Longitudinal analysis of Prototheca zopfii-specific immune responses: correlation with disease progression and carriage in dairy cows

Abstract

In order to characterize the humoral and cellular immune responses to bovine mammary protothecosis, serum and whey samples obtained from 72 dairy cows assigned to four different clinical stages of infection were examined for specific antibodies by indirect enzyme-linked immunosorbent assay techniques. Milk samples were analyzed for the total numbers of excreted algal cells and somatic cells. After characterization of the course of immune induction in bovine protothecal mastitis, a long-term sentinel study was performed in an affected herd in order to investigate disease progression. A total of 61 dairy cows with protothecal mastitis were examined for shedding of algae cells and for local immune responses three times in 6-month intervals. During acute and chronic stages of protothecosis, significantly elevated specific antibody activities in sera were detected. A strong correlation of whey immunoglobulin A (IgA) and whey IgG1 antibody activity with the total counts of somatic cells in milk was observed, whereas only a weak correlation of whey IgA and whey IgG1 concentrations to the number of algal cells excreted with the milk was seen. Our results from the sentinel long-term study of infected cows revealed that 70.5% of the persistently infected animals were continuously shedding the pathogen. About 4.9% of the animals showed an intermittent shedding, whereas 18% of the cows were tested culturally negative throughout the study. It can be assumed that Prototheca zopfii mastitis in dairy cows is maintained on the herd level by subclinically infected alga-shedding cows.

FIG. 1.

Numbers of algae (A) and somatic cells (B) excreted with milk dependent upon the clinical stages of bovine P. zopfii mastitis. Group 1 (MA, P⁺) included cows with clinical signs of acute mastitis and positive cultural P. zopfii findings. Group 2 (MC, P⁺) included cows with subacute to chronic clinical symptoms of mastitis and positive cultural P. zopfii findings. Group 3 (MC, P⁻) included cows with chronic clinical signs and an earlier positive cultural P. zopfii finding but culture negative for P. zopfii at the time of the present study. Group 4 (M⁻, P⁻) included cows without clinical signs of mastitis and culture negative for P. zopfii. A significant discrimination (P < 0.05) of the MA, P⁺ group versus the MC, P⁺ group is indicated by the symbol “#.” The dagger symbol (†) indicates significant differences (P < 0.05) between the MC, P⁺ group and MC, P⁻ group. A significant discrimination (P < 0.05) of chronically infected cultural positive versus noninfected cows is indicated by the “§” symbol. Asterisks indicate that P. zopfii could not be detected by plate culturing.

FIG. 2.

Correlation between specific antibody responses against P. zopfii in serum and whey samples of infected dairy cows on the number of CFU (left) and of the somatic cell counts in milk (right). Regression lines are depicted. (A and D) Serum IgG; (B and E) whey IgA; (C and F) whey IgG1.