Identifikacija i karakterizacija bakterija Bacillus cereus izoliranih iz hrane brzim pirosekvenciranjem i analizom masnih kiselina

In this study rapid pyrosequencing was used for identification and fatty acid analysis of enterotoxic and non-enterotoxic Bacillus cereus. Presumptive B. cereus strains were isolated from ready-to-eat foods on mannitol-egg yolk-polymyxin agar and the identification was made based on the characteristics of B. cereus sp. Pyrosequencing was carried out on amplicons derived from 3 different 16S gene regions of rDNA using commercial reagent kits. All 30 presumptive Bacillus cereus isolates were identified as B. cereus. At least two sequencing reads of 3 different 16S rDNR gene regions were specific for identification of individual B. cereus isolates. Among B. cereus isolates, 53.3 % were found to be enterotoxic (determined by BCET-RPLA immunoassay kit). Fatty acids produced by more than 50 % of the investigated B. cereus were assumed as typical for these bacteria. The analyzed B. cereus produced a total of 25 typical fatty acids. The strains were highly homogeneous with dominant C4:0, C14:0, C16:0, C16:1, C18:0, C18:1, C18:2 and C22:6n3 fatty acids. Enterotoxic B. cereus was differentiated from the non-enterotoxic by significantly lower amounts of C18:0 and the absence of C18:4 fatty acid. Non-enterotoxic B. cereus did not produce C21:0, C17:0, C17:1, C20:0 and C15:1 fatty acids. The observed differences of individual fatty acid amounts and similar composition of fatty acids within all investigated B. cereus strains allowed the characterization of these bacteria isolated from ready-to-eat foods.U ovom su radu brzim pirosekvenciranjem identificirane bakterije vrste Bacillus cereus izolirane iz gotove hrane, te su analizom masnih kiselina izdvojeni enterotoksični izolati. Bakterije su izolirane na hranjivoj podlozi od manitola, polimiksina i žutanjka, te okarakterizirane na osnovu značajki vrste B. cereus. Pirosekvenciranje je provedeno na umnošcima iz tri različite regije gena 16S rDNA pomoću komercijalnih setova reagensa. Svih je 30 izolata identificirano kao vrsta Bacillus cereus. Najmanje su dvije sekvence iz tri različite regije gena 16S rDNA bile specifične za određivanje izolata B. cereus. Pomoću imunoenzimskog testa BCET-RPLA utvrđeno je da 53,3 % izolata bakterije B. cereus luči enterotoksine. Više od 50 % ispitanih bakterija proizvelo je masne kiseline karakteristične za vrstu B. cereus. Sojevi su bili vrlo homogeni, a od 25 ukupno proizvedenih masnih kiselina prevladavale su sljedeće: C4:0, C14:0, C16:1, C18:0, C18:1, C18:2 i C22:6n3. Enterotoksične su bakterije sadržavale bitno manju količinu masne kiseline C18:0, te u svom sastavu nisu imale masnu kiselinu C18:4. Izolati bakterije B. cereus što ne luče enterotoksin nisu sadržavali masne kiseline C21:0, C17:0, C17:1, C20:0 i C15:1. Razlika u količini pojedinih masnih kiselina te njihov sličan sastav u svim ispitanim izolatima omogućili su karakterizaciju bakterija B. cereus izoliranih iz gotove hrane

Utjecaj novih fermentiranih proizvoda, koji sadržavaju ekstrudiranu pšenicu, na kakvoću pšeničnoga kruha

