Overcoming the Constraints of Emu Egg Laying Season Coordinated Emu Research Program

A report for the Rural Industries Research and Development Corporation by Graeme Martin University of Western Australia

March, 1999 RIRDC Publication No 99/23 RIRDC Project No UWA-25A

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© 1999 Rural Industries Research and Development Corporation. All rights reserved. ISBN 0 642 57828 1 ISSN 1440-6845 Overcoming the Constraints of the Egg Laying Season - Coordinated Emu Research Program Publication No. 99/23 Project No. UWA-25A The views expressed and the conclusions reached in this publication are those of the author and not necessarily those of persons consulted. RIRDC shall not be responsible in any way whatsoever to any person who relies in whole or in part on the contents of this report. This publication is copyright. However, RIRDC encourages wide dissemination of its research, providing the Corporation is clearly acknowledged. For any other enquiries concerning reproduction, contact the Publications Manager on phone 02 6272 3186.

Researcher Contact Details Associate Professor Graeme Martin University of Western Australia Faculty of Agriculture NEDLANDS WA 6907

Phone: (08) 9380 2528 Fax: (08) 9380 1040 Email: gmartin@agric.uwa.edu.au

RIRDC Contact Details Rural Industries Research and Development Corporation Level 1, AMA House 42 Macquarie Street BARTON ACT 2600 PO Box 4776 KINGSTON ACT 2604 Phone: 02 6272 4539 Fax: 02 6272 5877 Email: rirdc@netinfo.com.au Website: http://www.rirdc.gov.au

Published in March, 1999 Printed on environmentally friendly paper by Canprint

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Foreword Much has to be learnt on farming emus efficiently in the production of meat, oil and leather. Increased productivity and profitability will partially rely on the introduction of innovation in management and husbandry. Reproduction systems which exist in the wild will need to be adopted and in part replaced by new technologies which fit into the farming environment. Increased knowledge of the biology of the emu is the foundation upon which to build new technologies for commercial application. The research findings described in this report provide a greater understanding of mechanisms that affect reproduction in the emu. The report also recommends a body condition scoring system which relates well to ultrasonic measurements - a technique which can be used by the farmer without having to restrain the emu. This report, a new addition to RIRDC’s diverse range of over 250 research publications, forms part of our New Animal Products R&D program which aims to facilitate the development of new industries based in animal or animal products that have commercial potential for Australia. This report, and others, can be viewed or purchased online at www.rirdc.gov.au Peter Core Managing Director Rural Industries Research and Development Corporation

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Contents

FOREWORD III

EXECUTIVE SUMMARY VI

1. Background 1 Objective 1 1 Objective 2 1

2. Findings 3 Objective 1 3 Objective 2a 5 Objective 2b 6 Objective 2c 10 Objective 2d 12

3. Future Research & Development 14

4. Publications 15 Scientific papers 15 Media articles 16

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Executive Summary This project was focussed on the major costs in emu production that are caused by the seasonal pattern of reproduction. These include restriction to the number of eggs produced per year and an inability to use of incubation facilities efficiently throughout the year. In addition, the aggressiveness and loss of appetite at the start of the breeding season, a period that precedes slaughter, cause skin and carcase damage and loss of oil production. We carried out an extensive studies on large farms to quantify the seasonal patterns of lay and hatching, the year-to-year variations in those patterns, and relationships between those patterns and the environmental factors that may control them. This was matched with a series of intensive experiments designed to reveal the physiological mechanisms involved and to develop systems for overcoming the problems. The project has successfully provided useful outcomes for industry and major scientific findings that might lead to future industrial applications. Importantly, because the RIRDC lent the project a solid foundation, we were also able to attract other funding, notably from the Australian Research Council, and other personnel, including a research fellow and three PhD students. The emu industry has thus benefitted from the input of an excellent and sizeable team of people. Major outcomes The laying season is very tightly restricted to the period between late autumn and early spring, as emu farmers will attest. We have proven beyond doubt that this is primarily due to changes in daylength. In contrast with mammals, where melatonin can be used to overcome the effects of daylength, science has not yet provided us a solution for birds so, in the short term at least, we can do little to avoid the laying season. For the long term, there are avenues for scientific exploration: first, by developing an understanding of the physiological process involved we might reveal ways of avoiding seasonality; second, we can use genetic selection to move and lengthen the season, as has been done for other domestic birds and mammals. At this stage, we have no idea of the degree of between-bird variability in laying seasons within our emu populations. The overall seasonal pattern is not yet avoidable, but we still have some scope for control. The start of the season and the length of the season (and thus the number of eggs laid) differs between locations and between years, but this is only partly due to the differences in daylength or latitude. Summer rainfall is involved too, with good summer rains advancing the start of lay. On the other hand, the total amount of rain (that which falls before the laying season plus that which fall during the laying season) was related to the length of the season and to egg production. More rain leads to a longer season and more eggs. The reason for these relationships is not known, although we have been able to dismiss nutritional factors. Is it due to the rain itself or to some other factor related to rainfall (such as barometric pressure)? These phenomena require further investigation before we will know whether or not we can exploit them. We targetted the behavioural problems associated with the clash between breeding seasons and abattoir seasons by testing whether they could be solved through castration, as is done for mammal-based animal industries. Surgical castration is very difficult in birds and not likely to pass ethical approval, so we decided to test hormonal methods based on vaccine technology.

