We thank the Mississippi Department of Wildlife, Fisheries and Parks (MDWFP) for financial support using resources from the Federal Aid in Wildlife Restoration Act (W-48-61). We thank MDWFP biologists W. McKinley, A. Blaylock, A. Gary, and L. Wilf for their extensive involvement in data collection. We also thank S. Tucker as facility coordinator and multiple graduate students and technicians for their help collecting data. This manuscript is contribution WFA427 of the Mississippi State University Forest and Wildlife Research Center.

Ethics Statement: The Mississippi State University Institutional Animal Care and Use Committee approved all capture, handling, and marking techniques under protocols 04-068, 07-036, 10-033 and 13-034.
1. Establish Capture Sites, Immobilize and Transport wild White-tailed Deer
<ol>
	<li>Identify public and private properties that are enrolled in the Deer Management Assistance Program22 and establish &#8805;29 capture sites throughout three source regions in Mississippi, USA.</stron...

<ol>
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There are several steps associated with our protocol; however, there are four critical steps that must be taken to ensure success with this protocol. First, during capture of wild deer, there must be several capture locations throughout a single source region (step 1.1.1). Having multiple capture locations ensures that any genetic variability associated with the source region will be represented among deer. Second, deer must be kept separated by source region during breeding season (steps 1.4.9 and 5.2.1). Ensuring anima...

Environmental factors a mother experiences during gestation and lactation may influence her offspring&#39;s phenotype, independent of genotype1,2,3. Mothers who inhabit high-quality environments likely will produce offspring that exhibit a luxury phenotype (large antler and body size4), whereas mothers who inhabit a low-quality environment may produce offspring that exhibit an efficiency phenotype (small antler and body size4). Therefore, persisting in a high-quality environment may allow a mother to produce male offspring with large phenotypic characteristics, which may directly influence the offspring&#39;s reproductive opportunities5,6,7,8 and indirectly influence the mother&#39;s inclusive fitness.
Although nutrition directly influences phenotypic characteristics across taxa (Ursus americanus, Ursus arctos9; Liasis fuscusI10; Larus michahellis11), several factors may affect white-tailed deer phenotypes in Mississippi, USA. Antler and body size are about one-third larger for some populations compared to others12. This variation is strongly correlated with forage quality13,14; the largest males are found in areas with the greatest quality of forage. However, historic restoration efforts of white-tailed deer in Mississippi may have led to genetic bottlenecks and/or founder effects15,16, which may also partially explain some of the observed regional variation in white-tailed deer phenotype.
We provide the protocol we used to control nutritional quality of wild-caught white-tailed deer, which allowed us to assess whether male phenotype is restricted by population-level genetics. This protocol also allowed us to assess whether lagging maternal effects were present in our populations. Our controlled design is preferential to studies conducted on free ranging populations which are limited to using environmental variables as a proxy for nutritional restriction3,17. Our controlled design also allows for other variables such as potential chronic stress related to social interactions to be held constant as all individuals are subjected to similar housing and husbandry practices. Additionally, because nutrition directly influences other life history aspects ranging from reproduction to survival18,19, controlling nutrition allows for investigators to assess other variables that affect mammalian life history aspects. Similar protocols have been described to assess questions related to life history aspects for other ungulates across North America (e.g., 20,21).

