Geometrical Frustration in Interleukin-33 Decouples the Dynamics of the Functional Element from the Folding Transition State Ensemble

Interleukin-33 (IL-33) is currently the focus of multiple investigations into targeting pernicious inflammatory disorders. This mediator of inflammation plays a prevalent role in chronic disorders such as asthma, rheumatoid arthritis, and progressive heart disease. In order to better understand the possible link between the folding free energy landscape and functional regions in IL-33, a combined experimental and theoretical approach was applied. IL-33 is a pseudo- symmetrical protein composed of three distinct structural elements that complicate the folding mechanism due to competition for nucleation on the dominant folding route. Trefoil 1 constitutes the majority of the binding interface with the receptor whereas Trefoils 2 and 3 provide the stable scaffold to anchor Trefoil 1. We identified that IL-33 folds with a three-state mechanism, leading to a rollover in the refolding arm of its chevron plots in strongly native conditions. In addition, there is a second slower refolding phase that exhibits the same rollover suggesting similar limitations in folding along parallel routes. Characterization of the intermediate state and the rate limiting steps required for folding suggests that the rollover is attributable to a moving transition state, shifting from a post- to pre-intermediate transition state as you move from strongly native conditions to the midpoint of the transition. On a structural level, we found that initially, all independent Trefoil units fold equally well until a Q_CA of 0.35 when Trefoil 1 will backtrack in order to allow Trefoils 2 and 3 to fold in the intermediate state, creating a stable scaffold for Trefoil 1 to fold onto during the final folding transition. The formation of this intermediate state and subsequent moving transition state is a result of balancing the difficulty in folding the functionally important Trefoil 1 onto the remainder of the protein. Taken together our results indicate that the functional element of the protein is geometrically frustrated, requiring the more stable elements to fold first, acting as a scaffold for docking of the functional element to allow productive folding to the native state.     

Shellfish subsidies along the Pacific coast of North America

Spatial subsidies are associated with pronounced ecosystem responses, as nutrients cross ecological boundaries and cascade through food webs. While the importance of subsidies is known, the role of shellfish, specifically molluscs, as a marine subsidy has not been formally described. Focusing primarily on the Pacific coast of North America, we identify vectors that transport shellfish-derived nutrients into coastal terrestrial environments, including birds, mammals, and over 13 000 yr of marine resource use by people. Evidence from recipient ecosystems suggests shellfish drastically influence soil chemistry, forest productivity and the diversity of primary producers at the regional and landscape level. Responses in higher trophic levels have not yet been investigated, but given documented responses in lower trophic levels, this may be due to a lack of examination. To determine if the processes we describe within the northeast Pacific are pertinent to coastal environments worldwide, we also explore shellfish subsidies globally, with a specific focus on temperate and tropical islands. As shellfish are not as spatially or temporally constrained as other subsidies, our examination suggests our findings are applicable to many other geographical regions along the marine–terrestrial interface.     

Changes in the Alyki Kitrous wetland in northern Greece: 1990–1999, and future prospects

The Alyki wetland in northern Greece is a site of notable herpetological and ornithological importance. A large population of the tortoise Testudo hermanni began to recover slowly (r = 0.016) between 1990 and 1999, following catastrophic habitat destruction in 1980 and slow decline from 1980–1990. Heathland vegetation showed signs of desiccation, probably due to increased drainage by the salt works, including deterioration of Crataegus, Ruscus and Asphodelus, and loss of formerly important food plants of tortoises. Activity and body mass condition of tortoises were reduced in summer in grassy heath habitats. Predation of artificial (chicken egg) tortoise nests increased, and the relative frequency of juveniles (about 1 % of samples) was lower than after severe habitat destruction or pesticide damage. These changes were attributed to an increasing population of badgers, Meles meles. Population levels of waders were lower in 1998/1999 than in 1988/1989, especially avocets (Recurvirostra avosetta) for which the site was internationally important. Breeding populations of most waders and larids also declined. Mediterranean gulls, for which Alyki was formerly the most important breeding area outside Russia, did not nest from 1990 to 1998, and nests were unsuccessful in 1999. Changes in breeding populations were due to falling lagoon levels, exposure to predators and vegetation encroachment on islet breeding sites. Midwinter waterfowl populations remained high or increased, however, as alternative feeding areas were available in the lagoon. Preservation of the habitats against development of the salt works and increasing desiccation are necessary to maintain the long-term conservation value of the Alyki site.     

