Date of Award


Document Type


Degree Name

Doctor of Philosophy (PhD)




Studies to investigate possible endosymbiotic relationships within the southern green stink bug, Nezara viridula (L.), a major pest of soybeans in Louisiana, were conducted. Primary emphasis was placed on the caecum, a region of the digestive tract previously associated with microbial endosymbiosis in insects. By SEM, a monoculture of rod-shaped bacteria measuring 2.5 by 0.5 (mu)m was observed adhering to the caecal crypt walls. Examination of additional specimens revealed various morphological variations including cocci, elongated rods, and rods which were apparently budding, often found within the same specimen. Diapausing specimens appeared to be no different from reproducing individuals. TEM of thin sections provided further evidence of the structure of the bacteria and their association with the caecal wall. Subsequently, two distinct bacteria of different colonial types were isolated from the caecum; both were rods. One was coccoid in appearance (designated type-A), while the other consisted of longer rods (designated type-B). Both were Gram negative and motile. They were identified as Enterobacter aerogenes (type-A) and provisionally as Aeromonas sp. (type-B). Ferritin-antibody labelling of the caecum, using antisera prepared against type-A and type-B bacteria, was used to confirm that the isolates were identical to those bacteria observed within the caecum. Production of aposymbiotic specimens, to determine the value of the symbiosis to the host, was attempted. Stink bugs were fed antibiotic treated soybeans, sampled at 6, 12, 24, 48, 72, and 96 h, and the processed caeca were examined in the SEM. The antibiotics effects could be visualized as the bacteria became elongated, 5 to 8 (mu)m long, and distinct detachment sites remained in the walls of the caecum following bacterial elimination. Their effectiveness peaked at 24 to 48 h and there was no evidence to indicate that prolonged treatment beyond 96 h would completely eliminate the flora. Therefore, the method was considered unsuitable for producing aposymbiotic specimens. Confirmation that N. viridula transmitted its symbionts by smearing its eggs at deposition was obtained by two approaches: (1) bacterial isolations from eggs were tested by slide agglutination using the previously produced antisera; (2) eggs were processed and examined in the SEM. Both species of caecal bacteria isolated (type-A and type-B) were identified from the surface of the eggs. The bacteria were also visualized on an egg in the SEM. Both rods and coccoid forms were present on the eggs and their dimensions were comparable to those previously observed within the caecum. A limited field survey of N. viridula was conducted to confirm the consistent presence of both symbionts within the caecum. Isolates from the insects' caeca were tested by slide agglutination as previously done. Both type-A and type-B bacteria were identified from some field-collected specimens, although only a single type was isolated in the majority of caeca. These results were acceptable since it is not uncommon for one bacterial species to outgrow and mask another in such isolations. However, the caecal bacteria were unexpectedly absent in the remaining specimens, possibly indicating that the symbiosis may be of marginal, if any, benefit to the stink bug. Nevertheless, the cumulative data presented indicates an endosymbiotic relationship between N. viridula and its caecal bacteria.