Identifisering og karakterisering av bakteriocinet amylocyclicin fra Bacillus velezensis, og dets potensial mot Listeria monocytogenes

Stadig flere bakterier blir resistente mot antibiotika som brukes for å behandle bakterielle infeksjoner. Verdens helseorganisasjon har erklært antimikrobiell resistens som et av verdens største globale helseutfordringer som menneskeheten står ovenfor. For å kunne løse problemet er det avgjørende å finne alternativer til dagens bruk av antibiotika. En del av løsningen kan være å benytte bakteriers egenproduserte antimikrobielle peptider, også kalt bakteriociner. Bakteriociner er små ribosomalt syntetiserte peptider med stor variasjon i både struktur, aktivitet og målorganismer. Bakteriociner produseres av en rekke ulike bakterier, og syntesen av bakteriociner krever som regel minst to ekstra gener som koder for dedikerte transport- og immunitetsproteiner. Immunitet gjør at produsenten ikke blir hemmet av sitt eget bakteriocin, og på den måten utnyttes bakteriociner i konkurranse om næring og nisjer. Målet for denne oppgaven var å identifisere og karakterisere et antimikrobielt stoff produsert av en Bacillus velezensis stamme. Ved bruk av helgenomsekvensering, revers-fase kromatografi og massespektrometri ble dette stoffet identifisert til å være det sirkulære bakteriocinet amylocyclicin. Amylocyclicin var allerede beskrevet, men det var mye som ikke var kjent om peptidets antimikrobielle aktivitet, virkemåte og immunitet. Dette bakteriocinet ble derfor undersøkt videre. Amylocyclicin viste seg å være et poredannende bakteriocin med høy stabilitet mot varme- og pH-forandringer, og hadde et bredt inhiberingsspekter mot nesten alle Gram-positive bakterier. Det ble observert spesielt god aktivitet mot den patogene bakterien Listeria monocytogenes, en robust bakterie som kan forårsake den alvorlige sykdommen listeriose. L. monocytogenes kan også danne biofilm, som er et problem i mat- og helseindustrien. Flere ulike matprodukter, som røkt laks, har tidligere vært smittekilde for denne bakterien. Amylocyclicin var effektiv mot biofilm produsert av L. monocytogenes, og konfokal mikroskopi viste at bakteriocinet reduserte både celletettheten og levende celler i biofilmen. I tillegg var amylocyclicin effektiv som preventiv behandling for å hemme vekst av L. monocytogenes inokulert på røkt laks. Denne oppgaven viser at amylocyclicin har flere egenskaper som gjør at det kan være egnet for videre utvikling til bruk i mat- og helseindustrien. Videre forskning anbefales derfor på amylocyclicin og bakteriociner generelt, spesielt med hensyn til deres virkemåte og toksisitet, før de eventuelt kan utnyttes i kampen mot antibiotikaresistens.An increasingly number of bacteria are becoming resistant to antibiotics commonly used to treat infections. The World Health Organization has declared antimicrobial resistance as one of the greatest public threats facing humanity. To solve this problem, it is crucial to find alternatives to how antibiotics are presently used. A part of the solution could be the use of the antimicrobial peptides produced by the bacteria themselves, molecules also known as bacteriocins. Bacteriocins are small ribosomally synthesized peptides that vary considerably in structures, activity, and target organisms. Bacteriocins are produced by many different bacteria, and their biosynthesis most often requires two extra genes encoding dedicated transport and immunity proteins. Immunity enables the producing organisms to inhibit other bacteria in competition for nutrients and niches. The aim of this thesis was to identify and characterize an antimicrobial substance produced by a Bacillus velezensis strains. By the use of whole-genome sequencing, reversed-phase chromatography and mass spectrometry, the antimicrobial substance was identified as the circular bacteriocin amylocyclicin. Amylocyclicin have been previously described, but little is known about its antimicrobial activity, mode of action, and immunity. Amylocyclicin was shown to have a mode of action involving pore-formation and to have high thermal and pH-stability. Additionally, amylocyclicin exhibited broad-spectrum activity towards most Gram-positive bacteria. The activity was particularly potent against Listeria monocytogenes, a robust pathogen that causes the serious illness listeriosis. L. monocytogenes is also a strong biofilm producer, which is a problem in the food and health industry. Several ready-to-eat food products, such as smoked salmon, have been the source of numerous listeriosis outbreaks. Amylocyclicin was effective at disrupting biofilms produced by L. monocytogenes, and confocal microscopy revealed that the amylocyclicin treatment reduced the cell density and living cells in the biofilm. Additionally, pre-treatment of smoked salmon with amylocyclicin was shown to effectively inhibit the growth of L. monocytogenes. This thesis demonstrates that amylocyclicin has potential for applied use in the food and health industry. Further research on amylocyclicin and bacteriocins in general is thus needed, particularly regarding their mode of action and toxicity, prior to their effective use as a supplement or alternative to antibiotics

