Desenvolvimento de bioprocessos para produção de inoculante de cana-de-açúcar através do co-cultivo das espécies Herbaspirillum seropedicae BR11335 e H. rubrisubalbicans BR11504

Abstract

The objective of this work was the establishment of a growth system fitted for the co-cultivation of endophytic bacteria Herbaspirillum seropedicae (strain BR11335) and Herbaspirillum rubrisubalbicans (BR11504), which make up the recommended inoculant for a sugarcane. Individual growth curves were constructed in an orbital shaker (floor incubator) and bench bioreactor, evaluating two temperature levels, 30oC and 35oC°, and air flow from 1.0 to 6.0 L min-1. It was obtained the process conditions that favored both strains, and they were reproduced in co-cultivation systems either in shaker or in bench bioreactor. To follow the behavior of the strains in co-cultivation, samples of fermentative broth were taken at regular intervals of 4 hours and performed real-time PCR (qPCR) to identify a concentration of each bacterium in the system. For this, pairs of species-specific primers were designed for each of the strains. The conditions that favored both strains were: 35° C temperature and 5.0 to 6.0 L min-1 of aeration for. Evaluating temporal samples of the co-cultivations under the previous conditions by means of qPCR, the presence of the two strains in the co-cultures was observed, showing that it was possible to apply this system to the bacteria. There was significant difference (α = 0.05) between the average growthof strains BR11335 and BR11504, with BR11504 growth being higher in both aeration conditions. However, the temporal results of the qPCR did not follow the growth curves from which the aliquots were removed, evidencing that the DNA extraction method probably influenced the results and needs adjustments, mainly due to the occurrence of a high concentration of viscous material during extraction , Which may be evidencing large production of polysaccharides by the strains during the simultaneous cultivation of them, thus altering the results of the qPCR.