Biology subjects

Chandler, C. E.

Publications and source records attributed to Chandler, C. E..

4 recordsLinked to original sources

bioRxiv2023

Structural determination of Rickettsia lipid A without chemical extraction confirms shorter acyl chains in later-evolving Spotted Fever Group pathogens

Rickettsiae are Gram-negative obligate intracellular parasites of numerous eukaryotes. Human pathogens of the Transitional Group (TRG), Typhus Group (TG), and Spotted Fever Group (SFG) rickettsiae infect blood-feeding arthropods, have dissimilar clinical manifestations, and possess unique genomic and morphological attributes. Lacking glycolysis, rickettsiae pilfer numerous metabolites from host cytosol to synthesize peptidoglycan and lipopolysaccharide (LPS). For LPS, O-antigen immunogenicity varies between SFG and TG pathogens; however, lipid A proinflammatory potential is unknown. We previously demonstrated that R. akari (TRG), R. typhi (TG), and R. montanensis (SFG) produce lipid A with long 2 secondary acyl chains (C16 or C18) compared to short 2 secondary acyl chains (C12) in R. rickettsii (SFG) lipid A. To further probe this structural heterogeneity and estimate a time point when shorter 2 secondary acyl chains originated, we generated lipid A structures for two additional SFG rickettsiae (R. rhipicephali and R. parkeri) utilizing Fast Lipid Analysis Technique adopted for use with tandem mass spectrometry (FLATn). FLATn allowed analysis of lipid A structure directly from host cell-purified bacteria, providing substantial improvement over lipid A chemical extraction. FLATn-derived structures indicate SFG rickettsiae diverging after R. rhipicephali evolved shorter 2 secondary acyl chains. Bioinformatics analysis of Rickettsia LpxL late acyltransferases revealed discrete active sites and hydrocarbon rulers for long versus short 2 secondary acyl chain addition. While the significance of different lipid A structures for diverse Rickettsia pathogens is unknown, our success using FLATn will facilitate determining how structural heterogeneity impacts interactions with host lipid A receptors and overall inflammatory potential. IMPORTANCEDeforestation, urbanization, and homelessness lead to spikes in Rickettsioses. Vector-borne human pathogens of Transitional Group (TRG), Typhus Group (TG), and Spotted Fever Group (SFG) rickettsiae differ by clinical manifestations, immunopathology, genome composition, and morphology. We previously showed that lipid A (or endotoxin), the membrane anchor of Gram-negative bacterial lipopolysaccharide (LPS), structurally differs in R. rickettsii (later-evolving SFG) relative to R. montanensis (basal SFG), R. typhi (TG), and R. akari (TRG). As lipid A structure influences recognition potential in vertebrate LPS sensors, further assessment of Rickettsia lipid A structural heterogeneity is needed. Here, we sidestepped the difficulty of ex vivo lipid A chemical extraction by utilizing FLATn, a new procedure for generating lipid A structures directly from host cell-purified bacteria. These data confirm later-evolving SFG pathogens synthesize structurally distinct lipid A. Our findings impact interpreting immune responses to different Rickettsia pathogens and utilizing lipid A adjuvant or anti-inflammatory properties in vaccinology.

microbiology↗

bioRxiv2022

Surface anchoring of the Kingella kingae galactan is dependent on the lipopolysaccharide O-antigen

Kingella kingae is a leading cause of bone and joint infections and other invasive diseases in young children. A key K. kingae virulence determinant is a secreted exopolysaccharide that mediates resistance to serum complement and neutrophils and is required for full pathogenicity. The K. kingae exopolysaccharide is a galactofuranose homopolymer called galactan and is encoded by the pamABC genes in the pamABCDE locus. In this study, we sought to define the mechanism by which galactan is tethered on the bacterial surface, a prerequisite for mediating evasion of host immune mechanisms. We found that the pamD and pamE genes are glycosyltransferases and are required for synthesis of an atypical lipopolysaccharide (LPS) O-antigen. The LPS O-antigen in turn is required for anchoring of galactan, a novel mechanism for association of an exopolysaccharide with the bacterial surface. SignificanceKingella kingae is an emerging pediatric pathogen and produces invasive disease by colonizing the oropharynx, invading the bloodstream, and disseminating to distant sites. This organism produces a uniquely multifunctional exopolysaccharide called galactan that is critical for virulence and promotes intravascular survival by mediating resistance to serum and neutrophils. In this study, we established that at least some galactan is anchored to the bacterial surface via a novel structural interaction with an atypical lipopolysaccharide O-antigen. Additionally, we demonstrated that the atypical O-antigen is synthesized by the pamD and pamE genes, located downstream of the gene cluster responsible for galactan biosynthesis. This work addresses how the K. kingae exopolysaccharide can mediate innate immune resistance and advances understanding of bacterial exopolysaccharides and lipopolysaccharides.

microbiology↗

bioRxiv2021

Enhanced longitudinal differential expression detection in proteomics with robust reproducibility optimization regression

Quantitative proteomics has matured into an established tool and longitudinal proteomic experiments have begun to emerge. However, no effective, simple-to-use differential expression method for longitudinal proteomics data has been released. Typically, such data is noisy, contains missing values, has only few time points and biological replicates. To address this need, we provide a comprehensive evaluation of several existing differential expression methods for high-throughput longitudinal omics data and introduce a new method, Robust longitudinal Differential Expression (RolDE). The methods were evaluated using nearly 2000 semi-simulated spike-in proteomic datasets and a large experimental dataset. The RolDE method performed overall best; it was most tolerant to missing values, displayed good reproducibility and was the top method in ranking the results in a biologically meaningful way. Furthermore, contrary to many approaches, the open source RolDE does not require prior knowledge concerning the types of differences searched, but can easily be applied even by non-experienced users.

bioinformatics↗

bioRxiv2020

Host Adaptation Predisposes Pseudomonas aeruginosa to Type VI Secretion System-Mediated Predation by the Burkholderia cepacia Complex

Pseudomonas aeruginosa (Pa) and Burkholderia cepacia complex (Bcc) species are opportunistic lung pathogens of individuals with cystic fibrosis (CF). While Pa can initiate long-term infections in younger CF patients, Bcc infections only arise in teenagers and adults. Both Pa and Bcc use type VI secretion systems (T6SS) to mediate interbacterial competition. Here, we show that Pa isolates from teenage/adult CF patients, but not those from young CF patients, are outcompeted by the epidemic Bcc isolate Burkholderia cenocepacia strain AU1054 (BcAU1054) in a T6SS-dependent manner. The genomes of susceptible Pa isolates harbor T6SS-abrogating mutations, the repair of which, in some cases, rendered the isolates resistant. Moreover, seven of eight Bcc strains outcompeted Pa strains isolated from the same patients. Our findings suggest that certain mutations that arise as Pa adapts to the CF lung abrogate T6SS activity, making Pa and its human host susceptible to potentially fatal Bcc superinfection.

microbiology↗