Nocardiosis in HIV seropositive clinically suspected pulmonary tuberculosis patients

Introduction

Nocardiae are branching Gram-positive and weakly acid-fast bacteria. ¹ They are frequently found in soil. Nocardia asteroides is the most common species to be isolated from clinical specimens. Human infections are rare. The infection is contracted through inhalation and is more common among immunocompromised patients, especially those with an impaired cell-mediated immunity. ²

Nocardia generally originate as a pulmonary infection, varying from mild and slowly progressive to fulminant and fatal. The involvement of the brain, meninges and spinal cord is said to be more common than in other parts of the body such as skin, subcutaneous tissues, eye and liver. ³

The diagnosis is confirmed by the demonstration of branching filaments, either microscopically or by culture. ¹ Nocardia infection is serious: approximately 40% of the diagnoses are made at autopsy. The mortality rate is high, and those who survive often suffer significant end organ damage. ³

There are many reports of nocardiosis associated with HIV infections in the industrialized and developing countries, but its true prevalence is unknown. ⁴ Recent increases in the reported frequency of human nocardial infections can be attributed to the use of improved selective isolation procedures and increased clinical and microbiological awareness. However, in some developing countries where other chronic lung diseases, particularly tuberculosis (TB), are prevalent, nocardiae are either missed or misidentified in clinical specimens. ⁵

The aim of this study was to determine the frequency of nocardiosis in HIV-seropositive patients clinically suspected to have pulmonary TB.

Materials and methods

Sputum samples were collected from 140 HIV-seropositive clinically suspected pulmonary TB patients at Jimma University Specialized Hospital, Jimma, Ethiopia. The study was conducted between May and July 2005.

All samples were first screened for nocardia by Gram and modified Ziehl Neelsen (ZN) staining. ¹ Sputum samples were digested by N-acetyl cysteine without NaOH, ⁶ centrifuged, and the sediments were then inoculated on Lowenstein Jensen egg medium slopes.^1,6

Inoculated slopes were incubated at 37°C on 5% CO₂ for one week, thereafter at 37°C in air for another two weeks and were checked alternate days for nocardia growth. ²

Growth of the nocardia was confirmed by:

Ethical considerations

Oral consent was obtained and samples were only collected from consenting patients. Each respondent was interviewed separately to maintain privacy.

Results

Of the 140 patients, four (2.85%) patients tested positive for nocardia by microscopy using the Gram and modified ZN staining techniques; five (3.6%: two women and three men), all aged between 30 and 39 years old, had a positive culture for N. asterioides.

The growth of nocardia on Löwenstein-Jensen medium was confirmed by the appearance of typical colony morphology, by microscopy and by biochemical reactions. From colony morphology, colonies of four isolates appeared visible around the 11th day and in one around the 16th day of incubation. The colonies were small orange-red, wrinkled, granular and rough – typical of nocardia.^1,7

All isolates appeared as Gram-positive but unevenly stained, highly-branched filaments, fragmented to form bacillary and coccoid elements suggestive of the nocardia species.^1,2

Nocardia are acid fast to 1% sulphuric acid, which in smears were stained by modified ZN staining technique.

All the five isolates were negative for casein, tyrosine and xanthine hydroxylase, and all were positive for urease. The reactions confirmed the isolates as N asteroids. ⁶

Discussion

Nocardiae isolation from sputum samples of patients with lung infections is highly indicative of pulmonary nocardiosis.^5,8 In this study, nocardiae were isolated from the sputum of five HIV-seropositive patients with a suspected clinical diagnosis of pulmonary TB. No mycobacterium TB growth occurred on five positive slopes of nocardial growth after eight weeks of reincubation. The patients had symptoms such as a cough of three or more weeks’ duration, shortness of breath, chest pain, intermittent fever, loss of weight and appetite and tiredness.

The pulmonary manifestation of nocardiosis is often confused with TB. ² Similar findings have been reported in previous studies, suggesting that a considerable percentage of patients presenting symptoms of chronic lung disease in African countries could be suffering from pulmonary nocardiosis.^5,7,8

Clinical, radiological and histopathological findings are not sufficient for the recognition of nocardiosis. Definitive diagnosis depends upon the isolation and identification of the causal agent from clinical material. ⁹

The incidence of pulmonary nocardiosis in HIV- seropositive patients is increasing. ¹⁰ The actual extent of nocardial infection among HIV-positive patients is unknown as it is not an AIDS-defining illness as per the current criteria of the Centers for Disease Control. ²

A correct diagnosis is dependent upon the laboratory at which the specimens are analysed. Most laboratories discard bacterial cultures which are negative after 48 h, and TB laboratories do not process sputum specimens without decontaminating non-mycobacterial TB species.

Nocardiosis is a treatable lung disease which may be more common in developing countries than is currently recognized. ⁸ The prevalence of nocardiosis varies geographically and, in zones where HIV-associated TB is common. ⁴ It is possible that some patients diagnosed as smear-negative pulmonary TB actually have nocardiosis. Identification plays an important part in the management of the patient. To be considered negative for nocardia, inoculated culture media should be held for three weeks ⁶ ; furthermore, direct Gram or modified ZN staining of sputum samples is crucial in order to make a rapid and economical diagnosis.

As per the autopsy study conducted in Ivory Coast, 10 instances of (4%) nocardiosis were seen in HIV-positive cadavers, of which six had nocardiosis as the main cause of death. No nocardiosis was seen in HIV-negative cadavers. ⁴

The high mortality rate in undiagnosed patients should alert physicians caring for HIV-seropositive patients. ¹⁰ It is therefore imperative that clinicians in African and other developing countries should be aware of this entity when dealing with HIV/immunocompromised patients with respiratory infections which fail to respond to antitubercular treatments.