Variation in growth and drug susceptibility amongGiardia duodenalisassemblages A, B and E in axenicin vitroculture and in the gerbil model

Parasitology ◽  
2011 ◽  
Vol 138 (11) ◽  
pp. 1354-1361 ◽  
Author(s):  
E. BÉNÉRÉ ◽  
T. VAN ASSCHE ◽  
P. COS ◽  
L. MAES
Keyword(s):  
Oral Treatment ◽  
Field Isolate ◽  
Giardia Duodenalis ◽  
Laboratory Strain ◽  
Drug Susceptibility ◽  
Field Isolates ◽  
Generation Times ◽  
Subtype A ◽  
Assemblage B

SUMMARYThis study investigated the molecular and biological variation among differentGiardia duodenalisassemblages.In vitrogrowth and susceptibility to albendazole, fenbendazole, flubendazole, metronidazole, tinidazole and furazolidone was studied for laboratory (AI: WB, AII: G1 and B: GS/M-83-H7) and 6 field isolates of assemblage subtype AI, AII, B and EIII. Additionally, isolates of the 3 assemblages were evaluated in the gerbil upon 3-day oral treatment with albendazole (6 mg/kg), flubendazole (5 mg/kg) and metronidazole (20 mg/kg). Assemblage AIgrew significantly faster than all other assemblage subtypes, which showed comparable generation times. The assemblage A laboratory strains displayed alteredin vitrodrug susceptibilities compared to their matching AIor AIIfield isolate. No variation in drug susceptibility was observed between field isolates of assemblages A and E. However, assemblage A laboratory strains were more susceptible to the benzimidazoles and less susceptible to the nitro-imidazoles and furazolidone than the assemblage B laboratory strain. In the gerbil, no markedly different drug susceptibilities were observed. In conclusion, theGiardiaassemblage subtype can be associated with differences in growth characteristics rather than in drug susceptibility.

scholarly journals Piperaquine resistant Cambodian Plasmodium falciparum clinical isolates: in vitro genotypic and phenotypic characterization

2020 ◽  
Author(s):  
Nonlawat Boonyalai ◽  
Brian A Vesely ◽  
Chatchadaporn Thamnurak ◽  
Chantida Praditpol ◽  
Watcharintorn Fagnark ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Plasmodium Falciparum ◽  
Copy Number ◽  
Drug Susceptibility ◽  
Antagonistic Activity ◽  
Drug Combinations ◽  
Chloroquine Resistance ◽  
Phenotypic Characterization ◽  
Field Isolates

Abstract Background High rates of dihydroartemisinin-piperaquine (DHA-PPQ) treatment failures have been documented for uncomplicated Plasmodium falciparum in Cambodia. The genetic markers plasmepsin 2 ( pfpm2 ), exonuclease ( pfexo ) and chloroquine resistance transporter ( pfcrt ) genes are associated with PPQ resistance and are used for monitoring the prevalence of drug resistance and guiding malaria drug treatment policy.Methods To examine the relative contribution of each marker to PPQ resistance, in vitro culture and the PPQ survival assay were performed on seventeen P. falciparum isolates from northern Cambodia, and the presence of E415G-Exo and pfcrt mutations (T93S, H97Y, F145I, I218F, M343L, C350R, and G353V) as well as pfpm2 copy number polymorphisms were determined. Parasites were then cloned by limiting dilution and the cloned parasites were tested for drug susceptibility. Isobolographic analysis of several drug combinations for standard clones and newly cloned P. falciparum Cambodian isolates was also determined.Results The characterization of culture-adapted isolates revealed that the presence of novel pfcrt mutations (T93S, H97Y, F145I, and I218F) with E415G-Exo mutation can confer PPQ-resistance, in the absence of pfpm2 amplification. In vitro testing of PPQ resistant parasites demonstrated a bimodal dose-response, the existence of a swollen digestive vacuole phenotype, and an increased susceptibility to quinine, chloroquine, mefloquine and lumefantrine. To further characterize drug sensitivity, parental parasites were cloned in which a clonal line, 14-B5, was identified as sensitive to artemisinin and piperaquine, but resistant to chloroquine. Assessment of the clone against a panel of drug combinations revealed antagonistic activity for six different drug combinations. However, mefloquine-proguanil and atovoquone-proguanil combinations revealed synergistic antimalarial activity.Conclusions Surveillance for PPQ resistance in regions relying on DHA-PPQ as the first-line treatment is dependent on the monitoring of molecular markers of drug resistance. P. falciparum harbouring novel pfcrt mutations with E415G-exo mutations displayed PPQ resistant phenotype. The presence of pfpm2 amplification was not required to render parasites PPQ resistant suggesting that the increase in pfpm2 copy number alone is not the sole modulator of PPQ resistance. Genetic background of circulating field isolates appear to play a role in drug susceptibility and biological responses induced by drug combinations. The use of latest field isolates may be necessary for assessment of relevant drug combinations against P. falciparum strains and when down-selecting novel drug candidates.

