Transformation of raw ewes' milk applying “Grana” type pressed cheese technology: development of extra-hard “Gran Ovino” cheese

Abstract This work was carried out to pursue a double objective: to improve the hygienic safety of cheeses produced from raw ewes' milk; and to produce a new typology of raw ewes' milk through the application of “Grana” technology for which the name “Gran Ovino” was chosen. With this in mind, raw milk from an individual farm was transformed under controlled conditions at a dairy pilot plant. The production technology included the partial skimming of the evening and morning milk mixture by cream surfacing and the addition of a natural whey starter cultures (NWSC) prepared with four selected Streptococcus thermophilus strains (PON6, PON244, PON261 e PON413). Ten microbial groups were investig…

Development of multiplex-PCR protocol to amplify 12S and 16S rRNA genes of mtDNA for traceability of Sicilian mono-species dairy products.

Species identification in dairy sector is important not only to safeguard public health but also to verify compliance with the Production Regulations of many typical dairy products (PDO/PGI). The most common fraud in dairy sector is represented by the mixture of milks from different species, resulting in mislabeling of protected designation of origin (PDO) products. For a rapid, specific and sensitive identification of cattle’s, sheep’s and goat’s milk in mono-species Sicilian dairy products, species-specific multiplex-PCR protocol was developed. DNA from blood and experimental cheeses of Sicilian autochthonous breeds was used to amplify the 12S and 16S rRNA genes of the mitochondrial DNA. …

Enteric bacteria of food ice and their survival in alcoholic beverages and soft drinks

This study aimed to evaluate the levels of enteric bacteria in ice cubes produced in different environments (home-made, prepared in bars and pubs with ice machines and produced in industrial plants) and to determine their survival in different alcoholic beverages and soft drinks. Members of the Enterobacteriaceae family were found in almost all samples analysed. All industrial and the majority of home-made samples did not contain coliforms. Enterococci were not identified in domestic samples while they were detected in two industrial and three bar/pub samples. The samples collected from bars and pubs were characterized by the highest levels of enteric bacteria. Fourteen strains representing…

Evolution of indigenous starter microorganisms and physicochemical parameters in spontaneously fermented beef, horse, wild boar and pork salamis produced under controlled conditions

Abstract The present work was carried out to evaluate the microbiological and physicochemical composition of salamis produced with the meat of beef, horse, wild boar and pork. Salami productions occurred under controlled laboratory conditions to exclude butchery environmental contaminations, without the addition of nitrate and nitrite. All trials were monitored during the ripening (13 °C and 90% relative humidity) extended until 45 d. The evolution of physicochemical parameters showed that beef and pork salamis were characterized by a higher content of branched chain fatty acids (FA) and rumenic acid than horse and wild boar salamis, whereas the last two productions showed higher values of …

Development of an ad hoc natural whey starter culture for the production of Vastedda della valle del Belìce cheese

This work was aimed to characterize the autochthonous lactic acid bacteria (LAB) of PDO Vastedda della valle del Belìce cheeses, produced in several dairy factories, for the development of an ad hoc starter culture preparation for the production of this cheese. To this purpose, winter and spring productions were analysed, in order to isolate LAB adapted to perform the fermentation at low temperatures. Plate counts showed the total microbial counts (TMC) till levels of 9 -1almost 10 CFU g and all cheese samples were dominated by coccus LAB. Not all samples were positive for the presence of enterobacteria, but when they were found their concentrations were at similar levels in both seasons. A…

APPLICATION OF SELECTED LACTIC ACID BACTERIA FOR THE YEAR-ROUND PRODUCTION OF VASTEDDALIKE CHEESE

12S rRNA mitochondrial gene as marker to trace Sicilian mono-species dairy products

Abstract For a rapid, specific and sensitive identification of cows', ewes' and goats' milk in mono-species Sicilian dairy products, species-specific duplex-PCR protocol was applied. DNA samples from blood and experimental cheeses of Sicilian autochthonous breeds were extracted to amplify the 12S rRNA (and part of 16S rRNA in case of Ovis aries ) mitochondrial species-specific gene fragment. The use of species-specific primers for Bos taurus , Capra hircus and Ovis aries species, after electrophoresis on agarose gel, yielded fragments of 256 bp, 326 bp and 172 bp, respectively. Amplification by duplex - PCR of DNA pools from two species showed detection thresholds of 0.1% of “contaminant” D…

ANALYSIS OF MICROSATELLITE MARKERS IN SICILIAN GOATS BREED FOR TRACEABILITY OF GIRGENTANA TYPICAL DAIRY PRODUCTS

The establishment of useful analytical methods able to ensure the origin of the products, including the breed used, are important in maintaining the reliability of these products in order to develop a market segment. Traceability, obtained by molecular analysis, could be a reliable proposal for the authentication and valorization of animal products. In Sicily, the three most important dairy goat breeds are Girgentana (GR), Maltese (ML) and Derivata di Siria (DS). The GR is an endangered autochthonous goat breed. Preservation of endangered breeds could be achieved by establishing economic reasons for their survival. The aim of this work was to verify the use of microsatellite markers to asse…

Polyphasic Characterization of Microbiota of “Mastredda”, a Traditional Wooden Tool Used during the Production of PDO Provola dei Nebrodi Cheese

The biofilms of the wooden tables used for the acidification of the curd were investigated for PDO Provola dei Nebrodi cheese, a traditional stretched cheese made in eastern Sicily (southern Italy) from raw cows’ milk. To this purpose the wooden tables of four dairy facilities were analysed for their microbiota by scanning electron microscopy (SEM) analysis and a combined culture-independent and -dependent microbiological approach. SEM inspection showed an almost continuous biofilm formation. MiSeq Illumina analysis identified 8 phyla, 16 classes, 25 orders, 47 families and 50 genera. Corynebacterium, Bifidobacterium and lactic acid bacteria (LAB) were detected in all samples. In particular…

