فصلنامه پژوهشهای تولیدات دامی
پیاپی 44 (تابستان 1403)

To date, one of the main areas of research is the creation of breeds with inherent resistance to digestive nematodes in sheep. In this context, resistance to digestive nematodes has exhibited significant phenotypic differences within and between sheep breeds, suggesting a genetic basis for these differences. According to the list of effective candidate genes and the identified gene network for parasite resistance in sheep, there are genes involved in the immune system, such as the interferon-γ gene (IFN-γ). Cytokinin is involved in biochemical immune system signaling pathways, parasite resistance, and immunological responses. In addition, certain mutations in this gene impair the ability of specialized cells of the immune system to fight off parasite invasion. One of the most popular approaches to generating gene expression data for genome function studies is DNA microarray technology, which allows the simultaneous expression of thousands of genes. Proteomics and genomics are two application areas of microarray technology. This research aims to identify and categorize some of the genes involved in the relative genetic resistance of sheep to nematode infection using microarray data.

In this context, the GEO-Bank, part of the NCBI, was searched for access to open-access databases. Downloaded microarray data corresponded to infection with parasitic nematodes with the best replication (e.g., data in two resistant and sensitive groups), and the set of genes with differential expression was identified using R -based appropriate software packages (Biobase, GEOquery, limma, affy, Genfilter, Pheatmap, Plyr, Reshape2, and Ggplot2). Raw data were measured on a logarithmic scale, and the P-value fit statistics were used for expression comparisons between gene groups.

The main analysis was performed after precorrection and processing of the raw data because of the high intragroup variance of the data, as evidenced by the observed quality control results of the raw data and the quality control-related results of the integrated data. After pre-processing of the raw data, correlation analysis revealed a strong relationship between genes in the pre-Inf group (Pre-Inf) and the infected group (Inf) compared to the control group. Three heatmap reference charts, a PCA chart, and a volcano plot were used to verify data quality, and by verifying these charts, samples of unfavorable quality were removed from the next stages of analysis. After a bioinformatics analysis, the results showed significantly increased expression patterns (NACA, RPL4, NAGS, CTCF, GBP1, BHLHE, YTHDF3, PDHA1, and MXI1) and diminished expression patterns of PDHA1 and MXI1 genes. According to the results of this study, these genes play a role in the cellular metabolism process, molecular function, the formation of genetic connections, and cell life. Therefore, they were significant (p-value < 0.05) according to the magnitude of the change. The N-acryl glutamate synthetase (NAGS) gene is the cofactor of the first enzyme involved in the urea cycle in mammals. The functional role of this gene has been identified in many neurological diseases. The CTCF gene has a positive effect on the cells of the immune system and plays a special role in defending against viruses and pathogens that invade the host body. The guanylate binding protein 1 (GBP1) gene plays a role in the body's defense against many infectious pathogens. This gene causes oxidative reactions and autophagy of the host immune system as a barrier to invading pathogens. The methyl adenosine RNA binding 3 gene is more effective in antiviral immunity and is closely linked to the GBP1 gene, which plays a role in the body's resistance to many infectious agents. This gene causes oxidative responses and autophagy of the host immune system against invading pathogens. The PDHA1 pyruvate dehydrogenase gene is involved in immune system metabolism. This gene plays a role in allosteric factor-induced regulation, repair of covalent bonds, and relatively rapid changes in the level of expressed proteins, either through altered gene expression or proteolytic degradation. The MXI1 gene helps control the entry of invading pathogens into the host's body and promotes recovery and healing from infections. The results can be explained by several reasons, including the use of different sample breeds and laboratory techniques, the level of parasite contamination, the age of the test material, the variety of tested nematodes, and inconsistent techniques and tools of the current study with those of previous studies. Other reasons include the climatic differences between the test region and its physical location as well as the use of different RNA and microarray methods. However, the results of the study may be helpful under the related circumstances.

The results of microarrays and differential expression patterns can be of great help to molecularly modify livestock to resist internal parasites. Using more advanced tools, such as next-gene sequencing, will provide more accurate and relevant information.

Using genetic enhancement techniques has led to the development of poultry capable of fulfilling significant portions of human dietary requirements through meat and egg production. Maximizing genetic potential necessitates the provision of conducive environmental conditions. Heat stress poses a substantial challenge within the poultry sector, detrimentally impacting poultry production and growth, there by diminishing breeders' economic returns. Adaptation to heat stress is contingent upon a multitude of factors. The capacity to establish homeostasis through alterations in gene expression serves to mitigate extracellular damage and restore physiological equilibrium upon reversion to favorable temperature ranges. Domestic birds thrive within a temperature spectrum of 18-24 °C, with any deviation beyond this range precipitating elevated mortality rates. Elevated mortality is attributed to heat-induced suppression of the immune system and compromised resilience. Moreover, heat stress amplifies the secretion of the corticotropin-releasing hormone, subsequently elevating blood corticosterone levels, thereby instigating molecular alterations under stress conditions. Hence, a comprehensive examination of this challenge from diverse perspectives holds significant promise. Consequently, the present study endeavors to delineate protein complexes and pivotal signaling pathways regulated by distinct genes expressed in response to heat stress within the hypothalamus of broiler chickens.

The systemic response of chickens' hypothalamus transcripts to heat stress was comprehensively examined in this investigation. Transcript expression profiles from eight chicken hypothalamus tissues, including four subjected to thermal stress and four under non-stress conditions, were retrieved from the GEO database (accession number GSE37400). The raw microarray data underwent rigorous quality control measures and normalization using GEO2R software to ensure reliability and consistency. Genes with discernible expression changes were identified based on a stringent criterion of P-value > 0.05 and (LogFC < 0.5-0.5). Subsequently, a gene expression regulatory network was constructed to elucidate the interplay among genes implicated in broiler heat stress, employing Cytoscape software in conjunction with the STRING database. The network visualization facilitated the identification of protein clusters, representing areas of high density, which were further analyzed using the CytoCluster plugin renowned for its efficacy in detecting such densely interconnected regions. Following the delineation and characterization of genes within these clusters, the DAVID database was utilized to elucidate the relevant biological pathways associated with the identified genes. This comprehensive approach enabled a detailed exploration of the molecular mechanisms underlying the hypothalamic response to heat stress in chickens.

