Diagnostic Criteria of Nontuberculous Mycobacterial Lung Disease Key Laboratory Features of NTM Health Careand Hygiene-associated Disease Prevention Prophylaxis and Treatment of NTM Disease Introduction Methods Taxonomy Epidemiology Pathogenesis Host Defense and Immune Defects Pulmonary Disease Body Morphotype Tumor Necrosis Factor Inhibition Laboratory Procedures Collection, Digestion, Decontamination, and Staining of Specimens Respiratory Specimens Body Fluids, Abscesses, and Tissues Blood Specimen Processing Smear Microscopy Culture Techniques Incubation of NTM Cultures NTM Identification Antimicrobial Susceptibility Testing for NTM Molecular Typing Methods of NTM Clinical Presentations and Diagnostic Criteria Pulmonary Disease Cystic Fibrosis Hypersensitivity-like Disease Transplant Recipients Disseminated Disease Lymphatic Disease Skin, Soft Tissue, and Bone Disease

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An official ATS/IDSA statement: diagnosis, treatment, and prevention of nontuberculous mycobacterial diseases.

Detailed restriction analyses of many samples often require substantial amounts of time and effort for DNA extraction, restriction digests, Southern blotting, and hybridization. We describe a novel approach that uses the polymerase chain reaction (PCR) for rapid simplified restriction typing and mapping of DNA from many different isolates. DNA fragments up to 2 kilobase pairs in length were efficiently amplified from crude DNA samples of several pathogenic Cryptococcus species, including C. neoformans, C. albidus, C. laurentii, and C. uniguttulatus. Digestion and electrophoresis of the PCR products by using frequent-cutting restriction enzymes produced complex restriction phenotypes (fingerprints) that were often unique for each strain or species. We used the PCR to amplify and analyze restriction pattern variation within three major portions of the ribosomal DNA (rDNA) repeats from these fungi. Detailed mapping of many restriction sites within the rDNA locus was determined by fingerprint analysis of progressively larger PCR fragments sharing a common primer site at one end. As judged by PCR fingerprints, the rDNA of 19 C. neoformans isolates showed no variation for four restriction enzymes that we surveyed. Other Cryptococcus spp. showed varying levels of restriction pattern variation within their rDNAs and were shown to be genetically distinct from C. neoformans. The PCR primers used in this study have also been successfully applied for amplification of rDNAs from other pathogenic and nonpathogenic fungi, including Candida spp., and ought to have wide applicability for clinical detection and other studies.

Rapid genetic identification and mapping of enzymatically amplified ribosomal DNA from several Cryptococcus species.

Molecular chaperones, including the heat-shock proteins (Hsps), are a ubiquitous feature of cells in which these proteins cope with stress-induced denaturation of other proteins. Hsps have received the most attention in model organisms undergoing experimental stress in the laboratory, and the function of Hsps at the molecular and cellular level is becoming well understood in this context. A complementary focus is now emerging on the Hsps of both model and nonmodel organisms undergoing stress in nature, on the roles of Hsps in the stress physiology of whole multicellular eukaryotes and the tissues and organs they comprise, and on the ecological and evolutionary correlates of variation in Hsps and the genes that encode them. This focus discloses that (a) expression of Hsps can occur in nature, (b) all species have hsp genes but they vary in the patterns of their expression, (c) Hsp expression can be correlated with resistance to stress, and (d) species' thresholds for Hsp expression are correlated with levels of stress that they naturally undergo. These conclusions are now well established and may require little additional confirmation; many significant questions remain unanswered concerning both the mechanisms of Hsp-mediated stress tolerance at the organismal level and the evolutionary mechanisms that have diversified the hsp genes.

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Heat-shock proteins, molecular chaperones, and the stress response: evolutionary and ecological physiology

Invasive candidiasis (IC) is a leading cause of mycosis-associated mortality in the United States. We examined data from the National Center for Health Statistics and reviewed recent literature in order to update the epidemiology of IC. IC-associated mortality has remained stable, at approximately 0.4 deaths per 100,000 population, since 1997, while mortality associated with invasive aspergillosis has continued to decline. Candida albicans remains the predominant cause of IC, accounting for over half of all cases, but Candida glabrata has emerged as the second most common cause of IC in the United States, and several less common Candida species may be emerging, some of which can exhibit resistance to triazoles and/or amphotericin B. Crude and attributable rates of mortality due to IC remain unacceptably high and unchanged for the past 2 decades. Nonpharmacologic preventive strategies should be emphasized, including hand hygiene; appropriate use, placement, and care of central venous catheters; and prudent use of antimicrobial therapy. Given that delays in appropriate antifungal therapy are associated with increased mortality, improved use of early empirical, preemptive, and prophylactic therapies should also help reduce IC-associated mortality. Several studies have now identified important variables that can be used to predict risk of IC and to help guide preventive strategies such as antifungal prophylaxis and early empirical therapy. However, improved non-culture-based diagnostics are needed to expand the potential for preemptive (or early directed) therapy. Further research to improve diagnostic, preventive, and therapeutic strategies is necessary to reduce the considerable morbidity and mortality associated with IC.

