Κ‐CN gene polymorphism in Nili‐ravi buffalo,Achai and Sahiwal cattle of Pakistan

Kappa‐casein (κ‐CN) is the subtype of casein protein, an important constituent of bovine milk protein. The current study was undertaken to investigate the genetic polymorphism in κ‐CN gene of Nili‐ravi buffalo, Achai and Sahiwal cattle of Pakistan using polymerase chain reaction–restriction fragment length polymorphism (PCR‐RFLP) technique. The Nili‐ravi buffalo was found to be monomorphic (genotype BB only) for κ‐CN gene. Achai cattle were polymorphic for κ‐CN (having three genotypes AA, AB and BB) with a frequency of 0.70, 0.18 and 0.12, respectively, while in Sahiwal cattle, both the genotypes AA and AB were found with genotypic frequencies of 0.92 and 0.08, respectively. The presence of genotype BB in Achai cattle is surprising as it is absent in most of the cattle breeds worldwide.     

Genetic variability in the kappa‐casein gene in Sahiwal,Cholistani and Red Sindhi cattle breeds

Polymorphism of kappa‐casein (κ‐CN) gene in three Bos indicus cattle breeds was investigated using a polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) technique. Only genotypes AA and AB were observed, and no BB, AE, BE, EE, AC or BC genotypes were detected in the three cattle breeds. In the Sahiwal, Cholistani and Red Sindhi breeds, the frequencies of the allele A were 0.69, 0.90 and 0.86, and the frequencies of the allele B were 0.31, 0.10 and 0.14, respectively. The Sahiwal breed was found to have a higher frequency of the AB genotype as compared to the Cholistani and Red Sindhi breeds. The frequency of the κ‐CN alleles noted in the three cattle breeds was similar to those observed in other cattle breeds of B. indicus origin.     

The influence of time elapsed after milking and lactation stage on the lactoperoxidase system of milk in Sahiwal cattle and Nili‐Ravi buffaloes

The effects of time elapsed after milking and lactation stage were assessed on the lactoperoxidase (LP) enzyme activity and concentrations of thiocyanate in Sahiwal cattle (N = 15) and Nili‐Ravi buffaloes (N = 15). The LP activity and thiocyanate concentrations decreased as the time elapsed after milking (P < 0.05) but the concentration of these components increased as the lactation advanced. The LP enzyme activity was sufficient until 12 h post milking but the thiocyanate concentration after 8 h decreased to an insufficient level to activate the LP System, hence additional thiocyanate would be beneficial from external sources. No difference was noted regarding the LPS activity in cattle and buffaloes at the time of collection and subsequently after storage. However, at the end of lactation, the thiocyanate concentration was lower in buffaloes than that in cattle. (P < 0.05).     

Determination of allele frequencies of growth hormone AluI polymorphism in Brown Swiss,Holstein, native East Anatolian Red and Turkish Grey breeds

A total of 177 cattle of four breeds were genotyped for the bovine growth hormone (BGH)‐AluI polymorphism by polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP). The genotype and gene frequencies for each breed were determined and tested to be in Hardy–Weinberg equilibrium. According to breeds, frequencies of allele L gene were 0.905 for Brown Swiss, 0.898 for Holstein, 0.976 for East Anatolian Red and 0.893 for Turkish Grey Breeds. The allele L was predominant and variant VV was not detected in the breeds studied. BGH‐AluI genotypes were found to be in equilibrium within and among breeds.     

Short communication: Stall occupancy behavior of Nili Ravi buffaloes (Bubalus bubalis) when first introduced to freestall housing

