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Frontiers in Microbiology 2023Spores are important as dispersal and survival propagules in fungi. In this study we investigated the variation in number, shape, size and germination mode of ascospores...
Spores are important as dispersal and survival propagules in fungi. In this study we investigated the variation in number, shape, size and germination mode of ascospores in , the only species of the genus known to fruit in the autumn. Based on the observation of five samples, we first discovered significant variation in the shape and size of ascospores in . One to sixteen ascospores were found in the asci. Ascospore size correlated negatively with ascospore number, but positively with ascus size, and ascus size was positively correlated with ascospore number. We noted that ascospores, both from fresh collections and dried specimens, germinated terminally or laterally either by extended germ tubes, or via the production of conidia that were formed directly from ascospores at one, two or multiple sites. The direct formation of conidia from ascospores takes place within asci or after ascospores are discharged. Using laser confocal microscopy, we recorded the number of nuclei in ascospores and in conidia produced from ascospores. In most ascospores of , several nuclei were observed, as is typical of species of . However, nuclear number varied from zero to around 20 in this species, and larger ascospores harbored more nuclei. One to six nuclei were present in the conidia. Nuclear migration from ascospores to conidia was observed. Conidia forming directly from ascospores has been observed in few species of Pezizomycetes; this is the first report of the phenomenon in species. Morphological and molecular data show that conidial formation from ascospores is not found in all the specimens of this species and, hence, is not an informative taxonomic character in . Our data suggest that conidia produced from ascospores and successive mitosis within the ascus may contribute to asci with more than eight spores. The absence of mitosis and/or nuclear degeneration, as well as cytokinesis defect, likely results in asci with fewer than eight ascospores. This study provides new insights into the poorly understood life cycle of species and more broadly improves knowledge of conidia formation and reproductive strategies in Pezizomycetes.
PubMed: 38045031
DOI: 10.3389/fmicb.2023.1286501 -
Microorganisms Nov 2023This study aimed to investigate the optimal conditions for Papanicolaou (Pap) smear to increase the success rate of target cell isolation through manual microdissection...
This study aimed to investigate the optimal conditions for Papanicolaou (Pap) smear to increase the success rate of target cell isolation through manual microdissection (MMD) and prevent cell spread. Pap smears were prepared using an HPV42-positive SurePath™ liquid-based cytology case, and 46 and 50 koilocytes were used in wet and dried Pap smears, respectively, to verify the success rate of target cell isolation using MMD based on the HPV detection rate. During MMD, the microscopic examination of both specimens revealed that cells in dried smears could be easily identified; however, cell debris remained in the surrounding area after MMD. Although it was difficult to observe cells in wet smears, there was no cell debris. When the needle tip was immersed in DNA lysate after cell isolation through MMD, a difference in cell solubility was found between dry and wet smears. HPV42 was detected in 94.7% and 97.4% of dried and wet Pap smears, respectively, via polymerase chain reaction genotyping using lysed cell solution; the detection rates were not significantly different. The isolation of target cells from wet Pap smears using MMD reduced the risk of contamination and increased the success rate of HPV detection. This study might facilitate the identification of new CIN-derived HPV-infected cells using MMD with wet Pap smears.
PubMed: 38004711
DOI: 10.3390/microorganisms11112700 -
F1000Research 2022The objective of this anatomical study was to perform the morphometry of dried lumbar vertebrae in human cadavers.
BACKGROUND
The objective of this anatomical study was to perform the morphometry of dried lumbar vertebrae in human cadavers.
METHODS
This study utilized 200 adult human cadaveric dried lumbar vertebrae. The digital Vernier calipers was used to perform the measurements. The height, antero-posterior length, transverse length of the body of the vertebrae, interpedicular distance at the lateral ends, lamina length, height and thickness, superior and inferior articular facet height and width, mid sagittal and transverse diameter of vertebral foramen, height, width and thickness of the pars inter-articularis were measured.
