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. 1996 Jan;40(1):179–185. doi: 10.1128/aac.40.1.179

Effects of nifedipine, metronidazole, and nitric oxide donors on spore germination and cell culture infection of the microsporidia Encephalitozoon hellem and Encephalitozoon intestinalis.

Q He 1, G J Leitch 1, G S Visvesvara 1, S Wallace 1
PMCID: PMC163079  PMID: 8787902

Abstract

Two species of microsporidia, Encephalitozoon hellem and Encephalitozoon intestinalis, were isolated from AIDS patients and cultured in green monkey kidney cells. A spore germination assay and a cultured-cell infection assay were used to test the efficacy of candidate antiparasitic agents. The calcium channel blocker nifedipine, metronidazole, and two nitric oxide (NO) donors, S-nitroso-N-acetylpenicillamine and sodium nitroprusside, were tested in the two assays. Nifedipine (10(-8) M) significantly inhibited E. hellem spore germination in three of four germination media. Metronidazole (10(-5) M) inhibited germination weakly and significantly inhibited E. intestinalis germination in a single germination medium. The inhibitory effect of nifedipine and metronidazole used together was greater than the sum of the effects of the drugs used alone in all E. hellem germination assays. The NO donors also inhibited spore germination. The inhibitory effect of nifedipine and metronidazole could be reversed by washing the spores, while that of the NO donors was not reversible. In early cultured-cell infections, both nifedipine (10(-8) M) and metronidazole (10(-5) M) significantly reduced the number of cells being infected. As the infection spread, these agents were less effective. Some inhibition of the spread of the infection was also demonstrated with the NO donors at a concentration (10(-5) M) not obviously toxic to the cultured cells. These data suggest that combination drug therapy targeting spore germination and intracellular parasite development is promising.

