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CD14 is an important player in host innate immunity in that it confers lipopolysaccharide sensitivity to cell types like neutrophils, monocytes and macrophages. The study was aimed at characterizing the CD14 gene of cattle for sequence variations and to determine the effect of variations on the expression of the protein on the surfaces of monocytes...
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Context 1
... of the CD14 sequences of 106 Canadian Holsteins and 46 Jersey cows with published sequences (GenBank Nos. NW_001495367 and D84509) revealed a total of five SNPs including one in the 5' untranslated region (UTR) (g.C1291T, numbering is according to Gen- Bank No. EU148609), two in the coding regions (g.A1908G and g.A2318G) and two in the 3' UTR (g.A2601G and g.G2621T) ( Table 1). Four of the SNPs are transitional mutations while SNP 2621 involves the trans- version of guanine to thymine. ...
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... was absent in Jersey while CGAAG was absent in both populations. The gene alleles A1 and B were observed from the sequencing data to be associated with SNPs T 1291 , G 2601 and T 2621 while allele A was associated with C 1291 , A 2601 and G 2621 therefore giving rise to three actual haplotypes (TAGGT, TGGGT and CAAAG) in the analyzed populations (Table 1). The frequency of the haplotype associated with allele A (CAAAG) was highest (65.8%) in Holsteins while the haplotype associated with allele A 1 (TAGGT) was highest in Jerseys (80.2%) ( Table 1). ...
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... gene alleles A1 and B were observed from the sequencing data to be associated with SNPs T 1291 , G 2601 and T 2621 while allele A was associated with C 1291 , A 2601 and G 2621 therefore giving rise to three actual haplotypes (TAGGT, TGGGT and CAAAG) in the analyzed populations (Table 1). The frequency of the haplotype associated with allele A (CAAAG) was highest (65.8%) in Holsteins while the haplotype associated with allele A 1 (TAGGT) was highest in Jerseys (80.2%) ( Table 1). ...
Citations
... Polymorphisms in the bovine CD14 gene have been identified, some of them potentially linked to increased resistance against mastitis [33][34][35]. Such polymorphisms could have consequences on the activity of CD14 but also, when located in the promoter region for example, on the level of sCD14 in milk as has been observed in women breast milk [36]. ...
Bovine mastitis remains a major disease in cattle world-wide. In the mammary gland, mammary epithelial cells (MEC) are sentinels equipped with receptors allowing them to detect and respond to the invasion by bacterial pathogens, in particular Escherichia coli . Lipopolysaccharide (LPS) is the major E. coli motif recognized by MEC through its interaction with the TLR4 receptor and the CD14 co-receptor. Previous studies have highlighted the role of soluble CD14 (sCD14) in the efficient recognition of LPS molecules possessing a full-length O-antigen (LPSS). We demonstrate here that MEC are able to secrete CD14 and are likely to contribute to the presence of sCD14 in milk. We then investigated how sCD14 modulates and is required for the response of MEC to LPSS. This study highlights the key role of sCD14 for the full activation of the Myd88-independent pathway by LPSS. We also identified several lncRNA that are activated in MEC in response to LPS, including one lncRNA showing homologies with the mir-99a-let-7c gene (MIR99AHG). Altogether, our results show that a full response to LPS by mammary epithelial cells requires sCD14 and provide detailed information on how milk sCD14 can contribute to an efficient recognition of LPS from coliform pathogens.
... Polymorphisms in the bovine CD14 gene have been identi ed, some of them potentially linked to increased resistance against mastitis [31][32][33] . Such polymorphisms could have consequences on the activity of CD14 but also, when located in the promoter region for example, on the level of soluble CD14 in milk as has been observed in women breast milk 34 . ...