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One method involved immunising birds against a key reproductive hormone produced by the brain, gonadotrophin-releasing hormone (GnRH). This approach has been successful for controlling fertility in rangeland cattle. A second method involved the use of a long-lasting subcutaneous implant which blocked GnRH action. This approach has been successful for controlling fertility in dogs and horses. We compared the growth and abattoir performance of normal birds and vaccinated birds with birds that were surgically castrated and, we thought, would set the ultimate goal. We had initially some problems with the vaccine technology because emus proved to be resistant to standard antigens, but finally developed a combination that was very effective. Surprisingly, castrated birds had no advantage over normal birds, in terms of growth, or meat or fat yields when they were slaughtered at two years of age. This suggests that there is little scope for development here, at least with the current practice of slaughtering birds when they are less than two years old. On the other hand, if that practice were to change so that older birds were slaughtered, there may be an advantage because both hormonally- and surgically-castrated birds showed better growth in their third year. Will the industry ever move to the slaughter of older birds? Under the current system, this is not likely because the extra 12 months feed yields too little extra product. However, if the industry were to move to low-input rangeland practices, based on slower, cheaper growth over a longer period, castration technology will be useful. This applies to both sexes. However, the equality ends there because, in non-castrated birds, females show major advantages over males in terms of yield at the abattoir. They produce 25% more fat. Under current practices, farmers raise equal numbers of both sexes, feeding them the same way, and slaughter them all at the same age. Consideration should be given to a) slaughter of males as juveniles, perhaps with the development of an emu equivalent of the

capretto goat-meat market; b) focussing on females for fat yield. A problem we faced in these studies, a problem we share with farmers, is reliable, quantitative assessment of the fatness of live birds. We needed this to measure the progress of our birds during experiments, and farmers need it to decide whether or not to birds can be sent to the abattoir. We therefore tested whether ultrasound or body condition scoring (BCS) would be useful. The results were very clear. Weighing birds of a similar (and known) age and similar up-bringing gives a very good indication of the relative amounts of fat and muscle that they will yield. Ultrasound also seems feasible, with further development. However, both of these methods require restraining the birds, to the point of keeping them still, and neither are likely to be practical on the farm. This contrasts with BCS, which can be readily learned and can be done ‘on the fly’, often without any restraint of the birds. It gives very good estimates of fat yield and also has the advantage of being independent of frame size (and hence age). Conclusion We see little reason why the industry should not adopt body condition scoring. The other findings, such as the potential advantages of hormonal castration, of the effect of rainfall on the laying season, may become useful in the future, as we test new and varied management practices. The results of our photoperiod studies are scientifically very valuable in that they challenge the dogma that surrounds the physiology of seasonal breeding. The peculiarities of emus may ultimately lead to solutions to the problems posed by the laying season.

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1. Background This Program involved work at the WA Department of Agriculture (now ‘Agriculture WA’) and The University of WA. It focussed primarily on problems caused by the seasonal pattern of reproductive activity, including reproductive hormonal problems that add major costs at many levels of the production system, including:

• incubation facilities that are under-used early in the season but are insufficient for the peak of the laying period, so egg incubation has to be delayed with serious losses in hatchability;

• the loss of appetite during the period leading up to slaughter, reducing fat and oil production; • the restriction of the number of eggs produced, increasing the production cost per egg; • the periods of aggressive behaviour, leading to skin and carcase damage.

We developed this project in association with our commercial partner, Dromaius Australia Ltd, who provided access to the facilities and personnel on their farm for collection of hatching data and for field trials, and also supplied experimental birds for intensive studies at the University. These intensive studies also required a new emu facility on campus, and this was built with RIRDC funds (for materials only) and skills and labour from Dromaius Australia and The University. Dromaius Australia also donated a microchip tagging system to the project and paid for a working visit from the UK by Professor PJ Sharp, who helped us with the surgery. We originally planned to pursue two major objectives: Objective 1

A long-term extensive study on a large commercial enterprise (1500 breeders; about 750 pairs) to describe the seasonal patterns of lay and hatching in detail and relate these patterns to environmental factors.

