Application of Whole Genome Sequencing and MALDI-TOF to the Identifcation of Bacillus Species Isolated from Cleanrooms at NASA Johnson Space Center

Astromaterial cleanrooms at NASA Johnson Space Center are built environments that
hold samples, such as lunar rocks, from different space exploration missions. Bacillus sp.
are frequently detected in routine microbial monitoring of these facilities. Since this, and
related genera, can form endospores that can withstand harsh conditions, they could
contaminate astromaterials. This could confound searches for extraterrestrial life. Whole
genome sequencing (WGS) is widely used for identifying bacterial strains and tracking
their source; however, WGS is expensive and time consuming. Matrix-assisted laser
desorption ionization– time of flight mass spectrometry (MALDI-TOF) shows promise as
a low-cost, rapid method of identifying strains of bacteria, but few studies have compared
this proteomics method to WGS. To evaluate a high throughput method of tracking the
source of contamination of this built environment, WGS and MALDI-TOF was
conducted on 18 bacterial strains isolated from surfaces in astromaterials cleanrooms.
WGS identified 14 Bacillus, 2 Paenibacillus, 1 Solibacillus and 1 Alcaligenes strains. These isolates showed similarity to strains commonly observed in spacecraft assembly
cleanrooms at other facilities. Cluster analysis of mass spectra generated by MALDI-TOF
grouped strains together that were greater than 94% similar to each other in terms of
amino acid sequences of single copy core genes, as assessed by WGS. This suggests that
MALDI-TOF and WGS results are consistent with each other and MALDI-TOF can
rapidly identify strains of Bacillus sp. isolated from cleanroom environments with a
resolution comparable to WGS. Based on phylogenomic analysis, these results also
suggest the presence of a cosmopolitan class of Bacillus sp. that are more likely to be
found in cleanrooms and similar built environments than in natural systems.