Last Stage Of Gestation Research Articles

Molecular detection of Ovine listeric abortion in Nineveh governorate, Iraq

Listeriosis is a important abortive diseases in sheep , and is considered one of the most risky bacterial zoonotic disease worldwide.The study was carried out in 50 sheep flocks placed in the Nineveh governorate, Iraq during November and December 2022, a total of 300 specimens of blood , abomasum content and brain ( 100 each)were collected from local breed ovine aborted fetues(in the last stage of gestation) were obtained to be tested for molecular detection of Listeria monocytogenes . L monocytogenes DNA was detected in a total of 61(20.3%) specimens , distributed as: 35(57.4%) ,15(24.6% and 11(18.0%) strains from fetal brain, abomasum and blood specimens respectively using direct genus-specific conventional polymerase chain reaction (prfA gene) C- PCR. Two L. monocytogenes strains ( HMB1 listeriolysin, HMB 2 listeriolysin ) deposited in GenBank under accession numbers LC769365.1,and LC769366.1. All L. monocytogenes strains were positive for three genes (InlJ, InlA, and hlyA) except actA gene was detected in 46 (75.4%) strains. In conclusion , L. monocytogenes is one of the important causal agents of abortion in sheep flocks in Nineveh province , Iraq, and most of the brain specimens were positive for listeric infection compared to the other specimens.

The morphofunctional state of pregnant sheep in the last stages of gestation is normal and with symptoms of eclampsia

Sheep in the last stages of pregnancy showed signs of eclampsia, such as proteinuria, hypertension, swelling of the abdominal wall, and coma. Based on the diagnoses, two groups of 20 sheep with eclampsia were created: typical and atypical signs, in each, and one group of clinically healthy animals. From each group, 5 animals were slaughtered. In sheep suffering from eclampsia, a decrease in the total mass of the placenta and its cotyledons, as well as an increase in the length of the umbilical cord, was also found. Standard histological and histochemical studies were performed on the obtained placental samples. As a result, it was found that the average thickness of the integumentary epithelium of the caruncles in ewes with a typical form of eclampsia was 14.3 ± 1.7 μm, and in sheep with an atypical form - 15.5 ± 1.9 μm, compared with 27.9 ± 1 .9 µm in the physiological course of gestation. This leads to a general decrease in the thickness of the layers of connective tissue separating the crypt and villi of the cotyledons. In addition, the cytoplasm of cells exhibits dystrophic modifications along with a decrease in the ability to store glycogen. Dystrophic changes are present in the cytoplasm of cells, which contributes to a decrease in glycogen storage.

Heterologous Challenge with PRRSV-1 MLV in Pregnant Vaccinated Gilts: Potential Risk on Health and Immunity of Piglets.

Simple SummaryModified live virus (MLV) vaccines are considered as the key component to control the Porcine Reproductive and Respiratory Syndrome Virus (PRRSV). The majority of pig farms apply the ‘mass’ vaccination strategy in breeding female animals. However, this PRRS MLV vaccination protocol involves the risk of inoculation of sows in the last stage of gestation, resulting in possible infection of the fetus as the virus can efficiently cross the placenta during the last period of pregnancy. Thus, we evaluated the ability of the vaccine virus to act as a pathogenic strain, to be transmitted to fetuses and to affect the health status of neonatal piglets. The results indicated that the study gilts transmitted the vaccine virus to their offspring, as well as that the PRRSV-infected piglets showed a poor clinical performance. Consequently, the pig farms that apply PRRS MLV vaccination in a routine blanket vaccination strategy must avoid inoculating pregnant gilts the last week before their parturition.The objective of the present study was to evaluate the potential risks of the four commercial PRRS-1 MLV vaccines in pregnant vaccinated gilts at the last stage of gestation under field conditions. The study was conducted at four pig farms, including 25 gilts from each farm (25 × 4 = 100 gilts), which were equally allocated to five different study groups. A PRRS-1 MLV vaccination was applied on the 100th day of their pregnancy with the different commercial vaccines that are available in the Greek market. The results indicated virus congenital infection and viremia in piglets (20/200 = 10% PRRSV infected piglets), and detection of PRRSV-specific antibodies (181/200 = 90.5% piglets found with PRRSV antibodies). The subsequent phylogenetic analyses revealed high percentages of similarity between the PRRSV-1 strain detected in infected litters and the PRRSV-1 vaccine strain to which the study gilts had been previously exposed to. Health status analyses of trial piglets resulted in differences between litters from vaccinated sows and litters from non-vaccinated sows at 110th day of gestation as regards the number of weak-born piglets, mummies, and piglets with splay-leg and/or respiratory symptoms. The current study’s results indicate several potential dangers of the PRRS MLV vaccination in late gestation.

