Isolation, characterization and optimization of indigenous acetic acid bacteria and evaluation of their preservation methods
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Abstract
Background and Objectives: Acetic acid bacteria (AAB) are useful in industrial production of vinegar. The present study aims at isolation and identification of acetic acid bacteria with characterization, optimization, and evaluation of their acetic acid productivity.
Materials and Methods: Samples from various fruits were screened for presence of acetic acid bacteria on glucose, yeast extract, calcium carbonate (GYC) medium. Carr medium supplemented with bromocresol green was used for distinguishing Acetobacter from Gluconobacter. The isolates were cultured in basal medium to find the highest acetic acid producer. Biochemical tests followed by 16S rRNA and restriction analyses were employed for identification of the isolate and phylogenic tree was constructed. Bacterial growth and acid production conditions were optimized based on optimal inoculum size, pH, temperature, agitation, aeration and medium composition.
Results: Thirty-seven acetic acid bacteria from acetobacter and gluconobacter members were isolated. Acetic acid productivity yielded 4 isolates that produced higher amounts of acid. The highest producer of acid (10.03%) was selected for identification. The sequencing and restriction analyses of 16S rRNA revealed a divergent strain of Acetobacter pasteurianus (Gene bank accession number # GU059865). The optimum condition for acid production was a medium composed of 2% glucose, 2% yeast extract, 3% ethanol and 3% acid acetic at inoculum size of 4% at 3L/Min aeration level in the production medium. The isolate was best preserved in GYC medium at 12oC for more than a month. Longer preservation was possible at -70oC.
Conclusion: The results are suggestive of isolation of an indigenous acetic acid bacteria. Pilot plan is suggested to study applicability of the isolated strain in acetic acid production.
Matsushita K, Toyama H, Adachi O: Respiratory Chains and Bioenergetics of Acetic Acid Bacteria; in Rose and AH, (ed): Advances in Microbial Physiology. Academic Press, 1994, pp 247-301.
Adachi O, Moonmangmee D, Toyama H, Yamada M, Shinagawa E, Matsushita K: New developments in oxi-dative fermentation. Appl Microbiol Biotechnol 2003;60: 643-653.
De Ley J, Gillis M, Swings J: Bergey>s Manual of Sys- tematic Bacteriology; Baltimore, Williams and Wilkins Baltimore, 1984, pp 267-278.
Yamada Y, Hosono R, Lisdyanti P, Widyastuti Y, Sa- ono S, Uchimura T, Komagata K: Identification of ace- tic acid bacteria isolated from Indonesian sources, espe- cially of isolates classified in the genus Gluconobacter. J Gen Appl Microbiol 1999; 45: 23-28.
Ndoye B, Lebecque S, Dubois-Dauphin R, Tounkara L, Guiro AT, Kere C, Diawara B, Thonart P: Thermore- sistant properties of acetic acids bacteria isolated from tropical products of Sub-Saharan Africa and destined to industrial vinegar. Enzyme Microb Technol 2006; 39:916-923.
Pulvirenti A, Solieri L, Gullo M, De Vero L, Giudici P: Occurrence and dominance of yeast species in sour- dough. Lett Appl Microbiol 2004; 38: 113-117.
Sokollek SJ, Hertel C, Hammes WP: Description of Ac- etobacter oboediens sp. nov. and Acetobacter pomorum sp. nov., two new species isolated from industrial vinegar fermentations. Int J Syst Bacteriol 1998; 48: 935-940.
Lisdiyanti P, Kawasaki H, Seki T, Yamada Y, Uchimu- ra T, Komagata K: Identification of Acetobacter strains isolated from Indonesian sources, and proposals of Ac- etobacter syzygii sp. nov., Acetobacter cibinongensis sp. nov., and Acetobacter orientalis sp. nov. J Gen Appl Microbiol 2001; 47: 119-131.
Franke IH, Fegan M, Hayward C, Leonard G, Stack- ebrandt E, Sly LI: Description of Gluconacetobacter sacchari sp. nov., a new species of acetic acid bacterium isolated from the leaf sheath of sugar cane and from the pink sugar-cane mealy bug. Int J Syst Bacteriol 1999;49: 1681-1693.
Sambrook J, Fritsch EF, Maniatis T. Molecular Clon- ing, A Laboratory Manual (M). 2 nd edition, New York: Cold Spring Harbor Laboratory Press 1989; p: 1.21-1.52, 2. 60-2. 80, 7. 3-7. 35, 9. 14-9.22.
Gullo M, Romano AD, Pulvirenti A, Giudici P: Can- dida humilis - Dominant species in sourdoughs for the production of durum wheat bran flour bread. Int J Food Microbiol 2003; 80: 55-59.
Du Toit WJ, Pretorius IS: The occurrence, control and esoteric effect of acetic acid bacteria in winemaking. Ann Microbiol 2002; 52: 155-179.
Frateur J: Essai sur la systematique des acetobacters.Cellule 1950; 53: 392.
Carr JG: Methods for identifying acetic acid bacteria; in Gibbs BM, Shapton DA, (eds): Identification Methods for Microbiologists. London, Academic Press, 1968, pp1-8.
