1a.Objectives (from AD-416):
Study colonization of seed coats by 'Ca. Liberibacter asiaticus'and devise methods of extracting the bacteria for research.
1b.Approach (from AD-416):
We have identified seed coats of several citrus varieties as having high levels of 'Ca. Liberibacter asiaticus' DNA and in immature seeds and use microscopy to demonstrate viability of bacteria. We will determine if there are optimal stages of development of the seed for extraction of bacteria by sampling populations of seeds collected from fruit at various stages of development. We will test various mechanical methods to purify intact cells from the seed coats.
This project is related to inhouse project objective 1a: Characterize the etiology, molecular biology and genetics of ‘Candidatus Liberibacter asiaticus (Las),’ the bacterium associated with citrus huanglongbing (HLB).
The bacterium ‘Ca. Liberibacter asiaticus’ (Las) causes a damaging disease of citrus called greening. The bacterium cannot be grown on an artificial culture medium, only in citrus plants. We want to purify the bacterium from plant tissue so we can use these purified bacteria for laboratory experiments, such as experiments to culture the bacterium and for greenhouse experiments with citrus plants. We devised and tested experimental methods to extract the bacteria from the plant tissue and separate the bacterial cells from plant tissue. Our current protocols can extract a small quantity of bacteria from the tissue which we can see using a microscope. We are modifying our experimental methods to increase the efficiency of our extraction so we can recover more bacterial cells from different plant tissues.