PROC GENMOD Research Articles

Relationship of Stunting and Overweight in Egyptian children under five years of age: Trends and associated risk factors

BackgroundIn Egypt, rates of child overweight and obesity are consistently increasing. While progress has been made in the reduction of stunting, Egypt's rate of 21% is still higher than countries in the region that have the same GDP.ObjectivesThe objectives were to to examine and understand the trends and variability of malnutrition in all its forms (over and under nutrition) in Egyptian children under five, across time, regions and socio‐economic status and examine associated factors.MethodsDHS data from 1988 to 2014 were downloaded cleaned and analyzed using SAS (Statistical Analysis Software). Anthropometric indicators for children under five were calculated using the WHO 2006 growth reference. Descriptive statistics of dependent variables (stunting, overweight, overweight in stunted children, wasting, Z‐scores of HAZ, WAZ and WHZ) and independent variables (maternal BMI, occupation, education, birth size, birth order, maternal age at first birth, wealth index, infant and young child feeding practices) were computed. Bivariate analysis was conducted using logistic regression models. Multi‐variate analysis utilized the log‐binomial regression model (PROC GENMOD) to adjust for complex variation structures in the data.ResultsRates of stunting steadily decreased from 36% in 1988 to 20% in 2014. Severe stunting reduced from 16.5% to 9.6%. Overweight went from 7 % in 1988 to 13% in 2014. The percentage of stunted children who are overweight went from 9% in 1988 to 34% in 2014. Stunting across wealth categories changed over time with rates being higher in the low socio‐economic categories until 2005 while 2008 and 2014, the data show a flat line across wealth index. Multivariate analysis shows that boys were more likely to be stunted (Prevalence ratio PR= 1.11, p <0.001). Factors associated with stunting were birth order (PR =1.02, p<0.05) and number of children under five. Factors protective against stunting included location (rural PR=0.86, p<0.001), secondary school education in the mother (PR=0.90, P<0.001), obese BMI category (compared to underweight), birth size (very large to smaller than average compared to very small), and household size (p<0.05).Factors that were protective against being overweight as a stunted child included being a boy (PR=0.88, p<0.001) and age of the child (PR=0.995, p<0.0001). Factors that increased the risk of being overweight as a stunted child included secondary education in mothers (PR=1.11, p<0.05), maternal occupation (PR=1.01, p<0.001), BMI category (all categories compared to underweight), household size, very large to normal birth size compared to very small birth size and ever being breast fed.ConclusionIn Egypt, stunting has decreased but overweight has increased in the population over time. More concerning is the increased prevalence of overweight in a stunted child. In recent surveys, wealth is not a factor associated with either stunting or being overweight in a stunted child. Key factors such as birth size and maternal BMI that have traditionally been used in policy initiatives, as markers of reducing stunting on the other hand are associated with increased overweight.Support or Funding InformationThis study is supported by the USAID Feed the Future Innovation Lab for Nutrition, USAID Bureau for Food Security and USAID Egypt.

Effects of interdisciplinary teamwork on patient-reported experience of cancer care

BackgroundInterdisciplinary teamwork (ITW) is deemed necessary for quality cancer care practices. Nevertheless, variation in ITW intensity among cancer teams is understudied, and quantitative evidence of the effect of different ITW intensities among cancer teams on patients’ perceived experience of care is limited. This study aims to compare patient-reported experience measures (PREMs) of cancer outpatients followed by teams characterized by high vs. low ITW intensity.MethodsThe study is designed as an ex post facto quasi-experimental study. Participants (n = 1379) were recruited in nine outpatient oncology clinics characterized by different ITW intensities. ITW intensities were evaluated using the characteristics of structure (team composition and size) and process (interactions among team members), as per West’s seminal work on team effectiveness. ITW intensity was dichotomized (high vs. low ITW intensity). PREMs were classified and measured using validated scales corresponding to six dimensions: Prompt access to care, Person-centred response, Quality of patient-professional communication, Quality of the care environment, Continuity of care, and Results of care. Dichotomous variables were created for each dimension (positive vs. less positive experience). Multiple logistic regression analyses were performed to assess the association between ITW intensities and the six PREMs dimensions, while controlling for patient and organizational characteristics. PROC GENMOD was used to fit logistic models for categorical variables.ResultsOutpatients treated by teams characterized by high ITW intensity reported almost four times more positive perceptions of Prompt access to care compared to patients treated by low ITW intensity teams (OR = 3.99; CI = 1.89–8.41). High ITW intensity also positively affected patients’ perceptions of Quality of patient-professional communication (OR = 2.37; CI = 1.25–4.51), Person-centred response (OR = 2.11; CI = 1.05–4.24], and Continuity of care (OR = 2.18; CI = 1.07–4.45). No significant association was found between ITW intensity and perceived Results of care (OR = 1.31; CI = 0.68–2.52) or Quality of the care environment (OR = 0.66; CI = 0.31–1.39).ConclusionsThis study provides empirical evidence, from the patient’s perspective, that ITW intensity affects some critical aspects of patient-reported quality of care. Future research will allow explaining how and why ITW structure and processes may contribute to positive cancer care experiences.

Impact of a Child Life and Music Therapy Procedural Support Intervention on Parental Perception of Their Child's Distress During Intravenous Placement.

