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Joseph J. Gillespie, PhD

Academic Title:

Assistant Professor

Primary Appointment:

Microbiology and Immunology


HSF1, 363

Education and Training


Dr. Gillespie is an evolutionary biologist with broad interests in organismal and molecular evolution.  The major focus of his current research is deciphering the mechanisms by which obligate intracellular species of Rickettsiales (Alphaproteobacteria) invade, survive and replicate within eukaryotic cells.

In research funded by the National Institutes of Health, Dr. Gillespie utilizes phylogenetics, comparative genomics and bioinformatics to guide experimental studies on various pathogenic species of Rickettsia and their associated arthropod vectors.  His early research resulted in the reclassification of Rickettsia species and the identification of many lineage-specific pathogenicity factors.  Through years of intense scrutinization of dozens of diverse rickettsial genomes, Dr. Gillespie and colleagues have described a large, dynamic mobilome for Rickettsia species, resulting in the identification of integrative conjugative elements as the vehicles for seeding Rickettsia genomes with many of the factors underlying obligate intracellular biology and pathogenesis.  Via an iterative process of genome sequencing, phylogenomics, bioinformatics, and classical molecular biology and microbiology, Dr. Gillespie continues to lead and assist research projects on the characterization of rickettsial gene and protein function, as well as the description of cell envelope glycoconjugates, such as peptidoglycan and lipopolysaccharide.

A particular focal area of Dr. Gillespie’s research is the Rickettsia secretome, which includes the secretion systems and their cognate substrates, many of which directly engage arthropod and vertebrate molecules throughout the rickettsial infection process.  Dr. Gillespie led a study that identified across all genera of Rickettsiales the composition of an enigmatic type IV secretion system (T4SS), termed rvh (Rickettsiales vir homolog).  Several collaborative efforts have begun elucidating the odd architecture of the rvh T4SS, as well as the protein substrates (e.g., RalFRARP-2, and Risk1) it translocates into host cells during infection. A recent analysis of rickettsial metagenomes provided substantial insight on the evolution of effector architectures.

Research/Clinical Keywords

Evolutionary Biology, Rickettsiology, Rickettsia, Microbiology, Molecular Biology, Genetics, Bioinformatics, Phylogenetics, Phylogenomics, Metabolomics, Structural Biology, Lateral Gene Transfer, Type IV Secretion Systems, Secretome, Pathogenesis, Genome Sequencing, Arthropods, Ticks, Fleas, Body Lice, Epidemic Typhus, Murine Typhus, Rocky Mountain Spotted Fever, Transitional Group rickettsiae, Wolbachia

Highlighted Publications


Oyler B.L., Salje J., Rennoll-Bankert K.E., Verhoeve V.I., Rahman M.S., Skerry C., Azad A.F., Goodlett D.R., Gillespie J.J.     (2024)     Evidence for canonical Gram-negative bacterial peptidoglycan structure and murein sacculi in Rickettsia pathogens. View.

Lehman, S.S., Verhoeve, V.I., Driscoll, T.P.,  Beckmann, J.F., Gillespie, J.J.      (2024)       Metagenome diversity illuminates origins of pathogen effectorsmBio 2:e0075923. [EDITOR'S PICK].

Hofstaedter, C.E., Chandler, C.E., Met, C.M., Gillespie, J.J., Harro, J.M., Goodlett, D.R., Rasko, D.A., Ernst, R.K.     (2024)     Divergent  Pseudomonas aeruginosa  LpxO enzymes perform site-specific lipid A 2-hydroxylationmBio. 15:e0282323.

Yang, H., Verhoeve, V.I., Chandler, C.E., Nallar, S., Snyder, G.A., Ernst, R.K., Gillespie, J.J.     (2024)     Structural determination of Rickettsia lipid A without chemical extraction confirms shorter acyl chains in later-evolving Spotted Fever Group pathogens. mSphere 9:e0060923.

Giengkam, S., Kullapanich, C. Wongsantichon, J., Adcox H.E., Gillespie, J.J., Salje, J.       (2023)      Orientia tsutsugamushi: analysis of the mobilome of a highly fragmented and repetitive genome reveals ongoing lateral gene transfer in an obligate intracellular bacterium. mSphere 18:e0026823.

Beckmann, J.F.,   Gillespie, J.J., Tauritz, D.       (2023)       Modelling emergence of Wolbachia toxin-antidote protein functions with an evolutionary algorithm.  Frontiers in Microbiology (Microbial Symbioses) 14.