Bovine mastitis remains a major disease in cattle world-wide. In the mammary gland, epithelial cells are sentinels equipped with receptors allowing them to detect and respond to the invasion by bacterial pathogens, in particular Escherichia coli . Lipopolysaccharide (LPS) is the major E. coli motif recognized by MEC through its interaction with the TLR4 receptor and the CD14 co-receptor. Previous studies have highlighted the role of soluble CD14 in the efficient recognition of LPS molecules possessing a full-length O-antigen (LPSS). We demonstrate here that MEC are able to secrete CD14 and are likely to contribute to the presence of soluble CD14 in milk. We then investigated how CD14 modulates and is required for the response of MEC to LPSS. This study highlights the key role of soluble CD14 for the full activation of the Myd88-independent pathway by LPSS. We also identified several long non-coding RNA that are activated in MEC in response to LPS, including one lncRNA showing homologies with the mir-99a-let-7c gene (MIR99AHG). Altogether, our results provide detailed information on how milk soluble CD14 can contribute to an efficient recognition of LPS from coliform pathogens.
... Polymorphisms in the bovine CD14 gene have been identi ed, some of them potentially linked to increased resistance against mastitis [31][32][33] . Such polymorphisms could have consequences on the activity of CD14 but also, when located in the promoter region for example, on the level of soluble CD14 in milk as has been observed in women breast milk 34 . ...
Bovine mastitis remains a major disease in cattle world-wide. In the mammary gland, epithelial cells are sentinels equipped with receptors allowing them to detect and respond to the invasion by bacterial pathogens, in particular Escherichia coli . Lipopolysaccharide (LPS) is the major E. coli motif recognized by MEC through its interaction with the TLR4 receptor and the CD14 co-receptor. Previous studies have highlighted the role of soluble CD14 in the efficient recognition of LPS molecules possessing a full-length O-antigen (LPSS). We demonstrate here that MEC are able to secrete CD14 and are likely to contribute to the presence of soluble CD14 in milk. We then investigated how CD14 modulates and is required for the response of MEC to LPSS. This study highlights the key role of soluble CD14 for the full activation of the Myd88-independent pathway by LPSS. We also identified several long non-coding RNA that are activated in MEC in response to LPS, including one lncRNA showing homologies with the mir-99a-let-7c gene (MIR99AHG). Altogether, our results provide detailed information on how milk soluble CD14 can contribute to an efficient recognition of LPS from coliform pathogens.
... Chen et al., [16], found in a Chinese Holstein population polymorphism in the coding region of the LYZ gene associated with SCS, proposing its use as a candidate marker for mastitis. Ibeagha-Awemu et al. [17] reported a significant association of genotype in 1908 non-synonym locus to a higher percentage of neutrophils expressing CD14 molecules on their surfaces and suggesting an important role of CD14 in mediating bacterial infections. On this basis, Kumar et al. [18] demonstrated a significant association between polymorphisms in CL14 and mastitis frequency in Sahiwal cows. ...
Background
Lactoferrin (LTF) is an iron-binding glycoprotein found in milk and other exocrine secretion with antibacterial activity proposed as an alternative to mastitis treatment or prevention. LTF has been proposed as a candidate gene for mastitis resistance selection. The aim of this paper was to assess LTF promotor to explore variations with potential association to mastitis resistance in dairy cows from Honduras.MethodsA resequencing of promotor and Exon I of LTF gene in extreme mastitis susceptibility cows (126 Holstein and Holstein crossbred) was performed.ResultsEight polymorphisms were found in promotor region, four of them were novel variations. Two were important by frequency among extreme groups, but a polymorphism in − 421 A/T position was significantly (P = 0.0188) associated to mastitis susceptibility.Conclusion
Results support the key role of regulatory region of LTF gene. Some candidate genes are proposed in association with mastitis traits and implications are discussed.
... The CD14 gene product is an important part of host innate immunity. It provides sensitivity to lipopolysaccharides to individual cell types, such as neutrophils, monocytes, and macrophages (Wang et al., 2006;Ibeagha-Awemu et al., 2008;Wu et al., 2019b). The CD14 gene product mediates host defences against Gram-negative bacterial infections, provides immunity against viral infections, and determines the effect of changes in protein expression on the surface of monocytes and neutrophils in healthy dairy cows. ...
... It occurs on the cell membrane of monocytes and, to a lesser extent, on the membrane of neutrophils. CD14 ensures the sensitivity of cells to lipopolysaccharide (LPS), including epithelial cells and endothelial cells (Wang et al., 2002;Ibeagha-Awemu et al., 2008;Wu et al., 2019b). LPS, when bound to host membrane proteins such as CD14, causes the release of proinflammatory cytokines that recruit neutrophils as an early innate immune response. ...
