Kaikki aineistot
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Wild berries are interesting research subjects due to their rich sources of health-beneficial phenolic compounds. However, the internal microbial communities, endophytes, associated with the wild berry fruits are currently unknown. Endophytes are bacteria or fungi inhabiting inside plant tissues, and their functions vary depending on the host species and environmental parameters. The present study aimed to examine community composition of fungal and bacterial endophytes in fruits of three wild berry species (bilberry Vaccinium myrtillus L., lingonberry Vaccinium vitis-idaea L. and crowberry Empetrum nigrum L.) and the effects of host plant species and their growth sites on shaping the endophytic communities. We found that the endophytic community structures differed between the berry species, and fungi were predominant over bacteria in the total endophytic taxa. We identified previously unknown endophytic fungal taxa including Angustimassarina, Dothidea, Fellozyma, Pseudohyphozyma, Hannaella coprosmae and Oberwinklerozyma straminea. A role of soluble phenolic compounds, the intracellular components in wild berry fruits, in shaping the endophytic communities is proposed. Overall, our study demonstrates that each berry species harbors a unique endophytic community of microbes.
Abstract The aim of this work was to assess and improve the enrichment cultivation of food-contaminating bacteria prior to detection by means of RNA-based sandwich hybridization assay (SHA). The examples of beer-spoiling lactic acid bacteria (LAB) and food-borne Salmonella Typhimurium were selected based on their relevance in Finnish food industry. Also universal challenges affecting on the selection of the enrichment cultivation procedure are discussed, including some potential possibilities for improved enrichment cultivation. The results of this study may therefore be used for the assessment of the efficiency of bacterial cultivation in other applications. The evaluation of the enrichment cultivation procedures prior to SHA lead to following conclusions: i) the enrichment cultivation procedure is necessary prior to rRNA-based SHA, and it directly influences the accuracy of SHA; ii) the improvement of the enrichment cultivation may allow faster recovery and growth of bacteria; iii) the improved recovery of bacteria can be achieved by reducing environmental stress factors in the enrichment culture; and iv) the growth of bacteria may be accelerated by assuring the selectivity of medium and allowing accessibility to growth factors. Several growth factors were studied by means of full factorial design and response surface modeling. Measured cell densities, as well as predicted lag-times and maximum growth rates in the bacterial cultures were used as responses. The results show that small shifts in the cultivation conditions extend the lag-time and decrease the growth rate of both LAB and Salmonella. Besides adjusting the temperature and pH, the growth of LAB was facilitated by reducing osmotic and oxidative stresses in the enrichment medium. In this study, a novel enzyme controlled glucose delivery system was used for the first time in the enrichment cultivation of food-contaminating bacteria. The glucose delivery system improved the growth of LAB in single strain cultures and in actual brewing process samples. The recovery of injured Salmonella was also enhanced by using the glucose delivery system together with selective siderophore ferrioxamine E, both in terms of reduced lag-times and increased growth rates. Based on the SHA, the adjusted BPW broth enhanced the molecular detection of heat-injured Salmonella in meat.
Abstract This thesis aimed at developing methods for tracking the environmental origins of microbial contaminants of the food chain. We worked on three targets: i) environmental mycobacteria ii) toxinogenic Bacillus species iii) post-harvest fungi in strawberry jam. Our aim was to develop methods for early detection of the above contaminants, which have the potential to endanger consumer health. We developed a novel method based on 16S rRNA hybridization for tracking the reservoirs of potentially pathogenic environmental mycobacteria in piggeries and soil. From 1010 to 1012 16S rRNA molecules of environmental mycobacteria were found per gram of peat, wood shavings and straw in piggeries with a high prevalence of infections. These beddings may thus be a source of mycobacteria for pigs. We found 1010–1011 of mycobacterial 16S rRNA molecules per gram of Finnish forest soil, indicating that the soil contained 107–109 mycobacteria per gram. These numbers exceed the previous cultivation-based estimates of mycobacterial content in Finnish soils. To elucidate the role of mastitis in the input of toxinogenic Bacillus into the dairy production chain, milks were sampled from mastitic cows. Twenty-three Bacillus isolates were screened for toxins using the sperm cell motility inhibition assay. Four of the six toxinogenic isolates found were identified as Bacillus pumilus and two as Bacillus licheniformis. The isolates produced toxic substances that were heat-stable (100 °C) and soluble in methanol, thus being of non-protein nature. The extracts prepared from the toxin-producing isolates disrupted the plasma membrane of exposed sperm cells at concentrations 1–15 μg ml-1 (B. pumilus) 20–30 μg ml-1 (B. licheniformis). The toxic action of the mastitis-associated B. licheniformis strains was similar to that of the lipopeptide lichenysin A. The genes for lichenysin synthetase were found in these strains by PCR. This study revealed that heat-stable toxin-producing strains of B. pumilus and B. licheniformis occur in milk of mastitic milking cows. They may enter the dairy production chain when milk of clinically healthy cows recovered from mastitis is sent to dairies. Many foodborne contaminant fungi are known to produce volatile organic compounds. We investigated the suitability of such metabolites as early indicators of fungal contamination of strawberry jam. We found that volatile organic compounds commonly produced by the contaminant fungi in strawberry jam were 2-pentanone, styrene, 3-methyl-1-butanol, 1,3-pentadiene and ethanol. The results indicate that these compounds could be used to detect fungal contamination of jam.