Occurrence and Associated Risk Factors of Porcine Reproductive and Respiratory Syndrome Virus and Porcine Circovirus Type 2 Infections in Greece.

The objective of the present study was to identify factors associated with the probability of being polymerase chain reaction (PCR) positive and the level of porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus type 2 (PCV2) occurrence in Greek farms. The study included 56 pig farms, with a total population of 22.500 sows, which represent about 40% of the entire capacity of the Greek swine production. A total of 896 blood samples (224 pools*4 samples/pool = 896 samples) from breeding stock, weaners, growers, and finishers were collected from each farm and organized in pools of 4 samples size. Further, data regarding herd health management protocols were collected. The sera were tested for PRRSV and PCV2, using real-time PCR (RT-PCR). The results indicated that both viruses remain a major challenge for the Greek swine industry. Main risk factors involved in the infection process by these viruses were identified. In particular, vaccination programs such as the mass PRRSV vaccination with modified-live virus (MLV) in breeding stock during the last stages of gestation or with killed-virus (KV) during the middle of gestation are more likely to be associated with PRRSV PCR-positivity. Farms with low biosecurity level are associated with higher PRRSV circulation. It has also been revealed that breeding stock is more likely to be associated with PCV2 circulation compared to weaners and growers. In conclusion, our results could be the basis of the development of surveillance protocols for a national monitoring system for PRRSV and PCV2, which could prevent future infection of Greek farms.

LOCALIZATION OF EPITHELIAL AND MESENCHYMAL ELEMENTS IN THE PLACENTAL STRUCTURES OF COWS IN THE NORM AND DURING THE DEVELOPMENT OF THE STATE OF ECLAMPSIAS

The results of pathomorphological and morphometric studies showed that the histostructure of the normal placenta in cows in the last stages of gestation was characterized by the following features: - free location of the chorionic villi in the crypts formed by the branches of the uterine septa; - the composition of the intercellular substance of the uterine septa includes neutral mucopolysaccharides, chorionic villi - acid mucopolysaccharides. Histological examination of the placenta of cows during pregnancy complicated by preeclampsia revealed certain patterns in the structure of the organ, which can be taken into account in differential diagnosis: - a decrease in the mass of fetal membranes, as well as a decrease in the number and surface area of cotyledons; - hyperplasia of the crypts with simultaneous reduction of the villi, the crypts merged with each other, forming structures resembling the endometrium in structure; - vacuolar degeneration of chorionic villi in the fetal part of the placenta; - premature degradation of the extracellular matrix, not associated with labor activity.

Oxidative stress in suyagny sheep at the end of gestation, as a factor in pathogenesis development of eclampsia

