Clinical and Epidemiological Correlates of Genotypes within the Mycobacterium avium Complex Defined by Restriction and Sequence Analysis of hsp65

Species identification of isolates of the Mycobacterium avium complex (MAC) remains a difficult task. Although M. avium and Mycobacterium intracellulare can be identified with expensive, commercially available probes, many MAC isolates remain unresolved, including those representing Mycobacterium lentiflavum as well as other potentially undefined species. PCR restriction analysis (PRA) of the hsp65 gene has been proposed as a rapid and inexpensive approach. We applied PRA to 278 MAC isolates, including 126 from blood of human immunodeficiency virus (HIV)-infected patients, 59 from sputum of HIV-negative patients with chronic obstructive pulmonary disease, 88 from environmental sources, and 5 pulmonary isolates from a different study. A total of 15 different PRA patterns were observed. For 27 representative isolates, a 441-bp fragment of the hsp65 gene was sequenced; based on 54 polymorphic sites, 18 different alleles were defined, including 12 alleles not previously reported. Species and phylogenetic relationships were more accurately defined by sequencing than by PRA or commercial probe. The distribution of PRA types and, by implication, phylogenetic lineages among blood isolates was significantly different from that for pulmonary and environmental isolates, suggesting that particular lineages have appreciably greater virulence and invasive potential

Polyclonal Infections Due to Mycobacterium Avium Complex in Patients with AIDS Detected by Pulsed-field Gel Electrophoresis of Sequential Clinical Isolates.

Invasive infection with organisms of the Mycobacterium avium complex (MAC) is common among patients with advanced human immunodeficiency virus infection. In previous studies, we analyzed multiple individual colonies of MAC isolated from specimens obtained at the same time and observed that 14 to 20% of patients are simultaneously infected with more than one strain. In this study, we examined sequential isolates from 12 patients with AIDS who had two or more MAC isolates available from clinical specimens collected more than 1 week apart; the intervals between the first and last specimens ranged from 8 to 192 (median, 46) days. For each isolate, restriction digests of genomic DNA were analyzed by pulsed-field gel electrophoresis; DNA was prepared by using a protocol, described here in detail, which had been optimized for conditions of bacterial growth and lysis. The pulsed-field gel electrophoresis analysis identified four patients (33%) infected with two different MAC strains. Both M. avium and M. intracellulare were cultured from blood specimens from two patients. In each of the four patients, the second strain was identified from a culture taken within 14 days of the initial study isolate, and in three of these patients, the first strain was detected again in a subsequent culture. These observations suggest that the presence of two different strains among isolates from sequential cultures may reflect ongoing polyclonal infection. We conclude that polyclonal infection with MAC is common among patients with AIDS. The identification of such infections may be critical in the development of effective treatments

Polyclonal Mycobacterium Avium Infections in Patients with AIDS: Variations in Antimicrobial Susceptibilities of Different Strains of M. Avium Isolated from the Same Patient.

Broth microdilution MICs were determined for pairs of strains isolated from five AIDS patients with polyclonal Mycobacterium avium infection. Four (80%) of the five patients were infected simultaneously with strains having different antimicrobial susceptibility patterns. These findings have implications for the interpretation of susceptibility data in M. avium prophylaxis and treatment trials

Isolation of Mycobacterium avium from Potable Water in Homes and Institutions of Patients with HIV Infection in Finland and the United States

Symptomatic disease by nontuberculous mycobacteria has been linked to potable water from institutional and domestic potable water systems. Potable water samples were collected from homes and institutions of patients with AIDS. Colonization of potable water with nontuberculous mycobacteria was demonstrated in 230 (15%) of 1489 samples collected from domestic and institutional water systems of patients with HIV infection in the United States and Finland. Mycobacterium avium was the most common species and colonization was favored at temperatures of 40-50 degrees C in recirculating hot water systems. Such systems are a plausible source of human infection and disease.Peer reviewe

Basis for Treatment of Tuberculosis among HIV-Infected Patients in Tanzania: The Role of Chest X-Ray and Sputum Culture

Active tuberculosis (TB) is common among HIV-infected persons living in tuberculosis endemic countries, and screening for tuberculosis (TB) is recommended routinely. We sought to determine the role of chest x-ray and sputum culture in the decision to treat for presumptive TB using active case finding in a large cohort of HIV-infected patients. Ambulatory HIV-positive subjects with CD4 counts ≥ 200/mm3 entering a Phase III TB vaccine study in Tanzania were screened for TB with a physical examination, standard interview, CD4 count, chest x-ray (CXR), blood culture for TB, and three sputum samples for acid fast bacillus (AFB) smear and culture

Reliable identification of mycobacterial species by PCR-restriction enzyme analysis (PRA)-hsp65 in a reference laboratory and elaboration of a sequence-based extended algorithm of PRA-hsp65 patterns

