Onderzoeksvoorstel master Diergeneeskunde
Onderzoeksplan te verdelen in:
1. Background
2. Aim of the study
3. Materials and methods
4. Schedule
5. Literature
Personalia
Studentnummer: 3515389
Naam: Lisanne Francien Hageman
Curriculum: Master (LH/VV)
Telefoonnummer: 0613945730
E-mailadres: l.f.hageman@students.uu.nl
Onderwerp en plaats van uitvoering
Titel van onderzoek: ‘Serum and faecal Zinc concentrations following low dose oral
ZnO administration in sheep’
Geplande aanvangsdatum: Februari 2014
Departement: Landbouwhuisdieren
Begeleider op de faculteit: Henk Everts
Begeleiders externe stage: Stefan Smith en Chris Anderson, Massey University, Palmerston
North, New-Zealand
E-mailadres begeleider(s): S.L.Smith@massey.ac.nz , C.W.N.Anderson@massey.ac.nz
Background of the study
Facial Eczema is a fungal disease of ruminants in New Zealand caused by ingestion of spores.
These spores are produced by the saprophytic fungus Pithomyces Chartarum. P. Chartarum
contains a mycotoxin called sporidesmin, ingestion of this mycotoxin causes liver damage.
Clinical signs become visible when bile ducts are blocked. This leads to secondary
photosensitization, mainly visible in the face of the animal. (1) The disease is not fatal, but is
causes discomfort that results in less feed intake and decreased production (3) More often
there are no signs at all, but high activities of liver enzymes, such as high gamma-glutamyl
transpeptidase (GGT) in the blood are indicative of liver damage. GGT is a biochemical
indicator of FE and is elevates in animals with chronic or acute exposure to the disease. P.
Chartarum severely affects animal production and has animal welfare implication.
Prevention of Facial Eczema (FE) disease in livestock is currently achieved by regular dosing
of Zn salt (oxide and sulphates). The exact mechanism of Zn is still unknown. It is thought
that is prevents auto-oxidation of the reduced dithiol form op sporidesmin, so it prevents the
release of hepatotoxic free radicals. (1), (2)
Zinc is a heavy metal that is found in high concentrations in animal effluent, especially pig
effluent which, when sprayed onto soil can build up into very high levels. Plants such as
willow can selectively absorb Zn from these contaminated soils helping to de-contaminate the
soil but also these plants may then be a source of dietary Zn for prevention of Facial eczema.
We know that dietary Zn from plants does not elevate serum levels but does elevate faecal Zn
levels.
Current Zn prevention programmes involve the oral dosing of high doses of Zn Oxide which
elevates both the serum and the faecal levels of Zn. Using Zinc dosing in the form of long
acting Zn oral boluses or oral drenching with high doses (25mg/kg live weight) of ZnO. The
mechanism for Zn providing FE protection is unknown and it either comes via systemic
serum protection at the liver or is a gut protection effect at the level of the intestine epithelial
cell or both - We know that animals with 250mgZn / kg fecal weight or 30umol / l serum Zn
are protected from Facial Eczema but we do not know which one provides protection.(3)
Aim of the study
The aim of the study will be to determine an oral dose rate of Zn Oxide that only elevates the
fecal Zn concentration to 250mg/kg fresh weight without elevating serum Zn concentrations.
Once we have determined this oral dose rate then we can perform a challenge study to
determine whether faecal zinc levels provide the mechanism of action for preventing Facial
Eczema.
In 2013, a pilot study was undertaken to determine the efficacy of dosing sheep with either
intravenous ZnCl or oral ZnO
The results of this study include;
a) Intravenous ZnCl (5.1mg Zn) can elevate serum Zn levels to above 30umol / l although
these levels are only maintained for 2-3 hours post dosing.
Conclusion: ZnCl is unlikely to be used successfully to maintain serum Zn levels for future
sporidesmin intervention studies.
b) A single oral dose of ZnO (10mg / kg liveweight) successfully elevated the faecal Zn
concentration to > 250mg / kg fresh weight (FW) and that this level of faecal Zn lasted for 24
hours. This dose rate of oral ZnO also elevated the serum Zn concentration to 30umol / l for
up to 22 hours post dosing.
