Epiphytic fitness of a biological control agent of fire blight in apple and pear orchards under Mediterranean weather conditions

Abstract

The behaviour of Pseudomonas fluorescens EPS62e was investigated in apple and pear orchards under Mediterranean climatic conditions. The trials studied the influence of weather conditions, plant host species, presence of indigenous microbial community and spread from treated to nontreated trees on colonization and survival. Population dynamics were assessed by real-time PCR and CFU-counting methods. With inoculated flowers, weather conditions were optimal for colonization, and EPS62e established high and stable population levels around 10(8) CFU per organ, according to both methods of analysis. The plant host species did not influence the colonization rate, and the biocontrol agent dominated the microbial communities of blossoms, representing up to 100% of the total cultivable population. With inoculated leaves, the EPS62e population decreased to nondetectable levels 30 days after treatment according to both methods used. EPS62e spread moderately in the orchard, being detected in nontreated flowers of trees 15-35 m from the inoculation site. The combined use of real-time PCR and CFU-counting methods of analysis permitted the identification of three physiological states for EPS62e in the field, which consisted of active colonization, survival and entry into a viable but nonculturable state, and cell death.

Improvement of Fitness and Efficacy of a Fire Blight Biocontrol Agent via Nutritional Enhancement Combined with Osmoadaptation

“…Notably, Tween 80 is used as an adjuvant in the formulation of several biopesticides (2,17,31). Following a single field application during bloom, EPS62e colonized blossoms and reached population levels near the carrying capacity of apple and pear flowers (6,24,25); this behavior on blossoms is similar to that of other biocontrol agents of fire blight (16,27,37) that efficiently colonize blossoms when phenoclimatic conditions during spring are optimal (14). However, under limiting environmental and/or nutritional conditions, growth and colonization of biocontrol agents are not optimal.…”

“…However, under limiting environmental and/or nutritional conditions, growth and colonization of biocontrol agents are not optimal. Also, during delivery of the biocontrol agents to the orchard a sharp change in the environment conditions from laboratory to field occurs, affecting survival and colonization (25). The monitoring of the population of the introduced EPS62e strain into orchards using real-time PCR and CFU-counting methods provided information on the population viability.…”

“…However, field studies on fitness of biocontrol agents are difficult because of the need for methods to estimate population dynamics at strain level. The development of methods for the specific quantitative analysis of strain EPS62e by real-time quantitative PCR (Q-PCR) in combination with plate counting methods (24) provided a tool to analyze its population behavior in terms of the proportion of viable cells (24,25). Using this approach, cell death of the biocontrol agent was observed immediately after field delivery due to the sharp change in the biological and physical environmental conditions from the optimal laboratory culture medium to the growth-limiting tree surface (25).…”

“…The development of methods for the specific quantitative analysis of strain EPS62e by real-time quantitative PCR (Q-PCR) in combination with plate counting methods (24) provided a tool to analyze its population behavior in terms of the proportion of viable cells (24,25). Using this approach, cell death of the biocontrol agent was observed immediately after field delivery due to the sharp change in the biological and physical environmental conditions from the optimal laboratory culture medium to the growth-limiting tree surface (25). It has also been reported that aerial surfaces of plants can be transiently inadequate for bacterial growth, because they are exposed to rapid and wide changes in water availability and temperature, UV light, and limitation in nutrients (18,19,32).…”

Characterization of selected Gram-negative non-fermenting bacteria isolated from honey bees (Apis mellifera carnica)

“…During the last two decades (Klingauf 2006), there has been a growing interest to use antagonistic microorganisms (e.g., Pantoea agglomerans, Pseudomonas fluorescens, and fungus Aureobasidium pullulans (Cañamás et al 2007;Pujol et al 2007;Loncaric et al 2008)) for the biological control of some plant pathogenic bacteria (e.g., Erwinia amylovora, fire blight). The honey bees have been used as a vector for establishment of these three antagonistic of E. amylovora in apple and pear orchards (Thomson et al 1992;Loncaric et al 2008).…”