RESULTS
The vertebral body's anteroposterior length was more at the lower border than at the superior border ( 0.01). The length of lamina was higher over the right in comparison to the left (p < 0.001). The height of lamina, width of inferior articular facet, diameter of lateral recess and thickness of pars inter-articularis were greater for the left sided specimens ( 0.01). The statistical significance was not observed for the comparison of the remaining parameters ( 0.05).
CONCLUSION
This anatomical study offered several dimensions of lumbar vertebrae, which are essential in the surgical practice. The implants at the lumbar vertebrae need to be manufactured based on the anatomical dimensions of that particular sample population.
PubMed: 37990689
DOI: 10.12688/f1000research.126879.2 -
Emerging Infectious Diseases Dec 2023We conducted a cross-sectional study to determine the prevalence of soil-transmitted helminthiases (STH) in areas of rural Alabama, USA, that have sanitation deficits....
We conducted a cross-sectional study to determine the prevalence of soil-transmitted helminthiases (STH) in areas of rural Alabama, USA, that have sanitation deficits. We enrolled 777 children; 704 submitted stool specimens and 227 a dried blood spot sample. We microscopically examined stool specimens from all 704 children by using Mini-FLOTAC for helminth eggs. We tested a subset by using molecular techniques: real-time PCR analysis for 5 STH species, TaqMan Array Cards for enteric helminths, and digital PCR for Necator americanus hookworm. We analyzed dried blood spots for Strongyloides stercoralis and Toxocara spp. roundworms by using serologic testing. Despite 12% of our cohort reporting living in homes that directly discharge untreated domestic wastewater, stool testing for STH was negative; however, 5% of dried blood spots were positive for Toxocara spp. roundworms. Survey data suggests substantial numbers of children in this region may be exposed to raw sewage, which is itself a major public health concern.
Topics: Child; Animals; Humans; Cross-Sectional Studies; Soil; Alabama; Helminthiasis; Helminths; Feces; Prevalence
PubMed: 37987581
DOI: 10.3201/eid2912.230751 -
Se Pu = Chinese Journal of... Nov 2023The discovery and identification of mushroom toxins has long been an important area in the fields of toxicology and food safety. Mushrooms are widely favored for their...
The discovery and identification of mushroom toxins has long been an important area in the fields of toxicology and food safety. Mushrooms are widely favored for their culinary and medicinal value; however, the presence of potentially lethal toxins in some species poses a substantial challenge in ensuring their safe consumption. Therefore, the development of a robust and sensitive analytical method is necessary for accurately identifying the risks associated with mushroom consumption. The study of mushroom toxins, which are characterized by their diversity and substantial variations in chemical structures, present a considerable challenge for achieving precise and high-throughput analysis. To address this issue, the present study employed a robust approach combining a solid-phase extraction (SPE) purification technique with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) to establish an analytical method for the detection and quantification of five amatoxins and two tryptamines (psilocybin and bufotenine) present in some mushrooms. Several optimization procedures were undertaken, including optimizing the chromatographic conditions, mass spectrometric parameters, and sample extraction and purification. The procedure involved the extraction of dry mushroom powder with methanol containing 0.3% formic acid, followed by purification using a strong cation exchange cartridge (SCX). The analytes were separated on a T3 chromatographic column (100 mm×2.1 mm, 1.8 μm) using mobile phases of acetonitrile and 5 mmol/L ammonium acetate solution containing 0.1% formic acid. The multiple reaction monitoring (MRM) mode was employed for data acquisition. Amatoxins were quantified using matrix-matched standard calibration curves, whereas isotopic internal standards were used to quantify tryptamine. The results showed that all seven toxins exhibited good linearities (>0.99) within the optimized concentration range. The limits of detection (LODs) for bufotenine, psilocybin, and amatoxins were determined as 2.0, 5.0, and 10 μg/kg, respectively, while the