Tämä insinöörityö tehtiin Teknologian tutkimuskeskus VTT Oy:n toimeksiantona. Työn tärkeimpänä tavoitteena oli selvittää tiettyjen hiivalajikkeiden turvallisuutta oluen valmistuksessa sekä sitä, kuinka näistä lajikkeista tarvittaessa päästään eroon mahdollisen kontaminaation sattuessa. Eri hiivalajikkeet mahdollistavat uusien olutmakujen kehittämisen ja tästä syystä niiden lisätutkimus muun muassa turvallisuuden osalta on tärkeää. Tutkimuksen taustalla on muun muassa VTT:n pilottipanimon aiemmat havainnot Kazachstania servazzi-kannan aiheuttamista kontaminaatioista. Tästä syystä eri hiivalajikkeiden tarkemmalle tutkimiselle nähtiin tarvetta ja niiden käyttäytymisestä eri olosuhteissa haluttiin lisätietoa. Tässä insinöörityössä tutkittiin 25:tä eri hiivaa, joista suurin osa on villihiivoja sekä tunnettuja kontaminantteja ruoka- ja juomateollisuudessa. Tutkimus toteutettiin VTT:n panimon laboratoriossa, jossa testattiin biofilmin tuotantoa, desinfiointiainetoleranssia, säilöntäaineiden sietokykyä sekä hiivojen kykyä kasvaa eri lämpötiloissa. Käytettäessä uusia hiivoja tulisi niiden käytöstä ja kehitystä tarkkailla sekä varmistua riittävän hygieenisistä olosuhteista tuotannossa. Insinöörityössä huomattiin, että suurin osa tutkituista hiivoista voisi tulosten perusteella olla turvallisia oluen panemiseen. Tutkimuksessa kävi ilmi, että testatuista hiivalajikkeista K. servazzi oli kaikkein hankalimmin hallittavissa oleva hiivakanta ja sen käyttö oluen valmistuksessa vaikuttaisi sisältävän suurimmat riskit. Jatkotutkimusta tarvittaisiin etenkin niille hiivoille jotka tuottivat kokeissa paljon biofilmiä. Jatkokokeissa olisi hyvä testata niiden sietokykyä mm. puhdistusaineille. Lisäksi tulisi jatkotutkimuksen avulla selvittää joidenkin hiivojen kliininen merkitys esimerkiksi antibioottitesteillä.
Abstract Wild berries are interesting research subjects due to their rich sources of health-beneficial phenolic compounds. However, the internal microbial communities, endophytes, associated with the wild berry fruits are currently unknown. Endophytes are bacteria or fungi inhabiting inside plant tissues, and their functions vary depending on the host species and environmental parameters. The present study aimed to examine community composition of fungal and bacterial endophytes in fruits of three wild berry species (bilberry Vaccinium myrtillus L., lingonberry Vaccinium vitis-idaea L. and crowberry Empetrum nigrum L.) and the effects of host plant species and their growth sites on shaping the endophytic communities. We found that the endophytic community structures differed between the berry species, and fungi were predominant over bacteria in the total endophytic taxa. We identified previously unknown endophytic fungal taxa including Angustimassarina, Dothidea, Fellozyma, Pseudohyphozyma, Hannaella coprosmae and Oberwinklerozyma straminea. A role of soluble phenolic compounds, the intracellular components in wild berry fruits, in shaping the endophytic communities is proposed. Overall, our study demonstrates that each berry species harbors a unique endophytic community of microbes.