Towards a standardized method for broth microdilution susceptibility testing of Haemophilus parasuis

Pruller, Sandra, Turni, Conny, Blackall, Patrick J., Beyerbach, Martin, Klein, Guenter, Kreienbrock, Lothar, Strutzberg-Minder, Katrin, Kaspar, Heike, Meemken, Diana and Kehrenberg, Corinna (2017) Towards a standardized method for broth microdilution susceptibility testing of Haemophilus parasuis. Journal of Clinical Microbiology, 55 1: 264-273. doi:10.1128/JCM.01403-16

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Author Pruller, Sandra
Turni, Conny
Blackall, Patrick J.
Beyerbach, Martin
Klein, Guenter
Kreienbrock, Lothar
Strutzberg-Minder, Katrin
Kaspar, Heike
Meemken, Diana
Kehrenberg, Corinna
Title Towards a standardized method for broth microdilution susceptibility testing of Haemophilus parasuis
Formatted title
Towards a standardized method for broth microdilution susceptibility testing of Haemophilus parasuis
Journal name Journal of Clinical Microbiology   Check publisher's open access policy
ISSN 1098-660X
0095-1137
Publication date 2017-01-01
Year available 2016
Sub-type Article (original research)
DOI 10.1128/JCM.01403-16
Open Access Status File (Publisher version)
Volume 55
Issue 1
Start page 264
End page 273
Total pages 10
Place of publication Washington, DC, United States
Publisher American Society for Microbiology
Language eng
Subject 2726 Microbiology (medical)
Abstract Currently, there is no agreed method available for broth microdilution susceptibility testing of Haemophilus parasuis, one of the most important bacterial pathogens in pig production. Therefore, the aim of this study was to develop a method that could be easily performed by diagnostic laboratories and that appears suitable for a harmonized susceptibility testing. Growth determinations using one type strain and three field isolates revealed no visible growth of H. parasuis in media which have proven to be suitable for susceptibility testing of fastidious organisms. Therefore, a new medium, cation-adjusted Mueller-Hinton broth (CAMHB) plus NADH and sterile filtered heat-inactivated chicken serum, was developed. The reproducibility of MICs obtained in this medium was evaluated and statistically analyzed, considering a model with two different variables (precondition of five identical MICs and MIC mode accepting a deviation of ±1 dilution step, respectively). No significant differences for both variables were seen between two time points investigated and between results obtained with the recently proposed test medium broth (TMB). Nearly all MICs of quality control strains were in the acceptable range. Subsequently, 47 H. parasuis isolates representing 13 serovars were tested with the newly developed medium and TMB. Statistical analysis of all isolates and 15 antimicrobial agents and antimicrobial combinations showed no significant difference between MICs obtained in supplemented CAMHB and TMB. Because of a simplified implementation in routine diagnostic and a lower chance of interference between medium components and antimicrobial agents, supplemented CAMHB is recommended with an incubation time of 24 h.
Formatted abstract
Currently, there is no agreed method available for broth microdilution susceptibility testing of Haemophilus parasuis, one of the most important bacterial pathogens in pig production. Therefore, the aim of this study was to develop a method that could be easily performed by diagnostic laboratories and that appears suitable for a harmonized susceptibility testing. Growth determinations using one type strain and three field isolates revealed no visible growth of H. parasuis in media which have proven to be suitable for susceptibility testing of fastidious organisms. Therefore, a new medium, cation-adjusted Mueller-Hinton broth (CAMHB) plus NADH and sterile filtered heat-inactivated chicken serum, was developed. The reproducibility of MICs obtained in this medium was evaluated and statistically analyzed, considering a model with two different variables (precondition of five identical MICs and MIC mode accepting a deviation of ±1 dilution step, respectively). No significant differences for both variables were seen between two time points investigated and between results obtained with the recently proposed test medium broth (TMB). Nearly all MICs of quality control strains were in the acceptable range. Subsequently, 47 H. parasuis isolates representing 13 serovars were tested with the newly developed medium and TMB. Statistical analysis of all isolates and 15 antimicrobial agents and antimicrobial combinations showed no significant difference between MICs obtained in supplemented CAMHB and TMB. Because of a simplified implementation in routine diagnostic and a lower chance of interference between medium components and antimicrobial agents, supplemented CAMHB is recommended with an incubation time of 24 h.
Keyword Broth microdilution
Fastidious organism
Haemophilus parasuis
MICs
Supplemented CAMHB
Susceptibility testing
TMB
Q-Index Code C1
Q-Index Status Provisional Code
Grant ID 2811HS010
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: HERDC Pre-Audit
Queensland Alliance for Agriculture and Food Innovation
 
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