Reports

Research on thermophilic microorganisms in high-temperature areas in Reykjanes, Hengill area and Fremrinámur. Prepared for Expert Group 1 in Framework Plan 3 / Thermophilic microorganisms from geothermal areas at Reykjanes, Hengill and Fremrinámar.

Published:

30/05/2016

Authors:

Edda Olgudóttir, Sólveig K. Pétursdóttir

Supported by:

Framework Program 3 (RÁ3)

Research on thermophilic microorganisms in high-temperature areas in Reykjanes, Hengill area and Fremrinámur. Prepared for Expert Group 1 in Framework Plan 3 / Thermophilic microorganisms from geothermal areas at Reykjanes, Hengill and Fremrinámar.

The current study was carried out under the auspices of Framework Plan 3 and included high-temperature areas that declined in the waiting category in RÁ2. The areas were Trölladyngja and Austurengjahver in Reykjanes, Fremrinámar and Þverárdalur and Innstidalur in the Hengill area. The aim of the study was to assess the diversity and rarity of thermophilic microorganisms in the above areas. The methodology was based on DNA analysis independent of culture. DNA was isolated from samples and species-identifying genes amplified and sequenced and the sequences compared to sequences in gene banks and from previous studies. A total of 118 samples were collected in 2015 and were able to be sequenced approx. Their 59%. A total of 10 million 16S gene sequences were obtained from sequencing, which dropped to almost six million after all quality and length conditions had been met. Most rows were obtained from Innstadalur, or 2,176,174, but fewest 286,039 from Trölladyngja. The diversity of microorganisms in each region was assessed by comparing the number of arrays, number of species, collection curves, and Shannon's diversity coefficient. The total number of samples and sequences of each area was very different and reflect its diversity. When the series were classified as species with 97% matching, it was found that most species came from Þverárdalur, or about 42 thousand, but the fewest from Trölladyngja, or about 9 thousand. More than 12 thousand species were found in samples from Fremrinámur, which came as a surprise as the area and the samples seemed homogeneous and such diversity was not expected. All major thermal bacteria were found in the samples, both Ancient Bacteria and Bacteria. Special groups were found especially within the Thaumarchaeota faction. The assessment of diversity in individual areas with collection curves and diversity coefficient was based on the smallest number of rows or 286,039 from Trölladyngja. The greatest diversity of species was in Þverárdalur and Innstadalur and the least at Trölladyngja and Austurengjahver, but Fremrinámar was in between. Collection curves gave the same result. Biodiversity (H) gave a different ranking. There were Þverárdalur and Fremrinámar with the most diversity (H = 8 and 7.7), then Innstidalur but Trölladyngja and Austurengjahver (H = 6) ran the train. Rarity was assessed on the basis of the number and proportion of unknown species in comparison with the Silva database. The number of unknown species was greatest in Þverárdalur and Innstadalur, both with over 1000, Fremrinámar with 756 and Trölladyngja and Austurengjahver with over four hundred unknown species. National rarity was assessed by comparing data from individual regions in previous studies and examining whether a match occurred. It turned out that the geothermal area in Fremrinámur contains a large number of species that have no equivalent in other thermal areas.