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	<tbody>
		<tr height="20">
			<td height="20" style="width:416px;height:20px;">Shelled Corn</td>
			<td style="width:155px;"></td>
			<td style="width:163px;"></td>
			<td style="width:64px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Elevated Stand</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Safety Harness</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ground Blind</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Model 196 Projector</td>
			<td style="width:316px;">Pneu-Dart, Pennsylvania, USA</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">3cc Radio-Telemetry Darts</td>
			<td style="width:316px;">(Pneu-Dart, Pennsylvania, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Various Sized Darts&#160;</td>
			<td style="width:316px;">(Pneu-Dart, Pennsylvania, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Teletamine HCl&#160;</td>
			<td style="width:316px;">(Telazol, Fort Dodge Animal Health, Iowa, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Xylazine HCl&#160;</td>
			<td style="width:316px;">(West Texas Rx Pharmacy, Amarillo, Texas, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Yhoimbine HCl</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tolazoline HCl</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Military Style Gurney</td>
			<td></td>
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			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Rectal Thermometer</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Shade Cloth</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">20% Crude Protein Deer Pellets&#160;</td>
			<td style="width:380px;">(Purina AntlerMax Professional High Energy Breeder 59UB, Purina, Missouri, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Trough Style Feeders</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Commercial Clover&#160;</td>
			<td style="width:380px;">(Durana Clover, Pennington Seed Co., Georgia, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Commercial Fescue&#160;</td>
			<td style="width:380px;">(Max-Q Fescue, Pennington Seed Co., Georgia, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Blankets</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ice Packs</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Broadleaf Weed Control (2, 4-DB Herbacide, Butyrac 200)</td>
			<td></td>
			<td></td>
			<td></td>
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		<tr height="20">
			<td height="20" style="height:20px;">Grass Control&#160;</td>
			<td style="width:316px;">(Poast Herbacide, BASF Co.)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Pelleted Wormer</td>
			<td>Safeguard Co.,&#160;</td>
			<td></td>
			<td style="width:227px;">active ingredient fenbendazole</td>
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			<td height="20" style="height:20px;">Parasite Pour-on Treatment&#160;</td>
			<td>(Ivomec, Merial Co.)</td>
			<td></td>
			<td></td>
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		<tr height="20">
			<td height="20" style="height:20px;">Insecticide</td>
			<td style="width:316px;">Riptide, McLaughlin Gormley King Co.)&#160;</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Medium and Large Plastic Ear Tags&#160;</td>
			<td>(Allflex, Texas, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Remote site that assigned parentage</td>
			<td style="width:380px;">DNA Solutions Animal Solutions Manager (DNA Solutions, Oklahoma, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Digital Hanging Scale&#160;</td>
			<td style="width:316px;">(Moultrie, EBSCO Industries, Inc.)&#160;</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tape Measure</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Clostridium Perfringens Types C and D Toxoid Essential 3&#160;</td>
			<td>(Colorado Serum Co.)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Clostridium Perfringens Types C and D Antitoxin Equine Origin</td>
			<td>(Colorado Serum Co.)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ivermectin in propylene glycol</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Antibiotic</td>
			<td style="width:380px;">(Nuflor, Schuering-Plough Animal Health Corp., New Jersey, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ivermectin&#160;</td>
			<td style="width:380px;">(Norbrook Labratories, LTD., Down, Northern Ireland, UK)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Clostidrial vaccine</td>
			<td style="width:316px;">(Vision 7 with SPUR, Ivesco LLC, Iowa, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Leptospirosis vaccine&#160;</td>
			<td style="width:316px;">(Leptoferm-5, Pfizer, Inc., New York, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Trailer for transport</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Reciprocating saw&#160;</td>
			<td style="width:316px;">(DEWALT, Maryland, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Scientific Digital Scale&#160;</td>
			<td style="width:316px;">(Global Industrail, Global Equipment Company Inc)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Antler Measuring Tape</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Fogger</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Plastic Ear Tags&#160;</td>
			<td>(Allflex, Texas, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Plastic Ear Tagger</td>
			<td>(Allflex, Texas, USA)</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

protocol for assessing the relative effects of environment and genetics on antler and body growth for a long-lived cervid

Cervid phenotype can be placed into one of two categories: efficiency, which promotes survival over extravagant morphometric growth, and luxury, which promotes growth of large weaponry and body size. Populations of the same species display each phenotype depending on environmental conditions. Although antler and body size of male white-tailed deer (Odocoileus virginianus) varies by physiographic region in Mississippi, USA and is strongly correlated with regional variation in nutritional quality, the effects of population-level genetics from native stocks and previous re-stocking efforts cannot be disregarded. This protocol describes how we designed a controlled study, where other factors that influence phenotype, such as age and nutrition, are controlled. We brought wild-caught pregnant females and six-month-old fawns from three distinct physiographic regions in Mississippi, USA to the Mississippi State University Rusty Dawkins Memorial Deer Unit. Deer from the same region were bred to produce a second generation of offspring, allowing us to assess generational responses and maternal effects. All deer ate the same high-quality (20% crude protein deer pellet) diet ad libitum. We uniquely marked each neonate and recorded body mass, hind foot, and total body length. Each subsequent fall, we sedated individuals via remote injection and sampled the same morphometrics plus antlers of adults. We found that all morphometrics increased in size from first to second generation, with full compensation of antler size (regional variation no longer present) and partial compensation of body mass (some evidence of regional variation) evident in the second generation. Second generation males that originated from our poorest quality soil region displayed about a 40% increase in antler size and about a 25% increase in body mass when compared to their wild harvested counterparts. Our results suggest phenotypic variation of wild male white-tailed deer in Mississippi are more related to differences in nutritional quality than population-level genetics.

Individual age, nutritional quality, and genetics influence male white-tailed deer phenotype. Our study design allowed us to control the quality of nutrition deer were consuming and allowed us to identify the age of each deer for valid comparisons within year classes. By controlling nutrition and age with our study design, we were better able to understand whether population-level genetics were restricting the phenotype of males from two study populations. Improved nutrition had a positiv...

Watch this Scientific Journal Video about Protocol for Assessing the Relative Effects of Environment and Genetics on Antler and Body Growth for a Long-lived Cervid at JoVE.com

Protocol for Assessing the Relative Effects of Environment and Genetics on Antler and Body Growth for a Long-lived Cervid

Phenotypic differences among cervid populations may be related to population-level genetics or nutrition; discerning which is difficult in the wild. This protocol describes how we designed a controlled study where nutritional variation was eliminated. We found that phenotypic variation of male white-tailed deer was more limited by nutrition than genetics.

Cervid phenotype can be placed into one of two categories: efficiency, which promotes survival over extravagant morphometric growth, and luxury, which ...