... There are two forms of the gene product, the membranebound form of mCD14 and the soluble form of sCD14 (Ibeagha-Awemu et al., 2008;Hirsch et al., 2021). sCD14 is thought to be derived from monocytes by direct exocytosis and from proteolytic cleavage of mCD14 on the cell surface. ...
Mastitis is one of the most important diseases of the mammary gland. The increased incidence of this disease in cows is due to the breeding of dairy cattle for higher yields, which is accompanied by an increased susceptibility to mastitis. Therefore, the difficulty involved with preventing this disease has increased. An integral part of current research is the elimination of mastitis in order to reduce the consumption of antibiotic drugs, thereby reducing the resistance of microorganisms and decreasing companies' economic losses due to mastitis (i.e. decreased milk yield, increased drug costs, and reduced milk supply). Susceptibility to mastitis is based on dairy cows' immunity, health, nutrition, and welfare. Thus, it is important to understand the immune processes in the body in order to increase the resistance of animals. Recently, various studies have focused on the selection of mastitis resistance genes. An important point is also the prevention of mastitis. This publication aims to describe the physiology of the mammary gland along with its immune mechanisms and to approximate their connection with potential mastitis resistance genes. This work describes various options for mastitis elimination and focuses on genetic selection and a closer specification of resistance genes to mastitis. Among the most promising resistance genes for mastitis, we consider CD14, CXCR1, lactoferrin, and lactoglobulin.
... For PY1, the cluster differentiation antigen 14 (CD14) located on BBU9 encodes a membrane-associated protein on the surface of cells which is involved in the recognition of LPS, playing a role in mediating signals from toll-like receptors (Ibeagha-Awemu et al., 2008;Jin and Lee, 2008). In addition, the recombinant bovine soluble (sCD14) form can sensitize mammary epithelial cells, indicating an important role in initiating host responses to gram-negative bacterial infections (Wang et al., 2002). ...
Genomic selection has been widely implemented in many livestock breeding programs, but it remains incipient in buffalo. Therefore, this study aimed to (1) estimate variance components incorporating genomic information in Murrah buffalo; (2) evaluate the performance of genomic prediction for milk-related traits using single-and multitrait random regression models (RRM) and the single-step genomic best linear unbi-ased prediction approach; and (3) estimate longitudinal SNP effects and candidate genes potentially associated with time-dependent variation in milk, fat, and protein yields, as well as somatic cell score (SCS) in multiple parities. The data used to estimate the genetic parameters consisted of a total of 323,140 test-day records. The average daily heritability estimates were moderate (0.35 ± 0.02 for milk yield, 0.22 ± 0.03 for fat yield, 0.42 ± 0.03 for protein yield, and 0.16 ± 0.03 for SCS). The highest heritability estimates, considering all traits studied, were observed between 20 and 280 d in milk (DIM). The genetic correlation estimates at different DIM among the evaluated traits ranged from −0.10 (156 to 185 DIM for SCS) to 0.61 (36 to 65 DIM for fat yield). In general, direct selection for any of the traits evaluated is expected to result in indirect genetic gains for milk yield, fat yield, and protein yield but also increase SCS at certain lactation stages, which is undesirable. The predicted RRM coefficients were used to derive the genomic estimated breeding values (GEBV) for each time point (from 5 to 305 DIM). In general, the tuning parameters evaluated when constructing the hybrid genomic relationship matrices had a small effect on the GEBV accuracy and a greater effect on the bias estimates. The SNP solutions were back-solved from the GEBV predicted from the Legendre random regression coefficients, which were then used to estimate the longitudinal SNP effects (from 5 to 305 DIM). The daily SNP effect for 3 different lactation stages were performed considering 3 different lactation stages for each trait and parity: from 5 to 70, from 71 to 150, and from 151 to 305 DIM. Important genomic regions related to the analyzed traits and parities that explain more than 0.50% of the total additive genetic variance were selected for further analyses of candidate genes. In general, similar potential candidate genes were found between traits, but our results suggest evidence of differential sets of candidate genes underlying the phe-notypic expression of the traits across parities. These results contribute to a better understanding of the genetic architecture of milk production traits in dairy buffalo and reinforce the relevance of incorporating genomic information to genetically evaluate longitudinal traits in dairy buffalo. Furthermore, the candidate genes identified can be used as target genes in future functional genomics studies.