scholarly journals Optimizing the HRP-2 in vitro malaria drug susceptibility assay using a reference clone to improve comparisons of Plasmodium falciparum field isolates

2012 ◽  
Vol 11 (1) ◽  
pp. 325 ◽  
Author(s):  
Wiriya Rutvisuttinunt ◽  
Suwanna Chaorattanakawee ◽  
Stuart D Tyner ◽  
Paktiya Teja-isavadharm ◽  
Youry Se ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Drug Susceptibility ◽  
Field Isolates ◽  
Susceptibility Assay

scholarly journals Bactericidal efficacy of two disinfectants against Brachyspira hyodysenteriae and one feed supplement against B. hyodysenteriae and B. pilosicoli

2012 ◽  
Vol 49 (No. 5) ◽  
pp. 156-160 ◽  
Author(s):  
D. Lobova ◽  
A. Cizek
Keyword(s):  
Type Strain ◽  
Field Isolate ◽  
Citrus Fruit ◽  
In Vitro Tests ◽  
Feed Supplement ◽  
Brachyspira Hyodysenteriae ◽  
Field Isolates ◽  
Bactericidal Efficacy ◽  
Type Strains

In vitro tests were used to evaluate bactericidal efficacy of two disinfectants on the basis of peroxygen compounds against one type strain and one field isolate of B. hyodysenteriae. Mean bactericidal concentrations (MBCs) of the two products ascertained with and without the load of organic matter of sterile pig faeces were several times lower than the recommended application concentrations. Bactericidal efficacy of an extract of citric seeds (feed supplement) against type strains of B. hyodysenteriae and B. pilosicoli, with six field isolates of B. hyodysenteriae and three field isolates of B. pilosicoli was also demonstrated, and its MBCs were determined. It was further determined that after 10 minutes exposure of 10% sterile pig faeces to field isolate and type strain of B. hyodysenteriae the efficacy of both disinfectants was 16 times higher than the concentration recommended by the manufacturer. The bactericide effect of citrus fruit extracts was exhibited at 0.05% concentrations after 5 min exposure, which is the same as recommended by the manufacturer.

scholarly journals Variability and variation in drug susceptibility among Giardia duodenalis isolates and clones exposed to 5-nitroimidazoles and benzimidazoles in vitro

2004 ◽  
Vol 54 (4) ◽  
pp. 711-721 ◽  
Author(s):  
Raúl Argüello-García ◽  
Maricela Cruz-Soto ◽  
Lydia Romero-Montoya ◽  
Guadalupe Ortega-Pierres
Keyword(s):  
Giardia Duodenalis ◽  
Drug Susceptibility

Investigation of effects ofGiardia duodenalison transcellular and paracellular transport in enterocytes usingin vitroUssing chamber experiments

Parasitology ◽  
2014 ◽  
Vol 142 (5) ◽  
pp. 691-697 ◽  
Author(s):  
KRISTOFFER R. TYSNES ◽  
LUCY J. ROBERTSON
Keyword(s):  
Epithelial Transport ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Field Isolate ◽  
Giardia Duodenalis ◽  
Short Circuit Current ◽  
Paracellular Transport ◽  
Detailed Knowledge ◽  
Transport Mechanisms

