Comparative analysis of plant and animal models for characterization of Burkholderia cepacia virulence

Abstract

A simple alfalfa model was developed as an alternative infection model for virulence studies of the Burkholderia cepacia complex. Symptoms of disease were observed in wounded alfalfa seedlings within 7 days following inoculation of 10(1) to 10(5) CFU of most strains of the B. cepacia complex. Strains from seven genomovars of the B. cepacia complex were tested for virulence in the alfalfa model, and the degree of virulence was generally similar in strains belonging to the same genomovar. Strains of Burkholderia multivorans and some strains of Burkholderia stabilis did not cause symptoms of disease in alfalfa seedlings. Representative strains were also tested for virulence using the rat agar bead model. Most of the strains tested were able to establish chronic lung infections; B. stabilis strains were the exception. Most of the strains that were virulent in the alfalfa infection model were also virulent in the lung infection model. The B. cepacia genomovar III mutants K56pvdA::tp and K56-H15 were significantly less virulent in the alfalfa infection model than their parent strain. Therefore, this alfalfa infection model may be a useful tool for assessing virulence of strains of the B. cepacia complex and identifying new virulence-associated genes.

FIG. 1.

Effect of temperature on visual symptoms of pathology on alfalfa. Photographs of wounded alfalfa seedlings after inoculation with 10⁵ CFU of B. cepacia K56-2 are shown. Seedlings were incubated at 30°C (A) and 37°C (B). In each panel, the seedling on the left is the negative control (inoculated with saline), and the seedling on the right was inoculated with K56-2.

FIG. 2.

Effects of temperature and inoculum on alfalfa infections. Groups of 20 seedlings were inoculated with doses of B. cepacia K56-2 ranging from 10¹ to 10⁵ CFU and incubated at three different temperatures: room temperature (RT), 30°C, and 37°C. The results shown are the percentages of seedlings with disease symptoms visible on day 7 p.i.

FIG. 3.

Comparison of the ability of B. cepacia complex genomovars to cause infections in alfalfa. Seedlings were inoculated with 10⁵ CFU of the strains shown in Table 1 and incubated at 37°C for 7 days. Data from Table 1 for the percentage of seedlings with symptoms of disease for each genomovar (A) and for the number of bacteria recovered (10⁷) from individual homogenized alfalfa sprouts for each genomovar (B) are summarized. The means ± standard deviations (error bars) of the values in Table 1 are shown.