Genetic characterisation of CSN2 gene in Girgentana goat breed

Among calcium sensitive caseins, β-casein is the most abundant in goat milk, representing up to 50% of total casein content. The goat β-casein locus has been widely investigated and at least ten alleles have been identified in different goat breeds. The aim of this work was to investigate the polymorphisms of β-casein gene in Girgentana dairy goat breed in order to assess the genotype distribution and evaluate how frequencies have changed during the last 10 years, as genotype is known to influence technological and nutritional milk properties. Sequencing analysis and alignment of the obtained sequences of β-casein exon 7, showed the presence of C, C1, and A strong alleles, and 0' null allel…

High-density single nucleotide polymorphism markers reveal the population structure of 2 local chicken genetic resources

Italy counts a large number of local chicken populations, some without a recognized genetic structure, such as Val Platani (VPL) and Cornuta (COS), which represent noteworthy local genetic resources. In this study, the genotype data of 34 COS and 42 VPL, obtained with the Affymetrix Axiom600KChicken Genotyping Array, were used with the aim to investigate the genetic diversity, the runs of homozygosity (ROH) pattern, as well as the population structure and relationship within the framework of other local Italian and commercial chickens. The genetic diversity indices, estimated using different approaches, displayed moderate levels of genetic diversity in both populations. The identified ROH h…

Genomic Tools for the Characterization of Local Animal Genetic Resources: Application in Mascaruna Goat

Italy contains a large number of local goat populations, some of which do not have a recognized genetic structure. The “Mascaruna” is a goat population reared for milk production in Sicily. In this study, a total of 72 individuals were genotyped with the Illumina Goat_IGGC_65K_v2 BeadChip with the aim to characterize the genetic diversity, population structure and relatedness with another 31 Italian goat populations. The results displayed a moderate level of genetic variability for Mascaruna, in concordance with the estimated values for Italian goats. Runs of homozygosity islands are linked to genes involved in milk production, immune response and local adaptation. Population structure anal…

Two-dimensional difference in gel electrophoresis (2D-DIGE) for milk proteins characherization in Girgentana goat breed

Milk proteomics has rapidly become an eligible approach, since the protein fraction constitutes the most biologically relevant component in milk. Proteomic applications can vary greatly from straightforward protein identification to complex characterization of post-translational modifications and protein-protein interactions. The aim of this work was to compare protein profile of Girgentana whole milk samples between lactation periods and geographical areas by two dimensional difference in gel electrophoresis (2D-DIGE). Milk samples were collected in two flocks located in two different areas of Sicily at three different time points from calving: 30, 55, and 120 days, respectively. Protein p…

Determination of milk production losses and variations of fat and protein percentages according to different levels of somatic cell count in Valle del Belice dairy sheep

The somatic cell count (SCC) of milk is widely used to monitor udder health and the milk quality and because of its positive genetic correlation with mastitis this trait was included in breeding schemes of dairy sheep. The aim of this study was to estimate the loss in milk yield (MY) and related composition resulting from different levels of somatic cell count in Valle del Belice dairy sheep. Data were collected between 2006 and 2016 in 15 flocks following an A4 recording scheme. Somatic cell count (SCC), fat and protein percentage (F% and P%) were determined using mid-infrared spectroscopy. To evaluate loss in test day MY, F% and P%, five different classes of SCC were arbitrarily defined: …

Identification, typing, and investigation of the dairy characteristics of lactic acid bacteria isolated from 'Vastedda della valle del Belìce' cheese

Traditional cheeses made without starter cultures can be characterised by the attribute of instability. The addition of autochthonous starter cultures can ensure stability without compromising the characteristics of the final product. This study aimed to characterise the autochthonous lactic acid bacteria (LAB) population in “Vastedda della valle del Belice” cheeses, which have a protected designation of origin (PDO) status, in order to develop an ad hoc starter culture to be used in its future production. Winter and spring productions were analysed to ensure isolation of specific LAB that had adapted to perform fermentation at low temperatures. Plate counts revealed total microbial numbers…

Genome-wide scan for runs of homozygosity identifies potential candidate genes associated with local adaptation in Valle del Belice sheep

Background Because very large numbers of single nucleotide polymorphisms (SNPs) are now available throughout the genome, they are particularly suitable for the detection of genomic regions where a reduction in heterozygosity has occurred and they offer new opportunities to improve the accuracy of inbreeding (\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$F$$\end{document}F) estimates. Runs of homozygosity (ROH) are contiguous lengths of homozygous segments of the genome where the two haplotypes inherited from t…

Microbial dynamics in durum wheat kernels during aging

In the present work the microbial dynamics in wheat kernels were evaluated over time. The main aim of this research was to study the resistance of lactic acid bacteria (LAB) and yeasts associated to unprocessed cereals used for bread making during long term conservation. To this purpose four Triticum durum Desf. genotypes including two modern varieties (Claudio and Simeto) and two Sicilian wheat landraces (Russello and Timilia) were analysed by a combined culture-independent and -dependent microbiological approach after one, two or three years from cultivation and threshing. DNA based MiSeq Illumina technology was applied to reveal the entire bacterial composition of all semolina samples. T…

Microbial Activation of Wooden Vats Used for Traditional Cheese Production and Evolution of Neoformed Biofilms.

