This review describes the exploitation of exclusively optical surface plasmon resonance (SPR) biosensors for the direct and indirect detection of pathogenic microorganisms in food chains and the environment. Direct detection is, in most cases, facilitated by the use of defined monoclonal or polyclonal antibodies raised against (a part of) the target pathogenic microorganisms. The antibodies were immobilized to a solid phase of the sensor to capture the microbe from the sample. Alternatively, antibodies were used in an inhibition-like assay involving incubation with the target organism prior to analysis of nonbound antibodies. The free immunoglobins were screened on a sensor surface coated with either purified antigens or with Fc or Fab binding antibodies. Discussed examples of these approaches are the determination of Escherichia coli O1 57:H7, Salmonella spp., and Listeria monocytogenes. Another direct detection strategy involved SPR analysis of polymerase chain reaction products of Shiga toxin-2 genes reporting the presence of E. coli O157:H7 in human stool. Metabolic products have been exploited as biomarkers for the presence of a microbial agent, such as enterotoxin B and a virulence factor for the occurrence of Staphylococcus aureus and Streptococcus suis, respectively. Indirect detection, on the other hand, is performed by analysis of a humoral immune response of the infected animal or human. By immobilization of specific antigenic structures, infections with Herpes simplex and human immunodeficiency viruses, Salmonella and Treponema pallidum bacteria, and Schistosoma spp. parasites were revealed using human, avian, and porcine sera and avian eggs. Bound antibodies were easily isotyped using an SPR biosensor to reveal the infection history of the individual. Discussed studies show the recent recognition of the suitability of this type of instrument for (rapid) detection of health-threatening microbes to food and environmental microbial safety.