Evaluation of Road Safety Education Program with Virtual Reality Eye Tracking

Traffic rules are essential and part of today’s road safety education programs at school. However, learning how to properly use attention in complex traffic situations has never before been a focus when teaching pupils. A new road safety education program based on latest findings in Neuro-Education has been developed. In the program, pupil learned three new concepts: risk, orientation and attention. This to stimulate the pupil’s reflection about own behavior on school roads, before attended a session at a traffic center, where teachers encouraged pupil’s reflection about how to practically use their new knowledge about the three concepts. This new program design was evaluated in a Virtual Reality laboratory. This paper presents the approach for evaluation using VR and eye-tracking, including scenario building, measurement approach, and procedure for measurements in addition to a look at the actual evaluation results. The results showed that pupils managed to cycle in a safer way by putting their attention at the right places after the course compared to student following existing curriculum. As such the selected validation approach utilizing VR gave clear support for the proposed education program.publishedVersio

Iodine content in bulk biomass of wild-harvested and cultivated edible seaweeds: Inherent variations determine species-specific daily allowable consumption

This study represents a large-scale investigation into iodine contents in three commercially important and edible seaweed species from the North Atlantic: the brown algae Saccharina latissima and Alaria esculenta, and the red alga Palmaria palmata. Variability among and within species were explored in terms of temporal and spatial variations in addition to biomass source. Mean iodine concentration in bulk seaweed biomass was speciesspecific: Saccharina > Alaria > Palmaria. Iodine contents of Saccharina biomass were similar between years and seasons, but varied significantly between sampling locations and biomass sources. In Alaria and Palmaria, none of the independent variables examined contributed significantly to the small variations observed. Our data suggest that all three species are rich sources of iodine, and only 32, 283, or 2149 mg dry weight of unprocessed dry biomass of Saccharina, Alaria, or Palmaria, respectively, meets the recommended daily intake levels for most healthy humans.publishedVersio

NordAqua, a Nordic Center of Excellence to develop an algae-based photosynthetic production platform

NordAqua is a multidisciplinary Nordic Center of Excellence funded by NordForsk Bioeconomy program (2017-2022). The research center promotes Blue Bioeconomy and endeavours to reform the use of natural resources in a environmentally sustainable way. In this short communication, we summarize particular outcomes of the consortium. The key research progress of NordAqua includes (1) improving of photosynthetisis, (2) developing novel photosynthetic cell factories that function in a "solar-driven direct CO2 capture to target bioproducts" mode, (3) promoting the diversity of Nordic cyanobacteria and algae as an abundant and resilient alternative for less sustainable forest biomass and for innovative production of biochemicals, and (4) improving the bio-based wastewater purification and nutrient recycling technologies to provide new tools for integrative circular economy platforms.Peer reviewe

The Unique Lipidomic Signatures of Saccharina latissima Can Be Used to Pinpoint Their Geographic Origin

The aquaculture of macroalgae for human consumption and other high-end applications is experiencing unprecedented development in European countries, with the brown algae Saccharina latissima being the flag species. However, environmental conditions in open sea culture sites are often unique, which may impact the biochemical composition of cultured macroalgae. The present study compared the elemental compositions (CHNS), fatty acid profiles, and lipidomes of S. latissima originating from three distinct locations (France, Norway, and the United Kingdom). Significant differences were found in the elemental composition, with Norwegian samples displaying twice the lipid content of the others, and significantly less protein (2.6%, while French and UK samples contained 6.3% and 9.1%, respectively). The fatty acid profiles also differed considerably, with UK samples displaying a lower content of n-3 fatty acids (21.6%), resulting in a higher n-6/n-3 ratio. Regarding the lipidomic profile, samples from France were enriched in lyso lipids, while those from Norway displayed a particular signature of phosphatidylglycerol, phosphatidylinositol, and phosphatidylcholine. Samples from the UK featured higher levels of phosphatidylethanolamine and, in general, a lower content of galactolipids. These differences highlight the influence of site-specific environmental conditions in the shaping of macroalgae biochemical phenotypes and nutritional value. It is also important to highlight that differences recorded in the lipidome of S. latissima make it possible to pinpoint specific lipid species that are likely to represent origin biomarkers. This finding is relevant for future applications in the field of geographic origin traceability and food controlpublishedVersio

Dietary b-glucan (MacroGard®) enhances survival of first feeding turbot (Scophthalmus maximus) larvae by altering immunity, metabolism and microbiota

Reflecting the natural biology of mass spawning fish aquaculture production of fish larvae is often hampered by high and unpredictable mortality rates. The present study aimed to enhance larval performance and immunity via the oral administration of an immunomodulator, β-glucan (MacroGard®) in turbot (Scophthalmus maximus). Rotifers (Brachionus plicatilis) were incubated with or without yeast β-1,3/1,6-glucan in form of MacroGard® at a concentration of 0.5 g/L. Rotifers were fed to first feeding turbot larvae once a day. From day 13 dph onwards all tanks were additionally fed untreated Artemia sp. nauplii (1 nauplius ml/L). Daily mortality was monitored and larvae were sampled at 11 and 24 dph for expression of 30 genes, microbiota analysis, trypsin activity and size measurements. Along with the feeding of β-glucan daily mortality was significantly reduced by ca. 15% and an alteration of the larval microbiota was observed. At 11 dph gene expression of trypsin and chymotrypsin was elevated in the MacroGard® fed fish, which resulted in heightened tryptic enzyme activity. No effect on genes encoding antioxidative proteins was observed, whilst the immune response was clearly modulated by β-glucan. At 11 dph complement component c3 was elevated whilst cytokines, antimicrobial peptides, toll like receptor 3 and heat shock protein 70 were not affected. At the later time point (24 dph) an anti-inflammatory effect in form of a down-regulation of hsp 70, tnf-α and il-1β was observed. We conclude that the administration of MacroGard® induced an immunomodulatory response and could be used as an effective measure to increase survival in rearing of turbot