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Selected References

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  1. Ahlner J., Andersson R. G., Torfgård K., Axelsson K. L. Organic nitrate esters: clinical use and mechanisms of actions. Pharmacol Rev. 1991 Sep;43(3):351–423. [PubMed] [Google Scholar]
  2. Alspaugh J. A., Granger D. L. Inhibition of Cryptococcus neoformans replication by nitrogen oxides supports the role of these molecules as effectors of macrophage-mediated cytostasis. Infect Immun. 1991 Jul;59(7):2291–2296. doi: 10.1128/iai.59.7.2291-2296.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blanshard C., Hollister W. S., Peacock C. S., Tovey D. G., Ellis D. S., Canning E. U., Gazzard B. G. Simultaneous infection with two types of intestinal microsporidia in a patient with AIDS. Gut. 1992 Mar;33(3):418–420. doi: 10.1136/gut.33.3.418. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Boockvar K. S., Granger D. L., Poston R. M., Maybodi M., Washington M. K., Hibbs J. B., Jr, Kurlander R. L. Nitric oxide produced during murine listeriosis is protective. Infect Immun. 1994 Mar;62(3):1089–1100. doi: 10.1128/iai.62.3.1089-1100.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Boreham P. F., Phillips R. E., Shepherd R. W. A comparison of the in-vitro activity of some 5-nitroimidazoles and other compounds against Giardia intestinalis. J Antimicrob Chemother. 1985 Nov;16(5):589–595. doi: 10.1093/jac/16.5.589. [DOI] [PubMed] [Google Scholar]
  6. Bryan R. T., Cali A., Owen R. L., Spencer H. C. Microsporidia: opportunistic pathogens in patients with AIDS. Prog Clin Parasitol. 1991;2:1–26. [PubMed] [Google Scholar]
  7. Cali A., Kotler D. P., Orenstein J. M. Septata intestinalis N. G., N. Sp., an intestinal microsporidian associated with chronic diarrhea and dissemination in AIDS patients. J Eukaryot Microbiol. 1993 Jan-Feb;40(1):101–112. doi: 10.1111/j.1550-7408.1993.tb04889.x. [DOI] [PubMed] [Google Scholar]
  8. Croen K. D. Evidence for antiviral effect of nitric oxide. Inhibition of herpes simplex virus type 1 replication. J Clin Invest. 1993 Jun;91(6):2446–2452. doi: 10.1172/JCI116479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Curry A., Canning E. U. Human microsporidiosis. J Infect. 1993 Nov;27(3):229–236. doi: 10.1016/0163-4453(93)91923-d. [DOI] [PubMed] [Google Scholar]
  10. De Groote M. A., Visvesvara G., Wilson M. L., Pieniazek N. J., Slemenda S. B., daSilva A. J., Leitch G. J., Bryan R. T., Reves R. Polymerase chain reaction and culture confirmation of disseminated Encephalitozoon cuniculi in a patient with AIDS: successful therapy with albendazole. J Infect Dis. 1995 May;171(5):1375–1378. doi: 10.1093/infdis/171.5.1375. [DOI] [PubMed] [Google Scholar]
  11. Desportes I., Le Charpentier Y., Galian A., Bernard F., Cochand-Priollet B., Lavergne A., Ravisse P., Modigliani R. Occurrence of a new microsporidan: Enterocytozoon bieneusi n.g., n. sp., in the enterocytes of a human patient with AIDS. J Protozool. 1985 May;32(2):250–254. doi: 10.1111/j.1550-7408.1985.tb03046.x. [DOI] [PubMed] [Google Scholar]
  12. Didier E. S., Didier P. J., Friedberg D. N., Stenson S. M., Orenstein J. M., Yee R. W., Tio F. O., Davis R. M., Vossbrinck C., Millichamp N. Isolation and characterization of a new human microsporidian, Encephalitozoon hellem (n. sp.), from three AIDS patients with keratoconjunctivitis. J Infect Dis. 1991 Mar;163(3):617–621. doi: 10.1093/infdis/163.3.617. [DOI] [PubMed] [Google Scholar]
  13. Didier E. S., Shadduck J. A. IFN-gamma and LPS induce murine macrophages to kill Encephalitozoon cuniculi in vitro. J Eukaryot Microbiol. 1994 Sep-Oct;41(5):34S–34S. [PubMed] [Google Scholar]
  14. Dieterich D. T., Lew E. A., Kotler D. P., Poles M. A., Orenstein J. M. Treatment with albendazole for intestinal disease due to Enterocytozoon bieneusi in patients with AIDS. J Infect Dis. 1994 Jan;169(1):178–183. doi: 10.1093/infdis/169.1.178. [DOI] [PubMed] [Google Scholar]
  15. Eeftinck Schattenkerk J. K., van Gool T., van Ketel R. J., Bartelsman J. F., Kuiken C. L., Terpstra W. J., Reiss P. Clinical significance of small-intestinal microsporidiosis in HIV-1-infected individuals. Lancet. 1991 Apr 13;337(8746):895–898. doi: 10.1016/0140-6736(91)90215-b. [DOI] [PubMed] [Google Scholar]
