Human parasitic diseases

This is a list of topics related to human parasitic diseases. See also the categories shown below.

Diseases

Pathogens

Vectors and hosts

Antiparasitic drugs

SCREENING METHODS FOR ANTHELMINTICS

Test parasite should be taxonomically and biologically close to target parasite in humans. Screening models in suitable small animal hosts. 1. Antifilarial activity

^* In vitro methods:

Adult worms: - Litomosoides carinii, Acanthocheilonema viteae. Onchocerca gibsoni Setaria cervi Microfilareae: -L. carinii, Dirofilaria, Conspiculum guidense, S. cervi

(1) Setaria cervi

SINGHAL, K. C., SAXENA. P. N. and JOHRI. M. B. L. (1973): Studies on the use of Setaria cervi for in vitro antifilarial screening, Jap. J. Pharmacology, 23, 793-797.

Whole worm preparation Adult S. cervi (Nematoda: Filarioidea) obtained from peritoneal cavity of the freshly slaughtered cattle Transported to the laboratory in a vacuum flask containing modified Ringer’s solution, (NaCl-9g, CaCl2- 0.24g, KCl- 0.42g, NaHCO3- 0.5g,Glucose- 0.25g/Litre) 20 ml capacity isolated organ bath containing modified Ringer’s solution at 37% used for suspending the worm. Posterior end tied to hook at the bottom of the bath. The anterior end was attached to a frontal writing lever. At least 15 min allowed for a worm to stabilize Drug added in increasing conc., movements recorded on slow moving drum. Drug -- inactive if it does not modify movements in 10 minutes. Fresh worm used to test each dose of a drug.

Nerve – muscle preparation: Worm placed in petri dish containing modified ringer (370 C). 2 dissecting needles inserted at one end, cuticle split longitudinally. Intestine, uterus removed. Ant. 1 cm of worm removed to eliminate influence of nerve ring, cephalic ganglia Remaining part tied, suspended, tested as above.

(2) Onchocerca gibsoni Trop Med Parasitol. 1987 Jun; 38(2): 128-30 In vitro drug screening in isolated male Onchocerca gibsoni using motility suppression. Nowak M, Hutchinson GW, Copeman DB.

A primary in vitro screen was developed to screen for drug activity against Onchocerca volvulus. Assay estimates variation in motility through motility meter. Results compared favorably with reported in vivo tertiary screens for activity against Onchocerca species. Effects of these drugs were not reversible. Thus reduction in motility regarded as indicating significant metabolic damage.

(3) Acanthocheilonema viteae.

Jpn J Exp Med. 1990 Dec; 60(6): 303-9. A new technique of in vitro assay of antifilarials using different life forms of Acanthocheilonema viteae. Singh DP, Misra S, Chatterjee RK.

In vitro experiments conducted using three life-forms (adult, microfilariae and infective larva) of Acanthocheilonema viteae using different antifilarial agents.

Study indicated that this in vitro screening system can be used for primary screening of potential antifilarial agents provided three life forms of A. viteae are used simultaneously to avoid false negative results.

^* In vivo models:

(1) Litomosoides carinu – cotton rat system

Mites (Ornithonyssus bacoti) allowed to feed on Litomosoides infected cotton rats (1 week) Then on clean cotton rats 2 weeks (development in mites takes 2 wks.) De-mited – microfilareae in blood after 50 days Rats with at least 250 microfilareae/mm3 blood used Drug in 1/5 MTD given i.p. for 6 days Blood examined for microfilareae weekly Rats showing disappearance of microfilareae after 7 wks autopsied Absence of adult worm in pleural & peritoneal cavities observed.

(2) Setaria cervi – rat system:

Adult Setaria cervi collected from freshly slaughtered cattle. Rats (100- 150 gms), anaesthetized with ether, incision ½” long in abdominal wall 2 male & 2 female worms put in peritoneal cavity Peritoneum, body wall stitched, Antiseptic applied daily Microfilareae in blood after 10±3 days, last for 54± 6 days Rats showing Microfilareae for 3 consecutive days used Treated with test drug orally/ i.p. Complete disappearance of Microfilareae from blood for 3 consecutive days – evidence of antifilarial action.

(3) B. pahangi in Mongolian jirds (Meriones unguiculatus) G W Jeffers et al Infect Immun. 1984

Mongolian Jirds < 1 yr. used. s.c. inoculation of 100 infective larvae of B. pahangi obtained from infected mosquitoes. Animals sacrificed at 90,140,200 days post infection Total adult worm recovery Location of worms—heart, lymphatics Microfilareae in peripheral blood (4) Brugia malayi in Jirds (Maeda R et al Jpn J Exp Med. 1988)

Jirds infected subcutaneously with infective stage larvae (L3) of Brugia malayi evaluated as animal model for assessing macrofilaricides

Animals treated with a test compound

Change in microfilaria density observed

DEC at 50 mg/kg for 5 consecutive days for clearing the existing mf from the blood stream.