Gillespie, J.J., Salje, J.       (2023)     Orientia and Rickettsia: different flowers from the same gardenCurrent Opinion in Microbiology 74: 102318.

Verhoeve, V.I.,   Gillespie, J.J.       (2022)        Origin of rickettsial host dependency unravelled.     Nature Microbiology 7.

Verhoeve VI, Brammer JA, Driscoll TP, Kambouris AR, Rasko DA, Cross AS, Gillespie, J.J.       (2022)         Genome sequencing of Pseudomonas aeruginosa strain M2 illuminates traits of an opportunistic pathogen of burn wounds. G3 12.

Verhoeve V.I., Fauntleroy T.D., Risteen R.G., Driscoll, T.P.,    Gillespie, J.J.       (2022)         Cryptic Genes for Interbacterial Antagonism Distinguish Rickettsia Species Infecting Blacklegged Ticks From Other Rickettsia Pathogens.       Front Cell Infect Microbiol12: 880813.

Atwal, S., Chuenklin, S., Bonder, E.M., Flores, J., Gillespie, J.J., Driscoll, T.P., Salje, J.     (2021)     Discovery of a Diverse Set of Bacteria That Build Their Cell Walls without the Canonical Peptidoglycan Polymerase aPBP.   mBio. 12: e0134221.
Guillotte, M.L., Chandler, C.E., Verhoeve, V.I.,   Gillespie, J.J., Driscoll, T.P., Rahman, M.S., Ernst, R.K., Azad, A.F.      (2021)          Lipid A Structural Divergence in Rickettsia Pathogens.   mSphere 6: e00184-21.


Gillespie, J.J., Driscoll, T.P., Verhoeve, V.I., Rahman, M.S., Macaluso, K.R., Azad, A.F.  (2018)   A tangled web: origins of reproductive parasitism. Genome Biology and Evolution 10: 2292-2309.
Driscoll, T.P., Verhoeve, V.I., Guillotte, M.L., Lehman, S.S., Rennoll, S.A., Beier, M.S., Rahman, M.S., Azad, A.F., Gillespie, J.J.  (2017)  Wholly Rickettsia! Reconstructed Metabolic Profile of the Quintessential Bacterial Parasite of Eukaryotic CellsMBio 8: e00859-17.
Gillespie, J.J., Phan, I.Q., Driscoll, T., Guillotte, M.L., Lehman, S.S., Rennoll-Bankert, K.E., Subramanian, S., Beier-Sexton, M., Myler, P.J., Rahman, M.S., Azad, A.F.  (2016)  The Rickettsia type IV secretion system: unrealized complexity mired by gene family expansionPathogens and Disease 74: ftw058.
Gillespie, J.J., Kaur, S.J., Rahman, M.S., Rennoll-Bankert, K., Sears, K.T., Beier-Sexton, M. & Azad, A.F.  (2015)  Secretome of obligate intracellular RickettsiaFEMS Microbiology Reviews 39: 47-80.
Gillespie, J.J., Phan IQ, Scheib H, Subramanian, S., Edwards, T.E., Lehman, S.S., Piitulainen, H., Rahman, M.S., Rennoll-Bankert, K., Staker, B.L., Taira, S., Stacy, R., Myler, P.J., Azad, A.F., & Pulliainen, A.T.  (2015)  Structural Insight into How Bacteria Prevent Interference between Multiple Divergent Type IV Secretion SystemsMBio 6: e01867-15.

Gillespie, J.J., Joardar, V., Williams, K.P., Driscoll, T., Hostetler, J.B., Nordberg, E.K., Shukla, M., Walenz, B., Hill, C.A., Nene, V.M., Azad, A.F., Sobral, B.W. & Caler, E.  (2012)  A Rickettsia genome overrun by mobile genetic elements provides insight into the acquisition of genes characteristic of an obligate intracellular lifestyleJournal of Bacteriology 194: 376-394.

Gillespie, J.J., Beier, M.S., Rahman, M.S., Ammerman, N.C., Purkayastha, A., Shallom, J.M., Sobral, B.S., Azad, A.F.  (2007)  Plasmids and rickettsial evolution: insight from Rickettsia felis. PLoS ONE 2: e266.