... Milk neutrophils and monocyte/macrophages were differentiated from other cells by indirect fluorescent labeling by incubating for 30 min at room temperature with unlabeled primary mAb (Table 1). A gating strategy was used to differentiate granulocytes and monocytes ( Figure 1) because CD14, although abundantly expressed on the surface of monocytes, can also be expressed to a lesser extent on bovine PMN (Ibeagha-Awemu et al., 2008;Piepers et al., 2009), as previously described . Next, 1 mL of PBS was added to the cell suspension and centrifuged at 400 × g for 8 min. ...
... We accurately identified PMN, monocyte/ macrophage, and lymphocyte subsets, whereas others evaluating milk DCC by flow cytometry did not use CD14 and CH138A mAb in combination (Pillai et al., 2001;Rivas et al., 2001;Dosogne et al., 2003;Koess and Hamann, 2008;Schwarz et al., 2011;Pilla et al., 2013). Not doing so can lead erroneous identification of some PMN as monocyte/macrophages, considering that PMN can also express CD14 on their surface (Paape et al., 1996;Sladek et al., 2002;Ibeagha-Awemu et al., 2008). Even worse, some studies did not use a specific mAb to accurately differentiate and identify many milk cell types (Pillai et al., 2001;Dosogne et al., 2003). ...
We analyzed a large number of immune response parameters from quarter milk samples with distinct bacteriological and quarter somatic cell count (qSCC) statuses. Furthermore, we sought to explore and identify displayed immune response patterns in milk samples from mammary glands with nonspecific mastitis. Thus, 92 quarter milk samples from 28 cows were stratified into 4 groups, as follows: (1) 49 culture-negative control quarters with a low qSCC (<1 × 10⁵ cells/mL) from 19 dairy cows (so-called healthy quarters); (2) 15 culture-negative quarters with high qSCC (>2 × 10⁵ cells/mL; so-called quarters with nonspecific mastitis) from 10 dairy cows; (3) 8 culture-positive quarters with low qSCC (noninflammatory quarters with low qSCC) from 5 dairy cows; and (4) 20 culture-positive quarters with high qSCC (so-called truly infected quarters) from 8 dairy cows. Using flow cytometry, we evaluated the percentage of milk neutrophils and their viability, intracellular reactive oxygen species production, phagocytosis, and the expression of CD62L, CD11b, and CD44 for each of the 4 quarter strata. Furthermore, the percentage of monocyte/macrophages, B cells, and T lymphocyte subsets were evaluated by flow cytometry. Milk samples from bacteriologically negative quarters (both with a low and elevated qSCC) had a lower qSCC than those with bacteriologically positive outcomes (both with a low and elevated qSCC). As expected, the healthy quarters showed the lowest percentage of neutrophils and also showed a higher percentage of milk monocytes/macrophages and lower percentage of T lymphocytes than truly infected quarters. The most prominent result of the present study is that quarters with nonspecific mastitis showed the highest percentage of milk CD4⁺ T lymphocytes. The healthy quarters had a lower percentage of apoptotic neutrophils than noninflammatory and truly infected quarters, although it did not differ from those from the quarters with nonspecific mastitis. Our study supports the role of differential cell counting in the diagnosis of mastitis, as the milk leukocyte populations markedly fluctuate under healthy and inflammatory conditions. Furthermore, an increase in milk CD4⁺ T cells was associated with nonspecific mastitis, suggesting an increase in this leukocyte subpopulation is correlated with low bacterial shedding. Our study allows us to go further in our understanding of mammary gland immunity, providing further insights on potential protective mammary gland immunity, which we hypothesize can open new avenues for the development of novel targets that can promote bovine udder health.
