Examining the Nutrient Exchange in All Incubation Periods in Chukar Partridges

Background: This study was conducted to examine the development of the embryo during incubation and the composition, amount and consumption of the egg nutrients among Chukars (Alectoris chukar). Methods: A total of 1210 eggs were carefully separated into their components (albumen, yolk, shell, amniotic-allantoic fluid and embryo) on day 0 (fresh), in the first (day 7), second (day 14) and third (day 21) weeks of incubation and on the hatching date. The moisture, crude protein, crude fat, carbohydrate, crude ash and energy level were determined in egg components. Result: The albumen, yolk, shell, fluid and embryo rates were calculated to be 53.62, 34.84, 11.53, 0.0 and 0.0% in fresh egg, 16.74, 33.49, 16.22, 31.76 and 2.06% in the first week of incubation, 15.65, 28.32, 15.58, 23.35 and 17.10% in the second week of incubation and 0.0, 23.33, 19.38, 0.0 and 57.29% in the third week of incubation, respectively. The hatching weight of the chick was 16.16 g. The nutrients of the yolk, albumen, fluid and shell proportionally varied significantly in different periods of incubation. According to the nutrient changes calculated based on the weight differences of egg components, carbohydrates were mostly used for nutrients during incubation. It was found that even though the nutrients in all the egg components were used in the different periods of incubation, egg yolk was an important source for the embryo throughout the incubation period.

Detection of infectious bronchitis-like virus isolated from psittacines at breeding facilities

Suryaman GK, Soejoedono RD, Poetri ON, Setiyono A, Handharyani E. 2021. Detection of infectious bronchitis-like virus isolated from psittacines at breeding facilities. Biodiversitas 22: 4261-4265. Captive breeding of parrots has been carried out in Indonesia to fulfill market demand for pets and support conservation efforts. However, there is limited information about Infectious Bronchitis Virus (IBV) in captive parrots. Therefore, this study examines IBV virus in psittacines and observes the pathological changes in inoculated chicken embryos. Cloacal swabs were obtained from 31 psittacine birds consisting of 15 sulfur-crested cockatoos (Cacatua sulphurea), 10 palm cockatoos (Probosciger aterrimus), two blue and yellow macaws (Ara ararauna), two green-winged macaws (Ara chloropterus), and two scarlet macaws (Ara macao). Swabs were also injected for 9 days to obtain specific pathogen-free (SPF) embryonated chicken eggs, and allantoic fluids were collected after 7 days. The residual cloacal swabs and allantoic fluid samples were tested for IBV by reverse transcription polymerase chain reaction (RT-PCR) targeting spike 1 glycoprotein (S1) and nucleoprotein (N) gene by primer set XCE2+/XCE2- and IBVN+/IBVN-, while the harvested inoculated embryos were examined for histopathological lesions by hematoxylin-eosin staining. Three and one allantoic fluid of C. sulphurea and P. aterrimus respectively were found positive for IBV by primer set IBVN+IBVN-. However, there was no significant difference in the inoculated embryo's gross anatomy, and the slight histopathological changes observed were not caused by the virus. Therefore, it is unclear whether the IB-like virus causes clinical disease manifestation in both Psittacine species or the chicken embryo. This is the first study carried out on IB-like viruses for both species.

Characterization of the microbiota associated with 12-week-old bovine fetuses exposed to divergent in utero nutrition

A recent study reported the existence of a diverse microbiota in 5-to-7-month-old calf fetuses, suggesting that colonization of the bovine gut with so-called 'pioneer' microbiota may begin during mid-gestation. In the present study, we investigated 1) the presence of microbiota in bovine fetuses at early gestation (12 weeks), and 2) whether the fetal microbiota is influenced by the maternal rate of gain or dietary supplementation with vitamins and minerals (VTM) during early gestation. Amniotic and allantoic fluids, and intestinal and placental (cotyledon) tissue samples obtained from fetuses (n = 33) on day 83 of gestation were processed for the assessment of fetal microbiota using 16S rRNA gene sequencing. The sequencing results revealed that a diverse and complex microbial community was present in each of these fetal compartments evaluated. Allantoic and amniotic fluids, and fetal intestinal and placenta microbiota each had distinctly different (0.047 ≥ R2 ≥ 0.019, P ≤ 0.031) microbial community structures. Allantoic fluid had a greater (P < 0.05) microbial richness (number of OTUs) (Mean 122) compared to amniotic fluid (84), intestine (63) and placenta (66). Microbial diversity (Shannon index) was similar for the intestinal and placental samples, and both were less diverse compared with fetal fluid microbiota (P < 0.05). Thirty-nine different archaeal and bacterial phyla were detected across all fetal samples, with Proteobacteria (55%), Firmicutes (16.2%), Acidobacteriota (13.6%) and Bacteroidota (5%) predominating. Among the 20 most relatively abundant bacterial genera, Acidovorax, Acinetobacter, Brucella, Corynebacterium, Enterococcus, Exiguobacterium and Stenotrophomonas differed by fetal sample type (P < 0.05). A total of 55 taxa were shared among the four different microbial communities. qPCR of bacteria in the intestine and placenta samples as well as scanning electron microscopy imaging of fetal fluids provided additional evidence for the presence of a microbiota in these samples. Minor effects of maternal rate of gain and VTM supplementation, and their interactions on microbial richness and composition were detected. Overall, the results of this study indicate that colonization with pioneer microbiota may occur during early gestation in bovine fetuses, and that that the maternal nutritional regime during gestation may influence the early fetal microbiota.

