Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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    Risk assessment of Listeria monocytogenes in Gouda cheese
    Wemmenhove, Ellen - \ 2019
    Wageningen University. Promotor(en): M.H. Zwietering; A.C.M. van Hooijdonk, co-promotor(en): M.H.J. Wells-Bennik. - Wageningen : Wageningen University - ISBN 9789463435628 - 197

    In this PhD thesis, the potential of outgrowth of L. monocytogenes was assessed in Dutch-type Gouda cheese. It was demonstrated that L. monocytogenes, which can cause listeriosis, is not able to grow in or on Dutch-type Gouda. The factors present in Gouda cheese that can lead to full inhibition of growth of L. monocytogenes were identified and safety criteria aiming for complete inhibition of growth of L. monocytogenes in Dutch-type Gouda cheese are suggested.

    Dutch-type Gouda cheese is a semi-hard cheese made from bovine milk that is pasteurized when produced at an industrial scale. It is a ready-to-eat food with a pH > 5.0 and water activity aw > 0.94. In absence of scientific evidence that this product does not support growth, Dutch-type Gouda is classified by the European legislation as a ready-to-eat food product able to support growth of L. monocytogenes.

    In two challenge studies described in this thesis, it was demonstrated that Dutch-type Gouda cheese does not support growth of L. monocytogenes when inoculated in and on the product. During the cheese making process, entrapment but no growth of L. monocytogenes in the curd was observed. During subsequent ripening of the cheeses, no growth was observed, and upon prolonged ripening periods (>2 months) inactivation was found. In the second challenge study, a limited transfer of L. monocytogenes from brine to the outer layers of cheeses was observed, and during brining and ripening viable numbers of L. monocytogenes did not increase.

    The variation in aw inside Gouda cheese was assessed by determining the profiles of water and NaCl and the resulting aw in nature-ripened and foil-ripened Gouda cheese during brining and ripening. An empirical model was derived for Gouda cheese in which aw is expressed as a function of the NaCl-in-moisture content.

    Dutch-type cheeses contain organic acids that are known to have potential inhibitory effects on L. monocytogenes. The MICs of organic acids for 6 different L. monocytogenes strains were established at pH values that are relevant to Dutch-type Gouda. The MICs were established for lactic acid (which is the main organic acid in Gouda), acetic acid, propionic acid, and citric acid. Variations in MICs between strains were observed.

    Ifn an overall review of the factors present in Gouda cheese that are relevant to growth inhibition of L. monocytogenes, undissociated lactic acid was evaluated as the primary growth-inhibiting factor that can lead to full growth inhibition in Gouda. Additionally, low aw in the cheese rind and after prolonged ripening times can cause full growth inhibition.

    This thesis lends support to categorizing Gouda as a ready-to-eat food product that does not support growth of L. monocytogenes. Furthermore, it is justifiable to include undissociated lactic acid (together with pH and aw) in future food safety criteria for ready-to-eat products related to absence of growth of L. monocytogenes.