The aim of the present study was to assess stall occupancy of Nili Ravi buffaloes when first introduced to freestall housing. Thirty Nili Ravi buffaloes of different parity, weight, and pregnancy status were enrolled from the dairy farm of the University of Veterinary and Animal Sciences, Lahore, Pakistan. All of the enrolled animals had no previous experience with freestalls. The selected buffaloes were introduced to the freestall shed. During an adaptation period of 7 d, all buffaloes had 24 h of free access to lie down in 1 of 3 areas: (1) freestalls, (2) alleys, and (3) an outside open area. After the adaptation period, all buffaloes were housed indoors during nighttime (restricted period) to ensure that only freestalls or alleys were available as lying areas. An observer monitored animals at 0100 h and recorded the number of animals lying in freestalls, in alleys, and in the outside open area. The results indicated that during the adaptation period, herd-level freestall occupancy in buffaloes remained below 10%. During the restricted period, herd-level stall occupancy was 0, 13, 50, and 90% on d 1, 2, 5, and 17, respectively. Buffaloes with parity ≥3 took fewer days (6.6 ± 1.0; mean ± standard error of the mean) to occupy stalls than buffaloes with parity 2, 1, and heifers (13.2 ± 1.5, 16.7 ± 2.1, and 13.5 ± 2.1 d, respectively). Body weight and pregnancy status of buffaloes did not have any effect on stall occupancy. Buffaloes showed adjacent stall occupancy, filling opposite stalls first. The results indicated that older buffaloes occupied freestalls earlier than younger buffaloes. Our findings would help better manage buffaloes in freestall housing.     

Genetic polymorphism of kappa‐casein,growth hormone and prolactin genes in Turkish native cattle breeds

A total of 259 cattle of four Turkish native cattle breeds, East Anatolian Red (EAR), South Anatolian Red (SAR), Turkish Grey (TG) and Anatolian Black (AB), Holstein and Brown Swiss (BS) breeds were genotyped for kappa‐casein (CSN3), bovine growth hormone (GH1) and prolactin (PRL) polymorphism by the polymerase chain reaction and restriction length polymorphism (PCR‐RFLP). The degree of genetic differentiation between populations F_ST was calculated as 0.053 and was found to be significant (P < 0.001). According to the genetic distance values (Nei), the highest genetic difference was found between SAR and EAR in four Turkish native cattle breeds and this difference was significant.     

Evaluation of nutritional quality of fat obtained from different breeds of cow and buffalo with respect to fatty acids and in vitro digestibility

A comparative fatty acid analysis of raw milk and during in vitro digestion of milk from different breeds of cow (Gir, Sahiwal, Tharparkar, and Karan fries) and buffalo (Murrah) maintained under similar feeding regimes was conducted. Saturated fatty acids were higher in Murrah and Karan Fries, whereas monounsaturated fatty acids were higher in Gir, Sahiwal and Tharparkar. Based on the fatty acid profile, atherogenic, thrombogenic index and hypercholesterolemic saturated fatty acids were lower, and desired fatty acids were higher in milk from indigenous breeds than in crossbred and Murrah. The size of milk fat globules was also lower in indigenous cattle breeds, which has allowed for faster in vitro digestibility of milk fat.     

Role of TNF‐α polymorphism ‐308 in neurosensory irritation

Neurosensory cutaneous discomfort in response to topical products is common, yet the relationship between symptoms such as stinging and visible irritation is currently unclear. The presence of a polymorphism at position ‐308 on the TNF‐α gene has been associated with skin irritation, i.e., erythema, dryness. Individuals with a G to A transition (AA/GA genotypes) have a lower threshold to experimentally induced irritation than those with the wild type (G allele, GG genotype). We investigated the effect of this polymorphism on neurosensory irritation (NSI). DNA genotyping was used to determine the allele type amongst a population of health care workers. The neurosensory response to lactic acid and water on the nasolabial folds and hands was assessed using a quantitative lactic acid sting test. Both genotypes had a more intense response to lactic acid compared with water on the face. The AA/GA genotypes had directionally higher scores from lactic acid (P = 0.1) and significantly higher stinging intensities from water (P = 0.001) on the face. For the hands, stinging intensities were higher for lactic acid and water amongst the AA/GA genotypes (P = 0.03 and 0.006 respectively). NSI to lactic acid was significantly higher on the face than on the hands (P < 0.05). Our findings indicate that subjects with the A transition at position ‐308 on the TNF‐α gene experience more intense NSI with common ingredients, i.e., lactic acid and water, than those with the wild type. TNF‐α polymorphism ‐308 may account for some of the inter‐individual variability in response to skin care practices.     