Objective 2 A series of intensive experiments to test the ideas gained on-farm and to develop systems for overcoming the problems. Experiments were planned to test whether:

a) The reproductive season is induced by short or long days; b) Gonadectomy will prevent the seasonal loss of appetite, body mass and fat; c) Immunisation against gonadotrophin releasing hormone (GnRH) or prolactin will block

the laying season, prevent the seasonal loss of appetite, body mass and fat, and reduce skin damage;

d) Immunisation against prolactin will influence the seasonal patterns in appetite and broodiness;

e) Isolation of the sexes will prevent or delay the onset of the breeding season. We have successfully completed most of these studies. Our original plans were, however, adjusted following various commercial and scientific ‘events’: First, we had more trouble than expected in designing an effective GnRH antigen for “immunocastration”, so we had to repeat this study and also test whether a GnRH superagonist might not be better. These factors also led us to drop some of the more esoteric studies that we had tentatively planned for the final year (eg, Objective 2e); Second, our commercial partner, Dromaius Australia, fell into financial difficulties during the second year of the project. This led us to curtail the on-farm observational work and compromised some of the data we were collecting. As shall be detailed below, the overall impact of this event on the project was minimal, through both serendipity and management. For example, in the final year

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of the project, we had planned to carry out field trials at the Mount Gibson farm for some of our technologies. This would have been impossible but was ultimately not necessary because the results of the ‘castration studies’ suggested that there would be little commercial gain from this approach. The on-campus studies were not affected because already had the birds and pens in place. Despite these setbacks, the project has been very successful and we can offer some solid advice to industry about future developments. The project also attracted other funding, notably from the Australian Research Council (Main Grants) who we asked to support some of the more basic investigations that were needed to explain the field observations. As a result of the funding and the on-campus emu facility, the project attracted an excellent team of people. Thus, in addition to the personnel formally linked with this project (Martin, Frapple, O’Malley), RIRDC and the emu industry have benefitted greatly due to inputs from: • Dr Dominique Blache, research fellow — reproductive behaviour and endocrinology, feeding

behaviour and metabolic physiology, co-supervisor of the UWA emu team; • Ms Margaret Blackberry, senior technician — specialist in hormone techniques, including

radioimmunoassay development; • Ms Karen Williams, PhD student — a veterinarian who joined the program early on and carried

out most of the RIRDC-funded research; • Mr Irek Malecki, PhD student — bird expert who was working on reproductive technology for

the emu industry and also took part in the studies of emu seasonality described below. • Ms Judy van Cleeff, PhD student — a reproductive physiologist who joined the team late and is

progressing with the studies of metabolic and reproductive endocrinology; • Mr Ryan Mincham, Honours student — established a method for measuring fat content of emus

before slaughter.

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2. Findings Objective 1 The long-term, extensive study on the large commercial enterprise provided data for only two years during the project, however we persuaded Dromaius to allow us to use their records for our analysis. These beautiful data clearly describe the laying season (Fig. 1).

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Figure 1 Egg production by 1500 breeders during 3 consecutive laying seasons at Mount Gibson (30°S). The seasonal pattern is evident, with the lay beginning in April-May and ending in August-September. Our main hypothesis was that this pattern is controlled primarily by changes in daylength, and we subsequently investigated this (see below). However, there is clearly a great degree of variation between the seasons in the precise timing of onset of lay, in the shape of the laying pattern, and in the duration of the season. To investigate this further, we also used data from the Agriculture WA research station at Medina. The egg laying data and rainfall records from the two emu farms were compared. The two locations differ significantly: (A) Mount Gibson, 29° 45' South, 350 km north east of Perth, is in a semi-arid area subject to summer cyclones, with 1500 breeding birds recorded from 1992 to 1996; (B) The Medina Research Station, 32° South, 40 km south of Perth, with a Mediterranean environment, where 70 breeding birds were observed in 1988 and 1989, and 140 breeding birds were observed from 1990 to 1996. The factors examined were Length of Laying Season (days), the Delay to Start of Lay (number of days after the Autumn Equinox, March 22), and Total Eggs Laid. These were analysed by linear regression against the Rainfall Before the Laying Season, and Total Rainfall (before plus during the laying season).

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Figure 2 Relationships between rainfall pattern and the egg-laying season in emus at Medina (32°S: a,b) and Mount

Gibson (30°S: c,d). These studies suggest that the emu responds to rainfall (Fig. 2). There was a significant positive correlation between Length of Laying Season and Total Rainfall at Medina (r2 = 0.42; p = 0.024; Fig 1a) which was markedly improved when it was fitted to a binomial curve (r2 = 0.76; Fig 1a). At Mount Gibson, the correlation between Total Rainfall and Length of Laying Season was also significant (r2 = 0.91; p = 0.044; Fig 1c). There was a significant correlation between the Total Rainfall and the Total Eggs Laid at Medina (r2 = 0.75; p = 0.002; Fig 1b) but not at Mount Gibson. However, there was a negative relationship between Rain Before the Season and the Delay to Start of Lay after the Autumn Equinox (March 22) at Mount Gibson (p = 0.092; Fig 1d). This was not the case at Medina (data not shown). It is important to realise that these comparisons are within site (ie, at the same latitude and thus photoperiod) and that all birds were fed and watered ad libitum, so that variation in these external factors are not involved. Nevertheless, it seems likely that rainfall advances the onset of egg-laying at Mount Gibson, and extends the laying season (thus leading to production of more eggs) at both sites. The two locations differ fundamentally in terms of rainfall pattern because Mount Gibson is in a semi-arid area with a great deal of between-year variation in pre-season rainfall (Fig. 2d). In contrast, Medina is in a mediterranean region with a very little variation, making it difficult to test the correlation between rainfall and time of onset of season.

