Academic journal article Bulletin of the World Health Organization

Diagnosis of Disseminated Mycobacterial Infection: Testing a Simple and Inexpensive Method for Use in Developing Countries

Academic journal article Bulletin of the World Health Organization

Diagnosis of Disseminated Mycobacterial Infection: Testing a Simple and Inexpensive Method for Use in Developing Countries

Article excerpt

Introduction

Following the development of the acquired immuunodeficiency syndrome (AIDS) epidemic, an increasing number of secondary disseminated mycobacterial infections has prompted the search for a sensitive method of recovering mycobacteria from blood. The highest diagnostic yield for Mycobacterium avium complex organisms is achieved by lysis-centrifugation culture methods (1, 2). Such systems are commercially available and are commonly used in the USA and in Western Europe (3, 4). Diagnosis with the radiometric BACTEC culture medium is more rapid and has a similar sensitivity but requires a radioactive substrate and does not yield isolated colonies (5, 6). Both methods are too expensive for routine use in developing countries. This prospective study compares the efficiency of two different methods for detecting mycobacterial growth from the peripheral blood of AIDS patients: (1) direct inoculation onto a biphasic medium and (2) an inexpensive non-commercial lysis-centrifugation method.

Methods

Patients

During the period December 1993 to August 1994, blood specimens were obtained from 50 AIDS patients with suspected disseminated mycobacterial disease. The main characteristic used to select patients was fever of unknown origin plus one of the following: hepatosplenomegaly, lung infiltrate, anaemia, or elevated alkaline phosphatase. Furthermore, when possible, a CD4 lymphocyte T cell count (cells per [micro] l) was performed.

Blood specimens

A total of 3 blood samples were taken from each patient at 15-minute intervals by a trained phlebotomist. Each sample was collected in 2 Vacutainer tubes (Becton Dickinson, MD, USA) containing ethylenediaminetetraacetic acid (EDTA) as an anticoagulant. The temperature of the patients was measured at each sample. A total of 138 blood samples were obtained from the patients studied. Other specimens (e.g. sputum) were obtained for routine mycobacterial culture according to clinical indications.

Procedure

Upon arrival in the laboratory the contents of one Vacutainer tube were mixed with 30 ml of 0.3% (w/v) sodium deoxicholate solution and incubated for 15 min at room temperature. After centrifugation for 25 min at 3000 g (4 [degrees] C), the pellet was suspended in 1 ml of 0.2% bovine albumin saline (Fraction V. Sigma. St. Louis, MO, USA), and 0.2 ml of the suspension was added to one tube with Middlebrook 7H9 broth and to two Lowenstein-Jensen slants.

The contents of the second Vacutainer were directly added to biphasic medium containing 5 ml of Middlebrook 7H9 broth and 15ml of Middlebrook 7H10 agar. All mycobacterial isolates were identified using standard biochemical methods.

Statistical analysis

The [chi square] test (with Yates' correction for 2 x 2 tables) and Fisher's exact test were used to compare statistics between the patient groups. For continuous variables Student's t-test was used. In each case, a two-sided test with a 0.05 significance level was used.

Results

Mycobacterial growth was obtained from 70/138 specimens (from 30/50 (60%) patients). Positive cultures yielded M. tuberculosis in 19 (63%) and M. avium complex organisms in 11 (37%) patients. Only 4 patients with M. avium complex cultured from blood had mycobacteria isolated from cultures obtained from other clinical specimens. Of these, one patient had M. avium complex isolated from a liver sample (patient No. 3). Two patients with M. avium complex isolated from blood showed M. tuberculosis in cultures from either sputum (patient No. 46) or a lymph node biopsy (patient No. 1). One patient had Mycobacterium gordonae isolated from faeces (patient No. 13). The majority of patients with positive blood cultures for M. tuberculosis had the same mycobacteria species isolated from other clinical samples. However, 5 patients could be diagnosed with tuberculosis only after the results of blood culture (patients No. …

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