Joseph J. Gillespie, PhD

Dr. Gillespie's publications and related information can be found on the following web sites:

NCBI

Google Scholar

ORCID

Sequencing of the deer tick (Ixodes scapularis) genome revealed the nature of a Rickettsia endosymbiont that is frequently found in deer tick populations. Genomic analysis of this species, now called Rickettsia buchneri, unearthed a large and diverse cache of mobile genetic elements. Prominent among these is a distinct integrative and conjugative element named RAGE (Rickettsiales Amplified Genetic Element). Dr. Gillespie and colleagues determined that RAGEs are the vehicles for seeding Rickettsia genomes with many of the factors underlying obligate intracellular biology and pathogenesis. Recent analyses discovered substantial diversity among R. buchneri RAGEs, both within individual ticks as well as in the I. scapularis population at large, indicating that RAGEs are actively mobilizing across Rickettsia species.

This RAGE-mediated lateral gene transfer has steered research and discovery in three main focal areas:

1. Virulence Factors

The search for Rickettsia secreted effectors that target host cell molecules begins with detecting probable products of lateral gene transfer (e.g., piggybacking on RAGEs, phylogenetic signal discordant with “housekeeping” genes) that may contain eukaryotic-like characteristics (e.g., ankyrin repeats, Sec7-like domains, phosphatidylinositol kinase motifs). Dr. Gillespie and colleagues then aim to identify the rickettsial secretion pathway involved in translocating such effectors. A major focus is the rvh T4SS, wherein effectors must bind the rvh coupling protein (RvhD4) prior to secretion. Several approaches are used to capture RvhD4-effector interactions, with subsequent characterization of effectors using in vitro and in vivo experiments. The effectors RalF, RARP-2 and Risk1 have been shown to be translocated into host cells by Rickettsia typh during infection, and other candidate effectors are currently being characterized.

2. Metabolism

Many of the RAGE cargo genes encode for proteins that are seemingly important for intracellular survival (e.g., osmoregulatory transporters, lipopolysaccharide-modifying enzymes, peptidoglycan recycling factors) and host-dependent metabolism (stringent response components, metabolite transporters, metabolic enzymes). As rickettsiae are subject to reductive genome evolution, the RAGEs appear to offset this deleterious process by shuttling across the rickettsial mobilome genes that are critical for intracellular parasitism. The importance for maintaining genes underpinning metabolism was realized through reconstructing the Rickettsia-host metabolic network, which revealed 51 host cell metabolites required to compensate the patchwork rickettsial metabolism. Dr. Gillespie and colleagues are studying the novel mechanisms utilized by rickettsiae to pilfer host metabolites from the eukaryotic cytoplasm. A major focus entails solving how rickettsiae steal amino sugars to supply building blocks for cell envelope synthesis (rickettsiae lack glycolysis and have quite a sweet tooth!).

3. Reproductive Parasitism

RAGEs have also been shown to shuttle genes implicated in reproductive parasitism (RP), which is the ability of microbial parasites to influence the sexual reproduction of their hosts (via processes such as male-killing, feminization, parthenogenesis, and cytoplasmic incompatibility). Parasites induce RP to drive their retention in host populations via the female germline. RP, particularly cytoplasmic incompatibility (CI), is well-studied for Wolbachia parasites, which are close cousins to Rickettsia species and infect over half the world’s described arthropod species, as well as many nematode species. While the molecular basis for Wolbachia-mediated CI was discovered by Wolbachia researchers (classical toxin-antidote operons), Dr. Gillespie and colleagues identified divergent orthologs to these factors in a range of different intracellular parasites, including human pathogens (i.e., Rickettsia felis, Rickettsia gravesii and Orientia tsutsugamushi). Current studies are focused on elucidating the mechanisms of RP induction by these diverse toxin-antidote operons, as well as identifying novel reproductive parasites of the cat flea, a vector that transmits pathogenic rickettsiae that scourge the homeless population.

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Genome sequencing of the cat flea (Ctenocephalides felis) revealed a bizarre “genome in flux” characterized by inordinate copy number variation (~38% of proteins) and a broad range of genome size estimates (433-551 Mb) for individual fleas.  Surprisingly, the flea genome exhibits neither inflation due to rampant gene duplication nor reduction due to the flea parasitic lifestyle.  Based on these current data, Dr. Gillespie and colleagues posit that a dual mechanism of unequal crossing-over and gene conversion underpins flea genome variability (although the biological significance remains unknown).  Further investigations of flea genome maintenance, regulation and evolution are on-going.  The cat flea mitochondrial genome has no copy number variation, limiting this phenomenon to siphonapteran nuclear genomes.

Dr. Gillespie and colleagues also determined that cat fleas are often co-infected with divergent strains of Wolbachia (an intracellular bacterium related to Rickettsiae).  From the generated C. felis sequence reads, closed genomes for two novel Wolbachia strains were assembled, annotated and analyzed.  Both genomes contain laterally transferred genes that inform on the evolution of Wolbachia host associations.  wCfeT carries biotin synthesis genes, while wCfeJ carries a CI-inducing toxin-antidote operon.  Analyses of these genes highlight their mobility across the Wolbachia phylogeny and source to other intracellular bacteria.  Additional screening of geographically diverse cat fleas revealed predominant co-infection (wCfeT/wCfeJ) amongst C. felis colonies, though single wCfeT infection is the norm in wild populations.  Collectively, genomes of wCfeT and wCfeJ supply instances of lateral gene transfers that could drive transitions between parasitism and mutualism.