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Objective 2a The hypothesis was that the onset of the reproductive season is induced by changes in photoperiod. Here, we carried out two experiments, one with juvenile chicks in the period leading up to puberty, and one with adults. We used male birds because the sex steroid patterns are not complicated by ovulatory cycles. Experiment 1 The emu has a deferred puberty, suggesting that the juvenile is not responsive to changes in photoperiod. Emu chicks were raised on a farm from hatching until 14 weeks of age, (i.e. under increasing daylength) and then penned in groups in photoperiod-control rooms in January (mid-Summer). They were subjected to 10 h light (SD; n = 8) or 14 h of light per day (LD; n = 5), or natural photoperiod (ND; n = 7). Blood was sampled weekly up to 66 weeks of age. On Weeks 50, 52, 58, 62 and 66, phallic enlargement was assessed by cloacal palpation. Food and water were provided ad libitum and the total feed intake of each group was measured weekly. Light treatments did not affect feed intake, growth rate, or plasma testosterone. In all LD birds, the phallus became palpable by age 52 weeks. In SD and ND birds, this was delayed until 62-66 weeks of age (Table 1). Table 1 Effect of photoperiod (over 14 to 66 weeks of age) on mean (± sem) plasma concentrations of prolactin, LH

and testosterone, and identifiable phallus, in young male emus. Different superscripts within a row P < 0.05.

Natural light Short day Long day Testosterone (pg/ml) 90 ± 20 80 ± 10 80 ± 30 % of bird with identifiable phallus

50 weeks 0 0 40 52 weeks 29a 25a 100b 58 weeks 57a 63ab 100b 62 weeks 71 63 100 66 weeks 100 88 100

Thus, photoperiod affected phallic development in the absence of changes in the concentrations of testosterone. This suggests that our endocrine techniques are not sufficiently sensitive, or that we need to measure other gonadal hormones in future studies. Normally, the breeding season (in adults) begins under short days, so the most striking observation here was the accelerated development of the phallus under long days, a photoperiod that we would have expected to be inhibitory, and the complete lack of effect of short days, a photoperiod that we would have expected to be stimulatory. Clearly, to understand puberty in emus, we need to develop better hormone techniques and completely re-think the photoperiodic mechanisms involved. These birds do not follow the rules. Experiment 2 Most birds living at non-equatorial latitudes are long day breeders, but the emu breeds during autumn and winter. The timing of breeding in the emu may be influenced by a range of environmental factors including food availability, rainfall and social and behavioural cues, but seasonal change in photoperiod is probably the major cue, as has been demonstrated for long-day breeding birds. At the beginning of February, 8 mature males (120 weeks old) were moved from external enclosures into individual pens in two photoperiod-control rooms (4 birds per room). They were

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subjected to 10 hours light per day (short days, SD) or 14 hours light per day (long day, LD) for 21 weeks then the photoperiods were reversed for another period of 21 weeks. Food and water were provided ad libitum. Birds were weighed, blood sampled, and sexual behaviour was assessed.

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Figure 3 Feed intake (a) and plasma concentrations of testosterone (b) in mature male emus subjected to long or short

days. Closed circles: birds on SD then LD. Open circles: birds on LD then SD. *P<0.05. During the first 6 weeks, the birds were probably stressed by the change from outdoors to a confined environment, so feed intake was low in both groups, but SD birds had the lowest (Fig. 3a). This was significant during the last 7 weeks of Period 1 and then again, but in the reverse direction, when the photoperiod was reversed. Plasma testosterone increased over time in birds exposed to SD during both treatment periods, while birds exposed to LD had low levels (Fig. 3b). More birds expressed sexual behaviour on SD than on LD (7/8 vs 2/8; P < 0.05). Photoperiod clearly is the major time-keeper for the annual changes in testosterone production, sexual behaviour and appetite. Rainfall appears to be able to modulate the response to photoperiod, but we have no idea of the physiological mechanisms involved. We have not yet tested whether other factors (eg, nutrition, cues from opposite sex) also modulate the annual pattern. Objective 2b The commercial importance of body fat means that techniques which allow fat content to be manipulated or assessed in the months leading up to slaughter are likely to be key management tools. Here, we tested two approaches for assessment of the body fat content of live birds using the

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principles of ultrasound and body condition scoring (BCS), and then tested whether castration will prevent seasonal losses of body weight and body fat, and improve carcase yields.

