Quantitative PCR study on the mode of action of oligosaccharide elicitors on penicillin G production by Penicillium chrysogenum

Abstract

Aim: To investigate the effects of single and multiple additions of the oligosaccharide elicitors, obtained from alginate and locust bean gum, on penicillin G production and the transcript level of penicillin G biosynthetic genes. Methods and Results: The transcript copy numbers and penicillin G concentration in liquid cultures of Penicillium chrysogenum grown under control and elicited conditions were compared using quantitative PCR and HPLC assay respectively. An increase in the penicillin G production rate and transcript copy numbers of the three major penicillin G biosynthetic genes pcbAB, pcbC and penDE was observed in the elicited cultures compared to control cultures. The effects were observed to be higher in multiple elicitor added cultures compared to single elicitor supplemented and control cultures. Conclusions: The results show, for the first time in bioreactor cultures, the enhancement of penicillin G transcript copy number of the penicillin biosynthetic genes using qPCR with a corresponding increase in the penicillin G production upon multiple elicitor addition of two different types of elicitors. Significance and Impact of the Study: Establishment of the effect of multiple elicitor addition on penicillin G production and investigating the role of oligosaccharide elicitors as transcriptional activators has wide spread impact for antibiotic industry.

Enhanced intracellular Ca2+ concentrations in Escherichia coli and Bacillus subtilis after addition of oligosaccharide elicitors

“…signatures were observed upon addition of OG or MO to E. coli pHCM05-Aq and B. subtilis pHCM05-Aq, suggesting that different calcium messages might be transmitted probably due to the difference in the three-dimensional structure between MO and OG. This may explain why higher antibiotic levels are achieved when P. chrysogenum and Bacillus licheniformis are stimulated with multiple elicitor additions using different elicitors (OG and MO) in comparison to repeated addition of the same elicitor to the culture (Murphy et al 2007b;Nair et al 2009). …”

“…Enhancement of penicillin G in elicited cultures may be related to the morphological changes caused by the elicitation (Radman et al 2004). Elicitors also effect the transcription of genes for antibiotic biosynthesis (Murphy et al 2007b;Nair et al 2009). However, a possible mechanism by which elicitors cause these changes has yet to be defined.…”

Application of fluorescent indicators to analyse intracellular calcium and morphology in filamentous fungi

“…The reasons for these morphological changes are still speculative and require further investigation (Radman et al 2004). Recent studies have reported the enhancement of transcript copy number of the penicillin biosynthetic genes with a corresponding increase in the penicillin G production upon oligosaccharide elicitor (Nair et al 2009). There are initial reports of decrease in extracellular Ca 2þ levels on addition of mannan oligosaccharides (MO) as elicitors (Maria Teresa Saez Iturbe 2005).…”

“…A novel approach for enhancement of microbial metabolites is the use of elicitors. Elicitors are substances known to stimulate morphological and physiological responses when added in trace quantities to cultures (Radman et al 2003;Murphy et al 2007;Nair et al 2009). Previous reports of elicitation in Penicillium chrysogenum have reported an increase in hyphal-tip numbers, clump area and spore counts along with simultaneous increase in penicillin G production in elicitor-supplemented cultures.…”

Elicitation of secondary metabolism in actinomycetes

“…These concentration changes serve as internal signals to direct the regulation of secondary metabolites via yet unknown mechanisms. An elicitor can either directly influence the transcription of the secondary metabolite gene cluster or induce a transcriptional activator of the target gene cluster (Kawai, Wang, 2007, Nair et al, 2009, Rigali et al, 2008, Tanaka et al, 2010.…”