... Characterization of bovine CD14 and association studies were conducted to explore the surface expression on monocytes and polymorphonuclear neutrophils, an indicator of immune status 43 It is understood from the current study that the mutation in CD14 gene causes unstable and deleterious mutations, which ultimately causes depressed function as immunity leading to the occurrence of diseases like mastitis in the current case. The variability identi ed in the CD14 gene sequence and SNPs and variants may be identi ed at the day old stage and employed for marker-assisted selection to retain the wild-type for the production of a future disease-free stock of the farm. ...
... . The effect of CD14 in in ammatory responses during mastitis had been depicted44,45 . Reports indicate association of CD14 SNPs with SCC46,43 . SNPs in the CD14 Promoter decreased the ability of Sp Protein Binding and increases Transcriptional Activity47 (LeVan et al., 2001). ...
Background: CD14 is an important pattern recognition receptor having innate immune function and has antibacterial activity. It binds with LPS of gram-negative bacteria, arachidonic acid, and lipoteichoic acid. Being a receptor, it binds with the pathogen with the help of other cytokines. Mutations in CD14 affect the binding ability which in turn affects the biological potentiality.
Method: The present study was conducted on 228 nos. of buffaloes pertaining to four different breeds as Murrah, Mehsana, Surti and Bhadawari. CD14 gene was characterized and polymorphism was detected through Single nucleotide conformation polymorphism. Association study was conducted for different variants of CD14 with mastitis in buffalo, detected through somatic cell count, california mastitis test.
Result:Eight variants of CD14 were detected and mutational hotspots were detected in bubaline CD14 with 58 number of non-synonymous SNP, out of which 18 were observed to be deleterious and 34 as thermodynamically unstable. In the present study, we had detected the mutations in CD14 gene and its association with the somatic cell score and other indicators for mastitis. In-silico studies were conducted to understand the molecular mechanism how the mutations affect the biological potentiality by analyzing different domains and structural analysis along with various post-translational modification sites.
Conclusion: Deleterious mutations were observed in CD14 gene which have significant effect on mastitis of buffalo. It may be employed for marker assisted selection, therapeutic application of recombinant CD14, gene therapy, transgenic or gene edited animal production with wild type CD14 resistant to mastitis as future strategy.
... CD14 is a 55kD glycosyl phosphatidylinositol-anchored surface protein majorly expressed on monocytes, polymorphonuclear leucocytes, and macrophages [11,12]. Published reports have revealed the importance of CD14 polymorphisms as candidates for studying diseases such as bovine tuberculosis and mastitis [13][14][15]. Studies have localized bovine health-related quantitative trait loci (QTL) to the genomic region of the CD14 loci [13,15]. ...
... Published reports have revealed the importance of CD14 polymorphisms as candidates for studying diseases such as bovine tuberculosis and mastitis [13][14][15]. Studies have localized bovine health-related quantitative trait loci (QTL) to the genomic region of the CD14 loci [13,15]. In addition, results from malaria studies have shown CD14 promoter polymorphism to mediate an adaptive protective mechanism against severe disease [16,17], including regulation of parasitemia [18]. ...
... To initiate a proper innate immune response to infection, pattern recognition receptors such as CD14, and signaling molecules such as Toll-like receptors (TLR's), are of significant importance, with the former presenting either as mCD14, expressed primarily on macrophages, dendritic cells and neutrophils, or sCD14 expressed in serum [20]. The expression regulation of CD14 gene is therefore critical for the recognition of bacterial lipopolysaccharide during innate immunity [13,15], particularly since CD14 mutant variant (-159T/T) expresses significantly higher soluble CD14 levels in serum than the homozygous or heterozygous genotypes [21]. We have utilized qualitative data on CD14 gene promoter to decipher immune response and susceptibility to malaria infection [18], showing a defective response by mutant variants, driving a cascade that worsens disease and multiple deleterious outcomes [22][23][24]. ...
