H. Michael Kubisch

Sexual dimorphism among bovine embryos in their ability to make the transition to expanded blastocyst and in the expression of the signaling molecule IFN-τ

IFN-τ is a secretory product of trophectoderm of cattle, sheep, and their relatives and is expressed for a few days in early pregnancy after the blastocyst first forms. It serves to alert the mother that she is pregnant. A delayed or less than robust IFN-τ signal is a likely cause of embryonic loss. Here we have determined whether blastocyst production of IFN-τ, which is encoded by a cluster of genes on chromosome 9, differs between the sexes in cattle, as assessed by culture of in vitro-derived embryos on two different media, one complex (tissue culture medium 199 supplemented with serum) with coculture support, the other relatively simple (synthetic oviductal fluid plus albumin). With both media, female blastocysts produced approximately double the amount of IFN-τ as males, regardless of such variables as oocyte batch, blastocyst quality, hatching, and length of time in culture. However, in either tissue culture medium 199, which contains 5.5 mM d-glucose, or in synthetic oviductal fluid, in the presence but not in the absence of added glucose, significantly fewer female than male embryos were able to progress from the morula/early blastocyst stage to more advanced stages of development. It is possible that the differences between male and female embryos both in their production of IFN-τ and in their ability to progress in development in glucose-rich media are manifestations of phenomena that occur in vivo and provide plasticity in embryo selection during early pregnancy.

Male‐to‐male differences in post‐thaw motility of rhesus spermatozoa after cryopreservation of replicate ejaculates

Background  The efficiency of controlled propagation to produce rhesus monkeys of particular genotypes can be maximized by use of cryopreserved spermatozoa collected from specific males to inseminate appropriate females. But this assumes that semen from males with different genotypes can be cryopreserved with equal effectiveness.

Control of interferon-τ secretion by in vitro-derived bovine blastocysts during extended culture and outgrowth formation

A series of experiments was conducted to examine the pattern of interferon‐τ (IFN‐τ) secretion by bovine blastocysts during extended culture in vitro. In the first experiment, blastocysts were cultured individually for three 48‐hour periods. The day of blastocyst formation affected how much IFN‐τ was produced during the first two culture periods, but not during the third period. The overall secretion of IFN‐τ during the 6‐day period increased significantly and well beyond what could be accounted for by the concomitant increase in cell numbers. In the second experiment, blastocysts were initially cultured in individual droplets for 48 hr, then plated into 48‐well plates. Medium concentrations of IFN‐τ were determined after 48 hr and again after 6 and 12 days of culture. Initial IFN‐τ secretion did not affect the ability to form outgrowths or their final size, and initial differences in secretion between groups of blastocysts had disappeared by the second and third analyses. In the third experiment, blastocysts were cultured individually for 48 hr in droplets containing the medium that had been flushed through the uteri of non‐pregnant sheep on days 10, 12, and 15 of the estrous cycle. Culture in the medium obtained from the Day 15 flush significantly increased the number of cells that blastocysts contained, as well as IFN‐τ secretion. Mol. Reprod. Dev. 58:390–397, 2001.

Age as a determinant of reproductive success among captive female rhesus macaques (Macaca mulatta).

A retrospective analysis was performed on fertility outcomes among a colony of captive Indian rhesus monkeys. The analysis covered over 30 years and was based on 1443 females with a total of 11,453 pregnancies. Various determinants of fertility were assessed including birth rates, pregnancy loss, infant survival, interbirth intervals, and interval from last birth to death. Binary variables were analyzed with generalized linear models with random intercepts, while linear mixed models were used for analysis of continuous variables. Age of the dam was a significant factor in determining whether a pregnancy resulted in a birth and whether an infant survived the first 30 days with primiparous or older mothers being less likely to produce an infant surviving to that age. In contrast, sex proved to be the only significant factor in determining whether an infant lived to 1 year, with females being more likely to survive. The interval between births proved to be affected primarily by dam age, while the late death of an infant depressed the likelihood of an extended time interval between her last birth and her death. Overall, these results demonstrate that maternal age contributes significantly to a decline in fertility and older females can live relatively long periods following birth of their last infant.

Resources for genetic management and genomics research on non-human primates at the National Primate Research Centers (NPRCs)

Abstract The National Primate Research Centers (NPRCs) established Working Groups (WGs) for developing resources and mechanisms to facilitate collaborations among non‐human primate (NHP) researchers. Here we report the progress of the Genome Banking and the Genetics and Genomics WGs in developing resources to advance the exchange, analysis and comparison of NHP genetic and genomic data across the NPRCs.

Development and validation of a SNP‐based assay for inferring the genetic ancestry of rhesus macaques (Macaca mulatta)

Rhesus macaques (Macaca mulatta) are an important primate model species in several areas of biomedical research. The wide geographic distribution of this species has led to significant genetic differentiation among local and regional populations. These regional differences can be important factors in the selection of the most appropriate subjects for particular research studies, as animals from different populations can respond differently to the same experimental treatment. Consequently, it is valuable to confirm the ancestry of individual rhesus monkeys from geographically distinct populations. Using DNA samples obtained from rhesus macaques from six National Primate Research Centers, we tested a set of 384 potential ancestry informative single nucleotide polymorphisms (SNPs) and identified a final panel of 91 SNPs that can reliably distinguish Indian‐origin from Chinese‐origin rhesus monkeys. This genetic test can be used to determine the ancestral origin of animals and to detect individuals that are hybrids between these two regional populations. To demonstrate use of the SNP panel, we investigated the ancestry of 480 animals from the Yerkes NPRC (YNPRC) for which the colony records were insufficient to clearly establish ancestry. Three of the YNPRC animals tested were determined to be hybrids. This SNP ancestry tool will be useful to researchers, colony managers, and others who wish to evaluate the ancestral origin of individual rhesus macaques, and therefore will facilitate more effective and efficient use of these animals in biomedical research. Am. J. Primatol. 76:1105–1113, 2014.