Child life specialists and music therapists have a unique and integral role in providing psychosocial care to pediatric patients and families. These professionals are trained to provide clinical interventions that support coping and adjustment and reduce the risk of psychological trauma related to hospital visits and health care encounters. The researchers devised a multimodal approach using a combined child life and music therapy intervention to address procedure-related distress in patients receiving intravenous (IV) placement in the pediatric emergency department. The aim of this study was to investigate the efficacy of this collaborative intervention by evaluating parental perception of their child's distress. This study was a prospective analysis investigating the impact of a child life and music therapy intervention on children aged 4 to 11 years old receiving an IV placement in the pediatric emergency department. Efficacy was evaluated by comparing scores between a 4-question pretest and subsequent 4-question posttest that asked the child's parent to evaluate how they anticipated their child would respond to the procedure, and then to evaluate how they perceived their child to have responded after the procedure. Qualitative data were collected in the form of open-ended comments, which were accommodated at the end of the posttest. Data were analyzed by the Cochran-Mantel-Haenszel method for testing repeated ordinal responses and the PROC GENMOD procedure in the SAS system software. A total of 41 participants were enrolled in this study. Results of the statistical analysis revealed significant differences between all pre- and posttest scores (P < 0.05), and significant likelihood that the patient would improve relative to the 4 questions, as a result of the child life and music therapy intervention. Improvement was demonstrated across all 4 questions, suggesting that the child life and music therapy intervention supported healthy, adaptive coping and helped to minimize distress experienced by patients during IV placement. These results underscore the importance and potential clinical impact of child life psychological preparation and psychotherapy-based music therapy interventions in reducing distress in pediatric patients during common medical procedures.

Stroke-related stigma among West Africans: Patterns and predictors

BackgroundDisability-adjusted life-years lost after stroke in Low & Middle-Income Countries (LMICs) is almost seven times those lost in High-income countries. Although individuals living with chronic neurological and mental disorders are prone to stigma, there is a striking paucity of literature on stroke-related stigma particularly from LMICs. ObjectiveTo assess the prevalence, severity, determinants and psycho-social consequences of stigma among LMIC stroke survivors. MethodsBetween November 2015 and February 2016, we conducted a cross-sectional survey of 200 consecutive stroke survivors attending a neurology clinic in a tertiary medical center in Ghana. The validated 8-Item Stigma Scale for Chronic Illness (SSCI-8) questionnaire was administered to study participants to assess internalized and enacted domains of stigma at the personal dimension with further adaptation to capture family and community stigma experienced by stroke participants. Responses on the SSCI-8 were scored from 1 to 5 for each item, where 1=never, 2=rarely, 3=sometimes, 4=often and 5=always with a score range of 8–40. Demographic and clinical data on stroke type and severity as well as depression and Health-Related Quality of Life indicators were also collected. Predictors of stroke-related stigma were assessed using Linear Models (GLM) via Proc GENMOD in SAS 9.4. Results105 (52.5%) subjects recruited were males and the mean±SD age of stroke survivors in this survey was 62.0±14.4years. Mean SSCI-8 score was highest for personal stigma (13.7±5.7), which was significantly higher than family stigma (11.9±4.6; p=0.0005) and social/community stigma (11.4±4.4; p<0.0001). Approximately 80% of the cohort reported experiencing mild-to-moderate degrees of stigma. A graded increase in scores on the Geriatric Depression Scale and Centre for Epidemiological Studies-Depression scale was observed across the three categories. Living in an urban setting was associated with higher SSCI-8 scores. Moreover, stroke subjects with more severe post-stroke residual symptom deficits reported a significantly higher frequency of stigma. ConclusionFour out of five stroke survivors in this Ghanaian cohort reported experiencing some form of stigma. Stigmatized individuals were also more likely to be depressed and have lower levels of quality of life. Further studies are required to assess the consequences of stigma from stroke in LMIC.

Influence of quality of life, self-perception, and self-esteem on orthodontic treatment need.

In this study, we aimed to assess the relationship between normative and perceived orthodontic treatment need associated with quality of life, self-esteem, and self-perception. The sample included 248 schoolchildren aged 12years. The normative aspect of orthodontic treatment was assessed by the Dental Health Component and the Aesthetic Component of the Index of Orthodontic Treatment Need. The subjects were further evaluated for their oral health-related quality of life, self-esteem, and self-perception of oral esthetics. The Aesthetic Component of the Index of Orthodontic Treatment Need was considered as the response variable, and generalized linear models estimated by the GENMOD procedure (release 9.3, 2010; SAS Institute, Cary, NC). Model 1 was estimated with only the intercept, providing the basis for evaluating the reduction in variance in the other models studied; then the variables were tested sequentially, considering P≤0.05 as the criterion for remaining in the model. In the model, self-perception and self-esteem were statistically significant in relation to the perceived need for treatment. The normative need was significantly associated with the outcome variable and was not influenced by independent variables. The normative need for orthodontics treatment was not overestimated by the perceived need, and the perceived need was not influenced by sex and the impact on quality of life.