    Monitoring klimaateffect van NL agroproductie : verkenning van behoeften en ideeën om het klimaateffect geïntegreerd te meten
    Blonk, Hans ; Reijs, Joan ; Vellinga, Theun - \ 2018
    Gouda : Wageningen Economic Research - 19
    Factors that inhibit growth of Listeria monocytogenes in nature-ripened Gouda cheese: A major role for undissociated lactic acid
    Wemmenhove, E. ; Valenberg, H.J.F. van; Hooijdonk, A.C.M. van; Wells-Bennik, M.H.J. ; Zwietering, M.H. - \ 2018
    Food Control 84 (2018). - ISSN 0956-7135 - p. 413 - 418.
    Acetic acid - Diacetyl - Free fatty acids - Gouda cheese - Growth inhibition - Growth limit - L. monocytogenes - Lactoferrin - Nisin - Nitrate - Nitrite - pH - Safety criteria - Temperature - Undissociated lactic acid - Water activity
    In this study, factors relevant to nature-ripened Gouda cheese were evaluated for their potential to inhibit growth of Listeria monocytogenes. Factors included water activity, pH, undissociated acetic and lactic acid, diacetyl, free fatty acids, lactoferrin, nitrate, nitrite and nisin. In addition, the effect of temperature was evaluated. For each factor, the actual concentrations and values relevant to Gouda cheese were obtained and the inhibitory effect of these individual factors on growth of L. monocytogenes was assessed. This evaluation revealed that undissociated lactic acid is the most important factor for growth inhibition of L. monocytogenes in Gouda cheese and that, additionally, low water activity as present in the cheese rind and after prolonged ripening times can also cause full growth inhibition. Gouda cheeses have a typical total lactic acid content of 1.47% w/w. In a 2-week old Gouda cheese, with a pH value of 5.25 and a moisture content of 42% w/w, the concentration of undissociated lactic acid in the water phase is 10.9 mM. Growth of L. monocytogenes is not supported when the undissociated lactic acid concentration is > 6.35 mM. Concentrations of undissociated lactic acid in the water phase of Gouda cheese will be higher than this value when the total lactic acid content is > 0.86% w/w at a pH < 5.25 (relevant to young Gouda cheese), or > 1.26% w/w at a pH < 5.50 for mature Gouda cheese (moisture content of 35% w/w). This study underlines the importance of undissociated lactic acid as growth inhibitor for L. monocytogenes in Gouda cheese.
    Measuring temporal liking simultaneously to Temporal Dominance of Sensations in several intakes. An application to Gouda cheeses in 6 Europeans countries
    Thomas, A. ; Chambault, M. ; Dreyfuss, L. ; Gilbert, C.C. ; Hegyi, A. ; Henneberg, S. ; Knippertz, A. ; Kostyra, E. ; Kremer, S. ; Silva, A.P. ; Schlich, P. - \ 2017
    Food Research International 99 (2017)1. - ISSN 0963-9969 - p. 426 - 434.
    Gouda cheese - Liking - Temporal Dominance of Sensations (TDS) - Temporal Drivers of Liking (TDL)
    The idea of having untrained consumers performing Temporal Dominance of Sensations (TDS) and dynamic liking in the same session was recently introduced (Thomas, van der Stelt, Prokop, Lawlor, & Schlich, 2016). In the present study, a variation of the data acquisition protocol was done, aiming to record TDS and liking simultaneously on the same screen in a single session during multiple product intakes. This method, called Simultaneous Temporal Drivers of Liking (S-TDL), was used to describe samples of Gouda cheese in an international experiment.To test this idea, consumers from six European countries (n = 667) assessed 4 Gouda cheeses with different ages and fat contents during one sensory evaluation session. Ten sensory attributes and a 9-point hedonic scale were presented simultaneously on the computer screen. While performing TDS, consumers could reassess their liking score as often as they wanted. This new type of sensory data was coded by individual average liking scores while a given attribute was perceived as dominant (Liking While Dominant; LWD).Although significant differences in preference were observed among countries, there were global preferences for a longer dominance of melting, fatty and tender textures. The cheese flavour attribute was the best positive TDL, whereas bitter was a strong negative TDL. A cluster analysis of the 667 consumers identified three significant liking clusters, each with different most and least preferred samples. For the TDL computation by cluster, significant specific TDL were observed. These results showed the importance of overall liking segmentation before TDL analysis to determine which attributes should have a longer dominance duration in order to please specific consumer targets.
    Mixed culture engineering for steering starter functionality
    Spuś, Maciej - \ 2016
    Wageningen University. Promotor(en): E.J. Smid; Tjakko Abee. - Wageningen : Wageningen University - ISBN 9789462578333 - 170
    bacteriophages - predation - microorganisms - starters - genetics - diversity - bacteriofagen - predatie - micro-organismen - zuursels - genetica - diversiteit

    Undefined mixed complex starter cultures are broadly used in Gouda-type cheese production due to their robustness to phage predation, resilience for changes in environmental conditions and aroma compounds production ability during ripening. These microbial communities of lactic acid bacteria prior their isolation and deposition in starter culture collections were continuously used at the farm-level production facilities. Thus, one can consider undefined mixed complex starters as domesticated microbial communities. The process of domestication was facilitated by humans who have been continuously repeating successful fermentations using part of previous batch as inoculum (i.e. back-slopping). Therefore, a term ‘community breeding’ can describe this human-driven domestication of microbial communities. Community breeding of a model complex starter Ur led to establishment of a simple two-species composition of Lactococcus lactis and Leuconostoc mesenteroides represented by, in total, 8 genetic lineages. At the same time, this simple microbial community displays a high degree of intraspecies diversity, presumably caused by evolutionary processes of horizontal gene transfer, genome decay and mutations. Such diversity at strain level is particularly interesting in the context of continuous bacteriophage predation pressure present in this microbial community. It is thought that constant-diversity (CD) dynamics, based on the ‘kill-the-winner’ principles, is operational in Ur starter at the strain level. According to CD model, the fittest strain(s), which feed on the most abundant substrate, will be selected against due to density-dependent phage predation. The control of the fittest strain abundance by bacteriophages opens space for differentiation of strains via eco-evolutionary feedbacks. In particular, strains of complex starter culture not only adapted to quickly acidify milk (via efficient consumption of lactose and protein to peptides degradation), but concurrently, to consume other substrates present in milk. In addition, throughout the process of community breeding microbe-microbe interactions between community members have evolved. These interactions have led to division of metabolic labor among strains present in the culture, and eventually to better starter microbial community functioning.

    The aim of this thesis was to investigate the factors impacting the formation of compositionally and functionally stable undefined mixed complex starter cultures to further use this knowledge in steering its functionality, and potentially in developing new strategies for robust starter culture design. To facilitate this study, well-characterized Ur culture strain isolates were used to systematically reconstitute the starter culture into multi-strain blends with increasing level of strain and genetic lineage diversity. The investigation of factors such as phage predation, level of strain and genetic lineage diversity as well as environmental conditions, was performed during experimental evolution studies in milk. The functionality of the (evolved) starter cultures was tested in an adapted lab-scale MicroCheese model system. The specific approach used in each of the research chapters is described below in more detail.

