Promoting L-theanine accumulation in Camellia sinensis through irrigation with Pseudomonas knackmussii

We isolated endophytic bacteria Pseudomonas knackmussii from Camellia sinensis and identified its L-theanine synthesis-related gene, CDF86904.1. Irrigation with endophytic bacteria significantly increased the L-theanine content of tea plants. The presence of L-theanine in tea enhances its sensory attributes and mitigates the bitterness caused by polyphenols and caffeine. The low L-theanine content in summer and autumn teas, which is < 1% of their dry weight, is a key factor in their inferior taste. Our study successfully enhanced L-theanine levels in tea plants by irrigating them with endophytic bacteria. The Pseudomonas knackmussii isolated from tea plants is notable for its outstanding ability to synthesize L-theanine, using ethylamine to produce 201.4 μM of L-theanine. Our study identified the key gene responsible for the biosynthesis of L-theanine in P. knackmussii to be CDF86904.1 (L-glutamine synthetase, GS). We found that P. knackmussii can penetrate nearly all tea plant tissues and has excellent colonization ability. Additionally, irrigation of Baiye No.1 tea seedlings with both ethylamine and endophytes for 7 days resulted in top leaves exhibiting an L-theanine content of 58.5 mg/g dry weight. This represents a notable increase compared to 37.8 mg/g dry weight with only ethylamine and 15.4 mg/g dry weight with solely water. Surprisingly, after irrigating the tea plants with a combination of endophytic bacteria and ethylamine, the L-theanine content of the third tea leaf was 131.7 mg/g dry weight after 14 days. This study reveals the significant potential of endophytic bacteria in tea plants to increase L-theanine content, providing empirical evidence for improving tea quality and demonstrating important theoretical and practical significance in tea production.