Preservation of honey bee spermatozoa using egg yolk and soybean lecithin-based semen extenders and a modified cryopreservation protocol

Better protocols and suitable extenders for semen handling and cryopreservation are required for honey bees to enhance sperm quality and fertility similar to that of other species. The purpose of the present research is the improvement of honey bee sperm quality using different semen extenders and a modified cryopreservation protocol. Three different extenders were used: (1) buffer and egg yolk (EY); (2) buffer and 0.5% soybean lecithin (SL0.5); and (3) buffer and 2% soybean lecithin (SL2). Semen was collected and then diluted with the experimental extenders. The diluted semen was gradually cooled in a refrigerator to 5 °C and immediately loaded into straws and frozen with liquid nitrogen. Motility and viability data analyzed using the GENMOD and GLM procedure of SAS software. The results demonstrate that the mean fresh sperm motility in EY (4.75 ± 0.14) and SL2 (4.5 ± 0.2) were significantly (p < 0.05) higher than the SL0.5 (4.12 ± 0.12). Also, the mean of cooled motile spermatozoa in EY and SL2 were significantly higher than the SL0.5 (3.87 ± 0.24). Post-thawed sperm motility in EY (3.6 ± 0.24) was significantly (p < 0.05) higher than the other extenders. Furthermore, percentage of viable spermatozoa in EY (69.75 ± 2.32%) was significantly higher than SL0.5 and SL2 (38.5 ± 2.32 and 45 ± 2.32% respectively. Therefore, according to the results of this study, use of an egg yolk-based extender could better maintain honey bee semen motility and viability after freeze-thawing process, compared to SL-based extenders. However, more advanced tests and fertility evaluation are needed to reveal the exact effects of egg yolk and lecithin based semen extenders on post-thawed drone sperm quality.

Sperm quality in naturally infected rams with Brucella ovis

Ovine epididymitis caused by Brucella ovis is a chronic disease, characterized by testicular and epididymal alterations, which cause loss of fertility in rams. However, infected rams without clinical signs, may have decreased fertility and be excreting the bacteria. The aim of this study was to evaluate sperm quality in serologically positive to B. ovis naturally infected rams. Seminal samples were obtained of 207 rams to evaluate seminal volume, sperm concentration, mass motility, vitality, presence of somatic cells and percentage of abnormalities. Clinical examination was performed by palpation. The serological response against B. ovis was evaluated by double immunodiffusion in gel. The percentage variables were analyzed by GENMOD procedure and numerical variables were analyzed by ANOVA and differences were assessed using Tukey's test. 18.84% of the rams were positive against B. ovis. There was no difference in scrotal circumference and volume (P>0.05) between positive and negative animals for B. ovis. Sperm concentration, mass motility, vitality, and the percentage of sperm with abnormalities, were modified in positive animals (P<0.05), regardless of the presence of palpable lesions (P<0.05). The percentage of rams with lesions in the epididymis and severe presence of somatic cells was higher in positive animals (P<0.05). B. ovis infection adversely affects semen characteristics, due to an increase in epididymal lesions and by the severe presence of inflammatory cells in the ejaculate. Serological analysis to determine possible infection by B. ovis, should be considered as a component of the evaluation of fertility in rams, considering that sperm quality was affected in rams without lesions, but seropositive for B. ovis.

Familiality of Co-existing ADHD and Tic Disorders: Evidence from a Large Sibling Study.

Background: The association of attention-deficit/hyperactivity disorder (ADHD) and tic disorder (TD) is frequent and clinically important. Very few and inconclusive attempts have been made to clarify if and how the combination of ADHD+TD runs in families.Aim: To determine the first time in a large-scale ADHD sample whether ADHD+TD increases the risk of ADHD+TD in siblings and, also the first time, if this is independent of their psychopathological vulnerability in general.Methods: The study is based on the International Multicenter ADHD Genetics (IMAGE) study. The present sub-sample of 2815 individuals included ADHD-index patients with co-existing TD (ADHD+TD, n = 262) and without TD (ADHD–TD, n = 947) as well as their 1606 full siblings (n = 358 of the ADHD+TD index patients and n = 1248 of the ADHD-TD index patients). We assessed psychopathological symptoms in index patients and siblings by using the Strength and Difficulties Questionnaire (SDQ) and the parent and teacher Conners' long version Rating Scales (CRS). For disorder classification the Parental Account of Childhood Symptoms (PACS-Interview) was applied in n = 271 children. Odds ratio with the GENMOD procedure (PROCGENMOD) was used to test if the risk for ADHD, TD, and ADHD+TD in siblings was associated with the related index patients' diagnoses. In order to get an estimate for specificity we compared the four groups for general psychopathological symptoms.Results: Co-existing ADHD+TD in index patients increased the risk of both comorbid ADHD+TD and TD in the siblings of these index patients. These effects did not extend to general psychopathology.Interpretation: Co-existence of ADHD+TD may segregate in families. The same holds true for TD (without ADHD). Hence, the segregation of TD (included in both groups) seems to be the determining factor, independent of further behavioral problems. This close relationship between ADHD and TD supports the clinical approach to carefully assess ADHD in any case of TD.

