Development of a Custom MALDI-TOF MS Database for Species-Level Identification of Bacterial Isolates Collected From Spacecraft and Associated Surfaces.

Arman Seuylemezian,James Tan,Wayne Schubert,Heidi S Aronson,Mandy Lin,Parag Vaishampayan

doi:10.3389/fmicb.2018.00780

Abstract

Since the 1970s, the Planetary Protection Group at the Jet Propulsion Laboratory (JPL) has maintained an archive of spacecraft-associated bacterial isolates. Identification of these isolates was routinely performed by sequencing the 16S rRNA gene. Although this technique is an industry standard, it is time consuming and has poor resolving power for some closely related taxa. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry is widely used in clinical diagnostics and is a promising method to substitute standard 16S rRNA sequencing. However, manufacturer-provided databases lack the bacterial diversity found in spacecraft-assembly cleanrooms. This study reports the development of the first custom database of MALDI-TOF MS profiles of bacterial isolates obtained from spacecraft and associated cleanroom environments. With the use of this in-house database, 454 bacterial isolates were successfully identified in concurrence with their 16S rRNA sequence-based classifications. Additionally, MALDI-TOF MS resolved strain-level variations, identified potential novel species and distinguished between members of taxonomic groups, which is not possible using conventional 16S rRNA sequencing. MALDI-TOF MS has proved to be an accurate, high-throughput approach for real-time identification of bacterial isolates during the spacecraft assembly process.

Highlights

The Committee on Space Research maintains the planetary protection policy for the protection of Earth and other planets from biological contamination
While 16S rRNA sequencing analyzes only a single ∼1,500 bp gene, MALDI-TOF mass spectrometry (MS) analyzes a larger spectrum of protein peptides present in a bacterial cell
MALDI-TOF MS has been implemented in diagnostic microbiology laboratories as a rapid, high-throughput bacterial identification technique (Bizzini et al, 2011)

Summary

Introduction

The Committee on Space Research maintains the planetary protection policy for the protection of Earth and other planets from biological contamination. Microbial bioburden requirements must be met to minimize forward contamination. The NASA Standard Assay (NSA) is the method employed to continually monitor the microbial bioburden present on spacecraft hardware and associated surfaces throughout the assembly, testing, and launch operations of Identification of Spacecraft Microorganisms the flight project. Surface samples obtained from spacecraft and spacecraft-assembly cleanrooms are heat-shocked, and bacterial survivors of this assay are used as a proxy for total microbial bioburden. Planetary protection engineers archive these isolated microorganisms for long-term storage to be used in research studies that may better assess forward contamination concerns

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Conclusion