A spontaneous mutation in kdsD, a biosynthesis gene for 3 Deoxy-D-manno-Octulosonic Acid, occurred in a ciprofloxacin resistant strain of Francisella tularensis and caused a high level of attenuation in murine models of tularemia.
Francisella tularensis, a gram-negative facultative intracellular bacterial pathogen, is the causative agent of tularemia and able to infect many mammalian species, including humans. Because of its ability to cause a lethal infection, low infectious dose, and aerosolizable nature, F. tularensis subspecies tularensis is considered a potential biowarfare agent. Due to its in vitro efficacy, ciprofloxacin is one of the antibiotics recommended for post-exposure prophylaxis of tularemia. In order to identify therapeutics that will be efficacious against infections caused by drug resistant select-agents and to better understand the threat, we sought to characterize an existing ciprofloxacin resistant (CipR) mutant in the Schu S4 strain of F. tularensis by determining its phenotypic characteristics and sequencing the chromosome to identify additional genetic alterations that may have occurred during the selection process. In addition to the previously described genetic alterations, the sequence of the CipR mutant strain revealed several additional mutations. Of particular interest was a frameshift mutation within kdsD which encodes for an enzyme necessary for the production of 3-Deoxy-D-manno-Octulosonic Acid (KDO), an integral component of the lipopolysaccharide (LPS). A kdsD mutant was constructed in the Schu S4 strain. Although it was not resistant to ciprofloxacin, the kdsD mutant shared many phenotypic characteristics with the CipR mutant, including growth defects under different conditions, sensitivity to hydrophobic agents, altered LPS profiles, and attenuation in multiple models of murine tularemia. This study demonstrates that the KdsD enzyme is essential for Francisella virulence and may be an attractive therapeutic target for developing novel medical countermeasures.
Pathology Division, United States Army Medical Research Institute of Infectious Diseases (USAMRIID), Fort Detrick, Frederick, MD, United States of America.
Taylor
T
Chance
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Jennifer
http://orcid.org/0000-0001-8156-5174
J
Chua
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Ronald G
RG
Toothman
Center for Genome Sciences, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Jason T
http://orcid.org/0000-0001-7751-4366
JT
Ladner
Department of Molecular and Translational Sciences, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Jonathan E
JE
Nuss
Pathology Division, United States Army Medical Research Institute of Infectious Diseases (USAMRIID), Fort Detrick, Frederick, MD, United States of America.
Jo Lynne
JL
Raymond
Institut de Recherche Biomédicale des Armées, Département de Biologie des Agents Transmissibles, Unité de Bactériologie/UMR_MD1, B.P. 73, Brétigny-sur-Orge, France.
Fabrice V
FV
Biot
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Samandra
S
Demons
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Lynda
L
Miller
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Stephanie
S
Halasohoris
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Sherry
S
Mou
Center for Genome Sciences, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Galina
G
Koroleva
Center for Genome Sciences, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Sean
S
Lovett
Center for Genome Sciences, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Gustavo
G
Palacios
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Nicholas J
NJ
Vietri
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Patricia L
PL
Worsham
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Christopher K
CK
Cote
Department of Molecular and Translational Sciences, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Todd M
TM
Kijek
Bacteriology Division, USAMRIID, Fort Detrick, Frederick, MD, United States of America.
Joel A
JA
Bozue
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics*; D002939:Ciprofloxacin / Q000494:pharmacology; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D005604:Francisella tularensis / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D009154:Mutation / Q000235:genetics*; D056990:Post-Exposure Prophylaxis / Q000379:methods; D013400:Sugar Acids / Q000378:metabolism*; D014406:Tularemia / Q000382:microbiology*; D014774:Virulence / Q000235:genetics
D001426:Bacterial Proteins; D008070:Lipopolysaccharides; D013400:Sugar Acids; C002532:2-keto-3-deoxyoctonate; D002939:Ciprofloxacin