A Randomized Controlled Trial of a School-Implemented School–Home Intervention for Attention-Deficit/Hyperactivity Disorder Symptoms and Impairment

This study evaluated the efficacy of a novel psychosocial intervention (Collaborative Life Skills [CLS]) for primary-school students with attention-deficit/hyperactivity disorder (ADHD) symptoms. CLS is a 12-week program consisting of integrated school, parent, and student treatments delivered by school-based mental health providers. Using a cluster randomized design, CLS was compared with usual school/community services on psychopathology and functional outcomes. Schools within a large urban public school district were randomly assigned to CLS (12 schools) or usual services (11 schools). Approximately 6 students participated at each school (N= 135, mean age 8.4 years, grade range 2-5, 71% boys). Using PROC GENMOD (SAS 9.4), the difference between the means of CLS and usual services for each outcome at posttreatment was tested. To account for clustering effects by school, the generalized estimating equation method was used. Students from schools assigned to CLS compared with those assigned to usual services had significantly greater improvement on parent and teacher ratings of ADHD symptom severity and organizational functioning, teacher-rated academic performance, and parent ratings of oppositional defiant disorder symptoms and social/interpersonal skills. These results support the efficacy of CLS compared with typical school and community practices for decreasing ADHD and oppositional defiant disorder symptoms and improving key areas of functional impairment. They further suggest that existing school-based mental health resources can be redeployed from non-empirically supported practices to those with documented efficacy. This model holds promise for improving access to efficient evidence-based treatment for inattentive and disruptive behavior beyond the clinic setting. Clinical Trial Registration Information-Study of the Collaborative Life Skills Program; http://clinicaltrials.gov/; NCT01686724.

Bayesian logistic regression in detection of gene-steroid interaction for cancer at PDLIM5 locus.

The PDZ and LIM domain 5 (PDLIM5) gene may play a role in cancer, bipolar disorder, major depression, alcohol dependence and schizophrenia; however, little is known about the interaction effect of steroid and PDLIM5 gene on cancer. This study examined 47 single-nucleotide polymorphisms (SNPs) within the PDLIM5 gene in the Marshfield sample with 716 cancer patients (any diagnosed cancer, excluding minor skin cancer) and 2848 noncancer controls. Multiple logistic regression model in PLINK software was used to examine the association of each SNP with cancer. Bayesian logistic regression in PROC GENMOD in SAS statistical software, ver. 9.4 was used to detect gene- steroid interactions influencing cancer. Single marker analysis using PLINK identified 12 SNPs associated with cancer (P< 0.05); especially, SNP rs6532496 revealed the strongest association with cancer (P = 6.84 × 10⁻³); while the next best signal was rs951613 (P = 7.46 × 10⁻³). Classic logistic regression in PROC GENMOD showed that both rs6532496 and rs951613 revealed strong gene-steroid interaction effects (OR=2.18, 95% CI=1.31-3.63 with P = 2.9 × 10⁻³ for rs6532496 and OR=2.07, 95% CI=1.24-3.45 with P = 5.43 × 10⁻³ for rs951613, respectively). Results from Bayesian logistic regression showed stronger interaction effects (OR=2.26, 95% CI=1.2-3.38 for rs6532496 and OR=2.14, 95% CI=1.14-3.2 for rs951613, respectively). All the 12 SNPs associated with cancer revealed significant gene-steroid interaction effects (P < 0.05); whereas 13 SNPs showed gene-steroid interaction effects without main effect on cancer. SNP rs4634230 revealed the strongest gene-steroid interaction effect (OR=2.49, 95% CI=1.5-4.13 with P = 4.0 × 10⁻⁴ based on the classic logistic regression and OR=2.59, 95% CI=1.4-3.97 from Bayesian logistic regression; respectively). This study provides evidence of common genetic variants within the PDLIM5 gene and interactions between PLDIM5 gene polymorphisms and steroid use influencing cancer.

Lambing rate and prolificacy in inseminated hair sheep treated with bovine somatotropin.

This study evaluated whether the administration of 50 and 100 mg bovine somatotropin (bST) at the start of estrous synchronization and at the time of artificial insemination improves lambing rate and prolificacy in hair sheep. Four hundred eighty adult hair ewes (Pelibuey, Blackbelly, Dorper, Katahdin, and their crosses) were synchronized with intravaginal sponge containing 40 mg of fluorogestone acetate. On the day of sponge insertion, ewes were assigned to three treatments: the bST-100 treatment (n = 156) received 100 mg bST at the start of synchronization (d 0) and at the time of insemination (d 14), the bST-50 treatment (n = 159) received 50 mg bST in the same schedule as the previous group, and the control (n = 165) did not receive any bST. Lambing rate and percentage of multiple births were analyzed using the GENMOD procedure of SAS. Prolificacy data were analyzed using the MIXED procedure of SAS. The IGF-1 and insulin concentrations were analyzed with ANOVA for repeated measures. The bST application did not affect the lambing rate (P = 0.06). The proportion of ewes with multiple births (P = 0.01) and prolificacy (P = 0.04) were higher in the bST-50 (54.3% and 1.57 ± 0.1) than the bST-100 (18.2% and 1.25 ± 0.1) and control (33.3% and 1.28 ± 0.1) groups. The IGF-1 and insulin concentrations were higher (P < 0.05) in the bST-treated groups, but the insulin concentration was higher (P = 0.001) in the bST-100 group than in the bST-50 group. The administration of 50 or 100 mg bST at the start of synchronization and at the time of artificial insemination does not increase lambing rate. However, the dose of 50 mg increased the proportion of multiple births and prolificacy.

Twin Studies: A Unique Epidemiological Tool.