The analysis of microarray data unveiled a comprehensive portrait of the molecular response to heat stress in chickens. Among the 43,607 initially examined probes, 593 differentially expressed genes were identified after rigorous curation involving the elimination of duplicate genes and application of a significance threshold. A network encompassing 149 genes with a minimum interaction score of 0.4 was constructed utilizing the STRING software, shedding light on the intricate interplay among these genes. Subsequent investigations honed in on genes exerting significant regulatory effects in response to heat stress, elucidating their pivotal roles in orchestrating physiological processes. Functional assessment of the networked genes revealed their involvement in diverse biological activities, including the positive regulation of T cell-mediated immunity, WNT and MAPK signaling pathways, as well as phagocytosis and natural killer (NK) cell activity. Particularly noteworthy was the observation of suppressed NK cell activity in broilers under heat stress, likely influenced by heightened corticosterone levels and subsequent inhibition of thyroid hormone secretion, thereby impacting temperature regulation and metabolism. Moreover, the elevation in blood glucose levels observed in stressed chicks may contribute to their adaptive survival response. Importantly, heat stress was observed to disrupt cellular homeostasis and molecular pathways, manifesting in alterations in biological processes such as the cell cycle. This disruption in the cell cycle may signify a novel mechanism of stress resistance in vertebrates, further underscoring the complexity of the physiological response to thermal stress in chickens.

The analysis of gene functionality within the network underscores the intricate nature of the innate immune response and cellular demise elicited by heat stress. Notably, certain genes identified in this analysis participate in positively regulating T cell-mediated immunity, vascular endothelial growth factor (VEGF), WNT signaling, and mitogen-activated protein kinase (MAPK) pathways. Introducing a biomarker panel comprising genes, such as MAPK14, NT3, SOX2, YAP1, FGF2, LRRK1, LEF1, and TLE1, holds promise for advancing our understanding of the underlying molecular mechanisms governing stress responses in poultry. Furthermore, this approach may facilitate the discovery of novel genes whose modulation could mitigate the effects of thermal stress.

Rapid growth of broilers leads to insufficient cardiac output, cardiac hypertrophy, and finally sudden death. When livestock are exposed to hot environments, they often try to dissipate heat by increasing blood flow to the skin. This physiological response is a way for animals to adapt to long-term thermal conditions. However, this thermoregulatory mechanism has its limitations. While different animals have varying degrees of tolerance to heat stress, the cardiovascular system can only compensate within a certain range before it starts to impact food production. Domestic poultry, such as chickens, lack sweat glands, which is a key mechanism for body cooling in many other animals. This deficiency forces the cardiovascular system to work harder, sending more blood to the skin to help regulate body temperature. Broilers, in particular, have a narrow thermoneutral zone (the range of temperatures where an animal can maintain its body temperature without expending extra energy) and a high metabolic rate, which places an even greater burden on their cardiac function. Therefore, it is an appropriate model for studying cardiac disease. Heat stress is among the stressors affecting cardiac performance. In accordance with the cardiac system susceptibility of poultry to heat stress, especially broilers because of their fast growth rate, to enhance the ability of modern broiler chickens to withstand heat stress through selective breeding, we need to identify the crucial genes and biological processes that lead to unbalanced heart development and the high incidence of heat-induced heart problems. Thus, the aim was to consider the gene expression of cardiac contractility proteins under heat stress conditions during the growing period of broilers.

Ross 708 broilers were subjected to heat stress (HS) of 35–37 °C for 8 hours daily for 21 days post-hatch. Initially, male broilers had unrestricted access to feed and water in spacious colony houses maintained at 33 °C, with the temperature gradually decreasing by 3 °C per week until it reached 24 °C by the day 21 post-hatch. At the conclusion of the growth period, the broilers were euthanized, and their left ventricles were isolated for mRNA extraction. RNA-seq data were obtained from NCBI’s with the accession number SRP082125. All the expressions of deferentially expressed genes (DEGs) were determined and analyzed by DAVID online bioinformatics tools. Gene ontology qualification, including biological processes (BP), cellular component (CC), and molecular role (MF), were achieved from DAVID. Heat stress induction was from days 21-42 for 8 hours every day for 21 days until to end of the growing period.

Gene expression changes were related to 35 genes of the muscular cardiocyte of the left ventricle. From seven genes related to the protein of troponin, TNN and TNNI1 genes, declined significantly, with an increase in the TNNT3 gene expression (P< 0.05). Of five genes related to the tropomyosin protein, the TPM1 gene showed a significant increase in expression (P < 0.05). Of the two genes related to the ryanodine receptor, RYR3 had significant gene upregulation (P < 0.05). From the ten genes related to DHPR, only CACNA2D2 had significant downregulation (P < 0.05). Among four calmodulin-related genes, CAMKK1 and CAMK1D showed significant downregulation, and CAMSAP2 revealed upregulation (P< 0.05). Significant reductions occurred in the expression of five associated genes of heavy-chain myosin (P< 0.05).

Heat stress reduced gene expression of ryanodine, DHPR, and troponin, those related to calcium release into the cell cytosol. Furthermore, it prevents cardiac hypertrophy by decreasing the expression of genes related to the heavy chain of myosin. Thus, the vulnerability of modern broilers to heart problems, when exposed to high temperatures, may be linked to their heart's reduced capacity, which is likely due to their relatively smaller heart size. According to genetic analysis, the smaller heart size in broilers under heat stress than those in a comfortable temperature environment is likely caused by slower cell growth and division. This research identifies specific genes and biological pathways that could be targeted through breeding to develop broilers that are more resistant to heat stress and have healthier hearts. This study has identified specific genes and biological processes linked to the reduction in heart size observed in broiler chickens subjected to heat stress. These insights provide new opportunities for developing targeted breeding strategies to address this issue. The findings indicate that selective breeding, by focusing on the identified genes and pathways, may enable the development of broiler chickens that are more resistant to heat stress and have healthier hearts. By concentrating on these genetic factors, breeders can work toward creating broiler populations that are more resilient to high temperatures and less susceptible to heart problems.

