Somatic Cell Nuclear Transfer (SCNT) is a technology still relatively early in its development. Cloning has been accomplished in relatively few species, with most of our current information stemming from studies in cattle, swine, goats, and mice. This Risk Assessment has addressed the hazards and potential risks that may be experienced by domestic livestock (i.e., cattle, swine, sheep, and goats) involved in the cloning process (Animal Health Risks) and whether edible products from animal clones or their progeny pose food consumption risks beyond those of their conventional counterparts (Food Consumption Risks).
This Risk Assessment used two complementary approaches, the Critical Biological Systems Approach (CBSA) and Compositional Analysis Approach, to identify and characterize potential animal health and food consumption hazards associated with cloning. It then employed a weight of evidence approach to draw conclusions regarding risks to animal health and for consumption of food products from clones and their progeny. This weight of evidence approach consisted of four steps:
- Evaluation of the empirical evidence (i.e., data on molecular mechanisms, physiological measurements, veterinary records, and observations of general health and behavior) for the species being considered;
- Consideration of biological assumptions predicated on our growing understanding of the molecular mechanisms involved in mammalian development;
- Evaluation of the coherence of the observations with predictions based on biological mechanisms; and
- Evaluation of the consistency of observations across all of the species considered, including the mouse model system.
The Risk Assessment also assumed that animal clones, their progeny, and all food products derived from either clones or progeny must meet the same federal, state, and local laws and regulations as food from conventionally bred animals.
Because no exogenous genes have been introduced into animals derived via SCNT, the underlying assumption has been that adverse outcomes observed in animal clones arise from epigenetic modifications due to incomplete reprogramming of the donor cell nucleus. Methodological and technological components (e.g., selection of donor cell, cell cycle stage, in vitro factors associated with the SCNT process) may also affect outcomes as they do for other ARTs.
B. Conclusions Regarding Risks to Animal Health
To assess the health of animal clones for both the animal health and food consumption risk portions of this risk assessment, we used the Critical Biological Systems Approach (CBSA), which divides the life cycle of clones into five distinct Developmental Nodes. Available data for each species were sorted into these Developmental Nodes to evaluate the data systematically and to determine whether there are common developmental difficulties among the livestock clones or whether animals "recover" from initial infirmities related to cloning.
The results of the CBSA indicated that significant adverse health outcomes have been reported for cattle and sheep clones and their surrogate dams. These tend to include dystocia and high gestational mortality. In cattle and sheep clones, post-natal mortality tends to be concentrated in the perinatal period, and is higher in clones than in animals produced using other assisted reproductive technologies (ARTs).
To date, no adverse outcomes have been noted in clones that have not been observed in animals derived via other ARTs or natural mating. Goats and swine appear to develop without significant abnormalities. The incidence of adverse outcomes in cattle and sheep clones, however, appears to be higher than in other forms of ARTs. Common adverse developmental outcomes that have been observed in cattle and sheep generally fall under the heading of Large Offspring Syndrome (LOS), although there may be others. Newborn animals with LOS tend to be bigger than average for their breed and species, may show edema or other abnormalities of the lungs and other parts of the body, and exhibit cardiovascular and respiratory problems.
Most animal clones that survive the critical perinatal period appear to grow and develop normally. Even animals with physiological perturbations, including less severe manifestations of LOS, seem to resolve them, usually within a period of weeks. More severe complications of LOS may persist into the juvenile period, but clones do not appear to develop any additional health risks unrelated to those that were observed during the perinatal period. Clones that reach reproductive age appear to be normal in all of the measures that have thus far been investigated, and appear to give rise to healthy, apparently normal progeny. Mature clones appear normal and healthy, and are virtually indistinguishable from their conventionally bred counterparts.
Studies that evaluated epigenetic reprogramming in live, healthy clones indicate that although there is some variability between clones and their sexually-derived counterparts, these clones have undergone sufficient epigenetic reprogramming to carry out the coordinated functions necessary for survival and normal functioning. Molecular analyses reveal relatively small methylation differences, and either the animals are tolerant of such differences, or the epigenetic differences are below the threshold that poses observable adverse health outcomes.
C. Conclusions Regarding Food Consumption Risks
In order to evaluate potential food consumption risks associated with healthy-appearing clones, we developed a two-pronged approach. The first part of the approach is based on the hypothesis that a healthy animal is likely to be safe to eat, and relied on the CBSA. The second component of our two-pronged approach, or the Compositional Analysis Approach, assumed that if there are no material differences between the composition of milk and meat from animal clones (and their progeny) and their non-clone counterparts, then edible products derived from clone meat or milk would be as safe to eat as corresponding products from non-clones. This assessment assumed that animal clones and their progeny would be subject to all of the same federal and state requirements for milk and meat from conventionally bred animals.
Because each clone arises from an independent event, identification and characterization of potential subtle hazards (e.g., alterations in gene expression, immune function, or hormone levels) is best accomplished by the evaluation of individual animals, at as fine a level of resolution as possible. Characterization of the overall functionality of clones, however, is likely best considered by evaluating the animal as a whole, in particular assessing the degree to which highly complex functions have been integrated, for example by demonstrating normal growth and successful reproduction.
