Evaluation of efficacy expectations for novel and non-chemical

helminth control strategies in ruminants

Jennifer K. Ketzis a, Jozef Vercruysse b,*, Bert E. Stromberg c, Michael Larsen d,Spiridoula Athanasiadou e, Jos G.M. Houdijk e

a Novartis Animal Health, Basel, Switzerlandb Ghent University, Faculty of Veterinary Medicine, Department of Virology, Parasitology and Immunology,

Salisburylaan 133, B9820 Merelbeke, Belgiumc Department of Veterinary and Biomedical Science, College of Veterinary Medicine, University of Minnesota,

1308 Gortner Avenue, St. Paul, MN 55108, USAd Department of Veterinary Pathobiology, Danish Centre for Experimental Parasitology,

The Royal Veterinary and Agricultural University, DK-1870 Frederiksberg Copenhagen,

Copenhagen, Denmarke Animal Nutrition and Health Department, Scottish Agricultural College, West Mains Road,

Edinburgh, EH13 9DQ, Scotland, UK

www.elsevier.com/locate/vetpar

Veterinary Parasitology 139 (2006) 321–335

Abstract

The interest in novel methods of controlling helminth infections in ruminants is driven primarily by the development of

parasite resistance to currently available anthelmintics. While the purpose of anthelmintics is to achieve high efficacy, i.e.>90%

reduction of adult and/or larval parasites in the target host animal, the purpose of novel parasite control methods is rather to assist

in maintaining parasite infections below the economic threshold. The ability to maintain parasite levels below the economic

threshold is related not only to the efficacy of the control method, but also to the epidemiology of the parasites, climatic

conditions, the livestock management program, and integration in a sustainable parasite control program. Because of this

fundamental difference, novel parasite control methods need to be evaluated using efficacy criteria different from that adopted

for anthelmintics. Although the efficacy of novel parasite control methods may be demonstrated in classic dose-confirmation

studies, the impact on livestock production parameters can only be evaluated when tested on-farm. In this paper, the rationale for

evaluating novel methods differently from anthelmintics is reviewed, potential performance expectations are presented, and four

novel parasite control methods (vaccines, nematophagous fungi, condensed tannins, and immunonutrition) are assessed based on

the potential performance criteria.

# 2006 Elsevier B.V. All rights reserved.

Keywords: Helminths; Novel control methods; Guidelines; Duddingtonia; Vaccines; Immunonutrition; Tannins; Ruminants

* Corresponding author. Tel.: +32 9 264 73 90; fax: +32 9 264 74 96.

E-mail address: jozef.vercruysse@UGent.be (J. Vercruysse).

0304-4017/$ – see front matter # 2006 Elsevier B.V. All rights reserved.

doi:10.1016/j.vetpar.2006.04.022

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335322

1. Introduction

With few exceptions, control of parasites in

livestock relies almost exclusively on multiple and

regular dosing with anthelmintics. However, the use

of anthelmintics has several drawbacks including:

(1) the negative effect of preventive treatments on

the development of natural immunity against

helminths; (2) consumer concerns regarding drug

residues in food products and in the environment;

and, last but not least, (3) the increasing incidence of

parasite resistance against the available anthelmin-

tics (Corwin, 1997; Shaw et al., 1997; Thamsborg

et al., 1999; Vercruysse and Dorny, 1999). Conse-

quently, there are strong incentives for livestock

producers to adopt alternative or novel helminth

control methods.

Novel control methods can be used to manage

parasite infections (i.e., assist in maintaining below

the economic threshold) within a livestock production

system. An instrumental component of such methods

is that the elimination of the maximum number of

parasites from the ruminant is not necessary, and, in

fact, survival of some parasites in refugia can be of

benefit (van Wyk, 2001; Vercruysse and Dorny, 1999;

Waller, 1999). In contrast, based on the VICH

guidelines, anthelmintics are only considered effective

in the control of a parasite species (adult or larval

stages) when �90% is eliminated from the host

(Vercruysse et al., 2001). Unlike anthelmintics, novel

control methods usually do not have a direct effect on

parasite infections in the host. Instead, the mode-of-

action is based, for example, on improving the

immunity of the host or decreasing exposure of the

host to the parasite. Consequently, the use of novel

control methods constitutes a different approach

towards the control of helminth infections in

ruminants.

Achieving maximum efficacy of a novel control

method is highly dependent on understanding the

epidemiology of the parasite in the context of a

particular livestock management system. They are a

component of a control program and use as the sole

control method is not envisioned. For example, a novel

control method would be used to lower overall parasite

infection levels, while anthelmintics would continue

to be used (at a reduced frequency) as ‘‘clean-up’’ or

targeted treatments (Thamsborg et al., 1999; Ver-

cruysse and Dorny, 1999). Anthelmintics, on the other

hand, can be used as the primary component in a

parasite treatment program in situations where

anthelmintic resistance management is not a concern.

When anthelmintic use is timed with epidemiology

factors or combined with pasture rotation and other

management tools, their use can be decreased and

maximum benefits gained. However, neither epide-

miology nor management impact the efficacy obtained

with a single anthelmintic treatment. This is reflected

in the VICH guidelines, where the only critical

evaluation factor is the percentage of adult or larval

parasites directly killed or eliminated in the host

animal (Vercruysse et al., 2001). Epidemiology of the

parasite, the livestock management system, and

impact on production parameters are of limited

importance in the evaluation process recommended

by the guidelines.