The overall goal of this experimental design is to assist whether population level genetics restrict white-tailed deer, odocoileus virginianus antler and body size by controlling for other variables that also influence antler and body size such as age and nutrition. This methodology can really help us understand the true cause of morphological variation in wildlife species across their range. And this is important because morphology is the basis for historical designation of sub-species and with Endangered Species Act codifying those sub-species differences it's important to understand are those differences really due to genetic differences that are meaningful and need to be preserved or are they just nutritional habitat quality differences that are associated with the habitat where those sub-species occur?The main advantage of this technique is that it allows us in a controlled environment to focus on a specific characteristic of the environment that the animals were raised in naturally and in this case we're removing the animals from their natural habitat and controlling for nutrition to see if nutrition is the cause for the morphological differences. This technique can actually improve the implementation of the Endangered Species Act because it's gonna allow state resource agencies with limited resources to identify meaningful genetic variation. Demonstrating the technique today will be Dan Morina.A graduate student here at the MSU Deer Lab. Begin by insuring that side fencing has shade cloth to act as a visual and physical barrier between pens. Install elevated box blinds at one end of each pen to facilitate darting events during data collection.Keep two trough style feeders at separate ends of each pen to reduce competition for food among deer. Finally, provide deer with a high quality diet add Lupitum. Begin by walking the technician who'll be darting to the end of the pen where the elevated blinds are located.Post a single technician into a blind. Then have the individual who walked the technician to the blind walk back to the opposite end of the pen. After confirming sedation of the deer by checking for eye reflexes apply ophthalmic ointment to the eyes.And blindfold deer to reduce stress. Load the deer on a military-style gurney. Next use a rectal thermometer to assess body temperature after recovery.Warm deer with heated blankets if the animals temperature is below 37.7 degrees Celsius. Or cool the deer with ice packs if the animal's temperature is above 40 degrees Celsius. And transport it via utility task vehicle to a predetermined data collection area.Once transported, measure body mass to the nearest hundredth of a kilogram with a digital hanging scale. As well as hindfoot length and total body length to the nearest millimeter. Then administer size-appropriate amounts of antibiotic ivermectin, a custodial vaccine and a leptospirosis vaccine.Next, while the animal is sedated take three antler measurements from adult males using an antler measuring tape. Measure the inside spread, basal circumference and the main beam length of antlers prior to antler removal. Then remove antlers about three centimeters above the burr using a reciprocating saw.After the data is collected place the deer into the appropriate pen. And administer either 125 milligrams per kilogram of yohimbine hydrochloride or 4.0 milligrams per kilogram of tylosin hydrochloride to reverse the effects of xylazine hydrochloride. Finally, monitor the deer to ensure they remain in a sternal position until they come out of sedation and are fully alert.After acquiring the antler samples measure each individual tine protruding from the main beam and additional abnormal points by aid of wire. Wrap the wire around where the tine intersects the main beam and mark that point for reference. Then measure from this reference point to the tip of the tine and repeat for each tine.Next collect the remaining circumference measurements by identifying the smallest point between the G1 and G2 tines the G2 and G3 tines and the G3 and G4 tine if present. If the G4 tine is not present measure the distance between the mid-point of the G3 tine at the end of the main beam. And measure the H4 circumference at the midway point.Finally, weigh antlers to the nearest tenth of a gram using a scientific digital scale. And design a minimal, critical antler mass of one gram for first year animals with antler shorter than three centimeters. Results from this study indicated that improved nutrition had a positive effect on all morphometric measurements on a 3.5 year old male white-tailed deer from each source region.Antler mass and antler size did not vary among the three source regions after two generations of improved nutrition. Which suggest that antler size is not restricted by population level genetics. Further, body mass greatly increased from first to second generation.Suggesting there is a nutritional limitation in the wild for all populations. However, there were still regional variation among second generation males. Once mastered, this technique only takes about an hour per animal.But sample size is really important so you wanna have as many animals in your study as possible so you could end up spending several days if needed to process all of your animals. While attempting this procedure it's really important that everything within the deer pens is set up for success. The blind needs to be properly placed at the correct distance from the fence.Shooting lanes need to be cut if there are branches in the way. And everything that you can control needs to be controlled in advance because the one thing you cannot control is how the animal will behave under the particular stressful situation you're putting them into. Using this procedure you could also modify the nutritional components of the experiment.And look at how protein or energy intake might alter that genetic potential and the morphological expression of the animal. This same methodology can be used by other researchers to vary the type of treatment that's supplied to the animals and look at other things beside morphological development. You could look at how nutritional intake affects life history traits such as reproductive traits and recovery from the demand of gestation and lactation.After watching this video you should have a good idea of how to control nutrition and age to see how genetics and nutrition interact in the morphological development of an animal within a captive facility. Don't forget that whenever you sedate a wild animal even in a captive facility there's a chance for injury to the animal so we have to take care and monitor them closely.

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7.2K vues.  Mississippi State University. The overall goal of this experimental design is to assist whether population level genetics restrict white-tailed deer, odocoileus virginianus antler and body size by controlling for other variables that also influence antler and body size such as age and nutrition. This methodology can really help us understand the true cause of morphological variation in wildlife species across their range. And this is important because morphology is the basis for historical designation of sub-species and w...