Marine metagenomics: strategies for the discovery of novel enzymes with biotechnological applications from marine environments

Metagenomic based strategies have previously been successfully employed as powerful tools to isolate and identify enzymes with novel biocatalytic activities from the unculturable component of microbial communities from various terrestrial environmental niches. Both sequence based and function based screening approaches have been employed to identify genes encoding novel biocatalytic activities and metabolic pathways from metagenomic libraries. While much of the focus to date has centred on terrestrial based microbial ecosystems, it is clear that the marine environment has enormous microbial biodiversity that remains largely unstudied. Marine microbes are both extremely abundant and diverse; the environments they occupy likewise consist of very diverse niches. As culture-dependent methods have thus far resulted in the isolation of only a tiny percentage of the marine microbiota the application of metagenomic strategies holds great potential to study and exploit the enormous microbial biodiversity which is present within these marine environments.     

Strain persistence of invasive Candida albicans in chronic hyperplastic candidosis that underwent malignant change

Objectives: The aim of this study was to assess persistence and tissue invasion of Candida albicans strains isolated from a 65 year‐old patient with chronic hyperplastic candidosis (CHC), that subsequently developed into squamous cell carcinoma (SCC). Materials and Methods: C. albicans (n=7) were recovered from the oral cavity of the patient over seven years. Confirmation of CHC and SCC in this patient was achieved by histopathological examination of incisional biopsy tissue. DNA fingerprinting was performed on the seven isolates from the CHC patient together with a further eight isolates from patients with normal oral mucosa (n=2), chronic atrophic candidosis (n=1), SCC (n=1) and CHC (n=4). Genotyping involved the use of inter‐repeat PCR using the eukaryotic repeat primer 1251. Characterisation of the tissue invasive abilities of the isolates was achieved by infecting a commercially available reconstituted human oral epithelium (RHE; SkinEthic, Nice, France). After 24 h. C. albicans tissue invasion was assessed by histopathological examination. Results: DNA fingerprinting demonstrated strain persistence of C. albicans in the CHC patient over a seven year period despite provision of systemic antifungal therapy. The strain of C. albicans isolated from this patient was categorised as a high invader within the RHE compared to other isolates. Conclusions: Candidal strain persistence was evident in a patient with CHC over seven years. This persistence may be due to incomplete eradication from the oral cavity following antifungal therapy or subsequent recolonisation from other body sites or separate exogenous sources. The demonstration of enhanced in vitro tissue invasion by this particular strain may, in part, explain the progression to carcinoma.     

SEALNET: Facial recognition software for ecological studies of harbor seals

Methods for long‐term monitoring of coastal species such as harbor seals (Phoca vitulina) are often costly, time‐consuming, and highly invasive, underscoring the need for improved techniques for data collection and analysis. Here, we propose the use of automated facial recognition technology for identification of individual seals and demonstrate its utility in ecological and population studies. We created a software package, SealNet, that automates photo identification of seals, using a graphical user interface (GUI) software to detect, align, and chip seal faces from photographs and a deep convolutional neural network (CNN) suitable for small datasets (e.g., 100 seals with five photos per seal) to classify individual seals. We piloted the SealNet technology with a population of harbor seals located within Casco Bay on the coast of Maine, USA. Across two years of sampling, 2019 and 2020, at seven haul‐out sites in Middle Bay, we obtained a dataset optimized for the development and testing of SealNet. We processed 1752 images representing 408 individual seals and achieved 88% Rank‐1 and 96% Rank‐5 accuracy in closed set seal identification. In identifying individual seals, SealNet software outperformed a similar face recognition method, PrimNet, developed for primates but retrained on seals. The ease and wealth of image data that can be processed using SealNet software contributes a vital tool for ecological and behavioral studies of marine mammals in the developing field of conservation technology.

In this paper, we describe the successful application of our newly developed automated facial recognition software as a tool for ecological analysis of harbor seals (Phoca vitulina). We outline an emerging method of data collection and analysis that facilitates rapid interpretation of large photo datasets over wide temporal and geographic scales. In addition, we use this machine learning‐based technology in a preliminary ecological study in a wild population of seals in the Casco Bay region of Maine to demonstrate the effectiveness of this non‐invasive method for use in mark‐recapture and site fidelity studies in the field.