Given these fundamental differences, products

related to novel control methods cannot be evaluated

using the same criteria as anthelmintics. Hence, to

better enable adaptation of the technologies by the

end-users (e.g., livestock producers) and ease the

process of registration, a set of guidelines and

standards must be established. Such standards and

guidelines also would significantly increase the ability

to compare different novel control methods and assist

in identifying and understanding the limitations of

novel control methods. A first step in developing these

standards is to determine the efficacy (or suggested

minimal performance) to be expected from the use of

novel control methods or products. However, measur-

ing the relative success of novel control approaches is

a very complex task.

In order to initiate a discussion on the develop-

ment of guidelines for novel control methods,

criteria for evaluating novel control methods are

suggested and reviewed in this paper. These criteria

should not serve as definitive guidelines, but provide

a basis for further dialogue on the subject. In Section

2, the impact of parasite epidemiology and the

economic threshold on efficacy is briefly reviewed.

In Section 3, proposed minimal performance

expectations are discussed. In Section 4, the efficacy

of four novel approaches (nematophagous fungi,

condensed tannins, immunonutrition, and vaccines)

is reviewed against the proposed minimal perfor-

mance expectations criteria.

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335 323

2. Epidemiology and the economic threshold

Parasite epidemiology determines when the eco-

nomic threshold is reached as well as the level of

economic impact. Both influence the use patterns of

novel control methods, as well as the criteria for

evaluating their impact. The concept of an economic

threshold and the influence of parasite epidemiology on

that threshold are extremely complex (several review

papers are available, e.g., Corwin, 1997; Vercruysse and

Claerebout, 2001). It is not the intention of this paper to

cover either subject in detail, but to consider them in

relation to how they influence (and perhaps limit) the

evaluation of novel control methods.

The economic threshold is the maximum number of

adult and/or larval parasites that a host can accom-

modate without experiencing a decrease in production

parameters. This threshold is dependent on, e.g. host

species (cow, sheep, goat), host physiological status

(immune status, prior exposure, nutritional level, and

production stage) and parasite species. The economic

threshold of a single species infection will change

when an additional parasite species is present. When

two or more parasite species are present, the number

of interactions dramatically increases as do the

possibilities for variations in economic thresholds.

In addition to the influence of the parasite species

present and their interactions, the economic threshold

is influenced by the infection level of each species

present, their epidemiology, environmental stress

(such as nutrition and climatic conditions) and the

livestock management system (Corwin, 1997; Strom-

berg, 1997; Stromberg and Averbeck, 1999; Ver-

cruysse and Claerebout, 2001).

Defining and determining the economic threshold

(predicting the level at which the animal will begin to

experience an impact on productivity) can be difficult

because of the number of variables that influence it

and the manner in which they interact with each other.

This is compounded by the fact that satisfactory,

standardized parameters for measuring the effect on

productivity (e.g., morbidity markers) are either

unavailable or not routinely used (Shaw et al.,

1997). For example, with first-season grazing calves,

methods proposed for distinguishing ‘sub-clinical’

infections include gastrin and pepsinogen levels

(Dorny et al., 1999; Hilderson et al., 1989; Vercruysse

and Claerebout, 2001). For adult dairy cows, bulk tank

milk Ostertagia ostertagi ELISA values have been

proposed as a method of predicting when infection

rates will be high enough to impact on milk production

(Charlier et al., 2005). The FAMACHA system (van

Wyk and Bath, 2002) can be used to identify sheep

suffering from anaemia (likely caused by H. con-

tortus). A diarrhoea index (DISCO) can be a good

indicator of actual nematode infection during the

summer and autumn in a temperate climate (Cabaret,

2004), and body condition scoring (BODCON) is a

potential method for identifying sheep requiring

anthelmintic treatment (van Wyk and Bath, 2002).

However, the value of these methods in different

climates and the relationship of the methods to animal

productivity need further research.

Once the economic threshold of a parasite infection

has been reached, the impact can be measured using a

variety of indicators. These can include weight loss or

reduced weight gain, reduced reproductive perfor-

mance (pregnancy rate and calving/lambing rate) and

milk production (Stuedeman et al., 1989; Stromberg

et al., 1997).

In general, the purpose of novel control methods is

to assist in maintaining parasite levels for a flock or

herd below the economic threshold and/or counteract

the negative impact of parasite infections. This may be

achieved by improving the host’s immune response to

the parasite, direct action on ingested larvae or adult

parasites in the host, or by decreasing exposure to

infective larvae (L3) on pasture. Decreasing exposure

can be realized by reducing the numbers of infective

larvae either by direct action on eggs, direct action on

L3 on the pasture, or decreasing fecundity of adult

parasites in the early grazing season. For each of these

‘‘mode-of-actions’’ a different evaluation method

might be required, e.g., faecal egg counts, the viability

and hatchability of eggs, the percentage of L3 that

develop and how long they survive on pasture. Given

the purpose of novel control methods, the decision of

when such a method should be used is based on its

mode-of-action, the epidemiology of the parasite and

how well it will work within a management system.

3. Proposed general performance expectations

A single efficacy criteria (e.g., >90% reduction of

parasite burden) cannot be applied to novel control

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335324

methods, as it is with anthelmintics. Also, given the

variety of mode-of-actions, a single method cannot be

adopted for measuring efficacy. However, novel

control methods should ideally fulfil minimal perfor-

mance expectations that can be quantified. These

should reflect the purpose of using novel control

methods, and therefore, include not only their likely

impact on parasites but also their economic impact and

the relationship between the two.

3.1. Efficacy

It should be possible to demonstrate the efficacy of

a novel control method in vivo, using the target

animal, both in a controlled (laboratory) and field (on-

farm) setting. The efficacy obtained should be

measurable and be possible to achieve with consistent

results. While larger on-farm efficacy variations are to

be expected, such variations should be explainable

based on climatic, epidemiology, and/or management

system factors. This analysis should help livestock

producers, who adopt a particular method, to under-

stand how and why performance varies.