    Strains isolated from Ur starter culture were characterized in terms of their resistance against bacteriophages isolated from the same starter (Chapter 2). This test confirmed high diversity in phage resistance among strains belonging to different genetic lineages as well as among strains of the same lineage. Next, selected strains, which represented different levels of bacteriophage predation: resistant, moderately resistant, sensitive and no detectable sensitivity, were mixed in simple blends containing 4 strains representing 3 genetic lineages of Ur starter (3 such blends were designed). These blends were exposed to phage predation (one phage per blend) at the onset of prolonged sequential propagation experiment or propagated without phage addition (control). Throughout the serial propagation the genetic lineage composition was monitored. During the propagation of control blends we detected quick domination of a single lineage. This dominating lineage contained strains sensitive to phages. Genetic lineage level composition of the phage-challenged blends was much more dynamic suggesting the impact of phage predation. The relatively low strain diversity introduced in these blends was not high enough to sustain maximal diversity at the level of lineages.

    Chapter 3 describes a study using defined blends with higher complexity by extending the number of strains used. In total, 24 strains representing all 8 Ur starter lineages were exposed in sequential propagation experiment to a cocktail of 3 phages isolated from Ur starter. The propagation in milk of this multi-strain blend was executed for more than 500 generations and the abundance of genetic lineages was monitored throughout. Similarly as in the simple blends experiment, control blends were not exposed to bacteriophages. In control blends we observed a domination of one genetic lineage upon serial propagation, which resembles a periodic-selection-like (PS) behavior, where the fittest strains are dominating the microbial community and in result genetic-lineage diversity is being substantially reduced. In contrast, the composition of phage-challenged blends was again more dynamic than in control blends. In one of the phage-challenged blends behavior characteristic for a constant-diversity dynamics model was observed; throughout the serial transfer experiment, genetic lineage diversity was maintained by the presence of phage predation at relatively high level. In case of the second phage-challenged blend, due to a stochastic event, which likely caused a reduction in phage pressure, we observed a gradual recovery of the fittest strains, which again resembled a periodic-selection behavior. Therefore, phage predation, among other factors, can lead to shifts in microbial community population dynamics resulting in alternative stable states.

    The experimental evolution approach, resembling traditional process of back-slopping, was used in a Long-term experimental evolution of Undefined Mixed Starter Culture (LUMSC) study described in Chapter 4. The aim of this study was to investigate the compositional and functional stability ascribed to the undefined mixed Ur starter during enclosed prolonged propagation without any possible external influx of bacterial or phage material. Surprisingly, during this 1000-generation long experiment the enforced conditions of specific incubation temperature and propagation regime resulted in enrichment of previously not detected strain of Lactococcus laudensis. This strain was found to consume a by-product of metabolism of another strain present in the community, in particular, D-mannitol produced by Le. mesenteroides. Thus, a new putative interaction in the microbial community of the complex starter culture was found. This new interaction and the possible ability of L. laudensis to efficiently use peptides released by caseinolytic L. lactis ssp. cremoris resulted in a relatively high abundance of L. laudensis in all evolved LUMSC cultures. The high abundance of L. laudensis had a certain effect on the functionality of the cultures. The aroma profiles of model lab-scale milli-cheeses manufactured with LUMSC cultures, showed significant differences in formation of esters and alcohols when compared to cheeses produced with the original Ur starter. Moreover, L. laudensis strain was not only under the radar of previously used culture-dependent and culture-independent methods, but as well, under the radar of phage predation continuously present throughout the LUMSC experiment. This observation sheds new light on the possibility of how a strain can emerge to relatively high abundance in an enclosed serially propagated microbial community operating in accordance with CD dynamics model.

    Finally, the aspect of adaptation to environmental conditions was addressed by the study of an adjunct strain of Lactobacillus helveticus DSM 20075 described in Chapter 5. The aim was to develop a strain with increased autolytic capacity in conditions resembling the cheese matrix to possibly improve cheese ripening. The approach used here was based on a previously reported study, where the incubation of Lactococcus lactis MG1363 at high temperature resulted in spontaneous mutations causing stable heat-resistant and, in some cases, salt-hypersensitive phenotypes. In present study, after incubation of the Lb. helveticus DSM 20075 adjunct at different high temperatures (45-50 °C), heat-sensitive variants were recovered from plates. These variants were further characterized in terms of their growth rates at elevated temperatures (42-45 °C) and their autolytic capacity in low pH buffer with addition of NaCl. One of the variants (V50) showed substantially increased intracellular lactate dehydrogenase enzyme activity in the buffer suggesting its increased autolytic capacity. Next, both wild type and variant V50 were tested as adjuncts in lab-scale model milli-cheeses to determine their possible impact on the cheese aroma profiles. Indeed, adjunct strains, both WT and the variant, impacted the aroma profiles by producing benzaldehyde. In case of the variant strain the relative abundance of this compound was 3-fold higher. The applied strategy of incubating Lb. helveticus DSM20075 at high temperature resulted in specific, but different than in case of L. lactis MG1363, mutations suggesting another, yet to be elucidated, mechanisms for increasing the autolytic capacity of industrially-relevant strains. The approach of high-temperature incubation can be applied in dairy industry for the selection of (adjunct) cultures targeted at accelerated cheese ripening and aroma formation.

