Cinnamic acids (i.e., ferulic and caffeic acids) that are esterified to the vegetable cell walls should be enzymatically released to be absorbed in a mammal's intestines. A low dosage of ferulic acid in rodent diets stimulates insulin production and alleviates symptoms caused by diabetes (M. Sri Balasubashini, R. Rukkumani, and V. P. Menon, Acta Diabetol. 40:118-122, 2003). Several lactic acid bacteria are able to display ferulic acid esterase (FAE) activity, suggesting that their probiotic activity could be, in part, mediated by the slow release of ferulic acid. In the present work, we describe the isolation of one strain identified as being Lactobacillus johnsonii that displayed strong FAE activity in stool samples from diabetes-resistant biobreeding rats. These animals are genetically susceptible to becoming diabetic but do not develop the disease. By using genomic analysis coupled to protein purification and catalytic screening, we were able to purify two proteins with FAE activity. The enzymes displayed 42% sequence identity and a broad range of substrate preferences. High affinities and catalytic efficiencies toward aromatic compounds such as ethyl ferulate (K m ‫؍‬ 20 to 60 M) and chlorogenic acid (K m ‫؍‬ 10 to 50 M) were observed. The strain isolated herein as well as the enzymes studied could be potentially useful for the formulation of probiotics to ameliorate diabetes symptoms.According to the Centers for Disease Control and Prevention (CDC), approximately 23.6 million Americans suffer from diabetes. In the year 2000, this metabolic disease was directly or indirectly responsible for 200,000 deaths. On average, diabetes costs more than 170 billion dollars annually to the American economy. Diets abundant in carbohydrates, increasing rates of obesity, and sedentary lifestyles in combination with genetic predisposition are key factors associated with the development of diabetes. In addition to these environmental factors, notable changes in the composition of the gut microbial ecosystem of diabetic patients were recently described (36).A clever analysis utilized to measure microbially diverse populations based on genomic principles reported previously by Furrie (17) was used by Roesch and coworkers (30) to prove that rodents with diabetes have a specifically associated gut microflora. A deep characterization of the intestinal microflora of biobreeding diabetes-resistant (BB-DR) rats showed that the presence of Lactobacillus reuteri, Lactobacillus johnsonii, and Bifidobacterium species is predominant, compared with the bacterial species observed in biobreeding diabetes-prone (BB-DP) rats (30). In agreement with the large presence of probiotic-like bacteria, it was previously reported that the oral administration of lactic acid bacteria (LAB) can help to reduce blood glucose levels (25-27, 40) by stimulating insulin secretion via changes in autonomic neurotransmission (34). However, it is not clear how LAB species are able to induce these changes. Neither genetic nor physiological characteristics of the st...

Here Tn5 random transposon mutagenesis was used to identify the essential elements for culturing Liberibacter crescens BT-1 that can serve as antimicrobial targets for the closely related pathogens of citrus, Candidatus Liberibacter asiaticus (Las) and tomato and potato, Candidatus Liberibacter solanacearum (Lso). In order to gain insight on the virulence, metabolism, and culturability of the pathogens within the genus Liberibacter, a mini-Tn5 transposon derivative system consisting of a gene specifying resistance to kanamycin, flanked by a 19-base-pair terminal repeat sequence of Tn5, was used for the genome-wide mutagenesis of L. crescens BT-1 and created an insertion mutant library. By analyzing the location of insertions using Sanger and Illumina Mi-Seq sequencing, 314 genes are proposed as essential for the culture of L. crescens BT-1 on BM-7 medium. Of those genes, 76 are not present in the uncultured Liberibacter pathogens and, as a result, suggest molecules necessary for the culturing these pathogens. Those molecules include the aromatic amino acids, several vitamins, histidine, cysteine, lipopolysaccharides, and fatty acids. In addition, the 238 essential genes of L. crescens in common with L. asiaticus are potential targets for the development of therapeutics against the disease.