limits of quantification (LOQs) were determined as 5.0, 10, and 20 μg/kg, respectively. The LOD and LOQ values further underscore the ability of the method to detect minute quantities of toxins, making it particularly well suited for screening food samples for potential contamination. Using dried shiitake mushroom powder as the matrix, the recoveries of the two tryptamines ranged from 80.6% to 117%, with relative standard deviations (RSDs) ranging from 1.73% to 5.98%, while the recoveries of amatoxins ranged from 71.8% to 115%, with RSDs varying from 2.14% to 9.92% at the three concentration levels. The consistent and satisfactory recoveries of amatoxins and tryptamines demonstrated the ability of this method to accurately quantify the target analytes even in a complex matrix. Comparison with the results of supplementary test method recognized by State Administration for Market Regulation for food (BJS 202008) demonstrated comparable results, indicating no significant differences (>0.05) in amatoxin contents. The newly developed method is rapid, accurate, precise, meets the required standards, and is suitable for the detection of seven toxins in wild mushrooms. As part of the application of this method, a comprehensive investigation of the distribution of toxins in wild mushrooms from Fujian Province was undertaken. In this study, 59 wild mushroom samples from nine cities were collected in the Fujian province. Species identification was conducted using rDNA-internal transcribed space (rDNA-ITS) molecular barcode technology, which revealed the presence of toxins in the two samples. Notably, one specimen named contained -amanitin, -amanitin, and phalloidin in quantities of 607, 377, and 69.0 mg/kg, respectively. Additionally, another sample, identified as Tricholomataceae, had a psilocybin concentration of 12.6 mg/kg.
Topics: Chromatography, High Pressure Liquid; Amanita; Tandem Mass Spectrometry; Psilocybin; Bufotenin; Powders; Mycotoxins; Tryptamines; DNA, Ribosomal
PubMed: 37968816
DOI: 10.3724/SP.J.1123.2023.07013 -
Plant Disease Nov 2023Cucumbers have great economic and social importance. Annual worldwide production is approximately 80 million tons (FAOSTAT, 2019), 184 thousand tons of which are...
Cucumbers have great economic and social importance. Annual worldwide production is approximately 80 million tons (FAOSTAT, 2019), 184 thousand tons of which are produced in Brazil (IBGE, 2020). Leaves with symptoms of anthracnose (necrotic brown or angular spots) were observed on cucumber plants grown in organic systems in September 2021, Pernambuco, Brazil (8°7'45''S, 35°16'167''W). About 40% of the plants fields were infected. Samples were collected and fragments were cut from the margins of the symptomatic tissue. The fragments were superficially disinfected with 70% ethanol (30 s) and 2% sodium hypochlorite (2 min), then washed three times with sterile distilled H2O and dried on sterile filter paper. The fragments were placed on potato dextrose agar (PDA) containing chloramphenicol (50 mg/L) and incubated at 28 ± 2 °C for 3 days. From the fungal isolates obtained, a representative specimen of Colletotrichum spp. was isolated, purified by subculturing from emergent hyphae tips and used for morphological characterization, phylogenetic analysis, and pathogenicity testing. The fungus isolated on PDA formed gray to grayish-black colonies with white aerial mycelia after 7 days. Ascomata were globose to subglobose, 120-200 × 100-150 μm in size (n = 10). Setae formed directly on the hyphae. Asci were 50-70 × 10-12 μm in size, 8-spored, unitunicate, thin-walled, and clavate. Ascospores were 14-22 × 4-5 μm in size (n = 30), hyaline, slightly curved to curved with obtuse to slightly rounded ends. Conidia were hyaline, smooth-walled, aseptate, straight, cylindrical, the apex and base rounded, and 12-15 × 5 μm in size, (n = 30). For molecular identification, the nuclear ribosomal internal transcribed spacers (nrITS), actin (ACT), beta-tubulin (TUB), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes were sequenced (Damm et al. 2019). The sequences obtained were deposited in GenBank (nrITS: OP720945, ACT: OP723523, TUB: OP723525, and GAPDH: OP723524). The sequences from the nrITS region, ACT, TUB2, and GAPDH were highly similar to those from C. plurivorum: nrITS - CBS 125474 (539/539 - 100%; NR_160828); ACT - CBS 125474 (270/271 - 99%; MG600925), TUB2 - CBS 125474 (517/518 - 99%; MG600985); and GAPDH - CBS 