Shrayana Ghosh, Amity University Kolkata
Widely spread in the environment Klebsiella species are a multi-drug resistant pathogens that are increasing rapidly and affects the health of both animals and humans. The main types consist of Klebsiella oxytoca species complex (KoSC) which is a cause of hospital infections and also antibiotic-associated haemorrhagic colitis and the Klebsiella pneumoniae species complex (KpSC), which are a reason of hospital and community infections. The inability to classify each of these species complexes through commonly used clinical microbiological methods has contributed to high errors in the identification rates for these pathogens that obscure not only the true clinical value but also the possible epidemiological characteristics of different species.
Owing to the problems faced in the microbiology laboratory in identifying the individual members of the Klebsiella species, a web-based tool has been developed known as the Klebsiella MALDI TypeR which serves as a user-friendly independent platform that accepts uploaded raw data from MALDI-TOF mass spectrometer and identifies Klebsiella isolates at species complex and its phylogroup levels as well. This tool is built on a database of biomarkers that have been identified and which are unique to the complex organisms, individual phylogroups or associated phylogroups, available at https://maldityper.pasteur.fr.
The web interface Klebsiella MALDI TypeR designed by R and R Shiny enables a user to upload either multiple spectra or a single spectrum, in a zip archive format. Each archive can respond either to a single raw spectrum or several raw spectra of the similar isolate. It’s recommended to use at least 3 technical replicates for every isolate for a better result. After the upload, a panel appears for a quick visual inspection and a table highlighting the presence of any empty spectra.
The process of identification basically has two steps- first is the differentiation of isolates at the species complex level using generally two biomarkers, 57 biomarkers for Klebsiella oxytoca species complex and 29 biomarkers for the Klebsiella pneumoniae species complex. This step is a user choose one with two alternation algorithms, one being the R package MALDI-quant (runs faster) and the other being MassSpecWavelet package (runs slower). Species complex identification is based on the Identification Value which is the ratio between the common peaks between the reference peak list and sample peak list and the number of peaks in the corresponding reference.
The second step involves the identification of the phylogroup specific to the species or subspecies.
This is carried out through processing the raw spectrum by Multi Peak Shift Typing (MPST) with the usage of biomarkers corresponding to the assigned species complex. The tool tries to define the isoform for each individual biomarker and assigns the spectrum to the phylogroup analogous to the applicable MALDI-Type. In cases where automatic identification is not possible manual examination and data-evaluation are required.
Thus, the tool enables fast analysis and visualisation of single or multiple spectra. It demonstrates that Klebsiella ‘s main Klebsiella species complex found in clinical samples can be correctly described by MALDI-TOF MS. Therefore, Klebsiella MALDI TypeR is an effective method for identifying the members of the KpSC, which are difficult to identify in the regular practice of microbiology.
Also read: Molecular Paddling of White Blood Cells
References-
- Brisse, S. et al. (2004) Development of a rapid identification method for Klebsiella pneumoniae phylogenetic groups and analysis of 420 clinical isolates. Clin. Microbiol. Infect., 10, 942–945.
- Du, P. et al. (2006) Improved peak detection in mass spectrum by incorporating continuous wavelet transform-based pattern matching. Bioinforma. Oxf. Engl., 22, 2059–2065.
- https://www.biorxiv.org/content/10.1101/2020.10.13.337162v1.full.pdf
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