MycofluidicsThe Fluid Mechanics of Fungal Chimerism and Spore Dispersal
Presenters: Devin Bowers, Trevor Caldwell, Francesca Grogan, LisaYamada
Mentors: Prof. Marcus Roper, Prof. Emilie Dressaire
UCLA Applied Math REU
August 8th, 2012
Introduction to Mycology
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Notable Fungi
Rice blast
Cordyceps
White nose syndrome
Figure: Cordyceps versus an ant
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Introduction to Mycology
A chimera is an organism that is made up of cells with geneticallydifferent nuclei.
Can reproduce asexually via spores
Most fungi grow as hyphae, which are filamentous structures thatgrow at their tips.
D. Bowers, T. Caldwell, F. Grogan, L. Yamada (REU) Mycofluidics August 8th, 2012 4 / 25
Moran Process
A Moran process is a stochastic process that models processes affectinggenetic diversity.The dynamics of two different populations of nuclei withina fungi can be modeled as a two species Moran process.
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Roper M, Simonin A, Hickey PC, Leeder A, Glass NL (2012) Nuclear dynamics in afungal chimera (preprint).
Previous Work
Fungal chimerism gives rise to the internal genetic diversity of fungi.It has been found that nuclear mixing within the mycelium keepsgenetically different nuclei well-mixed and in stable proportionsthroughout the fungus.
D. Bowers, T. Caldwell, F. Grogan, L. Yamada (REU) Mycofluidics August 8th, 2012 6 / 25
Roper M, Simonin A, Hickey PC, Leeder A, Glass NL (2012) Nuclear dynamics in afungal chimera (preprint).
Our Experiment: Question
Is a fungus able to maintain its genetic diversity? If so, how?
D. Bowers, T. Caldwell, F. Grogan, L. Yamada (REU) Mycofluidics August 8th, 2012 7 / 25
Our Experiment: Method
Neurospora crassa (N. crassa), atype of bread mold, with GFP orDsRed proteins
GFP and DsRed cause nuclei toemit a green and redfluorescence, respectively.
Figure: N. crassa Cultures
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Our Experiment: Method
Figure: Race Tubes
We built and innoculatedrace tubes with bothGFP and DsRed.
When the coloniesstarted sporulating, wetook images with amicroscope.
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Our Experiment:Method
Figure: DIC Images of Spores
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Our Experiment:Method
Figure: Fluorescent DsRed and GFP
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Our Experiment: Method
Image Analysis
Find the percentage of spores that have only red nuclei (pR), onlygreen nuclei (pG ), and both (pY ).
Find the frequency of the number of nuclei present in a spore to getP(Nnucl = n).
From this, we can solve∑n
P(Nnucl = n)pnr = pR
∑n
P(Nnucl = n)(1− pr )n = pG
for pr , the probability that a nucleus is red.
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Our Experiment: Method
Pr Fitting
Estimate pr from likelihood function
L(NR ,NG ,NY ) =N!
NR !NG !NY !pNRR pNG
G pNYY
Find maximum of log-likelihood
NR
pR
∂pR∂pr
+NG
pG
∂pG∂pr− NY
pY
(∂pR∂pr
+∂pG∂pr
)= 0
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Our Experiment: MethodImage Analysis
Otsu’s Method–histogram-based thresholdingI Finds optimum value in a bi-modal histogram such that the intra-class
variance is minimal.
Figure: Initial Intensity Histogram
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Our Experiment: MethodImage Analysis
Modified Otsu’s Method to take in the original histogramUsed this second threshold to identify nuclei within spores
Figure: Normalized Intensity Histogram
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Thresholding
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Thresholding
Figure: Otsu’s thresholding
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Our Experiment: Results
Figure: pr values along race tube
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Our Experiment: Results
Figure: pr values along race tube
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Our Experiment: Results
Figure: pr values along race tube
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Our Experiment: Results
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Our Experiment: Results
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Our Experiment: Results
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Our Experiment: Results
Interpretation of pr Fluctuations
Moran process
Effect of shaking and accelerated growth
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Our Experiment: Future Work
Improve pr fittingI Multinomial distribution with dependency parameters
Segmentation issuesI ClumpingI Illumination/Saturation sensitivity
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