The food consumption portion of the risk assessment postulated that because the only hazards that may be present in clones would arise from epigenetic dysregulation, and because only healthy animals meeting the same standards that conventional food animals or their edible products meet would be permitted for use as food, the only hazards that could be present in these animals would be subtle. Part of the purpose of the CBSA approach was to determine whether any such subtle hazards could be identified. Following a detailed analysis of a wide range of health data, we identified only a few examples of altered physiological parameters in clones, and these were limited to young clone calves and swine. None of these alterations were correlated with any discernable adverse effects on animal health. We therefore concluded that no subtle hazards were identified that could pose food consumption risks from cattle, swine, or goat clones. The lack of allergenic and mutagenic effects in studies designed to detect those outcomes also indicated there were no food consumption risks.
Analyses of the composition of meat from bovine and swine clones and milk from bovine clones consistently indicate that there are no biologically relevant differences between the composition of food from clones, or their close comparators. In addition, there is no material difference, based on these studies, between the composition of meat and milk from clones and historical reference ranges of the composition of food from conventionally-bred animals.
D. Conclusions Regarding Food Consumption Risks from Clone Progeny
Progeny of animal clones are not anticipated to pose special animal health or food consumption concerns, as they are the product of sexual reproduction. The production of gametes by clones is expected to reset even those residual epigenetic reprogramming errors that could persist in healthy, reproducing clones. Studies in cattle and swine indicate that the progeny of clones are healthy and indistinguishable from other sexually-derived swine comparators. Because the value of clones lies in their genes, they are most likely to be used as breeding stock, and their use as food would be incidental. Almost all of the production animals (i.e., sources of meat and milk) from the overall SCNT process are therefore likely to be sexually-reproduced progeny of clones. An extensive dataset on the progeny of swine clones indicates that the composition of meat from those animals does not differ materially from that of comparator animals or historical reference ranges.
E. Weight of Evidence Evaluations
As is the case for all risk assessments, the amount of data and information available on endpoints varied in quantity, with respect to both number of studies and number of data points available to evaluate. In some cases, evaluation of one endpoint indicated different outcomes in different laboratories. This Risk Assessment took such differences into account by applying the criteria listed above to consider the overall weight of evidence— that is, the extent to which the data supported each other, and where they did not, to provide a framework for determining the underlying basis for the differences. Because the weight of evidence considers all of the information in one framework, it does not require any particular number of studies on any particular endpoint in order to be valid; rather, by considering all of the information together, it is able to develop a coherent perspective that takes into account biological assumptions, mechanisms, and empirical evidence. It also allows for the identification of uncertainties, and provides a science-based path for further investigations to resolve those uncertainties.
The judgment that cattle, swine, and goat clones meeting the same federal and state requirements as conventionally bred food animals would not likely pose food consumption risks, of course, contains some residual uncertainty. The source(s) of the uncertainty may be sorted into three categories:
- Uncertainties associated with empirical observations. Uncertainties are lowest for those individual clones whose health has been thoroughly evaluated and, by inference, other clones produced using the same methodology. The uncertainties associated with the evaluation of empirical observations can be a function of the size, consistency, and quality of the data being evaluated. For example, the degree of confidence that can be placed in judgments arising from a well-conducted, consistent, and extensive dataset is much higher than from a small, poorly designed, and highly variable dataset. Further, because datasets tend to arise from an individual laboratory or producer, the uncertainties associated with that producer and method are lower than for other laboratories or producers for which less information is available.
- Uncertainty stemming from biological sources can be minimized by the evaluation of the clones themselves. The most important factor in this evaluation is the healthy survival and functionality of individual clones, indicating that either the animal has minimal epigenetic dysregulation, or that any initial epigenetic dysregulation has been resolved. Uncertainty would be the lowest for individual clones demonstrating successful reproduction.
- Uncertainties stemming from technological or methodological grounds encompass the degree to which judgments regarding clones arising from technologies in use when this risk assessment was conducted can be applied to modifications of the technology. These may only be resolved by the evaluation of the outcomes of those technological changes (i.e., the actual clones).
Thus, our overall conclusions are:
For Animal Health: SCNT results in an increased frequency of health risks to animals involved in the cloning process, but these do not differ qualitatively from those observed in other ARTs or natural breeding. Cattle and sheep exhibit a set of clinical signs collectively referred to as LOS that do not appear to be present in swine or goats. Surrogate dams are at risk of complications from birth if the fetus suffers from LOS, or from accumulation of fluid in the placenta (hydrops). Clones exhibiting LOS may require additional supportive care at birth, but can recover and mature into normal, healthy animals. Most clones that survive the perinatal period are normal and healthy as determined by physiological measurements, behavior, and veterinary examinations. Progeny of animal clones also have been reported as normal and healthy.
For Food Consumption Risks: Extensive evaluation of the available data has not identified any food consumption risks or subtle hazards in healthy clones of cattle, swine, or goats. Thus, edible products from healthy clones that meet existing requirements for meat and milk in commerce pose no increased food consumption risk(s) relative to comparable products from sexually-derived animals. The uncertainties associated with this judgment are a function of the empirical observations and underlying biological processes contributing to the production of clones. Uncertainty about the health of clones decreases as they age and have more time to exhibit the full range of functionality expected of breeding stock. Edible products derived from the progeny of clones pose no additional food consumption risk(s) relative to corresponding products from other animals based on consistent empirical observations, underlying biological assumptions, and evidence from model systems.