Potential Enterotoxicity of Phylogenetically Diverse Bacillus cereus Sensu Lato Soil Isolates from Different Geographical Locations

Bacillus cereus sensu lato comprises Gram-positive spore-forming bacteria producing toxins associated with foodborne diseases. Three pore-forming enterotoxins, nonhemolytic enterotoxin (Nhe), hemolysin BL (Hbl), and cytotoxin K (CytK), are considered the primary factors in B. cereus sensu lato diarrhea. The aim of this study was to determine the potential risk of enterotoxicity among soil B. cereus sensu lato isolates representing diverse phylogroups and originated from different geographic locations with various climates (Burkina Faso, Kenya, Argentina, Kazakhstan, and Poland). While nheA- and hblA-positive isolates were present among all B. cereus sensu lato populations and distributed across all phylogenetic groups, cytK-2-positive strains predominated in geographic regions with an arid hot climate (Africa) and clustered together on a phylogenetic tree mainly within mesophilic groups III and IV. The highest in vitro cytotoxicity to Caco-2 and HeLa cells was demonstrated by the strains clustered within phylogroups II and IV. Overall, our results suggest that B. cereus sensu lato pathogenicity is a comprehensive process conditioned by many intracellular factors and diverse environmental conditions.Izabela Święcicka - [email protected] Drewnowska - Department of Microbiology, Faculty of Biology, University of Bialystok, Bialystok, PolandNatalia Stefańska - Department of Microbiology, Faculty of Biology, University of Bialystok, Bialystok, PolandMagdalena Czerniecka - Department of Cytobiochemistry, Faculty of Biology, University of Bialystok, Bialystok, Poland; Laboratory of Tissue Culture, Faculty of Biology, University of Bialystok, Bialystok, PolandGrzegorz Zambrowski - Laboratory of Applied Microbiology, University of Bialystok, Bialystok, PolandIzabela Święcicka - Department of Microbiology, Faculty of Biology, University of Bialystok, Bialystok, Poland; Laboratory of Applied Microbiology, University of Bialystok, Bialystok, PolandMock M, Fouet A. 2001. Anthrax. 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Characterization of emetic Bacillus weihenstephanensis, a new cereulide-producing bacterium. Appl Environ Microbiol 72:5118 –5121.https://doi.org/10.1128/AEM.00170-06.Jung MY, Kim JS, Paek WK, Lim J, Lee H, Kim PI, Ma JY, Kim W, Chan YH. 2011. Bacillus manliponensis sp. nov., a new member of the Bacillus cereus group isolated from foreshore tidal flat sediment. J Microbiol 49:1027–1032. https://doi.org/10.1007/s12275-011-1049-6.Jung MY, Paek WK, Park IS, Han JR, Sin Y, Paek J, Rhee MS, Kim H, Song HS, Chang YH. 2010. Bacillus gaemokensis sp. nov., isolated from foreshore tidal flat sediment from the Yellow Sea. J Microbiol 48:867– 871. https://doi.org/10.1007/s12275-010-0148-0.Cheng T, Lin P, Jin S, Wu Y, Fu B, Long R, Liu D, Guo Y, Peng L, Xia Q. 2014. Complete genome sequence of Bacillus bombysepticus, a pathogen leading to Bombyx mori Black Chest Septicemia. 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A new cytotoxin from Bacillus cereus that may cause necrotic enteritis. Mol Microbiol 38:254 –261. https://doi.org/10.1046/j.1365-2958.2000.02147.xFagerlund A, Ween O, Lund T, Hardy SP, Granum PE. 2004. Genetic and functional analysis of the cytK family of genes in Bacillus cereus. Microbiology 150:2689 –2697. https://doi.org/10.1099/mic.0.26975-0Gohar M, Faegri K, Perchat S, Ravnum S, Økstad OA, Gominet M, Kolstø AB, Lereclus D. 2008. The PlcR virulence regulon of Bacillus cereus. PLoS One 3:e2793. https://doi.org/10.1371/journal.pone.0002793Grenha R, Slamti L, Nicaise M, Refes Y, Lereclus D, Nessler S. 2013. Structural basis for the activation mechanism of the PlcR virulence regulator by the quorum-sensing signal peptide PapR. Proc Natl Acad Sci U S A 110:1047–1052. https://doi.org/10.1073/pnas.1213770110.Böhm ME, Huptas C, Krey VM, Scherer S. 2015. 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Struktura genetyczna środowiskowych izolatów Bacillus cereus sensu lato z północno-wschodniej Polski

