Alicyclobacillus

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Alicyclobacillus
Systematics
Domain : Bacteria (bacteria)
Department : Firmicutes
Class : Bacilli
Order : Bacillales
Family : Alicyclobacillaceae
Genre : Alicyclobacillus
Scientific name
Alicyclobacillus
Wisotzkey 1992 emend. Karavaiko et al. 2005

Alicyclobacillus is a genus of gram-positive , endospore-forming soil bacteria . Several species containunusual fatty acids called omega- alicyclic in the lipids of the biomembrane . The generic name refers to these fatty acids. Alicyclobacillus species are thermophilic and they grow even at low pH levels . They are involved in the bacterial spoilage of fruit juices and thus represent a possible problem in their production.

features

Appearance

Alicyclobacillus is a genus of gram-positive bacteria, in older cultures they are gram- variable. The cells are rod- shaped with a diameter of 0.3 to 0.8  µm and a length of 2.0 to 4.5 µm. They can move actively, they are motile . Under unfavorable environmental conditions, they develop endospores as persistent forms. The formation of the endospores takes place regardless of whether there is enough oxygen. The endospores of some Alicyclobacillus species are very heat-resistant and survive brief heating to 92 ° C. This is problematic for the fruit juice industry.

Growth and metabolism

Alicyclobacillus grows strictly aerobically , so it can only reproduce when oxygen is present. Alicyclobacillus species are thermophilic (“warmth-loving”) and acidophilic (“acid-loving”). They grow at temperatures between 20 and 70 ° C, with the optimal growth temperature between 35 and 60 ° C. The pH optimum is between 2.0 and 6.0.

Chemotaxonomic Features

Omega alicyclic fatty acids in the cell membrane of Alicyclobacillus , with reference to the names Alpha and Omega .
13-Cyclohexyltridecanoic acid.svg
Omega-cyclohexyltridecanoic acid (ω-cyclohexane C 19: 0 )
11-Cyclohexylundecanoic acid (svg) .svg
Omega-cyclohexylundecanoic acid (ω-cyclohexane C 17: 0 )
2-Hydroxy-11-Cycloheptylundecanoic acid. Svg
Omega-cycloheptyl-alpha-hydroxyundecanoic acid (ω-cycloheptane C 18: 0 2-OH)

Although a low GC content in the bacterial DNA is typical for Firmicutes , it is rather high in Alicyclobacillus species, between 48.7 and 62.7 mol percent. The main quinone is MK-7 type menaquinone . The lipids in the biomembrane contain unusual fatty acids , which are referred to as omega-alicyclic (ω- alicyclic ) and which have been taken into account in the generic name. The Greek letter omega refers to the last position in the carbon chain. The use of lowercase Greek letters as a locant is common with fatty acids. Alicyclic compounds are cyclic hydrocarbons . The rings in the structure of these fatty acids usually consist of six or seven carbon atoms; such ring systems are referred to as cyclohexane or cycloheptane .

This peculiarity is referred to in the species name Alicyclobacillus cycloheptanicus . In this type, fatty acids with a terminal cycloheptane are predominantly found in the cell membrane: Omega-cycloheptylundecanoic acid (ω-cycloheptane C 18: 0 ), omega-cycloheptyltridecanoic acid (ω-cycloheptane C 20: 0 ) and omega-cycloheptyl-alpha-hydroxyundecanoic acid (ω -Cycloheptane C 18: 0 2-OH). In A. acidocaldarius and A. acidiphilus the omega-alicyclic fatty acids with a ring of six carbon atoms predominate: omega-cyclohexylundecanoic acid (ω-cyclohexane C 17: 0 ) and omega-cyclohexyltridecanoic acid (ω-cyclohexane C 19: 0 ).

With the discovery of the species A. pomorum the description of typical fatty acids 2003 has been expanded: In addition to omega-alicyclic fatty acids and straight-chain and branched , saturated before fatty acids. Typical representatives of the latter are fatty acids with the abbreviations iso -C 15: 0 ( iso- pentadecanoic acid), anteiso -C 15: 0 ( anteiso- pentadecanoic acid), iso -C 16: 0 ( iso- hexadecanoic acid), iso -C 17 : 0 ( iso- heptadecanoic acid) and anteiso -C 17: 0 ( anteiso- heptadecanoic acid). The fatty acids with a total of 15 or 17 carbon atoms also belong to the odd-numbered fatty acids.