Analysis of the results of medical examination of pregnant sheep 30...15...5 days before the expected term of lambing showed that in the urine of 12.11% of ewes, the concentration of ketone bodies in the blood plasma was 1.43 times higher than the physiological norm. In addition, there is a tendency to decrease the buffer bases to 18.41±1.53 mmol/l, as well as the glucose content below 2.12 ± 0.12 mmol/l. At the same time, the ratio of VN/ ACAS is up to 1.47 ± 0.12 mmol / L. It was found that the albumin fraction of proteins decreased by 1.51 times. Edema in the abdominal wall and underbelly, 9.5% lack of reaction to external stimuli, 15.0% decreased appetite, 10.0% jaundice of mucous membranes and 9.0% of sheep had a comatose state were found in 15.0% of animals. In a clinical study in sheep with eclampsia, hypertension over 136.1 mmHg and the presence of protein in urine over 3.2 g/l were recorded. These symptoms indicated classic signs of eclampsia, which occurred depending on the condition of sick animals in atypical and typical forms of manifestation. In animals with typical clinical signs of eclampsia, an increase in the level of diene conjugates was found to be 1.87 times, and the concentration of ketodiene intermediates and conjugated trienes was 1.75 times, the concentration of stable nitric oxide metabolites was 38.0%, and the vitamin E content decreased by 13.1%. At the same time, there was a 20.46% increase in the concentration of double bonds in the atypical form of eclampsia. In the blood plasma of patients with eclampsia of pregnant ewes, the activity of superoxide dismutase (1,736±0.37 units) is lower than in the comparison group (2,146±0.56 mmol/l). The obtained material of this work should be taken into account in the future when studying the problem of complicated pregnancy in the last stages of gestation in sheep and when developing therapeutic and preventive measures to ensure the long-term functioning of the reproductive potential and obtaining a viable newborn offspring.

A computer vision-based approach for behavior recognition of gestating sows fed different fiber levels during high ambient temperature.

The objectives of this study were to evaluate convolutional neural network models and computer vision techniques for the classification of swine posture with high accuracy and to use the derived result in the investigation of the effect of dietary fiber level on the behavioral characteristics of the pregnant sow under low and high ambient temperatures during the last stage of gestation. A total of 27 crossbred sows (Yorkshire × Landrace; average body weight, 192.2 ± 4.8 kg) were assigned to three treatments in a randomized complete block design during the last stage of gestation (days 90 to 114). The sows in group 1 were fed a 3% fiber diet under neutral ambient temperature; the sows in group 2 were fed a diet with 3% fiber under high ambient temperature (HT); the sows in group 3 were fed a 6% fiber diet under HT. Eight popular deep learning-based feature extraction frameworks (DenseNet121, DenseNet201, InceptionResNetV2, InceptionV3, MobileNet, VGG16, VGG19, and Xception) used for automatic swine posture classification were selected and compared using the swine posture image dataset that was constructed under real swine farm conditions. The neural network models showed excellent performance on previously unseen data (ability to generalize). The DenseNet121 feature extractor achieved the best performance with 99.83% accuracy, and both DenseNet201 and MobileNet showed an accuracy of 99.77% for the classification of the image dataset. The behavior of sows classified by the DenseNet121 feature extractor showed that the HT in our study reduced (p < 0.05) the standing behavior of sows and also has a tendency to increase (p = 0.082) lying behavior. High dietary fiber treatment tended to increase (p = 0.064) lying and decrease (p < 0.05) the standing behavior of sows, but there was no change in sitting under HT conditions.

