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This study provides a comparative assessment of fresh semen quality and sensitivity to cryopreservation in several Italian chicken breeds. The research involved 145 roosters from 13 breeds. The results showed a wide variability in all the parameters considered among the different breeds, especially in the quantitative variables of fresh semen, such as volume and concentration. For the qualitative characteristics (sperm membrane integrity and motility parameters), the variability across breeds was more pronounced for frozen than fresh semen. Interestingly, apart from total motility in fresh semen, breed had a significant effect on all semen quality parameters in both fresh and thawed ejaculates. Considering the overall qualitative characteristics, the Robusta maculata, Siciliana, and Mericanel della Brianza breeds produced ejaculates with better semen quality compared to other Italian breeds. By evaluating the main parameters of semen quality, our results underline the potential of these traits to influence the reproductive success and genetic conservation. The Bionda piemontese, Bianca di Saluzzo, Livorno bianca, Pepoi, and Siciliana breeds showed better resilience to cryopreservation, suggesting the need for breed-specific protocols to optimise semen quality after thawing. Importantly, the research highlights the central role of semen quality for both immediate fertilisation success and long-term conservation efforts. Future studies integrating OMICS technologies could elucidate molecular markers influencing breed-specific differences, helping to refine cryopreservation techniques and improve conservation strategies for indigenous Italian chicken breeds. This work contributes valuable insights to global efforts aimed at safeguarding poultry genetic diversity and sustainability.

There is a paucity of information about milk composition and quality at the quarter level from complete milking of the udder, mainly because of the difficulties of sampling procedures which imply the adoption of a vacuum system connected to four independent buckets. The objective of this study was to evaluate the effect of quarter position, intramammary infection status, and somatic cells on milk yield and composition at the mammary quarter level of Simmental cows. Milk samples were collected during three consecutive days, after milking individual mammary quarters of 10 lactating cows separately, for a total of 120 quarter milk samples. Samples were analysed for composition, somatic cell count (SCC), differential SCC, and bacterial culture. Somatic cell count was log-transformed to somatic cell score (SCS) to achieve normality of distribution. Sources of variation of milk yield and quality traits were investigated using a linear mixed model which included the fixed effects of quarter position (1 to 4), pathogen presence (positive or negative), and SCS class (low, medium-low, medium-high, and high). The interaction between cow and quarter position was included as random effect. Quarter position significantly affected milk yield, with rear udder quarters yielding more milk compared to their front counterparts, but with negligible effects on milk composition. Classes of SCS influenced milk composition, in a way that quarters with high SCS had lower lactose content and higher differential SCC. This study is a contribution towards the possibility to better understand physiological processes at cow udder quarter level.

A tool to help Italian dairy farmers choosing the most suitable replacement strategy has been developed. The approach aimed to identify yearly female replacement needs based on herd performance level and combination of different semen type (conventional, sex-sorted, and beef semen), with the ultimate goal of enhancing farm profit. A case study based on a 350-cow Holstein herd was used and three levels of herd fertility (high, medium, and low) were simulated to define the yearly number of dairy female replacements needed and the number of females yielded under different semen utilisation scenario. The number of annual dairy replacements was obtained as the number of cows multiplied by the replacement rate and adjusted by the age at first calving. Number of animals yielded was used to evaluate the replacement cost per 100 L of milk. Then, four strategies of sexed semen utilisation were combined with five strategies of beef semen use. Animals that were not inseminated with sexed or beef semen were bred with conventional semen. Regardless of fertility level, the number of dairy female replacement heifers that the farm needs are 110. Increasing beef semen use allows farmer to yield less replacement heifers. Furthermore, as beef semen use increases and the number of replacement heifers decreases, replacement cost per 100 L of milk reduces. Therefore, our results highlighted that replacement costs increase with increasing number of yielded heifers. Hence, combining beef and sexed semen to reach heifer balance close to zero, decreased the replacement cost.