ABSTRACT Three Lactococcus lactis subsp. cremoris strains were used to develop ad hoc biofilms on the surfaces of virgin wooden vats used for cheese production. Two vats (TZ) were tested under controlled conditions (pilot plant), and two vats (TA) were tested under uncontrolled conditions (industrial plant). In each plant, one vat (TA1 and TZ1) was used for the control, traditional production of PDO Vastedda della Valle del Belìce (Vastedda) cheese, and one (TA2 and TZ2) was used for experimental production performed after lactococcal biofilm activation and the daily addition of a natural whey starter culture (NWSC). Microbiological and scanning electron microscopy analyses showed differenc…

Identificazione di una nuova variante alla κ-caseina nella razza caprina Girgentana

La κ-caseina è la lattoproteina che determina la grandezza e la funzione specifica delle micelle nel latte e la sua idrolisi, chimosina dipendente, è responsabile della coagulazione del latte stesso. Il gene della κ-caseina comprende 5 esoni. Ad oggi sono state identificate 16 varianti alleliche, di cui 13 sono varianti proteiche e 3 mutazioni silenti, per un totale di 15 siti polimorfici.Lo scopo di questo lavoro è stato la caratterizzazione dell’esone 4 del gene della κ-caseina nella razza caprina Girgentana. Un nuova variante alleliche, denominata X, è stata riscontrata con una frequenza relativamente bassa (0,04).

Genome-Wide Patterns of Homozygosity Reveal the Conservation Status in Five Italian Goat Populations.

The application of genomic technologies has facilitated the assessment of genomic inbreeding based on single nucleotide polymorphisms (SNPs). In this study, we computed several runs of homozygosity (ROH) parameters to investigate the patterns of homozygosity using Illumina Goat SNP50 in five Italian local populations: Argentata dell’Etna (N = 48), Derivata di Siria (N = 32), Girgentana (N = 59), Maltese (N = 16) and Messinese (N = 22). The ROH results showed well-defined differences among the populations. A total of 3687 ROH segments &gt

Genome-wide association study between CNVs and milk production traits in Valle del Belice sheep.

Copy number variation (CNV) is a major source of genomic structural variation. The aim of this study was to detect genomic CNV regions (CNVR) in Valle del Belice dairy sheep population and to identify those affecting milk production traits. The GO analysis identified possible candidate genes and pathways related to the selected traits. We identified CNVs in 416 individuals genotyped using the Illumina OvineSNP50 BeadChip array. The CNV association using a correlation-trend test model was examined with the Golden Helix SVS 8.7.0 tool. Significant CNVs were detected when their adjusted p-value was <0.01 after false discovery rate (FDR) correction. We identified 7,208 CNVs, which gave 365 C…

Monitoring of wheat lactic acid bacteria from the field until the first step of dough fermentation.

The present work was carried out to retrieve the origin of lactic acid bacteria (LAB) in sourdough. To this purpose, wheat LAB were monitored from ear harvest until the first step of fermentation for sourdough development. The influence of the geographical area and variety on LAB species/strain composition was also determined. The ears of four Triticum durum varieties (Duilio, Iride, Saragolla and Simeto) were collected from several fields located within the Palermo province (Sicily, Italy) and microbiologically investigated. In order to trace the transfer of LAB during the consecutive steps of manipulation, ears were transformed aseptically and, after threshing, milling and fermentation, s…

Identification of two new alleles at k-casein (CSN3) gene in Girgetana goat breed

In the milk of ruminants, more than 95% of proteins are synthesized by six structural genes, four caseins and two whey proteins. The κ-casein fraction plays an important role in the formation, stabilization and aggregation on casein micelles and thus affects technological and nutritional properties of milk. Goat κ-casein (CSN3) gene comprises five exons with the mRNA coding region for mature protein spanning from exon 3 to exon 4. The κ-casein locus has been widely investigated and several genetic variants have been found in different goat breeds. At protein level, AIEF and BIEF groups corresponding to isoelectric point 5.53 and 5.78, respectively, have been identified. In this study, exon …

Biological diversity of the microbial film associated to the central-Mediterranean Dendropoma cristatum (Biondi, 1859) reefs.

Microbial films may provide physical and bio-chemical cues which positively affect the settlement dynamic of a variety of benthic marine organisms, driving the development of ecosystems. Also for the Mediterranean intertidal reef-builder Dendropoma cristatum (Biondi, 1859), biofilm maturity has been found to enhance the settlement pattern. However, the microbial diversity associated with these bioconstructions has never been described. This study investigates the D. cristatum reef bacterial assemblage composition and temporal evolution in two localities in the northwest of Sicily. Biological diversity of the reef-associated biofilm and of 3 progressively older biofims obtained on artificial…

Genomic structural diversity in local goats: Analysis of copy-number variations

Copy-number variations (CNVs) are one of the widely dispersed forms of structural variations in mammalian genomes, and are present as deletions, insertions, or duplications. Only few studies have been conducted in goats on CNVs derived from SNP array data, and many local breeds still remain uncharacterized, e.g., the Sicilian goat dairy breeds. In this study, CNV detection was performed, starting from the genotypic data of 120 individuals, belonging to four local breeds (Argentata dell&rsquo

Identification of Copy Number Variations and Genetic Diversity in Italian Insular Sheep Breeds

Simple Summary The aim of this work is to identify CNVs and perform a CNV-based population genetics analysis in five Italian sheep breeds (Barbaresca, Comisana, Pinzirita, Sarda, and Valle del Belìce). We identified 10,207 CNVs and 365 CNV regions (CNVRs). The different approaches used to disclose the genetic relationship showed that the five breeds tend to cluster in distinct groups. Gene enrichment was described for the 37 CNVRs selected considering the top 10%. Gene Ontology analysis showed that several of these genes are involved in lipid metabolism, immune response, and the olfactory pathway. CNVs represent valuable molecular resources to provide useful information for separating the p…

Genetic characterization of the Mascaruna goat, a Sicilian autochthonous population, using molecular markers

