Elisa versus immunolocalization to determine the association of Erwinia tracheiphila in Acalymma vittatum (Coleoptera: Chrysomelidae)

DAS-ELISA, immunohistochemistry and electron microscopy were used to investigate the association of the causal agent of bacterial wilt, Erwinia tracheiphila (Smith) within the period, a high percentage of individuals beetle Acalymma vittatum (F.). After a 24-h acquisition tested positive for E. tracheiphila antigen using both immunohistochemistry (100%) and DAS-ELISA (70-60%). Both assays showed that the antigen remained in beetles long after the initial acquisition, with the percentage declining during incubation. Using ELISA, the percentage decreased to 4.7% within 3 d after acquistion then increased to 10% within 10 d and remained at 10% for 30 d. Immunoperoxidase assays of paraffin embedded gut sections were more sensitive, and showed that 95% of the beetles harbored the pathogen after 10 d and 20% after 30 d. E. tracheiphila antigen was present throughout the digestive tract soon after acquisition but only small clusters of E. tracheiphila were observed along the alimentary canal 3 d after transfer onto clean plants. After 10 and 30 d on clean plants, E. tracheiphila antigen reaction was stronger and clusters of bacteria were more numerous primarily in the posterior midgut and anterior portion of the hindgut. Scanning electron microscopy and TEM photomicrographs confirmed the presence of bacterial cells resembling E. tracheiphila associated with the intima of the hindgut 1 and 30 d after acquisition. This demonstrated the sensitivity of immunohistochemisty for detecting E. tracheiphila within its vector, and suggests a long-term extracellular endosymbiotic association of E. tracbeiphila with the alimentary anal of A. vittatum.