Epidemiology of Invasive Candidiasis: a Persistent Public Health Problem

Infectious Diseases Society of America.

ALTHOUGH osteomyelitis often falls into the purview of the orthopedic surgeon, the internist and pediatrician are also very frequently involved in the initial diagnosis and the subsequent antibioti...

Osteomyelitis: a review of clinical features, therapeutic considerations and unusual aspects.

Demographic information, risk factors, therapy, and outcome for all patients who had candidemia at Barnes Hospital, St. Louis, between 1 September 1988 and 1 September 1989 were retrospectively reviewed. One hundred six candidemic patients were identified, representing 0.5% of all medical and surgical discharges and 0.33% of total patient discharges. These percentages represent a 20-fold increase in the incidence of candidemia at our hospital in comparison with that during 1976-1979. Candida albicans was the most frequently isolated species (63%), followed by Candida tropicalis (17%), Candida glabrata (13%), Candida parapsilosis (6.5%), and Candida krusei (0.9%). Overall mortality was 57%, and 14 (23%) of 60 deaths occurred within 48 hours of the detection of candidemia. Mortality was associated with higher APACHE II scores (25 for nonsurvivors vs. 16 for survivors; P = .0001), the presence of a rapidly fatal underlying illness (P = .0009), and sustained positivity of blood cultures (P = .02). In cases of sustained candidemia, the isolation of non-albicans Candida species also correlated with increased mortality (8 of 8 vs. 10 of 21; P = .005). Thirty candidemic patients (28%) did not receive any antifungal therapy, and 19 (63%) of these untreated patients died. Eleven untreated patients (37%) survived without sequelae. There has been a marked increase in the incidence of candidemia in our institution that is associated with a high overall mortality. Candidemia lasting less than 24 hours was associated with a lower mortality than was that of longer duration. Severity of illness and duration of candidemia should be used as stratifying factors in prospective studies to determine optimum therapy.

Candidemia in a Tertiary Care Hospital: Epidemiology, Risk Factors, and Predictors of Mortality

This minireview describes the problems associated with the use of Amphotericin B and what is known about the mechanism(s) of its action. The efforts to design a more efficient vehicle for AmB are summarized in the following minireview (8)

Amphotericin B: current understanding of mechanisms of action.

One hundred ninety-four patients with cryptococcal meningitis were enrolled in a multicenter, prospective, randomized clinical trial to compare the efficacy and toxicity of four as compared with six weeks of combination amphotericin B and flucytosine therapy. Among 91 patients who met preestablished criteria for randomization, cure or improvement was noted in 75 percent of those treated for four weeks and in 85 percent of those treated for six weeks. The estimated relapse rate for the four-week regimen was higher--27 as compared with 16 percent--whereas the incidence of toxic effects for the two regimens was similar--44 as compared with 43 percent. Among 23 transplant recipients, 4 of 5 treated for four weeks relapsed, leading to the decision to treat the rest of the group for six weeks. Only 3 of the 18 treated for six weeks relapsed. In a third group of 80 patients, the protocol was not followed during the initial four weeks, and these patients were not randomized. Thirty-eight died or relapsed. Multifactorial analysis of pretreatment factors for all 194 patients identified three significant predictors (P less than 0.05) of a favorable response: headache as a symptom, normal mental status, and a cerebrospinal fluid white-cell count above 20 per cubic millimeter. These and other findings in this study are consistent with the view that the four-week regimen should be reserved for patients who have meningitis without neurologic complications, underlying disease, or immunosuppressive therapy; a pretreatment cerebrospinal fluid white-cell count above 20 per cubic millimeter and a serum cryptococcal antigen titer below 1:32; and at four weeks of therapy, a negative cerebrospinal fluid India ink preparation and serum and cerebrospinal fluid cryptococcal-antigen titers below 1:8. Patients who do not meet these criteria should receive at least six weeks of therapy.

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Treatment of Cryptococcal Meningitis with Combination Amphotericin B and Flucytosine for Four as Compared with Six Weeks

IN the past few years, there has been an increase in the number of systemic fungal infections seen in clinical practice. These infections have been of two types: those due to primary pathogens such...

Gerald Medoff

Papers

Osteomyelitis: a review of clinical features, therapeutic considerations and unusual aspects.

Candidemia in a Tertiary Care Hospital: Epidemiology, Risk Factors, and Predictors of Mortality

Amphotericin B: current understanding of mechanisms of action.

Treatment of Cryptococcal Meningitis with Combination Amphotericin B and Flucytosine for Four as Compared with Six Weeks

Strategies in the treatment of systemic fungal infections.