Occurrence

The habitat of several Alicyclobacillus species can be found in extreme locations. The first Alicyclobacillus strain was isolated from hot water springs in Japan in 1967 . In the following years, both the water and the soil were investigated at locations where thermophilic and acidophilic bacteria can grow. The discovered bacteria were later assigned to the genus Alicyclobacillus . Alicyclobacillus species could also be isolated from soils that are not in contact with thermal springs , e.g. B. from arable soils. Only in 1982 was Alicyclobacillus with spoilage of fruit juices associated since several species in acidic been found drinks, such as A. acidoterrestris in apple juice and passion fruit juice and A. acidiphilus . The species A. contaminans and A. fastidiosus , discovered in 2007, were isolated from orange juice and apple juice, respectively. Microbiological tests carried out in Brazil on orange trees and the soil in which they grow showed the presence of Alicyclobacillus and the bacterial spores, respectively. They can be transferred from the soil, as a natural habitat, to the fruit and, under certain circumstances, enter the fruit juice production process. A species ( A. sacchari ) has even been discovered in liquid sugar .

Systematics

The first Alicyclobacillus strain was isolated from hot water sources in 1967 and, after further research, was named as the new species Bacillus acidocaldarius in 1971 . Other newly discovered species were initially assigned to the genus Bacillus . In 1992, Alicyclobacillus was proposed as a new genus based on 16S rRNA studies . Two bacterial strains that were previously assigned to the genus Sulfobacillus were added to the Alicyclobacilli in 2005 as A. tolerans and A. disulfidooxidans . In 2007, phylogenetic studies on 146 bacterial strains isolated from fruit juices, their raw materials and environmental samples, led to the first description of six new species.

The genus Alicyclobacillus in the family Alicyclobacillaceae is placed in the order Bacillales in the department of Firmicutes . The following species are known of the genus (status 2014):

Industrial importance

Alicyclobacilli are of particular interest to the fruit juice industry, as common pasteurization processes (92 ° C for 10 seconds) do not deactivate the spores. The high temperatures lead to spore germination. If Alicyclobacillus is present as an impurity in a product, a disinfectant-like smell and / or taste develops due to the metabolite guaiacol . Similar compounds, the halogenated phenols, are also produced by them as undesirable sensory components ( off-flavor ). The product remains optically perfect, i. H. no gases are formed that would expand the packaging and there is no discoloration. Alicyclobacillus contamination is not dangerous to humans. Alicyclobacilli have been associated with the spoilage of pears , mangoes and grape juice , as well as fruit juice mixes and products made from tomatoes .

The species A. acidoterrestris is considered to be the most important spoilage pathogen of the genus Alicyclobacillus , however A. acidocaldarius , A. pomorum and A. herbarius were also isolated from spoiled fruit juice products. Other species found in acidic beverages include A. acidiphilus , A. contaminans, and A. fastidiosus .

Fruit juice manufacturers work on the assumption that bacterial spores do not germinate at a pH below 4.6, and that acid-tolerant organisms are not very heat-resistant. With these assumptions, a pasteurization process can be carried out at low temperatures. Today it is possible to detect the pathogen early in the manufacturing process with the help of real-time PCR and to adapt the process to a positive result. Alicyclobacillus is therefore an important microorganism in quality control in the fruit juice industry. The importance of Alicyclobacillus as a spoilage agent has led some researchers to use A. acidoterrestris as the reference organism for developing a pasteurization process for highly acidic foods. Similarly, the decimal reduction time ("death time" at a certain temperature) of Clostridium botulinum was used to develop the sterilization process for low-acid, canned foods.