Angel wing is a developmental wing deformity that can influence breeding and reproduction in the commercial duck industry. Therefore, genetic diversity studies and detection of the genomic region related to angel wings in duck populations are essential. In this regard, powerful tools, such as next-generation sequencing technology, have made it possible to decode genome information in this species. The genome-wide association study (GWAS) has been a powerful tool in detecting loci associated with complex traits and diseases; however, it also has some limitations. Complex traits are controlled by many genes, and hence, significant SNPs in general represent only a small fraction of genetic variation. Moreover, studies often report only the most significant SNPs and their neighboring genes, hence some smaller genetic variants and disease risks are unlikely to be detected. Alternatively, pathway-based analysis has been proposed as a complementary approach to investigate complex traits from a genetic and biological perspective. In contrast to a GWAS, pathway-based analysis considers factors that contribute simultaneously to the complex trait and looks beyond the most significant SNPs and genes. To complement GWAS studies, it is becoming common to use gene-set enrichment and pathway analyses. Such an approach helps alleviate problems related to GWAS (e.g., GWAS ignores the fact that genes work together in networks in various biological pathways), and to deepen the understanding of the biological pathways affecting quantitative traits. Integration of F, GWAS, and pathways analyses might address some aforementioned issues and has been already used in human studies, whereas its potential application in livestock breeding and genetics remains still unexplored. In addition, studies are available that performed GWAS or GWAS plus pathway analysis.

A total of 63 adult purebred Pekin ducks from the same population were selected for this study, of which 33 were ducks that could be identified as having angel wings (case) and 30 were ducks with normal wings (control). Genomic DNA was extracted from blood samples by DNA extraction using a kit (QIAampR DNA Blood Mini Kit; QIAGEN), following the manufacturer's protocol. Whole-genome re-sequencing data were generated on the Illumina Hiseq 4000 platform with 150 bp paired-end reads. Single-nucleotide polymorphism (SNP) calling was performed using the GATK (v4.1), and all parameters were kept at default settings, except for stand_callconf 30. VCFTOOLS (v0.1.16) and plink (v 1.90) were used for the quality control of the data. The 14 064 984 SNPs passed quality control that excluded SNPs using the following criteria: --min- alleles 2, --max- alleles 2, --minDP 3 –minQ 30 with VCFtools, minor allele frequency >0.01, and SNP call rate ≥ 0.95 with plink. An independent SNP set was used via the plink command --indep- pairwise 50 5 0.2 for principal component analysis. After quality control, 686 449 SNPs were used for the GWAS. The gene set analysis consists basically of three different steps: (i) the assignment of SNPs to genes, (ii) the assignment of genes to functional categories, and (iii) an association analysis between each functional category and the phenotype of interest. 1. The SNPs were assigned to bovine genes based on the CAU_duck1.0 duck genome sequence assembly using the Bioconductor R package biomaRt2. A given SNP was assigned to a particular gene if it was located within the gene or at most 15 kb either upstream or downstream of the gene. An arbitrary threshold of P-value ≤ 0.005 was used to define significant SNPs (based on the results of the GWAS); in this context, significant genes were defined as those genes that contained at least one significant SNP. 2. The databases Gene Ontology (GO) and Medical Subject Headings (MeSH) were used to define functional categories of genes. The idea is that genes assigned to the same functional category can be considered the members of a group of genes that share some particular properties, typically their involvement in the same biological or molecular process. 3. The significant association of a given term with angle wing was analyzed using Fisher’s exact test. Finally, a gene enrichment analysis was performed with the GOstats Bioconductor from R software for the assignment of the genes to functional categories.

The random effect was estimated from the groups clustered based on the kinship among all accessions, and the first two PCs, including PC1 with normal wings and PC2 with angel wings derived from whole-genome SNPs, were used as fixed effects in the mixed model to correct for stratification. In this research, SNP markers were identified on chromosomes 1, 2, 3, 6, 8, 11, 18, 20, 27, and 31. Different sets of candidate genes related to the angle wing trait, namely ATP11A, UBE2E2, ITPR2, GUCA1C, ATP2C1, PLCG1, and BMPR1A, were also identified in ducks. Some of the found genes are consistent with some of the previous studies related to wing traits. According to pathway analysis, 21 pathways from GO and biological pathways were associated with the angle wing trait. Some of the detected genes are consistent with some previous studies and are involved in biological pathways related to skeletal muscle growth and development, calcium ion response, bone growth and development, bone minerals, and calcium signaling pathway activation.

The results of our research can be used to understand the genetic mechanism controlling the angle wing trait. This study supports previous results from the GWAS of reproductive traits, revealing additional regions. Using these findings could potentially be useful for genetic selection in ducks.

Artificial insemination is one of the important management tools to control reproductive activities and increase the productivity of the livestock industry. This technique can play an important role in genetic improvement even in small herds. Sperm preparation is of particular importance for reproductive techniques, such as artificial insemination, but many harmful factors, such as temperature shock and oxidative stress, reduce sperm quality and directly affect sperm fertility during sperm preparation and storage. Therefore, the addition of protective substances and antioxidants in the diluent is considered essential in maintaining sperm quality during the cooling and freezing stages. Curcumin is a yellow substance obtained from the root of turmeric, and its biochemical structure contains a phenolic ring and a beta-di-ketone part in its structure. It can neutralize free radicals, but it has low solubility in hydrophilic environments. To solve this problem, the nanoliposome method has been proposed. Using this method can solve the problem of curcumin's low solubility in aqueous solutions and, additionally, curcumin nanoparticles can be guided to exert their effects in a completely targeted manner inside the target cell. On the other hand, sucrose protects sperm from cold shock by first rinsing sperm cells and maintaining the osmotic pressure of the diluting medium. This study aimed to investigate the effects of separate and mixed use of different levels of curcumin nanoparticles and sucrose sugar in ram epididymal sperm diluent during storage at 5 ˚C at different cooling times.

The present study was conducted in the Animal Physiology Laboratory of the Department of Animal and Poultry Sciences, Aburihan Agricultural Campus, the University of Tehran. The testicular tissues were prepared from a slaughterhouse on the day of the experiment and transported to the laboratory with special flux in less than an hour. Sperm samples were collected from the epididymal tail of the ram testicular tissue in the laboratory. Sperm was sampled after initial evaluation, with progressive motility of more than 75% and abnormality of less than 10%, and were added to the diluent at 37 °C. In this study, the diluent consisted of Tris (27.1 g/L), citric acid (14 g/L), fructose (10 g/L), egg yolk (20%), and glycerol (7%), with a pH of around 6.2-7.8. The osmolarity was set at about 310-320 milliosmoles. Sucrose sugar was obtained from the Sigma Aldridge Company, and curcumin nanoparticles were prepared from pure curcumin powder (Sigma Aldridge) by the nanoliposome method and using rotary devices, a homogenizer, and a prop sonicator. A DLS device was used to examine the particle size of curcumin nanoparticles (less than 80 nanograms). The experimental groups included 25 and 50 μM of nanoparticles of curcumin (NC), sucrose (S) at 100 and 150 mM separately, combined levels (25NC100S, 25NC150S, 50NC100S, and 50NC150S), and a group without these additives (control), which were added to the diluent at 37 °C. The diluents containing different levels of the treatments were refrigerated in 15 ml falcones and inside a water container at the same temperature, which gradually reached a temperature of 5 ˚C in about 2 hours. After stabilization at this temperature, total motility parameters, progressive motility using the KASA system, the viability with the eosin-nigrosin method (16.7 g of eosin, 100 g of nigrosin, and 29 g of citrate buffer in 1 L of double-distilled water), membrane integrity by the HOST solution method (9 g of fructose and 4.9 g of sodium citrate in 1 L of double-distilled water with an osmolarity of 100 milliosmoles), and the percentage of sperm abnormalities using Hancock's solution (5 62/L of 37% formalin, 150 ml of saline solution, 150 ml of a buffer solution, and 500 ml of double distilled water) were evaluated at 6, 12, 24, and 48 hours.The experimental results were analyzed in a completely random design by SAS software (9.2) and the GLM procedure at a significance level of P < 0.05. The means of the treatments were compared with Tukey's test.

