In this study, we have retrieved a list of genes found to be involved in both positive and negative regulation of litter size in S. scrofa, O. aries, Mus musculus, Rattus norvegicus and Capra hircus from UniProt and NCBI repositories. Earlier studies have reported that the reproductive system of animals is regulated by an arsenal of hormones . The litter size gene list retrieved from public repositories majorly represented genes encoding for hormones. Interestingly, when we compared the list of litter size-regulating genes of C. hircus, S. scrofa and M. musculus retrieved from public repositories, it resulted in a total of 26 genes among the compared list of genes. Studies conducted in the past have mainly reported the involvement of the commonly observed genes mentioned above to be involved in regulation of litter size. We have also retrieved and compared the decreased, increased and abnormal litter size gene sets from Harmonizome databases. In comparison, all of the genes obtained from our literature analysis and differentially expressed genes obtained from our metadata analysis have shown 3 genes in common, which are HEXA, PTTG1 and TIMP1. Hexosaminidase-α (HEXA) and its isozymes HEXB and HEXS together have the capacity to breakdown a variety of substrates such as GM2 gangliosides, glycolipids, glycosaminoglycans and glycoproteins, which for the most part contain β-linked N-acetylglucosamine and N-acetyl galactosamine residues . According to Juneja (2002), the hexosaminidase knockout mice exhibited reduced fertility at a young age, which progressively decreased with increased age and ultimately lead to infertility . However, the HEXB knockout mice were found to develop normally and be fertile during the early stages of development. These results indicate that hexosaminidase is not required for sperm-ovum interactions and fertilization . PTTG1 gene encodes for pituitary tumor-transforming gene, an oncogene which is found to play a key role in cell cycle regulation and sister chromatid separation . Previous studies have reported that PTTG1 is highly expressed in various tumors, especially ovarian tumorigenesis. However, the exact involvement of PTTG1 in fertilization and with respect to litter size has not yet been reported. TIMP1 gene encodes for tissue inhibitor of metalloproteinase, it is found to play a key role in ovulation. Rosewell et al. (2013) demonstrated that gonadotropin-induced increase in TIMP protein in human periovulatory follicles could help regulate the follicular extracellular matrix and other TIMP associated processes with ovulation with an increase in TIMP inhibitor .
Biosynthesis and Metabolism of Lipids, Cholesterol and Steroidogenesis
Genes involved in steroid biogenesis and ovarian steroidogenesis pathways such as Cyp11A1, Msmo1, Star, Dhrs7/Hsd17b7, and Hmgcs1 were up- regulated in high litter group samples. Cyp11A1, also known as cholesterol side-chain cleavage enzyme, is a mitochondrial enzyme which helps with the biosynthesis of various steroid hormones  According to Gharani et al. (1997), allelic variants of Cyp11a1 might cause hyperadrogenaemia which may further lead to changes in ovarian morphology . This study also reported that Cyp11a plays a significant role in the progression of hirsutism in polycystic ovary syndrome conditions . Msmo1 gene is also involved in the biosynthesis of steroids and the production of zymosterol from lanosterol. The gene expression studies conducted by Dessie et al. (2015) have reported that the down-regulation of genes involved in the biosynthesis and metabolism of steroids, cholesterol and lipids in PCOS model rats (rats subjected to 5α-dihydrotestosterone (DHT)) was found to mimic a hyperandrogenic condition . Dessie et al. (2015) also reported that genes involved in the synthesis of steroid hormones, such as Cyp11A1, Star, Dhrs7/Hsd17b7, and Hmgcs1 were significantly repressed in PCOS model rats but expressed within the control group . Thus, expression of genes involved in the biosynthesis and metabolism of steroids, lipids and cholesterol in high litter group samples supports their involvement in fertility. Similarly, we also observed various other genes involved in the biosynthesis and metabolism of lipids, such as Aldh1a1, Dhrs7, Faxdc2, HEXA, and Lipin1, which were up regulated in high litter group samples. Bowles et al. (2016) reported that Aldh1a1 expressed in fetal ovaries played an important role by providing retinoic acid, and that a lack of Aldh1a1 in fetal ovaries lead to delayed germ cell meiosis . Dhrs7, also known as 17β-hydroxysteroid dehydrogenase, is involved in the conversion of estrone (E1) to estradiol (E2) and is highly expressed in the ovaries of pregnant animals. According to Pasi et al. (2000), spatial and temporal expression of Dhrs7/Hsd17b7 in the uterus indicates that locally-produced estradiol plays a crucial role in implantation . Faxdc2 (fatty acid hydroxylase domain 2), which is involved in cholesterol synthesis, had megakaryopoiesis highly up-regulated in all high litter samples . Lipin1 is a central metabolic regulator found to play an important role in lipid metabolism, especially the glycerolipid and glycerophospholipid metabolic pathways. Lipin1 gene and its polymorphisms were also found to be involved in the development of PCOS. Gowri et al. (2007) reported that Lipin1 is down-regulated by estradiol in the uterus and liver, and that the expression levels of Lipin1 is low and compromised in mouse models with diabetes and/or reduced fertility . Earlier studies have reported that Lipin1 deficient mice (fld/fld) have less body fat and exhibit symptoms of diabetes and impaired fertility [37–39]. Regulation of Lipin1 by estrogen plays a critical role between reproduction, growth and metabolism .