Characterization of Newcastle Disease Virus in Broiler Flocks with Respiratory symptoms in some Provinces of Iran

Background: Newcastle disease, is one of the most important diseases of the poultry industry, has many economic losses. The aim of this study was to isolate and determine the molecular identity of Newcastle disease virus in 40 broiler flocks with respiratory symptoms in four provinces of Iran.Methods and Results: Samples of farms with respiratory symptoms were collected from different regions of Isfahan, East Azerbaijan, Golestan, and Khuzestan provinces and inoculated into 9-day-old embryonated chicken eggs. The Reverse-transcription polymerase chain reaction (RT-PCR) was performed to detect the Newcastle disease virus on allantoic fluid. Of the 40 flocks, the virus was isolated and identified in 16 flocks. The PCR products of 16 isolates were sequenced and a phylogenetic tree was drawn. Accordingly, six isolates were in genotype II and ten isolates were in subgenotype VIId of class II. Conclusion: Both genotypes were present in all four provinces. The isolates of Khuzestan province showed the greatest diversity compared to the other three provinces. The similarity of isolates belonging to genotype II in this study was observed with Pakistan, China, and Nigeria and other isolates were similar to previous isolates in Iran. Also, the highest amino acid sequence in the F-protein cleavage site was 112RRQKR/F117 for VIId genotype isolates and 112GRQGR/L117 for II genotype isolates.

​Color Doppler Ultrasonographic Characteristics of Embryonic Development and Mortality in Dairy Cows

Background: Embryonic mortality is one of the major factors leading to reproductive failure, decreased fertility and delayed pregnancy in cattle. Method: The study was aimed for investigating the incidence of embryonic mortality(EM), differentiate embryonic development and mortality based on color Doppler ultrasonographic characteristics. Different characteristics correlated were corpus luteum(CL) vascularity, embryonic development and EM features, monitored on day 21 post-insemination. Result: The overall incidence of EM was recorded to be 25%, with 15% and 10% mortality occurred between days 21-28 and 28-35 of gestation. Color Doppler analysis stated that, the CL in pregnant cows had a higher percentage of luteal tissue with positive Doppler signals on initial examination (day 21) differing significantly (P less than 0.01) from non-pregnant and consistently increased till the end of the examination. However, cows with EM had a significantly lower CL vascularity than pregnant (P less than 0.05) but significantly higher (P less than 0.05) than non-pregnant cows on day 21. Embryos from pregnant cows (detected by 28 days) increased in size along the days of examination. However, cows diagnosed with EM showed signs (day 28), which included reduced intra-uterine allantoic fluid, absence of embryo proper with an embryonic heartbeat, hyperechoic free-floating embryonic debris in the uterus and distorted allanto-chorionic membrane. In conclusion, the present study firmly supports color Doppler ultrasonography in diagnosis of pregnant cows based on CL vascularity, visualizing the different characteristics signs of EM, and detecting non-pregnant animals undergoing EM as early as day 21.