• 1998-2005: Member, Entomological Society of America
• 2000-present: Member, American Association for the Advancement of Science
• 2000-present: Member, Society for Molecular Biology and Evolution
• 2000-2003: Member, Society for the Study of Evolution
• 2000-present: Member, Society of Systematic Biologists
• 2004-2005: Member, European Society for Evolutionary Biology
• 2006-present: Member, American Society for Microbiology
• 2006-present: Member, American Society for Rickettsiology

R21 AI126108
NIH/NIAID
Gillespie (PI), Rahman (PI)
5/23/16 - 4/30/19
Characterizing gene family expansion in an atypical bacterial secretion system

R21 AI146773
NIH/NIAID
Gillespie (PI), Rahman (PI)
5/23/19 - 4/30/21
Rickettsia cell envelope glycoconjugates are derived from the host cell amino sugar biosynthesis pathway

R21 AI156762
NIH/NIAID
Gillespie (PI), Driscoll (PI)
12/01/20 - 11/31/22
Characterizing the Pan-genome of a Rickettsia Infecting the Eastern Black-legged Tick

Dr. Gillespie an ardent supporter of youth athletics, realizing the importance for introducing exercise and sports to children in a healthy and fun atmosphere.  He coaches annually in various baseball, basketball and soccer programs in the Lutherville-Timonium area.  He serves the community as the In-House Recreation Coordinator of the Lutherville-Timonium Soccer Club, and is a board member and commissioner in the Lutherville-Timonium Basketball Association.  Dr. Gillespie is also a certified referee and judge for the Central Maryland Dive League.

• 2017-present: Editorial Board, PeerJ
• 2019-present: Reviewer, ZRG1 IDM-B 80 Study Section (Topics in Bacterial Pathogenesis), NIH
• 2019: Site Visit Team Member for the review of Intramural Research Programs in the Tumor Vaccines and Biotechnology Branch (TVBB), Division of Cellular and Gene Therapies (DCGT), Office of Tissues and Advanced Therapies (OTAT), Center for Biologics Evaluation and Research (CBER), U.S. Food and Drug Administration, DHHS
• 2017-present: Scientific Review Committee, American Society for Rickettsiology
• 2020-present: Editorial Board, PLoS Neglected Tropical Diseases

NCBI, The National Center for Biotechnology Information, advances science and health by providing access to biomedical and genomic information.  NCBI's Pubmed is a repository for peer-reviewed science articles.  Just search by key words or phrases; e.g., try searching with "anti-vaccine misinformation".

PATRIC, the Pathosystems Resource Integration Center, provides integrated data and analysis tools to support biomedical research on bacterial infectious diseases.  Dr. Gillespie was a team member and contributor to PATRIC from 2006 to 2013.

Collaborators

Dr. Tim Driscoll and his Vector-borne Pathogen Dynamics Laboratory at West Virginia University collaborate with Dr. Gillespie on many different projects, but especially those involving arthropod and bacteria genome sequencing, phylogenomics, and arthropod-bacteria interactions.

Dr. John Beckmann and his laboratory at Auburn University collaborate with Dr. Gillespie on several projects involving reproductive parasitism.  Check out John's very inspiring art work.

Dr. Bob Ernst and Dr. Dave Goodlett collaborate with Dr. Gillespie on determining Rickettsia cell envelope glycoconjugate composition.

Dr. Kevin Macaluso and his laboratory at the University of Southern Alabama collaborate with Dr. Gillespie on projects involving fleas and flea-associated Rickettsia species.

Dr. Jeanne Salje and her laboratories at Rutgers University and Mahidol-Oxford Tropical Medicine Research Unit (MORU) in Bangkok, Thailand collaborate with Dr. Gillespie on projects analyzing rickettsial cell envelope biology and overall rickettsial phylogenomcis.

Luke J. Tallon, Scientific Director of the Genomics Resource Center, Institute for Genome Sciences, collaborates with Dr. Gillespie on the genome sequencing of the cat flea (Ctenocephalides felis) and its Wolbachia parasites.

Dr. Gillespie acknowledges other rickettsiologists (mentors, friends and collaborators) in the University of Maryland system: Dr. Abdu Azad, Dr. Sayeed Rahman, Dr. Joao Pedra, Dr. Sean Riley, and Dr. Utpal Pal.

Utilized Informatics Resources

Designing primers for PCR
Reverse-complement tool
Six frames DNA translation
Generate a CDS alignment from aligned proteins
Calculate positive selection

Protein sequence and functional information database
Searching for protein similarity
Build orthologous protein groups
Computing multiple sequence alignment
View and edit alignments
Manipulate/convert sequence formats
Masking multiple sequence alignments

Calculate pairwise divergences
Estimating phylogenies
Drawing phylogenetic trees

Repository for protein structures
Searching for structural analogs
Threading proteins to structures
Compare protein structures in 3D

Predict SEC and lipoprotein signal sequences
Predict transmembrane spanning regions
Predict protein domains and other features

Making sequence logos
Draw gene/protein schemas
Visualize genomic data and information in a circular format
Bin and visualize metagenomic data

Metabolic networks

Tool for parasitology

Learn about the bacterial world

Dr. Gillespie Interviewed on Netflix Series

Dr. Gillespie shares the dangers posed by Chigoe Flea infestations in this video segment from the Netflix series, "72 Dangerous Animals: Latin America" (Season 2, Episode 7)

Lab Training

Dr. Gillespie and Dr. Mark Guillotte engage in advanced laboratory training techniques (filmed by Dr. Stephanie Lehman).