МЕТАБОЛИЧЕСКИЕ ИЗМЕНЕНИЯ В ОРГАНИЗМЕ ОВЕЦ НА ПОСЛЕДНИХ СРОКАХ ГЕСТАЦИИ ПРИ ОСЛОЖНЕНИИ СУЯГНОСТИ ЭКЛАМПСИЕЙ

During the prophylactic examination at the last stages of gestation, 27.691.79% of pregnant ewes were diagnosed with eclampsia of varying severity: arterial hypertension (BP 136.1 ± 2.85 mm Hg), proteinuria (protein content in urine more than 3 0±0.49 g/l), edema and coma. In urine, an increase in the concentration of ketone bodies above the physiological norm by 2.3 times, their fractions of AcAc and BH - by 5.9 and 1.5 times, respectively, was noted. In the blood, a decrease in buffer bases to 18.41 ± 1.53 mmol/l, a glucose concentration to 2.12 ± 0.12 mmol/l, a BH/AcAc coefficient to 1.47 ± 0.12 mmol/l were established. There was an increase in the concentration of the intermediate product of lipid peroxidation in the blood by 43.0%, the concentration of stable nitric oxide metabolites - by 38.0%, vitamin C - by 24.1%. The content of vitamin E decreased by 13.1%, and the concentration of double bonds increased by 20.46% with an atypical form of eclampsia and by 34.13% with the appearance of a typical form of eclampsia. The level of diene conjugates increased by 1.87 times. The concentration of intermediate products of ketodienes and conjugated trienes is increased by 1.75 times in comparison with the atypical form of the course of eclampsia and by 3.54 times with its typical course. In pregnant ewes, the activity of oxidized glutathione is increased (2.879 ± 0.32 µmol/l), and superoxide dismutase (1.736±0.37 arb. units) is lower than in the comparison group (2.146±0.56 µmol/l). Consequently, the metabolic parameters that are traditionally used in the diagnostic algorithm in pregnant sheep with metabolic stress with symptoms of eclampsia in some cases are less sensitive and specific than the indicators of the system "lipid peroxidation - antioxidant protection". In the future, the material obtained in this work should be taken into account when studying the problem of eclampsia in pregnant sheep, as a concept for the development of impaired functioning of the lipid peroxidation-antioxidant protection system in this metabolic pathology.

Trophic ecology of Mexican Pacific harbor seal colonies using carbon and nitrogen stable isotopes.

There is limited information that provides a comprehensive understanding of the trophic ecology of Mexican Pacific harbor seal (Phoca vitulina richardii) colonies. While scat analysis has been used to determine the diet of some colonies, the integrative characterization of its feeding habits on broader temporal and spatial scales remains limited. We examined potential feeding grounds, trophic niche width, and overlap, and inferred the degree of dietary specialization using stable carbon and nitrogen isotope ratios (δ13C and δ15N) in this subspecies. We analyzed δ13C and δ15N on fur samples from pups collected at five sites along the western coast of the Baja California Peninsula, Mexico. Fur of natal coat of Pacific harbor seal pups begins to grow during the seventh month in utero until the last stage of gestation. Therefore pup fur is a good proxy for the mother’s feeding habits in winter (~December to March), based on the timing of gestation for the subspecies in this region. Our results indicated that the δ13C and δ15N values differed significantly among sampling sites, with the highest mean δ15N value occurring at the southernmost site, reflecting a well-characterized north to south latitudinal 15N-enrichment in the food web. The tendency identified in δ13C values, in which the northern colonies showed the most enriched values, suggests nearshore and benthic-demersal feeding habits. A low variance in δ13C and δ15N values for each colony (<1‰) and relatively small standard ellipse areas suggest a specialized foraging behavior in adult female Pacific harbor seals in Mexican waters.

Evaluation of efficacy of a biofilm-embedded bacteria-based vaccine against staphylococcal mastitis in sheep—A randomized, placebo-controlled field study