The defined impact on parasites, measurement

method, and test method would vary with the novel

control method. Measurement methods could include

faecal egg counts (FEC), L3 in coprocultures, L3 on

pasture, parasite stages within the host, etc. In on-farm

studies, measurement methods which reflect field

conditions should be used in addition to the method

used in the laboratory. For example, if a novel control

method decreases L3, then in laboratory studies

coprocultures might be used to determine the number

of L3 developing per gram of faeces, while pasture

assessments of L3 and coprocultures would be

adopted for on-farm studies. The testing methods

used (e.g., pasture sampling, coprocultures) should be

standardized or the variability assessed to improve the

comparison of results from studies conducted by

different researchers.

3.2. Economic benefit

The use of a novel control method should result in

an expected and measurable economic effect when

used in defined settings (i.e., described climatic zones

or production systems such as cow–calf, year-round

lambing/grazing, etc.). Economic impacts could

include increased weight gain, improved feed con-

version, decreased host morbidity, decrease in specific

parasite-induced disease incidence, reduced use of

anthelmintics, etc. Variations in the type, consistency

and level of economic impact are likewise expected

due to differences in climate and management

systems. However, livestock producers should be

assured of the benefit if and when a novel control

method is used appropriately.

4. Efficacy of four novel approaches

4.1. Nematophagous fungi

Nematophagous fungi, such as Duddingtonia

flagrans, can colonize faeces and kill developing

parasite larvae. The fungus must be present in the

faeces at the same time as the parasite eggs and/or

developing larvae (Faedo et al., 2000). This novel

control method has no direct impact on pre-existing

infections in the ruminants. While several nemato-

phagous fungi have been examined (e.g., Harpospor-

ium anguillulae and Arthrobotrys spp.), D. flagrans is

the most investigated species (Larsen, 2000). Resting

spores (chlamydospores) of this fungus survive

passage through the gastrointestinal tract, and hence,

are present in freshly deposited faeces on pasture

together with parasite eggs. These spores germinate

and the fungus spreads, setting specialized trapping

mechanisms concomitantly with the hatching and

development of the free-living larval stages of the

parasites.

D. flagrans is used to prevent the build-up of L3

(third stage infective larvae) parasites on pastures

during mid- and late-grazing season. The objective is

to decrease the number of L3 to which grazing

ruminants are exposed. Hence, it should be adminis-

tered daily to animals contaminating the pasture

during the early season. This can vary from 6 to >10

weeks, dependent on the production system. It is

proposed that 8 weeks of continual use is sufficient in

climates with clearly defined winter and summer

weather cycles. Less is known about optimal use

patterns in climates with mild summers or long mild

falls and springs. However, various options have been

proposed for sheep and cattle under permanent

grazing situations (Waller, 2003).

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335 325

D. flagrans has been tested against several parasite

species in single and mixed infections in young

grazing sheep (Chandrawathani et al., 2004; Fontenot

et al., 2003; Githigia et al., 1997; Waghorn et al.,

2003), goats (Paraud and Chartier, 2003; Terrill et al.,

2004), and cattle (Dimander et al., 2003; Fernandez

et al., 1999; Larsen et al., 1995; Sarkunas et al., 2000).

Studies also have been performed with non-ruminants,

but are not discussed here (Baudena et al., 2000;

Larsen et al., 1996; Nansen et al., 1996; Petkevicius

et al., 1998). While many laboratory, plot, and field

studies have been conducted, only a few publications

are referenced here.

Based on publicly disclosed trial data, best results

are obtained when D. flagrans spores are fed daily to

cattle at a dose of 1 � 106 spores/kg body weight. The

recommended dose for sheep and goats is still

pending but appears to range from 0.25 � 106 to

0.5 � 106 spores/kg body weight.

D. flagrans meets the previously defined general

performance expectations as described in the follow-

ing sections.

4.1.1. Efficacy of fungi

Efficacy of D. flagrans is determined either by

showing (1) a decrease in L3 recovered from faeces or

around faecal pats in plot studies (for animals on

pasture or in controlled laboratory studies) or (2) a

decrease in the intensity of infections in tracer animals

(placed on pastures previously grazed by D. flagrans

fed animals). Properly conducted on-farm studies with

D. flagrans are complex and expensive, because the

treatment comparisons (ideally replicated) should be

conducted on pastures that are identical in regards to

prior use, contamination levels at study start, and

forage content. To better evaluate D. flagrans under

field conditions, more standardization in techniques

for collecting larvae from pastures and laboratory

procedures (e.g., coprocultures) would be beneficial.

Coprocultures show the most consistent results, while

other test methods are more influenced by sampling

method, environmental factors, and grazing patterns.

In coprocultures and plot studies, a consistent

decrease of L3 counts has been demonstrated with

single infections of a range of parasite species,

including Cooperia, Ostertagia, Teladorsagia, Hae-

monchus, Trichostrongylus, and Oesophagostomum.

Efficacy of >90% has been recorded with cattle

parasites and >70% with sheep parasites. D. flagrans

has been found to be less effective against parasites

with slow moving larvae, such as Dictyocaulus

viviparus, when it exists as a single species infection

(Henriksen et al., 1997). Also, it was found to have no

effect against Muellerius (Paraud and Chartier, 2003).

Efficacy, however, seems to improve against some of

the species with slow moving larvae in the presence of

mixed infections. It remains uncertain whether a

reliable reduction of Nematodirus can be obtained

(Faedo et al., 2000; Githigia et al., 1997).

