Pasquale Filannino, Raffaella Di Cagno, Rocco Addante, Erica Pontonio, Marco Gobbetti

Fructophilic lactic acid bacteria (FLAB) are strongly associated to the gastrointestinal tract (GIT) of Apis mellifera L. worker bees due to the consumption of fructose as a major carbohydrate. Seventy-seven presumptive lactic acid bacteria (LAB) were isolated from GIT of healthy A. mellifera L. adults, which were collected from 5 different geographical locations of Apulia region (Italy). Almost all the isolates showed fructophilic tendencies, which were identified as Lactobacillus kunkeei (69%) or Fructobacillus fructosus (31%). A high-throughput phenotypic microarray, targeting 190 carbon sources, was used to determine that 83 compounds were differentially consumed. Phenotyping grouped the strains into two clusters, reflecting growth performance. The utilization of phenolic acids, such as p-coumaric, caffeic, syringic or gallic acids, as electron acceptors was investigated in fructose based medium. Almost all FLAB strains showed tolerance to high phenolic acid concentrations. p-Coumaric acid and caffeic acid were consumed by all FLAB strains through reductases or decarboxylases. Syringic and gallic acids were partially metabolized. The data collected suggest that FLAB require external electron acceptors to regenerate NADH. The use of phenolic acids as external electron acceptors by 4 FLAB, showing the highest phenolic acid reductase activity, was investigated in glucose based medium supplemented with p-coumaric acid. Metabolic responses observed through phenotypic microarray suggested that FLAB may use p-coumaric acid as external electron acceptor, enhancing glucose dissimilation but less efficiently than other external acceptors such as fructose or pyruvic acid. Fructophilic lactic acid bacteria (FLAB) remain to be fully explored. This study intends to link unique biochemical features of FLAB with their habitat. The quite unique FLAB phenome within the lactic acid bacteria (LAB) group may have practical relevance in food fermentations. The FLAB phenome may have implications in the levels of hexose metabolism products in fermented foods, as well as in food probiotication. Due to the harsh conditions of honeybees' GIT, these bacteria had to develop specific physiological and biochemical characteristics, such as tolerance to phenolic acids. The screening of FLAB strains based on metabolic pathways involving phenolic acids may allow the selection of starter cultures with both technological and functional beneficial attributes. Bioconversion of phenolic compounds may contribute to the aroma attributes and biofunctionality of fermented foods. Thus, the selection of FLAB strains as starter cultures with specific enzymatic activities involving phenolic acids, may have a promising role in food fermentations.