The use of 25NC100S in the diluent improved the parameters of total motility, progressive motility, viability, and integrity of ram epididymal sperm membrane in all evaluated times compared to the other groups, especially the control group (p < 0.05). No significant differences were observed between the treatments in the percentage of sperm abnormalities at 6 and 12 hours (p > 0.05). At 24 and 48 hours, however, the 25NC100S concentration significantly reduced the percentages of sperm abnormalities compared to the other concentrations (p < 0.05). Compared to the control group, adding different levels of curcumin nanoparticles and sucrose sugar separately significantly improved the evaluated parameters, but the best performance was observed in the combined treatment, indicating the synergistic role of these two compounds.

The results of the present study demonstrate that using a mixture of curcumin nanoparticles with sucrose in the diluent can protect sperm against oxidative damage and cold shock. The antioxidant effects of curcumin nanoparticles and the protective effects of sucrose sugar can result in positive effects on the quality of ram epididymal sperm during cooling. Therefore, it is recommended to use the 25NC100S level in ram sperm diluents.

One of the goals of new programs for silkworm breeding is to investigate the ability to withstand environmental fluctuations. Silkworms are cold-blooded organisms, meaning that their body temperature and metabolic processes are affected by the temperature of the environment. Silkworms are sensitive to temperature changes, and exposure to cold temperatures can affect their growth, development, and overall health. So far, no special attention has been paid to improving compatibility traits in Iranian silkworms. The requirement of this goal is to improve the traits related to resistance to stresses and adaptation to new conditions. Achieving this regard requires expanding the country's silkworm gene bank, especially in the field of commercial lines. New genetic resources should also be made available while identifying the genetic capabilities of the existing lines. In the meantime, producing eggs that are resistant to cold stress is one of the proposed solutions to deal with this stress. For this purpose, it is necessary to determine the level of resistance or susceptibility to cold stress in commercial lines used in the production of commercial eggs, and the desired lines should be rated from this aspect. With this rating, recommendations can be made to create crosses between the mentioned lines aiming at producing commercial eggs resistant or sensitive to cold stress and distributing each batch of these eggs in different geographical areas with different climates. In the present study, the performance of commercial lines of Iranian silkworms including 31, 32, 103, 104, 151, 153, and 154, under cold stress was investigated in control and challenge treatments.

After the steps of preparation for breeding, silkworm eggs were kept in a hatching room under standard temperature and humidity conditions for 12 days. The standard conditions for breeding were temperature (25 ± 2 °C), relative humidity (75 ± 5%), and photoperiod (16 hours light/ 8 hours dark). Mulberry leaves of modified varieties were used to feed the larvae. To induce cold stress, 300 larvae on the third day of the fifth instar (in the form of three replicates of 100 larvae) were incubated at 0 °C for 12 hours and then returned to the standard rearing conditions at 25 °C. The production traits the characteristics of cocoon weight, cocoon shell weight, cocoon shell percentage, number of cocoons per liter, cocoon weight per liter, the average weight of a cocoon, the average weight of the best cocoon, the average weight of a middle cocoon, the  average weight of a weak cocoon, and the average weight of a double cocoon, as well as some traits related to longevity, including the percentage of live larvae, the percentage of dead larvae, the percentage of pupal losses, and the percentage of produced butterflies, were investigated in each cold stress and control treatments. For statistical analysis, the generalized linear model (GLM) procedure was used in SAS software, and means were compared with Tukey's statistical test at a significant level of 0.05.

In general, the highest and lowest values of traits were not the same among the genotypes and under applied stress conditions, but some genotypes showed higher performance for a larger number of traits. Among the studied traits, those related to cocoon weight, cocoon shell weight, and cocoon shell percentage are the most important traits for breeding purposes, which have high economic values and are used to improve cocoon performance. Meanwhile, the live and dead larvae percentage traits, along with death pupae percentage and productive moth percentage, are important indicators related to viability, which showed significant differences between the studied lines. The results of mean comparisons showed that the larvae of the control group generally had better performance in all the examined traits than the larvae under cold stress, with a significant difference (P < 0.001). However, two lines 153 and 154 from the cold stress group had higher performance in many production and longevity traits than the other lines under cold stress and even some lines of the control group.

The temperature of the breeding environment is one of the factors that, in the case of fluctuation, leads to the deviation of breeding conditions from the optimal state and damage. In general, cold stress and low temperatures have different effects on insects, considering the intensity and duration of the insect's exposure. In addition, the stage of life, the evolution degree of adaptation mechanisms, and adaptation to the environment greatly affect the insect's response to cold stress. The results of the present research show that the performance of commercial lines of Iranian silkworms is affected under cold stress conditions, and lines 153 and 154 have higher resistance than the other lines. Hence, they can be prioritized in the production of commercial hybrids in terms of resistance to cold stress and production and survival records. Further studies are suggested to measure the molecular and biochemical characteristics of some proteins related to temperature stress in different treatments.

So far, changes in the vaginal microbial population during the estrous cycle have been evaluated in several livestock species such as sheep. However, changes in the rumen microbial population have not been investigated during the estrous period in sheep. In a study, the results showed that the differences in the rumen microbial population in two reproductive seasons were due to the differences caused by the reaction to day length, and nutrition did not have much effect on it. It is possible that during the reproductive and non-reproductive seasons, the animal has a different microbial population in the digestive tract. The available evidence from clinical research in animals shows that microbes in the digestive system can alter brain activity and behavior through hormonal and neural pathways. Therefore, this study mainly aims to investigate changes in the rumen microbial population during the reproductive season in the estrous period in ewes with estrus and those without estrus for any reason or are so-called anestrus. The changes in the rumen microbial population during the breeding season in the estrous period were measured for the animals showing the occurrence of estrus and those not showing this trait or were anestrous.