Dynamic Changes of Metabolite Profiles in Maternal Biofluids During Gestation Period in Huanjiang Mini-Pigs

The biochemical parameters related to nitrogenous metabolism in maternal biofluids may be linked and even reflect the fetal metabolism and growth. The present study have measured the concentrations of various parameters related to amino acid (AA) and lipid metabolism, as well as different metabolites including the free AAs in maternal plasma and amniotic and allantoic fluid corresponding to fetuses with different body weight (BW) during different gestation periods, in order to identify the possible relationships between biochemical parameters and fetal growth. A total of 24 primiparous Huanjiang mini-pigs were fed with a standard diet. Data showed that, from day 45 to day 110 of gestation, the maternal plasma levels of alanine aminotransferase (ALT), albumin (ALB), Ile, Orn, Car, α-ABA, and β-AiBA increased (P &lt; 0.05); while the levels of ammonia (AMM), choline esterase (CHE), high density lipoprotein-cholesterol (HDL-C), Leu, Glu, Cys, Asp, and Hypro decreased (P &lt; 0.05). From day 45 to 110 of gestation, the amniotic fluid levels of aspartate transaminase (AST), CHE, total protein (TP), and urea nitrogen (UN) increased (P &lt; 0.05), as well as the level of CHE and TP and concentration of Pro in allantoic fluid; while the amniotic fluid concentrations of Arg, Glu, Orn, Pro, and Tau decreased (P &lt; 0.05), as well as allantoic fluid concentrations of Arg and Glu. At day 45 of gestation, the amniotic fluid concentrations of Arg, Orn, and Tau corresponding to the highest BW (HBW) fetuses were higher (P &lt; 0.05), whereas the allantoic fluid concentrations of His and Pro were lower (P &lt; 0.05) when compared with the lowest BW (LBW) fetuses. At day 110 of gestation, the amniotic fluid concentration of Tau corresponding to the HBW fetuses was higher (P &lt; 0.05) than the LBW fetuses. These findings show that the sows display increased protein utilization and decreased lipid metabolism and deposition from day 75 to 110 of gestation. In addition, our data are indicative of a likely stronger ability of HBW fetuses to metabolize protein; and finally of a possible key role of Arg, Gln, Glu, Pro, Tau, and His for the fetal growth and development.

122 A Mechanistic Model of Growth and Amino Acid Deposition in the Pregnant Sow: Model Development, Evaluation, and Application

A mechanistic model was developed with the objective to characterize weight gain and essential amino acid (EAA) deposition in the different tissue pools that make up the pregnant sow: placenta, allantoic fluid, amniotic fluid, fetus, uterus, mammary gland, and maternal body were considered. The data used in this modelling approach were obtained from published scientific articles reporting weights, crude protein (CP), and EAA composition in the previously mentioned tissues; studies reporting not less than 5 datapoints across gestation were considered. A total of 12 scientific articles published between 1977 and 2020 were selected for the development of the model and the model was validated using 11 separate scientific papers. The model consists of three connected sub-models: protein deposition (Pd) model, weight gain model, and EAA deposition model. Weight gain, Pd, and EAA deposition curves were developed with nonparametric statistics using splines regression. The validation of the model showed a strong agreement between observed and predicted growth (r2 = 0.92, root mean square error = 3%). The proposed model also offered descriptive insights into the weight gain and Pd during gestation. The model suggests that the definition of time-dependent Pd is more accurately described as an increase in fluid deposition during mid-gestation coinciding with a reduction in Pd. In addition, due to differences in CP composition between pregnancy-related tissues and maternal body, Pd by itself may not be the best measurement criteria for the estimation of EAA requirement in pregnant sows. The proposed model also captures the negative maternal Pd that occurs in late gestation and indicates that litter size influences maternal tissue mobilization more than parity. The model predicts that the EAA requirements in early and mid-gestation are 75, 55 and 50% lower for primiparous sows than parity 2, 3 and 4+ sows, respectively, which suggest the potential benefits of parity segregated feeding.

Pre-implantation exogenous progesterone and pregnancy in sheep. II. Effects on fetal-placental development and nutrient transporters in late pregnancy

Background Administration of progesterone (P4) to ewes during the first 9 to 12 days of pregnancy accelerates blastocyst development by day 12 of pregnancy, likely due to P4-induced up-regulation of key genes in uterine epithelia responsible for secretion and transport of components of histotroph into the uterine lumen. This study determined if acceleration of blastocyst development induced by exogenous P4 during the pre-implantation period affects fetal-placental development on day 125 of pregnancy. Suffolk ewes (n = 35) were mated to fertile rams and assigned randomly to receive daily intramuscular injections of either corn oil vehicle (CO, n = 18) or 25 mg progesterone in CO (P4, n = 17) for the first 8 days of pregnancy. All ewes were hysterectomized on day 125 of pregnancy and: 1) fetal and placental weights and measurements were recorded; 2) endometrial and placental tissues were analyzed for the expression of candidate mRNAs involved in nutrient transport and arginine metabolism; and 3) maternal plasma, fetal plasma, allantoic fluid, and amniotic fluid were analyzed for amino acids, agmatine, polyamines, glucose, and fructose. Results Treatment of ewes with exogenous P4 did not alter fetal or placental growth, but increased amounts of aspartate and arginine in allantoic fluid and amniotic fluid, respectively. Ewes that received exogenous P4 had greater expression of mRNAs for SLC7A1, SLC7A2, SLC2A1, AGMAT, and ODC1 in endometria, as well as SLC1A4, SLC2A5, SLC2A8 and ODC1 in placentomes. In addition, AZIN2 protein was immunolocalized to uterine luminal and glandular epithelia in P4-treated ewes, whereas AZIN2 localized only to uterine luminal epithelia in CO-treated ewes. Conclusions This study revealed that exogenous P4 administered in early pregnancy influenced expression of selected genes for nutrient transporters and the expression of a protein involved in polyamine synthesis on day 125 of pregnancy, suggesting a ‘programming’ effect of P4 on gene expression that affected the composition of nutrients in fetal-placental fluids.