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Experiment 1 Sixteen emus were assessed for fat content at an abattoir. Before slaughter, they were condition-scored by two assessors using a 5-point system (1 = thin to 5 = fat) similar to that used for sheep. The preacetabular ilium was used as a marker for condition scoring (Fig. 4) and birds were scored to the nearest full point.

The birds were given a score based on the build up of muscles and fat around the backbone: 1 Bones prominent and sharp; 2 Bones prominent but smooth; 3 Bones can be felt but are smooth and rounded; 4 Bones are detectable with pressure on the thumb; 5 Bones are not detectable or detectable only with firm

pressure.

Condition score region

Preacetabular ilium

Postacetabular ilium

Figure 4 Position used for condition scoring in live birds.

Ultrasound measurements We used an “A-mode Sonatest” (Meritronics Inc.) with vegetable oil as a contact medium between the probe and the skin. Any remnant feather shafts were removed if they restricted probe-skin contact. Five marker sites were chosen for ultrasound measurements, based on the largest subcutaneous and abdominal fat depots observed on emu carcases (Fig. 5).

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Figure 5 Points used for ultrasound measurement of fat depth on the emu carcase (Left and Centre: A1, A2, B, C1,

C2). Right: a cross-section along the C2 line (at right angles to the midline) showing the partitioning of subcutaneous (sc) fat, muscle and abdominal fat.

A1 halfway between C1 and C2, 6 cm left of mid-line, 25 cm anterior to tip of tail; A2 halfway between C1 and C2, 6 cm right of mid-line, 25 cm anterior to tip of tail; B 6 cm anterior of vent on the underside of the bird, on the midline; C1 20 cm anterior to tip of tail along mid-line; C2 30 cm anterior to tip of tail along mid-line, directly across from join of legs to body. After slaughter, ultrasound measurements of subcutaneous fat depth were taken from five pre-determined marker sites on the dorsal and ventral sides of the on the defeathered whole carcase. Ultrasound and BCS values were then related to body weight and fat yield. Fat weight was highly correlated with live weight and with body condition score (Table 2). The ultrasound measurements of fat depth at points B, C1 and C2 were also correlated with fat weight, but they were poor predictors compared to body condition score.

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Table 2 Linear regression data for the significant relationships between fat weight (kg) and live weight (kg), body

condition score (BCS) and ultrasound measurements (mm) in emus.

Compared variables R2 r F P Intercept Slope s.e.m.Fat weight Live weight 0.76 0.87 45.01 <0.001 -8.79 0.38 0.06BCS 0.67 0.82 28.90 <0.001 0.55 1.55 0.29Ultrasound measurements A1 0.12 0.34 1.87 NS - - - A2 0.04 0.19 0.53 NS - - - C1 0.48 0.70 13.13 <0.01 1.56 0.12 0.03C2 0.63 0.80 24.1 <0.001 0.84 0.19 0.04B 0.50 0.70 14.1 <0.01 1.26 0.12 0.03Live weight BCS 0.80 0.90 53.76 <0.001 25.47 3.92 0.53Ultrasound measurements A1 0.04 0.19 0.53 NS - - - A2 0.08 0.30 1.35 NS - - - C1 0.63 0.80 23.8 <0.001 27.6 0.31 0.06C2 0.51 0.71 14.3 <0.01 27.9 0.40 0.11B 0.19 0.44 3.33 NS - - - BCS Ultrasound measurements A1 0.07 0.26 1.03 NS - - - A2 0 0.05 0.04 NS - - - C1 0.57 0.75 18.3 <0.001 0.87 0.07 0.02C2 0.4 0.64 9.51 <0.01 1.02 0.08 0.03B 0.15 0.38 2.43 NS - - -

Live weight was also highly related with body condition score and with measurements of fat depth at points C1 and C2, whereas values at A1, A2 and B were not related to live weight. The body condition scoring (BCS) was only correlated with the fat depth at points C1 and C2. The prediction of the live weight and fat weight appears to be related to the greater fat depth at points C1, C2 and B (Table 3).

Table 3 Estimated fat depth by ultrasound measurement at various points on the emu carcase.

Point of measurement A1 A2 C1 C2 B Mean ± s.e.m. (mm) 16.8 ± 1.5 16.3 ± 1.8 26.1 ± 3.0 19.8 ± 2.1 28.0 ± 3.0

CV (%) 35.7 45.3 46.6 43.2 43.3 We concluded that both BCS and ultrasound can be used successfully to predict fat content and body composition in emus. At this stage, BCS appears to be more reliable than ultrasound. With some further development, these techniques should benefit emu management, particularly the areas of emu nutrition, pre-slaughter finishing and meat marketing. Experiment 2 Emus do not reach slaughter size until at least twelve months of age and, in some circumstances, are grown until they become sexually mature. This is associated with aggressive behaviour, which

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causes damage to the skins, and a loss of appetite which causes a loss of fat during the Winter breeding season. In other animal production systems, these behavioural problems are usually managed by sterilisation which also leads to deposition of extra fat. We therefore compared the body composition of surgically sterilised male and female emus. In November 1996, 17 female and 22 male emus were slaughtered. Of these, six females and seven males had been surgically sterilised at eight weeks of age, leaving eleven females and fifteen males as sham-operated, gonad-intact controls. At slaughter, birds were two years and two months old and had been raised by Dromaius Australia Ltd under standard commercial practises on their farm at Mount Gibson, WA. We recorded live weight, counted skin lesions, weighed the fat (subcutaneous plus abdominal), the carcase (skeleton plus muscles) and total meat (Table 4). Table 4 Comparison of Body Composition of Male and Female Emus.