(5) Molinema dessetae in Proechimys oris Brienne MJ et al (J Med Chem. 1987)

Evaluated drugs for antifilarial activity against Molinema dessetae in vivo in its natural host, the rodent Proechimys oris

(6) Wuchereria kalimantani in leaf monkeys

Trials in leaf monkeys (Presbytis cristatus) infected with Wuchereria kalimantani. Kurniawan A et al (Southeast Asian J Trop Med Public Health. 1992)

Filaricidal drugs in given for 5 consecutive days to leaf monkeys (Presbytis cristatus) infected with Wuchereria kalimantani.

Optimal microfilaricidal effect occurred at 200-mg/kg body weight of Ivermectin.

(7) Setaria digitata microfilaraemia in Mastomys coucha: an animal model for chemotherapeutic and immunobiological studies. Mukhopadhyay S et al (Parasitology. 1996)

Intraperitoneal implantation of adult gravid females of the bovine filarial parasite, Setaria digitata in Mastomys coucha (multimammate mice) was found to induce microfilaraemia lasting for about 125 days.

Microfilariae could be detected 4 days post-implantation

Peak levels (about 30 mf /20 μl blood observed by 21 days

The mf in circulation eliminated by oral adm. of DEC indicating usefulness of the model for screening potential anti-microfilarial drugs.

Induction of antibodies to various fractionated antigenic components of adult parasites demonstrated by enzyme immunoassay in M. coucha implanted with live or cold-stunned adult worms.

S. digitata-M. coucha model thus found amenable to perform chemotherapeutic and immunobiological investigations in experimental filariasis.

(8) O. volvulus in chimpanzees Primate model for onchocerciasis research. Greene BM. (Ciba Found Symp. 1987) Infected 18 chimpanzees by s.c. inj. of 250 third-stage larvae of O. volvulus. Six received test drug on day 1, another six received test drug on day 28 after infection, and six received no drug. Four control animals received no infective larvae and no drug. Developed a pattern of infection that closely resembles that seen in humans- formation of nodules by adult worms, the subcutaneous distribution of microfilariae Antibody responses have also been increasing with time.

2. ECHINOCOCCUS GRANULOSUS (1) BALB/c mice infected with secondary equine E. granulosus

Parasitology. 1988 Apr;96 ( Pt 2):323-36 Echinococcus granulosus: the effects of praziquantel, in vivo and in vitro, on the ultrastructure of equine strain murine cysts. Richards KS, Morris DL, Daniels D, Riley EM.

Praziquantel (500 mg/kg) administered orally to BALB/c mice infected with secondary equine E. granulosus daily for 21, 30 or 30 + 30 days without the drug Ultrastructural examination of cysts showed increased vesiculation of the germinal layer leading, in many, to the loss of its integrity. Increased mitochondrial numbers occurred frequently. Longer drug treatments appeared to have greater effects on germinal layer There was no detectable reestablishment of structural organization within 30 days after drug withdrawal. Tissue from collapsed cysts was necrotic. In an in vitro study at praziquantel concentrations of 1000 and 5000 micrograms/l over a 10-day period, most cysts showed ultrastructurally a time- and concentration-dependent loss of integrity identical to that seen in vivo

(2) Echinococcus granulosus in Gerbils Morris DL, Taylor DH. ( J Helminthol. 1990)

Gerbils with well-developed peritoneal cysts of E. granulosus randomized to albendazole 50 mg/kg/day or untreated control.

Treated animals had less disease at post mortem after 3 months of treatment.

Cysts were then taken from both albendazole-treated and control animals and cultured in vitro either with or without albendazole sulphoxide (Alb Sx) 500 micrograms/L for 14 days.

Viability of cysts was then established by implantation of whole cysts into gerbils. 3. Trichinella spiralis (1) An in vitro screening test for compounds active against the parenteral stages of Trichinella spiralis.

Tropenmed Parasitol. 1981 Mar; 32(1): 31-4. (Jenkins DC, Carrington TS.) A new in vitro screening test for compounds showing activity against the tissue stages of Trichinella spiralis is described Freshly decapsulated larvae of the parasite are exposed to low concentrations of experimental compound in a medium capable of supporting the partial development of the worms. The screen detects the activity of those compounds known to be effective against the parenteral stages of the parasite.

(2) Trichinella pseudospiralis as a model for the "in vitro" screening of anthelmintics. Wiad Parazytol. 1986; 32(3): 303-11. (Gomez-Barrio A, Bolas-Fernandez F, Martinez-Fernandez AR.)

