CSULB 5.3.04

Effects of M1G1 on the systemic dissemination of Candida albicans

By

David Nguyen

Mentor: Dr. Zhang

CSULB 5.3.04

Background Information

• Candida albicans is a dimorphic pathogenic yeast known to cause systemic infections within the human population

• C. albicans is a common saprophyte human commensal, being found in the skin, urogenital, and the alimentary tract

• Severe C. albicans infections are frequently observed in those whose immune system has been compromised such as advance stage HIV patients

CSULB 5.3.04

Background Information

• Recently, C. albicans, including other Candida species, have become the 4th leading cause of nosocomial bloodstream infection

• It has been observed that mortality correlates with the fungal burden of the kidney

• It has also been noted that viable yeasts can be recovered from kidneys 56 days post-infection, implying that the organism can evade the host’s defenses. This organ’s susceptibility has yet to be elucidated.

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Question

• Question:

How is C. albicans able to evade the host’s defenses 56 days post-infection, especially in the kidneys?

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Hypothesis

• It is hypothesized that upon infection, before native host antibodies can be produced at an effective protective level, C. albicans can establish itself within the kidney organs. Once the yeasts have been established, the innate immunity is unable to protect the host from the infection and C. albicans colonies will continue to persist in the host.

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Materials and Methods

• 40 six week old female BALB/c mice

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Materials and Methods

• Group A (15 mice) are infected and passively immunized

• Group B (15 mice) are infected and non-passively immunized

• Group C (10 mice) are non-infected and non-passively immunized

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Materials and Methods

• 1 mg/mouse recombinant anti-mannan human antibody known as M1G1

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Materials and Methods

• 1x106 cells of C. albicans 3156A (ATCC)

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Materials and Methods

• Every two weeks, 3 mice from group A, 3 mice from group B, and 2 mice from group C will be euthanized according to institutional regulations from an Association for Assessment and Accreditation of Laboratory Animal Care-Certified (AAALAC-certified) animal facility at California State University Long Beach

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Materials and Methods

• Organs will be extracted and fixed in 10% formaldehyde. Briefly, each organ from each time frame will be processed via histochemical techniques involving 2 types of stains.

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Materials and Methods

• First type of stain is known as Hematoxylin and Eosin stain (HE) (detects host cell nuclei)

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Materials and Methods

• Second type of stain is known as Grocott’s Method for Fungi stain (GMS) (detects fungal matter)

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Materials and Methods

• Quantitative analysis of the results will be performed using the GMS stains via a calibrated light microscope under 25x. Random areas of cell counts will be taken per 1 mm2.

• Qualitative analysis of the results will be performed using the HE stains.

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Materials and Methods

• Statistics: Analysis of data will be performed using SigmaPlot to analyze the variances of the means of each group as well as correlation analysis.

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Expected Results

• Expected results of Group A mice should show low amounts of 3153A in the organs due to 3153A being unable to establish itself within organs such as kidneys.

• Group C should show neither signs of infection and its use will be to account for any artifacts which might show up via the histochemical techniques.

CSULB 5.3.04

Expected Results

• Expected results should show high densities of 3153A at immunologically relevant organs such as kidneys and spleen in Group B mice. There should be observation of extensive growth of yeast hyphae and filaments. Persistant colonies should be observed in the kidneys.

CSULB 5.3.04

Expected Results

• Quantitative determination should indicate a correlation between high and low yeast dissemination in non-passive immunization and passive immunization, respectively. As time progresses, systemic yeast dissemination should be reduced due to the host’s immune system producing its own antibodies against the yeast.

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Expected Results

Expected Cell Densities

0

50

100

150

0 5 10 15

week

cells

/1m

m^

2

Series1

Series2

Series3

G

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Why does it matter?

• If it is shown that passive immunity can prevent or minimize any detectable traces of C. albicans within the kidneys as well as other organs then this will allow insight into how C. albicans is able to persist within the body many days post-infection.

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Why does it matter?

• To date no there are no license vaccines for the treatment/prevention of human fungal infections. It is hoped that evidence from this research will provide additional evidence that passive immunity via M1G1 can be a potential candidate for use in clinical trials to treat patients with severe C. albicans infections.

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Acknowledgement

• I would like to thank the Howard Hughes Medical Institute for providing the grants necessary for completing this project.

• I would like to thank my mentor Dr. Zhang for allowing me to work on this project and for providing the laboratory for the experiments.