GENETIC ANALYSIS OF FERTILITY AND SEASONALITY TRAITS IN BUBALUS BUBALIS

Water buffalo is an important livestock resource, which occupies a critical niche in many ecologically disadvantaged agricultural systems, providing milk, meat, and work power. In Italy, the most important production related to buffalo breeding is milk, traditionally processed into mozzarella cheese. Reproductive efficiency is the main factor affecting productivity in female buffalo, but is greatly penalized by late achievement of puberty, long postpartum anoestrus and seasonality of calving. The main environmental factor affecting the reproductive seasonality is photoperiod, which regulates changes in the daily melatonin secretion by the pineal gland. Under Mediterranean latitudes, the reproductive efficiency of buffaloes is negatively affected by increasing day-length. In the present research, a polymorphism detection and an association study in candidate genes involved in fertility and seasonality of reproduction in Mediterranean Italian Buffalo were performed. The candidate genes analyzed were: signal transducer and activator of transcription 5A (STAT5A), serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 14 (SERPINA14) and tumor necrosis factor alpha (TNFA) for fertility, and melatonin receptor 1A (MTNR1A) for seasonality. The SNP discovery analysis allowed the identification of a total of 12 SNPs. Of these, 11 were detected for the first time in present work, while a C>T substitution in MTNR1A gene was already found in literature and associated to seasonality in buffalo species. Seven SNPs among those found resulted polymorphic in a sample of 36 buffaloes, including the one already present in literature and six SNPs identified in this work for the first time. Of these, 2 are located in exon 4 of TNFA gene and 4 in STAT5A gene (1 in exon 8 and 3 in introns). All the coding SNPs are synonymous. These polymorphisms were genotyped on a total of 491 female Mediterranean Italian Buffaloes, characterized by the periodic milking recording data and the dates of calving, provided by the Italian Buffalo Breeders’ Association (ANASB). Population genetics parameters were calculated and associations with fertility, seasonality and milk production traits were tested. The marker at the MTNR1A gene and one at the STA5A gene deviated significantly from the Hardy-Weinberg equilibrium, with significantly different values for observed and expected heterozygosity. An interesting result emerged for the SNP located at the 3rd exon of TNFA gene, a candidate gene for fertility traits. A statistically significant association with calving interval was assessed in the fourth lactation subset, highlighting an improving effect for the A allele, which resulted associated to a reduction of the calving interval. Nevertheless, the frequency of A allele in the analyzed population is very low and decreases from the first lactation on. Concerning seasonality parameter, the C>T substitution in MTNR1A gene found in literature and associated to seasonality in buffalo species was confirmed in present work, but a relationship between this polymorphism and seasonal breeding was not completely established. The SNP at MTNR1A resulted also associated with milk, fat and protein yield (kg) and with protein percentage parameters. The associations with fat and protein yield were observed in the subset excluding data related to the first lactation. In all cases, animals homozygous for the minor allele T showed lower performances compared to the CC ones. For STAT5A gene, the TT genotype at the c.989+344C locus seems to be associated to a less seasonal reproductive pattern and to higher protein percentage performances. Concerning the latter, the worst mean value is shown by the heterozygous animals, for the phenomenon known as overdominance. Also the SNP STAT5Ac.1342+99A resulted associated with seasonality of reproduction, with GG animals showing a reduced sensitiveness to photoperiod. The G allele was the minor allele in the analyzed population and the homozygous genotype GG is very under-represented. No other statistically significant effects were detected for this polymorphism in the analyzed sample. Another polymorphism detected on the STAT5A gene, the C to G substitution at position c.128, showed a trend in respect to the protein yield (kg) in the subset including data related to classes of lactation number from 2 to 7. Also in this case, the lowest average protein yields were recorded for the heterozygous animals. The associations found, that could be tested in a larger sample, may offer useful indications for the genetic improvement of fertility, seasonality and production traits in buffalo species. Further researches on other genes and also including the SNP chip tool, which is now developing for buffalo species, should be performed to extend the knowledge about the molecular basis underlying the relationships between production and reproduction and the complex mechanisms behind seasonality and response to photoperiod. However, an accurate recording activity of phenotypic parameters, as well as genealogies registration, are critical elements in buffalo, and must be improved to enhance the genetic progress in this species. This could also allow obtaining further relevant results from the association analyses performed in present study.