Decreased L3 counts can be demonstrated on-farm

with the same parasites (single and mixed infections)

as in the laboratory. Efficacy variation increases (due

to fungal isolate used, host, parasite species, manage-

ment system, and environmental conditions), but the

reduction in herbage infectivity can exceed 90% for

cattle and 60% for sheep and goats (Faedo et al., 1998,

2000; Fernandez et al., 1999; Fontenot et al., 2003;

Larsen et al., 1995). In other cases, however, only

negligible reductions in pasture parasite contamina-

tion levels have been observed which could be

attributed to adverse environmental factors. Also, it

has been hypothesized that the higher efficacy of D.

flagrans often recorded against bovine parasites

compared to sheep and goats, is due to differences

in the faecal mass and/or composition. That is, cattle

faeces, acting as a microclimatic buffer, provide a

more ideal environment for the development and

activity of the fungi. Also, cattle dung pats provide

good conditions for larvae to develop and survive,

thereby providing an increased stimulus of trap

formation by the growing fungus.

4.1.2. Economic benefits of fungi

A decrease in L3 pasture levels has resulted in

many different measurable parameters, such as

reduced use of anthelmintics (Hertzberg personal

communication), lower host infections in late season

(Chandrawathani et al., 2004; Fontenot et al., 2003;

Githigia et al., 1997; Knox and Faedo, 2001; Larsen

et al., 1995; Waller et al., 2004), increased weight gain

compared to untreated control animals (Chandra-

wathani et al., 2004; Dimander et al., 2003; Fernandez

et al., 1999; Nansen et al., 1995; Waller et al., 2004),

and/or earlier time to market (Waller et al., 2004). The

economic benefits will depend on the farm or/and

management system. Studies are still needed to

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335326

confirm whether the use of fungi allows for a less

frequent use of anthelmintics in a variety of farm

settings, and the relationship of the economic benefits

achieved to the parasite species present.

Based on the data available, D. flagrans as a novel

control method seems to meet the criteria listed in

Section 3. However, a shortcoming that is likely to

limit widespread on farm adoption is the need to daily

dose the animals with fungal spores. The oral

administration of D. flagrans spores must occur when

contamination of pastures with parasite eggs is the

source of seasonal peaks in pasture infectivity. Often,

livestock are not offered supplements during these

critical time periods (e.g., when grazing on lush green

spring pastures). One way to overcome this obstacle

would be to deliver spores through sustained-release

delivery devices. The timing of such a device would be

critical, i.e. to coincide with the shedding of nematode

eggs onto the pasture. Hence, livestock producers

would need to give the sustained-release formulation

2–3 weeks after exposure of grazing animals to

infected pastures; application much earlier or later

than this critical period would reduce any potential

economic benefits. Also, livestock producers would

have to realize that such expected economic benefits

are likely to be reduced during extremely dry summers

(when parasite pressure is unusually low), or

extremely wet summers (when parasite pressure is

unusually high). Hence, the use of D. flagrans spores

within a parasite control program and the expectation

in terms of economic benefits need to be carefully

considered in relation to expected infection pressure.

4.2. Tannins

Plants produce a range of secondary metabolites

that may have anti-parasitic properties (for a recent

review, see Houdijk and Athanasiadou, 2003). Plants

with such anti-parasitic properties are increasingly

referred to as bioactive forage. Here, we will focus on

tanniferous plants, and more specifically on their

active components, the condensed tannins. Nemato-

cidal activity of such tannins has been reported as

early as the 1960s (Taylor and Murant, 1966) and more

recently, has gained renewed interest as potential

component of non-chemical parasite control strate-

gies. We focus on tanniferous plants and their

condensed tannins because, relative to other potential

bioactive forages, there is a large body of quantitative

in vitro and in vivo evidence on their anti-parasitic

properties, which is required in the context of this

paper. However, the concepts would be equally

applicable for other bio-active forages, which have

shown promising anti-parasitic properties but from

which active compounds are yet to be identified, such

as chicory (Hoskin et al., 2000; Tzamaloukas et al.,

2005).

The antiparasitic effects of condensed tannins are

believed to be mediated via either a direct anthelmintic

and/or an indirect nutritional mechanism. Direct

anthelmintic-like effects have been demonstrated in

in vitro assays, which have shown that incubation in

crude condensed tannins extracts reduced the devel-

opment, viability, motility, and migratory ability of

parasite larvae (Athanasiadou et al., 2001; Butter

et al., 2001; Molan et al., 2002). The direct action of

condensed tannins in most in vitro studies could be

defined as strong, dose related, and with efficacy levels

approaching 100%. Direct anthelmintic-like effects

have also been demonstrated in in vivo studies, where

the addition of condensed tannins to high quality foods

has resulted in significantly reduced worm burdens

and nematode egg counts with 50% or more

(Athanasiadou et al., 2000; Butter et al., 2001). In

these studies, high quality foods were used in order to

avoid confounding direct anthelmintic-like effects of

condensed tannins with their indirect nutritional

effects. Indirectly, condensed tannins can affect

parasitism through increasing protein availability in

the lower gastrointestinal tract of ruminants. Most

condensed tannins have the ability to achieve this

through the protection of dietary protein from rumenal

degradation. As such, effects of condensed tannins in

low quality foods may be mediated through increased

protein supply, which is known to enhance immunity

to parasites (see Section 4.3).

Because of this potential two-way action of

condensed tannins against parasites, supplementation

of parasitized animals with condensed tannins or

grazing on condensed tannin-rich forages, could result

in a reduced anthelmintic use for parasite control

(Ramirez-Restrepo et al., 2005). However, in addition

to their reported anti-parasitic activity, the consump-

tion of condensed tannins is well known to be

associated with anti-nutritional effects, such as

reduced food intake and digestibility (Dawson

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335 327

et al., 1999; Reed, 1995; Acamovic and Brooker,

2005). Therefore, the assessment of benefits of

condensed tannins ingestion on the performance of

parasitised hosts has to be considered as the outcome

of a trade off between their positive, anti-parasitic

properties and negative, anti-nutritional properties

(see below).