Sex Number Live Wt (Kg) Carcase Wt (Kg) Fat Wt (Kg) Meat Wt (Kg) Female controls 11 44.68 21.35 8.96 13.90 Female surgery 6 46.00 21.48 10.60 14.18 s.e.d. 1.191 1.073 1.022 0.576 Significance 0.50 0.899 0.131 0.624 Male controls 15 39.97 20.07 7.55 13.38 Male surgery 7 39.64 19.71 7.80 13.23 s.e.d. 1.885 0.882 0.951 0.561 Significance 0.865 0.688 0.797 0.794 All females 17 45.15 21.39 9.54 14.00 All males 22 39.86 19.96 7.63 13.33 s.e.d 1.282 0.649 0.662 0.383 Significance < 0.001 0.034 0.007 0.090

s.e.d.: standard error of difference between means Surgical sterilisation had no effect on any aspect of yield that we measured. The data were therefore pooled to allow comparison between the sexes. The females were heavier, had larger carcases and more fat than the males. The industrial implications of this study are fairly clear. There is little advantage in pursuing castration with a view to incorporating it into current farming practices. On the other hand, as with the studies of season, the emu has provided us with some startling scientific observations. Clearly, the classical view of the anabolic effects of sex steroids on muscle and fat deposition does not hold for these birds. The males are smaller than the females, and removal of ovaries or testes does not affect this, or the carcase characteristics. Objective 2c Hypothesis: Immunisation against avian gonadotrophin releasing hormone (aGnRH-I) will block the laying season, prevent the seasonal loss of appetite, body weight and body fat, and reduce skin damage. This hypothesis was based on studies in mammals, although the final outcome for the industry would clearly depend on the experiment with surgical castration. These studies were all done in parallel, so we did not to know the outcome of the surgery study which, as we have seen above, does not suggest a useful outcome here. We did three experiments, two using GnRH antigens and one using a subcutaneous implant of the GnRH superagonist, “Deslorelin”. In dogs, this substance over-stimulates the pituitary gland and completely switches off the reproductive system. Experiment 1

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Using birds that hatched in September 1994, we allocated 16 for vaccination against mammalian GnRH, 16 against avian GnRH and 16 for sham vaccination (controls). The GnRH was conjugated to bovine serum albumen. Vaccines were administered at 4 and 12 weeks, 6 and 11 months. Blood samples taken at 4 week intervals for GnRH antibody testing and hormone profiles. Body weight taken monthly from 4 weeks. Up to December 1995, there were no detectable effects on growth and no antibodies were detected in the blood. The birds were the returned to Mount Gibson emu farm. Experiment 2 Birds hatched in September 1995 were surgically sexed in December 1995. The males were then vaccinated against GnRH (n = 10) or sham vaccinated (n = 10) in March and December 1996, and again in March 1997. In this case, the GnRH was conjugated to keyhole limpet haemocyanin, a far more antigenic protein and it the produced readily detectable antibodies (Fig. 6). The experiment continued until mid-1997. Blood was sampled every 2 weeks and the birds were weighed monthly.

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The growth of the birds was not affected for the first 12-13 months, but thereafter the immunised birds showed an advantage over the controls (Fig. 7). These data are the first indication that “castration” can affect body growth in emus. Experiment 3 Birds hatched in September 1995 were surgically sexed in December 1995. In March 1996, 15 males were given GnRH agonist implants and 10 males were given sham implants. The birds were given a second implant in March 1997. There was no effect of the treatment until the birds were about 50 weeks old, after which the implanted birds showed superior growth. The re-implantation at 78 weeks led to a depression in growth probably because, on insertion, the GnRH superagonist caused a “testosterone flare” which exacerbated the seasonal depression in appetite. This is a recognised disadvantage of this approach.

