Abstract/Details

Characterization of <i>Listeria innocua</i> biofilm formation using TN<i>917</i> transposon mutagenesis


2005 2005

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Abstract (summary)

Listeria sp. can grow on surfaces and be a member of multi-species biofilms within food processing plants, rendering them more resistant to sanitizers and other antimicrobial agents. Little is known of the cellular physiological processes involved in Listeria sp. growth on surfaces. The purpose of this study was to generate random transposon mutants in order to identify genetic determinants contributing to biofilm formation. Listeria innocua ATCC 51742 serotype 4b was selected for this study since it produces high levels of biofilm in a laboratory assay. Transposon mutagenesis was performed using plasmid pLTV3 (Tn917) with a temperature sensitive origin of replication. Mutants were selected by growth in the presence of erythromycin and lincomysin at 41°C. A library, consisting of 2500 mutants, was further screened by use of a PVC microtiter plate assay at 32°C at low nutrient levels. Reduced surface growth (RSG) mutants were defined as having a mean stained biofilm at 2 std dev below the mean of the wild-type strain. A total of 36 RSG mutants were isolated and further characterized by testing cellular motility and initial adhesion phenotypes. RSG mutants exhibited various phenotype profiles, suggesting a number of genes involved in biofilm formation of L. innocua. RSG 24 mutant was further characterized, showing an increased negative surface charge. RSG 24 mutant scanning electron microscopy and epifluorescence microscopy results showed reduced surface growth area coverage compared to wild-type. An arbitrary PCR, PCR product cloning and sequencing revealed that Tn917 transposition took place in lin2619, a gene homologous to mutarotase. Surface growth characteristics were restored to RSG 24 by complemetation of the lin2619 mutarotase gene in trans to levels comparable to the wild-type strain. Complementation in trans and results indicating altered cell surface characteristics, our study indicates that lin2619 is involved in L. innocua adhesion and surface growth.

Indexing (details)


Subject
Food science
Classification
0359: Food science
Identifier / keyword
Biological sciences; Biofilm; Listeria innocua; Transposon
Title
Characterization of <i>Listeria innocua</i> biofilm formation using TN<i>917</i> transposon mutagenesis
Author
Shaw, William K., Jr.
Number of pages
96
Publication year
2005
Degree date
2005
School code
0118
Source
DAI-B 66/02, Dissertation Abstracts International
Place of publication
Ann Arbor
Country of publication
United States
ISBN
9780496976966, 0496976966
Advisor
McLandsborough, L. A.
University/institution
University of Massachusetts Amherst
University location
United States -- Massachusetts
Degree
Ph.D.
Source type
Dissertations & Theses
Language
English
Document type
Dissertation/Thesis
Dissertation/thesis number
3163708
ProQuest document ID
304996791
Copyright
Database copyright ProQuest LLC; ProQuest does not claim copyright in the individual underlying works.
Document URL
http://search.proquest.com/docview/304996791
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