The influence of Burkholderia endosymbionts on squash bugs’ (Anasa tristis) responses to Serratia marcescens, the plant pathogen that they vector Open Access

Xia Junyan (Spring 2018)

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Many animals form and maintain long-term, symbiotic relationships with microbes. Symbiotic microbes can be acquired vertically, passing down from parents to their offspring. Alternatively, hosts can acquire symbionts horizontally from the environment. Primary, or obligate, symbionts usually play pivotal roles in increasing host fitness, and thus are beneficial most of the time. In contrast, secondary, or facultative, symbionts can benefit, harm, or have no influence upon a host. Symbiotic microbes can influence many aspects of host biology, including developmental time, nutrient acquisition, and defense. In terms of defense, while many symbiotic microbes enhances host resistance and tolerance towards pathogens and parasites, symbionts can also weaken a host’s defenses by down-regulating host immunity. Because a host’s response towards foreign microbes can be influenced by the presence of symbionts, a insect’s vectoring ability might also be influenced by its symbiotic microbes. Because of this, there is increasing interest in developing symbiont-mediated biocontrol strategies. 


The squash bug (Anasa tristis) is a vector of Serratia marcescens, the plant pathogen that causes cucurbit yellow vine disease (CYVD). Previous studies in our lab have shown that A. tristis partners with bacteria in the genus Burkholderia spp., forming a symbiotic relationship. It is of particular interest to study symbiosis in A. tristis given its potential to be applied to create novel biocontrol strategies. In my investigations, I examined whether the presence of symbiotic Burkholderia influences the establishment of S. marcescens in A. tristis. I found that symbiont-containing A. tristis had lower S. marcescens establishment than symbiont-lacking A. tristis after being orally inoculated with S. marcescens. It was also found that symbiont-containing A. tristis had higher survivorship than symbiont-lacking A. tristis after a subsequent introduction of S. marcescens through injection. This indicates that symbiont-containing A. tristis may have a stronger general immune response towards S. marcescens infection.

Table of Contents

Abstract      ----------------------------------------------------------------------------------------- 1                                              

Introduction    -------------------------------------------------------------------------------------- 2                                                  

Chapter 1:

1.1Abstract    --------------------------------------------------------------------------------------- 6                                                   

1.2 Introduction    ---------------------------------------------------------------------------------- 7                                            

1.3 Methods and Material    ---------------------------------------------------------------------- 10                                        

1.4 Results    --------------------------------------------------------------------------------------- 15

Figure 1.1 Estimated Log CFUs of fluorescent S. marcescens remaining in A. tristis 20 hours after orally introducing A. tristis with fluorescent S. marcescens    ----------------------------- 15

Figure 1.2 Bacteria zone of inhibition assay    --------------------------------------------------- 17                                           

1.5 Discussion    ---------------------------------------------------------------------------------- 18                                               

Chapter 2: 

2.1 Abstract    ------------------------------------------------------------------------------------- 21                                                   

2.2 Introduction    --------------------------------------------------------------------------------- 22                                                

2.2 Methods and Material    ---------------------------------------------------------------------- 25                                        

2.3 Results    -------------------------------------------------------------------------------------- 29            

Figure 2.1 Survivorship of A. tristis after being infected by S. marcescens ZO1 bacteria with a minutin pin    -------------------------------------------------------------------------------------- 29

Figure 2.2 Estimated Log CFUs of fluorescent Burkholderia SQ4A remaining in A. tristis 0 hours after introducing A. tristis with fluorescent Burkholderia SQ4A through injection

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Figure 2.3 Estimated Log CFUs of fluorescent S.marcescens ZO1 remaining in A. tristis after introducing A. tristis with fluorescent S. marcescens ZO1 through injection

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Figure 2.4 Estimated Log CFUs of fluorescent S. marcescens Z01 remaining in A. tristis 6 hours after introducing A. tristis with fluorescent S. marcescens Z01 through injection

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2.4 Discussion    ----------------------------------------------------------------------------------- 33                                                  

Conclusions and Future Directions    ------------------------------------------------------------- 37                                

Supplemental Figures       

Supplemental figure 1 Burkholderia Phylogeny    ------------------------------------------------ 39

Supplemental figure 2 Symbiotic Burkholderia SQ4A significantly increases A. tristis’s survival

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Supplemental figure 3 Symbiotic Burkholderia SQ4A significantly increase the number of A. tristis that achieve later developmental stage.    ------------------------------------------------ 40

Supplemental figure 4 The estimated amount of S. marcescens remaining in Burkholderia symbiont-containing and symbiont-lacking A. tristis at 0 hour time point after orally introducing S. marcescens into A. tristis.   -------------------------------------------------------------------- 41

Supplemental figure 5 The estimated amount of symbiotic bacteria in adult A. tristis.

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Supplemental figure 6 Bacteria Zone of Inhibition Design    ------------------------------------ 42

Supplemental figure 7 The number of days for Burkholderia A33.M4.C. symbiont-containg and symbiont-lacking A. tristis to develop into adulthood    ---------------------------------------- 43

Supplemental figure 8. A. tristis rearing, symbiont establishment, and experiments    ------ 44  

Supplemental figure 9. Three mechanisms of symbiont-mediated host defense    ------------ 45                 

References    -------------------------------------------------------------------------------------- 46       


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