Comparative analysis of diagnostic methods for the detection of Cryptococcus neoformans meningitis

• Published in: PLoS neglected tropical diseases 2023-03, Vol.17 (3), p.e0011140-e0011140
• Main Authors: Dantas, Katia Cristina, de Freitas-Xavier, Roseli Santos, Spina Lombardi, Suzete Cleusa Ferreira, Júnior, Alfredo Mendroni, da Silva, Marcos Vinicius, Criado, Paulo Ricardo, de Freitas, Vera Lúcia Teixeira, de Almeida, Terezinha Morato Bastos
• Format: Article
• Language: English
• Subjects: Acquired immune deficiency syndrome; Acquired Immunodeficiency Syndrome; Agglutination; Agglutination tests; AIDS; Analysis; Assaying; Biology and Life Sciences; Causes of; Cerebrospinal fluid; Comparative analysis; Cryptococcal meningitis; Cryptococcosis; Cryptococcosis - diagnosis; Cryptococcus; Cryptococcus neoformans; Cryptococcus neoformans - genetics; Deoxyribonucleic acid; Detection; Diagnosis; Diagnostic systems; DNA; Fungal infections; Fungi; HIV; Human immunodeficiency virus; Humans; Identification and classification; Latex; Latex agglutination; Latex Fixation Tests; Medicine and Health Sciences; Meningitis; Meningitis, Cryptococcal - diagnosis; Molecular diagnostic techniques; Mortality; Nucleotide sequence; Opportunist infection; PCR; Polymerase chain reaction; Protocols; Research and Analysis Methods; Samples; Sensitivity; Sensitivity analysis; Serum; Specificity; Staining; Strains; Tropical diseases
• ISSN: 1935-2735; 1935-2727; 1935-2735
• DOI: 10.1371/journal.pntd.0011140

Cryptococcosis is a devastating opportunistic infection in immunocompromised individuals, primarily in people living with HIV/AIDS. This study evaluated a protocol for the early diagnosis of meningitis due to C. neoformans, utilizing established molecular techniques from serum and CSF samples. The 18S and 5.8S (rDNA-ITS) sequence-specific nested PCR assays were compared with direct India ink staining and the latex agglutination test for detection of C. neoformans in serum and cerebrospinal fluid (CSF) from 49 Brazilian suspected meningitis patients. Results were validated with samples obtained from 10 patients negative for cryptococcosis and HIV, and by analysis of standard C. neoformans strains. The 5.8S DNA-ITS PCR was more sensitive (89-100%) and specific (100%) than the 18S rDNA PCR and conventional tests (India ink staining and latex agglutination) for identification of C. neoformans. While the 18S PCR exhibited a sensitivity (72%) similar to that of the latex agglutination assay in serum samples, it was superior to the latex agglutination assay when testing CSF, with a sensitivity of 84%. However, the latex agglutination was superior to the 18SrDNA PCR in specificity in CSF (92%). The 5.8S DNA-ITS PCR yielded the highest levels of accuracy (96-100%) of any test for detection (serological and mycological) of C. neoformans in both serum and CSF. Use of the nested 5.8S PCR was superior to other techniques for the diagnosis of cryptococcosis. The possibility of using serum, a non-invasively collected material, in a targeted 5.8S PCR analysis to identify Cryptococcus spp. is recommended, especially in immunosuppressed patients. Our results indicate that nested 5.8S PCR can increase the diagnostic capability of cryptococcosis, and we suggest its use to monitor patients in the future.