UNASSIGNED: Sporotrichosis is a subcutaneous mycosis caused by fungi of the genus Sporothrix sp. Phenotypic and genotypic differences have been associated with their geographic distribution, virulence, or clinical manifestation of sporotrichosis. In the past decade, the interest in identifying species of the Sporothrix sp. has been increasing, due to its epidemiological importance and, in consequence, is important to know how to preserve them for future studies, in culture collection.
UNASSIGNED: The purposes of this study were to analyze the global distribution of environmental isolates and/or causal agents of sporotrichosis identified by polyphasic taxonomy, with mandatory use of molecular identification, and to evaluate the percentages and distribution of isolates stored in culture collections.
UNASSIGNED: A systematic review of articles on animal and human sporotrichosis and/or environmental isolation of the fungus, from 2007 to 2023, was done. Results: Our results demonstrated that, S. globosa, S. schenckii, and S. brasiliensis were the most identified species. With respect to the deposit and maintenance of species, we observed that only 17% of the strains of Sporothrix sp. isolated in the world are preserved in a culture collection.
UNASSIGNED: This systematic review confirmed a difficulty in obtaining the frequency of Sporothrix species stored in culture collection and insufficient data on the molecular identification mainly of animal sporotrichosis and isolation of Sporothrix sp. in environmental samples.

Collecting and preserving biological samples in the field, particularly in remote areas in tropical forests, prior to laboratory analysis is challenging. Blood samples in many cases are used for nucleic acid-based species determination, genomics or pathogen research. In most cases, maintaining a cold chain is impossible and samples remain at ambient temperature for extended periods of time before controlled storage conditions become available. Dried blood spot (DBS) storage, blood stored on cellulose-based paper, has been widely applied to facilitate sample collection and preservation in the field for decades. However, it is unclear how long-term storage on this substrate affects nucleic acid concentration and integrity. We analysed nucleic acid quality from DBS stored on Whatman filter paper no. 3 and FTA cards for up to 15 years in comparison to cold-chain stored samples using four nucleic acid extraction methods. We examined the ability to identify viral sequences from samples of 12 free-ranging primates in the Amazon forest, using targeted hybridization capture, and determined if mitochondrial genomes could be retrieved. The results suggest that even after extended periods of storage, DBS will be suitable for some genomic applications but may be of limited use for viral pathogen research, particularly RNA viruses.

Environmental DNA (eDNA) workflows contain many familiar molecular-lab techniques, but also employ several unique methodologies. When working with eDNA, it is essential to avoid contamination from the point of collection through preservation and select a meaningful negative control. As eDNA can be obtained from a variety of samples and habitats (e.g., soil, water, air, or tissue), protocols will vary depending on usage. Samples may require additional steps to dilute, block, or remove inhibitors or physically break up samples or filters. Thereafter, standard DNA isolation techniques (kit-based or phenol:chloroform:isoamyl [PCI]) are employed. Once DNA is extracted, it is typically quantified using a fluorometer. Yields vary greatly, but are important to know prior to amplification of the gene(s) of interest. Long-term storage of both the sampled material and the extracted DNA is encouraged, as it provides a backup for spilled/contaminated samples, lost data, reanalysis, and future studies using newer technology. Storage in a freezer is often ideal; however, some storage buffers (e.g., Longmires) require that filters or swabs are kept at room temperature to prevent precipitation of buffer-related solutes. These baseline methods for eDNA isolation, validation, and preservation are detailed in this protocol chapter. In addition, we outline a cost-effective, homebrew extraction protocol optimized to extract eDNA.

Impressions of vertebrate bodies or their parts, such as trace fossils and natural molds of bones, are a valuable source of information about ancient faunas which may supplement the standard fossil record based on skeletal elements. Whereas trace fossils of animal activity are relatively common and actively studied within the field of ichnology, and natural impressions of internal or external surfaces are a frequent preservation mode in fossil invertebrates, natural molds of bones are comparatively rare and less extensively documented and discussed. Among them, internal molds (steinkerns) of turtle shells are a relatively well-known form of preservation, but the mechanisms and taphonomic prerequisites leading to their formation are poorly studied. External shell molds are even less represented in the literature. Herein, we describe a historic specimen of a natural external turtle plastron mold from the Triassic (Norian) Löwenstein Formation of Germany-a formation which also yielded a number of turtle steinkerns. The specimen is significant not only because it represents an unusual form of preservation, but also due to its remarkably large size and the presence of a potential shell pathology. Although it was initially interpreted as Proterochersis sp., the recent progress in the knowledge of proterochersid turtles leading to an increase in the number of known taxa within that group allows us to verify that assessment. We confirm that the specimen is morphologically consistent with the genus and tentatively identify it as Proterochersis robusta, the only representative of that genus from the Löwenstein Formation. We note, however, that its size exceeds the size observed thus far in Proterochersis robusta and fits within the range of Proterochersis porebensis from the Grabowa Formation of Poland. The marks interpreted as shell pathology are morphologically consistent with Karethraichnus lakkos-an ichnotaxon interpreted as a trace of ectoparasites, such as leeches. This may support the previously proposed interpretation of Proterochersis spp. as a semiaquatic turtle. Moreover, if the identification is correct, the specimen may represent a very rare case of a negative preservation of a named ichnotaxon. Finally, we discuss the taphonomy of the Löwenstein Formation turtles in comparison with other Triassic turtle-yielding formations which show no potential for the preservation of internal or external shell molds and propose a taphonomic model for the formation of such fossils.