Additional Publication Citations

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


Google Scholar


Research Interests

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. Prior 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. Dr. Gillespie and a team led by colleague Dr. Jeanne Salje (University of Cambridge) recently determined that RAGEs are also actively transferring between genomes of the agent of Scrub Typhus, Orientia tsutsugamushi. Rickettsia and Orientia species are closely related rickettsial lineages that differ markedly in their biology and pathogenesis, allowing for many key insights through comparative analyses.

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 RalFRARP-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, or CI). Parasites induce RP to drive their retention in host populations via the female germline. RP, particularly 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. Dr. Gillespie collaborates with Dr. John Beckmann, one of the pioneers for discovering the genetic basis for CI. A recent study developed a model to better understand the evolution of CI genes in arthropod populations. Furthermore, the team is currently characterizing CI-like genes of Orientia tsutsugamushi, which induces parthenogenesis in trombiculid mites.


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 provide instances of lateral gene transfers that could drive transitions between parasitism and mutualism.

Awards and Affiliations

Grants and Contracts


R21 AI164730
Vadyvaloo (PI), Eckstrand (Co-investigator), Gillespie (Co-investigator)
2/01/23 - 2/01/25
Emerging understanding of the rat flea response to Yersinia pestis infection

R21 AI166832
Gillespie (PI), Ernst (PI)
7/01/22 - 6/30/24
Investigating Rickettsia Interspecies and Host-Specific Lipopolysaccharide Variation



R21 AI156762
Gillespie (PI), Driscoll (PI)
1/22/21 - 12/31/22
Characterizing the Pan-genome of a Rickettsia Infecting the Eastern Black-legged Tick

R21 AI146773
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 AI126108
Gillespie (PI), Rahman (PI)
5/23/16 - 4/30/19
Characterizing gene family expansion in an atypical bacterial secretion system

Community Service

  • University of Maryland School of Medicine Council
  • Chair, GPILS  Awards Committee (2022-present)
  • UMB Molecular Microbiology and Immunology Admissions Committee
  • UMB Molecular Microbiology and Immunology Curriculum Committee
  • UMB Education Technology Steering Committee
  • Course Director and Instructor for GPLS 601: Mechanisms in Biomedical Sciences (The Core Course)
  • Course Director and Instructor for GPLS 710: Principles of Microbial Pathogenesis
  • Former Course Director and Instructor for GPLS 725: Systems Level Research in Microbial Pathogenesis
  • Instructor for GPILS 693: Introduction to Molecular Microbiology and Immunology
  • Instructor for RCR, CIPP907: Responsible Conduct of Research
  • Judge for School of Medicine Annual Graduate Research Conference
  • Judge for Microbiology and Immunology Graduate Research Competition
  • Judge for School of Medicine Medical Student Research Day

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.

Professional Activity

Links of Interest

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".

Bacterial and Viral Bioinformatics Resource Center (BV-BRC), an information system designed to support research on bacterial and viral infectious diseases. The BV-BRC combines the data and tools from the Legacy BRC resources: PATRIC, the bacterial BRC, and IRD and ViPR, the viral BRCs. Previously, 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.



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

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 and entertainment.

Dr. Bob Ernst and his laboratory in the School of Dentistry, University of Maryland Baltimore, collaborate with Dr. Gillespie on deciphering Rickettsia lipid A structure, as well as on several phylogenomics projects analyzing lipid A modification systems.

Dr. Alan Cross and his laboratory in the Department of Medicine and Center for Vaccine Development and Global Health, University of Maryland Baltimore, collaborate with Dr. Gillespie on structural analysis of Rickettsia LPS and S-layer glycoconjugates.

Dr. Dave Goodlett and his laboratory at The University of Victoria collaborate with Dr. Gillespie on determining Rickettsia peptidoglycan structure and dynamics.

Dr. Greg Snyder (Institute of Human Virology) collaborates with Dr. Gillespie on protein structural biology.

Dr. Viveka Vadyvaloo and her laboratory at Washington State University collaborate with Dr. Gillespie on characterizing flea transcriptional responses to Yersinia and Rickettsia pathogens.

Dr. Anders Omsland and his laboratory at Washington State University collaborate with Dr. Gillespie on developing axenic culture media to support cell-free Rickettsia growth.

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.


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
AlphaFold Protein Structure Database

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).


Dr. Gillespie and Dr. Verhoeve recently participated in the American Cancer Society-Diversity in Cancer Research Internship Program and trained Trinity Soto, a rising senior at Towson University (link to video)