    In conclusion, the work presented in this thesis highlights the importance of co-evolution of strains in compositional and functional stability of the complex undefined mixed starter culture. In particular, the factors such as heterogeneity of bacteriophage resistance among highly related strains, microbe-microbe interactions and division of metabolic labor are crucial for optimal functioning of a complex starter microbial community. Further investigation of the factors impacting the composition of starter cultures is crucial to steer the functionality in a desired direction. With straightforward methods, such as changing the incubation temperature or the propagation regime it is possible to induce shifts in strain composition and thereby obtain cultures with new characteristics. Moreover, experimental evolution studies with microbial communities used in food fermentation can lead to the discovery of new strains with potentially new characteristics. Additionally, the study of microbial communities of starter cultures not only delivers industrially applicable knowledge but also reveals the action of basic principles in microbial ecology and evolution.

    How NaCl and water content determine water activity during ripening of Gouda cheese, and the predicted effect on inhibition of Listeria monocytogenes
    Wemmenhove, E. ; Wells-Bennik, M.H.J. ; Stara, A. ; Hooijdonk, A.C.M. van; Zwietering, M.H. - \ 2016
    Journal of Dairy Science 99 (2016)7. - ISSN 0022-0302 - p. 5192 - 5201.
    Gouda - Listeria monocytogenes - Salt content - Water activity

    This study describes the diffusion of NaCl and water in Gouda cheese during brining and ripening. Furthermore, we established water activity as a function of the NaCl-in-moisture content in Gouda cheese during ripening. We determined NaCl content, water content, and water activity in block-type Gouda cheeses that were brined for 3.8 d and foil-ripened for a period of 26 wk, and in wheel-type Gouda cheeses that were brined for 0.33, 2.1, or 8.9 d and subsequently nature-ripened for a period of 26 wk. The calculated diffusion coefficients of NaCl during brining were 3.6·10-10 m2s-1 in the block-type Gouda cheeses and 3.5·10-10 m2s-1 in the wheel-type Gouda cheeses. Immediately after brining, gradients of NaCl and water were observed throughout both types of cheese. During ripening, these gradients disappeared, except for the water gradient in nature-ripened cheeses. An empirical model was derived for Gouda cheese, in which water activity is expressed as a function of the NaCl-in-moisture content, as established for different brining times, locations and ripening times. Moreover, the effect of reduced water activity on inhibition of growth of Listeria monocytogenes in Gouda cheese was calculated. In addition to the presence of lactate and a pH of 5.2 to 5.3, the reduced water activity as seen in Gouda cheese can substantially contribute to inhibition of microbial growth and even to inactivation when cheeses are brined and ripened for extended times and subjected to nature-ripening.

    Use of propidium monoazide for selective profiling of viable microbial cells during Gouda cheese ripening
    Erkus, Oylum ; Jager, Victor C.L. de; Geene, Renske T.C.M. ; Alen-Boerrigter, Ingrid van; Hazelwood, Lucie ; Hijum, Sacha A.F.T. van; Kleerebezem, Michiel ; Smid, Eddy J. - \ 2016
    International Journal of Food Microbiology 228 (2016). - ISSN 0168-1605 - p. 1 - 9.
    Cheese - Dairy - Food fermentation - Metagenomics - Microbial community profiling - Propidium monoazide

    DNA based microbial community profiling of food samples is confounded by the presence of DNA derived from membrane compromised (dead or injured) cells. Selective amplification of DNA from viable (intact) fraction of the community by propidium monoazide (PMA) treatment could circumvent this problem. Gouda cheese manufacturing is a proper model to evaluate the use of PMA for selective detection of intact cells since large fraction of membrane compromised cells emerges as a background in the cheese matrix during ripening. In this study, the effect of PMA on cheese community profiles was evaluated throughout manufacturing and ripening using quantitative PCR (qPCR). PMA effectively inhibited the amplification of DNA derived from membrane compromised cells and enhanced the analysis of the intact fraction residing in the cheese samples. Furthermore, a two-step protocol, which involves whole genome amplification (WGA) to enrich the DNA not modified with PMA and subsequent sequencing, was developed for the selective metagenome sequencing of viable fraction in the Gouda cheese microbial community. The metagenome profile of PMA treated cheese sample reflected the viable community profile at that time point in the cheese manufacturing.