Improving the shelf lives of fruits is challenging. The biodegradable polysaccharide pullulan exhibits excellent film-forming ability, gas barrier performance, and natural decomposability, making it an optimal material for fruit preservation. To overcome problems of high cost and film porosity of existing packaging technologies, we aimed to develop pullulan-based packaging paper to enhance the shelf lives of fruits. A thin paper coating comprising a mixture of 15 wt.% pullulan solution at various standard viscosities (75.6, 77.8, and 108.5 mPa·s) with tea polyphenols (15:2) and/or vitamin C (150:1) improved the oxygen transmission rate (120-160 cm3 m-2·24 h·0.1 MPa), water vapor transmission rate (<5.44 g·mm-1 m-2·h·kPa), maximum free radical clearance rate (>87%), and antibacterial properties of base packaging paper. Grapes wrapped with these pullulan-based papers exhibited less weight loss (>4.41%) and improved hardness (>16.4%) after 10 days of storage compared to those of control grapes (wrapped in untreated/base paper). Grapes wrapped with pullulan-based paper had >12.6 wt.% total soluble solids, >1.5 mg/g soluble protein, >0.44 wt.% titratable acidity, and ≥4.5 mg 100 g-1 ascorbic acid. Thus, pullulan-based paper may prolong the shelf life of grapes with operational convenience, offering immense value for fruit preservation.

BACKGROUND: Alveolar ridge preservation (ARP) procedures are designed to lessen dimensional changes in the alveolar ridge after tooth extraction. Wound healing after ridge preservation involves the formation of new vital bone in the former socket, and this vital bone is important in the osseointegration of dental implants.
METHODS: A series of ARP studies have been performed to help clinicians better understand the wound-healing events that occur following tooth extraction and ridge preservation. Different protocols have been examined using various materials and periods of healing time prior to implant placement. The primary aim of these studies was to ascertain the relative percentage of vital bone formation, residual graft material, and connective tissue (CT)/other at the healing site using histomorphometric examination of bone core biopsies obtained during osteotomy preparation.
RESULTS: For allografts, the use of demineralized bone alone or in combination with mineralized is associated with more vital bone formation than the use of mineralized allograft alone. For mineralized allografts, the use of cortical versus cancellous bone has only minimal impact on new bone formation. Xenografts from bovine and porcine sources appear to have similar vital bone formation. Longer healing times prior to implant placement are associated with increased vital bone formation and decreased residual graft material. The most stable component in most studies is the percentage of CT/other.
CONCLUSIONS: The percentage of vital bone and residual graft at ARP sites is dependent on the materials used and the length of healing time prior to obtaining core biopsies.
CONCLUSIONS: What factors may affect the amount of new bone at the ARP site? At a time point about 4 months after ARP, the type of graft material used for ARP plays a large role in new bone formation. Studies focus on means and standard deviations, but patients often do not \"follow the mean.\" Even if a single ARP protocol is used for all patients, there is great interindividual variability in new bone formation, and there is often variability between sites within a single patient. How long after ARP with an allograft should I wait to place an implant? Longer healing times such as 4-5 months generally provide higher amounts of vital bone formation than shorter healing times like 2-3 months. Differences in vital bone formation between ARP protocols tend to decrease with longer healing time. FDBA that contains demineralized bone, either alone or combined with mineralized FDBA, often provides higher amounts of new bone formation than 100% mineralized allograft, especially at shorter healing periods. Even a year after ARP with an allograft, residual graft material is often still present at the ARP site.

Many biomolecular condensates are enriched in and depend on RNAs and RNA binding proteins (RBPs). So far, only a few studies have addressed the characterization of the intermolecular interactions responsible for liquid-liquid phase separation (LLPS) and the impact of condensation on RBPs and RNAs. Here, we present an approach to study protein-RNA interactions inside biomolecular condensates by applying cross-linking of isotope labeled RNA and tandem mass spectrometry to phase-separating systems (LLPS-CLIR-MS). LLPS-CLIR-MS enables the characterization of intermolecular interactions present within biomolecular condensates at residue-specific resolution and allows a comparison with the same complexes in the dispersed phase. We observe that sequence-specific RBP-RNA interactions present in the dispersed phase are generally maintained inside condensates. In addition, LLPS-CLIR-MS identifies structural alterations at the protein-RNA interfaces, including additional unspecific contacts in the condensed phase. Our approach offers a procedure to derive structural information of protein-RNA complexes within biomolecular condensates that could be critical for integrative structural modeling of ribonucleoproteins (RNPs) in this form.