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Figure 7 Live weights in untreated male emus, and male emus treated with GnRH superagonist implants

(“Deslorelin”) or the antigen, avian GnRH-KLH. When we were finally able to achieve hormonal castration and observe it over the long term, advantages for castration began to appear. GnRH immunity and Deslorelin both shows some promise, although the timing of Deslorelin implantation needs further investigation. Interestingly, if we can show that testosterone “flare-up” does indeed explain the depression of appetite after re-implantation, then this is even more evidence implicating sex steroids in the seasonal depression of appetite. Objective 2d Hypothesis: immunisation against prolactin will influence the seasonal patterns in appetite and broodiness. The original plan here was to clone emu prolactin, produce it through recombinant technology, and use it to make an antigen with which birds might be immunised. The process of cloning began when Ms K Williams took some mRNA isolated from emu pituitary glands to Prof Peter Sharp’s laboratory in Edinburgh. She spent 9 weeks there. Subsequently, the work was carried on by Prof Sharp’s team and they have recently completed the genetic sequencing. The rate of progress was too slow for us to complete the experiment with recombinant hormone, so we turned to chicken prolactin. Our work with the chicken prolactin radio-immunoassay in Perth, showing that it could measure the emu hormone, suggested that there is sufficient similarity between the chicken and emu hormones for this approach to be feasible. Accordingly, we immunised a group of male emus against chicken prolactin and observed the effects on broodiness. Some of our other studies had also suggested that testosterone might block the expression of broody behaviour, so we included a group of birds that were given subcutaneous testosterone implants. We have yet to complete all of the endocrine analyses (eg, concentrations of prolactin antibodies and testosterone), but it is clear that neither treatment exerted a significant effect on the onset of broody behaviour (Fig. 8).

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Birds sitting

(%)TestosteroneShamAnti-Prolactin

Figure 8 Effect of testosterone implants (n = 8), immunisation against chicken prolactin (n = 6 ) and sham

immunisation (n = 6) on onset of broodiness in male emus.

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3. Future Research & Development

“Hormonal castration” It is clear from both the surgical and hormonal castration studies, that there will be little benefit until the birds are entering their second breeding season when they are about 20 months old. Under current farming systems, meat birds have generally been slaughtered by this time. The technology would only become useful if farmers were to move away from the intensive system focussed on maximum bird growth. Dromaius Australia were beginning to explore this concept in 1995-6 in an effort to reduce feed costs. The plan was to retain intensive practices from laying until the chicks were about 4 months old, then place them into very large “foraging paddocks”, allowing them to grow more slowly under rangeland conditions, with supplementary feeding when needed. The birds would be fed intensively over the final months before slaughter, now delayed until about 20 months. Another reason for pursuing this line of enquiry, especially in association with studies of the physiology of puberty, is the particularly attractive possibility that a successful immunity to GnRH very early in life will cause permanent “castration” (as has been shown for sheep). With a workable antigen at our disposal, we are in a position to test this, and thus to test whether there is any possibility of preventing the partial puberty that birds undergo in their first breeding season. Seasonal breeding The role of rainfall clearly deserves some attention. At the risk of seeming excessively simplistic, we suggest that it will be worth testing whether artificial rain over the breeding pens during the dry periods can be used to advance and extend the breeding season. Also, the phenomenal appetite changes that are induced by changes in daylength must be investigated. If the hormonal mechanisms involved can be unravelled, then we should be able to short-circuit them and avoid the problem. The seasonal aspects of puberty are also an area of great potential. Emus show delayed puberty, itself an unusual phenomenon in birds. There are two potential avenues here, both of which depend on an understanding of the delaying mechanism. Advancing puberty in breeder birds would be advantageous, as they are very near full adult body weight in their first breeding season and thus should be able to cope with full reproductive competence. On the other hand, delaying puberty, even by 6 months, should help avoid some of the problems associated with reproductive activity that we have been attempting to overcome through castration. The ultimate solution to seasonal breeding is a long-term goal and this can only be achieved through a better understanding of the physiological processes involved. In itself, this is proving to be a major scientific conundrum. Interestingly, the emu may help to solve that conundrum because it does not follow the rules. In addition, we also need to survey the genetic diversity of the Australian emu population, with a focus on finding birds that can either lay all year round, or at least outside the normal seasonal boundaries. Selection away from strict seasonality has been shown to be very effective in sheep populations. Taking this concept further, it is clear that, in the very long term, Australia’s major advantage in any global emu industry will be found in the genetic diversity of our birds. We cannot hope to make use of that diversity if we have not quantified it.

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4. Publications This is the total output to date. Most of the work is in conference proceedings at this stage because we have yet to complete many of the laboratory measurements, and because Malecki and Williams are concentrating on the completion of their doctoral theses.

Scientific papers Blache, D., Malecki, I.A., Williams, K.M., Sharp, P.J. & Martin, G.B. (1997). Reproductive

responses of juvenile and adult emus (Dromaius novaehollandiae) to artificial photoperiod. In: Current Advances in Comparative Endocrinology — Proceedings of the 13th International Congress of Comparative Endocrinology, Japan [Monduzzi Editore: Bologna, Italy]. (in press).

Blache, D., Malecki, I.A., Williams, K.M., Van Cleeff, J.K., Sharp, P.J. & Martin, G.B. (1997).

