1408
Activity of the Anthelmintic Benzimidazoles against Giardia lamblia In Vitro Thomas D. Edlind, 10 L. Hang, and Prasanta R. Chakraborty
From the Department of Microbiology and Immunology, Medical College of Pennsylvania, Philadelphia, and the Department of Biochemical Parasitology, Merck Sharp & Dohme Research Laboratories, Rahway, New Jersey
The protozoan Giardia lamblia is a common parasitic agent of diarrheal disease in the USA and much of the world [1]. Chemotherapeutic treatment is recommended for all symptomatic cases and often for asymptomatic cyst-passagers. Currently, the most widely prescribed drugs are metronidazole, quinacrine, and furazolidone (for review, see [2]). However, problems associated with the use of these drugs include unpleasant side effects; activity extending to normal bacterial flora; high absorption, limiting activity against intestinal infection; and potential carcinogenicity. The aminoglycoside paromomycin has antigiardial activity [3] and has been recommended in pregnancy [2]. Its major advantage is lack of intestinal absorption; a disadvantage is its activity against normal intestinal bacteria. Lipophilic tetracyclines [4] and other nitroimidazoles [5] have activity but are deficient in one or more ways. Although G. lamblia is phylogenetically primitive by several criteria, in terms of cytoskeletal structures this organism appears highly evolved. In addition to its characteristic halfpear morphology, G. lamblia trophozoites have four pairs of flagella, a median body, and a ventral disk (involved in attachment to the intestinal microvilli). Microtubules are major components of all of these structures [6]. The l3-tubulin subunit of G. lamblia microtubules was recently characterized on the gene level [7]; there is extensive similarity to the l3-tubulins from unrelated organisms.
Received 5 February 1990; revised 11 June 1990. Grant support: Allegheny Singer Research Institute. Reprints or correspondence: Dr. Thomas Edlind, Department of Microbiology and Immunology, Medical College of Pennsylvania, Philadelphia, PA 19129. The Journal of Infectious Diseases 1990;162:1408-1411 © 1990 by The University of Chicago. All rights reserved. 0022-1899/90/6206-0033$01.00
An important group of anthelmintic agents, the benzimidazoles, target l3-tubulin [8]. Thiabendazole was introduced in 1961 and is still in use; the benzimidazole carbamates are more recent derivatives and include mebendazole and albendazole, currently approved for human use in many countries [9, 10]. Mebendazole is largely not absorbed from the gut and hence most useful for treating intestinal infections; albendazole is partly absorbed and is active against nonintestinal helminth forms. Mebendazole, in particular, has few side effects and no activity against the normal intestinal flora. Although fungi are susceptible to some benzimidazoles, activity against most protozoa either is lacking or has not been adequately investigated. One exception is Trichomonas vaginalis, susceptible in vitro to mebendazole and flubendazole but not thiabendazole [11]. Two small, uncontrolled clinical studies suggest, however, that G. lamblia may be another exception: all 5 giardiasis cases treated with albendazole were cured [12] and 38 of 40 were cured with mebendazole [13]. These studies, in conjunction with our interest in G. lamblia microtubule structure and genetics [7, 14], prompted us to examine the in vitro activity of benzimidazoles against this organism.