Rozdziały (3 załączniki) w postaci publikacji naukowych w czasopismach FEMS Microbiology Ecology (Appendix 1) oraz PLOS ONE (Appendix 2 oraz Appendix 3).Przedstawiciele B. cereus s.l. występują powszechnie w środowisku naturalnym i wywierają ogromny wpływ na zdrowie człowieka, przemysł spożywczy oraz rolnictwo. Te tlenowe laseczki z jednej strony produkują toksyny szkodliwe dla ludzi i zwierząt, ale znane są również z syntezy wtórnych metabolitów degradujących niebezpieczne związki chemiczne lub wspomagających wzrost roślin. Powyższe właściwości były intensywnie badane w odniesieniu do szczepów o szczególnym znaczeniu gospodarczym i medycznym. Tymczasem pokrewieństwo filogenetyczne i podłoże ekologicznej dywersyfikacji szczepów izolowanych z gleby nie jest dostatecznie poznane. Analizowałam strukturę genetyczną 297 szczepów B. cereus s.l. wyizolowanych z gleby (i) Narwiańskiego PN, (ii) Białowieskiego PN, (iii) Biebrzańskiego PN oraz (iv) gospodarstwa rolnego w Jasienówce. Zidentyfikowałam homogeniczną grupę bakterii (i) zdolną do wzrostu w niskiej temperaturze (ekotyp psychrotolerancyjny) oraz (ii) syntetyzującą ciemno-brązowy barwnik (ekotyp melaninowy). Analizy MLST wskazują na istnienie specyficznych genotypów wśród naturalnych populacji B. cereus s.l. Ponadto wykazałam istnienie trzech grup filogenetycznych, obejmujących zmienną liczbę B. cereus, B. thuringiensis i B. mycoides. Jednakże analizy typów sekwencyjnych i kompleksów klonalnych wskazują, iż środowiskowe izolaty B. cereus s.l. nie reprezentują jednego gatunku. Szczegółowe badania genetyczne, fenotypowe oraz biochemiczne środowiskowych szczepów B. cereus s.l., rzuciły nowe światło na ewolucję oraz ekologiczną adaptację tych bakterii.B. cereus s.l. are widespread in natural environments and have a significant impact on human health, food industry, and agriculture. On one hand, members of this group synthetize various toxins harmful to humans, herbivores and invertebrates. On the other hand, they are also known as producers of various secondary metabolites whereby they degrade pollutants and promote the growth of plants and animals. These aspects have been intensively studied especially with regard to the B. cereus group members with the highest impact on human health and economy. Meanwhile, the phylogenetic relationships and the basis of ecological diversification of soil B. cereus s.l. remains largely undescribed. I investigated the genetic structure of 297 B. cereus s. l. soil isolates from diverse habitats in Northeastern Poland, such as (i) the Narew NP, (ii) the Białowieża NP, (iii) Biebrza NP, and (iv) agricultural land in Jasienowka. I identified homogenous groups of bacteria able to (i) growth in low temperature (psychrotolerant ecotype) and (ii) synthesis a dark pigment (melanotype). The MLST analysis indicates the existence of specific genotypes within the natural B. cereus s.l. populations. Phylogenetic studies revealed three major clades, in which B. cereus, B. thuringiensis and B. mycoides were intermixed. However, analysis of sequence types and clonal complexes indicate that environmental B. cereus s.l. do not represent one species. Detailed genetic, phenotypic and biochemical analyses of the environmental B. cereus s.l. strains shed new light on the evolution and ecological adaptation of these bacteria to specific soil habitats differing in scope of human activity.Wydział Biologiczno-Chemiczny. Instytut Biologii

Detection and Removal of Priority Substances and Emerging Pollutants from Stormwater: Case Study of the Kołobrzeska Collector, Gdańsk, Poland

