Cathelicidins are antimicrobial peptides (AMPs), which together with defensins form a large group of cationic peptides. They are part of the immune system of many vertebrates, including humans and farm animals. Cathelicidins are primarily found in cells of the immune system, particularly in macrophage granules. This peptide family is important in the host immune system because it takes part in the immune response against pathogens, including bacteria (Gram-positive and Gram-negative), viruses and fungi. Because of their antimicrobial activity, cathelicidins play an essential role in immunomodulation,
Cathelicidins are secreted in epidermal cells and glands of various organs – on the skin and in the gastrointestinal oral and nasal cavity, and udder lining epithelia. Secretion of these proteins in sweat, milk, and saliva has also been noted (24).
Genes encoding cathelicidins have been found in many animals, both invertebrates and vertebrates, but especially in mammals, in which more than 30 members of the cathelicidin family have been identified (18). One cathelicidin has been described in humans, while in cattle seven cathelicidin genes have been identified on chromosome 22 (36). The genes encoding proteins from the cathelicidin family are composed of four exons divided by three introns. The structure of each bovine cathelicidin is similar. It consists of three main elements: the N-terminal signal peptide, the cathelin domain, and the C-terminal variable region, which is responsible for the functions of cathelicidins, particularly their antibacterial, antiviral, antifungal, and immunomodulatory activity. The first three exons encode both the signal peptide and the cathelin domain, while the variable region is encoded by exon 4. The variable region’s role validates investigating the potential influence of polymorphisms in exon 4 on selected performance traits in a group of animals (2).
Cathelicidins are increasingly attracting attention because of their functions in various animal species, including mammals, amphibians, and birds. Pig cathelicidin research by Ahn
In dairy cattle breeding, it is crucial to ensure the highest milk quality, to meet the expectations of consumers. Milk quality is influenced by both genetic and non-genetic factors, including nutrition, housing, hygiene and milking method (9). One of the most common problems in dairy cattle farming is mastitis, caused by numerous pathogenic bacteria. The infection causes a reduction in milk production, resulting in further economic losses in addition to those arising from the cost of veterinary treatment and from deaths in the herd. For this reason, it is essential to improve udder health by introducing tools for monitoring and prevention of infection (23).
Mastitis is one of the most commonly diagnosed diseases in dairy cattle. The most common aetiological agent of mastitis is bacteria of the
Mastitis is manifested by an increase in the somatic cell count (SCC) in milk. A study by Smolenski
The search for dairy cattle genes of which variants influence animal performance has intensified over the years. Scientists are attempting to identify SNPs of various genes which can have a significant impact on the production levels of livestock, including dairy cows, focusing mainly on their potential effects on milk yield, protein and fat content and on milk SCC. In addition to the use of genetic tests to assess dairy performance, it is worth emphasising their potential use to improve the health of farm animals, and thus the quality of the products obtained from them (26).
Gillenwaters
Marker-assisted selection is very useful due to its multi-faceted applications in breeding. It can be used to analyse traits that may be sex restricted, such as milk yield and performance traits, and can be a valuable tool supporting standard selection programmes with the potential to improve the genetic profile of the herd (34).
Essential techniques, such as the polymerase chain reaction (PCR) with its various modifications, are helpful tools enabling genotyping of animals in terms of selected genes and their potential impact on individual performance traits. The results can be used to implement marker-assisted selection. This makes it possible to perform a series of tests to search for and assess interesting genetic markers that could potentially be included in research aimed at improving the selection and breeding of animals (15).
Special attention should be focused on studies related to SCC, due to the cell count’s potential role in the selection of cows showing increased resistance to pathogens or in the development of rapid tests for identification of mastitis, including subclinical forms, in milk. Wollowski
There is a need for further knowledge on the potential positive impact of cathelicidins on the condition, health, and production characteristics of animals. The study aimed to estimate the effect of polymorphisms occurring within the
The material consisted of 279 Polish Black-and-White Holstein-Friesian dairy cows, including 56 heifers, all in their second lactation. The animals were kept on a farm located in the Opolskie voivodeship. They were housed in a stand-alone system and fed using total mixed rations. They were milked twice a day using an automatic milking system and provided the investigated milk samples in the summer of 2022. Data on milk yields of cows were obtained from records of dairy performance evaluation by the Polish Federation of Cattle Breeders and Milk Producers. None of the cows suffered from clinical mastitis during the study period, while subclinical mastitis was detected in 7% of animals.
The SCC was log-transformed (LnSCC) according to Ali and Shook (4) and expressed as thousands/mL of milk, so that the conditions of normal distribution were met for this parameter as well as for daily milk yield (kg) and protein and fat content (%) in milk.
The first step in the analysis was DNA extraction. Peripheral blood was collected from the jugular vein of each cow. Sterile tubes containing the K3 ethylenediaminetetraacetic acid anticoagulant were used to preserve the sample. Isolation of DNA was achieved using the whole blood MasterPure DNA isolation kit (Epicenter Technologies, Mumbai, India) according to the manufacturer’s recommendations.
The genotypes of individual animals were analysed by PCR-restriction fragment length polymorphism. Two polymorphic sites within exon 4 of the
The PCR reaction conditions, product length, restriction enzymes used for digestion, genotypes, and length of the fragments after restriction enzyme cleavage are shown in Table 1.