Materials and Methods Organisms and cultures. Trophozoites of G. Lamblia strains WB, LT, and PO were cultured at 37°C in TYI-S-33 supplemented with 10% fetal bovine serum, bile, and cysteine hydrochloride as previously described [3]. Penicillin (100 units/ml), streptomycin (100 ILg/ml), and amphotericin B (0.25 ILg/ml) were added during routine culture but omitted during experimental treatments. Growth inhibition assay. The susceptibility of G. Lamblia growth in vitro to the indicated drugs was determined as before [3]. Briefly. 2-ml cultures of log-phase organisms were exposed to drugs at various concentrations in fresh medium (minus antibiotics) for 48 h. Cell numbers, determined with a hemocytometer, are expressed as
Downloaded from http://jid.oxfordjournals.org/ at University of Iowa Libraries/Serials Acquisitions on March 18, 2015
In vitro growth of the protozoan parasite Giardia Lamblia was highly sensitive to certain anthelmintic benzimidazoles. Albendazole and mebendazole were 30- to SO-foldmore active than metronidazole and 4- to 40-fold more active than quinacrine. Thiabendazole, a noncarbamate benzimidazole, was less active. Since lack of intestinal absorption makes mebandazole an attractive new antigiardial agent, its in vitro activity was further characterized. At low concentrations (0.05 ILg/mI) mebendazole had a static effect on G. Lamblia growth; however, lethal activity was observed at a concentration fivefold lower (0.3 ILg/mI) than necessary for the cidal agent metronidazole. Two observations are consistent with a microtubule target for mebendazole. First, attachment of cells to the culture tube, mediated by the ventral disk and flagella, was rapidly disrupted by mebendazole treatment. Second, the characteristic cell structure was grossly distorted by treatment. No mebendazole-resistant G. Lamblia were detected in a population of 108 cells.
JID 1990;162 (December)
Concise Communications
Results
Inhibition ofgrowth in vitro. G. lamblia growth in vitro over 48 h in the presence ofdifferent drug concentrations was measured and expressed as a percentage of the growth in untreated cultures [3, 4]. The results in figure lA are for the prototype strain WB. The dose-response curves were used to estimate the IC so and IC 90 • The drug sensitivities of the geographically unrelated strains PO and LT were also examined (for quinacrine, strain WB only was tested). Metronidazole and quinacrine are the most widely prescribed antigiardial agents; mean IC so values of 1.5 (range, 1.3-1.6) and 0.2 J.tg/ml, respectively, were obtained for these agents. These values are similar to those reported by others using different assays (e.g. [5]). With quinacrine, a low percentage of cells remained viable even at high drug concentrations (figure lA); consequently, the IC 90 was relatively high (2.5 J.tg/ml). In contrast, metronidazole showed a steep dose-response curve (IC 90 = 2.8 J.tg/ml; range, 2.7-2.9). These differences prob-
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O+---r"TTT--.---.-....................-r----.----.Ii"...,:::;:::;:.,....,....-F=..-..-.-O;::::;::::;'.:;..,--. 0.1 10 0.01
Drug concentration (Jlg/ml) 0 - 0 albendazole
... -
. - . mebendazole 6 - 6 quinacrine ... metronidazole []-[] thiabendazole
Figure 1. A, Inhibition of Giardia Lamblia growth in vitro by selected benzimidazoles and, for comparison, quinacrine and metronidazole. B, Detachment of G. lamblia from the culture tube after 3 h in the presence of mebendazole, quinacrine, and metronidazole. All data points represent the average of two to four determinations from separate experiments.
ably reflect differences in cidal compared with static activity for these drugs (see below). The three benzimidazoles in current use for treatment of human helminth infections were tested for inhibition of G. lamblia growth in vitro. The dose-response curves for strain WB are shown in figure lA. Thiabendazole was only moderately active (IC so = 3.9 J.tg/ml; range, 3.0-5.0). In marked contrast, albendazole (IC so = 0.031 J.tg/ml; range, 0.0260.035) and mebendazole (IC so = 0.045 J.tg/ml; range, 0.0430.047) were highly active. These values are 30- to 50-fold lower than those achieved with metronidazole (60- to 75-fold on a molar basis). Quinacrine, at low concentrations, was nearly comparable in activity to mebendazole (ICso only fourfold higher). Unlike quinacrine, however, the benzimidazoles displayed steep dose-response curves (figure lA). Consequently, in terms of their IC 90 values, albendazole (0.059 J.tg/ml; range, 0.0460.066) and mebendazole (0.077 J.tg/ml; range, 0.075-0.078) were 30- to 40-fold more active than quinacrine. Although albendazole was slightly more active in vitro than mebendazole, it is also more highly absorbed from the gut. Consequently, its activity against a strictly intestinal infec-
Downloaded from http://jid.oxfordjournals.org/ at University of Iowa Libraries/Serials Acquisitions on March 18, 2015
a percentage of the cell number in control cultures. Mebendazole, metronidazole, quinacrine, and thiabendazole were obtained from Sigma Chemical (S1. Louis); albendazole was a gift from Smith Kline & French Laboratories (Philadelphia). Metronidazole and quinacrine were dissolved in water and the benzimidazoles in DMSO; the final vehicle concentration in the culture tubes was always48 h. Similar effects were seen with albendazole and thiabendazole at their inhibitory concentrations (not shown).