This research was done at the same time as the non-breeding season (late spring-beginning of July). The ewes were of Grey Shirazi breed (2-3 years old). To investigate changes in the rumen fluid microbial population in the estrus cycle in estrus and non-estrus animals, two groups of 10 animals were formed from estrus and anestrus animals with the use of teaser rams, and rumen fluid samples were collected for microbial culture and other investigations in both groups once per four days (5 times) during 17 days (like estrus cycle days). The colonies were cultured and separated using general culture media with different growth factors for anaerobic bacteria colonies, such as plate count agar, MRS agar (de Man-Rogosa - Sharpe), and starch agar. The colonies were subtracted by subtractive cultures or different diagnostic tests to identify colonies. Data were analyzed using SAS software version 1/9. Means were compared with the least square procedure and adjusted for Tukey's test.

The population difference between the two groups at 5 different times showed that the population of lactic acid bacteria (LAB) on the day of estrus showed a higher value and a significant difference with other times than the anestrus group. Lipolytic bacteria were higher on the day of estrus than on all days in both groups, with a significant difference. The comparison of different groups showed that the estrous group from times 1 to 5 in the breeding season had the highest number compared to the other groups at different times of the study (p < 0.05). Anestrous groups also had the lowest number at all times (p < 0.05). The comparison of the two estrus and anestrus groups showed that heat and ninth days had higher values than the other times. In comparing the average number of lipolytic colonies in the estrus group, the highest and the lowest numbers belonged to the 1st and the 4th times, respectively, which were significantly different from the other times (p < 0.05). The population of proteolytic bacteria had the highest number of bacteria at time 1 and showed the next values at times 2, 5, 3, and 4, respectively, compared to the estrus group at time 5 (p < 0.05). Time 5 was not significantly different from times 2 and 3 (p < 0.05). In this season, the estrus group was the highest and the lowest at times 1 and 4, respectively (p < 0.05). Times 3-5 were no different significantly (p < 0.05). In the anestrus group, times 3 and 1 had the highest number vs. the lowest numbers in times 2 and 5  (p < 0.05). Based on the comparison of the average population resulting from the counting of total bacterial colonies in the examination of different groups, the estrous group had the highest number of the average microbial population in all 5 times (p < 0.05). The lowest number belonged to two anestrus groups (p < 0.05). The population difference between the two groups at 5 different times showed that the population of LAB had a higher value only on the day of estrus, with a significant difference compared to the anestrous group. The population of lipolytic bacteria in two groups of estrus and anestrus in the 5 studied times showed that the estrus group had a higher and significant value than the anestrus group in the 4th time, i.e. the day of estrus and days 5, 6, and 17. The results showed that the total bacterial population was higher in the estrus group at all times, with a significant difference. Furthermore, anaerobic bacteria increased significantly in certain days of estrus (days 1-9) compared to the anestrous group. The population difference between the two groups at 5 different times indicated that the population of LAB had a higher value only on the day of estrus, with a significant difference compared to the anestrous group. The population of lipolytic bacteria in the two estrus and anestrus groups in the 5 studied times revealed that the estrus group had a higher and significant value than the anestrus group in 4 times, i.e. days of estrus, 5, 6, and 17. The total bacterial population in the estrous group was higher and significant at all times.

In general, the colony population of amylolytic and lipolytic bacteria and the total colonies of anaerobic bacteria are significantly different on days of the estrus cycle, which may affect reproduction. Accordingly, it is better to provide more understandable results of rumen microbial effects on estrus in ewes by increasing analyses such as hormonal changes and molecular investigations, along with microbial culture in estrus crops.

The main challenge of animal production in Iran is the lack of fodder, and corn is one of the important agricultural crops to provide fodder. The level of cultivation and performance of corn have increased significantly in recent decades, and it is predicted that the demand for corn may double the current demand by 2050. Many factors are involved in the increase or decrease of corn yield, but choosing a superior, compatible, and high-yielding hybrid in each region is one of the main factors in increasing the production and yield of the corn crop. One of the limiting factors is the lack of water resources and the occurrence of droughts in Iran in the last few years. In this regard, it is necessary to better understand the morphological characteristics of the plant, the performance of different hybrids, and their nutritional value in different stages of growth and different cultivars in livestock feeding. Therefore, this experiment was conducted to investigate the effects of the growth stages of some fodder corn cultivars on chemical compounds, parameters of gas production, and ruminal degradability in four hybrid varieties, namely corn single-cross Simon, single-cross Valbum, single-cross corn BC 678, and single-cross N.S 770 maize variety.

Four varieties of fodder corn were planted in a uniform field with similar crop management in the farm of Behdis Protein Nasr Company, Behshahr, Mazandaran province, Iran, with an area of about one hectare for each variety. After the operation, they were planted in two stages (August 2 and 24 2022). Harvesting and laboratory operations were carried out in the Animal Nutrition Laboratory of the Animal Science Department, Sari University of Agricultural Sciences and Natural Resources (SANRU). In this research, four fodder corn cultivars were used in a completely randomized design in four treatments with four replications and two harvesting stages. Chemical compounds, including dry and organic matter, crude ash, NDF and ADF, crude protein, crude fat, and non-fibrous carbohydrates, were determined in the cultivars. The potential of gas production, digestibility, metabolizable energy, and volatile fatty acids were assessed in vitro. In addition, rumen degradability and effective parameters of dry matter, crude protein, and neutral detergent fiber in rumen incubation at 0, 2, 4, 6, 12, 24, 36, 48, 72, and 96 hours were determined in situ using three rumen fistulated Zel sheep.