Genetic variants of beta-lactoglobulin gene and its association with milk composition traits in riverine buffalo

A study was carried out to determine genetic variants of beta-lactoglobulin gene and to explore associations between these and milk composition traits in riverine buffalo. Single strand conformation polymorphism was employed to detect the genetic variants of the gene. Two fragments of this gene i.e. 119 bp of exon I and 400 bp spanning exon IV and intron IV were included in the study. For 119 bp fragment, three alleles namely, A, B and C were observed in all the buffalo breeds whereas four alleles (A, B, C and D) were detected for 400 bp fragment. The frequency distribution of alleles was different in different breeds of buffaloes for both the fragments. For exon I fragment, the milk composition traits such as total SNF, protein, solid, fat and whey protein yield were found to be significantly (P<0.05) associated with genotypes in Murrah and Bhadawari buffalo whereas in Mehsana breed genotypes were significantly (P<0.05) co-related with total SNF, solid and fat yield. Genotypes of 400 bp fragment, only total fat yield in Mehsana buffalo was found to be significantly (P<0.05) associated with genotypes.     

Energy and protein requirements of growing Nili‐Ravi buffalo heifers in tropical environment

Thirty Nili‐Ravi buffalo heifers (average initial weight 119 kg) were used to assess energy and protein requirements for maintenance and growth. The calves were randomly divided into six equal groups and were fed six diets consisting of three levels of energy (55%, 60% and 65% total digestible nutrient—TDN) in ration and two levels of protein (10% and 12% crude protein—CP) in ration in factorial arrangement ad libitum until they attained 400 kg body weight. Energy and protein requirements at different stages of growth were estimated by partitioning of the metabolizable energy (ME), CP and metabolizable protein (MP) intake for maintenance and body weight gain by regression analysis. All of the multiple regression equations derived for different body weight ranges were highly significant (P < 0.01) and had high R² values. The ME (kJ), CP (g) and MP (g) requirements for maintenance at different body weights were 443–542, 5.89–9.38 and 4.03–6.30 kg^?0.75 d^?1. The respective values for average daily gain in body weight (ADG) were 26–53, 0.24–0.48 and 0.18–0.31 g^?1 ADG d^?1. The predicted requirements matched well with their corresponding actual intake values across the data range. Regressed values were used to develop new feeding standards for buffalo heifers. Copyright © 2007 Society of Chemical Industry     

Comparison of oil and fat supplementation on lactation performance of Nili Ravi buffaloes

The effects of feeding rumen-inert fat sources on production responses of lactating dairy cows have been well reported but less thoroughly described in lactating dairy buffalo. The objective of this study was to investigate the effect of oil and 2 different rumen-inert fat sources on dry matter intake, milk yield, milk composition, and milk fatty acid (FA) profile in Nili Ravi buffalo. Twelve multiparous mid-lactating Nili Ravi buffaloes received 4 treatments in a 4 × 4 Latin square design with a period length of 21 d. The treatments were (1) the basal diet without supplementation of oil or fats (CTRL), (2) the basal diet supplemented with canola oil (CO), (3) the basal diet supplemented with calcium salts of palm FA (Ca-FA), and (4) the basal diet supplemented with high palmitic acid (PA). Dry matter intake was decreased by 4.4% in the CO compared with Ca-FA and PA. Milk yield and milk fat yield were increased by 7.8 and 14.3%, respectively, in CO, Ca-FA, and PA compared with the CTRL. Milk fat content increased by 7.5%, whereas milk fat yield tended to increase with the supplementation of Ca-FA and PA compared with CO. No effect on milk yield and milk composition was observed in Ca-FA versus PA treatments. The yield of medium-chain FA was increased by Ca-FA and PA versus CO. The CO treatment increased the yield of long-chain FA compared with Ca-FA and PA treatments. Plasma glucose level was higher in CO, Ca-FA, and PA compared with the CTRL. In conclusion, feeding rumen-inert fats in the lactating buffalo diet proved to be a useful strategy to increase the 3.5% fat-corrected milk yield due to the higher milk fat content in this study.     