Swings J: The genera Acetobacter and Gluconobacter; in Balows A, Dworkin, G, M. H, Schleifer KH, (eds): The Prokaryotes. A Handbook on the Biology of Bac- teria: Ecophysiology, Isolation, Identi¢cation, Appli- cations. New York., Springer-Verlag, 1992, vol III, pp2268-2286.
Lisdiyanti P, Kawasaki H, Seki T, Yamada Y, Uchimu-ra T, Komagata K: Systematic study of the genus Ac- etobacter with descriptions of Acetobacter indonesien- sis sp. nov., Acetobacter tropicalis sp. nov., Acetobacter orleanensis (Henneberg 1906) comb. nov., Acetobacter lovaniensis (Frateur 1950) comb. nov., and Acetobacter estunensis (Carr 1958) comb. nov. J Gen Appl Micro- biol 2000; 46: 147-165.
Gullo M ,Caggia C, De Vero L, Giudici P: Characteriza- tion of acetic acid bacteria in «traditional balsamic vin- egar». Int J Food Microbiol 2006; 106: 209-212.
Cleenwerck I, Vandemeulebroecke K, Janssens D, Swings J: Re-examination of the genus Acetobacter, with descriptions of Acetobacter cerevisiae sp. nov., and Acetobacter malorum sp. nov. Int J Syst Evol Microbiol 2002; 52: 1551-1558.
Dellaglio F, Cleenwerck I, Felis GE, Engelbeen K, Jans- sens D, Marzotto M: Description of Gluconacetobacter swingsii sp. nov. and Gluconacetobacter rhaeticus sp. nov., isolated from Italian apple fruit. Int J Syst Evol Mi- crobiol 2005; 55: 2365-2370.
Gonzalez A, Hierro N, Poblet M, Rozes N, Mas A, Guiro AT: Application of molecular methods for the differen- tiation of acetic acid bacteria in a red wine fermentation. J Appl Microbiol 2004; 96: 853-860.
Gonzalez A, Guillamon JM, as A, oblet M: Application of molecular methods for routine identification of acetic acid bacteria. Int J Food Microbiol 2006; 108:141-146.
Poblet M, Rozes N, Guillamon JM, Mas A: Identifica- tion of acetic acid bacteria by restriction fragment length polymorphism analysis of a PCR-amplified fragment of the gene coding for 16S rRNA. Lett Appl Microbiol 2000; 31: 63-67.
Ruiz A, Poblet M, Mas A, Guillamon JM: Identification of acetic acid bacteria by RFLP of PCR-amplified 16S rDNA and 16S-23S rDNA intergenic spacer. Int J Syst Evol Microbiol 2000; 50: 1981-1987.
Trcek J, Teuber M: Genetic and restriction analysis of the 16S-23S rDNA internal transcribed spacer regions of the acetic acid bacteria. FEMS Microbiol Lett 2002;208: 69-75.
Trcek J: Quick identification of acetic acid bacteria based on nucleotide sequences of the 16S-23S rDNA internal transcribed spacer region and of the PQQ-de- pendent alcohol dehydrogenase gene. SYST APPL MI- CROBIOL 2005; 28: 735-745.
Yukphan P, Malimas T, Takahashi M, Potacharoen W, Busabun T, Tanasupawat S, Nakagawa Y, Tanticharoen M, Yamada Y: Re-identification of Gluconobacter strains based on restriction analysis of 16S - 23S rDNA internal transcribed spacer regions. J Gen Appl Micro- biol 2004; 50: 189-195.
Bartowsky EJ, Xia D, Gibson RL, Fleet GH, Henschke PA: Spoilage of bottled red wine by acetic acid bacteria. Lett Appl Microbiol 2003; 36:307-314.
Nanda K, Taniguchi M, Ujike S, Ishihara N, Mori H, Ono H, Murooka Y: Characterization of acetic acid bacteria in traditional acetic acid fermentation of rice vinegar (komesu) and unpolished rice vinegar (kurosu) produced in Japan. Appl Environ Microbiol 2001; 67:986-990.
Carlotti A, Funke G: Rapid distinction of Brevibacte- rium species by restriction analysis of rDNA generated by polymerase chain reaction. SYST APPL MICROBIOL 1994; 17: 380-386.
De Ory I, Romero LE, Cantero D: Modelling the ki- netics of growth of Acetobacter aceti in discontinuous culture: Influence of the temperature of operation. Appl Microbiol Biotechnol 1998; 49: 189-193.
Moonmangmee D, Adachi O, Ano Y, Shinagawa E, Toyama H, Theeragool G, Lotong N, Matsushita K: Isolation and Characterization of Thermotolerant Glu- conobacter Strains Catalyzing Oxidative Fermentation at Higher Temperatures. Biosci Biotechnol Biochem 2000; 64: 2306-2315.
Drysdale GS, Fleet GH: Acetic Acid Bacteria in Wine-making: A Review. Am J Enol Vitic 1988; 39: 143-154.
De Vero LGM, Gala E, Giudici P: Aspetti microbiologi- ci nella gestione di una batteria di Aceto Balsamico Tradizionale. Industrie Delle Bevande 2006; 3535:443-448.
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| Issue | Vol 2 No 1 (2010) | |
| Section | Articles | |
| Keywords | ||
| Acetic acid bacteria Acetobacter pasteurianus Vinegar Isolation Iran | ||
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