Chemical compounds were affected by cultivars and harvest time. In two stages of harvest, the crude protein content and percentage of dry matter were significantly higher in singlecross NS770 cultivar, and fat, insoluble fiber in acid detergent, and insoluble fiber in neutral detergent rose in the singlecross BC678 cultivar. The effect of corn varieties on the parameters of gas production at harvest times was different. In the first and second harvests, the rate of gas production and gas production in 96 hours, digestibility of organic matter, metabolizable energy, and the concentration of short-chain fatty acids (SCFAs) were higher in the singlecross NS770 variety. There was a significant difference in the gas production capacity of Single-Cross-Simon and Single-Cross NS770 cultivars in the first and second stages of harvesting. The gas production rate constant in the second stage of harvest was different between the treatments, with a higher rate in single-cross BC678 and single-cross NS770 than in single-cross-Simon and single-cross-Valbum varieties. The percentage of organic matter digestibility was different between the treatments in the first and second harvests. The organic matter digestibility was higher in Single-Cross-Simon and Single-Cross NS770 in the first harvest, and Single-Cross-Simon contained the highest amount of digestibility in the second harvest. The concentration of SCFAs was also significantly different between the treatments in the first and second harvests, with the highest concentration in Single-Cross-Simon in the first and second harvests. The degradability of dry matter, crude protein, insoluble fiber in neutral detergent, fast degradable, slow degradable, potentially degradable, and effective degradability fractions were higher in the singlecross NS770 silage at 2, 5, and 8% passage rates. The results of the gas test and determination of degradability showed that the Single Cross NS770 variety had gas production capacity, a constant rate of gas production, digestibility of organic matter, the parameters of degradability (dry matter, crude protein, and insoluble fibers in neutral detergent), and higher effective degradability in the first harvest.

The test results showed that chemical compounds were affected by cultivars and harvest time. In two stages of harvest, the crude protein content and percentage of dry matter were significantly higher in the single-cross NS770 cultivar, and fat, insoluble fiber in acid detergent, and insoluble fiber in neutral detergent rose in the single-cross BC678 cultivar. The results of the gas test and determination of degradability showed that the Single Cross NS770 variety had gas production capacity, a constant rate of gas production, digestibility of organic matter, a, b, and a + b, Kd as parameters of degradability of dry matter, crude protein, insoluble fibers in neutral detergent, and higher effective degradability in the first harvest.

Zinc (Zn) is one of the essential minerals for the health and productivity of growing calves because this mineral is important for metabolism, growth, immune and defense systems, and antioxidant status. This element is also known as an anti-inflammatory and anti-diarrheal agent. In addition, Zn in enzymes acts as a structural component away from the active site, a proton donor in the active site, and an atomic bridge between the substrate and the enzyme. Zn has an essential role in the regulation of many metabolic processes, and its deficiency results in low appetite and, consequently, decreased feed intake. Therefore, it is of strong interest for producers, feed manufacturers, veterinarians, and scientists. The National Research Council (2021) recommends 70 mg/kg of Zn for calves at 30 days of age while the amount of this element is commonly low in the soil of many regions of Iran. Therefore, plants that grow in these soils have a low level of Zn concentration, and when they are consumed as animal feed, can cause a wide range of complications due to Zn deficiency, among which growth abnormality is one of the most obvious signs. Zn supplementation may improve the health and performance of suckling calves. However, the use of high Zn concentrations in the diet may affect the digestion, absorption, and use of other nutrients in the diet and potentially lead to environmental pollution due to the excess excretion of Zn in feces. Thus, the use of Zn sources above bioavailability has a special place. Recently, organic and hydroxy forms of minerals in animal feed supplements have attracted substantial interest from feed manufacturers and animal producers because they have higher Zn bioavailability than inorganic salts. This study aimed to evaluate the effect of Zn sulfate, organic Zn, and Zn hydroxychloride on performance, growth, and blood parameters in Holstein suckling calves.

This study was conducted on 40 Holstein suckling calves from 7 to 77 days of age in a completely randomized design. Each treatment had the same number of male and female calves. Treatments were 1) a basal diet without Zn supplement (containing 53.29 mg/kg of Zn DM), 2) a basal diet + 20 mg/kg of Zn DM as organic Zn, 3) a basal diet + 20 mg/kg of Zn DM as Zn hydroxychloride, and 4) a basal diet + 20 mg/kg of Zn DM as Zn sulfate. Calves were kept in individual stalls and had ad libitum access to water and a starter. During the experiment, the calves were fed with two meals of milk at 8:00 and 16:00 at the rate of 2 kg per meal. The dry matter intake, daily gain, and feed conversion ratio (FCR) were determined in calves. Blood samples were taken from the jugular vein on 7, 43, and 77 days of age almost 4-5 hours after morning feeding. The collected blood was poured into two separate tubes, one containing heparin to obtain plasma and the other without heparin to obtain serum. Plasma and serum samples were kept at −80°C until the measurement of the studied parameters. Blood concentrations of calcium and phosphorus, aspartate aminotransferase, alkaline aminotransferase, alanine aminotransferase, superoxide dismutase, creatine phosphokinase, and lipid parameters (triglyceride, cholesterol, high-density lipoprotein, low-density lipoprotein, and very low-density lipoprotein) were measured using commercial kits. A Mindray BS-800 Chemistry Analyzer - BS-800 was used to measure the amounts of Zn, Fe, and Cu in blood. Data were analyzed in a completely randomized design using SAS.

There were no significant differences between treatments for daily weight gain and dry matter intake at 21, 35, 45, and 63 days of age (P > 0.05). At 77 days of age, daily weight gain and feed intake in calves received Zn supplements as Zn sulfate, Zn hydroxychloride, and organic Zn were significantly higher than the control group (P < 0.05). However, there were no  significant differences between different sources of Zn supplemented to the groups (P > 0.05). Body weight and FCR of the calves during the suckling period were not affected by Zn supplementation (P > 0.05). There were significant increases in the serum Zn and alkaline phosphatase concentrations of the calves supplemented with Zn sulfate, Zn methionine, and Zn organic compared to the control group (𝑃 < 0.05). Serum concentrations of Fe, Cu, Ca, P, aspartate aminotransferase, alanine aminotransferase, creatine phosphokinase, and lipid parameters (triglyceride, cholesterol, high-density lipoprotein, low-density lipoprotein, and very low-density lipoprotein) were not affected by the treatments (P > 0.05). However, the superoxide dismutase activity of the Zn-supplemented groups showed a numerical increase compared to the control group (P = 0.08).

The results showed that adding 20 mg/kg of Zn DM as Zn sulfate, Zn hydroxychloride, and organic Zn increased serum Zn and alkaline phosphatase concentrations and, finally, improved dry matter intake and daily weight gain. However, there were no significant differences between the Zn-supplemented groups for serum Zn and alkaline phosphatase concentrations, dry matter intake, and daily weight gain.