Comparative dry matter intake and nutrient utilisation efficiency in lactating cattle and buffaloes

An attempt was made to assess the comparative dry matter intake and nutrient utilisation efficiency of lactating buffaloes and cows based on results obtained from experimental feeding trials conducted in India. Data on dry matter (DM) intake, total digestible nutrient (TDN) intake, digestible crude protein (DCP) intake, body weight or metabolic body size (MBS), body weight change, milk yield or 4% fat‐corrected milk (FCM) yield, milk fat percentage and roughage/concentrate ratio in the diet of lactating buffaloes and cows were collected from published reports. The data were processed and analysed to assess the comparative dry matter intake and nutrient utilisation efficiency using suitable statistical analysis models. DM intake was significantly (P < 0.01) lower in buffaloes (2.57 kg DM per 100 kg body weight or 119.2 g kg^?1 MBS) than in cattle (3.09 kg DM per 100 kg body weight or 132.0 g kg^?1 MBS). Mean gross energetic efficiency (30.53 versus 27.83%; P < 0.01), gross protein efficiency (45.48 versus 37.06%; P < 0.01), net energetic efficiency (69.16 versus 64.10%; P < 0.05) and net protein efficiency (80.15 versus 59.59%; P < 0.001) were significantly higher in buffaloes than in cattle. Lactating buffaloes consumed significantly less (P < 0.001) protein (75.69 g DCP) and less (P < 0.01) energy (695.9 g TDN) than cows (93.89 g DCP, 774.8 g TDN) for production of 1 kg of 4% fat‐corrected milk. Buffaloes had higher energy and protein utilisation efficiencies as compared with cattle at similar fat‐corrected milk production level, plane of energy and protein nutrition, body size and body weight change. Buffaloes (1.189 kg DM kg^?1 4% FCM) consumed a similar (P > 0.05) amount of feed dry matter to that of cows (1.267 kg DM kg^?1 4% FCM). However, when DM intake kg^?1 FCM (4%) was compared at constant levels of metabolic body size, fat‐corrected milk, body weight change, dietary energy concentration and green forage percentage in the diet, lactating buffaloes consumed significantly less DM kg^?1 FCM yield as compared with cattle. It was concluded that DM intake was lower in lactating buffaloes. Moreover, lactating buffaloes utilised dietary dry matter, energy and protein for milk production more efficiently than cattle. © 2003 Society of Chemical Industry     

The determination of amino acids composition of the traditional food Potentilla anserina L. root by high‐performance liquid chromatography via fluorescent detection and mass spectrometry

In this study, we developed a method of liquid chromatography with fluoresce detector (LC‐FLD) and on‐line mass spectrometry identification (MSI) using 2‐[2‐(7H‐dibenzo [a, g] carbazol‐7‐yl)‐ethoxy] ethyl chloroformate (DBCEC‐Cl) as pre‐column derivatisation reagent for determination of the amino acids (AAs) in Potentilla anserina L. Separation of the derivatised AA exhibited a good baseline resolution in combination with a gradient elution. All AA derivatives give excellent linear responses with correlation coefficients of >0.9992. The detection limits of each AA were 2.60–24.3 fmol. Eighteen AAs, involving seven essential AAs, were detected in Potentilla anserina L. Quantitative recoveries of the AAs from the Potentilla anserina L. were 84–107%, and the relative standard deviation values were <1.45%. The established method in this study was sensitive and precision enough to separate and quantify AA composition of Potentilla anserina L. Good compositional data were obtained from the analysis of the AAs obtained from Potentilla anserina L. root.     

Characterization of the DGAT1 K232A and variable number of tandem repeat polymorphisms in French dairy cattle

A quantitative trait locus (QTL) underlying different milk production traits has been identified with a high significance threshold value in the genomic region containing the acylCoA:diacylglycerol acyltransferase (DGAT1) gene, in the 3 main French dairy cattle breeds: French Holstein, Normande, and Montbéliarde. Previous studies have confirmed that the K232A polymorphism in DGAT1 is responsible for a major QTL underlying several milk production traits in Holstein dairy cattle and several other bovine breeds. In this study, we estimate the frequency of the 2 alternative alleles, K and A, of the K232A polymorphism in French Holstein, Normande, and Montbéliarde breeds. Although the K allele segregates in French Holstein and Normande breeds with a similar effect on production traits, the existence of additional mutations contributing to the observed QTL effect is strongly suggested in both breeds by the existence of sires heterozygous at the QTL but homozygous at the K232A polymorphism. One allele at a variable number of tandem repeats (VNTR) locus in the 5′ noncoding region of DGAT1 has been recently proposed as a putative causative variant. In our study, this marker was found to present a high mutation rate of 0.8% per gamete and per generation, making the allele diversity observed compatible with that expected under neutrality. Moreover, among the sires homozygous at the K232A polymorphism, no allele at the VNTR can fully explain their QTL status. Finally, no allele at the VNTR was found to be significantly associated with the fat percentage variation in the 3 breeds simultaneously after correction for the effect of the K232A polymorphism. Therefore, our results suggest the existence of at least one other causative polymorphism not yet described. Because the A allele is nearly fixed in the Montbéliarde breed, this breed represents an interesting model to identify and confirm other mutations that have a strong effect on milk production traits.     