4. Hookworms. ^* Using Ancylostoma ceylanicum J Helminthol. 1981 Dec; 55(4): 273-8. (Misra A, Visen PK, Katiyar JC)

Test nematodes Ancylostoma ceylanicum in hamsters. Advantages: - Human infection - Easily maintained in lab animal (hamster) - Corresponds to human hookworms in sensitivity to drugs. Method: Hamsters (40- 60 gms) fed 60 ±5 larvae Mature – 13- 17 days Faeses examined for ova -- +ve used for screening Fasted overnight – test compound given orally – 3 consecutive days. Untreated – control, known antihookworm drug –std. Sacrificed on 3rd day of last dose – intestine examined for worms

% Activity= (N- n)/ N N= Avg no. of worms in control grp N= Avg no. of worms in treated grp. Simple micromotility recorder for rapid screening of potentially anthelmintic compounds.

Das AK, Bhattacharya S, Chatterjee GK, Chaudhuri SK. J Pharmacol Methods. 1988 Dec;20(4):323-7.

A micromotility recorder for monitoring the motility of small nematodes (adult) is described.

Normal motility of Ancylostoma ceylanicum and Nematospiroides dubius was recorded.

The time course to cause paralysis (paralysis time) was also observed in the presence of various anthelmintics at graded concentrations.

This is a simple in vitro model for screening of potential anthelmintic compounds.

^* Nippostrongylus brasiliense – rat system

Fertile eggs passed in faeces of infected rats

Faeces mixed with charcoal to creamy consistency –spread on central area of filter paper.

Incubated in dark at 270 C for 7 days.

Infective larvae develop- migrate to periphery – seen as fringe.

Rats infected with 200 larvae- s.c. inj. in neck

Eggs in faeces in 6-8 days

Drug adm. – 12th day.

Autopsy- 18th day-

Intestine examined between sheets of glass plates- red tinted worms seen

5. Tapeworms H. nana in mice

Used because - Human infection—easily maintained in mice - Armed scolex similar to other pathogenic tapeworms - Corresponds to other tapeworms in its sensitivity to standard anthelmintics

Methods Mature worms collected from infected mice Terminal gravid proglottids removed, crushed under coverslip—eggs removed Eggs containing hooklets (mature) counted 0.2 ml stock soln. containing 1000 eggs/ml given to each mouse. Adult worm develops- 15-17 days. Test drug given orally – autopsied on 3rd day Std. drug given Intestine examined under dissecting microscope for worms/ scolex Response – no. of mice cleared.

6. Anti-trichostrongyle activity . (1) A new primary screening test for anthelmintics utilizing the parasitic stages of Nippostrongylus brasiliensis, in vitro. Z Parasitenkd. 1980; 63(3): 261-9 Jenkins DC, Armitage R, Carrington TS.

Described in vitro test suitable for the large scale screening of chemical compounds for anthelmintic activity.

Utilizes fourth larval and adult stages of Nippostrongylus brasiliensis in medium capable of supporting its growth and development

Detects selectively those compounds that possess anti-trichostrongyle activity.

(2) Acetylcholinesterase secretion--a parameter for the interpretation of in vitro anthelmintic screens. Parasitology. 1986 Apr;92 ( Pt 2):425-30 Rapson EB, Chilwan AS, Jenkins DC

Novel biochemical parameter dependent upon spectrophotometric assay of acetylcholinesterase (AChE), secreted in large quantities by certain trichostrongylid nematodes, has been developed as index of activity.

Using Nippostrongylus brasiliensis, secretion of AChE in presence or absence of a large number of drugs in vitro was determined.

AChE was secreted by normal 4th larval and immature adult stages of the worm in a linear fashion.

Anthelmintics, dramatically reduced amount of enzyme secreted

7. Ascaris

(1) Mouse-ascaris suum test model. (Howes HL Jr. J Parasitol. 1971 Jun; 57(3): 487-93.)

(2) Ascaris suum in experimentally infected pigs.

Persistent activity of doramectin and ivermectin against Ascaris suum in experimentally infected pigs. Lichtensteiger CA et al; 1999

Study was conducted to investigate the persistent nematocidal activity of two avermectins against experimentally induced infections of Ascaris suum in swine.

Seventy-two nematode-free cross-bred pigs of similar bodyweight were randomly allotted to nine treatment groups of eight pigs each.

Eight of the groups were treated with injectable solutions containing 300 microg of doramectin/kg (IM) or 300 microg of ivermectin/kg (SC) either 0 (same day), 7, 14, or 21 days prior to an oral challenge of 50000 embryonated A. suum eggs.

The ninth group (control) was challenged in parallel without any avermectin treatment.

At 41 or 42 days after challenge, pigs euthanatized, adult & larval stages of A. suum were collected from the gastrointestinal tract of each pig and counted.

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Diseases

Pathogens

Vectors and hosts

Antiparasitic drugs

SCREENING METHODS FOR ANTHELMINTICS

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