Data on condensed tannins are available to meet

some of the criteria described in Section 3.

4.2.1. Efficacy of tannins

The direct efficacy of condensed tannins has been

demonstrated in pen-based drench and feed supple-

mentation studies. A condensed tannin extract

administered as a drench to parasitized sheep resulted

in a 50% reduction of egg excretion and 30%

reduction of parasite burden as compared to

undrenched sheep (Athanasiadou et al., 2000,

2001). Parasitized sheep offered foods that contain

a commercially available condensed tannin extract,

had reduced nematode egg excretion when compared

to sheep offered the same diet without condensed

tannins (Butter et al., 2000). Similarly, a 30%

reduction in FEC has been observed in goats offered

condensed tannin-containing forages (Kahiya et al.,

2003), or commercially available condensed tannins

(Paolini et al., 2003) as supplement.

Field studies have focused on the antiparasitic

effects of condensed tannins when animals are grazed

on condensed tannin-rich forages. For example, sheep

grazing on forages such as Lotus corniculatus and

Hedysarium coronarium, which contain moderate to

high concentrations of condensed tannins (20–60 g

condensed tannins per kg food), showed reduced level

of parasitism and improved performance when

compared to sheep grazing on pastures of comparable

nutrition, but without these forages (Marley et al.,

2003; Niezen et al., 2002). Goats grazing on Sericea

lespedeza, a tannin-rich forage, showed a 60% FEC

reduction and daily egg output compared to those

grazing on low tannin forages (Min et al., 2004). The

effect of condensed tannins does seem to vary between

ruminant hosts and parasite species. In sheep,

condensed tannins are active against intestinal but

not against abomasal nematodes, whilst in goats,

abomasal and intestinal nematodes appear to be

equally susceptible to condensed tannins (Athanasia-

dou et al., 2001; Paolini et al., 2003).

In general, the efficacy achieved with condensed

tannins is lower in grazing studies compared to pen

studies (for which up to 60% efficacy has been

reported). This is likely to be due to more variable and

complex conditions on-farm than those indoors, as

environmental and climatic conditions interfere with

the epidemiology of the parasite and the anthelmintic

properties of condensed tannin-rich forages. Also, the

efficacy obtained in supplementation studies is lower

than what is achieved in vitro. However, in all types of

studies, standard methods of assessing effects of

condensed tannins can be used. Relatively consistent

results are feasible in in vitro and controlled studies.

More research is needed on the percentage of

tanniferous forage required on pasture to obtain

consistent results.

4.2.2. Economic benefits of tannins

Although the consumption of condensed tannins

can reduce the level of parasitism in ruminants, the

observed reduction may not necessarily exceed the

economic threshold. In addition, the consumption of

condensed tannins may not result in economic benefits

because of the negative effects of excessive condensed

tannin consumption on the performance of the hosts.

For example, the consumption of tannins has been

associated with a reduction of food intake and food

digestibility, impairment of rumen metabolism and

mucosal toxicity (Dawson et al., 1999; Reed, 1995). In

supplementation studies, the performance of para-

sitized animals treated with condensed tannins was not

improved compared to that of unsupplemented

animals (Butter et al., 2000). Similarly, the adminis-

tration of a condensed tannin drench to parasitized

sheep resulted in a reduced level of parasitism but

performance remained unaffected (Athanasiadou

et al., 2000, 2001).

Economic benefits might be more pronounced in

grazing studies. For example, it has been demonstrated

that sheep grazing on condensed tannin-rich forages

showed a reduced level of parasitism and improved

performance when compared to sheep grazing on

tannin-free pastures (Marley et al., 2003; Niezen et al.,

2002). However, it is not certain whether this

improved performance was the direct consequence

of a reduced larval intake, an anthelmintic-like effect,

or of grazing high quality forages. The latter might

have been the case, as the majority of tanniferous

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335328

forages are leguminous and usually are of higher

nutritive value than conventional grazing pastures. It is

therefore possible that the improved performance

observed in animals grazing tanniferous forages is a

consequence of improved resilience (i.e., the outcome

of grazing high quality leguminous forages per se

whilst facing parasitism), rather than the consequence

of improved resistance (i.e., through anthelmintic-like

effects).

In conclusion, there is no strong evidence for

condensed tannin consumption improving the perfor-

mance of parasitized sheep to a level that results in an

economic benefit. One reason for this is the lack of

experiments where the direct effects derived from

tannin consumption are distinguished from those

associated with the nutritional profile of the grazing

forages that contain tannins. Once these potential

benefits have been eliminated, the antiparasitic and

negative effects of condensed tannin ingestion on

parasitized host performance should be considered and

evaluated simultaneously. More research is required on

the level of condensed tannins needed to lower the level

of parasitism and improve performance of parasitized

ruminants. Research on condensed tannins might not

necessarily result in a commercially available product,

but instead could increase the value placed by livestock

producers on incorporating tanniferous forages in

pastures and enhance the understanding of how pasture

management can impact parasite infections.

4.3. Immunonutrition

Whilst it has long been known that the adverse

effects of gastrointestinal nematode parasitism on

productivity of the host can be reduced through

improved nutrition, an increasing body of more recent

evidence shows that improved host nutrition may, in

turn, affect gastrointestinal nematodes by improving

host immunity (see Houdijk and Athanasiadou, 2003

for a recent review). Effects of nutrient supply on

resistance and resilience to gastrointestinal nematodes

can largely be accounted for through a nutrient-

partitioning framework (Coop and Kyriazakis, 1999).