Ruminants and their products play an important role in providing food for many people. They are able to convert plants that cannot be digested by humans into usable protein for humans. New animal feed has been commonly developed for long years. To improve food production from ruminants, it is important to explore new feed sources. New or less exploited types of feed can replace or supplement traditional feeds in ruminant diets. Sesbania is a multipurpose plant that is cultivated for consumption as fodder and green manure. As a fast-growing plant, Sesbania has been mentioned as a food source to reduce feed costs. The food value of some of its species is equal to alfalfa and several other valuable plants. Due to proper digestibility, low fiber, and relatively high protein, it is possible to replace it with hay for livestock, especially small ruminants. In addition, as a tropical legume, Sesbania can easily withstand temperatures above 35°C, which are limiting for most legume plants, and grow well under salinity stress conditions if there is sufficient moisture in the soil. Therefore, this research aimed to investigate the nutritional value of Sesbania sp. L. for ruminants.

The present research was carried out in the laboratories and teaching-research station of animal husbandry at the Agricultural Sciences and Natural Resources University of Khuzestan. The experimental treatments included a control diet (without the Sesbania plant), 25, 50, 75, and 100% replacements of the Sesbania plant with alfalfa. The chemical compositions of the Sesbania plant, alfalfa hay, treatments, and other experimental samples, including crude protein, dry matter, ash, acid detergent fiber, and total tannin, were measured by standard methods. The neutral detergent fiber was quantified with the usual method without using the alpha-amylase enzyme and sodium sulfite by removing ash. The amount of gas produced by the samples was recorded at 2, 4, 6, 8, 10, 12, 24, 48, 72, and 96 hours after incubation, and gas production parameters, digestibility of organic matter, their metabolizable energy, ammonia nitrogen concentration, pH, and protozoa population of culture medium were estimated in the treatments. In the gas production experiment, rumen fluid was taken from the rumen of three male sheep fed with a forage-based diet at the maintenance level and before morning feeding using a stomach tube. Ruminal fluid was filtered with a four-layer cheesecloth, mixed, and transferred to the laboratory in a flask containing hot water at 39 °C. To count protozoa at the end of 24 hours, a sample was prepared from the gas culture medium and mixed with 10 ml of 10% formaldehyde, and protozoa were counted using a hemocytometer slide. The resulting data were analyzed with a completely randomized design. In the present experiment, the chemical composition, digestion, and fermentation of the Sesbania plant were also compared with alfalfa hay.

Compared to the control, Sesbania-containing diets replaced with alfalfa had significantly the lowest potential and gas production rate at all levels (P < 0.05). The comparison of the Sesbania plant and alfalfa hay showed that the potential and gas production rate of alfalfa were higher than that of the Sesbania plant. In the experimental diets, the partitioning factor, microbial biomass production, and microbial biomass production efficiency were higher at all levels of Sesbania replaced with alfalfa than the control diet, and the levels of 75% and 100% of Sesbania replaced with alfalfa had a significantly higher partitioning factor than the control and other treatments (P < 0.05). However, the truly degradable organic matter was not affected by the experimental treatments. The results of the comparison of the Sesbania plant and alfalfa hay showed that the partitioning factor, microbial biomass production, and microbial biomass production efficiency of the Sesbania plant were significantly higher than alfalfa (P < 0.05). The results of fermentation parameters after 24 hours of incubation showed that the pH of the rumen culture medium was not affected by the experimental treatments, but the ammonia nitrogen concentration was affected by the experimental treatments (P < 0.05), and the control treatment contained the highest ammonia nitrogen concentration. In the comparison of the Sesbania plant with alfalfa hay, the pH of the rumen culture medium was not affected by the experimental treatments, and the ammonia nitrogen concentration of the Sesbania plant was significantly lower than that of alfalfa hay (P < 0.05). The helotrich, entodinomorph, and cellulolytic protozoa population, and the total rumen protozoa population were the highest in the control treatment (29.33) and the lowest in the 100% replacement of the Sesbania plant with alfalfa hay (9.66). The results of the comparison of the Sesbania plant with alfalfa hay showed an increase in the population of Helotrich, Entodinomorph, and Cellulolytic protozoa and the total population of rumen protozoa in alfalfa hay, with a decrease in the population of the Sesbania plant. The results of in vitro digestibility indicated the highest digestibility of dry matter, NDF, and, ADF in the control treatment. The comparison of the Sesbania plant with alfalfa hay revealed that the digestibility of dry matter, NDF, and ADF of alfalfa was higher than that of the Sesbania plant.

In general, the results of the present experiment demonstrate that the chemical composition and digestibility of the Sesbania plant are similar to alfalfa hay. Therefore, the Sesbania plant can be considered a suitable substitute for alfalfa hay.

In the poultry industry, additives are used to stimulate growth, improve production performance, and improve the health of the flock. On the other hand, reducing the quality of food items and their protein level, as well as the high level of anti-nutritional substances, disturbs the balance of the microbial ecosystem of the digestive system of birds and health and ultimately reduces their performance. The health and nutritional status of poultry is largely dependent on the microbial flora of the digestive system, which directly and indirectly affects the morphology of the digestive system, nutrition, and immune system. The microbial flora of the intestine is relatively unstable and is easily affected by various nutritional factors. Therefore, this research aimed to investigate the effect of the simultaneous use of organic acids and prebiotics in diets containing low-quality soybean meal on the performance of laying hens.

This experiment was conducted using a completely randomized design in a 2 x 2 factorial arrangement using two levels of mixed organic acids and prebiotics and two types of soybean meal in the diet of laying hens. Chickens were distributed among pens based on weight and production rates for 2 weeks. Then, 192 laying hens of the LSL strain at the end of 73 weeks of age, after weighing and entering the wing number, were placed in 48 experimental cages, including four treatments with 12 replications, and fed for 12 weeks. Experimental treatments were treatment I (without the combination of organic acids and probiotics and containing 42% protein soy), treatment II (without the combination of organic acids and probiotics and containing 38% protein soy), treatment III (the combination of organic acids and probiotics and containing 42% protein soy), and treatment IV (a combination of organic acids and probiotics with 38% protein soy). To add the combination of organic acids and prebiotics, organic acid was used at a level of 1 kg per ton. The composition of the product included 30% propionic acid, 10% butyric acid, and 15% acetate plus 20% prebiotic with the origin of the Saccharomyces cerevisiae yeast cell wall. Performance traits, including production percentage and egg weight, egg mass weight, and feed conversion ratio (FCR) were measured at the end of the experiment (age 85 weeks) when all the birds were killed using the cervical vertebrae displacement method. The abdominal fat response of the immune system, histomorphology of the small intestine, and microbial population of the cecum were the investigated parameters.

