Interpreting S. neurona antibody tests
This is Legend, he's not feeling well. Michelle has owned Legend for about 11 years. She says “He is basically my 1200 pound baby”. She moved to Arizona, Illinois, and back to Arizona over several years and then to Texas for 4 months. He never had any serious health problems until his EPM nightmare. And then Legend became Michelle’s full time occupation. She felt she had to move him back home despite worrying about stressing him with a trailer ride. She mentally made the decision that she’d never let him suffer.
Antibodies 101…antigens and epitopes
Antibodies are made by a host in response to an infection, and antibody responses are specific to an organism. Organisms display proteins that are available to the hosts immune system, these proteins are called antigens. Antigens are like fingerprints. An antibody is made by the body and binds individual amino acids of a protein antigen, the recognized sequence is called an “epitope”. Epitopes can be linear or conformational. Conformational epitopes are a huge issue in recognizing organisms in diagnostic tests, re-read the blog Western Blot, do’s and don’ts if you forgot why! The bottom line was that purifying antigens (immunoprecipitation) was a powerful tool to identify S. neurona in infected horses.
Sidebar: The host makes antibodies to any foreign protein that is not recognized as “self”. Antibodies that are made against “self” result in autoimmune disease. The basis of the Sidewinder test is measuring antibodies against myelin protein 2 in horses. It is relevant to EPM diagnosis because an autoimmune disease won’t be resolved with anti-parasitic agents.
Sarcocystis neurona displays antigens on its surface that directs the horses reaction to the invasion. Did you know that there are different genotypes of S. neurona? The horse can’t differentiate the organism at the DNA level. However, there are three serotypes of this parasite, this is the level the horse can distinguish. The serotypes represent the mutually exclusive, immunodominant surface antigens displayed by the three different strains of S. neurona. Our tests measure that specific reaction to the surface antigen.
S. neurona is called the ‘master of disguise’ because as the infection progresses in the horse, neurona changes the antigens it displays on its surface! S. neurona displays different antigens at different stages of development. Stage related antigen expression has important implications in detecting disease-causing strains. That deception, antigen switching, is an immune invasion tactic often employed by protozoa.
Sidebar: Also, organisms differ in their virulence. Virulence is related to genotype, and perhaps serotype. Another factor of infection versus disease is dose, how many organisms the host contacts. If you read much of the EPM animal model literature, you are aware that dose and route of infection are important in producing disease. The pandemic certainly made us all aware of infection (production of antibodies and an immune response) and disease (a morbid result of infection). Both situations produce antibodies. Only disease should be treated.
Our animal infection model studies taught us that the titer increases with duration of infection, not the degree of clinical signs displayed when the animal is diseased. The clinical signs are related to cytokine responses, remember cytokines are part of the innate immune response and occur earlier than antibody production. Innate responses are not specific but highly important in inflammation.
Antibodies were shown in an experiment to protect against challenge in the Trojan Horse model. We did show in a serum neutralization assay that a rSAG1 (recombinant) vaccine, and resulting immunity, did not neutralize SAG 5 organisms. However, following a natural infection with SAG 1, neutralizing antibodies were produced against a SAG 5 strain. Here’s why. The horse makes antibodies against the whole organism, proteins that are shared between serotypes. Antibodies to non-specific, shared antigens can be partially protective.
We developed three ELISA tests to monitor S. neurona infections in your horse. We use recombinant proteins representing SAG 1, SAG 5, and SAG 6 antigens. The tests measure antibodies against these immunodominant antigens of S. neurona in any body fluid. The three individual tests results distinguish the serotype of the infecting organism, or organisms, infecting a horse from one sample of serum. A titer is a number that indicates the level of antibody response to an infection at one point in time. The SAG ELISA tests give you three titers that give the antibody profile of infection with S. neurona.