Conclusion: Oral dosing sheep with 10mg/kg ZnO provided adequate Zn for both protective
mechanisms for 24 hours and repeat dosing at this dose rate every 24 hours should provide
therapeutic protection against facial eczema. However, this result does not allow for the
differentiation of the protective mode of action for Zn against sporidesmin toxin as both
faecal Zinc and serum Zn levels were elevated to levels considered to be in the preventative
range using an oral dose rate of 10mg/kg ZnO.
Further research is now required, as another pilot study, to determine whether a lower oral
dose of ZnO (5mg/kg liveweight) can be administered to maintain serum Zn below and
elevate faecal Zn concentrations above the therapeutic range for the prevention of facial
eczema in sheep, to determine the duration of these therapeutic levels, and whether changing
the dosing interval using this lowered ZnO dose rate has an effect on this therapeutic duration.
This pilot study will determine whether the faecal concentration of Zn alone can be
manipulated using low oral dose rates of ZnO and will enable a dose rate and administration
protocol to be developed to achieve faecal Zn concentrations of >250mg/kg FW in sheep.
Once this is established the faecal protective mode of action of Zn against sporidesmin
toxicity can be tested using intervention studies.
Should the Zn faecal protection mode of action against sporidesmin be confirmed, then an
entire research field will be developed regarding the use of plant bio-fortification and the use
of plants, such as willow, as a supplementary feed source with therapeutic benefits.
This data will underpin our ongoing research into the potential of bio-fortification to improve
animal health in New Zealand agriculture.
If we can show this then the use of Zn fortified plants as a means to provide dietary protection
then becomes a real possibility. (4)
Material & methods
Six hoggets (approx. 50 kg liveweight) will be ear tagged, randomly selected, divided into 2
groups of 3 animals and run together on pasture at VLATU, Old West Road, Massey
University.
Group 1 – Orally dosed with 5mg / kg ZnO at 24 hour intervals for 2 doses.
Group 2 – Orally dosed with 5mg/kg ZnO at 12 hour intervals for 4 doses.
All animals will be blood and faecal sampled at
T=0, T=4, 8, 12, 24, 28, 32, 48, 52, 56 and 72 hours.
(The initial pilot study showed oral dosing to have no effect on serum or faecal levels until 4
hours post dosing eliminating the need in this study to test in the earlier stages. Additionally,
the duration of sampling is also reduced to 72 hours from the initial pilot study results.)
Animals will be kept on pasture for the duration of the study and penned in the same paddock
for each sampling session.
ZnO will be mixed with a suspending agent and administered using a 10ml syringe.
All sheep will be clipped for blood sampling with all blood samples collected from the jugular
vein using a 20G x 1” vacutainer needle into a 10ml plain vacutainer tube for serum analysis.
Data from this pilot study will enable an oral ZnO dose rate to be established that will achieve
a faecal Zn concentration of >250mg / kg fresh weight without elevating the serum Zn levels.
Schedule
Period of 3 months, starts in February.
Literature
1. JJ Bennison, RM Nottingham, EL key, JJ Parks, 2010 The effect of zinc oxide and
elemental zinc boluses on the concentrations of Zn in serum and faeces, and on
proving protection from natural Pithomyces chartarum challenge sheep. New Zealand
Veterinary Journal 58:4, 201-206
2. Christopher W.N. Anderson, Brett Robinson, David M. West, Lynne Clucas, Denise
Portmann, 2012 Zinc-enriched and zinc-biofortified feed as a possible animal remedy
in pastoral agriculture: Animal health and environmental benefits. Elsevier 30-35
3. CA morris, NR Towers, WD Hohenboken, N Maqbool, BL Smith, SH Phua (2004),
Inheritance of resistance findings from sheep and cattle in New Zealand. New Zealdn
Veterinary Journal 52:2, 205-2.15
4. Massey University Study of 2011 by Chris Anderson (Institute of Agirculture and
Environment)