The aim of this work was to characterize a Sicilian autochthonous goat population using microsatellite markers and genetic polymorphisms at the casein genes. In order to investigate the genetic structure of the Mascaruna goat, a total of 60 (20 Girgentana, 20 mixed populations, and 20 Mascaruna) individuals were analyzed, using a panel of 18 microsatellite markers. Moreover, the Mascaruna goats were genotyped at casein loci using several molecular techniques. Based on the genetic structure at casein genes, the Mascaruna goat was similar to most goat breeds from the Mediterranean area, which are characterized by the predominance of strong alleles. The low value of genetic differentiation amo…

Use of microsatellite markers for genetic traceability of Girgentana dairy products

Genetic traceability is based on the identification of both animals and their products through the study of DNA. With the goal of developing a genetic traceability system for dairy products, the aim of this study was to identify specific microsatellite markers able to discriminate among the most important Sicilian dairy goat breeds, in order to detect possible adulteration in Girgentana dairy products. A total of 20 microsatellite markers were analyzed on a total of 338 individual samples from Girgentana (GIR), Maltese (MAL) and Derivate di Siria (DdS) goat breeds. The first step was to identify breed specific microsatellite markers that can be used for the traceability of dairy products. A…

A pipeline for variants discovery using next-generation DNA sequencing data

Recent advances in next generation sequencing (NGS) technology provide a cost-effective approach to large-scale resequencing of livestock samples in order to study several biological phenomena. NGS produces millions of short DNA sequences that require an unbiased way to make possible comprehensive searches for variation to identify putative causative mutations for economically important traits. The aim of this work was to present a bioinformatics pipeline analysis for variants discovery in ovine genome. A total of 30 individuals belonging to Valle del Belice dairy ewes was used for whole genome sequencing of pooled libraries prepared using Illumina Nextera Kit. Paired-end sequencing was car…

Genome structure in Sicilian cattle breeds

Genomic technologies provide background information concerning genome structure in domestic animals. The aim of this work was to investigate the genetic structure and the patterns of linkage disequilibrium (LD) in two Sicilian local cattle breeds, Cinisara and Modicana. Genotypes from animals of Italian Holstein breed were also used to investigate the relationship among breeds. Structure software was used to analyze the genetic structure and assign the individuals to each cluster. The genetic relationship between individuals was estimated by Principal Components Analysis (PCA) of genetic distance. A standard descriptive LD parameter (r2) was obtained between adjacent SNPs and for all pairwi…

Genome-wide detection of copy-number variations in local cattle breeds

The aim of the present study was to identify copy-number variations (CNVs) in Cinisara (CIN) and Modicana (MOD) cattle breeds on the basis of signal intensity (logR ratio) and B allele frequency of each marker, using Illumina’s BovineSNP50K Genotyping BeadChip. The CNVs were detected with the PennCNV and SVS 8.7.0 software and were aggregated into CNV regions (CNVRs). PennCNV identified 487 CNVs in CIN that aggregated into 86 CNVRs, and 424 CNVs in MOD that aggregated into 81 CNVRs. SVS identified a total of 207 CNVs in CIN that aggregated into 39 CNVRs, and 181 CNVs in MOD that aggregated into 41 CNVRs. The CNVRs identified with the two softwares contained 29 common CNVRs in CIN and 17 co…

In-Depth Investigation of the Safety of Wooden Shelves Used for Traditional Cheese Ripening

ABSTRACT The main goal of this research was to characterize the bacterial diversity of the wooden boards used for aging traditional Sicilian cheeses and to evaluate whether pathogenic bacteria are associated with these surfaces. Eighteen cheese dairy factories producing three traditional cheese typologies (PDO Pecorino Siciliano, PDO Piacentinu Ennese, and Caciocavallo Palermitano) were selected within the region of Sicily. The wooden shelf surfaces were sampled by a destructive method to detach wood splinters as well as by a nondestructive brushing to collect microbial cells. Scanning electron microscopy showed the presence of almost continuous bacterial formations on the majority of the s…

Genetic variability of Girgentana goat breed using molecular markers

Assessing genetic diversity of local breeds through the information provided by neutral molecular markers, allows determination of their extinction risk and to design strategies for their management and conservation. The aim of this work was to quantify the levels of genetic variability in Girgentana goat breed using microsatellite markers and genetic polymorphisms at the casein genes. A total of 264 individuals were genotyped for 20 microsatellites, selected as suggested by ISAG and FAO. Moreover, the same individuals were characterized for the casein loci (CSN1S1, CSN2, CSN1S2, and CSN3) using PCR and sequencing protocols. Several genetic diversity indexes were estimated. A total of 130 a…

Microbiological analysis and metagenomic profiling of the bacterial community of an anthropogenic soil modified from typic haploxererts

This work aimed to characterize the microbial communities of an anthropogenic soil originating from application of pedotechniques to Vertisols in a Mediterranean environment. Bare soil profiles were sampled at three depths (0–10 cm, 10–30 cm, and 30–50 cm) and compared with the original soil not transformed at the same depths. The anthropogenic soils were characterized by a higher CaCO3 concentration (360–640 g/kg) than control soil (190–200 g/kg), while an opposite trend was registered for clay, where control soil showed a higher concentration (465 g/kg on average) than anthropogenic soil (355 g/kg on average). Organic carbon content was much higher in the unt…

A combined genome-wide approach identifies a new potential candidate marker associated with the coat color sidedness in cattle

Abstract Coat color is one of the most important phenotypic features in livestock breeds. Cinisara is a local cattle breed generally of uniform black color which occasionally presents a particular phenotype, with animals typically display a white band along their spine, from the head to the tail, and on the ventral line (color sidedness). Therefore, this breed provides an ideal model to study the genetic components underlying phenotypic variation in coat color. A total of 63 animals, ten with sidedness phenotype and 53 with uniform black color were genotyped with Illumina Bovine 50 K. The comparison among genome-wide association study and FST analysis revealed a single nucleotide polymorphi…