Two recent studies suggested that albendazole [12] and mebendazole [13] had clinical efficacy against giardiasis. It was important, however, to confirm that these benzimidazole carbamates were directly antigiardial in vitro. Our studies indicate that albendazole and mebendazole are considerably more active, by criteria including growth, viability, and attachment, than the currently recommended antigiardial agents. Thus, there is now adequate rationale to support larger, controlled clinical studies. Although albendazole was 1.5-fold more active than mebendazole, the latter is poorly absorbed from the gut and therefore would be preferable for noninvasive G. lamblia infections. The antigiardial activity of additional benzimidazoles, many of which have important veterinary applications, should be evaluated. The three geographically unrelated isolates of G. lamblia studied here were comparable in their benzimidazole susceptibilities. A larger number of isolates should now be examined. Benzimidazole activity against protozoan parasites other than G. lamblia and T. vaginalis also deserves investigation. These studies have additional implications in regard to characterizing the mechanism of action of this important group of drugs. Similar to other antigiardial agents, but in contrast to metronidazole, mebendazole at low concentrations is static, not cidal. This implies that its mechanism of action involves a reversible binding to its target. Considerable evidence from helminth, mammalian, and fungal systems supports a microtubule target for this drug [8]. A microtubule target in G. lamblia is supported by two observations: Treatment caused the cells to detach, at concentrations and time points well below those causing detachment with other drugs. Also, within hours treatment induced gross morphologic changes not seen with any other drug. Microtubules are major components of the ventral disk, which mediates attachment (coordinated flagella movement may be involved as well [6]), and of the cytoskeleton, which confers structure to the cell. More direct evidence that benzimidazoles are targeting G. lamblia microtubules is, however, needed. In particular, it is puzzling that flagellar activity was not obviously affected, since these have microtubule components. One explanation would be lack of access
of mebendazole to its tubulin target within the flagella. An interesting alternative is that flagella l3-tubulin, previously shown to be an isoform of cytoskeletal and ventral disk l3-tubulin [15], lacks the specific mebendazole binding site. Comparing the sequences of the three G. lamblia l3-tubulin genes [7, 14] may shed light on this issue. Note Added in Proof In agreement with our results, Meloni et al. have recently reported the superior activity of albendazole over metronidazole and tinidazole in vitro (Meloni BP, Thompson RCA, Reynoldson lA, Seville P. Albendazole: a more effective antigiardial agent in vitro than metronidazole or tinidazole. Trans R Soc Trop Moo Hyg 1990;84: 375-379).
