MALDI-TOF lipidomics rapidly detects modification of 2-hydroxymyristate lipid A, a potential virulence trait in Enterobacter bugandensis

Abstract

ABSTRACT Enterobacter cloacae complex (ECC) comprises numerous species that can be difficult to identify. Among these species, Enterobacter bugandensis is recognized as a threat in neonatal intensive care units due to its involvement in septic shock. A potential virulence factor responsible for its virulence is the modification of the lipid A of the lipopolysaccharide (LPS). This modification corresponds to a hydroxylation of the myristate component of lipid A. A collection of 168 ECC, including 10 E. bugandensis and one isolate involved in fatal septic shock in the neonatal intensive care unit, were tested for the presence of 2-hydroxymyristate. All isolates were sequenced to determine the genetic diversity of this collection and detect the presence of the lpxO gene responsible for this hydroxylation. matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF) analysis revealed that only a fraction (4/11) of E. bugandensis were able to modify their lipid A, which correlates with the presence of the lpxO gene. Genomic analysis of E. bugandensis revealed that only half of the genomes carried the lpxO gene. Lpxo-negative isolates exhibited a deletion of 1,094 bp encompassing the lpxO gene. Lipid A analysis also revealed that hydroxylation of myristate was not specific to E. bugandensis since this modification was systematically found in several species including Enterobacter chengduensis, Enterobacter chuandaensis, or Enterobacter cloacae subsp cloacae. Here, we provide a simple detection test allowing the rapid identification of hydroxymyristate of lipid A involved in the virulence of E. bugandensis. Rapid identification of lipid A modification might help clinicians to adapt procedures when an Enterobacter spp. is identified in newborns.IMPORTANCEEnterobacter bugandensis is now recognized as a potential threat in neonatal wards due to its ability to survive in incubators and its virulence properties. Its increased virulence is linked to the addition of 2-hydroxymyristate on lipid A, resulting from the action of the LpxO enzyme. Here, we proposed a rapid and simple assay to decipher the presence of 2-hydroxymyristate on lipid A directly from the bacterial colony. This assay can be performed easily on MALDI-TOF routine machine using a commercial kit (MBT Lipid Xtract Kit, Bruker Daltonic). This test can accurately detect the 2-hydroxymyristate on lipid A of lpxO-positive ECC isolates, including E. bugandensis. In neonate wards, newborns are now screened for ECC carriage. Accordingly, the MALDIxin test might help clinicians detect the presence of lpxO-positive E. bugandensis, which can be considered a risk factor for the newborn, and implement measures to avoid potentially fatal septicemia.

Keywords

• detection test
• carbapenemase
• Enterobacterales
• Galleria mellonella