Heart failure with preserved ejection fraction (HFpEF) is increasingly recognised and diagnosed in clinical practice, a trend driven by an ageing population and a rise in contributing comorbidities, such as obesity and diabetes. Representing at least half of all heart failure cases, HFpEF is recognised as a complex clinical syndrome. Its diagnosis and management are challenging due to its diverse pathophysiology, varied epidemiological patterns, and evolving diagnostic and treatment approaches. This Seminar synthesises the latest insights on HFpEF, integrating findings from recent clinical trials, epidemiological research, and the latest guideline recommendations. We delve into the definition, pathogenesis, epidemiology, diagnostic criteria, and management strategies (non-pharmacological and pharmacological) for HFpEF. We highlight ongoing clinical trials and future developments in the field. Specifically, this Seminar offers practical guidance tailored for primary care practitioners, generalists, and cardiologists who do not specialise in heart failure, simplifying the complexities in the diagnosis and management of HFpEF. We provide practical, evidence-based recommendations, emphasising the importance of addressing comorbidities and integrating the latest pharmacological treatments, such as SGLT2 inhibitors.

Accurate minimum post-mortem interval (minPMI) estimations often rely on a precise age determination of insect developmental stages, which is significantly influenced by environmental temperature. An optimal preservation of the entomological samples collected at crime scenes is pivotal for a reliable aging of immature insect samples. For blow flies (Diptera: Calliphoridae), the most widely used insect indicators in forensic investigations, an appropriate preservation of tissues is particularly important in the case of puparial samples because aging methods for intra-puparial forms usually depend on morphological analyses; however, although informative soft tissues and structures could be discoloured and/or distorted if they are not properly fixed, there is a lack of studies to assess different methods for the optimal preservation of intra-puparial forms collected in forensic investigations. The present study compares three preservation methods for intra-puparial forms of the blow fly Calliphora vicina Robineau-Desvoidy, 1830: (i) direct immersion into 80% ethanol, (ii) puncturing of the puparium and hot water killing (HWK) prior to preservation in 80% ethanol, and (iii) HWK without puncturing before preservation in 80% ethanol. External and internal morphological analyses of intra-puparial forms of different ages were conducted to assess the quality of preservation. The results indicate that direct immersion in ethanol led to poor preservation, affecting both external and internal tissues. Both methods with HWK resulted in a better preservation, but puncturing resulted, in some cases, in physical damage of the specimens. HWK without puncturing emerged as the optimal preservation method, consistently yielding high preservation scores for both external and internal morphological analyses. These findings have practical implications for forensic practitioners and emphasise the need for updating some published guidelines and protocols in forensic entomology.

In the present study, the synergistic bactericidal effect and mechanism of ultrasound (US) combined with Lauroyl Arginate Ethyl (LAE) against Salmonella Typhimurium were investigated. On this basis, the effect of US+LAE treatment on the washing of S. Typhimurium on the surface of onions and on the physical and chemical properties of onion during fresh-cutting and storage were studied. The results showed that treatment with US+LAE could significantly (P < 0.05) reduce the number of S. Typhimurium compared to US and LAE treatments alone, especially the treatment of US+LAE (230 W/cm2, 8 min, 71 μM) reduced S. Typhimurium by 8.82 log CFU/mL. Confocal laser scanning microscopy (CLSM), flow cytometry (FCM), protein and nucleic acid release and N-phenyl-l-naphthylamine (NPN) assays demonstrated that US+LAE disrupted the integrity and permeability of S. Typhimurium cell membranes. Reactive oxygen species (ROS) and malondialdehyde (MDA) assays indicated that US+LAE exacerbated oxidative stress and lipid peroxidation in cell membranes. Field emission scanning electron microscopy (FESEM) demonstrated that US+LAE treatment caused loss of cellular contents and led to cell crumpling and even lost the original cell morphology. US+LAE treatment caused a significant (P < 0.05) decrease in the number of S. Typhimurium on onions, but there was no significant (P > 0.05) effect on the color, hardness, weight and ascorbic acid content of onions. This study elucidated the synergistic antibacterial mechanism of US+LAE and verified the feasibility of bactericidal effect on the surface of onions, providing a theoretical basis for improving the safety of fresh produce in the food industry and to propose a new way to achieve the desired results.