Date Published: February 13, 2008
Publisher: Public Library of Science
Author(s): Wendy R. Hansen, Daniel A. Fletcher, Anuradha Lohia
Abstract: BackgroundThe parasite Giardia lamblia must remain attached to the host small intestine in order to proliferate and subsequently cause disease. However, little is known about the factors that may cause detachment in vivo, such as changes in the aqueous environment. Osmolality within the proximal small intestine can vary by nearly an order of magnitude between host fed and fasted states, while pH can vary by several orders of magnitude. Giardia cells are known to regulate their volume when exposed to changes in osmolality, but the short-timescale effects of osmolality and pH on parasite attachment are not known.Methodology and Principal FindingsWe used a closed flow chamber assay to test the effects of rapid changes in media osmolality, tonicity, and pH on Giardia attachment to both glass and C2Bbe-1 intestinal cell monolayer surfaces. We found that Giardia detach from both surfaces in a tonicity-dependent manner, where tonicity is the effective osmolality experienced by the cell. Detachment occurs with a characteristic time constant of 25 seconds (SD = 10 sec, n = 17) in both hypo- and hypertonic media but is otherwise insensitive to physiologically relevant changes in media composition and pH. Interestingly, cells that remain attached are able to adapt to moderate changes in tonicity. By exposing cells to a timed pattern of tonicity variations and adjustment periods, we found that it is possible to maximize the tonicity change experienced by the cells, overcoming the adaptive response and resulting in extensive detachment.Conclusions and SignificanceThese results, conducted with human-infecting Giardia on human intestinal epithelial monolayers, highlight the ability of Giardia to adapt to the changing intestinal environment and suggest new possibilities for treatment of giardiasis by manipulation of tonicity in the intestinal lumen.
Partial Text: The parasitic protozoan Giardia lamblia is a major cause of diarrheal illness and infects millions of people per year, primarily via waterborne outbreaks in developed countries and long-term water contamination in developing countries ,. Giardia trophozoites infect many hosts, including humans, preferentially colonizing the proximal small intestine, which is sparsely populated by other microbes in comparison to the rest of the intestinal tract ,. Symptoms of giardiasis vary among patients, ranging from asymptomatic infections to malabsorption and severe chronic diarrhea. The health impacts of giardiasis can be severe, as chronic infection or reinfection may cause malnutrition and growth retardation ,. Giardiasis is generally treated with metronidazole or other nitroimidazoles, which target the parasite’s anaerobic metabolism , as well as with furazolidone or quinacrine. However, these chemotherapeutic agents are not always available, fully effective, or tolerated ,.
Giardia trophozoites rely on attachment to stay within the host and must accommodate a large range of perturbations in flow and chyme composition. In these experiments, we used live cell microscopy to monitor immediate detachment events upon exposure to test media in a flow chamber. Short-timescale detachment of Giardia has not been examined in the numerous studies of forced detachment, as most assays measure population attachment levels after a period of 2−24 hours incubation with a given drug or medium composition (e.g. –,). Such studies have shown that Giardia attachment and survival rates are inhibited by changes in pH and osmolality , but the long incubation prior to quantification of attachment makes it very difficult to separate direct effects on attachment from an overall effect on cell viability. Our results demonstrate that not only are Giardia susceptible to tonic shock, but that the response is rapid, with a detachment time constant of 25 seconds. This is consistent with the extremely rapid attachment and detachment behavior of trophozoites noted in the literature ,. Notably, remaining cells appear to adapt to the new tonicity and are not susceptible to further detachment unless subjected to a second tonic shock.