    Minimal inhibitory concentrations of undissociated lactic, acetic, citric and propionic acid for Listeria monocytogenes under conditions relevant to cheese
    Wemmenhove, Ellen ; Valenberg, Hein J.F. van; Zwietering, Marcel H. ; Hooijdonk, Toon C.M. van; Wells-Bennik, Marjon H.J. - \ 2016
    Food Microbiology 58 (2016). - ISSN 0740-0020 - p. 63 - 67.
    Growth inhibition - PH - Salt - Temperature - Undissociated organic acid

    Minimal inhibitory concentrations (MICs) of undissociated lactic acid were determined for six different Listeria monocytogenes strains at 30 °C and in a pH range of 4.2-5.8. Small increments in pH and acid concentrations were used to accurately establish the growth/no growth limits of L. monocytogenes for these acids. The MICs of undissociated lactic acid in the pH range of 5.2-5.8 were generally higher than at pH 4.6 for the different L. monocytogenes strains. The average MIC of undissociated lactic acid was 5.0 (SD 1.5) mM in the pH range 5.2-5.6, which is relevant to Gouda cheese. Significant differences in MICs of undissociated lactic acid were found between strains of L. monocytogenes at a given pH, with a maximum observed level of 9.0 mM. Variations in MICs were mostly due to strain variation. In the pH range 5.2-5.6, the MICs of undissociated lactic acid were not significantly different at 12 °C and 30 °C. The average MICs of undissociated acetic acid, citric acid, and propionic acid were 19.0 (SD 6.5) mM, 3.8 (SD 0.9) mM, and 11.0 (SD 6.3) mM, respectively, for the six L. monocytogenes strains tested in the pH range 5.2-5.6. Variations in MICs of these organic acids for L. monocytogenes were also mostly due to strain variation. The generated data contribute to improved predictions of growth/no growth of L. monocytogenes in cheese and other foods containing these organic acids.