Twin studies are a special type of epidemiological studies designed to measure the contribution of genetics as opposed to the environment, to a given trait. Despite the facts that the classical twin studies are still being guided by assumptions made back in the 1920s and that the inherent limitation lies in the study design itself, the results suggested by earlier twin studies have often been confirmed by molecular genetic studies later. Use of twin registries and various innovative yet complex software packages such as the (SAS) and their extensions (e.g., SAS PROC GENMOD and SAS PROC PHREG) has increased the potential of this epidemiological tool toward contributing significantly to the field of genetics and other life sciences.

The effect of parturition induction treatment on interval to calving, calving ease, postpartum uterine health, and resumption of ovarian cyclicity in beef heifers

The aim of this study was to compare the effects of two parturition induction protocols with a nontreated control group, on interval to calving, calving ease, postpartum uterine health, and ovarian cyclicity in beef heifers. At Day 285 of gestation, 81 crossbred recipient beef heifers carrying purebred Simmental fetuses, were blocked by live-weight, body condition score, expected calving date and fetal sex, and assigned to one of three groups: (1) control (CON; no induction treatment, n = 29); (2) induction with corticosteroids (CORT; n = 27); or (3) induction with corticosteroids plus prostaglandin (CORT + PG; n = 25). Interval from induction to calving in hours and calving ease on a scale of 1 to 5 were recorded. Vaginal mucus samples were collected on Day 21 and Day 42 after calving (Day 0) by means of a Metricheck and scored on a scale of 0 to 3. Reproductive tract examinations were conducted on Day 21 and Day 42 after calving, and uterine cytology samples were obtained on Day 21. A positive cytologic sample was defined as greater than 18% neutrophils in the sample obtained via a cytobrush technique. Cows were considered to have resumed ovarian cyclicity if the presence of the CL was confirmed. Data were analyzed using the Mixed (normally distributed data) and Genmod (nonparametric data) procedures of SAS (v. 9.3). The interval from treatment to calving was longer (P < 0.0001) for CON (161.9 ± 15.12 hours) animals compared with CORT (39.7 ± 11.64 hours) or CORT + PG (32.6 ± 12.10 hours), which did not differ. Treatment did not affect calving difficulty score. There was also no difference in incidence of retained placenta between the three groups. At Day 21 postpartum, cytology score tended to be higher for both induced groups (48%) compared with the control animals (24%), but this was not the case for vaginal mucus score (CON 52%, CORT 70%, and CORT + PG 52%). A higher proportion of CON had an involuted uterus by Day 21 postpartum (69%) compared with both induced groups (CORT 48%, CORT + PG 32%). Day 21 ovarian cyclicity was higher in both CON (52%) and CORT (59%) compared with CORT + PG (29%). By Day 42, there was no difference in ovarian cyclicity or uterine involution between CON and CORT; however, a positive relationship was observed between uterine involution score on Day 21 and return to cyclicity on Day 42 in these two groups. There was a negative relationship between uterine involution score and return to cyclicity in the CORT + PG group, and these animals were slower (P < 0.05) to resume cyclicity by Day 42 with a larger proportion animals having evidence of having resumed postpartum ovarian cyclicity in both CON (P = 0.03) and CORT compared with CORT + PG on Day 42. In conclusion, the use of corticosteroid-based treatments is an effective strategy to advance parturition in full term dams and does not have a negative effect on calving progress or dam health. However, when prostaglandin is also included in the protocol, these treatments may lead to greater delay in uterine involution with increased chance of uterine infection and slower resumption of ovarian cyclicity.

Dynamics of Sus scrofa semen quality in the western-central region of Colombia

In order to characterize the dynamics of Sus scrofa (order Artiodactyla, family Suidae) semen quality in the Western-central region of Colombia, information of 1,017 assessments from 177 males from 12 swine farms located between 700 and 2,600 meters above sea level in different inter-tropical systems was collected in the Laboratory of Reproductive Biology at Universidad de Caldas (Colombia) during seven years. The database included the seminal variables: concentration, morphology (head, midpiece and tail), membrane structural integrity (MSI), acrosome integrity (ACRI), and membrane functional integrity (MFI) proximal citoplasmic droplets (PCD), distal citoplasmic droplets (DCD), and total citoplasmic droplets (TCD). Additionally, the racial group of the male (grac): maternal cross (cm), paternal cross (cp), paternal pure (pp), maternal pure (pm) and age group (agegr): young (up to 18 months), mature (19 to 36 months) and old (over 37 months) were registered. Data was explored by estimating descriptive statistics; it was analyzed using a log-linear model consisting of a Poisson regression with a log link function using the PROC GENMOD of SAS (SAS Inst. Cary, NC). The effect of age group and racial group on semen quality variables using a mixed model for repeated measurements in the same boar over time were evaluated. The Spearman correlation coefficients between variables were calculated. The variables: macro, micro and pyriform head; double, eccentric, and thickened midpiece; tail whip; ladder bow; ball and distal reflex showed less than 1% prevalence. The highest values of FMI were found in cm and pp. Males cp, cm and pm do not differ from each other in MFI, but all had higher values than pp. The ACRI in cm was higher than cp, and these higher than pm. Presentation of TCD increased with age, with a higher percentage of PCD in older animals than in younger animals; mature animals showed intermediate values between them. The TCD exceeded 15% of presentation, being always higher for PCD than for GCP. The TCD is negatively correlated with MSI and ARCI, but positively correlated with MFI. The same trend is observed in the correlations with distal cytoplasmic droplets. However, the correlation of proximal droplets with the structural integrity loses significance, though they are negatively correlated with acrosome integrity. Values for MSI, MFI and ACRI, do not limit the quality of the samples in this population.

