Final Report for FNE11-734
Project Information
Beyond the everyday production costs of the livestock farmer, feed,overhead, veterinarian, transport, PARASITIC SUPPRESSION of animal performance can be devastating economically. Moreover, nematode worm infestation is historically a troublesome burden on optimal production for graziers. For cattle and small ruminants, effective chemical anthelmintics have been a reliable crutch for standard non-sustainable farmers to count on, but this therapy has been losing its effectiveness with increasing worm resistance.Organic graziers have had to emphasize critical management stategies like rotational grazing and genetic selection, because a parade of ineffective ‘wormers’ or agents with distorted or anecdotal efficacy continue to cloud the literature.However, the Organic community must continue to seek non-chemotherapeutic anthelmintics, BECAUSE their existence would be highly useful to the organic grazier during emergency situations(no loss of the animal’s organic certification) or integrated into a planned parasite control strategy.
We performed a scientific on-farm study to try to establish the anthelmintic efficacy of the natural chemical Plumbagin by using Plumbago zeylanica plant extract.Plumbagin has been shown to have anti-parasitic properties invitro , and we sought to establish its effectiveness and safety invivo both as a preventative and therapeutic agent in sheep. Haemonchus contortus ,who has a devastating impact on grass-fed sheep production,was chosen as the studied nematode target.
Three trial groups(control,preventative, therapeutic) containing 10 feeder lambs each were studied using the Fecal Egg Count(FEC),Packed Cell(PCV), and Total Solid(TS) indices to measure Plumbagin’s efficacy after a challenge of 5000 infective H.contortus L3 larvae. Unfortunately, we demonstrated that Plumbagin was ineffectual as a preventative and therapeutic organic anthelmintic.
After initial positive data showing lower fecal egg counts and higher PCV’s in the Therapeutic group, results were found to be statistically insignificant.
Introduction:
A key USDA research project was published by Raymond Fetterer and Michael Fleming in 1991. The title of the scientific paper was " The Effects of Plumbagin on the Development of the Parasitic Nematodes Haemonchus contortus and Ascaris suum." Previous studies have shown that Plumbagin(see Picture #1),a naphthoquinone, found in extracts of plants of the Plumbago spp, has antimicrobial,antitumor, and inhibited insect development by presumably interfering with moulting. Plumbagin also has been implicated as an antiparasitic agent against Leishmania.This USDA study showed that Plumbagin had a marked dose-dependent effect on motility of H. contortus L1 larvae.At levels of 10ug/ml, larval motility was completely inhibited. Plumbagin also was effective against egg-hatching and significantly decreased recovery of infective H.contortus L3 larvae from the fecal slurry assay. Plumbagin was larvicidal against A.suum and had a partial inhibition of embryonic development against the tough A.suum egg shell. The paper concludes that Plumbagin is not likely to have practical use to control nematodes because of the relative high doses required for maximum effect;
HOWEVER, the conclusion was based on simple stomached animals. Black walnuts contain Plumbagin and other napthoquinones, like Juglone, and toxicosis has been documented in all simple-stomached animals, especially the horse......... But not Ruminants!! Also,the dosing of Plumbagin in this trial for 5 days will still be below the LD50 in mice. Contact with Cornell University School of Animal Science and others suggests that Plumbagin will not be degradated in the sheep's rumen allowing its passage to the home of the targeted H.contortus.....in the sheep's abomasum.
Our objectives in this study were 2-fold.First, was to establish Plumbagin as an organic anthelmintic in the Preventative role.
Secondly, to demonstrate Plumbagin's efficacy as a Therapeutic anthelmintic, that is, as a treatment against patent adult H.contortus infestations.We performed the triple chemical worming initialy on the study animals to chemically "cleanse" all of the patent nematode infestations the lambs may have been carrying.This has been shown to be effective because it is extremely unlikely that nematodes would be resistent to 3 separate anthlmintic categories,each killing in a different way:Albendazole is a benzamidazole,Levamisole is a imidazothiazole, and moxidectin is a semi-synthetic derivative of nemadectin.
A minimum 2 week "rest" period was necessary because moxidectin is an effective larvicidal for up to 2 weeks post administrion. In the Preventative role, we would expect the Plumbagin-treated Preventative group to have statistically significant lower fecal egg counts(FEC) than the Control group at the 1 month(Day0) follow-up FEC.
The P.zeylanica extract was dosed 2 days prior to L3 dosing to make sure Plumbagin was "on-board" to kill the infective L3 larvae when they were dosed........mimicking a highly infected pasture.We would not expect a significant difference in PCV's at 1 month because the patent infection of H.contortus adults would be so short(19-21 day pre-patent period).To refresh, H.contorus adults are so deadly because they suck blood in the sheep's abomasum causing anemia ie.lower PCV's and lower TS's In the Therapeutic role, we would expect the Pumbagin-treated Treatment group to have statistically significant lower FEC's and higher PCV's/TS's than the Control group at day 7,14,and 21 days post p.zeylanica root extract dosing at day0-day4.