This framework introduces the concept of partial

prioritization for the allocation of scarce resources

toward bodily functions: the higher the priority, the

less likely nutrient scarcity would affect this bodily

function.

The framework puts forward the hypothesis that

scarce nutrient allocation to maintenance functions

has the highest priority, and that scarce nutrient supply

to growth and reproductive functions are prioritized

over expression of immunity to parasites. Therefore,

nutrient scarcity leads to reduced productivity in

parasitized animals, as scarce nutrients would be

allocated to the repair or replacement of parasite-

induced damaged tissue rather than to productive

tissues. In addition, nutrient scarcity would penalize

expression of immunity to parasites. Consequently,

improved nutrition would lead to enhanced immunity

(resistance) and increased productivity (resilience) in

parasitized animals. Most research on nutritional

control of parasitism has concentrated on protein

nutrition. However, the concept would be equally

applicable to any scarce nutrient, provided it is rate

limiting for growth and/or reproductive functions and

expression of immunity to parasites (Houdijk and

Athanasiadou, 2003).

4.3.1. Efficacy of immunonutrition

Controlled studies have shown that supplementa-

tion with a wide range of protein sources including

soybean meal, fish meal, cottonseed meal, sunflower

meal and urea, can reduce FEC and worm burdens in

growing and periparturient ruminant hosts. Some

studies are highlighted here to illustrate this point (for

a recent review see Houdijk and Athanasiadou, 2003).

Bown et al. (1991) infused casein in the abomasum of

growing T. colubriformis infected sheep, and worm

burdens were reduced by up to 55%. In other studies,

urea and fishmeal supplementation in T. colubriformis

infected sheep reduced the degree of parasitism by

30% (Knox and Steel, 1999) and 44–99% (van Houtert

et al., 1995), respectively. Protein supplementation

also reduced the number of nematodes by 40% in H.

placei infected calves (Gennari et al., 1995). Protein

supplementation to periparturient ewes, in the form of

xylose-treated soybean meal and fishmeal, reduced

FEC and worm burdens by more than 60% (Donaldson

et al., 2001; Houdijk et al., 2003a). Dose–response

experiments in parasitized growing lambs and

periparturient ewes indicated that the level of protein

nutrition required to improve resistance and/or

resilience is in the order of 20–25% above estimated

protein requirements for maintenance (Datta et al.,

1998; Donaldson et al., 2001; Houdijk et al., 2003a).

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335 329

The rate at which protein supply can improve

expression of immunity is important for determining

its viability as a novel approach. This has been

assessed by reducing protein demand at times of

reduced expression of immunity (Houdijk et al.,

2003b). Two groups of lactating ewes were fed at�0.6

times protein requirements, with one group twin-

rearing throughout and another group changed to

single-rearing at day 10 of lactation. The FEC of the

latter group of ewes rapidly decreased by 75% after

day 10, and were within a matter of days, similarly low

as that of ewes fed at �1.2 times protein requirements

and twin-rearing throughout. These data support the

view that FEC in ewes are relatively sensitive to

changes in the degree of protein availability and that

protein supplementation can rapidly contribute to

reductions in pasture infectivity.

The above discussion illustrates that, under well-

controlled conditions, nutritional control of gastro-

intestinal nematode parasitism may well be feasible,

and the development of a nutrient-partitioning frame-

work (Coop and Kyriazakis, 1999) has made its

outcome increasingly predictable. The reduced nema-

tode egg excretion and worm burdens observed could

contribute to reducing the degree of parasitism in the

direction of the economic threshold. Under conditions

where the ewe is the major source of infection to her

lambs, through her contribution to pasture infectivity,

such reduction will be expected to result in reduced

larval intake of the lambs, and, hence in reduced

penalties on growth performance.

Few studies have been published on nutritional

control of gastrointestinal nematode parasitism on a

larger scale, i.e. at farm (grazing) level. Papado-

poulos et al. (2000) supplemented naturally infected,

grazing goats with soybean meal and observed

reduced FEC and worm burdens. Keatinge et al.

(2003) supplemented soybean meal to naturally

infected, organically managed grazing ewes and

observed improved lamb weight gain and reduced

FEC during lactation. Similarly, protein supplemen-

tation reduced the degree of parasitism in peripar-

turient ewes (Kahn et al., 2003) and dairy goats

(Chartier et al., 2000). While these results are

generally in agreement with data from the controlled

studies, more such studies are needed to draw firm

conclusions on the efficacy of nutritional control of

parasitism at farm level.

4.3.2. Economic benefits of immunonutrition

Cost-benefit analysis of nutritional control of

gastrointestinal nematode parasitism has yet to be

given the attention it deserves. In our view, such a

cost-benefit assessment would need to take into

account at least the following points. Improved

protein nutrition at times of protein scarcity will

invariably improve resilience in the face of parasit-

ism. Compared to their unsupplemented counterparts,

supplemented, parasitized lambs and kids will grow

faster, and supplemented parasitized ewes and goats

will produce heavier offspring and more milk. The

higher milk production will contribute to higher pre-

weaning weight gain of the lambs and kids and

maternal body weight at weaning. In addition,

improved protein nutrition can reduce worm burdens

and FEC. The former may contribute to reduced

losses due to parasitism per se, whilst the reduced

FEC can contribute to reduced pasture infectivity.

Hence, protein supplementation to ewes and goats has

the potential to result in a two-sided advantage for the

farmer in terms of lamb and kid growth performance,

both contributing to finishing them earlier in time.

