Veterinary Microbiology 136 (2009) 403–407 Contents lists available at ScienceDirect Veterinary Microbiology journal homepage: www.elsevier.com/locate/vetmic Short communication In vitro assessment of the effectiveness of powder disinfectant (Stalosan 1 F) against Lawsonia intracellularis using two different assays Suphot Wattanaphansak a , Randall S. Singer a , Richard E. Isaacson a , John Deen b , Bradley R. Gramm c , Connie J. Gebhart a, * a Department of Veterinary and Biomedical Sciences, College of Veterinary Medicine, University of Minnesota, St Paul, MN 55108, USA b Department of Veterinary Population Medicine, College of Veterinary Medicine, University of Minnesota, St. Paul, MN 55108, USA c Phibro Animal Health, Bloomington, IL 61704, USA ARTICLE INFO Article history: Received 30 August 2008 Received in revised form 1 December 2008 Accepted 2 December 2008 Keywords: Proliferative enteropathy Lawsonia intracellularis Disinfectant effectiveness ABSTRACT The objective of this study was to determine the in vitro efficacy of Stalosan 1 F, a mixed chemical and heavy metal disinfectant, against two strains of Lawsonia intracellularis using both a modified tissue culture and a direct count method. For testing as a powder, 1 g, 0.5 g, or 0.25 g of Stalosan F was applied to bacterial solutions spread into sterile dishes. For use as an aqueous suspension, Stalosan F was prepared to final concentrations of 1%, 4%, 8%, 16%, and 32%. In both applications, L. intracellularis was exposed to Stalosan F for 0.5 h, 1 h, 2 h, and 4 h. The results showed that both strains were similar in their susceptibilities to Stalosan F. The modified tissue culture assay showed no detectable L. intracellularis in cell culture after exposure to all levels of Stalosan F powder for 0.5 h. Furthermore, the number of viable bacteria was markedly reduced in the aqueous concentration of 4% and no L. intracellularis was detected at concentrations of ! 8% for 0.5 h. Using the direct count method, detection of live bacteria was less than 1% after exposure to the powder for 0.5 h. After exposure to the aqueous form, the number of viable bacteria killed was over 99% in concentrations of ! 16% compared to controls. Our results indicate that Stalosan F in both powder and suspension forms is able to inactivate over 99% of L. intracellularis after 30 min of exposure. Furthermore, both laboratory methods can be used to determine the effect of disinfectants on L. intracellularis viability. ß 2008 Elsevier B.V. All rights reserved. 1. Introduction There are limited data on the effectiveness of disin- fectants against Lawsonia intracellularis , a Gram-negative obligately intracellular bacterium that causes proliferative enteropathy (PE) (Lawson and Gebhart, 2000). This is mainly due to the difficulty of finding good methods to measure the efficacy of disinfectants against an obligately * Corresponding author at: Department of Veterinary and Biomedical Sciences, College of Veterinary Medicine, 205 Veterinary Science Building, 1971 Commonwealth Ave, St Paul, MN 55108, USA. Tel.: +1 612 624 3444; fax: +1 612 625 5203. E-mail address: gebha001@umn.edu (C.J. Gebhart). 0378-1135/$ – see front matter ß 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.vetmic.2008.12.002 intracellular bacterium. One study used a conventional tissue culture method to measure the viability status of L. intracellularis after exposure to some disinfectants (Collins et al., 2000). That assay was time-consuming and incapable of distinguishing between proportions of viable or non- viable bacteria. Recently, a modified tissue culture method that cultured bacteria in 96-well tissue culture plates has been developed for use in serological diagnosis (Guedes et al., 2002) and for the determination of the in vitro antimicrobial activity (Wattanaphansak et al., 2009) of L. intracellularis . This method allows the testing of a large number of compounds over various concentrations at the same time. Additionally, the viability of L. intracellularis in a mixed bacterial population can be directly measured using a direct count method with specific fluorescence staining
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