<p>Abstract</p> <p>Background</p> <p>Identification of nontuberculous mycobacteria (NTM) based on phenotypic tests is time-consuming, labor-intensive, expensive and often provides erroneous or inconclusive results. In the molecular method referred to as PRA-<it>hsp65</it>, a fragment of the <it>hsp65 </it>gene is amplified by PCR and then analyzed by restriction digest; this rapid approach offers the promise of accurate, cost-effective species identification. The aim of this study was to determine whether species identification of NTM using PRA-<it>hsp65 </it>is sufficiently reliable to serve as the routine methodology in a reference laboratory.</p> <p>Results</p> <p>A total of 434 NTM isolates were obtained from 5019 cultures submitted to the Institute Adolpho Lutz, Sao Paulo Brazil, between January 2000 and January 2001. Species identification was performed for all isolates using conventional phenotypic methods and PRA-<it>hsp65</it>. For isolates for which these methods gave discordant results, definitive species identification was obtained by sequencing a 441 bp fragment of <it>hsp65</it>. Phenotypic evaluation and PRA-<it>hsp65 </it>were concordant for 321 (74%) isolates. These assignments were presumed to be correct. For the remaining 113 discordant isolates, definitive identification was based on sequencing a 441 bp fragment of <it>hsp65</it>. PRA-<it>hsp65 </it>identified 30 isolates with <it>hsp65 </it>alleles representing 13 previously unreported PRA-<it>hsp65 </it>patterns. Overall, species identification by PRA-<it>hsp65 </it>was significantly more accurate than by phenotype methods (392 (90.3%) vs. 338 (77.9%), respectively; p < .0001, Fisher's test). Among the 333 isolates representing the most common pathogenic species, PRA-<it>hsp65 </it>provided an incorrect result for only 1.2%.</p> <p>Conclusion</p> <p>PRA-<it>hsp65 </it>is a rapid and highly reliable method and deserves consideration by any clinical microbiology laboratory charged with performing species identification of NTM.</p

Campylobacteriosis, Eastern Townships, Québec

Independent risk factors for campylobacteriosis (eating raw, rare, or undercooked poultry; consuming raw milk or raw milk products; and eating chicken or turkey in a commercial establishment) account for <50% of cases in Québec. Substantial regional and seasonal variations in campylobacteriosis were not correlated with campylobacter in chickens and suggested environmental sources of infection, such as drinking water

Molecular Epidemiology of HIV-Associated Tuberculosis in Dar es Salaam, Tanzania: Strain Predominance, Clustering, and Polyclonal Disease

Molecular typing of Mycobacterium tuberculosis can be used to elucidate the epidemiology of tuberculosis, including the rates of clustering, the frequency of polyclonal disease, and the distribution of genotypic families. We performed IS6110 typing and spoligotyping on M. tuberculosis strains isolated from HIV-infected subjects at baseline or during follow-up in the DarDar Trial in Tanzania and on selected community isolates. Clustering occurred in 203 (74%) of 275 subjects: 124 (80%) of 155 HIV-infected subjects with baseline isolates, 56 (69%) of 81 HIV-infected subjects with endpoint isolates, and 23 (59%) of 39 community controls. Overall, 113 (41%) subjects had an isolate representing the East Indian “GD” family. The rate of clustering was similar among vaccine and placebo recipients and among subjects with or without cellular immune responses to mycobacterial antigens. Polyclonal disease was detected in 6 (43%) of 14 patients with multiple specimens typed. Most cases of HIV-associated tuberculosis among subjects from this study in Dar es Salaam resulted from recently acquired infection. Polyclonal infection was detected and isolates representing the East Indian GD strain family were the most common

Multi-institutional analysis shows that low PCAT-14 expression associates with poor outcomes in prostate cancer

AbstractBackgroundLong noncoding RNAs (lncRNAs) are an emerging class of relatively underexplored oncogenic molecules with biological and clinical significance. Current inadequacies for stratifying patients with aggressive disease presents a strong rationale to systematically identify lncRNAs as clinical predictors in localized prostate cancer.ObjectiveTo identify RNA biomarkers associated with aggressive prostate cancer.Design, setting, and participantsRadical prostatectomy microarray and clinical data was obtained from 910 patients in three published institutional cohorts: Mayo Clinic I (N=545, median follow-up 13.8 yr), Mayo Clinic II (N=235, median follow-up 6.7 yr), and Thomas Jefferson University (N=130, median follow-up 9.6 yr).Outcome measurements and statistical analysisThe primary clinical endpoint was distant metastasis-free survival. Secondary endpoints include prostate cancer-specific survival and overall survival. Univariate and multivariate Cox regression were used to evaluate the association of lncRNA expression and these endpoints.Results and limitationsAn integrative analysis revealed Prostate Cancer Associated Transcript-14 (PCAT-14) as the most prevalent lncRNA that is aberrantly expressed in prostate cancer patients. Down-regulation of PCAT-14 expression significantly associated with Gleason score and a greater probability of metastatic progression, overall survival, and prostate cancer-specific mortality across multiple independent datasets and ethnicities. Low PCAT-14 expression was implicated with genes involved in biological processes promoting aggressive disease. In-vitro analysis confirmed that low PCAT-14 expression increased migration while overexpressing PCAT-14 reduced cellular growth, migration, and invasion.ConclusionsWe discovered that androgen-regulated PCAT-14 is overexpressed in prostate cancer, suppresses invasive phenotypes, and lower expression is significantly prognostic for multiple clinical endpoints supporting its significance for predicting metastatic disease that could be used to improve patient management.Patient summaryWe discovered that aberrant prostate cancer associated transcript-14 expression during prostate cancer progression is prevalent across cancer patients. Prostate cancer associated transcript-14 is also prognostic for metastatic disease and survival highlighting its importance for stratifying patients that could benefit from treatment intensification