    Strain diversity and phage resistance in complex dairy starter cultures
    Spus, M. ; Alexeeva, S.V. ; Wolkers-Rooijackers, J.C.M. ; Zwietering, M.H. ; Abee, T. ; Smid, E.J. - \ 2015
    Journal of Dairy Science 98 (2015)8. - ISSN 0022-0302 - p. 5173 - 5182.
    streptococcus-cremoris - lactic streptococci - lactococcus-lactis - food fermentations - bacteriophages - competition - products - genomics - bacteria - plasmid
    The compositional stability of the complex Gouda cheese starter culture Ur is thought to be influenced by diversity in phage resistance of highly related strains that co-exist together with bacteriophages. To analyze the role of bacteriophages in maintaining culture diversity at the level of genetic lineages, simple blends of Lactococcus lactis strains were made and subsequently propagated for 152 generations in the absence and presence of selected bacteriophages. We first screened 102 single-colony isolates (strains) from the complex cheese starter for resistance to bacteriophages isolated from this starter. The collection of isolates represents all lactococcal genetic lineages present in the culture. Large differences were found in bacteriophage resistance among strains belonging to the same genetic lineage and among strains from different lineages. The blends of strains were designed such that 3 genetic lineages were represented by strains with different levels of phage resistance. The relative abundance of the lineages in blends with phages was not stable throughout propagation, leading to continuous changes in composition up to 152 generations. The individual resistance of strains to phage predation was confirmed as one of the factors influencing starter culture diversity. Furthermore, loss of proteolytic activity of initially proteolytic strains was found. Reconstituted blends with only 4 strains with a variable degree of phage resistance showed complex behavior during prolonged propagation. Key words: starter culture; bacteriophage; diversity; proteolytic activity
    Metabolic modeling of a single Leuconostoc mesenteroides and seven Lactococcus lactis strains identifies the possible metabolic dependencies within a complex bacterial starter culture
    Wels, M.W.W. ; Smid, E.J. ; Groof, B.M.J. de; Spus, M. - \ 2014
    In: Proceedings of the 11th International Symposium on Lactic Acid Bacteria. - - p. E051 - E051.
    The use of starter cultures is an essential aspect in cheese manufacturing. Starter cultures may consist of single strains or mixtures of different strains and species of mainly lactic acid bacteria. An undefined mixture of different strains and species can be regarded as a complex ecosystem. Starter culture “Ur”, used for the production of Gouda cheese is an example of a complex starter culture and it contains 7 genetic lineages of Lactococcus lactis and 1 of Leuconostoc mesenteroides [1]. Complex ecosystems are known for their stability in composition, even during the process of back-slopping, where new milk is inoculated with whey from the previous batch. We set out to investigate the compositional stability of starter culture Ur during propagation at different temperatures. In addition, Ur was reconstituted from the 8 defined genetic lineages in 1:1 starting ratio. Propagation was carried out in fat free milk and the inoculated milk was incubated at 20°C, 25°C and 30°C. These temperatures were selected to possibly favor strains that have different optimum growth temperatures. After 28 transfers (~200 generations) the composition of Ur and the reconstituted mixtures was determined using qPCR with lineage specific primers. Sequential propagation of Ur in milk at different temperatures induced changes in the relative abundance of different genomic lineages, but none of the lineages were lost. We also demonstrated that reconstituted Ur, in comparison with original Ur, leads to a similar end-point composition after 28 transfers as Ur. This demonstrates the robustness of the original Ur culture and suggests interactions between the different lineages. 1. Erkus O, de Jager VC, Spus M, van Alen-Boerrigter IJ, van Rijswijck IM, Hazelwood L, Janssen PW, van Hijum SA, Kleerebezem M, Smid EJ: Multifactorial diversity sustains microbial community stability. ISME J 2013, 7:2126-2136.
    Aeration reduces stress in Lactococcus lactis MG1363
    Brandsma, J.B. ; Wegkamp, A. ; Abee, T. ; Zwietering, M.H. ; Meijer, W. - \ 2014
    In: Book of abstracts 11th International Symposium on Lactic Acid Bacteria. - - p. E089 - E089.
    The use of starter cultures is an essential aspect in cheese manufacturing. Starter cultures may consist of single strains or mixtures of different strains and species of mainly lactic acid bacteria. An undefined mixture of different strains and species can be regarded as a complex ecosystem. Starter culture “Ur”, used for the production of Gouda cheese is an example of a complex starter culture and it contains 7 genetic lineages of Lactococcus lactis and 1 of Leuconostoc mesenteroides [1]. Complex ecosystems are known for their stability in composition, even during the process of back-slopping, where new milk is inoculated with whey from the previous batch. We set out to investigate the compositional stability of starter culture Ur during propagation at different temperatures. In addition, Ur was reconstituted from the 8 defined genetic lineages in 1:1 starting ratio. Propagation was carried out in fat free milk and the inoculated milk was incubated at 20°C, 25°C and 30°C. These temperatures were selected to possibly favor strains that have different optimum growth temperatures. After 28 transfers (~200 generations) the composition of Ur and the reconstituted mixtures was determined using qPCR with lineage specific primers. Sequential propagation of Ur in milk at different temperatures induced changes in the relative abundance of different genomic lineages, but none of the lineages were lost. We also demonstrated that reconstituted Ur, in comparison with original Ur, leads to a similar end-point composition after 28 transfers as Ur. This demonstrates the robustness of the original Ur culture and suggests interactions between the different lineages. 1. Erkus O, de Jager VC, Spus M, van Alen-Boerrigter IJ, van Rijswijck IM, Hazelwood L, Janssen PW, van Hijum SA, Kleerebezem M, Smid EJ: Multifactorial diversity sustains microbial community stability. ISME J 2013, 7:2126-2136.
    Engineering functionality of complex starter cultures
    Wolkers-Rooijackers, J.C.M. ; Groof, B.M.J. de; Spus, M. ; Smid, E.J. - \ 2014
    In: Abstract book 11th International Symposium on Lactic Acid Bacteria. - - p. E047 - E047.
    The use of starter cultures is an essential aspect in cheese manufacturing. Starter cultures may consist of single strains or mixtures of different strains and species of mainly lactic acid bacteria. An undefined mixture of different strains and species can be regarded as a complex ecosystem. Starter culture “Ur”, used for the production of Gouda cheese is an example of a complex starter culture and it contains 7 genetic lineages of Lactococcus lactis and 1 of Leuconostoc mesenteroides [1]. Complex ecosystems are known for their stability in composition, even during the process of back-slopping, where new milk is inoculated with whey from the previous batch. We set out to investigate the compositional stability of starter culture Ur during propagation at different temperatures. In addition, Ur was reconstituted from the 8 defined genetic lineages in 1:1 starting ratio. Propagation was carried out in fat free milk and the inoculated milk was incubated at 20°C, 25°C and 30°C. These temperatures were selected to possibly favor strains that have different optimum growth temperatures. After 28 transfers (~200 generations) the composition of Ur and the reconstituted mixtures was determined using qPCR with lineage specific primers. Sequential propagation of Ur in milk at different temperatures induced changes in the relative abundance of different genomic lineages, but none of the lineages were lost. We also demonstrated that reconstituted Ur, in comparison with original Ur, leads to a similar end-point composition after 28 transfers as Ur. This demonstrates the robustness of the original Ur culture and suggests interactions between the different lineages. 1. Erkus O, de Jager VC, Spus M, van Alen-Boerrigter IJ, van Rijswijck IM, Hazelwood L, Janssen PW, van Hijum SA, Kleerebezem M, Smid EJ: Multifactorial diversity sustains microbial community stability. ISME J 2013, 7:2126-2136.
    Kolonist in museum
    Jansman, Hugh - \ 2014
    Het stoffelijk overschot van de otter die werd doorgereden op de A12 bij Gouda gaat naar onderzoeksinstituut en natuurhistorisch museum Naturalis in Leiden.
    Otter was beresterk mannetje
    Jansman, Hugh - \ 2014
    De op 12 februari op de A12 bij Gouda doodgereden otter lijkt in Nieuwkoop gebivakkeerd te hebben. Dat zegt Hugh Jansman van het Animal Ecology Team van de onderzoeksgroep Alterra in Wageningen, waar het stofferlijk overschot van het dier werd ontleed en onderzocht. De onfortuinlijke otter blijkt een volwassen mannetje van anderhalf jaar oud.
    The fate of Listeria monocytogenes in and on Gouda cheese following artificial contamination during brining and artificial contamination of pasteurized cheese milk
    Wemmenhove, E. ; Stampelou, I. ; Beumer, R.R. ; Hooijdonk, A.C.M. van; Zwietering, M.H. ; Wells-Bennik, M.H.J. - \ 2014
    In: Abstract book ICFMH Conference Food Micro. - - p. 457 - 457.
    The fate of Listeria monocytogenes in brine and on Gouda cheese following artificial contamination during brining
    Wemmenhove, E. ; Beumer, R.R. ; Hooijdonk, A.C.M. van; Zwietering, M.H. ; Wells-Bennik, M.H.J. - \ 2014
    International Dairy Journal 39 (2014)2. - ISSN 0958-6946 - p. 253 - 258.
    inoculum size - growth - survival - temperature - pathogen - limits - milk - ph
    The fate of 3 different Listeria monocytogenes strains (Scott A, 2F and 6E) was studied independently in brine and on factory-scale Gouda cheeses that had been submerged in brine that was artificially contaminated with these individual strains. Viable numbers of L. monocytogenes in the brine decreased during brining (0, 1, 2.9 and 8.9 d). L. monocytogenes was enumerated on the surface of Gouda cheese directly after brining and over 26 weeks of ripening at 12.5 degrees C. Transfer of L. monocytogenes from brine to cheese during brining was limited. L. monocytogenes was detected in the outer layer of Gouda cheese but not in the centre directly after brining or during ripening. Throughout the ripening period, the viable numbers of L. monocytogenes declined significantly. This study adds to the understanding of the fate of L. monocytogenes in brine and on Gouda cheese, and demonstrates that growth of L. monocytogenes on Gouda cheese is not supported following contamination during brining.
    Community dynamics of complex starter cultures for Gouda-type cheeses and its functional consequences
    Erkus, O. - \ 2014
    Wageningen University. Promotor(en): Michiel Kleerebezem, co-promotor(en): Eddy Smid. - Wageningen UR : Wageningen - ISBN 9789462570108 - 215
    goudse kaas - lactococcus lactis - microbiële diversiteit - gouda cheese - lactococcus lactis - microbial diversity

