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Impact of Strain Varia on on the Ability of Biosensor Technology to Detect Salmonella enterica Jean Guard, U. S. Department of Agriculture, Agricultural Research Service, Athens GA
COLONY MORPHOLOGIES
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20125
20126
20127
20131
20136
21035
21036
21037
21038
21064
22079
28016
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28033
100616-97
100616-100
100616-102
100709-07
100723-04
. A. SefD‐ wt 22079 SE PT4, B. efD‐ 29108, C. SefD+ 100713.
Yellow indicates similar results. Red indicates significant varia on. A. SefD‐ wt 22079 SE PT4
vs SefD‐ 29108 B. SefD‐ 29108 vs SefD‐ wt 22079
with transforming plasmid but no sefD.
C. SefD+ 100713.
A
B C
FIG 1. Compara ve metabolic pa erns of
S. Enteri dis that varies in sefD
TRANSMISSION ELECTRON SPECTROPHY
FIG 2. Single cell and colony morphologies of
S. Enteri dis that varies in sefD
TRIAL 1 Impact of cell density on detec on of different
Salmonella enterica serotypes
TRIAL 2 Impact of the fimb r i al gene sefD on
detec on of S. Enteri dis
A
B
C
TRIAL 3 Evalua on of other mutants of
S. Enteri dis by biosensor
APPROACH [TRIAL 1] Analysis of various serotypes of Salmonella enterica. Salmonella enterica serovar Enteri dis (S. Enteri dis) Salmonella enterica serovar Gallinarum (S. Gallinarum) Salmonella enterica serovar Typhimurium (S. Typhimurium) Salmonella enterica 1,4,[5],12:I,‐ (S. Typhimurium variant) Salmonella enterica serovar Heidelberg (S. Heidelberg) [TRIAL 2] Analysis of strains from S. Enteri dis that vary in sefD. STRAIN 22079 (sefD‐) S. Enteri dis with sefD gene but nega ve for transcrip on. STRAIN 29108 (sefD‐) Mutant with sefD disrupted. STRAIN 100713 (sefD+) Mutant with sefD restored on a plasmid that has cons tu ve expression. [TRIAL 3] Analysis of knockout mutants of S. Enteri dis.
RESULTS [TRIAL 1] Detec on of Salmonella enterica occurs within an op mal range of approximately 102 to 107
CFU. Thus, too high of an organic load or too few cells are likely to give a nega ve result. The lower limit of detec on was between 30 to 300 cells. [TRIAL 2] Cell surface proper es has poten al to interfere with biosensor approaches. Cons tu ve expression by S. Enteridis of sefD, which encodes a fim
br i al protein, decreased biosensor signal. In
addi on, cons tu ve expression inhibited metabolic proper es (FIG 1) as well as changing cell size, shape and surface characteris cs (FIG 2). [TRIAL 3] Single gene knockout muta ons usually do not impede detec on of Salmonella by biosensor. A set of muta ons that knocked out flagella on, produc on of lipopolysaccharide O‐an gen and other func ons did not impede detec on.
INTRODUCTION
Methods that can quickly and accurately detect the presence of harmful bacteria associated with food borne disease are needed.
Salmonella enterica is a major food borne pathogen. It has over 2500 serotypes plus strains that vary in virulence within serotype.
Methods that target cell surface structures should be tested against many serotypes and strains to determine limits of detec on.
PURPOSE
Test the ability of an body‐based biosensor technology to detect serotypes and strains of Salmonella enterica known to vary in growth proper es and cell surface structures.
Inves gate limits of detec on for Salmonella enterica.
METHOD
CANARY® (Cellular Analysis and No fica on of An gen Risks and Yields) technology was developed by scien sts at Massachuse s Ins tute of Technology and published in Science.
CANARY® biosensors (PathSensors) are B‐cell lines that express monoclonal an bodies and the aequorin protein. Luminescence is generated when the an bodies bind to their an gen.
We tested CANARY® biosensor technology for the ability to detect nine serotypes of Salmonella enterica and 18 strains of Salmonella enterica serovar Enteri dis (S. Enteri dis), which is the world’s leading serotype associated with human food borne salmonellosis.
SIGNIFICANCE
Salmonella enterica cell concentra ons outside of an op mal range of 102 to 107 CFU will impede detec on. OF 27 strains tested only one evaded detec on and it was a laboratory construct not found in nature. CANARY® technology is otherwise rapid and accurate, and it collects living culture for genomic analyses. Further knowledge about specific
mo l ecul es tar get ed by bi os ens or is needed to ma ke it ser ot ype speci fic.
USE OF COMMERCIALLY AVAILABLE PRODUCTS OR TECHNOLOGY BY USDA DOES NOT CONSTITUTE OFFICIAL ENDORSEMENT