Seasonal behaviour, reproduction and endocrinology in the male Emu (Dromaius novaehollandiae). Proceedings of the Australian Society for Reproductive Biology 29, 1.

Malecki, I.A. & Martin, G.B. (1996). Photoperiod and puberty in the male emu, Dromaius

novaehollandiae. Poultry and Avian Biology Reviews 6, 331 (abstract 13.21). Malecki, I.A., Martin, G.B., O’Malley, P.J, Meyer, G.T., Talbot, R.T. & Sharp, P.J. (1997).

Endocrine and testicular changes in a short-day seasonally breeding bird, the male emu (Dromaius novaehollandiae), in south-western Australia. Animal Reproduction Science (in press)

Malecki, I.A., Williams, K.M., Martin, G.B. & Sharp, P.J. (1997). Effects of season and incubation

on reproduction in the male emu (Dromaius novaehollandiae). In: Current Advances in Comparative Endocrinology — Proceedings of the 13th International Congress of Comparative Endocrinology, Japan [Monduzzi Editore: Bologna, Italy]. (in press).

Martin, G.B. (1994). A newcomer's view of the emu industry. In: "Emu farming in Western

Australia — Proceedings of a Seminar". Eds.: G. Mata, G.B. Martin, M. Sanders and N. Chamberlain. [Australian Society of Animal Production; Perth).

Martin, G.B., Malecki, I. & Williams, K.M. (1994). Reproductive technology for the emu industry.

In: "Emu farming in Western Australia — Proceedings of a Seminar". Eds.: G. Mata, G.B. Martin, M. Sanders and N. Chamberlain. [Australian Society of Animal Production; Perth).

Martin, G.B., Tan, N.S., Malecki, I., O’Malley, P. & Sharp, P.J. (1993). Season, broodiness and

testosterone secretion in the male emu. Abstracts of The XII International Congress of Comparative Endocrinology, Toronto, Ontario, Canada (May 16-21, 1993). A-99 (abstract).

Mincham, R., Malecki, I.A., Williams, K.M., Blache, D., Williams, I.H. & Martin, G.B. (1998).

Assessment of fat content and body composition in the emu (Dromaius novaehollandiae). Proceedings of the Australian Society of Animal Production (in press)

Sharp, P.J., Talbot, R.T., O'Malley, P., Tan, N.S., Williams, K.M., Blackberry, M.A. & Martin,

G.B. (1996). Neuroendocrine control of incubation behaviour in the emu. In: Improving our understanding of ratites in a farming environment (Ed.: D.C. Deeming) pp. 162-164 [Ratite Conference; Oxford Print Centre].

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Williams, K.M., Martin, G.B. & Potter, E. (1996). The influence of rainfall on the timing of the breeding season of the emu, Dromaius novaehollandiae. Poultry and Avian Biology Reviews 6, 331 (abstract 13.22).

Williams, K.M., O'Malley, P. & Martin, G.B. (1997). Influence of rainfall on the length of the

breeding season of the emu (Dromaius novaehollandiae) at two latitudes. Proceedings of the Australian Society for Reproductive Biology 29, 46.

Williams, K.M., Sharp, P.J. & Martin, G.B. (1998). Effect of surgical sterilisation on body

composition of male and female Emus (Dromaius novaehollandiae). Proceedings of the Australian Society of Animal Production (in press)

Williams, K.M., Tan, N.S., Blackberry, M.A., O'Malley, P., Sharp, P.J. & Martin, G.B. (1995). Differences in serum concentrations of testosterone and prolactin in broody and non-broody male emus (Dromaius novaehollandiae). Proceedings of the Australian Society for Reproductive Biology 27, 111.

Media articles This is a reflection of our commitment to publicising our work to the community and to industry. In addition to the radio and newspaper items listed here, we organised a workshop on emu farming through the WA branch of the Australian Society of Animal Production (listed above), and presented our work to the International Emu Farmers Congress in Perth (1994, 1995). Bogle, Deborah. (1993). No sex ... we're emus. The Australian, Wednesday 23 June, 1993 (page 9). Campbell, Meredith. (1993). Radio interview: the 'super emu'. Interview on ABC Radio Darwin

(16:15, June 12, 1993). McIlwraith, J. (1993). The sensitive new age 'super emu'. Uniview Magazine (The University of

Western Australia) 12 (2), 6. Night-time Drive. (1993). Radio interview: no sex ... we're emus. Interview on Radio 6PR (18:10,

June 23, 1993). Richardson, Alan. (1993). Radio interview: the 'super emu'. Recorded interview on "The Country

Hour", ABC Radio 720 6WF (June 7, 1993). Schultz, Michael. (1993). Radio interview: the 'super emu'. Interview on ABC Radio 720 6WF

(20:15, May 31, 1993). Vaisey, Melissa (1995). Leading the race in emu breeding. The Countryman (November 30, 1995),

page 10. Vaisey, Melissa (1995). Search hots up for control of emu sex drive. The Countryman (December

14, 1995) page 17.