SUMMARYThe mechanisms by which different genotypes ofGiardia duodenalisresult in different symptoms remain unresolved. In particular, we lack detailed knowledge on which transport mechanisms (transcellular or paracellular) are affected by differentGiardiaisolates. Using horse radish peroxidase (HRP) and creatinine as transcellular and paracellular probes, respectively, we developed a robust assay that can be used with an Ussing chamber to investigate epithelial transport, as well as short-circuit current as an indicator of net ion transport. We investigated 2Giardiaisolates, both Assemblage A, one a lab-adapted strain and the other a field isolate. Results indicate that products from sonicatedGiardiatrophozoites increase both transcellular and paracellular transport. A non-significant increase in transepithelial electrical resistance (TEER) and short-circuit current were also noted. The paracellular transport was increased significantly more in the field isolate than in the lab-adapted strain. Our results indicate that while both transcellular and paracellular transport mechanisms may be increased following exposure of cells toGiardiatrophozoite sonicate, perhaps by inducing non-specific increases in cellular traffic, it is important thatin vitrostudies ofGiardiapathophysiology are conducted with differentGiardiaisolates, not just lab-attenuated strains.

scholarly journals Identification of compounds active against quiescent artemisinin-resistant Plasmodium falciparum parasites via the quiescent-stage survival assay (QSA)

2020 ◽  
Vol 75 (10) ◽  
pp. 2826-2834
Author(s):  
Thibaud Reyser ◽  
Lucie Paloque ◽  
Manel Ouji ◽  
Michel Nguyen ◽  
Sandie Ménard ◽  
...  
Keyword(s):  
Artemisinin Resistance ◽  
Laboratory Strain ◽  
Parasite Clearance ◽  
Quiescent State ◽  
Field Isolates ◽  
Pharmacological Targets ◽  
Survival Assay ◽  
Quiescent Stage ◽  
Selection Of

Abstract Background Quiescence is an unconventional mechanism of Plasmodium survival, mediating artemisinin resistance. This phenomenon increases the risk of clinical failures following artemisinin-based combination therapies (ACTs) by slowing parasite clearance and allowing the selection of parasites resistant to partner drugs. Objectives To thwart this multiresistance, the quiescent state of artemisinin-resistant parasites must be taken into consideration from the very early stages of the drug discovery process. Methods We designed a novel phenotypic assay we have named the quiescent-stage survival assay (QSA) to assess the antiplasmodial activity of drugs on quiescent parasites. This assay was first validated on quiescent forms from different artemisinin-resistant parasite lines (laboratory strain and field isolates), using two reference drugs with different mechanisms of action: chloroquine and atovaquone. Furthermore, the efficacies of different partner drugs of artemisinins used in ACTs were investigated against both laboratory strains and field isolates from Cambodia. Results Our results highlight that because of the mechanism of quiescence and the respective pharmacological targets of drugs, drug efficacies on artemisinin-resistant parasites may be different between quiescent parasites and their proliferating forms. Conclusions These data confirm the high relevance of adding the chemosensitivity evaluation of quiescent parasites by the specific in vitro QSA to the antiplasmodial drug development process in the current worrisome context of artemisinin resistance.

scholarly journals Inheritance of Carboxin Resistance in a European Field Isolate of Ustilago nuda

2000 ◽  
Vol 90 (2) ◽  
pp. 179-182 ◽  
Author(s):  
G. Newcombe ◽  
P. L. Thomas
Keyword(s):  
Mating Type ◽  
Host Plants ◽  
Single Gene ◽  
Field Isolate ◽  
In Planta ◽  
Tetrad Analysis ◽  
Field Isolates ◽  
The Past ◽  
Ustilago Nuda