NordAqua, a Nordic Center of Excellence to develop an algae-based photosynthetic production platform

NordAqua is a multidisciplinary Nordic Center of Excellence funded by NordForsk Bioeconomy program (2017-2022). The research center promotes Blue Bioeconomy and endeavours to reform the use of natural resources in a environmentally sustainable way. In this short communication, we summarize particular outcomes of the consortium. The key research progress of NordAqua includes (1) improving of photosynthetisis, (2) developing novel photosynthetic cell factories that function in a "solar-driven direct CO2 capture to target bioproducts" mode, (3) promoting the diversity of Nordic cyanobacteria and algae as an abundant and resilient alternative for less sustainable forest biomass and for innovative production of biochemicals, and (4) improving the bio-based wastewater purification and nutrient recycling technologies to provide new tools for integrative circular economy platforms.</p

Managing the microbial community of marine fish larvae: a holistic perspective for larviculture

The availability of high-quality juveniles is a bottleneck in the farming of many marine fish species. Detrimental larvae-microbe interactions are a main reason for poor viability and quality in larval rearing. In this review, we explore the microbial community of fish larvae from an ecological and eco-physiological perspective, with the aim to develop the knowledge basis for microbial management. The larvae are exposed to a huge number of microbes from external and internal sources in intensive aquaculture, but their relative importance depend on the rearing technology used (especially flow-through vs. recirculating systems) and the retention time of the water in the fish tanks. Generally, focus has been on microbes entering the system, but microbes from growth within the system is normally a substantial part of the microbes encountered by larvae. Culture independent methods have revealed an unexpected high richness of bacterial species associated with larvae, with 100–250 operational taxonomic units associated with one individual. The microbiota of larvae changes rapidly until metamorphosis, most likely due to changes in the selection pressure in the digestive tract caused by changes in host-microbe and microbe-microbe interactions. Even though the microbiota of larvae is distinctly different from the microbiota of the water and the live food, the microbiota of the water strongly affects the microbiota of the larvae. We are in the early phase of understanding larvae-microbe interactions in vivo, but some studies with other animals than fish emphasize that we so far have underestimated the complexity of these interactions. We present examples demonstrating the diversity of these interactions. A large variety of microbial management methods exist, focusing on non-selective reduction of microbes, selective enhancement of microbes, and on improvement of the resistance of larvae against microbes. However, relatively few methods have been studied extensively. We believe that there is a lot to gain by increasing the diversity of approaches for microbial management. As many microbial management methods are perturbations of the microbial community, we argue that ecological theory is needed to foresee and test for longer term consequences in microbe-microbe and microbe-larvae interactions. We finally make some recommendations for future research and development

Is Europe ready for integrated multi-trophic aquaculture? A survey on the perspectives of European farmers and scientists with IMTA experience

Integrated Multi-Trophic Aquaculture (IMTA) refers to the integrated farming of several species from different trophic levels in close proximity. In IMTA, one species is intended to complement another. The concept has long been in use in Asia and contributes significantly to the sustainability of aquaculture as it can potentially drive ecological efficiency, environmentally acceptability, product-diversity, profitability and benefit society. Although the concept has received increasing academic attention during the last two decades, it has not yet become a commercial reality in European mariculture. The reasons for this were explored, by interviewing farmers and scientists with previous experience on IMTA. The interviewing approach can be effective in identifying important omissions from the available literature and also possible exaggerations of positive results. For the purposes of this study, a qualitative survey was undertaken using a structured questionnaire with open-ended questions. As a result, the opinions of 34 farmers and scientists with substantial experience of IMTA from 12 European countries have been obtained. A broad spectrum of IMTA impediments has been identified. These have been separated into nine major categories; namely Biological, Conflicts, Environmental, Interest, Legislation, Market, Operational, R&D, and Vandalism. The importance of each category was found to vary among different locations and regions of Europe indicating the need for site-specific targeted approaches. Nevertheless, factors from several categories were raised in all countries/IMTA configurations which highlights that for IMTA to be further developed and adopted, the involvement of stakeholders and personnel from several disciplines is necessary (i.e. biologists, economists, engineers, farm managers, modellers, regulators, stakeholders and statisticians). This work identifies many of the challenges that European IMTA is likely to encounter, and proposes areas that are likely to benefit from focused research and development.PostprintPeer reviewe