Progressive urban development affects environmental balance and disrupts the hydrologic cycle, in which rainfall plays a significant role. Since rainwater is considered a valuable resource of the environment, many technical solutions are implemented that enable effective rainwater management. On the other hand, stormwater runoff from urban areas contains numerous (also toxic) substances, and therefore should be properly treated. In this study, a multistage constructed wetland (MCW) pilot installation was used to remove selected groups of priority substances and emerging pollutants from rainwater discharged from the urbanized catchment of the Kołobrzeska stormwater collector in Gdańsk, Poland. The obtained results show that rainwater runoff was characterized by a variable concentrations of heavy metals (Zn, Cd, Cu, Ni, Pb, Hg), polycyclic aromatic hydrocarbons (benzo(a)pyrene, benzo(b)fluoranthene, phenanthrene, fluoranthene and pyrene) and microplastics. Depending on the hydraulic load of the bed, the reduction efficiency for heavy metals ranged from 26.19 to 100%, and for microplastics from 77.16 to 100%, whereas for polycyclic aromatic hydrocarbons it was consistently high, and equaled 100%

Detection and Removal of Priority Substances and Emerging Pollutants from Stormwater: Case Study of the Kołobrzeska Collector, Gdańsk, Poland

Progressive urban development affects environmental balance and disrupts the hydrologic cycle, in which rainfall plays a significant role. Since rainwater is considered a valuable resource of the environment, many technical solutions are implemented that enable effective rainwater management. On the other hand, stormwater runoff from urban areas contains numerous (also toxic) substances, and therefore should be properly treated. In this study, a multistage constructed wetland (MCW) pilot installation was used to remove selected groups of priority substances and emerging pollutants from rainwater discharged from the urbanized catchment of the Kołobrzeska stormwater collector in Gdańsk, Poland. The obtained results show that rainwater runoff was characterized by a variable concentrations of heavy metals (Zn, Cd, Cu, Ni, Pb, Hg), polycyclic aromatic hydrocarbons (benzo(a)pyrene, benzo(b)fluoranthene, phenanthrene, fluoranthene and pyrene) and microplastics. Depending on the hydraulic load of the bed, the reduction efficiency for heavy metals ranged from 26.19 to 100%, and for microplastics from 77.16 to 100%, whereas for polycyclic aromatic hydrocarbons it was consistently high, and equaled 100%

Notes on selenium in mushrooms data determined by inductively coupled plasma atomic emission spectroscopy (ICP-AES) and hydride generation atomic absorption spectroscopy (HG-AAS) techniques

This manuscript discusses the credibility of the selenium in mushroom data generated using inductively coupled plasma atomic emission spectroscopy (ICP-AES) compared to that from hydride generation atomic absorption spectroscopy (HG-AAS). Selenium (Se) was determined by two methods: one was the widely applied and well validated hydride generation atomic absorption spectroscopy (HG-AAS) that was a reference method, while the validity of using the ICP-AES was tested. We found that Se determination in fungal and plant materials by ICP-AES gives inaccurate and imprecise results. Hence, reports of rather high concentrations of Se determined by ICP-AES for mushrooms that do not accumulate Se or that have not emerged at selenoferous areas should be treated with caution, because such data could be highly biased

Piloting technologies for microplastics removal from urban waters : FanpLESStic-sea project report

The FanpLESStic-sea – “Initiatives to remove microplastics before they enter the sea” project funded by the EU Interreg Baltic Sea Region programme aimed at decreasing microplastic (MP) sources and removing MPs before they enter the Baltic Sea. As part of the project, novel and innovative removal technologies and methods for MPs in urban waters were developed and tested in four pilot systems by three project partners: a common reed filter for treatment of urban snow meltwaters and stormwaters by Natural Resources Institute Finland (Luke), an urban snow melting and filtering unit by Clewat Ltd. and Luke, a constructed wetland system for wastewater effluent by Gdansk Water Utilities, Poland, and a constructed wetland system for stormwater treatment by Gdansk Water. The results obtained from the four MP removal pilots were very promising. The filter consisting of harvested and bundled common reed is cost-effective and locally adaptable. However, due the insufficient time available for maturation of the reed filter, its performance in MP removal remained uncertain. Novel snow melting and filtering technology developed by Clewat Ltd. solves the problem of dumping urban snow untreated into the sea in a sustainable way. In this project, the efficiency of the snow treatment unit in MP removal was optimized concerning the mesh size for the finer filter used. The constructed wetland systems piloted showed MP removal efficiencies up to 90% from urban stormwaters and wastewater treatment plant effluent. In addition to retention of MPs present in various urban waters, high removal rates were demonstrated for total suspended solids, nutrients and various pollutants. However, additional longer-time pilot experiments are needed to optimize the performance of the systems, especially for those based on vegetation and biofilm formation. Environmental impact and techno-economic feasibility assessments were also performed for the piloted technologies