An empirical comparison of short tandem repeats (STRs) and single nucleotide polymorphisms (SNPs) for relatedness estimation in Chinese rhesus macaques (Macaca mulatta)

We compare the effectiveness of short tandem repeat (STR) and single nucleotide polymorphism (SNP) genotypes for estimating pairwise relatedness, using molecular data and pedigree records from a captive Chinese rhesus macaque population at the California National Primate Research Center. We find that a panel of 81 SNPs is as effective at estimating first‐order kin relationships as a panel of 14 highly polymorphic STRs. We note, however, that the selected STRs provide more precise predictions of relatedness than the selected SNPs, and may be preferred in contexts that require the discrimination of kin related more distantly than first‐order relatives. Additionally, we compare the performance of three commonly used relatedness estimation algorithms, and find that the Wang [2002] algorithm outperforms other algorithms when analyzing STR data, while the Queller & Goodnight [1989] algorithm outperforms other algorithms when analyzing SNP data. Future research is needed to address the number of SNPs required to reach the discriminatory power of a standard STR panel in relatedness estimation for primate colony management. Am. J. Primatol. 76:313–324, 2014.

Estimates of heritability for reproductive traits in captive rhesus macaque females

Records from a colony of captive Indian rhesus macaques (Macaca mulatta) were used to estimate heritability for a number of reproductive traits. Records were based on a total of 7,816 births by 1,901 females from 1979 to 2007. Heritability was estimated with a linear animal model using a multiple trait derivative free REML set of programs. Because no male parents were identified, the numerator relationship matrix contained female kinships established over six generations. Reproductive traits included female age at the birth of the first, second and last infant, age at death, inter‐birth intervals, number of infants born per female and infant survival. Heritability for each trait was estimated as the ratio of the additive genetic variance to phenotypic variance adjusted for significant fixed effects. Estimates of heritability for early reproduction ranged from 0.000±0.072 for birth interval after the first reproduction to 0.171±0.062 for age of female at the first infant. Higher estimates of heritability were found for female longevity [0.325±0.143] and for productivity of deceased females born before 1991 [0.221±0.138]. Heritability for infant survival ranged from 0.061±0.018 for survival from 30 days to 1 year to 0.290±0.050 for survival from birth to 30 days when adjusted to an underlying normal distribution. Eight of the 13 estimates of heritability for reproductive traits in this study were different from zero [P<0.05]. Generally, heritability estimates reported in this study for reproductive traits of captive rhesus macaque females are similar to those reported in the literature for free‐ranging rhesus macaque females and for similar reproductive traits of other species. These estimates of heritability for reproductive traits appear to be among the first for a relatively large colony of captive rhesus macaque females. Am. J. Primatol. 72:811–819, 2010.

Postthaw survival of rhesus macaque sperm: variation in the response of individual males to different freezing protocols.

This study was designed to assess the effects of various cryopreservation protocols on the postthaw survival of rhesus macaque sperm. Multiple ejaculates of five mature males were collected and frozen by each of four different methods. Cryopreservation significantly reduced motility in all samples, regardless of the method, and the response of different ejaculates of the same male to each method proved to be consistent. The freezing method was a significant factor in determining the immediate postthaw motility, although this effect disappeared after 4 hr. Moreover, there was a significant interaction between individual males and freezing method suggesting that developing a single freezing protocol that is universally suitable may be difficult. Am. J. Primatol. 70:1093–1096, 2008.

Reproductive Efficiency of Captive Chinese- and Indian-Origin Rhesus Macaque (Macaca mulatta) Females

Reproductive and survival records (n = 2,913) from 313 Chinese‐origin and 365 Indian‐derived rhesus macaques at the Tulane National Primate Research Center (TNPRC) spanning three generations were studied. Least‐squares analysis of variance procedures were used to compare reproductive and infant survival traits while proportional hazards regression procedures were used to study female age at death, number of infants born per female, and time from last birth to death. Chinese females were older at first parturition than Indian females because they were older when placed with males, but the two subspecies had similar first postpartum birth interval (1st PPBI) and lifetime postpartum birth interval (LPPBI). Females that gave birth to stillborn infants had shorter first postpartum birth intervals (1st PPBI) than females giving birth to live infants. Postpartum birth intervals decreased in females from age 3 to 12 but then increased again with advancing age. Chinese infants had a greater survival rate than Indian infants at 30 days, 6 months, and 1 year of age. Five hundred and forty‐three females (80.01%) had uncensored, or true records for age at death, number of infants born per female, and time from the birth until death whereas 135 females (19.91%) had censored records for these traits. Low‐ and high‐uncensored observations for age at death were 3 and 26 years for Chinese, and 3 and 23 years for Indian females. Uncensored number of infants born per female ranged from 1 to 15 for Chinese females and 1 to 18 for Indian females. Each of these traits was significantly influenced by the origin × generation interaction in the proportional hazards regression analyses, indicating that probabilities associated with age at death, number of infants born per female, and time from last birth to death for Chinese and Indian females did not rank the same across generations.