Analytics

The cultural determination according to IFU method no. 12 uses the physiological peculiarities of Alicyclobacillus spp. out. It is prepared in parallel on BAT (" Bacillus acidoterrestris ") medium with a low pH value and on a conventional plate count agar (PCA). Alicyclobacillus spp. grow exclusively on BAT agar. Likewise, heat-tolerant acidophilic accompanying flora grows on both media. The slow growth of Alicyclobacillus species is problematic for the detection . For reliable negative detection in processed products, incubation times cannot be reduced any further despite advances in the polymerase chain reaction (PCR) in order to avoid false negative results. Supplementary confirmation reactions by means of guaiacol detection (biochemical) and species identification by means of MALDI-TOF MS can accelerate and additionally secure the classical cultural analysis in the positive case. To avoid the long generation times and matrix effects , an aptamer- based enrichment of the spores is possible. Using real-time quantitative PCR of the ribosomal DNA , the bacterial spores can then be counted before they have germinated in the product. A modern culture-based method enables growth curves to be recorded during 96 hours of incubation. For confirmation, a PCR can be used in combination with a lateral flow test to quickly confirm the findings.

Individual evidence

  1. ↑ a b c d e f g J. D. Wisotzkey, P. Jurtshuk et al .: Comparative sequence analyzes on the 16S rRNA (rDNA) of Bacillus acidocaldarius, Bacillus acidoterrestris, and Bacillus cycloheptanicus and proposal for creation of a new genus, Alicyclobacillus gen. Nov. In: International journal of systematic bacteriology. Volume 42, No. 2, April 1992, pp. 263-269, ISSN  0020-7713 . PMID 1374624 .
  2. ↑ a b c K. Goto, K. Mochida and others: Alicyclobacillus pomorum sp. nov., a novel thermo-acidophilic, endospore-forming bacterium that does not possess omega-alicyclic fatty acids, and emended description of the genus Alicyclobacillus. In: International journal of systematic and evolutionary microbiology. Volume 53, No. 5, September 2003, pp. 1537-1544, ISSN  1466-5026 . PMID 13130044 .
  3. ↑ a b c Ana Cláudia NF Spinelli, S. Anderson Sant'Ana Salatir Rodrigues Junior, Pilar R. Massaguer: Influence of different filling, cooling, and storage conditions on the growth of Alicyclobacillus acidoterrestris CRA7152 in orange juice. In: Applied and environmental microbiology. Volume 75, No. 23, December 2009, pp. 7409-7416, ISSN  1098-5336 . doi: 10.1128 / AEM.01400-09 . PMID 19801469 . PMC 2786409 (free full text).
  4. ↑ a b c G. I. Karavaiko, TI Bogdanova et al: Reclassification of 'Sulfobacillus thermosulfidooxidans subsp. thermotolerans' strain K1 as Alicyclobacillus tolerans sp. nov. and Sulfobacillus disulfidooxidans Dufresne et al. 1996 as Alicyclobacillus disulfidooxidans comb. nov., and emended description of the genus Alicyclobacillus. In: International journal of systematic and evolutionary microbiology. Volume 55, No. 2, March 2005, pp. 941-947, ISSN  1466-5026 . doi: 10.1099 / ijs.0.63300-0 . PMID 15774689 .
  5. ↑ a b Su-Sen Chang, Dong-Hyun Kang: Alicyclobacillus spp. in the fruit juice industry: history, characteristics, and current isolation / detection procedures. In: Critical reviews in microbiology. Volume 30, Number 2, 2004, pp. 55-74, ISSN  1040-841X . doi: 10.1080 / 10408410490435089 . PMID 15239380 . (Review).
  6. ^ A b c Jean Euzéby, Aidan C. Parte: Genus Alicyclobacillus. In: List of Prokaryotic names with Standing in Nomenclature ( LPSN ).
  7. ↑ a b c d e H. Matsubara, K. Goto et al: Alicyclobacillus acidiphilus sp. nov., a novel thermo-acidophilic, omega-alicyclic fatty acid-containing bacterium isolated from acidic beverages. In: International journal of systematic and evolutionary microbiology. Vol. 52, No. 5, September 2002, pp. 1681-1685, ISSN  1466-5026 . PMID 12361274 .
