Tritrichomonas foetus: Stable anaerobic resistance to metronidazole in vitro

Kulda, Jaroslav; Čerkasov, J.; Demĕs, P; Cerkasovova, A.

doi:10.1016/0014-4894(84)90068-7

Cited by 38 publications

(26 citation statements)

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“…It is detectable under anaerobic conditions and characterized by very high minimum lethal concentration (MLC) values in vitro (over 1000 µg metronidazole ml −" ). This type of resistance has been demonstrated in Tritrichomonas foetus Kulda et al, 1984). The loss of PFOR activity holds for a key attribute of the anaerobic resistance.…”

Section: Introductionmentioning

confidence: 99%

“…The anaerobic resistance resides in elimination of pathways responsible for the reductive activation of the drug. So far it has been demonstrated only in laboratory-developed strains (Brown et al, 1999 ;Kulda et al, 1984Kulda et al, , 1993. It is detectable under anaerobic conditions and characterized by very high minimum lethal concentration (MLC) values in vitro (over 1000 µg metronidazole ml −" ).…”

Section: Introductionmentioning

confidence: 99%

“…Resistance to metronidazole and related 5-nitroimidazole drugs has been demonstrated both in field isolates of Trichomonas vaginalis from patients refractory to treatment (Kulda et al, 1982 ;Lossick et al, 1986 ;Meingassner & Thurner, 1979 ;Meri et al, 2000 ;Mu$ ller et al, 1980) and in laboratory-developed strains obtained by exposing trichomonads to sublethal pressure of the drug either in vitro (Brown et al, 1999 ;Kulda et al, 1984Kulda et al, , 1993Tachezy et al, 1993) or in vivo (de Carneri et al, 1969 ;Meingassner et al, 1978). Two distinct types of resistance have been recognized and named aerobic and anaerobic according to conditions required for demonstration of resistance by susceptibility assays in vitro (Kulda, 1999).…”

Section: Introductionmentioning

confidence: 99%

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Mechanisms of in vitro development of resistance to metronidazole in Trichomonas vaginalis

et al. 2002

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Mechanisms of in vitro development of resistance to metronidazole in Trichomonas vaginalisDominique Rasoloson, SB te) pa! nka Van) a! c) ova! , Eva Tomkova! , Jakub Ra! zga, Ivan Hrdy! , Jan Tachezy and Jaroslav Kulda Development of resistance against metronidazole and mechanisms responsible for this process were studied in a sexually transmitted pathogen of humans, Trichomonas vaginalis. Monitoring of changes in metabolism and protein expression that accompanied increasing resistance of strains derived from a common drug-susceptible parent (TV 10-02) showed the multistep character of the process. The aerobic type of resistance known to occur in isolates from patients non-responsive to treatment appeared at the earliest stage, followed by development of the anaerobic type of resistance which was accompanied by gradual loss of hydrogenosomal proteins associated with drug-activating pathways [pyruvate:ferredoxin oxidoreductase (PFOR), hydrogenase, ferredoxin]. Unexpectedly, the loss of PFOR did not result in acquisition of full anaerobic resistance, thus indicating an alternative source of electrons required for the drug activation. These data suggest involvement of the oxidative decarboxylation of malate in hydrogenosomes, catalysed by NAD Mdependent malic enzyme and subsequent transfer of reduced equivalents to the drug via NADH :ferredoxin oxidoreductase and ferredoxin. Accordingly, all components of this pathway were eliminated before the resistance was fully developed. Resistant Trichomonas vaginalis compensated the impaired function of hydrogenosomes by enhanced conversion of pyruvate to lactate in the cytosol. Further analysis of the two key enzymes involved in metronidazole activation by Northern blotting and assay for nascent mRNA showed that the insufficient expression of the PFOR protein results from decreased gene transcription, while down-regulation of malic enzyme is controlled at the mRNA level.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Mechanisms of in vitro development of resistance to metronidazole in Trichomonas vaginalis

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“…T. foetus strain KV-1 (Kulda et al, 1984) was maintained in Trypticase-yeast extract-maltose (TYM) medium (Diamond, 1957) supplemented with 10 % heat-inactivated horse serum at pH 7?2.…”

