Creation of an In-House Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry Corynebacterineae Database Overcomes Difficulties in Identification of Nocardia farcinica Clinical Isolates

Abstract

e Nocardiosis is a rare disease that is caused by Gram-positive actinobacteria of the Nocardia genus and affects predominantly immunocompromised patients. In its disseminated form, it has a predilection for the central nervous system and is associated with high mortality rates. Therefore, prompt identification of the pathogen is critical. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry is a relatively novel technique used for identification of microorganisms. In this work, an upgraded MALDI-TOF Biotyper database containing Corynebacterineae representatives of strains deposited in the Polish Collection of Microorganisms was created and used for identification of the strain isolated from a nocardial brain abscess, mimicking a brain tumor, in an immunocompetent patient. Testing with the API Coryne system initially incorrectly identified Rhodococcus sp., while chemotaxonomic tests, especially mycolic acid analysis, enabled correct Nocardia identification only at the genus level. Subsequent sequence analysis of 16S rRNA and secA1 genes confirmed the identification. To improve the accuracy of the results, an in-house database was constructed using optimized parameters; with the use of the database, the strain was eventually identified as Nocardia farcinica. Clinical laboratories processing various clinical strains can upgrade a commercial database to improve and to accelerate the results obtained. This is especially important in the case of Nocardia, for which valid microbial diagnosis remains challenging; reference laboratories are often required to identify and to survey these rare actinobacteria.N ocardia represents Gram-positive actinobacteria that are etiological agents of the rare disease nocardiosis, which affects predominantly immunocompromised patients. Nocardiosis usually presents itself as a solitary lesion located in lung, skin, or brain (primary nocardiosis), but sometimes disseminated nocardiosis with multiple sites is observed. Nocardia spp. show a marked predilection for the central nervous system (CNS) (1).Nocardiosis of the CNS constitutes about 40% of all cases of disseminated nocardioses and is associated with high mortality rates. The true incidence of Nocardia infections is difficult to assess, because of problems with the identification of these bacteria. Many different phenotypic, chemotaxonomic, or genotyping methods are available, but each method possesses drawbacks; for this reason, fast reliable identification of Nocardia isolates is very important. Because Nocardia spp. differ in antibiotic susceptibility, correct identification of clinical strains is critical for adequate treatment.Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) has been introduced for microbial identification in the past few years. Its reliability, accuracy, and cost-effectiveness have been well described in the literature (2-4). This method can be used to identify microorganisms from all domains of life, i.e., Archaea, Bacteria, and E...

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