References 1. Pickering LK, Engelkirk PG. Giardia lamblia. Pediatr Clin North Am 1988;35:565-577 2. Davidson RA. Issues in clinical parasitology: the treatment of giardiasis. Am J Gastroenterol 1984;79:256-261 3. Edlind TO. Susceptibility of Giardia lamblia to aminoglycoside protein synthesis inhibitors: correlation with rRNA structure. Antimicrob Agents Chemother 1989;33:484-488 4. Edlind TO. Tetracyclines as antiparasitic agents: lipophilic derivatives are highly active against Giardia lamblia in vitro. Antimicrob Agents Chemother 1989;33:2144-2145 5. Boreham PFL, Phillips RE, Shepherd RW. A comparison of the in vitro activity of some 5-nitroimidazoles and other compounds against Giardia intestinalis. J Antimicrob Chemother 1985;16:589-595 6. Erlandsen SL, Meyer EA, OOs. Giardia and giardiasis. New York: Plenum Press, 1984 7. Kirk-Mason KE, Thrner MJ, Chakraborty PRo Cloning and sequence of {3-tubulincDNA from Giardia lamblia. Nucleic Acids Res 1988; 16:2733 8. Lacey E. The role of the cytoskeletal protein, tubulin, in the mode of action and mechanism of drug resistance to benzimidazoles. Int J Parasitol 1988;18:885-936 9. Campbell WC, Rew RS. Chemotherapy of parasitic diseases. New York: Plenum Press, 1986 10. Okelo GBA. Albendazole, mebendazole and levarnisole. In: Peterson PK, Verhoef J, OOs. The antimicrobial agents annual/3. New York: Elsevier, 1988:266-274 11. Juliano C, Martinotti MG, Cappuccinelli P. In vitro effect of microtubule inhibitors on Trichomonas vaginalis. Microbiologica 1985;8: 31-42 12. Zhong HL, Cao WJ, Rossignol JF, Feng ML, Hu RY, Gan SB, Tan W. Albendazole in nematode, cestode, trematode and protozoan (Giardia) infections. Chin Moo J [Engl] 1986;99:912-915 13. AI-Waili NS, Al-Waili BH, Saloom KY. Therapeutic use of mebendazole in giardial infections. Trans R Soc Trop Moo Hyg 1988;82:438 14. Kirk-Mason KE, Thrner MJ, Chakraborty PRo Evidence for unusually short tubulin mRNA leaders and characterization of tubulin genes in Giardia lamblia. Mol Biochem Parasitol 1989;36:87-100 15. Torian BE, Barnes RC, Stephens RS, Stibbs HH. Thbulin and highmolecular-weight polypeptides as Giardia lamblia antigens. Infect Immun 1984;46:152-158
Downloaded from http://jid.oxfordjournals.org/ at University of Iowa Libraries/Serials Acquisitions on March 18, 2015
Discussion
1411
Activity of the Anthelmintic Benzimidazoles against Giardia lamblia In Vitro Thomas D. Edlind, 10 L. Hang, and Prasanta R. Chakraborty
From the Department of Microbiology and Immunology, Medical College of Pennsylvania, Philadelphia, and the Department of Biochemical Parasitology, Merck Sharp & Dohme Research Laboratories, Rahway, New Jersey
The protozoan Giardia lamblia is a common parasitic agent of diarrheal disease in the USA and much of the world [1]. Chemotherapeutic treatment is recommended for all symptomatic cases and often for asymptomatic cyst-passagers. Currently, the most widely prescribed drugs are metronidazole, quinacrine, and furazolidone (for review, see [2]). However, problems associated with the use of these drugs include unpleasant side effects; activity extending to normal bacterial flora; high absorption, limiting activity against intestinal infection; and potential carcinogenicity. The aminoglycoside paromomycin has antigiardial activity [3] and has been recommended in pregnancy [2]. Its major advantage is lack of intestinal absorption; a disadvantage is its activity against normal intestinal bacteria. Lipophilic tetracyclines [4] and other nitroimidazoles [5] have activity but are deficient in one or more ways. Although G. lamblia is phylogenetically primitive by several criteria, in terms of cytoskeletal structures this organism appears highly evolved. In addition to its characteristic halfpear morphology, G. lamblia trophozoites have four pairs of flagella, a median body, and a ventral disk (involved in attachment to the intestinal microvilli). Microtubules are major components of all of these structures [6]. The l3-tubulin subunit of G. lamblia microtubules was recently characterized on the gene level [7]; there is extensive similarity to the l3-tubulins from unrelated organisms.