Quantitative determination of casein genetic variants in goat milk: Application in Girgentana dairy goat breed

The study was conducted to develop a high-performance liquid chromatographic (HPLC) method to quantify casein genetic variants (αs2-, β-, and κ-casein) in milk of homozygous individuals of Girgentana goat breed. For calibration experiments, pure genetic variants were extracted from individual milk samples of animals with known genotypes. The described HPLC approach was precise, accurate and highly suitable for quantification of goat casein genetic variants of homozygous individuals. The amount of each casein per allele was: αs2-casein A = 2.9 ± 0.8 g/L and F = 1.8 ± 0.4 g/L; β-casein C = 3.0 ± 0.8 g/L and C1 = 2.0 ± 0.7 g/L and κ-casein A = 1.6 ± 0.3 g/L and B = 1.1 ± 0.2 g/L. A good correl…

Detection of genomic regions underlying milk production traits in Valle del Belice dairy sheep using regional heritability mapping

The aim of this study was to identify genomic regions underlying milk production traits in the Valle del Belice dairy sheep using regional heritability mapping (RHM). Repeated measurements for milk yield (MY), fat percentage and yield (F% and FY) and protein percentage and yield (P% and PY), collected over a period of six years (2006-2012) on 481 Valle del Belice ewes, were used for the analysis. Animals were genotyped with the Illumina 50k SNP chip. Variance components, heritabilities, and repeatability within and across lactations were estimated, fitting parity, litter size, season of lambing, and fortnights in milk, as fixed; and additive genetic, permanent environment within and across …

Weighted Single-Step Genome-Wide Association Study Uncovers Known and Novel Candidate Genomic Regions for Milk Production Traits and Somatic Cell Score in Valle del Belice Dairy Sheep

The objective of this study was to uncover genomic regions explaining a substantial proportion of the genetic variance in milk production traits and somatic cell score in a Valle del Belice dairy sheep. Weighted single-step genome-wide association studies (WssGWAS) were conducted for milk yield (MY), fat yield (FY), fat percentage (FAT%), protein yield (PY), protein percentage (PROT%), and somatic cell score (SCS). In addition, our aim was also to identify candidate genes within genomic regions that explained the highest proportions of genetic variance. Overall, the full pedigree consists of 5534 animals, of which 1813 ewes had milk data (15,008 records), and 481 ewes were genotyped with a …

Variation of proteomic profile during lactation in Girgentana goat milk: a preliminary study

The knowledge of milk proteome has been greatly enhanced by technological advances in the proteomics field as the use of the two-dimensional differential in-gel electrophoresis, a gel-based approach which allowed the analysis of proteins from complex mixtures and the comparing of several protein samples in the same experiment. The aim of this study was to characterise the whole milk proteomic profile in Girgentana dairy goat breed by two-dimensional differential in-gel elecrophoresis. The obtained representative 2D whole milk proteomic map showed a general picture of the protein distributions over the pH 3–10 NL including about 100 spots, most of them organised like a spot train. Among diff…

Full-length sequencing and identification of novel polymorphisms in the ACACA gene of Valle del Belice sheep breed

The essential role of the acetyl-CoA carboxylase (ACACA) enzyme in milk fatty acid (FA) synthesis suggests that it may be responsible for the phenotypic variability observed in milk. Before attempting association analyses between this gene and/or enzyme and phenotypic traits, a study on the genetic variability within this locus is required. The aim of this work was to sequence the entire coding region of ACACA gene in Valle del Belice sheep breed to identify polymorphic sites. A total of 51 coding exons of ACACA gene were sequenced in 32 individuals of Valle del Belice sheep breed. Sequencing analysis and alignment of obtained sequences showed the presence of 23 polymorphic sites. The most …

Biodiversity and dairy traits of indigenous milk lactic acid bacteria grown in presence of the main grape polyphenols

ABSTRACT The present work was developed to select lactic acid bacteria (LAB) to be used as starter cultures in functional cheese production. The indigenous milk LAB populations were isolated from fermented raw ewes’ milks (four bulks) added with 0.5 mg/mL of nine polyphenols commonly found in winery by-products. After 48 h of fermentation, all milks were characterized by an increase of LAB levels of about 3–4 Log cycles. All different colonies were purified and characterized for the main physiological and biochemical traits and then differentiated genetically at strain level and identified. Ten species belonging to the LAB genera Lactobacillus, Streptococcus, Enterococcus, Leuconostoc and L…

Application of microsatellite markers as potential tools for traceability of Girgentana goat breed dairy products

In livestock, breed assignment may play a key role in the certification of products linked to specific breeds. Traceability of farm animals and authentication of their products can contribute to improve breed profitability and sustainability of animal productions with significant impact on the rural economy of particular geographic areas and on breed and biodiversity conservation. With the goal of developing a breed genetic traceability system for Girgentana dairy products, the aim of this study was to identify specific microsatellite markers able to discriminate among the most important Sicilian dairy goat breeds, in order to detect possible adulteration in Girgentana dairy products. A tot…

Genome-wide association study for milk production traits in Valle del Belice sheep

Over the last years, high-throughput technologies have provided the opportunity to explore the genomes of livestock species to identify regions influencing traits of economic interest. Genome wide association studies (GWAS) have been widely used to disentangle the genetic variation in complex phenotypes, such as milk production traits in sheep. The study presented here investigated the possible association of genome-wide SNPs and breeding values for milk production traits in the Valle del Belice sheep. The studied population consisted of a total of 481 ewes belonging to 9 half-sib families with available records for milk yield (MY), protein and fat percentage (P% and F%, respectively). All …

