Endophytic Methylobacterium extorquens expresses a heterologous beta-1,4-endoglucanase A (EglA) in Catharanthus roseus seedlings, a model host plant for Xylella fastidiosa

Based on the premise of symbiotic control, we genetically modified the citrus endophytic bacterium Methylobacterium extorquens, strain AR1.6/2, and evaluated its capacity to colonize a model plant and its interaction with Xylella fastidiosa, the causative agent of Citrus Variegated Chlorosis (CVC). AR1.6/2 was genetically transformed to express heterologous GFP (Green Fluorescent Protein) and an endoglucanase A (EglA), generating the strains ARGFP and AREglA, respectively. By fluorescence microscopy, it was shown that ARGFP was able to colonize xylem vessels of the Catharanthus roseus seedlings. Using scanning electron microscopy, it was observed that AREglA and X. fastidiosa may co-inhabit the C. roseus vessels. M. extorquens was observed in the xylem with the phytopathogen X. fastidiosa, and appeared to cause a decrease in biofilm formation. AREglA stimulated the production of resistance protein, catalase, in the inoculated plants. This paper reports the successful transformation of AR1.6/2 to generate two different strains with a different gene each, and also indicates that AREglA and X. fastidiosa could interact inside the host plant, suggesting a possible strategy for the symbiotic control of CVC disease. Our results provide an enhanced understanding of the M. extorquens-X. fastidiosa interaction, suggesting the application of AR1.6/2 as an agent of symbiotic control.

CD8(+) T-cell-mediated immunity against malaria: a novel heterologous prime-boost strategy

Evaluation of: Rodriguez D, Gonzalez-Aseguinolaza G, Rodriguez JR et al. Vaccine efficacy against malaria by the combination of porcine parvovirus-like particles and vaccinia virus vectors expressing CS of Plasmodium. PLoS ONE 7(4), e34445 (2012). Recently, a vaccine against malaria was successfully tested in a human Phase III trial. The efficacy of this vaccine formulation, based on the Plasmodium falciparum circumsporozoite protein, was approximately 50% and correlated with the presence of antibodies specific to the infective stages of the malaria parasites. Different strategies are being pursued to improve vaccine efficacy levels. One such strategy is the induction of specific cytotoxic T cells that can destroy the intracellular hepatocyte stages of the malaria parasite. In this study, a novel vaccination protocol was developed to elicit strong immune responses mediated by CD8(+) cytotoxic cells specific to the circumsporozoite protein. As proof-of-concept, the authors used the rodent malaria Plasmodium yoelii parasite. The vaccination strategy consisted of a heterologous prime-boost vaccination regimen involving porcine parvovirus-like particles for priming and the modified vaccinia virus Ankara for the booster immunization, both of which expressed the immunodominant CD8 epitope of the P. yoelii circumsporozoite protein. Results from this experimental model were extremely meaningful. This vaccination strategy led to a significant T-cell immune response mediated by CD8(+) multifunctional T effector and effector-memory cells. However, most importantly for the malaria vaccine development was the fact that following a sporozoite challenge, immunized mice eliminated more than 97% of the malaria parasites during the hepatocyte stages. These results confirm and extend a vast body of knowledge showing that a heterologous prime-boost vaccination strategy can elicit strong CD8(+) T-cell-mediated protective immunity and may increase the efficacy of malaria vaccines.