69 AGE AT PUBERTY AND REPRODUCTIVE DEVELOPMENT OF LAMBS FED SUNFLOWER CAKE

The objective of this study was to evaluate the effects of lipid inclusion, represented by sunflower cake (SC), on the performance, age at puberty, ovarian follicular development, and metabolic panel of prepubertal lambs. Thirty-five crossbred Santa Ines lambs were used [initial body weight (BW) 21.5 ± 1.6 kg and age 138 ± 3 days]. Four SC inclusions were tested: 0% SC, n = 9; 15% SC, n = 9; 30% SC, n = 9; and 45% SC, n = 8, within the dry matter of the diet. The diets were offered to the lambs until they reached 43.6 ± 0.8 kg; after reaching this BW they were slaughtered. Dry matter intake and performance were analysed. The number of follicles, the diameter of the larger follicle, and the total cholesterol serum concentration were evaluated one day before slaughtering. The serum progesterone concentration was analysed every 6 days after the animals reached 30 kg. Puberty was determined when the progesterone concentration reached = 1 ng mL–1 for the first time. The animals were randomly confined in individual pens. To study the significant variable and its effects on the treatments, regression analysis was performed. The number of follicles visualised in the ovary through ultrasound and after slaughtering was analysed by Proc Genmod. Spearman correlation coefficient of the number of follicles obtained through the in vivo ultrasound technique and by the count of follicles in the ovary postmortem. SAS software was used (SAS Institute Inc., Cary, NC, USA). Daily dry matter intake (1.3 ± 0.5, 1.4 ± 0.5, 1.3 ± 0.5, and 1.3 ± 0.6 kg) and weight gain (0.19 ± 0.02, 0.23 ± 0.02, 0.19 ± 0.02, and 0.18 ± 0.02 kg) in the treatments of 0, 15, 30, and 45% of SC, respectively, presented no significant differences (P &gt; 0.05). Total cholesterol serum concentration linearly increased (y = 73.1784 + 0.7206x), (R2 = 0.8477; P &lt; 0.001) with the SC inclusion. The number of follicles &lt;3 mm was not correlated with pre- and post-slaughtering (r = –0.097; P &gt; 0.05), whereas the number of follicles = 3 mm of diameter was correlated (r = 0.56; P &lt; 0.001). The number of follicles = 3 mm (2.4 ± 0.1, 2.3 ± 0.1, 2.4 ± 0.1, and 2.2 ± 0.1), &gt;5 mm (0.5 ± 0.3, 0.7 ± 0.3, 0.8 ± 0.2, and 0.6 ± 0.3), and the diameter of the largest follicle (0.45 ± 0.03, 0.45 ± 0.04, 0.43 ± 0.03, and 0.40 ± 0.04 cm) verified through ultrasound, age at puberty (204 ± 11, 211 ± 13, 211 ± 13, 196 ± 11, 218 ± 11 days), and the weight at puberty (33.5 ± 1.5, 33.5 ± 1.7, 32.2 ± 1.5, and 33.5 ± 1.5 kg) in the treatments 0, 15, 30, and 45% SC, respectively, did not differ (P &gt; 0.05), although the progesterone concentration presented a linear increase (y = 1.4783 + 0.02826x; P &lt; 0.10). The number of follicles &lt;3 mm observed postmortem was influenced (y = 3.0687 + 0.0288*x – 0.0006*x2; P &lt; 0.001) by the levels of SC inclusion (i.e. a higher number of follicles were observed with the SC inclusion until 23.4% SC); follicles &gt; 3 mm (0.91 ± 0.2, 0.91 ± 0.2, 0.97 ± 0.2, and 0.88 ± 0.2) did not differ (P &gt; 0.05). In conclusion, the use of SC in the lamb’s diet positively affected the progesterone concentration and the development of follicles &lt;3 mm of diameter, which presented an optimal level at 23.4% SC inclusion.