PCR conditions, product size, restriction enzyme, and fragment size after restriction endonuclease digestion
SNP | PCR Conditions | Product size | Restriction enzyme | Genotype | Fragment size after digestion |
---|---|---|---|---|---|
initial: 95ºC/5 min | 324, 23 bp |
||||
2,383 |
347, 324, 23 bp |
||||
35 cycles: denaturation:95ºC/30 s | 347 bp |
||||
annealing: 52ºC/45 s extension:72ºC/30 s | 347 bp | 161, 81, 58, 34, 14 bp |
|||
2,468 |
219, 161, 81, 58, 34, 13 bp |
||||
final extension: 72ºC/7 min | 219, 81, 34, 13 bp |
bp – base pairs
The next stage of the analysis was digestion of the PCR products with restriction enzymes
Agarose gel electrophoresis (3%) of PCR-restriction fragment length polymorphism result for
Agarose gel electrophoresis (3%) of PCR-restriction fragment length polymorphism result for
Statistical analysis of the results included the relationship between different genotypes, different combinations of genotypes, and selected performance parameters,
Statistical analysis of the relationship between
The following general linear model was used: Y
where Y
Three genotypes were identified for
Genotype and allele frequencies of
N | Genotype frequencies | Allele frequencies | χ2 | |||
---|---|---|---|---|---|---|
169 | 0.606 | |||||
91 | 0.326 | 0.769 | 1.897 |
|||
19 |
0.068 |
0.231 |
||||
144 | 0.516 | |||||
115 | 0.412 | 0.722 | 0.208 | |||
20 | 0.072 | 0.278 |
N – number of cows
The results obtained for the relationship between the genotypes of the polymorphisms and selected dairy performance parameters are presented in Table 3. In the case of the
Means and standard deviation of traits in relation to genotypes
N | Genotype | Milk yield kg | Fat content % | Protein content % | Lactose content % | LnSCC |
---|---|---|---|---|---|---|
138 | 33.93A ± 7.86 | 3.84A ± 0.72 | 3.52A ± 0.33 | 4.95AB ± 0.16 | 4.18A ± 0.82 | |
68 | 33.81B ± 8.18 | 3.80B ± 0.75 | 3.52B ± 0.35 | 4.89B ± 0.17 | 4.28 ± 0.84 | |
17 |
30.42AB ± 6.29 |
4.07AB ± 0.70 |
3.59AB ± 0.37 |
4.90A ± 0.16 |
4.34A ± 0.75 |
|
117 | 33.48 ± 7.59 | 3.87 ± 0.71 | 3.53A ± 0.34 | 4.94a ± 0.16 | 4.18 ± 0.85 | |
91 | 33.87 ± 8.34 | 3.81 ± 0.77 | 3.51a ± 0.34 | 4.91a ± 0.16 | 4.27 ± 0.81 | |
15 | 34.42 ± 7.89 | 3.80 ± 0.67 | 3.47Aa ± 0.31 | 4.92 ± 0.17 | 4.25 ± 0.73 |
N – number of cows; LnSCC – natural log somatic cell count; a – values in a row with different letters differ significantly (P ≤ 0.05); A, B – values in a row with different letters differ highly significantly (P ≤ 0.01)
In the case of the
Research to identify candidate genes associated with dairy performance parameters in cattle is carried out in order to improve animal welfare and selection programmes (29). The SCC in the milk of dairy animals is a good indicator of quality and udder health (30). An increase in SCC is a symptom of clinical or subclinical mastitis, which can cause significant economic losses and have a long-term impact on the health of the entire herd (12).
To increase the efficiency of breeding for dairy performance, researchers are exploring the potential uses of genetic testing for genes encoding proteins that may affect milk performance parameters (1). A study conducted on the
The results of the present study suggest a relationship between the polymorphisms tested and parameters of milk performance. In the case of
These polymorphisms are mapped in exon 4, which is responsible for encoding the variable region of the BMAP-34 protein. The
Research on the potential use of individual SNPs as markers has been conducted using various cattle breeds to study various performance traits. Ateya
Khan
There are relatively few scientific reports on the potential link between cathelicidins and performance parameters. There is great interest in the development of tests for the rapid diagnosis of mastitis in cows by detecting cathelicidins in their milk, as the concentration of cathelicidins increases during the course of mastitis. BMAP proteins have been shown to have antimicrobial functions,
Identification of genes affecting resistance to mastitis has been of interest to researchers for many years. Griffin
The characteristics of the proteins encoded by cathelicidin family genes make them potential targets in the search for genetic markers. There are studies of the relationships between polymorphisms within these proteins and studies of various parameters and relationships, focusing mainly on the relationship between the host immune system and cattle performance, including the influence of polymorphisms on meat performance parameters.
The search for genetic markers for use in livestock breeding programmes is a priority task for researchers. Cathelicidins are proteins performing numerous functions in the body and may be genetic markers which lend themselves to use in such programmes. Due to their antimicrobial activity, it is essential to learn more about their potential effects. Particularly noteworthy are potential relationships between polymorphisms within the