Received 5 February 1990; revised 11 June 1990. Grant support: Allegheny Singer Research Institute. Reprints or correspondence: Dr. Thomas Edlind, Department of Microbiology and Immunology, Medical College of Pennsylvania, Philadelphia, PA 19129. The Journal of Infectious Diseases 1990;162:1408-1411 © 1990 by The University of Chicago. All rights reserved. 0022-1899/90/6206-0033$01.00
An important group of anthelmintic agents, the benzimidazoles, target l3-tubulin [8]. Thiabendazole was introduced in 1961 and is still in use; the benzimidazole carbamates are more recent derivatives and include mebendazole and albendazole, currently approved for human use in many countries [9, 10]. Mebendazole is largely not absorbed from the gut and hence most useful for treating intestinal infections; albendazole is partly absorbed and is active against nonintestinal helminth forms. Mebendazole, in particular, has few side effects and no activity against the normal intestinal flora. Although fungi are susceptible to some benzimidazoles, activity against most protozoa either is lacking or has not been adequately investigated. One exception is Trichomonas vaginalis, susceptible in vitro to mebendazole and flubendazole but not thiabendazole [11]. Two small, uncontrolled clinical studies suggest, however, that G. lamblia may be another exception: all 5 giardiasis cases treated with albendazole were cured [12] and 38 of 40 were cured with mebendazole [13]. These studies, in conjunction with our interest in G. lamblia microtubule structure and genetics [7, 14], prompted us to examine the in vitro activity of benzimidazoles against this organism.
Materials and Methods Organisms and cultures. Trophozoites of G. Lamblia strains WB, LT, and PO were cultured at 37°C in TYI-S-33 supplemented with 10% fetal bovine serum, bile, and cysteine hydrochloride as previously described [3]. Penicillin (100 units/ml), streptomycin (100 ILg/ml), and amphotericin B (0.25 ILg/ml) were added during routine culture but omitted during experimental treatments. Growth inhibition assay. The susceptibility of G. Lamblia growth in vitro to the indicated drugs was determined as before [3]. Briefly. 2-ml cultures of log-phase organisms were exposed to drugs at various concentrations in fresh medium (minus antibiotics) for 48 h. Cell numbers, determined with a hemocytometer, are expressed as
Downloaded from http://jid.oxfordjournals.org/ at University of Iowa Libraries/Serials Acquisitions on March 18, 2015
In vitro growth of the protozoan parasite Giardia Lamblia was highly sensitive to certain anthelmintic benzimidazoles. Albendazole and mebendazole were 30- to SO-foldmore active than metronidazole and 4- to 40-fold more active than quinacrine. Thiabendazole, a noncarbamate benzimidazole, was less active. Since lack of intestinal absorption makes mebandazole an attractive new antigiardial agent, its in vitro activity was further characterized. At low concentrations (0.05 ILg/mI) mebendazole had a static effect on G. Lamblia growth; however, lethal activity was observed at a concentration fivefold lower (0.3 ILg/mI) than necessary for the cidal agent metronidazole. Two observations are consistent with a microtubule target for mebendazole. First, attachment of cells to the culture tube, mediated by the ventral disk and flagella, was rapidly disrupted by mebendazole treatment. Second, the characteristic cell structure was grossly distorted by treatment. No mebendazole-resistant G. Lamblia were detected in a population of 108 cells.
JID 1990;162 (December)
Concise Communications
Results
Inhibition ofgrowth in vitro. G. lamblia growth in vitro over 48 h in the presence ofdifferent drug concentrations was measured and expressed as a percentage of the growth in untreated cultures [3, 4]. The results in figure lA are for the prototype strain WB. The dose-response curves were used to estimate the IC so and IC 90 • The drug sensitivities of the geographically unrelated strains PO and LT were also examined (for quinacrine, strain WB only was tested). Metronidazole and quinacrine are the most widely prescribed antigiardial agents; mean IC so values of 1.5 (range, 1.3-1.6) and 0.2 J.tg/ml, respectively, were obtained for these agents. These values are similar to those reported by others using different assays (e.g. [5]). With quinacrine, a low percentage of cells remained viable even at high drug concentrations (figure lA); consequently, the IC 90 was relatively high (2.5 J.tg/ml). In contrast, metronidazole showed a steep dose-response curve (IC 90 = 2.8 J.tg/ml; range, 2.7-2.9). These differences prob-
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CD
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c:
"ii
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-
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.!!l "ii u
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40 20
.B ~
O+---r"TTT--.---.-....................-r----.----.Ii"...,:::;:::;:.,....,....-F=..-..-.-O;::::;::::;'.:;..,--. 0.1 10 0.01
Drug concentration (Jlg/ml) 0 - 0 albendazole
... -
. - . mebendazole 6 - 6 quinacrine ... metronidazole []-[] thiabendazole
Figure 1. A, Inhibition of Giardia Lamblia growth in vitro by selected benzimidazoles and, for comparison, quinacrine and metronidazole. B, Detachment of G. lamblia from the culture tube after 3 h in the presence of mebendazole, quinacrine, and metronidazole. All data points represent the average of two to four determinations from separate experiments.