Genes exhibiting a role in the progression of polycystic ovary syndrome
The above obtained list of differentially expressed genes were found to be play a minor to major role in the development and progression of PCOS. Previous studies have reported that genes encoding for CYP11A1, HSD17B7, STAR, INHA, PARM1, SCARB1, PTGFR, SLC22A4, and SLC35F5 were found to take part in the development and progression of PCOS. The gene expression studies conducted by Dessie et al. (2015) reported that down-regulation of genes involved in the biosynthesis and metabolism of steroids, cholesterol and lipids in PCOS model rats (rats subjected to 5α-dihydrotestosterone (DHT)) was found to mimic a hyperandrogenic condition . Dessie et al. (2015) also reported that genes involved in the synthesis of steroid hormones such as Cyp11A1, Star, Dhrs7/Hsd17b7, and Hmgcs1 were significantly repressed in PCOS model rats but expressed in the control group . Francisco et al. (2009) hypothesized that haptoglobin (HP2) polymorphisms may contribute to the conditions, such as oxidative stress and chronic inflammation, which are associated with polycystic ovary syndrome, obesity and glucose tolerance . All the genes mentioned above that are involved in the metabolism of steroids, cholesterol, and lipids were found to be significantly down regulated in low litter group samples.
Genes involved in the immune response, cancer, cell growth and death-related pathways
Genes involved in immune responsive pathways such as CD55 (decay-accelerating factor for complement-55) and OAS1 (2'-5'-oligoadenylate synthetase 1) were highly up-regulated in high litter group samples. OAS1D is a cytoplasmic protein expressed in growing oocytes and early embryos (Wei et al., 2005). Mutant mice without OAS1D exhibited reduced fertility as they possessed defects in ovarian follicle development . For the first time, Wei et al. (2005) revealed that OAS1D controls female fertility in mice, and that OAS1D non-enzymatic OAS1 proteins may suppress IFN/OAS/RNaseL and protect oocytes and early embryos from cell death . Decay-accelerating factor (CD55) is a complement regulatory protein which protects the host cells through the innate immune response . The function of CD55 in reproduction has been hypothesized based on its up-regulation in the fetoplacental trophoblast, which protects the fetus from maternal complement injury . Kim et al. (2017) reported that CD55 was down regulated in the endometrium of subjects with repeated implantation failure. Similarly, genes encoding for CCNG1 (participates in p53-dependent G1–S and G2 checkpoints and might function as an oncogenic protein in the initiation and metastasis of ovarian carcinoma)  are PTGFR and ADHFE1 (breast cancer oncogene which induces metabolic re-programming) . All the genes above were up-regulated in high litter group samples, whereas genes encoding for CCNA2, GLI1, and SLC16A3 involved in cancer progression were up-regulated in low litter group samples.
Potential genes involved in prolificacy: The quantitative trait locus (QTL) studies conducted in the past [4, 46] have tried to understand the list of candidate genes affecting litter size in pigs. These studies have reported a list of 18 genes as significant QTL: CYP19A1, C5, PTGDS, NOV, TST, KRT8, HP, CES1, SULT2A1, CD83, FKBP5, DHRS4, SERPINA1/3, FGA/B/G, SPP1, HPX, MSRB2, SLC16A3, SPHK1, VTN, AHSG, OAS1, RBP4, CYP2E1, NCKAP5, EPHA4, and HOXA9 [4, 46]. Amanda et al. (2011) reported that genes encoding for OAS1, CD55, and SERPINA1 were up-regulated in high litter samples, and genes encoding for FAM46C, SPP1, RBP4, TST, and VTN were highly up-regulated in low litter group samples . Results obtained were in accordance with the findings of Amanda et al. (2015) , except genes encoding for FAM46C and FAM167B were found to be differentially up-regulated in high litter groups of the GSE23985 dataset. Genes encoding for CD55 and OAS1 were highly up-regulated in high litter samples and genes encoding for RASSF2, NEXN, and SLC16A3 were up-regulated in low litter samples of GSE21383 found in common with other datasets, respectively. Sun et al. (2011) reported genes involved in the p53 and Wnt signaling pathways such as CCNG1 (GSE23985 and Zhang et al., ), GTSE1 (Zhang et al., ) and WLS (GSE23985 and Zhang et al., ). Zhang et al. (2015) has reported about 10 genes encoding for CO1, GPX3, MSMB, COX3, TIMP1, CYTB, STAR, HSD3B, CYP11A1, SCARB1, and HSD17B2 that were found to be differentially expressed between the high and low litter group samples. Results obtained from our metadata analysis are also in accordance with the findings of Zhang et al. (2015).
Finally, results obtained from the metadata analysis of prolificacy-based gene expression datasets of pig and sheep has revealed a list of 42 genes differentially expressed in high litter sow groups and 20 genes expressed in low litter sow groups. Previous reports have proposed the involvement of genes such as CYP11A1, HSD17B2, STAR, SCARB1, IGSF8, MSMB, and SERPINA1 in regulating fecundity. The functional involvement of several other genes such as HEXA, PTTG1, TIMP1, FAM46C, FAM167B, CCNG1, FAXDC2, HMGCS1, L2HGDH, Lipin1, MME, MSMO1, PARM1, PTGFR, SLC22A4, SLC35F5, CCNA2, CENPU, CEP55, RASSF2, and SLC16A3 were reported to play a role in fertilization and embryo development. However, their exact function regarding prolificacy rate must be uncovered in future studies.