Except for the first week of the experiment, the addition of the combination of organic acids and probiotics and the use of high-quality soybean meal in the diet significantly increased the egg-laying percentage during the rest of the experiment (p < 0.01). In the first 4 weeks of the experiment (weeks 75-78), the percentage of egg production was under the significant influence of the interaction effects of the experimental factors. The treatment with organic acids plus prebiotics and high-quality soybean meal (treatment 3) presented the highest egg production rate (p< 0.05). From the second week to the end of the experiment, the consumption of diets containing the combination of organic acids and probiotics and high-quality soybean meal significantly increased the weight of the produced eggs (p< 0.05). The interaction effects of experimental treatments during the experimental period did not show a constant trend in the weight of produced eggs (p≥ 0.05). Adding the combination of organic acids and probiotics to the diet elevated egg mass in the entire experimental period, the difference of which was significant (p< 0.05). The use of soybean meal with low-protein soybean meal in the diet during the experimental period decreased the weight of the egg mass compared to the group consuming the diet containing high-quality soybean meal (p< 0.05). Adding the combination of organic acids and probiotics and the use of high-quality soybean meal in the diet of the studied laying hens improved the FCR during the test period (p< 0.05). The FCR in the first 5 weeks of the experiment was under the interaction effects of the experimental factors (p< 0.05). Abdominal fat at the beginning (age 73 weeks) and the end of the experiment (age 85 weeks) did not show significant differences between different treatments (p≥ 0.05). Cellular immune response to DNCB injection significantly increased using the organic acids and prebiotic mixture and decreased with the presence of low-protein soybean meal in the diet (p < 0.05). The microbial population of the cecum changed significantly under the influence of the interaction effects of the mixture of organic acids and prebiotics and the quality of the feed soup (p < 0.05). The use of a mixture of organic acids and prebiotics caused a significant increase in the height of the duodenal villi (p< 0.05). The presence of low-protein soybean meal in the diet significantly elevated the depth of crypts (p< 0.05). The ratio of villus height to crypt depth changed under the significant influence of the interaction effects of the experimental factors. The highest ratio was recorded in the treatment with a mixture of organic acids and prebiotics and high-quality soybeans in the diet while the lowest ratio was measured in the treatment without a mixture of organic acids and prebiotics and low-protein soybeans in the diet (p< 0.05). The width of the duodenal villi did not change under the influence of experimental factors, with no significant differences (p≥ 0.05).

In general, the results showed that using a combination of organic acids and probiotics in the diet of laying hens improved their productive performance and the health of their digestive system, as well as the performance of chickens fed with low-protein soybean meal.

Milk is a complex physiological and biological liquid that contains water, protein, lactose, fat, vitamins, and minerals. Milk proteins (lactoferrin and lactoperoxidase) are known to have antimicrobial and immunomodulatory effects. More than half of the world's population consumes buffalo milk and milk products. Due to its unique properties, buffalo milk is highly regarded in developing countries and has been cultured in Iran for a long time. The main proteins of milk are casein and whey. Casein exists in the form of micelles, and 80% of milk proteins include alpha S1 casein, alpha S2 casein, beta casein, and kappa casein. Whey proteins comprise about 20% of milk proteins, which have four main components, including betalactoglobulin, alphalactalbumin, bovine serum albumin, and immunoglobulin. Lactoferrin, peptone protease, calmodulin, prolactin, and folate-binding proteins are also found in milk and are considered accessory proteins. Milk proteins have positive effects on different structures of the body, such as the immune, nervous, digestive, and cardiovascular systems. In addition, milk proteins are considered to contain a wide variety of biologically active peptides with superior nutritional and immunological activities. Therefore, obtaining complete information about the physicochemical properties of milk from different species and its nutritional value seems essential for liquid milk technology. Milk proteins have been analyzed continuously, and the information about this very important and complex system has been increasingly rising in recent years, mainly due to technological advances. In addition, the scope of research work on the analysis of proteins in milk has increased significantly due to the increased knowledge of bioactive proteins in milk. The present study aimed to measure the total protein values of cow and buffalo milk samples in the climatic conditions of Khuzestan province. The electrophoretic pattern of buffalo milk proteins was also compared with cow milk.

Milk samples were collected from 30 apparently healthy cows and 30 buffaloes from the surrounding villages of Shushtar City and cattle farms of Masjid Suleiman City. Raw milk proteins were then evaluated by the Kjeldahl method. To measure milk proteins, milk was first digested with the addition of sulfuric acid with gentle heat. Complete digestion continued until the milk became colorless. After cooling, distillation was done by adding boric acid and methyl red, and the contents were titrated with 0.1 normal tetrazole sulfuric acid. The first color change marked the end of the titration, followed by calculating the protein content. After separating milk serum, the milk protein content in these two types of animals was compared with the sodium dodecyl sulfate (SDS) plate method. In this technique, milk protein molecules are linearized using SDS, and their movement is based on weight, that is, the larger the protein, the shorter the traveled distance. The relationship between the distance traveled by proteins and the logarithm of their molecular weight was investigated and milk proteins were identified in the samples. T-test statistical analysis was used to compare the average total protein in cows and buffaloes at a 95% confidence level as the statistical significance (p < 0.05).

In sodium dodecyl sulfate plates, standard samples were able to create 14, 18, 22, and 70 kD bands for alpha-lactalbumin, beta-lactoglobulin, casein, and albumin proteins according to  the protein marker. The average protein contents of buffalo and cow milk samples were equal to 4.71±1.79 and 3.91±1.19 mg/liter, respectively, meaning that the total protein content in buffalo milk is significantly higher than that of the cow (p < 0.05). In addition, cow's milk contained lower levels of beta-lactoglobulin and casein than buffalo's milk while the amount of alpha-lactalbumin in cow's milk was higher than in buffalo's milk. The bands obtained from the electrophoresis of casein and alpha-lactalbumin on the gel showed genetic diversity among the subunits of casein and alpha-lactalbumin in cow and buffalo milk samples.

In the comparison of the protein profile in animal milk, various reports show that protein profiles of milk are different in animal species. So far, the protein profile of buffalo milk has not been compared in the literature, which was investigated in this study. In general, the present study shows that cow and buffalo milk are different in terms of the protein profile due to the higher total protein content, especially the casein and beta-lactoglobulin levels in buffalo's milk than in cow's milk. It seems that the nutritional value of buffalo milk is higher than cow milk.