Our objective was to evaluate the efficacy of a vaccine against staphylococcal mastitis in 5 dairy sheep farms, with 316 ewes in the vaccinated (V) group and 307 in the control (C) group studied throughout a lactation period. Two administrations of the vaccine were performed during the last stage of gestation of ewes. Starting 15 d after lambing and at monthly intervals thereafter, up to 9 milk samplings were performed for bacteriological and cytological examinations. Staphylococcal isolates recovered were examined for biofilm formation. Blood samples were collected for measurement of IgG poly-N-acetylglucosamine-specific antibodies. The most frequently isolated bacteria were staphylococci: 56.4 and 76.1%, respectively, of total isolates recovered from ewes of group V and C, respectively; staphylococci as causal agents of mastitis were isolated less frequently from V (5.3%) than in ewes in C (10.3%). Among mastitis-associated staphylococcal isolates recovered from V ewes, a smaller proportion was biofilm-forming than among ones from C: 53.2% versus 74.9% of isolates; biofilm-forming staphylococci as causal agents of mastitis were isolated less frequently from ewes in group V (2.3%) than in ewes in group C (6.0%). Anti-poly-N-acetylglucosamine-specific antibody values increased in V ewes and were higher than in C; a greater proportion of ewes with low antibody titers developed staphylococcal mastitis (41.4%) than of V ewes with high antibody titers (17.0%). Incidence risk of mastitis, staphylococcal mastitis, and biofilm-associated staphylococcal mastitis was smaller in V than in C: 36.7, 17.1, and 8.0% versus 44.3, 30.9, and 18.9%, respectively. The first case of staphylococcal mastitis occurred later in V than in C: third versus second sampling point. Overall, efficacy of the vaccine was 44.6% for staphylococcal mastitis, 57.7% for biofilm-associated staphylococcal mastitis, 33.1% for staphylococcal intramammary infection, and 51.5% for biofilm-associated staphylococcal intramammary infection. Nevertheless, vaccination should not be the only means for controlling mastitis; other udder health management measures should be included therein to improve control of the infection.

Influence of maternal infection caused by Klebsialle pneumonia on morphologic and functional state of fetuses’ adrenal glands cortex (experimental research).

Background. Infections of fetuses and newborns caused by Klebsiella themselves and in polymicrobial associations have short incubation period, as well as a high percentage of deaths. Intrauterine infection of fetus and fetal chronic hypoxia should be considered as conditions that require a lot of adaptive changes where one of the most important roles is performed by adrenal glands. Objective: to identify the influence of mother infection caused by K.pneumoniae on fetal adrenal cortex. Methods: During the experiment, laboratory rats were divided into 3 groups. Group 1 - control - pregnant rats lived in physiological conditions. Group 2 - modeling of chronic intrauterine hypoxia - pregnant rats throughout pregnancy (21 days) were subjected to a daily high altitude hypoxia. Group 3 – modeling of infection – using the infective dose of Klebsiella pneumonia was modeled. Then male were tucked to infected female, females got pregnant. All rats were removed from the experiment on the last stages of gestation in order to extract fetus for further study. The adrenal glands of fetuses were examined using histological, morphometric and immunohistochemical methods. Results. Infectious pathology of mother has a more detrimental effect on fetal adrenal glands, than "pure" chronic intrauterine hypoxia. In group 3 hypoplasia is more pronounced in zona glomeruloza. Severe lesions of cytolysis and resorption of cell with increasing of cortisol production are seen in zone fasciculata. In fetal zone there are depletion of functional activity, zone is decreasing and has a tendency to decrease hormone production. Conclusion: when mother is infected by Klebsiella pneumonia in constant and fetal adrenal cortical areas of fetus changes manifested by hypoplasia of zona glomeruloza, hyperactivity of zona fasciculata and depletion of functional activity in fetal zone.

Possibilities of antenatal prevention of food allergy in young children

Objective: to determine the efficiency of antenatal prevention of food allergy in children. A total of 248 mother-child pairs were followed up. According to their maternal feeding pattern during pregnancy, the newborn infants were divided into 3 subgroups. Subgroup 1 consisted of 37 infants whose mothers had ingested a hypoallergenic diet during pregnancy. Subgroup 2 entered 29 babies, whose mothers had adhered to a hypoallergenic diet and received the probiotic Lactobacillus reuteri Protectis in the last stages of gestation. Subgroup 3 comprised 82 infants whose mothers had received a diet with a hypoallergenic substitute of cow’s milk for the New Zealand goat’s milk Amalthea in combination with the probiotic during pregnancy. There were no differences between the neonatal groups in anthropometric indicators. Analysis of intestinal microbiota showed that significantly more children had an increasing amount of Klebsiella, enterococci, Candida albicans, and Staphylococcus aureus in the comparison group (p&lt;0.05). Subgroup 3 babies whose mother had received lactobacilli and the whole goat’s milk Amalthea were more frequently found to have normal levels of bifidobacteria and lactobacilli (p&lt;0.05) and a smaller number of opportunistic pathogens (p&lt;0.05) in the enteric flora. In addition, the hypoallergenic diet including goat’s milk and/or Lactobacillus reuteri Pro￾tectis in mothers in the last stages of gestation reduced the frequency and degree of sensitization to cow’s and goat’s milk in the infants.