Novel Mineral Regulatory Pathways in Ovine Pregnancy: II. Calcium Binding Proteins, Calcium Transporters, and Vitamin D Signaling

Mineralization of the fetal mammalian skeleton requires a hypercalcemic gradient across the placenta from mother to fetus. However, the mechanisms responsible for maintaining the placental transport of calcium remain poorly understood. This study aimed to identify calcium and vitamin D regulatory pathway components in ovine endometria and placentae across gestation. Suffolk ewes were bred with fertile rams upon detection of estrus (Day 0). On Days 9, 12, 17, 30, 70, 90, 110, and 125 of pregnancy (n = 3–14/Day), ewes were euthanized and hysterectomized. Calcium abundance was influenced by gestational day in uterine flushings and allantoic fluid (P &lt; 0.05). The expression of S100G, S100A9, S100A12, ATP2B3, ATP2B4, TRPV5, TRPV6, CYP11A1, CYP2R1, CYP24, and VDR mRNAs known to be involved in calcium binding, calcium transport, and vitamin D metabolism were quantified by qPCR. Mediators of calcium and vitamin D signaling were expressed by Day 17 conceptus tissue, and endometria and placentae across gestation. Gestational day influenced the expression of S100G, S100A9, S100A12, TRPV6, VDR, and CYP24 mRNAs in endometria and placentae (P &lt; 0.05). Gestational day influenced endometrial expression of ATP2B3, and placental expression of TRPV5, ATP2B4, and CYP11A1 (P &lt; 0.05). VDR protein localized to the endoderm and trophectoderm (Day 17 conceptus) and was expressed in endometria and placentae throughout gestation. The observed spatiotemporal profile suggests a potential role of calcium and vitamin D in the establishment of pregnancy and regulation of fetal and placental growth, providing a platform for further mechanistic investigation.

Development and pre-clinical evaluation of Newcastle disease virus-vectored SARS-CoV-2 intranasal vaccine candidate

ABSTRACTThe COVID-19 pandemic has claimed the lives of millions of people worldwide and threatens to become an endemic problem, therefore the need for as many types of vaccines as possible is of high importance.Because of the millions of doses required, it is desirable that vaccines are not only safe and effective, but also easy to administer, store, and inexpensive to produce.Newcastle Disease Virus (NDV) is responsible for a respiratory disease in chickens. It has no pathogenic homologue in humans. NDV is recognized as an oncolytic virus, and its use in humans for oncological treatment is being evaluated.In the present work, we have developed two types of NDV-vectored candidate vaccines, which carry the surface-exposed RBD and S1 antigens of SARS-CoV-2, respectively. These vaccine candidates were produced in specific-pathogen-free embryonating chicken eggs, and purified from allantoic fluid before lyophilization. These vaccines were administered intranasally to three different animal models: mice, rats and hamsters, and evaluated for safety, toxicity, immunogenicity, stability and efficacy. Efficacy was evaluated in a challenge assay against active SARS-CoV-2 virus in the Golden Syrian hamster model.The NDV-vectored vaccine based on the S1 antigen was shown to be safe and highly immunogenic, with the ability to neutralize SARS-CoV-2 in-vitro, even with an extreme dilution of 1/640. Our results reveal that this vaccine candidate protects the lungs of the animals, preventing cellular damage in this tissue. In addition, this vaccine reduces the viral load in the lungs, suggesting that it may significantly reduce the likelihood of transmission. Being lyophilized, this vaccine candidate is very stable and can be stored for several months at 4-8⁰C.In conclusion, our NDV-based vaccine candidate has shown a very favorable performance in the pre-clinical study, serving as evidence for a future evaluation in a Phase-I human clinical trial. This candidate represents a promising tool in the fight against COVID-19.