Abstract Background Milk is essential for mammalian nutrition because it provides vital nutrients for growth and development. Milk composition, which is influenced by genetic and environmental factors, supports lactation, a complex process crucial for milk production and quality. Recent research has focused on noncoding RNAs, particularly microRNAs (miRNAs), which are present in body fluids and regulate gene expression post-transcriptionally. This study comprehensively characterizes miRNAs in milk of four livestock species, namely Bubalus bubalis, Capra hircus, Equus asinus, and Ovis aries and identifies potential target genes. Results High-throughput sequencing of milk RNA resulted in distinct read counts across species: B. bubalis (8,790,441 reads), C. hircus (12,976,275 reads), E. asinus (9,385,067 reads), and O. aries (7,295,297 reads). E. asinus had the highest RNA mapping rate (94.6%) and O. aries the lowest (84.8%). A substantially greater proportion of miRNAs over other small RNAs was observed for the donkey milk sample (7.74%) compared to buffalo (0.87%), goat (1.57%), and sheep (1.12%). Shared miRNAs, which included miR-200a, miR-200b, miR-200c, and miR-23a among others, showed varying expression levels across species, confirmed by qPCR analysis. Functional annotation of predicted miRNA target genes highlighted diverse roles, with an enrichment in functions linked to metabolism and immunity. Pathway analysis identified immune response pathways as significant, with several miRNAs targeting specific genes across species, suggesting their regulatory function in milk. Conclusions Both conserved and species-specific miRNAs were detected in milk of the investigated species. The identified target genes of these miRNAs have important roles in neonatal development, adaptation, growth, and immune response. Furthermore, they influence milk and meat production traits in livestock.

Abstract Background Intramammary infection is the result of invasion and multiplication of microorganisms in the mammary gland and commonly leads to mastitis in dairy animals. Although much has been done to improve cows’ udder health, mastitis remains a significant and costly health issue for dairy farmers, especially if subclinical. In this study, quarter milk samples from clinically healthy cows were harvested to detect pathogens via quantitative PCR (qPCR) and evaluate changes in individual milk traits according to the number of quarters infected and the type of microorganism(s). A commercial qPCR kit was used for detection of Mycoplasma bovis, Mycoplasma spp., Staphylococcus aureus, coagulase-negative staphylococci (CNS), Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis, Prototheca spp., Escherichia coli, Klebsiella spp., Enterococcus spp. and Lactococcus lactis ssp. lactis. Quarter and pooled milk information of 383 Holstein, 132 Simmental, 129 Rendena, and 112 Jersey cows in 9 Italian single-breed herds was available. Results Among the cows with pathogen(s) present in at least 1 quarter, CNS was the most commonly detected DNA, followed by Streptococcus uberis, Mycoplasma bovis, and Streptococcus agalactiae. Cows negative to qPCR were 206 and had the lowest milk somatic cell count. Viceversa, cows with DNA isolated in ≥ 3 quarters were those with the highest somatic cell count. Moreover, when major pathogens were isolated in ≥ 3 quarters, milk had the lowest casein index and lactose content. In animals with pathogen(s) DNA isolated, the extent with whom milk yield and major solids were impaired did not significantly differ between major and minor pathogens. Conclusions The effect of the number of affected quarters on the pool milk quality traits was investigated in clinically healthy cows using a commercial kit. Results remark the important negative effect of subclinical udder inflammations on milk yield and quality, but more efforts should be made to investigate the presence of untargeted microorganisms, as they may be potentially dangerous for cows. For a smarter use of antimicrobials, analysis of milk via qPCR is advisable – especially in cows at dry off - to identify quarters at high risk of inflammation and thus apply a targeted/tailored treatment.

