and at varying intensity by both routes of infection, the strain of type O:1a being the most invasive. Moreover, clinical and pathological alterations occurred only in animals inoculated with bacteria carrying the virulence plasmid, regardless of the route of infection.

Immune response in cervical dysplasia induced by human papillomavirus: the influence of human immunodeficiency virus-1 co-infection - review

Human immunodeficiency virus (HIV-1) has become an important risk factor for human papillomavirus (HPV) infection and the development of HPV associated lesions in the female genital tract. HIV-1 may also increase the oncogenicity of high risk HPV types and the activation of low risk types. The Center for Disease Control and Prevention declared invasive cervical cancer an acquired immunodeficience virus (AIDS) defining illness in HIV positive women. Furthermore, cervical cancer happens to be the second most common female cancer worldwide. The host's local immune response plays a critical factor in controlling these conditions, as well as in changes in the number of professional antigen-presenting cells, cytokine, and MHC molecules expression. Also, the production of cytokines may determine which arm of the immune response will be stimulated and may influence the magnitude of immune protection. Although there are many studies describing the inflammatory response in HPV infection, few data are available to demonstrate the influence of the HIV infection and several questions regarding the cervical immune response are still unknown. In this review we present a brief account of the current understanding of HIV/HPV co-infection, emphasizing cervical immune response.