Conservation status and historical relatedness of Italian cattle breeds

Background In the last 50 years, the diversity of cattle breeds has experienced a severe contraction. However, in spite of the growing diffusion of cosmopolite specialized breeds, several local cattle breeds are still farmed in Italy. Genetic characterization of breeds represents an essential step to guide decisions in the management of farm animal genetic resources. The aim of this work was to provide a high-resolution representation of the genome-wide diversity and population structure of Italian local cattle breeds using a medium-density single nucleotide polymorphism (SNP) array. Results After quality control filtering, the dataset included 31,013 SNPs for 800 samples from 32 breeds. Ou…

Association study between beta-defensin gene polymorphisms and mastitis resistance in Valle del Belice dairy sheep breed

Abstract Mastitis is generally caused by bacteria, and it is the most common disease in livestock species. Defensins are peptides with a broad spectrum of antimicrobial activity and β-defensin genes have been studied in several livestock species due to their important role in the innate immune response. The aim of this study was to establish an association between polymorphisms in the β-defensin 1 and 2 genes and mastitis resistance in the Valle del Belice dairy sheep. Data consisted of 1855 and 2804 observations for case and control group, respectively. Six single nucleotide polymorphisms and seven haplotypes were selected for association studies with mastitis. In particular, polymorphism …

Genomic inbreeding estimation in small populations: evaluation of runs of homozygosity in local cattle breeds

The availability of high throughput genotyping has facilitated the quantification of inbreeding by genomic markers in farm animals. Run of homozygosity (ROH) are contiguous lengths of homozygous genotypes and represent an estimate of the degree of autozygosity at genome-wide level. The current study aims to quantify the genomic inbreeding derived from ROH (FROH) in three Italian local cattle breeds. Individuals of Cinisara (71), Modicana (72), Reggiana (168) were genotyped with the 50K v2 Illumina BeadChip. Genotypes from 96 animals of the Italian Holstein cattle breed were included in the analysis. The following criteria were used to define a ROH: two missing SNPs; one heterozygous SNP; mi…

Genome wide Copy Number Variation (CNV) detection in Cinisara cattle breed

Copy Number Variations (CNVs) are classes of polymorphic genomic regions including deletions, duplications and insertions of DNA fragments from at least 0.5 kb up to several Mb. CNV represents an important source of genetic variability that provides genomics structural information complementary to the single nucleotide polymorphism (SNP) data. Some CNVs have been shown to be important in both normal phenotypic variability and disease susceptibility in livestock. Several approaches to identify CNVs including FISH, aCGH, SNP array or NGS, were proposed and among these SNP genotyping is relatively low cost, high-throughput and high coverage method. The aim of this study was to identify the CNV…

MOESM2 of Genome-wide scan for runs of homozygosity identifies potential candidate genes associated with local adaptation in Valle del Belice sheep

Additional file 2: Figure S2. Total number of runs of homozygosity (ROH) longer than 1Â Mb and total length of genome (Mb) covered by ROH segments per individual. Observed (black) vs simulated (red) data.

Production of the Sicilian distillate “ Spiritu re fascitrari ” from honey by-products: An interesting source of yeast diversity

The “Spiritu re fascitrari” (SRF) is a typical Sicilian distillate obtained from the by-products of traditional process of honey production. Although some alcoholic fermentation of honey based products have been described, the present research represents the first investigation on the yeast ecology and the physico-chemical characteristics of honey by-products subjected to an alcoholic fermentation followed by distillation. All samples collected during manufacturing process were analysed for the count of total, osmophilic and osmotolerant yeasts. The honeycombs and equipment surfaces showed the presence of yeasts that was 1.7 and 1.1 Log (CFU/mL), respectively. After enrichment, yeast popula…

Activation of wooden vats with selected lactic acid bacteria for the year-round production of traditional Vastedda-like cheese

Vastedda is a pasta filata cheese made from raw ewes’ milk without the addition of starter cultures. Cheese production is carried out in wooden vats characterized by microbial biofilms hosting lactic acid bacteria (LAB) responsible for the acidification of the curds. Vastedda cheese was traditionally produced during the summer season, but it is currently produced throughout the year. In order to minimize production variability, several LAB were isolated from Vastedda PDO cheeses, characterized for their technological potential (Gaglio et al., 2014a) and selected to obtain the best strain combination for the production (Gaglio et al., 2014b). In this study, the multiple strain culture compos…

Genome-Wide Association Study Identifies New Candidate Markers for Somatic Cells Score in a Local Dairy Sheep

In the Mediterranean basin countries, the dairy sheep production is usually based on local breeds, which are very well-adapted to their production systems and environments and can indeed guarantee income, employment, and economic viability in areas where production alternatives are scarce or non-existent. Mastitis is still one of the greatest problems affecting commercial milk production. However, genetic evaluation of mastitis is particularly difficult because of its low heritability and the categorical nature of the trait. The aim of this study was to identify genomic regions putatively associated with somatic cells count (SCC) in the local economically important Valle del Belice sheep br…

Refining the genetic structure and relationships of European cattle breeds through meta-analysis of worldwide genomic SNP data, focusing on Italian cattle

AbstractThe availability of genotyping assays has allowed the detailed evaluation of cattle genetic diversity worldwide. However, these comprehensive studies did not include some local European populations, including autochthonous Italian cattle. In this study, we assembled a large-scale, genome-wide dataset of single nucleotide polymorphisms scored in 3,283 individuals from 205 cattle populations worldwide to assess genome-wide autozygosity and understand better the genetic relationships among these populations. We prioritized European cattle, with a special focus on Italian breeds. Moderate differences in estimates of molecular inbreeding calculated from runs of homozygosity (FROH) were o…

Diversity and technological potential of lactic acid bacteria of wheat flours

Abstract Lactic acid bacteria (LAB) were analysed from wheat flours used in traditional bread making throughout Sicily (southern Italy). Plate counts, carried out in three different media commonly used to detect food and sourdough LAB, revealed a maximal LAB concentration of approximately 4.75 Log CFU g−1. Colonies representing various morphological appearances were isolated and differentiated based on phenotypic characteristics and genetic analysis by randomly amplified polymorphic DNA (RAPD)-PCR. Fifty unique strains were identified. Analysis by 16S rRNA gene sequencing grouped the strains into 11 LAB species, which belonged to six genera: Enterococcus, Lactobacillus, Lactococcus, Leucono…

Italian Goat Consortium: a collaborative project to study the Italian caprine biodiversity.