125474 (197/197 - 100%; MG600781), respectively. Multilocus phylogenetic analysis was performed using Bayesian inference, which showed that the isolate C. plurivorum FPO04 clustered in the same clade as the ex-type of C. plurivorum (CBS 125474). In the pathogenicity test, leaves of five healthy cucumber plants, previously injured in the middle region with sterile needles, were inoculated with 50 µl of a conidial suspension (1 × 106 spores mL -1) prepared from 7-day-old of colonies of C. plurivorum. Sterile distilled water was used as negative controls. The inoculated plants were maintained in a humid greenhouse chamber for 24 hours. After 7 days, the same anthracnose symptoms seen in the field were observed on the inoculated plants. Control plants remained healthy. Colletotrichum plurivorum was reisolated from symptomatic leaves, fulfilling Koch's postulates. This species has been reported from several crops, including Abelmoschus esculentus (okra) (Damm et al. 2019) and Glycine max (soybeans) (Zaw et al. 2019). To our knowledge, this is the first report of C. plurivorum causing anthracnose on cucumber leaves in Brazil. This report lays the groundwork for future studies to determine management practices for control of this disease in C. sativus.
PubMed: 37953227
DOI: 10.1094/PDIS-10-23-2245-PDN -
JIMD Reports Nov 2023Measurement of plasma and dried blood spot (DBS) phenylalanine (Phe) is key to monitoring patients with phenylketonuria (PKU). The relationship between plasma and...
Measurement of plasma and dried blood spot (DBS) phenylalanine (Phe) is key to monitoring patients with phenylketonuria (PKU). The relationship between plasma and capillary DBS Phe concentrations has been investigated previously, however, differences in methodology, calibration approach and assumptions about the volume of blood in a DBS sub-punch has complicated this. Volumetric blood collection devices (VBCDs) provide an opportunity to re-evaluate this relationship. Paired venous and capillary samples were collected from patients with PKU ( = 51). Capillary blood was collected onto both conventional newborn screening (NBS) cards and VBCDs. Specimens were analysed by liquid-chromatography tandem mass-spectrometry (LC-MS/MS) using a common calibrator. Use of VBCDs was evaluated qualitatively by patients. Mean bias between plasma and volumetrically collected capillary DBS Phe was -13%. Mean recovery (SD) of Phe from DBS was 89.4% (4.6). VBCDs confirmed that the volume of blood typically assumed to be present in a 3.2 mm sub-punch is over-estimated by 9.7%. Determination of the relationship between plasma and capillary DBS Phe, using a single analytical method, common calibration and VBCDs, demonstrated that once the under-recovery of Phe from DBS has been taken into account, there is no significant difference in the concentration of Phe in plasma and capillary blood. Conversely, comparison of plasma Phe with capillary DBS Phe collected on a NBS card highlighted the limitations of this approach. Introducing VBCDs for the routine monitoring of patients with PKU would provide a simple, acceptable specimen collection technique that ensures consistent sample quality and produces accurate and precise blood Phe results which are interchangeable with plasma Phe.
PubMed: 37927487
DOI: 10.1002/jmd2.12398 -
Applications in Plant Sciences 2023Current methods for maceration of plant tissue use hazardous chemicals. The new method described here improves the safety of dissection and maceration of soft plant...
PREMISE
Current methods for maceration of plant tissue use hazardous chemicals. The new method described here improves the safety of dissection and maceration of soft plant tissues for microscopic imaging by using the harmless enzyme pectinase.
METHODS AND RESULTS
Leaf material from a variety of land plants was obtained from living plants and dried herbarium specimens. Concentrations of aqueous pectinase and soaking schedules were optimized, and tissues were manually dissected while submerged in fresh solution following a soaking period. Most leaves required 2-4 h of soaking; however, delicate leaves could be macerated after 30 min while tougher leaves required 12 h to 3 days of soaking. Staining techniques can also be used with this method, and permanent or semi-permanent slides can be prepared. The epidermis, vascular tissue, and individual cells were imaged at magnifications of 10× to 400×. Only basic safety precautions were needed.
