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Metabolic studies of genetically heterogeneous isolates of the protozoan parasite Giardia duodenalis

Hall, Martin Leonard (1993) Metabolic studies of genetically heterogeneous isolates of the protozoan parasite Giardia duodenalis. PhD thesis, Murdoch University.

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Giardia duodenalis, a protozoan flagellate, is the causative agent of the debilitating disease giardiasis. Variation in symptomatology, course of infection and response to chemotherapy result in difficulties in the diagnosis and treatment of the disease. Though morphologically identical, variation in antigenicity, growth and metabolism between isolates could contribute to variable clinical manifestations.

In this study the ultrastructure, metabolism, response to chemotherapeutic agents and the effects, if any, of these, agents on metabolism were investigated in order to determine whether variation amongst these parameters con-elated with predetermined genetic heterogeneity in a number of isolates of G. duodenalis. The aims of this study were to investigate a threefold hypothesis: That genetic variation amongst isolates of G. duodenalis is reflected in i) metabolic variation ii) .variation in sensitivity to chemotherapeutic agents and iii) the effect of these chemotherapeutic agents on the metabolism of isolates.

Two genetically heterogeneous isolates of G. duodenalis were selected for observation using transmission electron microscopy during in vitro growth in the presence and absence of glucose. Neither of these isolates was found to contain structures resembling either endosymbiotic bacteria or viruses. Trophozoites of both isolates appeared morphologically normal when cultured in both the absence and presence of glucose, at all stages of growth. The ultrastructure of the two isolates differed in only one aspect: the protrusion of granular material through the disk in isolate BAH 12, but not in the Portland 1 isolate.

Metabolic studies were undertaken on three distinct isolates of G. duodenalis, Portland 1, BAC 1 and BAH 12. These isolates were compared to determine whether proven genetic differences were reflected in the carbohydrate metabolism of the parasite. The metabolic endproducts, alanine, acetate and ethanol were assayed during exponential growth of all three isolates. Studies were undertaken in both BIS-33 medium in the presence of glucose, and in serum-free medium where glucose was either present or absent. Results showed that the genetic differences between isolates are associated with quantitative differences in their metabolism.

Results of metabolite assays and transmission electron microscopy indicated that glucose and glucose-equivalent glycogen did not appear to be consumed by any of the three isolates. However, the BAH 12 isolate did not increase in cell numbers in the absence of glucose. As the levels of ethanol and acetate produced, in all isolates, were low in comparison to the error involved in glucose and glucose-equivalent glycogen determination, radioisotope studies were undertaken. These studies showed that glucose was being utilized, albeit at extremely low concentrations.

The effect of three chemotherapeutic agents, albendazole, metronidazole and chloroquine, on the growth and metabolism of five genetically and metabolically heterogeneous isolates of Giardia was studied. Albendazole was found to have a higher efficacy against the five isolates than either metronidazole or chloroquine. Variation existed between isolates in their sensitivity to all three drugs. However, no correlation was found between sensitivity and either rate of growth or genetic profile of the isolates.

All three drugs affected the energy production of isolates by diverting carbon flow from the production of ethanol toward the production of acetate and alanine to varying degrees. The only exception was the BAH 15 isolate.

This study has shown that genetic variation amongst isolates of G. duodenalis was reflected in metabolic variation and variation in sensitivity to chemotherapeutic agents between isolates. However, although variation was found, in the effect of chemotherapeutic agents on the metabolism of isolates, this variation could not be correlated with genetic diversity.

Item Type: Thesis (PhD)
Murdoch Affiliation: School of Veterinary Studies
Notes: Note to the author: If you would like to make your thesis openly available on Murdoch University Library's Research Repository, please contact: Thank you.
Supervisor(s): Thompson, Andrew and Costa, Nick
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