Gene expression profiles of estrogen receptors α and β in the fetal bovine hypothalamus and immunohistochemical characterization during development.

Steroid hormones intervene in the structural and functional regulation of neuronal processes during development and thus determine brain differentiation. The effects of estrogens are mediated by two transcription factors, namely estrogen receptor α (ER-α) and estrogen receptor β (ER-β), that regulate the expression of target genes through their binding to specific DNA target sequences. We describe the mRNA expression of ER-α and ER-β in the hypothalamus of developing male and female bovines as revealed by quantitative real-time polymerase chain reaction, and the distribution of the two ERs in hypothalamic sections of all fetal stages as shown by immunohistochemistry. The expression profiles of the mRNAs of both ERs are mutually correlated throughout the gestation period, and their levels increase significantly in the last stages of gestation. No sexual differences in the mRNA expression of either ER-α or ER-β have been found in our fetal specimens. The use of specific antisera against ER-α and ER-β has allowed us to characterize and confirm the distribution of these receptors in the hypothalami of all fetal stages considered. Our results offer detailed information concerning the distribution of ER-α and ER-β in the developing bovine hypothalamus and provide additional insights into the processes involved in the hypothalamic development of a mammal with a long gestation and a highly gyrencephalic brain.

Transcriptomics of the late gestation ovine fetal brain: modeling the co-expression of immune marker genes.

BackgroundMajor changes in gene expression occur in the fetal brain to modulate the function of this organ postnatally. Thus, factors can alter the genomics of the fetal brain, predisposing to neurological disorders later in life. We hypothesized that the physiological dynamics of the immune system transcriptome of the fetal brain during the last stage of gestation will reveal patterns of immune function and development in the developing brain. In this study we applied weighted gene co-expression analysis of microarrays performed on ovine fetal brain samples, to model the changes in gene expression throughout the second half of gestation.ResultsClusters of co-expressed genes that strongly increase in expression toward the first day of extra-uterine life are related to the hematopoietic lineage, while activation of immune pathways is induced after birth. Moreover, the pattern of gene expression suggests induction of tolerance mechanisms, probably necessary to protect highly produced proteins –such as myelin basic protein- from an autoimmune attack.ConclusionsThis study provides insight into the dramatic changes in gene expression that take place in the brain during the fetal life, especially during the last stage of gestation, and suggests that the immune system may have an important role in maturation of the fetal brain, which if disrupted or altered, could have negative consequences in postnatal life.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-1001) contains supplementary material, which is available to authorized users.

The effect of dietary organic and inorganic selenium supplementation on serum Se, Cu, Fe and Zn status during the late pregnancy in Merghoz goats and their kids

The present study aims to determine the effect of dietary Se supplement on Merghoz goats at the last stage of gestation on serum Se, Cu, Zn and Fe status of goats and their kids. Thirty Merghoz does (30±3kg) were fed the same basal diet containing 0.11mgSekg−1 DM and randomly assigned to one of three dietary treatments in a randomized complete block design. Three weeks before the expected kidding, the basal diet was supplemented with 0 (control) or 0.3mgSehead−1d−1 in the form of l-selenomethionin (Se-Met) and 0.3mgSehead−1d−1 in the form of sodium selenite (SS). Blood samples were collected from the does three weeks before the expected kidding and on the kidding day. Blood samples of newborn kids were also taken from the jugular vein at birth and 7 days of age. The results indicated that the serum concentration of Se increased in supplemented does compared with the control ones. The selenium concentration was also significantly increased in colostrum of treated does (P<0.05). There was no significant difference between Se sources on the Se status although there was higher efficiency of Se-Met than sodium selenite supplement. The serum concentration of copper increased in supplemented doses, which was concomitant with a reduction in Zn concentration in serum and colostrums. It seems that an increase in serum concentration of Cu at the end of pregnancy in goats given Se supplement could disturb the Zn:Cu ratios which in turn lead to Zn reduction. Serum Se, Cu and Fe concentrations of kids of treated doses increased at birth and first week of age, while the serum concentration of Zn decreased in newborn kids. It seemed that there was a positive correlation between serum Se concentration with Cu and Fe levels and also a negative correlation between Se and Zn in treated dose (P<0.05).