We performed a day21 FEC to rule out a possible egg suppressive effect of Plumbagin vs a killing effect of H.contortus adults. We would expect to see an egg "Rebound" at day 21 if Plumbagin only suppressed H.contortus adult egg output.
Cooperators
Research
31 feeder lambs of similar ages were purchased by Green Alchemy Farm LLC in May 17, 2011.On May 21st after purchase , all lambs were weighed and given a thorough physical exam and mini blood panel to establish health for the trial. This step was performed by a veterinarian at Silver Maple Veterinary Clinic,Kuztown,Pa. Any sick lambs would have been rejected and replaced, but none were excluded.
All lambs were immediately treated by the veterinarian with the 3 chemical wormer combination of albendazole(Valbazen drench 0.75ml/25#BW),moxidectin(Cydectin drench at 1ml/11#BW), and levamisole(11.7grams/1qtH20 at 1ml/100#BW) to "cleanse" all lambs of all nematodes.This has been shown to be highly effective and was directed by Dr. Ray Kaplan at the University of Georgia Veterinary School and would focus the study on H.contortus alone and eliminate other confounding nematode infestations.
The lambs were subsequently split ito 3 study groups of 10 lambs a piece: Control, Preventative,and Therapeutic. Each group contained an equally distributed number of different sexes and a similar average weight per study group.
Three weeks later on June/11/2011, all three groups were dosed orally with 5000 infective L3 H.contortus larvae. We obtained these larvae from the University of Georgia School of Veterinary Medicine. This will simulate a highly contaminated pasture.
The Preventative group was simultaneously treated orally with a 33.3mg/kg dose of Plumbago zeylanica Root Extract 2 days prior and 3 days post treatment in dosed form attaining a 10ug/ml concentration in the abomasum.We utilized a soluble solution of 100mg/ml of standrdized Plumbago zeylanica root extract(obtained from Jinan Jiaquan International Trade co,LTD,China) containing 3% Plumbagin.This would establish a dose of a 1mg/kg dose daily of 100%Plumbagin over 5 days, or total dose of 5mg/kg of Plumbagin ,staying well below the 8mg/kg LD50 in mice.
All 3 groups were then released onto a "Clean" pasture and grazed for 4 weeks at which time H.contortus egg counts were established, demonstrating patent infestations.The 3 groups were allocated randomly to 3 equivalent areas on a hay field of equivalent plant distribution and which had never been grazed by small ruminants over the past decades. 7/10/2011,day1, fecal egg counts,PCV's,and TS's were performed by the veterinarian on all 3 groups and only the therapeutic group was dosed orally for 5 days in a row with 3% P.zeylanica Root Extract attaining a total dose of 5mg/kg of Plumbagin and reaching a 10ug/ml concentration in the abomasum.
Subsequently,fecal egg counts, PCV's,and TS's were performed on all 3 groups at day7 ,day 14, and day21. Its important to do the 21day fecal egg count to make sure fecal egg counts did not rebound post treatment. If high egg counts or dangerously low PCV's were found in any individuals of the 3 groups, Copper Oxide Particle capsules or moxidectin would be administered to endangered individuals to kill established H.contortus infestations,but eliminating them from the study. Luckily, no lambs developed low PCV's to be treated, but #20 lamb died acutely in the treatment group and was replaced by the extra control lamb #31.
Fecal egg counts(FEC) as eggs per gram , Packed Cell Volumes(PCV)and TS"s were established for all 3 groups on 7/11/2011, 7/17/2011, 7/26/2011, and 8/2/2011. FEC's , PCV's,and TS's were not performed on the Preventative group on 7/26/2011 and 8/2/2011 because this data was not required to substantiate Plumbagin's potential preventative function.
See Tables 1-3 to see Fecal Egg Counts. Graph #1 shows the comparison between the average fecal egg count between the Control and treatment groups(Multiply average count by 50 to get EPG).
This graph shows our initial indication that Plumbagin was efficacious(a 58% lower average FEC increase in Treatment vs Control groups &/10-8/2) as a therapeutic agent, BUT data was not statistically significant (Kaplan,2012).
Table # 4 shows the comparison of the the control and preventative FEC's.This demonstrated that the Preventative group did have lower FEC's thus eliminating Plumbagin as an effective preventative anthelmintic.
Tables 5-6 shows the Packed Cell Volume data for the Control and Treatment(Therapeutic) groups.Graph #2 shows the relationship of Average PCV for Control vs Treatment groups over time.