82 EARLY PORCINE EMBRYO ENERGY PREFERENCE AND SUBSEQUENT DEVELOPMENT

Early porcine embryo metabolism in vitro is not completely understood. It has been suggested that before embryo genome activation (4-cell stage), the preferred energy source of the embryo is pyruvate. In our porcine zygote culture medium (MU1), the energy sources are 0.2 mM pyruvate and 2.0 mM calcium lactate. Three experiments were performed with in vitro-matured and IVF embryos to examine the effect on blastocyst development after withholding pyruvate and/or lactate during the first 48 h of culture. In Experiment 1, embryos were cultured without lactate for 48 and then cultured to Day 6 in control medium containing lactate. Control embryos were cultured in medium with lactate starting after fertilization to Day 6. All data were analysed by using SAS 9.3 with a GENMOD procedure used for the blastocyst data and a GLM procedure used for the cell number data. On Day 6, the percentage of embryos that formed blastocysts was 30.2% for control and 26.5% for embryos cultured for 48 h without lactate (n = 490, 4 replications). The difference was not significant P &gt; 0.05. In Experiment 2, embryos were cultured without pyruvate for 48 and then cultured to Day 6 in control medium containing pyruvate. Control embryos were cultured in medium with pyruvate starting after fertilization to Day 6. On Day 6, the percentage of embryos that formed blastocysts was 31.1% for control and 30.5% for embryos cultured for 48 h without pyruvate (n = 385, 3 replications). In Experiment 3, embryos were cultured in control medium for the first 48 h and then cultured to Day 6 in medium without pyruvate, thus forcing the embryos to use lactate instead of pyruvate. On Day 6, the percentage of embryos that formed blastocysts in the pyruvate free medium increased from 28.6%a ± 1.0 to 33.9%b ± 1.0; P ≤ 0.05 (n = 490, 4 replications) compared with the control and total cell number increased from 30.7a ± 1.5 to 41.3b ± 1.8 cells, respectively; P ≤ 0.05 (n = 65, 4 replications). The results from Experiments 2 and 3 were unanticipated as it was believed that the embryo would be more dependent on pyruvate for energy up to the blastocyst stage. We believed in Experiment 2 that from zygote to 4 cells the embryos were not as capable of using lactate and that removing the pyruvate would hinder further development. In Experiment 3, forcing the embryo to use lactate from Day 2 to Day 6 significantly improved blastocyst development and total cell number, suggesting that the embryo is not dependent on a specific energy source or that there are adequate pyruvate stores in the oocyte to 4-cell stage, to promote development to blastocyst. Funding was provided by Food for the 21st Century, the University of Missouri, and the National Institutes of Health (OD011140).

83 IN LOW OXYGEN CULTURE, IS HYPOTAURINE NECESSARY FOR IN VITRO DEVELOPMENT OF PORCINE EMBRYOS?

For decades it has been known that reactive oxidative species (ROS) form during in vitro embryo culture. A buildup of ROS can be detrimental to individual cells in the embryo and lead to a decrease in development and quality. To overcome oxidative stress in culture systems, additives, such as taurine and/or hypotaurine, have been used. In the pig, taurine or hypotaurine addition is deemed necessary for normal in vitro development. Another commonly used technique to reduce ROS is to culture embryos in a lowered oxygen environment (e.g. 5%). Porcine zygote medium 3 (PZM3) base culture medium is used in the following experiments and contains 5 mM hypotaurine, which is one of the most costly additives in the medium. The objective of this experiment was to determine if hypotaurine is still necessary if the embryos were cultured in 5% O2 from the zygote to the Day 6 blastocyst stage. In Experiment 1, oocytes were matured for 44 h and fertilized in vitro. After fertilization, presumptive zygotes were then transferred to 500 µL of MU-1 medium (PZM3 with 1.69 mM arginine) that either contained or did not contain hypotaurine for overnight culture at 20% O2. On Day 1, the same embryo culture plates were moved to 5% O2, 5% CO2, and 90% N2 and cultured to Day 6. The percent blastocyst stage was determined, and total cell number was counted in 3 of the 5 replicates in order to give us an indication of the embryo quality. The percent blastocyst in the controls (+hypotaurine) was 34.4% ± 2.8 and not different from the no hypotaurine (32.9% ± 2.2; N = 830; 5 replications; P &gt; 0.10). Furthermore, total cell number was not different between the two groups (30.8 ± 1.5 v. 33.6 ± 1.8, respectively, N = 146; 3 replications; P &gt; 0.10). In Experiment 2, the same experiment was repeated in somatic cell nuclear transfer derived embryos, which may be more sensitive to ROS due to the micromanipulation procedure. Wild type fetal fibroblast cells were used as donor cells. There was no significant difference in development to the blastocyst stage due to the presence or absence of hypotaurine (17.7% ± 2.5 v. 11.8% ± 2.3, respectively; N = 454; 4 replications; P = 0.07). All blastocyst data were analysed using the GENMOD procedure in SAS 9.4 (SAS Institute Inc., Cary, NC, USA), and cell number data were analysed using the PROC GLM also with SAS 9.4. These data show that porcine embryos can be efficiently cultured to the blastocyst stage without adding any oxygen free radical scavengers to the media when culturing in reduced oxygen atmosphere. Further studies include evaluating term development via embryo transfers and measuring ROS production of these embryos. Funding was provided by Food for the 21st Century and the National Institutes of Health (U42 OD011140).