Effect of dietary vitamins E and C supplementation on performance of sows and piglets

The aim of this study was to evaluate the effect of increased vitamins E and C doses in the feeding of sows at the last stage of gestation (90 day) and during lactation on the performance of sows and their piglets. The study was carried out in two seasons (winter and summer) on 139 multiparous sows, divided into three groups: control diet – vitamin E 60 mg/kg, experimental diet – vitamin E 200 mg/kg, experimental diet – vitamin E 200 mg/kg + vitamin C 500 mg/kg. In spite of an increased concentration of these vitamins in the serum of piglets, no clear effect on the rearing results was found. However, an advantageous post-effect of the vitamins fed to sows was shown. The addition of vitamin E together with vitamin C significantly reduced the body temperature of sows after farrowing and considerably reduced the number of sows culled after rearing.

Porcine reproductive and respiratory syndrome virus (PRRSV) causes apoptosis during its replication in fetal implantation sites

Reproductive failure due to porcine reproductive and respiratory syndrome virus (PRRSV) is characterized by late-term abortions, early farrowing and an increase of dead and mummified fetuses and weak-born piglets. The mechanism of PRRSV-induced reproductive failure is poorly understood. Human pregnancies, complicated by some pathogens leading to reproductive disorders exhibit increased apoptosis in the fetal membranes. Because PRRSV-target cells are present in endometrium/fetal placentas from healthy sows and PRRSV-infected macrophages in other organs die by apoptosis, we hypothesized that PRRSV can replicate and induce apoptosis in the fetal implantation sites at the last stage of gestation. In the present study, identification, localization and quantification of the PRRSV-positive and apoptotic cells were performed in the fetal implantation sites. Three dams were inoculated intranasally with 10 5 TCID 50 PRRSV 07V063 at 90 days of gestation and sampled at 10 days post-inoculation. Two non-inoculated dams that were euthanized at 100 days of gestation served as control animals. Inoculation of the dams resulted in a viremia that lasted until the end of the study. Transplacental PRRSV spread was detected in all inoculated dams. Using immunofluorescence staining, single PRRSV-positive cells were found in the endometrial connective tissues adjacent to both PRRSV-positive and PRRSV-negative fetuses. In the fetal placental mesenchyme of the PRRSV-positive fetuses, infected cells were more abundant and spread focally. Double staining showed that all PRRSV-positive cells in the fetal implantation sites were positive for sialoadhesin and CD163. Apoptotic cells (TUNEL+) were detected in endometrium and fetal placentas of both non- and PRRSV-inoculated dams. The number of apoptotic cells was significantly higher in PRRSV-positive endometrium/fetal placentas. PRRSV caused apoptosis in infected cells since 20–61% of PRRSV-positive cells were apoptotic and in surrounding cells since 43–91% of the apoptotic cells were virus-negative. The main conclusion obtained from the present study is that PRRSV replicates in the fetal implantation sites and causes apoptosis in infected macrophages and surrounding cells at the last stage of gestation. The possible mode of PRRSV replication in the fetal implantation sites and the events that might contribute to the reproductive disorders are discussed.