Table#7 shows the same comparison with raw data.There was a higher average PCV(5%) in the Treatment group from day 1 thru day 21 , but was considered statistically insignificant.Tables 8-10 show Total Solid data for the Control,Preventative and Treatment groups.Again,results were statistically insignificant.
- Table#1 Control Fecal Egg Counts
- Table#3 Treatment Fecal Egg Counts
- Table#8 Control TS’s
- Table#9 Preventative TS’s
- Table#10 Treatment TS’s
- Table #2 Preventative Fecal Egg Counts
- Graph #1 Average FEC Control vs Treatment over Time
- Table#4 Control vs Preventative FEC’s
- Table#5 Control PCV’s
- Graph#2 Control vs Treatment Average PCV’s over Time
- Table#7 Control vs treatment PCV’s
- Table #6 Treatment PCV’s
In this study we have striven to find an effective "organic" anthelmintic to add to the tools used by an organic farmer against GI nematodes. Unfortunately, we were unsuccessful in showing the efficacy of Plumbagin in a root extract of Plumbago zeylanica as either a preventative or therapeutic wormer against Hemonchus contortus in sheep.However, we have scientifically dismissed its potential anthelmintic function and that in itself is an accomplishment for the organic community.
Education & Outreach Activities and Participation Summary
Participation Summary:
NOFA-NY Research Symposium 2/2012 NOFA-Vt Winter Annual Conference 2012 Alvernia College Honor's Society 10/2011
Project Outcomes
Potential Contributions
The world of Organic Agriculture is both challenging and rewarding in it's desire to eschew the Industrial Farming that poisons our lives with chemicals,GMO's, and petroleum fertilizers that are all unsustainable! The quest for an effective Organic wormer /"Holy Grail" has been difficult based on literature review.There has been a renewed search for alternative dewormers mainly because of a number of factors.First,the increased multiple resistence of GI nematodes to chemical /commercial anthelmintics.Second,there has not been a push for new dewormers for small ruminants, esp goats, because there is not the economic pressure because of their representive small market .Thirdly, these agents are no longer cheap.Forthly, goats have very few LEGAL options for controlling these parasites.Fifthy, the burgeoning sustainable food market.The rule to be learned is that these chemical agents are a precious resource that should be used prudently and should only be reserved for recue situations.
So, based on literature review,what are an organic farmers choices for organic anthelmintics?? Garlic on the whole appears to have little response to nematodes based on a majority of the studies(B.Burke et al 2009,C. Strickland et al 2009 vs J.Noon SARE FNE032-482 2003). Commercial organic dewormers are also ineffective with studies: Molly's Complete Herbal Wormer(Burke et al .....Kaplan2009),Farmstead Wormer(Lugenbuhl et al 2006),Hoegger's Supply wormer and DE,garlic cloves and pyrethrum(1998 British Columbia study),Commercial mix of garlic powder,DE and Silica(SARE 2008 FNE05-230),Crystal Creek and Farmstead Health Supply dewormers(Noon SARE 2003).Wormwood spp have shown some efficacy in some studies (Eurasian wormwood MacCleod,1997;herba -alba wormwood Idris et al,1982;Cooper /Gordan,1996)Cucurbits showed potential in Shama et al 1976 using squash extracts and Forgacs et al 1970.Tansy seed showed potential in Papkanchov 1968.Papaya latex had positive effects against Ascaris suum in Satrija et al 1994.Evidence showing Diatomacious Earth(DE) as having anti-nematode activity has been contradictory at best.Copper Oxide Wire Particles(COWP) have shown scientific promise against H.contortus in small ruminants(Burke and Miller,2006;Burke et al 2004;Burke et al2007a;Soli....Burke, 2009;Burke,Soli,Miller et al 2009).Commercially available as Copasure boluses for cattle(Animax Vet),the boluses can be repackaged for use in small ruminants;however, copper blood levels would be wise before use in sheep.Can they be certified as organic SOON???They can be used with FAMACHA score>3 and response monitored with FEC's.
Because of the failure of many organic dewormers or lack of scientific research, the organic/sustainable grazier must incorporate a more comprehensive worming program using Genetics,FAMACHA scoring,Nutrition,High Condensed Tannin plants,Pasture management and rotation,proper administration of wormers,and finally selective vs whole herd treatment of organic wormers.Here the Holy Grail wormer represents only a small part of your Parasite Management Plan. Unfortunately, this study seems to eliminate Plumbagin from the list of potential organic compounds to be used as an anthelmintic,eventhough, earlier studies seemed to suggest promise.This should only encourage the further search for such agents scientifically........and thanks to SARE such studies are possible.