The current project was requested by the Master Plan for Nature Protection and Energy Utilization and aimed at geothermal areas which had not been classified for preservation or utilization during Masterplan 2. The geothermal areas investigated were Trölladyngja and Austurengjahver at Reykjanes, Fremrinámar and Þverárdalur and Innstidalur at Hengill. The goal of the project was to estimate biodiversity and rarity of thermophiles inhabiting the areas mentioned. The methods used were DNA based and were performed on DNA extracted from primary samples (culture independent). Microbial species identification was performed by amplification and sequencing of 16S rRNA genes and comparison with sequence databases and previous research. A total of 118 samples were collected in 2015 of which 59% were sequenced. The total sequencing yield was 10 million reads, of which 6 million passed quality assessment and were used for downstream analyzes. The largest proportion of the reads were obtained from Innstidalur samples, 2,176,174 reads, and the lowest proportion from Trölladyngja, 286,036 reads. The biodiversity of microorganisms within each area was estimated using the number of phyla and species, rarefaction curves and Shannons' biodiversity index. The total number of species identified varied between sites and reflected the diversity of the geothermal area and the total amount of sequences obtained. Using a cut-off value of 97% similarity, the sequences were classified to the species level. The highest number of species, approximately 42.000, were identified in samples from Þverárdalur and the lowest number, approximately 9.000, in samples from Trölladyngja. Roughly 12,000 species were found in samples from Fremrinámar, which was surprising as the area and the samples appeared rather homogenous and such diversity was therefore not expected. All the main thermophilic taxa of the Bacteria and Archaea domains were identified in the samples. Unknown groups were found especially within the phylum of Thaumarchaeota. For the rarefaction and biodiversity index estimates the lowest number of sequence reads, Trölladyngja, was used as reference. The species diversity was found to be highest in Þverárdalur and Innstidalur, the lowest in Trölladyngja and Austurengjahver, and intermediate in Fremrinámar. Rarefaction curves showed the same results. The calculated biodiversity index (H) gave different results, indicating highest diversity in Þverárdalur and Fremrinámar (H = 8 and 7,7 respectively), intermediate in Innstidalur (H = 7.0) and lowest in Trölladyngja and Austurengjahver (H = 6). Rarity was estimated as the number of species which could not be identified by comparison to the Silva database. The highest number of unidentified species was roughly 1000 in Þverárdalur and Innstidalur, 756 in Fremrinámar and between 300- 400 in Trölladyngja and Austurengjahver. The rarity was also estimated by comparing data obtained in the current project with data from previous projects. The analysis revealed a particularly high number of unique species in Fremrinámar that have not been identified in other geothermal areas in Iceland.

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Reports

Bacterial diversity in the processing environment of fish products

Published:

01/03/2010

Authors:

Eyjólfur Reynisson, Sveinn Haukur Magnússon, Árni Rafn Rúnarsson, Viggó Þór Marteinsson

Supported by:

Tækniþróunarsjóður, AVS

Contact

Viggó Marteinsson

Research Group Leader

viggo.th.marteinsson@matis.is

Bacterial diversity in the processing environment of fish products

The report seeks to address the diversity and species composition of micro-organisms in fish processing environments. The research work began with the installation and development of methods for scanning microbial composition using molecular biological methods, and then at a later stage, work began on examining selected environments from the fishing industry. Two fish processing plants were visited, each twice where an evaluation was made of the processing and approx. 20 samples taken in each trip. A diverse community of bacteria was found, where known harmful bacteria were usually in a high proportion along with various other species. Microbial counts showed high levels of bacteria on the surface of production lines during processing with few bacterial groups in excess but also numerous other species in smaller quantities. The main groups of bacteria found belong to Photobacterium phosphoreum, which was in the highest proportion overall throughout the study, along with Flavobacterium, Psychrobacter, Chryseobacter, Acinetobacter and Pseudoalteromonas. All of these species are known fish bacteria that live in the redness and intestines of live fish. This is the first known project where molecular biological methods are used to scan the bacterial ecosystem of fish processing plants. A knowledge base has therefore been laid here for bacterial ecosystems in different conditions in fish processing, which will be used permanently in research and development of improved processing processes and storage methods for fish.

In this report we seek answers on diversity and species composition of bacteria in fish processing environment. The study initiated method development to screen microbial systems using molecular methods followed by analysis of samples from 2 fish processing plants. This research shows the presence of a diverse microbial community in fish processing environment where known spoilage microorganisms are typically in high relative numbers along with various other bacterial species. Total viable counts showed the presence of bacteria in high numbers on processing surfaces during fish processing where few species typically dominated the community. Photobacterium phosphoreum was the most apparent species followed by genera such as Flavobacterium, Psychrobacter, Chryseobacter, Acinetobacter and Pseudoalteromonas. All these species are known fish associated bacteria that live on the skin and in the digestive tract of a living animal. To our knowledge, this is the first study where molecular methods are used to screen microbial communities in fish processing plants. This research has therefore contributed a database on bacterial diversity in fish processing plants that will be used in the future to improve processing and storage methods in the fish industry.

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