Stanko F. Boboš

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

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.

Prevalence and Molecular Characterization of Enterotoxin-Producing Strains of Staphylococcus Aureus Isolated from Serbian Dairy Cows

Abstract Staphylococcus aureus is known worldwide as a frequent cause of mastitis in dairy cattle. Due to the production of heath resistant enterotoxins, this pathogen is also a major cause of food poisoning among humans, with symptoms of often severe vomiting and diarrhea. The aim of our study was to determine the prevalence of enterotoxinproducing strains of S. aureus originating from samples of cows with subclinical and clinical mastitis in the Republic of Serbia. Furthermore, we analyzed the type of staphylococcal enterotoxin they produce and phylogenetic relatedness among the S. aureus isolates recovered from milk in this study. Production of staphylococcal enterotoxins A, B, C, D and E was determined by commercial immunoenzyme assay VIDAS® SET2, and presence of corresponding genes encoding enterotoxin synthesis in positive isolates confi rmed by Polymerase Chain Reaction. Enterotoxin production was determined in 5 out of 75 (6.67%) isolates of S. aureus and all of them produced staphylococcal enterotoxins C. After analyzing the nucleotide sequence of the gene encoding the synthesis of staphylococcal protein A, S. aureus isolates were assigned into 2 phylogenetic groups, including 7 clusters. All S. aureus isolates with the presence of sec gene formed one cluster even dough they originated from milk samples from different farms.

Available control measures for Q fever in sheep.

Q fever is a worldwide zoonosis caused by Gram-negative bacteria, Coxiella burnetii. This antropozoonosis is characterized by a wide spectrum of hosts and vectors. Primary role of domestic animals as reservoirs of human infections emphasizes the accurate prompt detection of Q fever in domestic animals. This microbe can survive for months and even years in dust or soil. Sanitation of endemic foci of Q fever is practically pretty close to impossible because of the high resistance of the causative agent, small infectious dose and various epidemiologies. Within the group of zooantroponoses, Q fever takes a leading position in the region of Vojvodina. This is why Q-fever is considered a specific problem of this province. Control programs against Q fever in sheep and cattle, most frequently recommend serological examination and vaccination of animals. In animals, the most effective vaccines are those composed of inactivated whole phase I bacteria. Bacterial shedding in placental tissue and milk was reduced in experimental infection or in natural C.burnetii infection of sheep and cows vaccinated by phase I vaccines. One of the recommended measures is excluding positive reactors from the flock followed by continuous monitoring and separation of seropositive animals. Milk from seropositive cows must be pasteurized. The aim of the vaccination after lambing season, is to prevent new infections until next lambing of young animals, which were not pregnant. Well-timed sequential vaccination before pregnancy reduces the risk of C. burnetii infection, highly prevents the abortions decreases the shedding rate of C. burnetii after the abortion. [Projekat Ministarstva nauke Republike Srbije, br. TR31084]

Leptospira interrogans serotype hardjo in dairy cows

Data on L. hardjo infection of dairy cows in the world pint out its important role in the occurrence of health and economic problem. L. interrogans serotype hardjo has been described as the cause of miscarriages, stillbirts, or the birhs of poorly vital calves, agalactia, mastitis, and low fertility in cows. Two L. hardjo genotypes have been identified in cows, namely, hardjopraitno and hardjobovis. Serological investigations have established a drastic increase in this leptospiral infection in cows. L. hardjo has become adapted to cattle as the primary host, so that an infection is maintained in herds and becomes deeply rooted because of the permanent presence of the source of infection. It was believed that sheep were accidental hosts, but the latest research suggest that they are yet another, transitory, host for maintining this leptospira serotype. L. hardjo is also important from the aspect of human health, especially of persons who are professionally exposed to this infection. L. hardjo infection is detected using serological tests and by proving the presence of leptospira. The medicine of choice in the therapy of leptospiral infections is streptomycin (DSM). Therapy using oxytetracyclines for clinical mastitis was also proven effective. Treatment is most successful in the early stage of the disease. A single dose of streptomycin administered in infected herds reduces the duration period of leptospira excretion through urine, thus preventing the spread of infection thorugh contaminated urine. The basic components of the plan to contain leptospira are the following: serological investigations, sanitary-higiene measures, the elimination of animals which excrete leptospira through urine, therapy, vaccination, quarantine.