The latter is usually associated with higher carcass

prices.

All these benefits need to be evaluated against the

costs of the additional feeding, which includes the cost

of the protein supplements themselves as well as the

extra time spent on feeding. In the short term, these

costs are larger than those using anthelmintics.

However, the benefits of nutritional control of

parasites in terms of productivity are often manifested

over a longer period of time. Hence, this novel

approach requires a cost benefit analysis with a

relative long perspective. Recently, a study has been

published in which effects of supplementary feeding

in goats on resistance and resilience at the farm level

have been evaluated for its costs and benefits, relative

to the use of anthelmintics (Torres-Acosta et al.,

2004). The authors concluded that supplementary

feeding did not greatly improve resistance but did

improve resilience, and was as such economically

viable. This cost benefit analysis was conducted in

accordance with the above suggestions and applied to

this specific farm situation. Further reports of this

nature are required to confirm a possible economic

feasibility of nutritional control of gastrointestinal

nematode parasitism in grazing livestock.

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335330

4.4. Vaccines

Similar to other control methods, the degree of

protection needed by parasitic vaccines is difficult to

estimate due to the numerous variables involved, e.g.

parasite species, climate, management (Vercruysse

and Claerebout, 2003). Furthermore all variables

which may affect the performance of helminth

vaccines, following exposure to L3 on pasture, have

not been studied. Helminth vaccine efficacy and the

levels of protection deemed necessary for commercial

viability have, until now, mostly been estimated by the

percentage reduction in number of parasites shortly

after vaccination. However, the major goal in

controlling helminth infection by vaccination should

aim at changing the host–parasite relationship by

reducing the infection level (i.e., morbidity) and also

reducing transmission by decreasing the number of

viable nematode eggs that are deposited in faeces on

pasture. Both of these potential attributes of vaccines

need to be evaluated differently with different efficacy

and length of action expectations.

4.4.1. Efficacy of vaccines

The efficacy expectation for O. ostertagi, Hae-

monchus contortus, D. viviparus and Fasciola

hepatica, four economically important helminths of

ruminants, will be briefly discussed as examples. For

each helminth, parameters that can be used to evaluate

efficacy and the likely minimum percentage efficacy

needed are described (within the limits of available

information).

O. ostertagi of cattle. To control O. ostertagi by

vaccination, is essentially to immunize first-season

grazing calves before turn-out (i.e., worm-free when

vaccinated), thus protecting them against a relatively

low over-wintered parasite challenge (Shaw et al.,

1998). Reducing infection levels is, therefore, not a

priority as these low over-wintering larval populations

rarely induce disease. In contrast, reduced egg excretion

should be the target, as the number of worm eggs shed

during the first part of the grazing season determines the

number of infective larvae on the pasture in the second

half of the grazing season. Duration of protection

should be approximately 2–3 months, since in Western

Europe, egg output peaks at around 2 months after

turnout (Shaw et al., 1997, 1998). An Ostertagia

vaccine in cattle should thus mainly be an anti-

fecundity vaccine. Because fecundity of Ostertagia is

highly density dependent, an anti-infection vaccine, i.e.

a vaccine that reduces the number of worms, would only

be successful if it had an extremely high efficacy level.

Efficacy in practical terms should, therefore, mainly be

based on faecal egg count and/or egg fertility reduction

over a period of at least 2 months.

For Western Europe, efficacy expectations will

depend on (1) the level of over-wintering infections

and (2) the length of the pasture season. Shaw et al.

(1998) showed that when faecal egg counts were

higher than 200 epg on day 56 post turn-out, at least

89% of calves showed parasitic gastroenteritis later in

the first grazing season. In contrast, the mean faecal

egg counts of chemoprophylactically treated calves, or

calves without obvious parasitic gastroenteritis

remained mostly below 100 epg. It should be stressed

that these threshold counts are based on data from

naturally infected calves, where less than 50% of the

eggs were from O. ostertagi, while the majority of

eggs excreted in the beginning of the pasture season

are predominately from Cooperia spp. Hence, it can

be estimated that for calves turned-out in May and

housed in autumn, efficacy expectations should be

approximately 50–60% reduction of the cumulative

faecal egg counts (over a 2-month period) compared to

a control group where the peak mean faecal egg count

is at least 100 epg (O. ostertagi infection). Efficacy

expectations considered useful in Western Europe

may be different on other continents and should be

adapted according to local epidemiological situations

(Vercruysse and Claerebout, 2003).

H. contortus of sheep. In contrast to O. ostertagi,

animals may well encounter high pasture infestations

with H. contortus at turn-out, leading to clinical

disease. Also, the fecundity of H. contortus is not

regulated by the intensity or duration of the infection,

and there is a very good correlation between total daily

faecal egg counts and mature female worm burdens

(Coyne and Smith, 1994). Consequently, to protect

animals against high infections and to prevent the

build-up of high pasture infestations, a vaccine needs

to reduce the number of adult worms present in the

animals, either by reducing the establishment of

infective larvae (immune exclusion), or by increasing

the mortality of established worms. Efficacy expecta-

tions for a vaccine against H. contortus will thus differ

from O. ostertagi (Vercruysse and Claerebout, 2003).

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335 331

Different effects of vaccination against H. con-

tortus in a grazing population of lambs were simulated

in a model using a larval vaccine affording 50% or

80% protection against larval stages and an adult

vaccine giving 80% protection against adult worms.

As expected, the model predicted that the use of

vaccines that confer higher levels of immunity will

have greater effects than those conferring lower levels

of immunity, but a threshold of protection that is

needed to protect the animals from acquiring harmful

worm burdens during the entire grazing season was

not determined (Meeusen and Maddox, 1999).