The Italian Goat Consortium (IGC), joined the effort of many Universities and Research Institutes, in a comprehensive study of the Italian goat population genetic makeup using a medium density (54K) SNPs chip. Currently IGC has genotyped more than 1,000 animals from more than 30 goat breeds and populations from all Italian geographical and agroecological areas of goat rearing. The aim of this work is to obtain a clear picture of the Italian caprine biodiversity, to reconstruct the ancestry, to disentangle the genetic background and to assess the relationships among and within the investigated breeds. To date, the IGC dataset includes about 50 million genotypes. The data were quality checked…

Analysis of acetyl-CoA carboxylase gene polymorphisms in Valle del Belice dairy breeds

Acetyl-CoA carboxylase is the flux-determining enzyme in the regulation of fatty acid synthesis within animal tissues. The expression of the mammary gland isoform of this enzyme, during lactation, is regulated by the acetyl-CoA carboxylase-α (ACACA) gene. The ovine ACACA gene, with 51 coding exons, is responsible for phenotypic variability observed in milk fat content and milk fatty acids (FAs) composition. However, before attempting association analyses between this enzyme and phenotypic traits of interest, a study on the genetic variability within this locus is required. The aim of this work was to sequence the encoding region of ACACA gene in Valle del Belice dairy ewes in order to ident…

Polymorphisms at β-defensin genes and mastitis in Valle del Belice dairy sheep: a case control study

Mastitis is the most common and costly pathology affecting dairy animals. The identification of genetic markers that allow the inclusion of mastitis resistance in selection programs would help to reduce the costs due to the disease and the use of antibiotics. In recent years, β-defensin genes have been studied in several domestic species due to their important role in the innate immune response. The aim of this study was to determine whether β-defensin genes variability, β-defensin 1 (SBD1) and β-defensin 2 (SBD2), could distinguish between infected cases and healthy controls in Valle del Belice dairy sheep. Somatic cell count (SCC) is a powerful tool to identify animals with intramammary i…

Genome wide linkage disequilibrium and genetic structure in Sicilian dairy sheep breeds

Background The recent availability of sheep genome-wide SNP panels allows providing background information concerning genome structure in domestic animals. The aim of this work was to investigate the patterns of linkage disequilibrium (LD), the genetic diversity and population structure in Valle del Belice, Comisana, and Pinzirita dairy sheep breeds using the Illumina Ovine SNP50K Genotyping array. Results Average r2 between adjacent SNPs across all chromosomes was 0.155 ± 0.204 for Valle del Belice, 0.156 ± 0.208 for Comisana, and 0.128 ± 0.188 for Pinzirita breeds, and some variations in LD value across chromosomes were observed, in particular for Valle del Belice and Comisana breeds. Ave…

MOESM3 of Genome-wide scan for runs of homozygosity identifies potential candidate genes associated with local adaptation in Valle del Belice sheep

Additional file 3: Figures S3, S4, S5, S6. Plot of SNP occurrences (%) in ROH against the genomic regions of QTL for OAR chromosomes with the highest inbreeding coefficient (OAR 2, 4, 11, 23).

Shelf life evaluation of fresh-cut red chicory subjected to different minimal processes

Abstract Microbiological, chemical and physical parameters of minimally processed red chicory (Cichorium intybus L.) subjected to two different transformation processes were investigated. A classic ready-to-eat (RTE) process (P1) and a production without cutting (P2) were monitored during refrigerated (4 °C) storage (15 d). Total mesophilic microorganisms, total psychrotrophic microorganisms and pseudomonads were detected at the highest cell densities in all samples. Presumptive Pseudomonas population dominated the cultivable microbial community of RTE red chicory and were characterized genetically. Twenty-two randomly amplified polymorphic DNA (RAPD) types were investigated by 16S rRNA gen…

Distribution of functional variants within Runs of Homozygosity in four Italian cattle breeds

Several methods have been used to identify genomic regions subjected to selective sweeps. An alternative method can make use of runs of homozygosity (ROH), defined as stretches of homozygous regions in a genome. The aim of the present study was to detect selective sweeps using ROH in four cattle breeds. Individuals of Cinisara (71), Modicana (72),Reggiana (168) and Italian Holstein (96) were genotyped with the Illumina Bovine50SNP v2 BeadChip. To identify genomic regions that were most commonly associated with ROH within each breed, the percentage of occurrences of a single nucleotide polymorphism (SNP) in ROH was calculated across animals. The genomic regions most commonly associated with …

Linkage disequilibrium and genetic diversity estimation in three Sicilian autochthonous dairy sheep breeds

Understanding genetic structure is essential for achieving genetic improvement through genome-wide association studies, genomic selection and the dissection of quantitative traits. In the present study, we used the OvineSNP50K BeadChip to characterize LD, identify haplotype blocks and to analyze genetic diversity in the Valle del Belice (VDB), Comisana (COM) and Pinzirita (PIN) dairy sheep breeds. LD between adjacent SNPs and for all pairwise combinations of SNPs on each chromosome was measured using r2. Haplotype blocks were estimated using D’ based method. Genetic diversity and Principal Component Analysis (PCA) were calculated using PLINK. Small differences in average LD value for adjace…