Lactobacillus sakei MI806, Pediococcus pentosaceus MI810 and Pediococcus acidilactici MI807, able to produce bacteriocin-like inhibitory substances, were originally isolated from Lithuanian spontaneous rye sourdough and adapted in the novel fermentation medium containing extruded wheat material. The novel fermented products (50 and 65 % moisture content) were stored at the temperatures used in bakeries (15 days at 30–35 °C in the summer period or 20 days under refrigeration conditions at 0–6 °C). The number of lactic acid bacteria (LAB) was determined during the storage of fermented products for 15–20 days. Furthermore, the effect of novel fermented products stored under different conditions on wheat bread quality was examined. Extruded wheat material was found to have a higher positive effect on LAB growth compared to the control medium by lowering the reduction of LAB populations in fermented products with the extension of storage time and increase of temperature. During storage, lower variation and lower decrease in LAB count were measured in the novel fermented products with a moisture content of 65 % compared to those with 50 %. Furthermore, this humidity allows for the production of a product with higher moisture content in continuous production processes. The addition of the new fermented products with 65 % humidity to the wheat bread recipe (10 % of the quantity of flour) had a significant effect on bread quality: it increased the acidity of the crumb and specific volume of the bread, and decreased the fractal dimension of the crumb pores and crumb firmness. Based on the microbiological investigations of fermented products during storage and baking tests, the conditions of LAB cultivation in novel fermentation media were optimized (time of cultivation approx. 20 days at 0–6 °C and approx. 10 days at 30–35 °C).Lactobacillus sakei MI806, Pediococcus pentosaceus MI810 i Pediococcus acidilactici MI807, bakterije koje proizvode inhibitore slične bakteriocinu, izolirane su iz litvanskoga kiselog tijesta što se proizvodi od raženoga brašna, te prilagođene za uporabu u novoj fermentacijskoj podlozi s ekstrudiranom pšenicom. Da bi se ispitala mogućnost njihove primjene u pekarama, ti novi fermentirani proizvodi (s 50 i 65 % vlažnosti) skladišteni su 15 dana pri ljetnim temperaturama od 30 do 35 °C i 20 dana u hladnjaku pri 0 do 6 °C. Tijekom tih 15 do 20 dana ispitivan je broj mliječno-kiselih bakterija u fermentiranim proizvodima, te njihov utjecaj na kakvoću kruha. Dodatak ekstrudirane pšenice bitno je utjecao na rast mliječno-kiselih bakterija, u usporedbi s kontrolnom podlogom, smanjujući njihovo odumiranje u fermentiranim proizvodima tijekom duljeg skladištenja pri povišenoj temperaturi. Tijekom skladištenja primijećene su manje promjene broja mliječno-kiselih bakterija u novim fermentiranim proizvodima sa 65 % vlažnosti nego u onima s 50 % vlažnosti, pa je zaključeno da se njihovim dodatkom u kontinuiranom procesu može proizvesti kruh s većim udjelom vlage. Dodatak 10 % novih fermentiranih proizvoda sa 65 % vlažnosti pšeničnom brašnu bitno je utjecao na kakvoću kruha: povećali su se kiselost krušnih mrvica i specifični volumen kruha, a smanjila fraktalna dimenzija pora te čvrstoća mrvica. Na osnovi mikrobioloških istraživanja fermentiranih proizvoda tijekom skladištenja te pokusnog pečenja kruha optimirani su uvjeti uzgoja mliječno-kiselih bakterija u novim fermentiranim proizvodima (optimalno vrijeme uzgoja bilo je otprilike 20 dana pri 0-6 °C i 10 dana pri 30-35 °C)

Application of Edible Coating Based on Liquid Acid Whey Protein Concentrate with Indigenous Lactobacillus helveticus for Acid-Curd Cheese Quality Improvement

publishedVersio

Rapid Pyrosequencing and Fatty Acid Analysis for Characterization of Bacillus cereus Isolates from Food

In this study rapid pyrosequencing was used for identification and fatty acid analysis of enterotoxic and non-enterotoxic Bacillus cereus. Presumptive B. cereus strains were isolated from ready-to-eat foods on mannitol-egg yolk-polymyxin agar and the identification was made based on the characteristics of B. cereus sp. Pyrosequencing was carried out on amplicons derived from 3 different 16S gene regions of rDNA using commercial reagent kits. All 30 presumptive Bacillus cereus isolates were identified as B. cereus. At least two sequencing reads of 3 different 16S rDNR gene regions were specific for identification of individual B. cereus isolates. Among B. cereus isolates, 53.3 % were found to be enterotoxic (determined by BCET-RPLA immunoassay kit). Fatty acids produced by more than 50 % of the investigated B. cereus were assumed as typical for these bacteria. The analyzed B. cereus produced a total of 25 typical fatty acids. The strains were highly homogeneous with dominant C4:0, C14:0, C16:0, C16:1, C18:0, C18:1, C18:2 and C22:6n3 fatty acids. Enterotoxic B. cereus was differentiated from the non-enterotoxic by significantly lower amounts of C18:0 and the absence of C18:4 fatty acid. Non-enterotoxic B. cereus did not produce C21:0, C17:0, C17:1, C20:0 and C15:1 fatty acids. The observed differences of individual fatty acid amounts and similar composition of fatty acids within all investigated B. cereus strains allowed the characterization of these bacteria isolated from ready-to-eat foods

The Impact of Novel Fermented Products Containing Extruded Wheat Material on the Quality of Wheat Bread

Lactobacillus sakei MI806, Pediococcus pentosaceus MI810 and Pediococcus acidilactici MI807, able to produce bacteriocin-like inhibitory substances, were originally isolated from Lithuanian spontaneous rye sourdough and adapted in the novel fermentation medium containing extruded wheat material. The novel fermented products (50 and 65 % moisture content) were stored at the temperatures used in bakeries (15 days at 30–35 °C in the summer period or 20 days under refrigeration conditions at 0–6 °C). The number of lactic acid bacteria (LAB) was determined during the storage of fermented products for 15–20 days. Furthermore, the effect of novel fermented products stored under different conditions on wheat bread quality was examined. Extruded wheat material was found to have a higher positive effect on LAB growth compared to the control medium by lowering the reduction of LAB populations in fermented products with the extension of storage time and increase of temperature. During storage, lower variation and lower decrease in LAB count were measured in the novel fermented products with a moisture content of 65 % compared to those with 50 %. Furthermore, this humidity allows for the production of a product with higher moisture content in continuous production processes. The addition of the new fermented products with 65 % humidity to the wheat bread recipe (10 % of the quantity of flour) had a significant effect on bread quality: it increased the acidity of the crumb and specific volume of the bread, and decreased the fractal dimension of the crumb pores and crumb firmness. Based on the microbiological investigations of fermented products during storage and baking tests, the conditions of LAB cultivation in novel fermentation media were optimized (time of cultivation approx. 20 days at 0–6 °C and approx. 10 days at 30–35 °C)