  16. Hartskeerl R. A., Van Gool T., Schuitema A. R., Didier E. S., Terpstra W. J. Genetic and immunological characterization of the microsporidian Septata intestinalis Cali, Kotler and Orenstein, 1993: reclassification to Encephalitozoon intestinalis. Parasitology. 1995 Apr;110(Pt 3):277–285. doi: 10.1017/s0031182000080860. [DOI] [PubMed] [Google Scholar]
  17. Ishihara R. Stimuli causing extrusion of polar filaments of Glugea fumiferanae spores. Can J Microbiol. 1967 Oct;13(10):1321–1332. doi: 10.1139/m67-178. [DOI] [PubMed] [Google Scholar]
  18. Johnson P. J. Metronidazole and drug resistance. Parasitol Today. 1993 May;9(5):183–186. doi: 10.1016/0169-4758(93)90143-4. [DOI] [PubMed] [Google Scholar]
  19. Leitch G. J., He Q. Arginine-derived nitric oxide reduces fecal oocyst shedding in nude mice infected with Cryptosporidium parvum. Infect Immun. 1994 Nov;62(11):5173–5176. doi: 10.1128/iai.62.11.5173-5176.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Leitch G. J., He Q., Wallace S., Visvesvara G. S. Inhibition of the spore polar filament extrusion of the microsporidium, Encephalitozoon hellem, isolated from an AIDS patient. J Eukaryot Microbiol. 1993 Nov-Dec;40(6):711–717. doi: 10.1111/j.1550-7408.1993.tb04463.x. [DOI] [PubMed] [Google Scholar]
  21. Metcalfe T. W., Doran R. M., Rowlands P. L., Curry A., Lacey C. J. Microsporidial keratoconjunctivitis in a patient with AIDS. Br J Ophthalmol. 1992 Mar;76(3):177–178. doi: 10.1136/bjo.76.3.177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Orenstein J. M. Microsporidiosis in the acquired immunodeficiency syndrome. J Parasitol. 1991 Dec;77(6):843–864. [PubMed] [Google Scholar]
  23. Parker J. O. Update on nitrate tolerance. Br J Clin Pharmacol. 1992;34 (Suppl 1):11S–14S. doi: 10.1111/j.1365-2125.1992.tb04142.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Pleshinger J., Weidner E. The microsporidian spore invasion tube. IV. Discharge activation begins with pH-triggered Ca2+ influx. J Cell Biol. 1985 Jun;100(6):1834–1838. doi: 10.1083/jcb.100.6.1834. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Ruf B., Sandfort J. Human microsporidiosis. J Antimicrob Chemother. 1994 Nov;34(5):609–611. doi: 10.1093/jac/34.5.609. [DOI] [PubMed] [Google Scholar]
  26. Sandfort J., Hannemann A., Gelderblom H., Stark K., Owen R. L., Ruf B. Enterocytozoon bieneusi infection in an immunocompetent patient who had acute diarrhea and who was not infected with the human immunodeficiency virus. Clin Infect Dis. 1994 Sep;19(3):514–516. doi: 10.1093/clinids/19.3.514. [DOI] [PubMed] [Google Scholar]
  27. Shadduck J. A. Human microsporidiosis and AIDS. Rev Infect Dis. 1989 Mar-Apr;11(2):203–207. doi: 10.1093/clinids/11.2.203. [DOI] [PubMed] [Google Scholar]
  28. Spedding M., Paoletti R. Classification of calcium channels and the sites of action of drugs modifying channel function. Pharmacol Rev. 1992 Sep;44(3):363–376. [PubMed] [Google Scholar]
  29. Undeen A. H., Frixione E. The role of osmotic pressure in the germination of Nosema algerae spores. J Protozool. 1990 Nov-Dec;37(6):561–567. doi: 10.1111/j.1550-7408.1990.tb01265.x. [DOI] [PubMed] [Google Scholar]
  30. Vincendeau P., Daulouède S., Veyret B., Darde M. L., Bouteille B., Lemesre J. L. Nitric oxide-mediated cytostatic activity on Trypanosoma brucei gambiense and Trypanosoma brucei brucei. Exp Parasitol. 1992 Nov;75(3):353–360. doi: 10.1016/0014-4894(92)90220-5. [DOI] [PubMed] [Google Scholar]
  31. Visvesvara G. S., da Silva A. J., Croppo G. P., Pieniazek N. J., Leitch G. J., Ferguson D., de Moura H., Wallace S., Slemenda S. B., Tyrrell I. In vitro culture and serologic and molecular identification of Septata intestinalis isolated from urine of a patient with AIDS. J Clin Microbiol. 1995 Apr;33(4):930–936. doi: 10.1128/jcm.33.4.930-936.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Weber R., Bryan R. T. Microsporidial infections in immunodeficient and immunocompetent patients. Clin Infect Dis. 1994 Sep;19(3):517–521. doi: 10.1093/clinids/19.3.517. [DOI] [PubMed] [Google Scholar]
  33. Weidner E., Byrd W. The microsporidian spore invasion tube. II. Role of calcium in the activation of invasion tube discharge. J Cell Biol. 1982 Jun;93(3):970–975. doi: 10.1083/jcb.93.3.970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Zhu L., Gunn C., Beckman J. S. Bactericidal activity of peroxynitrite. Arch Biochem Biophys. 1992 Nov 1;298(2):452–457. doi: 10.1016/0003-9861(92)90434-x. [DOI] [PubMed] [Google Scholar]

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