title: A Burkholderia pseudomallei deltapurM mutant is avirulent in immunocompetent and immunodeficient animals: candidate strain for exclusion from select-agent lists.
abstract: Burkholderia pseudomallei causes the disease melioidosis in humans and is classified as a category B select agent. Research utilizing this pathogen is highly regulated in the United States, and even basic studies must be conducted in biosafety level 3 (BSL-3) facilities. There is currently no attenuated B. pseudomallei strain available that is excluded from select-agent regulations and can be safely handled at BSL-2 facilities. To address this need, we created Bp82 and Bp190, which are DeltapurM derivatives of B. pseudomallei strains 1026b and K96243 that are deficient in adenine and thiamine biosynthesis but replication competent in vitro in rich medium. A series of animal challenge studies was conducted to ensure that these strains were fully attenuated. Whereas the parental strains 1026b and K96243 and the complemented mutants Bp410 and Bp454 were virulent in BALB/c mice following intranasal inoculation, the DeltapurM mutants Bp82 and Bp190 were avirulent even when they were administered at doses 4 logs higher than the doses used for the parental strains. Animals challenged with high doses of the DeltapurM mutants rapidly cleared the bacterium from tissues (lung, liver, and spleen) and remained free of culturable bacteria for the duration of the experiments (up to 60 days postinfection). Moreover, highly susceptible 129/SvEv mice and immune incompetent mice (IFN-gamma-/-, SCID) were resistant to challenges with DeltapurM mutant Bp82. This strain was also avirulent in the Syrian hamster challenge model. We concluded that DeltapurM mutant Bp82 is fully attenuated and safe for use under BSL-2 laboratory conditions and thus is a candidate for exclusion from the select-agent list.
authors.affiliation: Department of Microbiology, Immunology and Pathology, Rocky Mountain Regional Center of Excellence for Biodefense and Emerging Infectious Diseases Research, Colorado State University, Fort Collins, Colorado 80523-2025, USA.
authors.forename: Katie L
authors.initials: KL
authors.lastname: Propst
authors.forename: Takehiko
authors.initials: T
authors.lastname: Mima
authors.forename: Kyoung-Hee
authors.initials: KH
authors.lastname: Choi
authors.forename: Steven W
authors.initials: SW
authors.lastname: Dow
authors.forename: Herbert P
authors.initials: HP
authors.lastname: Schweizer
mesh_terms: D000818:Animals; D016957:Burkholderia pseudomallei / Q000235:genetics / Q000276:immunology / Q000472:pathogenicity*; D006224:Cricetinae; D005260:Female; D017353:Gene Deletion; D005798:Genes, Bacterial / Q000235:genetics / Q000502:physiology; D008099:Liver / Q000276:immunology / Q000382:microbiology; D008168:Lung / Q000276:immunology / Q000382:microbiology; D008554:Melioidosis / Q000276:immunology / Q000382:microbiology*; D008647:Mesocricetus; D051379:Mice; D008807:Mice, Inbred BALB C; D016513:Mice, SCID; D016296:Mutagenesis / Q000235:genetics; D013154:Spleen / Q000276:immunology / Q000382:microbiology
references.citation: Infect Immun. 2006 Sep;74(9):5333-40
references.pmid: 16926428
references.citation: Infect Immun. 2005 Feb;73(2):1106-15
references.pmid: 15664954
references.citation: Emerg Infect Dis. 2005 Oct;11(10):1638-9
references.pmid: 16355505
references.citation: Appl Environ Microbiol. 2008 Feb;74(4):1064-75
references.pmid: 18156318
references.citation: Nat Rev Microbiol. 2006 Apr;4(4):272-82
references.pmid: 16541135
references.citation: MMWR Morb Mortal Wkly Rep. 2006 Aug 18;55(32):873-6
references.pmid: 16915220
references.citation: Infect Immun. 2009 Apr;77(4):1579-88
references.pmid: 19179415
references.citation: Infect Immun. 2005 Feb;73(2):1260-4
references.pmid: 15664977
references.citation: Infect Immun. 1999 Jul;67(7):3593-600
references.pmid: 10377144
references.citation: Emerg Infect Dis. 2008 Jul;14(7):e2
references.pmid: 18598617
references.citation: J Immunol. 2010 Feb 1;184(3):1445-54
references.pmid: 20042590
references.citation: Infect Immun. 1999 Jun;67(6):2867-73
references.pmid: 10338493
references.citation: Microbiology (Reading). 2004 Aug;150(Pt 8):2669-2676
references.pmid: 15289563
references.citation: Appl Environ Microbiol. 2008 May;74(10):2985-9
references.pmid: 18310423
references.citation: J Bacteriol. 1997 Apr;179(7):2116-25
references.pmid: 9079894
references.citation: Proc Natl Acad Sci U S A. 2004 Sep 28;101(39):14240-5
references.pmid: 15377794
references.citation: Infect Immun. 2003 Dec;71(12):7002-13
references.pmid: 14638790
references.citation: Trans R Soc Trop Med Hyg. 2008 Dec;102 Suppl 1:S89-94
references.pmid: 19121696
references.citation: Clin Infect Dis. 2006 Oct 15;43(8):979-86
references.pmid: 16983608
references.citation: J Med Microbiol. 2002 Jul;51(7):539-553
references.pmid: 12132769
references.citation: N Engl J Med. 1981 Nov 5;305(19):1133-5
references.pmid: 7290119
references.citation: PLoS One. 2008 Apr 30;3(4):e2058
references.pmid: 18446194
references.citation: Southeast Asian J Trop Med Public Health. 2009 Jan;40(1):123-30
references.pmid: 19323044
references.citation: Expert Rev Anti Infect Ther. 2006 Jun;4(3):445-55
references.pmid: 16771621
references.citation: Trans R Soc Trop Med Hyg. 2008 Dec;102 Suppl 1:S1-4
references.pmid: 19121666
references.citation: Scand J Infect Dis. 2006;38(1):71-4
references.pmid: 16338844
references.citation: Clin Microbiol Rev. 2005 Apr;18(2):383-416
references.pmid: 15831829
references.citation: Infect Immun. 2003 Jul;71(7):3960-70
references.pmid: 12819083
references.citation: Infect Immun. 2004 Aug;72(8):4911-7
references.pmid: 15271960
references.citation: Clin Infect Dis. 2005 Oct 1;41(7):982-90
references.pmid: 16142663
references.citation: Infect Immun. 2006 Aug;74(8):4452-61
references.pmid: 16861631
references.citation: Infect Immun. 2001 Jan;69(1):34-44
references.pmid: 11119486
references.citation: Appl Environ Microbiol. 2009 Oct;75(20):6496-503
references.pmid: 19700544
references.citation: Infect Immun. 2002 Sep;70(9):5290-4
references.pmid: 12183585
references.citation: Infect Immun. 2006 Jun;74(6):3576-86
references.pmid: 16714590
references.citation: Infect Immun. 2006 Oct;74(10):5465-76
references.pmid: 16988221
references.citation: Emerg Infect Dis. 2008 Nov;14(11):1689-92
references.pmid: 18976550
references.citation: J Chemother. 2006 Aug;18(4):443-4
references.pmid: 17024805
references.citation: Eur Respir J. 2003 Sep;22(3):542-50
references.pmid: 14516149