Section: Methodsmentioning

confidence: 99%

Siderophore and haem iron use by Tritrichomonas foetus

Šuták

2004

Microbiology

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The ability of the parasitic flagellate Tritrichomonas foetus to use various iron sources for its physiological requirements was studied. The siderophores ferrioxamine B, ferrichrome, triacetylfusarinine, coprogen, enterobactin and pyoverdine sustained growth of the cells under iron-limited conditions, and siderophore iron was incorporated into the major iron protein of T. foetus, ferredoxin. The kinetics of siderophore uptake by the cells indicated that a non-saturable transport is involved, unlike the uptake of a ferrous salt. Siderophore uptake by the cells did not involve extracellular reductive dissociation of the ferric chelates, although T. foetus cells had some ferrireductase activity on ferric citrate. Fluorescent analogues of siderophores were used to show that the siderophores taken up by the cells were in small intracellular vesicles. The fluorescence emission maximum of pyoverdine in these intracellular vesicles shifted from 460 nm to 530 nm, indicating a very acidic environment. The results suggest that a wide range of chemically unrelated siderophores can be taken up non-specifically and efficiently used by T. foetus; the mechanism involved may be pinocytosis and removal of the iron from the siderophores in acidic intracellular vesicles. Haemin also sustained the growth of T. foetus cells under iron-limited conditions. The use of haemin iron by the cells probably involves haem oxygenase, since traces of biliverdin were found in the medium when haemin was the iron source. The iron uptake and ferrireductase activities of the cells do not seem to be regulated by the amounts of iron and copper in the growth medium.

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“…Prolonged cultivation of trichomonads with sublethal concentrations of metronidazole can result in the development of stable drug resistance accompanied by the loss of pyruvate: ferredoxin oxidoreductase activity (14,15). Metronidazole-resistant trichomonads show an altered carbohydrate metabolism.…”

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Pyruvate Decarboxylase, the Target for Omeprazole in Metronidazole-Resistant and Iron-Restricted Tritrichomonas foetus

Šuták

Tachezy

Kulda

2004

Antimicrob Agents Chemother

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The substituted benzimidazole omeprazole, used for the treatment of human peptic ulcer disease, inhibits the growth of the metronidazole-resistant bovine pathogen Tritrichomonas foetus in vitro (MIC at which the growth of parasite cultures is inhibited by 50%, 22 g/ml [63 M]). The antitrichomonad activity appears to be due to the inhibition of pyruvate decarboxylase (PDC), which is the key enzyme responsible for ethanol production and which is strongly upregulated in metronidazole-resistant trichomonads. PDC was purified to homogeneity from the cytosol of metronidazole-resistant strain. The tetrameric enzyme of 60-kDa subunits is inhibited by omeprazole (50% inhibitory concentration, 16 g/ml). Metronidazole-susceptible T. foetus, which expresses very little PDC, is only slightly affected. Omeprazole has the same inhibitory effect on T. foetus cells grown under iron-limited conditions. Similarly to metronidazole-resistant cells, T. foetus cells grown under iron-limited conditions have nonfunctional hydrogenosomal metabolism and rely on cytosolic PDC-mediated ethanol fermentation.Tritrichomonas foetus is a parasitic flagellate that causes sexually transmitted diseases in cattle that can lead to abortion and infertility. For decades the drugs of choice for the treatment of urogenital trichomoniasis were 5-nitroimidazoles, such as metronidazole ( Fig. 1), dimetridazole, and related derivatives, until they were subjected to regulatory actions in the United States, the European Union, and some other countries. The antitrichomonad activity of metronidazole depends upon reduction of its nitro group to short-lived free radicals, which cause multiple forms of cellular damage, including DNA breakage and subsequent cell death. Reductive activation of the drug proceeds in typical trichomonad organelles called hydrogenosomes, whose main function is a substrate-level synthesis of ATP linked to the production of molecular hydrogen. Electrons required for metronidazole reduction are generated by pyruvate:ferredoxin oxidoreductase, the key hydrogenosomal enzyme catalyzing oxidative decarboxylation of pyruvate (6,16,18).Prolonged cultivation of trichomonads with sublethal concentrations of metronidazole can result in the development of stable drug resistance accompanied by the loss of pyruvate: ferredoxin oxidoreductase activity (14, 15). Metronidazole-resistant trichomonads show an altered carbohydrate metabolism. The pyruvate-oxidizing pathway in the hydrogenosomes disappears, and the loss of ATP is compensated for by an increased rate of glycolysis in the cytosol. The dominant end product of glucose breakdown becomes ethanol, the production of which is negligible in metronidazole-susceptible trichomonads. The ethanol production depends on the activities of pyruvate decarboxylase (PDC) and alcohol dehydrogenase (2). While the activity of the latter enzyme is not markedly changed in metronidazole-resistant strains, the activity of PDC is strongly upregulated, suggesting that PDC is the rate-limiting enzyme in the production of et...

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Tritrichomonas foetus: Stable anaerobic resistance to metronidazole in vitro

Cited by 38 publications

References 10 publications

Mechanisms of in vitro development of resistance to metronidazole in Trichomonas vaginalis

Mechanisms of in vitro development of resistance to metronidazole in Trichomonas vaginalis

Siderophore and haem iron use by Tritrichomonas foetus

Pyruvate Decarboxylase, the Target for Omeprazole in Metronidazole-Resistant and Iron-Restricted Tritrichomonas foetus

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