The Diel Rhythm that underlies the symbiotic relationship between

Euprymna scolopes and Vibrio fischeri

Mckayla BurnsSamantha Jackson

Key WordsDiel- A 24 hour periodCytoskeleton- A structural support system

made of microtubules and microfilamentsMicroarray-A technique used for

biochemical, genetic analysisChitin-a nitrogen-containing polysaccharideG3P-is a chemical compound used for the

anaerobic respiration of glycerolBlebbing- a blistering or swelling that

occurs in the cytoskeleton

About the AuthorsAndrew M. Wier• Associate professor at Pace University• Uses Euprymna scolopes and Vibrio fischeri as a general model to

study the effects of beneficial bacteria on animal host tissues.

Spencer V. Nyholm• Previous research: Dominance of Vibrio fischeri in secreted mucus

outside the light organ of Euprymna scolopes: the first site of symbiont specificity. Appl. Environ. Microbiol., 69(7), 3932-3937.

• Professor at the University of Connecticut• Also studies the symbiosis between tubeworm and sulfur oxidizing

bacteria in hydrothermal vents

Background InformationHorizontal Transmission:• Every generation of Euprymna scolopes and

Vibrio fischeri must establish the symbiosis, which is different from vertical transmission where the symbiosis is passed from parent to offspring

Mutualistic Relationship:• The squid provides nutrients for the bacteria

and the bacteria provides the squid with bioluminescence which is used for the squid’s protection

Background Information

Figure 1: The diel cycle of the squid/Vibrio symbiosis

A: Shows the position of the light organ within the squid, the expulsion of bacteria from the light organ, and crypt where bacteria symbionts resides

B: The blue line symbolizes the population of the bacteria throughout a 24 hour period. Bacteria’s populations are the highest when the nocturnal host is the most active.

- During expulsion the squid releases 95% of it’s bacteria symbionts, keeping only 5% to regenerate

General Gene ExpressionThe greatest changes in gene expression of

the host cells and symbiont cells occurred before and after dawn

The highest up regulation of host genes occurred before dawn

The highest percentage of up regulated genes in the symbionts occurred after dawn

During some intervals both partners changed the expression of genes specifically used for signaling and metabolism

PurposeTo describe the

transcriptional, metabolic, and physiological aspects of the symbiosis between Euprymna scolopes and Vibrio fischeri and determine if they follow a diel pattern

PhysiologicalMethods:• Used TEM (Transmission Electron Microscopy)• Examined the epithelium cells within the light organ of

Euprymna scolopes over 4 different time periods within 24 hours Results:• Major change in the morphology of the epithelial cells• Directly after dawn, the epithelial cells of the light organ are

obliterated• Following this there was an increase in the number of host

vesicles anticipating the creation of new bacterial symbionts• A few hours later the microvillar border is restored within the

epithelial cells• Next, the light organ is colonized by new bacteria cells and it is

ready for night fall and bioluminescence

How do the Bacteria Repopulate the light organ?Metabolize the lipid vesicles the host cells creates

after dawnHost changes its tissue organization in order to

provide the symbiont with glycerol and fatty acidsBacteria incorporate fatty acids from host lipids into

its membrane

Fatty Acid Composition of Bacterial Symbionts

Regulation of cytoskeleton genes

Up regulated just before dawn

Down regulated just after down

TEMS of the host crypt epithelia

Transcriptional Analysis Reveals Patterns in Symbiont MetabolismMethods:Examined changes in bacterial gene expression

during the day and nightMicroarray analysis of the RNA in the central cores of

the E. scolopes light organsResults:Diel Pattern of Symbiont MetabolismJust before expulsion there is an increase in the

expression of genes for the fermentation for chitinJust after dawn there is an increase in the expression

of genes for the anaerobic respiration of glycerol

Why do the symbionts shift their metabolism?At night during bioluminescence symbionts

use chitin fermentation to generate ATPAfter dawn and throughout the day the

symbionts use anaerobic respiration of glycerol to generate ATP

The form of anaerobic metabolism may be dependent on the substrates available at the time

When chitin is available 12% more energy is produced using fermentation

When glycerol is available more energy is produced using anaerobic respiration

Evidence of a diel pattern of symbiont metabolism.

Wier A M et al. PNAS 2010;107:2259-2264

©2010 by National Academy of Sciences

Conclusions:

There is a transformation of host tissue anatomy

There is a change in gene expression by both partners

These changes are controlled by a daily rhythm

Bioluminescence is maintained by daily interactions between the squid and the bacteria

Strengths/WeaknessesStrengths:• Results for anatomical changes are consistent

with diel rhythm hypothesis• Results support metabolic changes occur on a

diel rhythm Weaknesses:• The source of the nitrate for anaerobic

respiration is still unknown

Further Study:1. “Although the nutritional conditions within the light organ remain unknown, taken together, these calculations support the possibility that over the course of a 24 hour period, the host provides different substrates to its symbionts to optimize their performance under changing physiological conditions present in its tissue over the day-night cycle.” ^(Page 2262)

2. Do changes in the anatomy and gene expression present in other mammalian systems follow a diel rhythm?Tissues in Mouse Colon are believed to follow a circadian

rhythm 3. Parallels between beneficial and pathogenic bacteria induced effacement

References:-http://www.pnas.org/content/107/5/2259.figures-only

-Wier, Andrew M., and Spencer V. Nyholm. "Transcriptiona Patterns in Both Host and Bacterium Underlie a Daily Rhythm of Anatomical and Metabolic Change in a Beneficial Symbiosis." PNAS 107.5 (2010): 2259-264. Print.