ably reflect differences in cidal compared with static activity for these drugs (see below). The three benzimidazoles in current use for treatment of human helminth infections were tested for inhibition of G. lamblia growth in vitro. The dose-response curves for strain WB are shown in figure lA. Thiabendazole was only moderately active (IC so = 3.9 J.tg/ml; range, 3.0-5.0). In marked contrast, albendazole (IC so = 0.031 J.tg/ml; range, 0.0260.035) and mebendazole (IC so = 0.045 J.tg/ml; range, 0.0430.047) were highly active. These values are 30- to 50-fold lower than those achieved with metronidazole (60- to 75-fold on a molar basis). Quinacrine, at low concentrations, was nearly comparable in activity to mebendazole (ICso only fourfold higher). Unlike quinacrine, however, the benzimidazoles displayed steep dose-response curves (figure lA). Consequently, in terms of their IC 90 values, albendazole (0.059 J.tg/ml; range, 0.0460.066) and mebendazole (0.077 J.tg/ml; range, 0.075-0.078) were 30- to 40-fold more active than quinacrine. Although albendazole was slightly more active in vitro than mebendazole, it is also more highly absorbed from the gut. Consequently, its activity against a strictly intestinal infec-
Downloaded from http://jid.oxfordjournals.org/ at University of Iowa Libraries/Serials Acquisitions on March 18, 2015
a percentage of the cell number in control cultures. Mebendazole, metronidazole, quinacrine, and thiabendazole were obtained from Sigma Chemical (S1. Louis); albendazole was a gift from Smith Kline & French Laboratories (Philadelphia). Metronidazole and quinacrine were dissolved in water and the benzimidazoles in DMSO; the final vehicle concentration in the culture tubes was always48 h. Similar effects were seen with albendazole and thiabendazole at their inhibitory concentrations (not shown).
Two recent studies suggested that albendazole [12] and mebendazole [13] had clinical efficacy against giardiasis. It was important, however, to confirm that these benzimidazole carbamates were directly antigiardial in vitro. Our studies indicate that albendazole and mebendazole are considerably more active, by criteria including growth, viability, and attachment, than the currently recommended antigiardial agents. Thus, there is now adequate rationale to support larger, controlled clinical studies. Although albendazole was 1.5-fold more active than mebendazole, the latter is poorly absorbed from the gut and therefore would be preferable for noninvasive G. lamblia infections. The antigiardial activity of additional benzimidazoles, many of which have important veterinary applications, should be evaluated. The three geographically unrelated isolates of G. lamblia studied here were comparable in their benzimidazole susceptibilities. A larger number of isolates should now be examined. Benzimidazole activity against protozoan parasites other than G. lamblia and T. vaginalis also deserves investigation. These studies have additional implications in regard to characterizing the mechanism of action of this important group of drugs. Similar to other antigiardial agents, but in contrast to metronidazole, mebendazole at low concentrations is static, not cidal. This implies that its mechanism of action involves a reversible binding to its target. Considerable evidence from helminth, mammalian, and fungal systems supports a microtubule target for this drug [8]. A microtubule target in G. lamblia is supported by two observations: Treatment caused the cells to detach, at concentrations and time points well below those causing detachment with other drugs. Also, within hours treatment induced gross morphologic changes not seen with any other drug. Microtubules are major components of the ventral disk, which mediates attachment (coordinated flagella movement may be involved as well [6]), and of the cytoskeleton, which confers structure to the cell. More direct evidence that benzimidazoles are targeting G. lamblia microtubules is, however, needed. In particular, it is puzzling that flagellar activity was not obviously affected, since these have microtubule components. One explanation would be lack of access
of mebendazole to its tubulin target within the flagella. An interesting alternative is that flagella l3-tubulin, previously shown to be an isoform of cytoskeletal and ventral disk l3-tubulin [15], lacks the specific mebendazole binding site. Comparing the sequences of the three G. lamblia l3-tubulin genes [7, 14] may shed light on this issue. Note Added in Proof In agreement with our results, Meloni et al. have recently reported the superior activity of albendazole over metronidazole and tinidazole in vitro (Meloni BP, Thompson RCA, Reynoldson lA, Seville P. Albendazole: a more effective antigiardial agent in vitro than metronidazole or tinidazole. Trans R Soc Trop Moo Hyg 1990;84: 375-379).