Expression and localization of aromatase P450 AROM, estrogen receptor-α, and estrogen receptor-β in the developing fetal bovine frontal cortex

The enzyme aromatase (P450 AROM) converts testosterone (T) into 17-β estradiol (E 2) and is crucial for the control of development of the central nervous system during ontogenesis. The effects of E 2 in various brain areas are mediated by the estrogen receptor alpha (ER-α) and the estrogen receptor beta (ER-β). During fetal development, steroids are responsible for the sexual differentiation of the hypothalamus. Estrogens are also able to exert effects in other brain areas of the fetus including the frontal cortex, where they act through estrogen receptors (ERs) modulating cognitive function and affective behaviors. In this study we have determined the expression profiles of P450 AROM and ERs in the fetal bovine frontal cortex by quantitative Real-Time PCR (qRT-PCR) throughout the prenatal development. The data show that the patterns of expression of both ERs are strongly correlated during pregnancy and increase in the last stage of gestation. On the contrary, the expression of P450 AROM has no correlation with ERs expression and is not developmentally regulated. Moreover, we performed immunochemical studies showing that fetal neurons express P450 AROM and the ERs. P450 AROM is localized in the cytoplasm and only seldom present in the fine extensions of the cells; ER-α is detected predominantly in the soma whereas ER-β is only present in the nucleus of a few cells. This study provides new data on the development of the frontal cortex in a long gestation mammal with a large convoluted brain.

Vascular and apoptotic changes in the placode of myelomeningocele mice during the final stages of in utero development

Myelomeningocele (MMC) is a primary neurulation defect that is associated with devastating neurological disabilities in affected newborns. To better characterize the in utero neurodegenerative process of MMC, the authors investigated the changes in vascular organization, apoptosis, and the presence of inflammatory cells during gestation by using a mutant mouse model of MMC. The curly tail/loop tail (ct/lp) mutant mouse model of MMC was chosen to obtain fetuses at different stages of gestation. Mouse fetuses harboring MMC were harvested by caesarean section at embryonic Days 14.5, 16.5, and 18.5 (complete mouse gestation at 19 days, 6 mice/group); littermate fetuses with the same gestational age but without an MMC were used as controls. Samples of the MMC placode or normal spinal cord were stained for immunocytochemical labeling with caveolin antibody (endothelium marker) and activated caspase-3 antibody (apoptosis marker). Samples were morphometrically analyzed with a computer-assisted image analyzer. The MMC mice presented with an increase in vascular density from embryonic Days 16.5-18.5 and an enhanced number of apoptotic cells at embryonic Day 18.5, compared with controls. There were scarce signals of an inflammatory reaction in the MMC placode, as a few infiltrating neutrophils were seen only at embryonic Day 18.5. Fetal placodes in MMC mice showed evidence of increased vascular density since embryonic Day 16.5 and increased apoptosis at embryonic Day 18.5. These new data support the view that in utero changes of the MMC placode, occurring during the last stages of gestation, contribute to the neuropathological manifestations in full-term newborns with MMC.

Regulation of IGF-I and -II by Insulin in Primary Cultures of Fetal Rat Hepatocytes

During perinatal development, insulin and nutrients, rather than GH, regulate the IGF system. A selective primary culture of fetal rat hepatocytes has been established in our laboratory to elucidate the molecular mechanism of action of the above regulatory factors on IGF-I and -II gene expression during the late fetal period of the rat. In this model we have previously reported a regulatory role for glucose on IGF-I and -II synthesis and secretion. In the same experimental model, we now report that doses of insulin (0.1–5 μm) within the physiological range in rat fetuses during the last stages of gestation evoke an increase of IGF-I and -II mRNA abundance. Insulin regulated in a parallel manner IGF peptide secretion, and an excellent correlation was observed between IGF-I and -II mRNA and IGF-I and -II peptide levels in the conditioned media in response to the hormone. Finally, the insulin-induced rise in IGF-I and -II mRNA was not mediated by stimulation of gene transcription but by increased transcript stability. The results support the hypothesis that insulin plays a major role in IGF regulation at immature stages of development.