    Lactic acid bacteria (LAB) are used as starter and adjunct cultures for the production of artisanal and industrial fermented milk products such as yoghurt and cheese. Artisanal fermentations is propagated with the transfer of an inoculum from old batch of fermented food to the new batch (back-slopping) to initiate the fermentation with the activity of the indigenous microbiota present in the inoculum. In industrial production, these inocula with indigenous microbiota are replaced with the starter cultures that contain lower numbers of LAB species for better controlled fermentation process and consistent final product quality. Cheese manufacturing is still performed in both artisanal ways and with the use of starter cultures. Gouda cheese starter cultures constitute several strains from the subspecies of Lactococcus lactisand Leuconostocs mesenteroidesin different combinations. The mixed and undefined type of starter culture may harbour variable number of strains that contribute unique functionalities to the cheese manufacturing process. Therefore, understanding, controlling and predicting the cheese manufacturing processes require the determination of strain level diversity in the starter culture, their collective and specific metabolic complement, and their activity throughout the cheese manufacturing process, including the interactions between the strains. The first two studies that are covered in this thesis describes the development of a high resolution AFLP fingerprinting tool allowing the discrimination of closely related strains in the starter culture and the subsequent analysis of the microbial community of Gouda cheese starter with this implemented technique and with metagenomics. Furthermore, the thesis includes the development of another tool to selectively amplify DNA only from live fraction of the microbial community in cheese using propidium monoazide (PMA), which is required to study community dynamics with culture independent approaches. The last study in the thesis describes the effects of the variation in propagation regime on the community composition of a mixed starter culture and connects the composition change to the functionalities that impact on flavour development during cheese manufacturing. Overall, the approaches presented in this thesis are intended to eventually enable accurate prediction and control of the cheese manufacturing process using (un)defined starter cultures, but may also allow rational design and development of new starter cultures.

    Stories becoming sticky : how civic initiatives strive for connection to governmental spatial planning agendas
    Stoep, H. van der - \ 2014
    Wageningen University. Promotor(en): Adri van den Brink; Noelle Aarts. - Wageningen : Wageningen University - ISBN 9789461738295 - 282
    ruimtelijke ordening - publieke participatie - maatschappelijk middenveld - bestuurskunde - besluitvorming - maatschappelijke betrokkenheid - regionale planning - communicatie - verhoudingen tussen bevolking en staat - burgers - participatie - zuid-holland - krimpenerwaard - zuid-limburg - physical planning - public participation - civil society - public administration - decision making - community involvement - regional planning - communication - relations between people and state - citizens - participation - zuid-holland - krimpenerwaard - zuid-limburg

    This thesis aims to understand the phenomenon of self-organizing civic initiatives, how they engage in and connect to planning practices aimed at the improvement of the quality of places and why these connections lead to alteration or transformation of governmental planning agendas or not. By providing greater understanding about these processes the thesis aims to contribute to debates about how planners can improve connections with civil society initiatives and how a more responsive and adaptive attitude towards a dynamically changing society can be achieved.

    Conclusions were drawn from two in-depth case-studies of civic initiatives in two Dutch regions: 1) initiatives of business entrepreneurs and experts to develop New Markets which support the cultural landscape of the region Heuvelland, and 2) initiatives of citizens for the protection and development of landscape values in the urban-rural fringe Gouda-Krimpenerwaard. Building on agenda-setting and framing theory the analysis focused on how initiatives self-organized and connected to other stakeholders and how outcomes of their efforts in terms of their ambitions and government agendas could be understood.

