Micro in vitro test for chloroquine resistance of Plasmodium, falciparum: dicrepancies attributable to batch variation of plates

Increasing natural ingredient awareness and utilization has created an increased demand for sources of natural medicinal ingredients, including sources of compound used to treat malaria. Streptomyces is a genus of prokaryote well recognized for its production of antibiotics and other pharmaceutically useful compound. This study aimed to assess the ability of unpurified fermentation metabolites to inhibit Plasmodium parasites. A strain of bacteria identified as Streptomyces hygroscopicus subsp. hygroscopicus strain i18 were isolated from pineapple fields in Lampung province, and was cultured and fermented on liquid synthetic Gause medium for 10 days. The supernatant was separated from the cells and extracted with ethyl acetate-methanol (1:1). Plasmodium falciparum 3D7 was used for antiplasmodial testing. Metabolites were tested qualitatively using a phytochemical approach. Saponins and triterpenoids were found to be present in the extract. Parasite inhibition as measured using probit analysis and yielded an IC50 value of 11.07 g.m/L. These findings suggest further examinations of this extract (e.g. assessment of off-target effects) are warranted.

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A Rapid in Vitro Test for Chloroquine-Resistant Plasmodium falciparum

American Journal of Tropical Medicine and Hygiene ◽

10.4269/ajtmh.1990.42.521 ◽

1990 ◽

Vol 42 (6) ◽

pp. 521-526 ◽

Cited By ~ 3

Author(s):

Ilya Y. Gluzman ◽

Kano Nkangineme ◽

Augustine U. Orjih ◽

J. Tyler Martin ◽

Thomas E. Wellems ◽

...

Keyword(s):

Plasmodium Falciparum ◽

In Vitro Test ◽

Vitro Test

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VISUAL IN-VITRO TEST FOR DETERMINING THE DRUG SENSITIVITY OF PLASMODIUM FALCIPARUM

The Lancet ◽

10.1016/s0140-6736(82)92403-5 ◽

1982 ◽

Vol 319 (8285) ◽

pp. 1333-1335 ◽

Cited By ~ 8

Author(s):

KarlH. Rieckmann

Keyword(s):

Plasmodium Falciparum ◽

Drug Sensitivity ◽

In Vitro Test ◽

Vitro Test

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In vivo-in vitro test for chloroquine potentiation by cyproheptadine against Plasmodium falciparum

Transactions of the Royal Society of Tropical Medicine and Hygiene ◽

10.1016/0035-9203(91)90021-p ◽

1991 ◽

Vol 85 (2) ◽

pp. 206-207 ◽

Cited By ~ 2

Author(s):

Leonardo K. Basco ◽

Pascal Ringwald ◽

Jacques Le Bras

Keyword(s):

Plasmodium Falciparum ◽

In Vitro Test ◽

Vitro Test

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Simple, fast, and accurate fluorometric method to determine drug susceptibility of Plasmodium falciparum in 24-well suspension cultures.

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.40.4.835 ◽

1996 ◽

Vol 40 (4) ◽

pp. 835-838 ◽

Cited By ~ 23

Author(s):

L J Smeijsters ◽

N M Zijlstra ◽

F F Franssen ◽

J P Overdulve

Keyword(s):

Plasmodium Falciparum ◽

Drug Susceptibility ◽

Multidrug Resistant ◽

In Vitro Test ◽

Hoechst 33258 ◽

Ring Form ◽

Parasite Stage ◽

Drug Inhibition ◽

Vitro Test

An in vitro test which quantifies drug inhibition of Plasmodium falciparum replication by measuring the fluorescence intensity of Hoechst 33258 dye bound to DNA is described. The procedure does not require expensive reagents or equipment and can be completed in less than 10 min. The assay was highly accurate and sensitive: cultures with as few as 0.4% schizont-infected erythrocytes could reliably be analyzed. The method was not biased by the actual parasite stage used; i.e., the amount of fluorescence detected in a sample of a culture of mature schizonts equaled the amount detected with the ring form culture derived from these schizonts. Even the presence of large proportions of free merozoites, which are easily neglected in microscopic estimates, did not bias the results. Furthermore, measurement of the chloroquine susceptibility of the multidrug-resistant K1 strain and the chloroquine-susceptible NF54 strain showed that the method is most suitable for quantifying the drug resistance of P. falciparum.

