Lactoferrin and Immunoglobulin G Concentration in Bovine Milk from Cows with Subclinical Mastitis during the Late Lactation Period

Abstract

Background: Lactoferrin and immunoglobulin G in milk have an important role in udder resistance to infection in the involution period. Both proteins express antimicrobial activity- lactoferrin by the binding and sequestration of iron ion; and immunoglobulin G by complement activation, bacterial opsonization and agglutination. Many factors affect lactoferrin and immunoglobulin G concentrations in bovine milk, such as the stage of lactation, milk production, and intramammary infections. The aim of this study was to determine concentrations of lactoferrin and immunoglobulin G in milk from healthy cows and subclinical mastitic cows during the late lactation period, and to evaluate the relationship between them.Materials, Methods & Results: A total of 150 quarter milk samples from 41 cows (Holstein-Friesian breed) in late lactation period were reviewed in this study. Milk samples were collected during morning milking, using aseptic techniques in sterile test tubes. From each sample, 0.1 mL of milk was plated on Columbia blood agar base with 5% defibrinated ovine blood, MacConkey agar and Sabouraud dextrose agar and incubated for 24 h - 48 h (bacteria) and 5 days (yeasts, mould) at 37oC. Milk samples for detection lactoferrin and immunoglobulin G concentration were skimmed at 1,400 g for 45 min and stored at -20°C until analysis. Lactoferrin concentration in bovine milk was determined using the Bovine Lactoferrin ELISA Quantitation Set. Milk samples were diluted at a ratio of 1:10,000. Plates were read at 450 nm absorbence values. Immunoglobulin G concentration was determined by the immunodiffusion method using radial immunodiffusion (RID) plates. Milk samples were diluted in a ratio of 1:30. Reading of results was done after incubation for 48 h by measuring the diameter of the precipitation ring. The highest mean lactoferrin concentration was observed in udder quarters infected with contagious pathogens (Streptococcus agalactiae and Staphylococcus aureus), while the highest mean immunoglobulin G concentration was detected in milk samples where minor mastitis pathogens (coagulase-negative staphylococci and Corynebacterium spp.) were isolated. Milk samples where Staphylococcus aureus was isolated had the lowest immunoglobulin G concentration, and the lowest lactoferrin concentration was observed in samples infected with enviromental pathogens (Streptococcus dysgalactiae).Discussion: This study showed that lactoferrin and immunoglobulin G concentrations are higher in milk samples from subclinical mastitic cows than in milk from normal lactating cows. Lactoferrin concentrations in milk samples from udder quarters infected with major mastitis pathogens were significantly higher than in milk infected with minor mastitis pathogens. The lowest concentration of immunoglobulin G was detected in milk samples where Staphylococcus aureus was isolated, while the highest immunoglobulin G concentration was observed in milk samples from quarters infected with minor mastitis pathogens. Lactoferrin and immunoglobulin G concentrations were significantly and positively correlated in all milk samples. This means that cows with high lactoferrin concentrations have high immunoglobulin G concentrations. In quarter milk samples infected with Staphylococcus aureus, lactoferrin and immunoglobulin G concentrations were negatively correlated. The cause of these findings could be the suppression of local immune response of mammary gland.

Milk proteins as mastitis markers in dairy ruminants - a systematic review

“…Concerning test characteristics for goats and cows, two studies carried out a competitive ELISA by using a lactoferrin antiserum from rabbit, and goat lactoferrin was isolated and purified (Chen and Mao 2004 ; Chen et al 2004 ). In other studies, cow LF was quantified by a commercial sandwich LF ELISA kit (Bethyl Laboratories, Montgomery, TX) (Cheng et al 2008 ; Sobczuk-Szul et al 2014 ; Galfi et al 2016a , 2016b ). For water buffalo, a specific ELISA kit was produced for the study (Özenç et al 2019 ).…”

“…The presence of LF in milk is due to secretion by epithelial cells and degranulation of PMNs during inflammation (Lash et al 1983). Even though LF is not an APP, it increases remarkably during the inflammatory response due to its production by mammary epithelial cells (Galfi Galfi et al 2016aGalfi et al , 2016b. For water buffalo, a specific ELISA kit was produced for the study (Özenç et al 2019).…”

Comparison of Bovine Mammary Involution and Intramammary Infections Following Intramammary Treatment with Casein Hydrolysate and Other Conventional Treatments at Dry-Off

“…Increases in the concentration of bovine lactoferrin and bovine serum albumin in milk have been used as indicators of bovine mammary involution previously [ 29 , 33 , 34 , 36 ]. Besides being an indicator of mammary involution, bovine lactoferrin plays an immunologically important role during mammary involution, by binding available iron and making it unavailable for iron-dependent bacteria to grow [ 37 , 38 ]. It has been reported that lactoferrin presents a robust defense against IMI [ 36 , 37 , 38 ].…”

“…Besides being an indicator of mammary involution, bovine lactoferrin plays an immunologically important role during mammary involution, by binding available iron and making it unavailable for iron-dependent bacteria to grow [ 37 , 38 ]. It has been reported that lactoferrin presents a robust defense against IMI [ 36 , 37 , 38 ]. One week after drying off, the concentration of lactoferrin was higher in cows that were treated with casein hydrolysate compared with the controls and all but one other treatment.…”

Lactoferrin concentration and expression in New Zealand cows milked once or twice a day

“…A positive effect of OAD is a significant increase in the content of valuable bioactive molecules, such as lactoferrin (LF), and other immune-related proteins in milk, such as acute phase proteins and RNases (Farr et al, 2002;Stelwagen et al, 2011). LF is produced in bovine milk as part of a wide range of bioactive proteins and peptides, which have roles in immunity and immunoregulation functions (Galfi et al, 2016). LF has been found to have antibacterial activity for bacteria with high iron requirements like Escherichia coli and Staphylococcus aureus, coagulase negative Staphylococcus sp.…”