CONCLUSIONS
This pectinase method is a cost-effective and safe way to obtain images of epidermal peels, separated tissues, or isolated cells from a wide range of plant taxa.
PubMed: 37915428
DOI: 10.1002/aps3.11543 -
Brazilian Dental Journal 2023This study evaluated the effect of heating on the physicochemical properties and surface changes of tricalcium silicate sealers. Three tricalcium silicate root canal...
This study evaluated the effect of heating on the physicochemical properties and surface changes of tricalcium silicate sealers. Three tricalcium silicate root canal sealers (Bio-C Sealer, BioRoot-RCS, EndoSequence BC Sealer), and one epoxy resin-based sealer (AH Plus; control) were tested. The effect of heating on setting time (ST) and flowability were assessed according to ANSI/ADA 57 and ISO 6876 standards. Solubility and dimensional change (DC) of the set sealers were evaluated at 24 hours and after 30 days; the pH of the water used in the DC testing was also measured. Tests were repeated with heated sealers in an oven at 100 °C for 1 min. SEM and EDS analysis were performed. Data were analyzed using One-Way ANOVA and Tukey post-hoc tests (α=5%). Heating decreased the ST for AH Plus and EndoSequence (p<0.05). Heating reduced flowability (p<0.05) and increased pH for AH Plus (p<0.05). The solubility of Bio-C (dried specimens) was not in accordance with the ANSI/ADA standard. The solubility of EndoSequence was significantly higher (p<0.05) when it was heated and dried after 30 days. DC of Bio-C (24 h and 30 days), BioRoot-RCS (30 days) and AH Plus (24 h and 30 days) were not in accordance with the standards. SEM and EDS analysis showed significant changes in sealer microstructure after heating. In conclusion, heating decreased the ST and increased the solubility of EndoSequence BC sealer. No significant changes in flowability, DC, and pH were identified for all three tricalcium silicate sealers after heat application. However, all sealers had significant surface changes.
Topics: Root Canal Filling Materials; Heating; Dental Pulp Cavity; Calcium Compounds; Epoxy Resins; Silicates; Materials Testing
PubMed: 37909640
DOI: 10.1590/0103-6440202305237 -
G3 (Bethesda, Md.) Dec 2023Relict species, like coelacanth, gingko, tuatara, are the remnants of formerly more ecologically and taxonomically diverse lineages. It raises the questions of why they...
Relict species, like coelacanth, gingko, tuatara, are the remnants of formerly more ecologically and taxonomically diverse lineages. It raises the questions of why they are currently species-poor, have restrained ecology, and are often vulnerable to extinction. Estimating heterozygosity level and demographic history can guide our understanding of the evolutionary history and conservation status of relict species. However, few studies have focused on relict invertebrates compared to vertebrates. We sequenced the genome of Baronia brevicornis (Lepidoptera: Papilionidae), which is an endangered species, the sister species of all swallowtail butterflies, and is the oldest lineage of all extant butterflies. From a dried specimen, we were able to generate both long-read and short-read data and assembled a genome of 406 Mb for Baronia. We found a fairly high level of heterozygosity (0.58%) compared to other swallowtail butterflies, which contrasts with its endangered and relict status. Taking into account the high ratio of recombination over mutation, demographic analyses indicated a sharp decline of the effective population size initiated in the last million years. Moreover, the Baronia genome was used to study genome size variation in Papilionidae. Genome sizes are mostly explained by transposable elements activities, suggesting that large genomes appear to be a derived feature in swallowtail butterflies as transposable elements activity is recent and involves different transposable elements classes among species. This first Baronia genome provides a resource for assisting conservation in a flagship and relict insect species as well as for understanding swallowtail genome evolution.
Topics: Animals; Butterflies; Genome Size; Phylogeny; DNA Transposable Elements; Genomics; Demography
PubMed: 37847748
DOI: 10.1093/g3journal/jkad239