Analysis of bacterial community during the fermentation of pulque, a traditional Mexican alcoholic beverage, using a polyphasic approach

• Published in: International journal of food microbiology 2008-05, Vol.124 (2), p.126-134
• Main Authors: Escalante, Adelfo, Giles-Gómez, Martha, Hernández, Georgina, Córdova-Aguilar, María Soledad, López-Munguía, Agustín, Gosset, Guillermo, Bolívar, Francisco
• Format: Article
• Language: English
• Subjects: Acetobacter; Acinetobacter radioresistens; Agave; Agave - microbiology; Alcoholic Beverages - microbiology; Bacteria - classification; Bacteria - isolation & purification; Biological and medical sciences; Cloning, Molecular; DNA, Bacterial - analysis; DNA, Ribosomal - analysis; Ecosystem; Enterobacter; Erwinia rhapontici; Fermentation; Firmicutes; Food industries; Food microbiology; Fundamental and applied biological sciences. Psychology; Lactic acid bacteria; Lactobacillus acidophilus; Lactococcus lactis; Leuconostoc citreum; Mexico; Microbial diversity; Phylogeny; Pulque; Restriction Mapping; RNA, Ribosomal, 16S - genetics; Species Specificity; Traditional alcoholic fermentation; Zymomonas mobilis
• ISSN: 0168-1605; 1879-3460
• DOI: 10.1016/j.ijfoodmicro.2008.03.003

In this study, the characterization of the bacterial community present during the fermentation of pulque, a traditional Mexican alcoholic beverage from maguey (Agave), was determined for the first time by a polyphasic approach in which both culture and non-culture dependent methods were utilized. The work included the isolation of lactic acid bacteria (LAB), aerobic mesophiles, and 16S rDNA clone libraries from total DNA extracted from the maguey sap (aguamiel) used as substrate, after inoculation with a sample of previously produced pulque and followed by 6-h fermentation. Microbiological diversity results were correlated with fermentation process parameters such as sucrose, glucose, fructose and fermentation product concentrations. In addition, medium rheological behavior analysis and scanning electron microscopy in aguamiel and during pulque fermentation were also performed. Our results showed that both culture and non-culture dependent approaches allowed the detection of several new and previously reported species within the α-, γ-Proteobacteria and Firmicutes. Bacteria diversity in aguamiel was composed by the heterofermentative Leuconostoc citreum, L. mesenteroides, L. kimchi, the γ-Proteobacteria Erwinia rhapontici, Enterobacter spp. and Acinetobacter radioresistens. Inoculation with previously fermented pulque incorporated to the system microbiota, homofermentative lactobacilli related to Lactobacillus acidophilus, several α-Proteobacteria such as Zymomonas mobilis and Acetobacter malorum, other γ-Proteobacteria and an important amount of yeasts, creating a starting metabolic diversity composed by homofermentative and heterofermentative LAB, acetic and ethanol producing microorganisms. At the end of the fermentation process, the bacterial diversity was mainly composed by the homofermentative Lactobacillus acidophilus, the heterofermentative L. mesenteroides, Lactococcus lactis subsp. lactis and the α-Proteobacteria A. malorum. After a 6-h fermentation, 83.27% of total sugars detected after inoculation were consumed (228.4 mM hexose equivalents) and a carbon (C) recovery of 66.18% in fermentation products was estimated. They were produced 284.4 mM C as ethanol, 71.5 mM C as acetic acid and 19 mM C as lactic acid, demonstrating the presence of homo- and heterofermentative, acetic and alcoholic metabolisms in the final product. It was also found, after hydrolysis, that the exopolysaccharide produced during the fermentation was mainly composed by fruct