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Assessment of Three In Vitro Tests and an In Vivo Test for Chloroquine Resistance in Plasmodium falciparumClinical Isolates

Journal of Clinical Microbiology ◽

10.1128/jcm.36.1.243-247.1998 ◽

1998 ◽

Vol 36 (1) ◽

pp. 243-247 ◽

Cited By ~ 14

Author(s):

Jean Bickii ◽

Leonardo K. Basco ◽

Pascal Ringwald

Keyword(s):

Host Factors ◽

Chloroquine Resistance ◽

In Vitro Assays ◽

In Vitro Tests ◽

In Vitro Test ◽

In Vivo Evaluation ◽

In Vivo Test ◽

Vitro Test

Three in vitro assays (the isotopic semimicrotest [700 μl per well; 24-well plates], the isotopic microtest [200 μl per well; 96-well plates], and the rapid in vitro test) and the standard in vivo test for chloroquine resistance were compared for 99 clinical isolates of Plasmodium falciparum obtained from symptomatic African patients. The 50% inhibitory concentrations determined by the two isotopic tests were similar and were highly correlated (r = 0.965; P < 0.05), showing a high concordance between the semimicrotest and the microtest. There was a moderate agreement between these two isotopic tests and the in vivo test. Most of the discordant results were probably due to host factors, including reinfections, pharmacokinetic variations, and immunologic response, which are eliminated in in vitro assays. The rapid in vitro test based on the inhibition of chloroquine efflux in the presence of verapamil was poorly concordant with the other tests. Despite some discordant results, isotopic in vitro assays are useful to characterize the phenotypes of individual isolates without the interference of host factors and are complementary to in vivo evaluation of drug efficacy. However, in vitro assays need to be standardized to allow direct comparison of results between different laboratories.

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Haemostatic Platelet Plug Formation in the Isolated Rabbit Mesenteric Preparation - An Analysis of Red Blood Cell Participation

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650069 ◽

1980 ◽

Vol 44 (01) ◽

pp. 006-008 ◽

Cited By ~ 12

Author(s):

D Bergqvist ◽

K-E Arfors

Keyword(s):

Whole Blood ◽

Platelet Rich Plasma ◽

Adenosine Diphosphate ◽

In Vitro Test ◽

Pharmacological Agents ◽

Vitro Test ◽

Releasing Effect ◽

Plug Formation

SummaryIn a model using an isolated rabbit mesenteric preparation microvessels were transected and the time until haemostatic plugs formed was registered. Perfusion of platelet rich plasma gave no haemostasis whereas whole blood did. Addition of chlorpromazine or adenosine to the whole blood significantly prolonged the time for haemostasis, and addition of ADP to the platelet rich plasma significantly shortened it. It is concluded that red cells are necessary for a normal haemostasis in this model, probably by a combination of a haemodynamic and ADP releasing effect.The fundamental role of platelets in haemostatic plug formation is unquestionable but there are still problems concerning the stimulus for this process to start. Three platelet aggregating substances have been discussed – thrombin, adenosine diphosphate (ADP) and collagen. Evidence speaking in favour of thrombin is, however, very minimal, and the discussion has to be focused on collagen and ADP. In an in vitro system using polyethylene tubings we have shown that "haemostasis" can be obtained without the presence of collagen but against these results can be argued that it is only another in vitro test for platelet aggregation (1).To be able to induce haemostasis in this model, however, the presence of red blood cells is necessary. To further study this problem we have developed a model where haemostatic plug formation can be studied in the isolated rabbit mesentery and we have briefly reported on this (2).Thus, it is possible to perfuse the vessels with whole blood as well as with platelet rich plasma (PRP) and different pharmacological agents of importance.

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