    The research results point to the crucial role of storytelling and the day-to-day interactions in which stories emerge and become ‘sticky’. Sticky stories are strong ‘attractors’ that mobilize attention and support. The stickiness of a story was enhanced through discursive processes in which the story was connected to the self-referential frames of targeted supporters. Thus, sticky stories could not emerge without empathic listening, timing and patience. Three interplaying conditions were critical in the process of a story becoming sticky or fading away: 1) formal and informal ‘catalytic’ conversations as the medium of storytelling; 2) storytelling by people who perform as connectors and enable the travelling of stories through a wider network, and 3) signalling and incorporating focusing events into evolving stories in ways considered meaningful and relevant by targeted supporters. This results in a model that offers a way to understand dynamical change of policy and planning agendas by focusing on the interactive construction, connection, and subtle alteration of stories in day-to-day conversations, by the right people, at the right moments.

    Fate of Listeria monocytogenes in Gouda microcheese: No growth, and substantial inactivation after extended ripening times
    Wemmenhove, E. ; Stampelou, I. ; Hooijdonk, A.C.M. van; Zwietering, M.H. ; Wells-Bennik, M.H.J. - \ 2013
    International Dairy Journal 32 (2013)2. - ISSN 0958-6946 - p. 192 - 198.
    cheddar cheese - inoculum size - lactic-acid - behavior - manufacture - inhibition - survival - storage - milk - ph
    This challenge study demonstrates that Listeria monocytogenes does not grow in Gouda cheese: during the first 8 weeks of ripening no growth was observed and between 8 and 52 weeks viable numbers declined significantly in a well-established Gouda microcheese system. Cheese milk was artificially contaminated just prior to addition of the starter culture. Three individual L. monocytogenes strains were used, including strains originating from cheese, a cheese plant environment and a reference strain. During curd formation, viable numbers of L. monocytogenes increased by 0.5 log cfu g-1, resulting from entrapment in the curd. No growth was observed during the first 8 weeks of ripening. A significant decline in the viable numbers of L. monocytogenes was observed in Gouda cheese that was ripened for longer than 8 weeks. Two factors that could possibly control the fate of L. monocytogenes in Gouda cheese were lactic acid and water activity.
    Kasteelt (innovaties in de glastuinbouw, met Tycho Vermeulen en Leo Ammerlaan)
    Muusze, B. ; Vermeulen, T. ; Ammerlaan, L. - \ 2013
    Omgevingsdienst Midden-Holland
    glastuinbouw - energiebesparing - belichting - innovaties - algenteelt - warmte - duurzaamheid (sustainability) - biobased economy - greenhouse horticulture - energy saving - illumination - innovations - algae culture - heat - sustainability
    De Stichting Duurzame Stad organiseert in Zoetermeer en Gouda een ontwerpwedstrijd ‘duurzame stad van de toekomst’ voor leerlingen tussen de 14 en 18 jaar. Drie middelbare scholen uit Zoetermeer en drie uit Gouda doen aan mee aan de wedstrijd. De leerlingen kunnen als hoofdprijs zonnepanelen voor op hun school winnen. Ter inspiratie voor de wedstrijd bekijken leerlingen korte filmpjes over hoe verschillende sectoren in de maatschappij werken aan verduurzaming, om vervolgens zelf aan de slag te gaan. Duurzame Voedselvoorziening is één van de oDe Stichting Duurzame Stad organiseert in Zoetermeer en Gouda een ontwerpwedstrijd ‘duurzame stad van de toekomst’ voor leerlingen tussen de 14 en 18 jaar. Drie middelbare scholen uit Zoetermeer en drie uit Gouda doen aan mee aan de wedstrijd. De leerlingen kunnen als hoofdprijs zonnepanelen voor op hun school winnen. Ter inspiratie voor de wedstrijd bekijken leerlingen korte filmpjes over hoe verschillende sectoren in de maatschappij werken aan verduurzaming, om vervolgens zelf aan de slag te gaan. Duurzame Voedselvoorziening is één van de onderwerpen. In de voedselvoorziening speelt de glastuinbouw in Nederland een belangrijke rol. De Stichting Duurzame Stad heeft daarom samen met de Omgevingsdienst Midden-Holland, de gemeente Zoetermeer en Wageningen UR Glastuinbouw een inspiratiefilmpje gemaakt. Het filmpje toont de leerlingen interessante ontwikkelingen die op dit moment plaatsvinden in kassen. Vooral de energie-aspecten van de glastuinbouw komen aan bod: . de opkomst van het ledlicht . opwekken van warmte en energie uit kassen . algenteelt en bioplastics . het verbinden van de tuinbouw met de stad Voor het laatste onderwerp zijn opnames gemaakt bij potplantenkwekerij Ammerlaan The Green Innovator in Pijnacker-Nootdorp. Finales van de ontwerpwedstrijd zijn in Zoetermeer op 24 april en in Gouda op 30 mei 2013.
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