157 EFFECT OF MELATONIN ON EMBRYO QUALITY OF BOVINE OOCYTES SUBJECTED TO HEAT SHOCK

The aim of this study was to evaluate the effect of different concentrations of melatonin added to in vitro maturation (IVM) medium of oocytes subjected to heat shock on embryo quality. Immature oocytes aspirated from ovaries obtained from a slaughterhouse were selected and randomly allocated in factorial experiment design (3 × 2). Three concentrations of melatonin (0, 10–6, and 10–4 M; M5250, Sigma, St. Louis, MO, USA) were added to the IVM medium and 2 incubation conditions (conventional: 24 h at 38.5°C and 5% CO2; heat shock: 12 h at 41°C followed by 12 h at 38.5°C and 5% CO2) were tested, resulting in treatments: M1 (0 M; 38.5°C; n = 15), M2 (10–6 M; 38.5°C; n = 15), M3 (10–4 M; 38.5°C; n = 15), M4 (0 M; 41°C; n = 15), M5 (10–6 M; 41°C; n = 15), and M6 (10–4 M; 41°C; n = 15). The IVM was performed in a Nunc plate (144444 – Thermo, Fisher Scientific Inc., Pittsburgh, PA, USA) containing 400 µL of TCM-199 (Invitrogen, Carlsbad, CA, USA) supplemented with 20 µg mL–1 of FSH (Pluset®, Calier Laboratories, Barcelona, Spain) and 10% oestrus cow serum. Oocytes were IVF in FERT-TALP medium for 20–22 h and incubated at 38.5°C and 5% CO2. After IVF, the presumptive zygotes were denuded and cultured in CR2aa medium supplemented with 2.5% FCS (Nutricell, Campinas, Brazil) in an incubator at 38.5°C under 5% CO2, 5% O2, and 90% N2, and saturated humidity for 8 days. Blastocysts with 8 days post-fertilization from different treatments were fixed in 4% paraformaldehyde in PBS for 1 h and analysed by TUNEL assay (Deadend™ Fluorometric TUNEL System, Promega, Madison, WI, USA) to evaluate embryonic quality. Data were analysed by generalised linear models considering the Poisson distribution and using the Proc Genmod of SAS software (version 9.1; SAS Institute Inc., Cary, NC, USA) considering effects of melatonin concentration, incubation conditions, and interaction between the factors. Values shown are the mean ± s.e.m. The interaction between melatonin concentration and incubation conditions was no significant (P &gt; 0.05). The total number of cells was not affected (P &gt; 0.05) by melatonin, but it was decreased (P &lt; 0.05) by heat shock (M1 = 117 ± 6.7a; M2 = 118 ± 4.2a; M3 = 120 ± 6.3a; M4 = 102 ± 6.2b; M5 = 106 ± 5.7b; M6 = 108 ± 8.9b). Melatonin and heat shock did not affect (P &gt; 0.05) the index of embryo apoptotic cells (M1 = 15.3% ± 2.0; M2 = 15.5% ± 1.3; M3 = 13.6% ± 2.0; M4 = 14.9% ± 1.5; M5 = 13.3% ± 1.3; M6 = 13.5% ± 1.2) and the index of trophoblast cells (M1 = 74.6% ± 2.3; M2 = 75.0% ± 1.7; M3 = 75.2% ± 1.9; M4 = 78.4% ± 2.3; M5 = 76.4% ± 3.0; M6 = 75.2% ± 2.6). The melatonin and heat shock affected the index of the inner cell mass (ICM; P &lt; 0.05), and the heat shock reduced the index of the ICM of oocytes not treated with melatonin (M1 = 25.4% ± 2.3a; M2 = 25.0% ± 1.7a; M3 = 24.8% ± 1.8a; M4 = 21.6% ± 2.3b; M5 = 23.6% ± 3.0a; M6 = 24.8% ± 2.6a). In conclusion, melatonin supplementation to the medium IVM of oocytes subjected to heat shock had no effect on blastocyst total cell number, general apoptotic index, or index of the trophoblast cells, but increased index of the ICM. Research was supported by Fapemig, CNPq, Embrapa, and CAPES.

The relationship between growth hormone polymorphism and growth hormone receptor genes with milk yield and reproductive performance in Holstein dairy cows.

The aim of this study was to investigate the potential association between growth hormone GH/AluI and growth hormone receptor GHR/AluI polymorphisms with milk yield and reproductive performances in Holstein dairy cows in Iran. Blood samples of 150 Holstein cows were collected and their genomic DNA was extracted using Gene-Fanavaran DNA extracting kit. Fragments of the 428 bp of exon 5 growth hormone (GH) gene and the 342 bp of exon 10 growth hormone receptor (GHR) gene were amplified using the polymerase chain reaction (PCR) method. PCR products were digested by the AluI restriction enzyme and electrophoresed on 3% agarose gel. Continuous and categorical data were analyzed using linear mixed models through Proc MIXED and logistic regression models through Proc GENMOD of SAS software, respectively. The results showed no relationship between the examined traits and GH/AluI or GHR/AluI genes. A significant relationship was found between GH/AluI polymorphism and dystocia, but the presence of the GH-L allele reduced the incidence of dystocia. The results suggest that the GH-LL genotype reduces dystocia probably by affecting the release of growth hormone; nevertheless, further studies will be needed to examine the relationship between dystocia and GH genotypes.

&lt;b&gt;Reproduction performance of female Nile tilapia under different environments and age classes

Current assay verifies the reproductive performance of females of Nile tilapia in different age classes and reproductive environments. Four hundred and fifty-one females were evaluated in two different Brazilian environments, namely in Maringa, Parana State, Brazil (Site 1 - hapas of 10 m3 and Site 2 - hapas of 1 m3) and in one environment in Alfenas, Minas Gerais State, Brazil (Site 3 - hapas of 14 and 28 m3). The females were divided into three age classes, corresponding to animals under one year (age 1); between one to two years (age 2); and over three years old (age 3). Spawning was verified once a week, from October 2012 to March 2013, corresponding to a 23-week assessment. The statistical analyses were performed with Proc Genmod from the statistical program SAS. Site 2 had the lowest results when compared to the other sites for multiple spawning, spawning frequency and volume of eggs/female for animals of age 2. Statistical differences were not detected between ages 1 and 2 for the same assessment site, although females at age 3 in Site 3 provided higher results than those at age 2. Results show that, precluding the use of animals in individual hapas (1 m³) and maintaining older females in the stock, may improve reproduction results.