Immune response to infections has been shown to be mediated by genetic diversity in pattern recognition receptors, leading to disease tolerance or susceptibility. We elucidated naturally occurring variations within the bovine CD14 gene promoter in trypanosome-tolerant (N’Dama) and susceptible (White Fulani) cattle, with genomic and computational approaches. Blood samples were collected from White Fulani and N’Dama cattle, genomic DNA extracted and the entire promoter region of the CD14 gene amplified by PCR. We sequenced this region and performed in silico computation to identify SNP variants, transcription factor binding sites, as well as micro RNAs in the region. CD14 promoter sequences were compared with the reference bovine genome from the Ensembl database to identify various SNPs. Furthermore, we validated three selected N’Dama specific SNPs using custom Taqman SNP genotyping assay for genetic diversity. In all, we identified a total of 54 and 41 SNPs at the CD14 promoter for N’Dama and White Fulani respectively, including 13 unique SNPs present in N’Dama only. The significantly higher SNP density at the CD14 gene promoter region in N’Dama may be responsible for disease tolerance, possibly an evolutionary adaptation. Our genotype analysis of the three loci selected for validation show that mutant alleles (A/A, C/C, and A/A) were adaptation profiles within disease tolerant N’Dama. A similar observation was made for our haplotype analysis revealing that haplotypes H1 (ACA) and H2 (ACG) were significant combinations within the population. The SNP effect prediction revealed 101 and 89 new transcription factor binding sites in N’Dama and White Fulani, respectively. We conclude that disease tolerant N’Dama possessing higher SNP density at the CD14 gene promoter and the preponderance of mutant alleles potentially confirms the significance of this promoter in immune response, which is lacking in susceptible White Fulani. We, therefore, recommend further in vitro and in vivo study of this observation in infected animals, as the next step for understanding genetic diversity relating to varying disease phenotypes in both breeds.
... Missense SNP are those that result in a change in the codon and amino acid sequence in the protein which led to the change the neuclotide position which led to the formation of a new amino acid (Medicine, 2012), as well as it affected on the formation of heat shock proteins 70 and agreed with study by (Yudin and Voevod, 2015) that indicated that genetic mutations in the heat shock protein 70 increase cattle resistance to various stress conditions. Additionally, previous studies indicated that the mutation produced by the amino acid Aspartic makes livestock more resistant to pathogens (Ibeagha et al., 2008). While, Bhat et al., (2016) who recorded SNP (G149T) substitution leads to a change of amino acids from Aspartate to Tyrosine in gene transcript and concluded that gene have been more resistant than other genotype in Tharparker cattle. ...
This study was undertaken in cattle field in the private sector in Al-Qurna, north of Basra Governorate, where it was conducted in the period between July 2018 to February 2019 and included the following seasons, the monthly summer season(July and August of 2018), the monthly autumn season (October and November of 2018) and the winter season is monthly (January and February of 2019). This experiment was designed to detect the effect of the THI value on genetic polymorphism of Hsp70 member A (HSPA1A) gene in the blood calves, and their relation with serum level of Hsp70 of local Holstein calves during different months and seasons. Twenty blood samples were collected monthly from the same Holstein male calves (n = 20) and revealed that : After DNA extracted from calves blood, A Polymerase Chain Reaction (PCR) for the amplification product 2956bp was carried out. Nucleotide sequence analysis was done to detect polymorphism and compared with the genotypes of the Hsp70 gene in the Gene Bank (accession number NM_203322.3.) There are two Genotypes of Hsp70 gene were obtained in calves : Group A: This genotype exposed to different mutation on different sites which include : On the site (A302 del and G504 del) nucleotide Adenine and Guanine have been deleted, and three missense mutation on positions (A329G, C359G andG506T) encoding to new amino acid, while the genetic mutation at the position (G367C and G403A) are silent (calves, n = 12). Group B: This genotype is the closest to heat shock protein Hsp70(HSPA1A) gene than to genotype A in the Holstein calves in Gene Bank. This genotype was exposed to genetic mutations at different position: On the site (G506A) one missense mutation encoding to new amino acid was recorded, while the rest mutation at sites (G367C and A504G) are silent did not encoded to new amino acid (calves, n = 8). The result of Multiple Sequences Alignment (MSA) of Hsp70 calves showed that the genotype A has identities 97%, while genotype B has identities 99% with Bous taraus heat shock protein 70 Member A (HSPA1A1) in Gene Bank. Significantly higher (P 0.05) in serum level of HSP70 in genotype A than genotype B in same months and summer season than the rest months and seasons.