Marker discovery and associations with β-carotene content in Indian dairy cattle and buffalo breeds

Vitamin A is essential for human health, but current intake levels in many developing countries such as India are too low due to malnutrition. According to the World Health Organization, an estimated 250 million preschool children are vitamin A deficient globally. This number excludes pregnant women and nursing mothers, who are particularly vulnerable. Efforts to improve access to vitamin A are key because supplementation can reduce mortality rates in young children in developing countries by around 23%. Three key genes, BCMO1, BCO2, and SCARB1, have been shown to be associated with the amount of β-carotene (BC) in milk. Whole-genome sequencing reads from the coordinates of these 3 genes in 202 non-Indian cattle (141 Bos taurus, 61 Bos indicus) and 35 non-Indian buffalo (Bubalus bubalis) animals from several breeds were collected from data repositories. The number of SNP detected in the coding regions of these 3 genes ranged from 16 to 26 in the 3 species, with 5 overlapping SNP between B. taurus and B. indicus. All these SNP together with 2 SNP in the upstream part of the gene but already present in dbSNP (https://www.ncbi.nlm.nih.gov/projects/SNP/) were used to build a custom Sequenom array. Blood for DNA and milk samples for BC were obtained from 2,291 Indian cows of 5 different breeds (Gir, Holstein cross, Jersey Cross, Tharparkar, and Sahiwal) and 2,242 Indian buffaloes (Jafarabadi, Murrah, Pandharpuri, and Surti breeds). The DNA was extracted and genotyped with the Sequenom array. For each individual breed and the combined breeds, SNP with an association that had a P-value <0.3 in the first round of linear analysis were included in a second step of regression analyses to determine allele substitution effects to increase the content of BC in milk. Additionally, an F-test for all SNP within gene was performed with the objective of determining if overall the gene had a significant effect on the content of BC in milk. The analyses were repeated using a Bayesian approach to compare and validate the previous frequentist results. Multiple significant SNP were found using both methodologies with allele substitution effects ranging from 6.21 (3.13) to 9.10 (5.43) µg of BC per 100 mL of milk. Total gene effects exceeded the mean BC value for all breeds with both analysis approaches. The custom panel designed for genes related to BC production demonstrated applicability in genotyping of cattle and buffalo in India and may be used for cattle or buffalo from other developing countries. Moreover, the recommendation of selection for significant specific alleles of some gene markers provides a route to effectively increase the BC content in milk in the Indian cattle and buffalo populations.     

Goat acetyl-coenzyme A carboxylase alpha: molecular characterization, polymorphism, and association with milk traits

Acetyl-coenzyme A carboxylase α (ACACA), the major regulatory enzyme of fatty acid biosynthesis, catalyzes the conversion of acetyl-CoA to malonyl-CoA. We have sequenced 5.5 kb of the goat ACACA cDNA in 18 individuals. The translated cDNA sequence encodes 1,832 AA and shares a high AA identity (99%) with the ovine and bovine ACACA orthologous sequences. One silent single nucleotide polymorphism was identified at exon 45 (C5493T). The genotyping of this polymorphism in 4 goat breeds, Murciano-Granadina, Teramana, Majorera, and Malagueña, showed that it was segregating in all 4 breeds at variable frequencies. In addition, a mixed animal model was used to evaluate the association of C5493T genotypes with milk traits. Results from this study show that the C5493T mutation was suggestively associated with fat yield, lactose content, and somatic cell count.     