References 1. Pickering LK, Engelkirk PG. Giardia lamblia. Pediatr Clin North Am 1988;35:565-577 2. Davidson RA. Issues in clinical parasitology: the treatment of giardiasis. Am J Gastroenterol 1984;79:256-261 3. Edlind TO. Susceptibility of Giardia lamblia to aminoglycoside protein synthesis inhibitors: correlation with rRNA structure. Antimicrob Agents Chemother 1989;33:484-488 4. Edlind TO. Tetracyclines as antiparasitic agents: lipophilic derivatives are highly active against Giardia lamblia in vitro. Antimicrob Agents Chemother 1989;33:2144-2145 5. Boreham PFL, Phillips RE, Shepherd RW. A comparison of the in vitro activity of some 5-nitroimidazoles and other compounds against Giardia intestinalis. J Antimicrob Chemother 1985;16:589-595 6. Erlandsen SL, Meyer EA, OOs. Giardia and giardiasis. New York: Plenum Press, 1984 7. Kirk-Mason KE, Thrner MJ, Chakraborty PRo Cloning and sequence of {3-tubulincDNA from Giardia lamblia. Nucleic Acids Res 1988; 16:2733 8. Lacey E. The role of the cytoskeletal protein, tubulin, in the mode of action and mechanism of drug resistance to benzimidazoles. Int J Parasitol 1988;18:885-936 9. Campbell WC, Rew RS. Chemotherapy of parasitic diseases. New York: Plenum Press, 1986 10. Okelo GBA. Albendazole, mebendazole and levarnisole. In: Peterson PK, Verhoef J, OOs. The antimicrobial agents annual/3. New York: Elsevier, 1988:266-274 11. Juliano C, Martinotti MG, Cappuccinelli P. In vitro effect of microtubule inhibitors on Trichomonas vaginalis. Microbiologica 1985;8: 31-42 12. Zhong HL, Cao WJ, Rossignol JF, Feng ML, Hu RY, Gan SB, Tan W. Albendazole in nematode, cestode, trematode and protozoan (Giardia) infections. Chin Moo J [Engl] 1986;99:912-915 13. AI-Waili NS, Al-Waili BH, Saloom KY. Therapeutic use of mebendazole in giardial infections. Trans R Soc Trop Moo Hyg 1988;82:438 14. Kirk-Mason KE, Thrner MJ, Chakraborty PRo Evidence for unusually short tubulin mRNA leaders and characterization of tubulin genes in Giardia lamblia. Mol Biochem Parasitol 1989;36:87-100 15. Torian BE, Barnes RC, Stephens RS, Stibbs HH. Thbulin and highmolecular-weight polypeptides as Giardia lamblia antigens. Infect Immun 1984;46:152-158
Downloaded from http://jid.oxfordjournals.org/ at University of Iowa Libraries/Serials Acquisitions on March 18, 2015
Discussion
1411
