Trichomonas vaginalis: characterization of a 39-kDa cysteine proteinase found in patient vaginal secretions

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Trichomonas vaginalis characterization of a
39-kDa cysteine proteinase found in patient
vaginal secretions
Rodolfo Hernandez-Gutierrez, Leticia
Avila-Gonzalez, Jaime Ortega-Lopez,Fernando
Cruz-Talonia, Guillermo Gomez-Gutierrez, and
Rossana Arroyoa

• Mariska Petersen

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Trichomonaisis

• History
• Background Info
• Biological Mechanism
• Experimentation
• Data
• Results
• Conclusion

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Trichomonaisis

• Sexually transmitted infection
• Discovered in 1836
• 5 million cases reported every year
• 170 million people infected worldwide
• Resides in genitourinary tract of males and
  females
• 50 asymptomatic carriers

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Transmission

• Sexual intercourse
• Mutual masturbation
• Sharing sex toys
• Mother to child during vaginal delivery

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Signs and Symptoms

• In females there is
• inflammation of the vaginal canal
• vulvar itching leading to edema
• tenderness and chaffing
• redness
• yellow and green discharge
• painful urination
• pain during sex
• punctuate hemorrhages on the cervix known as
  colpitis macularis or strawberry cervix.

• In males there is
• whitish discharge although rare
• pain and burning during urination

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Treatment
Metronidazole (Flagyl) single two gram dose
(85-92 cure rate) OR 250 mg three time daily for
7-10 days. Tinidazole is an alternate drug
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Biological Mechanism

• T. vaginalis causitive agent
• Reproduction- binary fission
• Incubation period is 4-28 days
• Facultative anerobic parasite
• Adhesion proteins on surface of flagella
• Ligand/Receptor Cytoadherence
• 11-23 different CPs (cysteine proteinases)
• CPs play an important role in the pathogenicity
  of the parasite.

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Key Terms to know

• Cysteine Proteinase (CP) 39- an active surface
  proteinase secreted during the infection.
• Western Blot- a technique in molecular biology,
  used to separate and identify proteins.
• HeLa cells- Cervical Cancer cells
• VEC - Vaginal Epithelial Cells
• DU-145- Human prostatic cancer cells
• CaCo cell line- Human colon cancer cell line

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Characterization of CP39 (cysteine proteinase)
found in the vagina of women with trichomoniasis.
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Materials and Methods

• Determination of proteinases in vaginal
  secretions
• CP39 binding to different cell types
• Presence of CP39
• Heat stability and pH tolerance of CP39.
• Degradation of substrates by CP39
• Localization of CP39 on T. vaginalis
• Presence of antibodies to CP39 in sera from
  patients with T. vaginalis

• Patients were diagnosed with T. vaginalis
• Healthy patients used as negative controls
• The samples (vaginal secretions, urinary
  sediment, and sera) obtained were tested for the
  T. vaginalis and trichomonad proteinases.
• Samples used in several tests including ?

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Determination of proteinases in vaginal secretions

• Figure on the left shows proteinases can be
  secreted by viable and metabolically active
  trichomonads under in vitro controlled conditions.

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CP39 Cell Binding
Figure above shows that CP39 bound to the
surface of the HeLa and VEC. CP39 also bound to
the prostatic DU-145 cell line, but not to the
surface of the CaCo cells.
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Presence of CP39
CP39 present in different T. vaginalis isolates.
CP39 was present in all isolates varying in
intensities (despite loading same amount of cells
on the gel).
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Heat stability pH
A- CP39 active in temperatures ranging from 4 to
50 C. B- CP39 has maximum activity at pH 7.0.
C- CP39 range of activity increased from 3.6 to
7.0. (Most activity)
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Degradation of Substrates
CP39 analyzed on substrate gel electrophoresis
using Collagen I,II, III, IV, human Fibronectin,
Hemoglobin, IgA and IgG as subtrates. CP39
degraded the all proteins tested. CP39 was
strongest on Collagen IV, IgA and IgG. Other
tests were performed on vaginal cultures whereby
CP39 degraded the IgA-containing cells. Data
suggests that CP39 might be apart of immune
invasion mechanisms used by the parasite.
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Localization of CP39 on T. vaginalis
CP39 was determined to be another trichomonad
surface protein.
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Conclusion

• The characteristics of CP39 are
• It is present in vaginal secretions in patients
  with trichomoniasis
• Its optimal temperature is 37C and a pH range of
  3.6 to 7.0
• Suggested to be involved in tissue damage.
• Has broad substrate specifity
• Plays a role in parasite survival and immune
  evasion by degrading hemoglobin and
  immunoglobulins.
• Indicative that CP39 plays a role in trichomonal
  infection

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References

• Burgess, Donald and J.R. Schwebke .
  Trichomoniasis. Clinical Microbiology Reviews.
  Volume 17 pp. 794-803.
• Clenney, Timothy L. and M.K. Owen. Management of
  Vaginitis. American Family Physician. Volume 70
  pp.2125-2132. http//search.epnet.com/login.aspx?d
  irecttruedbaphan15224416
• Hernandez, G.R. L.A. Gonzalez, J. O. Lopez,F. C.
  Talonia, G. G. Gutierrez, and R. Arroyoa 2004.
  Trichomonas vaginalis characterization of a
  39-kDa cysteine proteinase found in patient
  vaginal secretions. Experimental Parasitology
  Volume 107(3-4)125-135.
• Singh B.N. 2004. Tritrichomonas foetus induces
  apoptotic cell death in bovine vaginal epithelial
  cells. Infect. Immunology. Volume 72(7)4151-4158
• Soper, David. Trichomoniasis under control or
  undercontrolled?. American Journal of Obstetrics
  and Gynecology, Volume 190 pp. 281-290.
• Unknown. Vaginitis. American Family Physician.
  Volume 70. p.1. http//search.epnet.com/login.aspx
  ?directtruedbaphan15227627

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Thank You