Effect of beta-lactoglobulin polymorphism and seasonality on bovine milk composition

The objective was to evaluate the effect of beta-lactoglobulin (beta-lg) polymorphism and seasonality on milk composition (fat, lactose, total solids, milk urea nitrogen, total protein, true protein, casein and somatic cell counts) of Holstein and Girolando cows. Milk and blood samples from 278 Holsteins cows and 156 Girolando cows were taken during two dry seasons and two rainy seasons, for milk composition analysis and to determine beta-lg genotypes, respectively. BB genotype was the most frequent for both breeds, followed by AA genotype for Holstein (BB>AA>AB) and by AB for Girolando cows (BB>AB>AA). No differences were found in milk compositional characteristics among genetic variants of beta-lg (AA, AB and BB) either between Holstein or Girolando cows. No association between milk composition and beta-lg genetic polymorphism was observed. During the dry season, independently of the breed considered, higher contents of lactose, true protein, casein and casein:true protein ratio were found.     

Metabolome profiling of plasma reveals different metabolic responses to acute cold challenge between Inner-Mongolia Sanhe and Holstein cattle

Low-temperature conditions influence cattle productivity and survivability. Understanding the metabolic regulations of specific cattle breeds and identifying potential biomarkers related to cold challenges are important for cattle management and optimization of genetic improvement programs. In this study, 28 Inner-Mongolia Sanhe and 22 Holstein heifers were exposed to ?25°C for 1 h to evaluate the differences in metabolic mechanisms of thermoregulation. In response to this acute cold challenge, altered rectal temperature was only observed in Holstein cattle. Further metabolome analyses showed a greater baseline of glycolytic activity and mobilization of AA in Sanhe cattle during normal conditions. Both breeds responded to the acute cold challenge by altering their metabolism of volatile fatty acids and AA for gluconeogenesis, which resulted in increased glucose levels. Furthermore, Sanhe cattle mobilized the citric acid cycle activity, and creatine and creatine phosphate metabolism to supply energy, whereas Holstein cattle used greater AA metabolism for this purpose. Altogether, we found that propionate and methanol are potential biomarkers of acute cold challenge response in cattle. Our findings provide novel insights into the biological mechanisms of acute cold response and climatic resilience, and will be used as the basis when developing breeding tools for genetically selecting for improved cold adaptation in cattle.     

Immunoglobulin E epitope mapping by microarray immunoassay reveals differences in immune response to genetic variants of caseins from different ruminant species

The allergenicity of the caseins (CN), one of the major allergens in cow milk, is well characterized and their immunoglobulin E (IgE)-binding epitopes have been identified. However, investigations about the allergenic potential of the genetic variants occurring in the caseins are lacking. Therefore, this study determined the influence of the genetic polymorphism on IgE binding to epitopes of bovine casein variants. Furthermore, differences in IgE binding between epitopes of goats and water buffaloes were analyzed. A set of 187 peptides, covering the previously identified sequential IgE-binding epitopes of α_S1-, α_S2-, β-, and κ-CN variants from cows and the corresponding homologous peptides of water buffaloes and goats, were synthesized and tested by means of peptide microarray for IgE binding, using sera from 16 cow milk-sensitized individuals. Seven of the 16 sera samples showed positive signals on microarrays and were included in this study. In 5 α_S1-CN variants (A, B, C, E, and I), the AA substitution or deletion affected the immunoreactivity of epitopes AA 4 to 23, AA 17 to 36, AA 83 to 102, AA 173 to 192, and AA 175 to 194, as well as of the variant-specific peptides AA 184 to 196, AA 187 to 199, AA 174 to 193, and AA 179 to 198, which were found to resist gastrointestinal digestion. Variation in IgE binding was further detected for peptides AA 103 to 123 and AA 108 to 129 of 3 β-CN variants (A¹, A², and B). The majority of sera showed IgE binding to α_S1-CN peptides of cows and the homologous counterpart of goats and water buffaloes. However, α_S1- and β-CN epitopes from goats and water buffaloes had lower immunoreactivity than those of cows, but, in some cases, higher or exclusive IgE binding was observed. The results of this study indicate that genetic variants of the caseins differ in their allergenicity. This might be useful in the search for a suitable protein source for cow milk-allergic patients. In addition, milk from water buffaloes and goats harbor an allergenic potential due to cross-reactivity of IgE antibodies with cow milk caseins and are, therefore, not an acceptable alternative in the nutrition of cow milk-allergic patients.