Abstract Background Copy number variants (CNV) hold significant functional and evolutionary importance. Numerous ongoing CNV studies aim to elucidate the etiology of human diseases and gain insights into the population structure of livestock. High-density chips have enabled the detection of CNV with increased resolution, leading to the identification of even small CNV. This study aimed to identify CNV in local Italian chicken breeds and investigate their distribution across the genome. Results Copy number variants were mainly distributed across the first six chromosomes and primarily associated with loss type CNV. The majority of CNV in the investigated breeds were of types 0 and 1, and the minimum length of CNV was significantly larger than that reported in previous studies. Interestingly, a high proportion of the length of chromosome 16 was covered by copy number variation regions (CNVR), with the major histocompatibility complex being the likely cause. Among the genes identified within CNVR, only those present in at least five animals across breeds (n = 95) were discussed to reduce the focus on redundant CNV. Some of these genes have been associated to functional traits in chickens. Notably, several CNVR on different chromosomes harbor genes related to muscle development, tissue-specific biological processes, heat stress resistance, and immune response. Quantitative trait loci (QTL) were also analyzed to investigate potential overlapping with the identified CNVR: 54 out of the 95 gene-containing regions overlapped with 428 QTL associated to body weight and size, carcass characteristics, egg production, egg components, fat deposition, and feed intake. Conclusions The genomic phenomena reported in this study that can cause changes in the distribution of CNV within the genome over time and the comparison of these differences in CNVR of the local chicken breeds could help in preserving these genetic resources.

ABSTRACT: Eggshell and shank color in poultry is an intriguing topic of research due to the roles in selection, breed recognition, and environmental adaptation. This study delves into the genomics foundations of shank and eggshell pigmentation in Italian local chickens through genome-wide association studies analysis to uncover the mechanisms governing these phenotypes. To this purpose, 483 animals from 20 local breeds (n = 466) and 2 commercial lines (n = 17) were considered and evaluated for shank and eggshell color. All animals were genotyped using the Affymetrix Axiom 600 K Chicken Genotyping Array. As regards shank color, the most interesting locus was detected on chromosome Z, close to the TYRP1 gene, known to play a key role in avian pigmentation. Additionally, several novel loci and genes associated with shank pigmentation, skin pigmentation, UV protection, and melanocyte regulation were identified (e.g., MTAP, CDKN2A, CDKN2B). In eggshell, fewer significant loci were identified, including SLC7A11 and MITF on chromosomes 4 and 12, respectively, associated with melanocyte processes and pigment synthesis. This comprehensive study shed light on the genetic architecture underlying shank and eggshell color in Italian native chicken breeds, contributing to a better understanding of this phenomenon which plays a role in breed identification and conservation, and has ecological and economic implications.

Crossbreeding has been employed to address environmental challenges. One successful example is the Siboney de Cuba, developed in response to economic challenges in the 1960s. The aim of this study was to perform the first genomic characterization of the Siboney de Cuba breed, a successful hybrid breed resulting from the crossbreeding of Cuban Zebu and Holstein, using SNP array chip. For this purpose, 48 Siboney de Cuba cattle samples were collected and genotyped with the GGP Bovine 100k BeadChip, resulting in 83,314 SNPs after quality control. The genetic diversity was investigated using observed and expected heterozygosity, inbreeding coefficient, and minor allele frequency. Runs of homozygosity (ROH) analysis provided insights into molecular inbreeding. Additionally, the study investigated copy number variants (CNV), identifying CNV regions and their distribution. The genetic relationship and population structure of Siboney de Cuba were analyzed in comparison with worldwide cattle populations using ADMIXTURE, multidimensional scaling, and phylogenetic analysis. Six ROH islands containing a total of 50 genes were discovered, some of which were uncharacterized loci. Furthermore, 792 CNV with higher occurrence of genetic material loss were observed. The overall genome coverage for CNV regions was 2.16%. The Siboney de Cuba exhibited a good level of genetic variability with high heterozygosity and low inbreeding when compared with other cattle breeds worldwide. Also, the breed shared genetic similarity to hybrids from America and Bos indicus from Africa and highlighted a moderate level of genetic isolation with some overlaps with Bos taurus from America. The breed showed a complex genetic composition, influenced by historical factors. Overall, findings of the present study contribute to the understanding of genomic structure of Siboney de Cuba cattle breed.