  8. ^ A b F. Uchino, S. Doi: Acido-thermophilic Bacteria from Thermal Waters. In: Agricultural and Biological Chemistry. Volume 31, No. 7, 1967, pp. 817-822, ISSN  0002-1369 . doi: 10.1271 / bbb1961.31.817 .
  9. ↑ a b G. Darland, TD Brock: Bacillus acidocaldarius sp. nov., an Acidophilic Thermophilic Spore-forming Bacterium. In: Journal of General Microbiology. Volume 67, No. 1, July 1971, pp. 9-15, ISSN  0022-1287 . doi: 10.1099 / 00221287-67-1-9 .
  10. ↑ a b G. Deinhard, P. Blanz, K. Poralla, E. Altan: Bacillus acidoterrestris sp. nov., a New Thermotolerant Acidophile Isolated from Different Soils. In: Systematic and Applied Microbiology. Volume 10, No. 1, November 1987, pp. 47-53, ISSN  0723-2020 . doi: 10.1016 / S0723-2020 (87) 80009-7 .
  11. ↑ a b c d e K. Goto, K. Mochida et al.: Proposal of six species of moderately thermophilic, acidophilic, endospore-forming bacteria: Alicyclobacillus contaminans sp. nov., Alicyclobacillus fastidiosus sp. nov., Alicyclobacillus kakegawensis sp. nov., Alicyclobacillus macrosporangiidus sp. nov., Alicyclobacillus sacchari sp. nov. and Alicyclobacillus shizuokensis sp. nov. In: International journal of systematic and evolutionary microbiology. Volume 57, No. 6, June 2007, pp. 1276-1285, ISSN  1466-5026 . doi: 10.1099 / ijs.0.64692-0 . PMID 17551043 .
  12. ^ A b I. C. McKnight, MNU Eiroa, AS Sant'Ana, PR Massaguer: Alicyclobacillus acidoterrestris in pasteurized exotic Brazilian fruit juices: isolation, genotypic characterization and heat resistance. In: Food microbiology. Volume 27, Number 8, December 2010, pp. 1016-1022, ISSN  1095-9998 . doi: 10.1016 / j.fm.2010.06.010 . PMID 20832679 .
  13. ^ A b Willem H. Groenewald, Pieter A. Gouws, R. Corli Witthuhn: Isolation, identification and typification of Alicyclobacillus acidoterrestris and Alicyclobacillus acidocaldarius strains from orchard soil and the fruit processing environment in South Africa. In: Food microbiology. Volume 26, No. 1, February 2009, pp. 71-76, ISSN  1095-9998 . doi: 10.1016 / j.fm.2008.07.008 . PMID 19028308 .
  14. ↑ Mirtha Nelly Uboldi Eiroa, Valéria Christina Amstalden Junqueira, Flaávio Schmidt: Alicyclobacillus in orange juice: occurrence and heat resistance of spores. In: Journal of food protection. Vol. 62, No. 8, August 1999, pp. 883-886, ISSN  0362-028X . PMID 10456741 .
  15. ↑ Filipa VM Silva, Paul Gibbs: Alicyclobacillus acidoterrestris spores in fruit products and design of pasteurization processes. In: Trends in Food Science & Technology. Volume 12, No. 2, February 2001, pp. 68-74, ISSN  0924-2244 . doi: 10.1016 / S0924-2244 (01) 00070-X .
  16. ↑ S. Kohl: Alicyclobacillus in fruit juice . In: Beverage Industry. Vol. 5, May 2011, pp. 26-27.
  17. ↑ Filipa VM Silva, Paul Gibbs: Target selection in designing pasteurization processes for shelf-stable high-acid fruit products. In: Critical reviews in food science and nutrition. Volume 44, No. 5, 2004, pp. 353-360, ISSN  1040-8398 . doi: 10.1080 / 10408690490489251 . PMID 15540648 . (Review).
  18. ^ A. Rau, M. Mailänder, B. Schütze: Detection of acidophilic beverage spoilers. In: Deutsche Lebensmittel-Rundschau . September 2016.
  19. ↑ Tim Hünniger, Christine Felbinger, Hauke ​​Wessels, Sophia Mast, Antonia Hoffmann: Food Targeting: A Real-Time PCR Assay Targeting 16S rDNA for Direct Quantification of Alicyclobacillus spp. Spores after Aptamer-Based Enrichment . In: Journal of Agricultural and Food Chemistry . tape 63 , no. 17 , May 6, 2015, ISSN  0021-8561 , p. 4291-4296 , doi : 10.1021 / acs.jafc.5b00874 .
  20. ↑ Detection of Alicyclobacillus possible within 96 hours , Liquid Fruit, July 17, 2019