In vivo application and dynamics of lactic acid bacteria for the four-season production of Vastedda-like cheese.

article Twelve lactic acid bacteria (LAB), previously selected in vitro (Gaglio et al., 2014), were evaluated in situ for their potential to act as starter cultures for the continuous four-season production of Vastedda-like cheese, made with raw ewes' milk. The strains belonged to Lactobacillus delbrueckii, Lactococcus lactis subsp. cremoris, Leuconostoc mesenteroides subsp. mesenteroides and Streptococcus thermophilus .L AB werefirst inoculated in multiple-strain combinations on the basis of their optimal growth temperatures in three process conditions which differed for milk treatment and medium for strain development: process 1, growth of strains in the optimal synthetic media and pasteu…

Genome-wide association study for milk production traits in an economically important local dairy sheep breed

In this study, we conducted a genome-wide association study (GWAS) for five milk production traits in the Valle del Belice sheep. Repeated measurements for milk yield (MY), fat percentage and yield (F% and FY) and protein percentage and yield (P% and PY) on 481 ewes, were available for the analysis. The animals were genotyped using the Illumina Ovine 50k BeadChip. Weighted deregressed breeding values (DEBVw) were used as phenotypes for GWAS analysis. A total of 23 genome-wide significant SNPs were identified: 3 associated with MY, 9 with FY, and 11 with P%. Several SNPs mapped within known candidate genes or previously reported QTL for milk production traits in livestock species. Additional…

MOESM1 of Genome-wide scan for runs of homozygosity identifies potential candidate genes associated with local adaptation in Valle del Belice sheep

Additional file 1: Figure S1. Mean sum of runs of homozygosity (ROH) per animal estimated within four different generation categories. ROH were mapped according to their genetic positions (i.e. linkage map positions). ROH length (l cM) within each category was determined using 100/2 g, replacing g with the number of generations of interest.

Molecular characterisation of k-casein gene in Girgentana dairy goat breed and identification of two new alleles

The k-casein fraction plays an important role in the formation, stabilisation and aggregation on casein micelles and thus affects technological and nutritional properties of milk. In this study, exon 4 of k-casein (CSN3) gene was sequenced and analysed in Girgentana goat breed. Analyses of the obtained sequences showed the presence of A, B, D, and G known alleles and two new genetic variants, named D’ and N. The new D’ allele differs from D in one transition, G284→A284, which did not cause amino acid change. The new N allele differs from A in five single nucleotide polymorphisms (SNPs): T245/C245, G284/A284, G309/A309, G471/A471 and T591/C591, while it differs from C in one transition, i.e.…

The Sicilian cattle breeds in a global context: genome-wide relationship with other worldwide cattle

Genomic technologies, such as high-throughput genotyping based on Single Nucleotide Polymorphism (SNP) arrays, provide background information concerning genome structure in domestic animals. The aims of this study were to cluster animals, to explore the relationships among and within breeds, and to place the Sicilian breeds, Cinisara and Modicana, in a global context. The Illumina Bovine SNP50K v2 BeadChip genotyping data from 144 animals of Sicilian breeds and from other 1,543 animals belonging to 134 domesticated bovid breeds (DRYAD) were used. These breeds arose from 3 domesticated (sub)species: Bos javanicus, B. taurus indicus, and B. t. taurus. Principal Component Analysis (PCA) genera…

MOESM4 of Genome-wide scan for runs of homozygosity identifies potential candidate genes associated with local adaptation in Valle del Belice sheep

Additional file 4: Figure S7. Plot of SNP occurrences in ROH against recombination rate. Recombination rate is the solid red line and the occurrence of a SNP in a ROH is represented by blue dots. (A) OAR2, (B) OAR4, (C) OAR11, and (D) OAR23. Recombination rate (cM/Mb) was estimated every 500Â kb.

Genome‐wide survey on three local horse populations with a focus on runs of homozygosity pattern

Purosangue Orientale Siciliano, Sanfratellano and Siciliano represent the Sicilian equine genetic resource. This study aimed to investigate the genetic diversity, population structure and the pattern of autozygosity of Sicilian horse populations using genome-wide single-nucleotide polymorphism (SNP) data generated with the Illumina Equine SNP70 array. The genotyping data of 17 European and Middle East populations were also included in the study. The patterns of genetic differentiation, model-based clustering and Neighbour-Net showed the expected positioning of Sicilian populations within the wide analysed framework and the close connections between the Purosangue Orientale Siciliano and the…

MOESM5 of Genome-wide scan for runs of homozygosity identifies potential candidate genes associated with local adaptation in Valle del Belice sheep

Additional file 5: Table S1. List of 239 SNPs considered as candidate markers under directional selection in the Valle del Belice sheep breed.

MOESM6 of Genome-wide scan for runs of homozygosity identifies potential candidate genes associated with local adaptation in Valle del Belice sheep

Additional file 6: Table S2. List of 107 potential candidate genes under directional selection in the Valle del Belice sheep breed.

Variation of proteomic profile during lactation in Girgentana goat milk: a preliminary study

The knowledge of milk proteome has been greatly enhanced by technological advances in the proteomics field as the use of the two-dimensional differential in-gel electrophoresis, a gel-based approach which allowed the analysis of proteins from complex mixtures and the comparing of several protein samples in the same experiment. The aim of this study was to characterise the whole milk proteomic profile in Girgentana dairy goat breed by two-dimensional differential in-gel elecrophoresis. The obtained representative 2D whole milk proteomic map showed a general picture of the protein distributions over the pH 3–10 NL including about 100 spots, most of them organised like a spot train. Among diff…