The selection of the commercial platform was based on the manufacturer's claims relating to high specificity, sensitivity and the availability of the test kits within the country. An immunochromatographic assay allows the detection of a target analyte in a specific sample. This was performed in accordance with the manufacturer's recommendations. Immunochromatographic assays were employed for the serological detection of cryptococcal capsular polysaccharide antigen in CSF. Characteristically, the attachment between the parent and daughter cell is extremely narrow, and a lattice formation is evident within a cluster of cryptococci. For the Gram stain, cryptococci appear as single, budding, thin-walled, oval to round cells containing gram-positive granular inclusions, surrounded by a capsule that varies in size and appears to be gram-negative. In the India ink test, the polysaccharide capsule does not take up the stain, resulting in the appearance of a refractile large white circle against a dark background. Microscopy techniques were then used to carry out morphological tests to identify cryptococcus directly from the CSF using India ink staining and Gram staining these tests were carried out in accordance with established standard operating procedures (SOPs) 14. CSF was collected from participants following aseptic standard procedures used in the hospital. OK, USA Dynamiker Biotechnology (Tianjin) Co., Ltd). Four types of diagnostic tests were conducted on the samples, including: the India ink test, Gram stain, and two types of commercial LFAs (Immy, Inc. The main objectives of the study were to: (1) establish the detection rate of positive-CM patients among HIV-infected patients presenting with features of meningitis in Mzuzu Central Hospital by using different diagnostic methods, and (2) compare the sensitivity and specificity of the Gram stain, India ink and two LFA tests for the diagnosis of CM.Ĭryptococcus species were identified by morphological, biochemical and serological characteristics. We therefore undertook a comparative evaluation of Gram stain, India ink stain, and two forms of lateral flow assay (LFA) (Immy and Dynamiker), for the rapid diagnosis of cryptococcal meningitis in HIV-infected patients in Mzuzu, Malawi. Furthermore, there is little published data available that compares microscopic and antigen-based CM diagnostic methods, particularly in resource-constrained settings such as Malawi. There are few published data on the utility of the Gram stain for the direct detection of cryptococci in CSF samples, particularly in patients infected with HIV. Therefore, microscopy continues to be widely adopted as a rapid, cheap and reliable diagnostic method in these settings. Antigen detection is rapid, and is associated with high sensitivity (93%–100%) and specificity (93%–98%) however, this form of detection is not available in low-resource countries due to its excessive costs 10. Even though India ink is rated poorly in terms of diagnostic ability for CM, and with regards to sensitivity and specificity 8 – 11, evidence indicates that India ink stain is still widely used for the detection of cryptococci in CSF, particularly in resource-limited countries such as Malawi 12.Īlthough cell culture is relatively slow, it is still considered to represent the gold standard for CM diagnosis. The laboratory diagnosis of CM includes direct visualization of cryptococci via microscopy, culture of the organism and/or the detection of cryptococcal antigens in cerebrospinal fluid (CSF). As such there is a need for a more accurate and robust diagnostic test for the diagnosis of CM 7. The clinical presentations of CM are often indistinguishable from other causes of meningitis. Consequently, early diagnosis and the timely initiation of antifungal therapy is essential if we are to reduce morbidity and mortality from cryptococcal disease. In Malawi, studies have reported that more than 40% of patients diagnosed with CM die within 4 weeks of the diagnosis, and that 55%–58% die by 10 weeks 5, 6. Despite the increased coverage of antiretroviral treatment (ART) programs in high burden countries, the case fatality rate of CM remains high at 35%–65% in SSA 2 – 4.

Globally, there are an estimated 181,100 estimated deaths from CM, with 75% of the deaths occurring in sub-Saharan Africa (SSA) 1. Cryptococcal meningitis (CM) is the most common systemic fungal infection in patients with HIV infection.

India ink stain wood