Two carboxin-resistant field isolates of Ustilago nuda from Europe were crossed with a carboxin-sensitive field isolate from North America. Meiotic tetrads isolated from germinating F1 teliospores of one of the hybrids were tested for carboxin resistance and mating type. Carboxin resistance was shown to be controlled by a single gene (CBX1R), because a 1:1 segregation of carboxin resistance was observed in all 27 tetrads. Tetrad analysis indicated that the loci for carboxin resistance (Cbx1) and mating type (MAT1) segregate independently but may be located on the same chromosome. Tetrad analysis was not possible with the F1 hybrid of he other field isolate, and its resistance cannot yet be attributed to CBX1R. Carboxin resistance was qualitatively dominant to sensitivity in vitro, as demonstrated by triad analysis of germinating F1 teliospores. Quantitative in planta infection percents supported the conclusion that CBX1R is dominant, although incompletely, in the F1 hybrid of one of the field isolates. Also, fewer than expected carboxin-sensitive F2 individuals were observed in planta. However, inoculations of host plants with U. nuda have resulted in similar, unexpected variation in the past.

scholarly journals Piperaquine resistant Cambodian Plasmodium falciparum clinical isolates: in vitro genotypic and phenotypic characterization

2020 ◽  
Author(s):  
Nonlawat Boonyalai ◽  
Brian A Vesely ◽  
Chatchadaporn Thamnurak ◽  
Chantida Praditpol ◽  
Watcharintorn Fagnark ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Plasmodium Falciparum ◽  
Copy Number ◽  
Antimalarial Activity ◽  
Drug Susceptibility ◽  
Drug Combinations ◽  
Chloroquine Resistance ◽  
Phenotypic Characterization ◽  
Field Isolates

Abstract Background: High rates of dihydroartemisinin-piperaquine (DHA-PPQ) treatment failures have been documented for uncomplicated Plasmodium falciparum in Cambodia. The genetic markers plasmepsin 2 (pfpm2), exonuclease (pfexo) and chloroquine resistance transporter (pfcrt) genes are associated with PPQ resistance and are used for monitoring the prevalence of drug resistance and guiding malaria drug treatment policy.Methods: To examine the relative contribution of each marker to PPQ resistance, in vitro culture and the PPQ survival assay were performed on seventeen P. falciparum isolates from northern Cambodia, and the presence of E415G-Exo and PfCRT mutations (T93S, H97Y, F145I, I218F, M343L, C350R, and G353V) as well as pfpm2 copy number polymorphisms were determined. Parasites were then cloned by limiting dilution and the cloned parasites were tested for drug susceptibility. Isobolographic analysis of several drug combinations for standard clones and newly cloned P. falciparum Cambodian isolates was also determined.Results: The characterization of culture-adapted isolates revealed that the presence of novel PfCRT mutations (T93S, H97Y, F145I, and I218F) with E415G-Exo mutation can confer PPQ-resistance, in the absence of pfpm2 amplification. In vitro testing of PPQ resistant parasites demonstrated a bimodal dose-response, the existence of a swollen digestive vacuole phenotype, and an increased susceptibility to quinine, chloroquine, mefloquine and lumefantrine . To further characterize drug sensitivity, parental parasites were cloned in which a clonal line, 14-B5, was identified as sensitive to arteminsinin and piperaquine but resistant to chloroquine. Assessment of the clone against a panel of drug combinations revealed antagonistic activity for six different drug combinations. However, mefloquine-proguanil and atovoquone-proguanil combinations revealed synergistic antimalarial activity. Conclusions: Surveillance for PPQ resistance in regions relying on DHA-PPQ as the first-line treatment is dependent on the monitoring of molecular markers of drug resistance. P. falciparum harboring novel PfCRT mutations with E415G-exo mutations displayed PPQ resistant phenotype. The presence of pfpm2 amplification was not required to render parasites PPQ resistant suggesting that the increase in pfpm2 copy number alone is not the sole modulator of PPQ resistance. Genetic background of circulating field isolates appear to play a role in drug susceptibility and biological responses induced by drug combinations. The use of latest field isolates may be necessary for assessment of relevant drug combinations against P. falciparum strains and when down-selecting novel drug candidates.

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