D. viviparus of cattle. D. viviparus is the only

helminth against which there is a commercially

available vaccine. The existing lungworm vaccine,

based on irradiated infective larvae, induces a

temporary protection against re-infection which

requires natural boosting within the same grazing

season before it results in solid immunity. The aim of

the vaccine is to prevent clinical outbreaks, but little

information on the reduction in number of worms and/

or faecal larvae required is available. Poyntner et al.

(1970) reviewed the experience gained with the

vaccine and recorded an efficiency (= prevention of

husk) in the field of around 98%. Breakdowns were

mainly due to faulty grazing practices, which led to the

sudden exposure of vaccinated calves to heavy pasture

contamination.

Lungworm vaccines should, therefore, prevent the

development of ingested larvae. To establish the

percentage efficacy of a lungworm vaccine remains

extremely difficult because of the unpredictable onset

and level of the initial infection an animal will

encounter. However, considering the low infection

levels able to produce disease (less than 100 worms), a

lungworm infection should have relatively high

efficacy (potentially >95%).

Fasciola spp. of sheep. The goal of Fasciola spp.

vaccines should be, as for H. contortus, the reduction

of worm burdens in order to reduce pathology and/or a

reduction of pasture contamination. In contrast to

gastro-intestinal nematodes, a continuous natural

boosting of immunity by field exposure to Fasciola

spp. following vaccination is rather unlikely. Thus,

repeated courses of vaccination may be necessary.

Sheep infections as low as 54 flukes per animal were

shown to reduce weight gain by 8–9%, even though this

level of infection resulted in no clinical signs of disease

(Hope Cawdery et al., 1977). Under natural conditions,

significant weight loss is observed with fluke burdens

above 30 (Dargie, 1986) to 40 (Hope Cawdery et al.,

1977). Therefore, Spithill et al. (1999), suggested that a

vaccine with mean efficacy (reduction of fluke burden)

as low as 43% would still reverse production losses in

animals infected with as few as 53–140 flukes, which is

the normal range of the fluke burdens in sheep reported

from endemic areas in different countries.

To achieve a reduction of pasture contamination by

vaccination, immunized animals should excrete as few

eggs as possible, because of the high reproductive

capacity of Fasciola spp. To reduce transmission, it

was shown that only regular treatments at 12- to 13-

week intervals with flukicides effective against both

mature and immature flukes (i.e., efficacy >90%)

were able to reduce the intensity of infection in a flock,

or herd over time (Fawcett, 1990; Maes et al., 1993).

Therefore, when a reduction of parasite transmission

is the objective of a Fasciola spp. vaccine, animals

need to be monitored for several months for reductions

in faecal egg counts and/or fertility of the eggs.

However, since the level and the duration of the

reduction of viable egg excretion required to reduce

parasite transmission to such a degree that production

losses are minimized are uncertain, it remains difficult

to design Fasciola spp. vaccine experiments and to set

criteria for the efficacy assessments (Vercruysse and

Claerebout, 2003).

4.4.2. Economic benefits of vaccines

Since the status of development of many of these

vaccines is unknown and/or publicly available data are

lacking, vaccines (with the exception of D. viviparus)

as novel control methods could not be fully assessed

using the criteria outlined in Section 3 of this paper.

However, it is clear that the models to be used to

evaluate the efficacy in laboratory and field settings

will differ according to the parasite and type of

vaccine. In addition, the efficacy levels required for a

significant economic impact also will differ for each of

the important parasites of livestock.

5. Conclusions

The efficacy required for novel control methods

depends not only on the parasite species and

J.K. Ketzis et al. / Veterinary Parasitology 139 (2006) 321–335332

epidemiological circumstances but also on other

measurements of efficacy. Parasitologists, epidemiol-

ogists, clinicians, nutritionists, economists, and live-

stock producers may all have different views about the

merits of a particular control program and its

requirements. The ultimate milestone of success of

a novel control method is its availability to livestock

producers and the ability of livestock producers to

adopt, adapt, and implement the new program.

In order to facilitate the commercialization and

enhance the acceptance of novel control methods, a

standard framework to measure and evaluate their

efficacy is required. It can be concluded from this

review that novel approaches are difficult, if not

impossible, to evaluate on the basis of a set of uniform

efficacy criteria. However, we have proposed and

tested some general efficacy criteria referred to as

minimal performance expectations. First, novel con-

trol methods must exhibit a significant level of efficacy

when used alone in controlled laboratory studies.

Second, the efficacy of novel control methods needs to

be confirmed in different environmental settings on a

large scale (e.g., on-farm). Third, the efficacy achieved

by the use of a novel control method should result in a

reliable economic benefit (on a herd/flock basis),

which needs to be interpreted within the context of a

particular a management system.

While these minimal performance expectations are

not as strict or as easy to evaluate as the established

requirements for anthelmintics, they do provide a

valuable framework for evaluating novel control

methods. While laboratory and on-farm studies provide

evidence for the effectiveness of a novel control

method, the inclusion of an economic factor as part of

the minimal performance expectations (when a control

method is used in a defined management system),

overcomes the difficulty of setting a minimum efficacy

threshold such as those adopted for anthelmintics. This

new approach may, however, have inherent geographic

uncertainties as economically viable options cannot

easily be extrapolated across different sites, countries,

and/or livestock management systems. Nevertheless,

the proposed efficacy expectations could be used to

evaluate very different novel control methods. Further

assessment of these proposed minimal performance

expectations, their applicability to other novel control

methods, and the practicality of adapting them as

guidelines are needed. However, this first attempt to

define efficacy guidelines for novel approaches

demonstrates the feasibility of developing criteria for

diverse control methods.

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