Light Microscopy Techniques Research Articles

Leaf secretory structures in Rosa lucieae (Rosaceae): two times of secretion-two ecological functions?

Secretory trichomes and colleters are two of the secretory structures whose exudates may cover thebody of the plant. Such secretions comprise resins or mucilages which are associated with an array of ecological roles. In Rosaceae, secretory trichomes have been reported for the leaves while colleters associated with leaf teeth. Our study aimed to identify the secretory structures of Rosa lucieae and understand the ecological role played by these glands as interpreted by morphoanatomical and histochemical studies. Samples from developing and fully mature leaves were collected, fixed, and processed according to usual techniques for light and scanning electron microscopy. In R. lucieae, colleters are restricted to the leaf and stipular margins and are associated with the teeth. They present a parenchymatous axis surrounded by a secretory palisade epidermis and usually fall off after the secretory activity is finished. Different from colleters, secretory trichomes are persistent. They present a multicellular secretory head and stalk. They are found at the base of the leaflet, petiolule, rachis, and petiole and occasionally on the stipular and leaf margins. The colleters predominantly secrete mucilages while the secretory trichomes secrete lipids and terpenes, both via cuticle rupture. The secretory activity of colleters is predominant in the leaf primordia, holding leaflets together and protecting meristems and leaves from desiccation, while the secretory trichomes maintain their secretory activity at different stages of leaf development, protecting different regions of the leaf against pathogens and herbivores.

Effect of barbotage refining time and recycled scrap content on the microstructure of EN AC 44200 alloy

The article presents the results of research on the effect of the barbotage refining time and recycled scrap content on selected properties, the microstructure and the degree of gasification of the EN AC 44200 alloy (EN AC-AlSi12). The input material consisted of various proportions of pure ingots and scrap; the scrap contents were 80%, 70% and 60%, respectively. Each batch of material was subjected to different refining times, which were 0, 7, 9 and 15 min, respectively. Microstructure studies were conducted with the techniques of light and scanning electron microscopy. Additionally, the distribution of pores in the volume of the material and their proportion were determined by means of the X-ray computed tomography method. In addition, the hardness and degree of gasification depending on the refining time were performed. It was proved that the refining process fosters homogenization of the microstructure and reduces porosity. Moreover, extending the refining time has a positive effect on reducing the porosity of thin-walled castings and reducing the level of outgassing of the alloy.

Interlaboratory study: Testing reproducibility of solid biofuels component identification using reflected light microscopy

Considering global market trends and concerns about climate change and sustainability, increased biomass use for energy is expected to continue. As more diverse materials are being utilized to manufacture solid biomass fuels, it is critical to implement quality assessment methods to analyze these fuels thoroughly. One such method is reflected light microscopy (RLM), which has the potential to complement and enhance current standard testing, leading to improving fuel quality assessment and, ultimately, preventing avoidable air pollution.An interlaboratory study (ILS) was conducted to test the reproducibility of biomass fuels component identification using a reflected light microscopy technique. The exercise was conducted on thirty photomicrographs showing biomass and various undesired components (like plastics or mineral matter), which were purposely added (by the ILS organizers) to contaminate wood pellets and charcoal-based grilling fuels.Forty-six participants had various levels of difficulty identifying the marked components, and as a result, the percentage of correct answers ranged from 52.2 to 94.4%. Among the most difficult components to distinguish were petroleum products and inorganic matter. Various reasons led to the misidentification, including insufficient morphological descriptions of the components provided to participants, ambiguities of the nomenclature, limitations of the analytical and exercise method, and insufficient experience of the participants.Overall, the results indicate that RLM has the potential to enhance the quality assessment of biomass fuels. However, they also demonstrate that the petrographic classification used in this exercise requires further refinement before it can be standardized. While a new simplified classification of solid biomass fuels components was created as an outcome of this study, future research is necessary to refine the nomenclature, develop a microscopic morphological description of the components, and verify the accuracy of component identification with a follow-up ILS.

Human health risk assessment for consumption of microplastics and plasticizing substances through marine species

A special characteristic of MP (microplastics) in the ocean is they may act as carriers of additives specific to the plastic materials used in their manufacture, such as plasticizers, among which Bisphenol-A (BPA), bis (2-ethylhexyl) phthalate (DEHP), dybuthyl phthalate (DBP). Both MP as the plasticizers were searched in composite samples of mangrove cockle (Anadara tuberculosa), Stolzmann's weakfish (Cynoscion stolzmanni) and arched swimming crab (Callinectes arcuatus). Extraction of MP was done through physical-chemical techniques and identification was carried out employing the techniques of light microscopy, energy dispersive spectrometer (EDS), scanning electron microscope (SEM) and Raman spectroscopy; the sizes of MP obtained were between 0.5 μm and 106 μm, the following average results being obtained: for Arched swimming crab 4.0 ± 1.0 MP/g; mangrove cockle 3.3 ± 2.9 MP/g; and for Stolzmann's weakfish, the average was 2.4 ± 1.3 MP/g; the most observed shapes were fibers and irregular segments; the most identified MP was polyethylene terephthalate (PET). Regarding extraction and quantification of plasticizers, the extraction stage was carried out using QuEChERS tubes; and the identification and quantification with gas chromatography coupled to a mass spectrometer (GC-MS). Regarding the plasticizing substances, DEHP was found in detectable levels in all the samples; BPA was found in 84% of the composite samples analyzed; DBP was found in 50% of them, of the analyzed samples 34% were positive for the 3 analytes. The dietary exposure of people to plasticizers was calculated and for BPA the exposure obtained was compared with respect to the TDI (tolerable intake dose) for pregnant women and the new TDI proposed by EFSA in 2021 according to the estrogenic effect of this substance in the fetus. The objective of the work was to determine if a relationship could be established between both PM and plasticizers, which gave a positive relationship.

Prevalence of Plasmodium Falciparum Gametocytes among Malaria Positive Cases at Korogwe District Hospital: The Use of Molecular Techniques in Comparison with Light Microscopy

BackgroundMonitoring gametocytes in the population can inform about the human infective reservoir, which greatly aids malaria transmission, and provide relevant data for transmission models. Using molecular techniques in preference to light microscopy to detect gametocytes may lead to most reliable results. Effective determination of gametocytes is inevitable to achieve the transmission-blocking interventions as a prime target to end malaria. We aimed to determine the prevalence of P. falciparum gametocytes in malaria-positive cases from Korogwe district hospital MethodologyArchived DNA samples collected from Korogwe district hospital collected in a cross-sectional study were used to determine the prevalence of P. falciparum gametocytes using specific primers for cPCR targeting a Pfg27 gene. Demographic data, including blood slides data were retrieved from the database for statistical analysis. ResultsWith light microscopy, prevalence of P. falciparum gametocytes was 9.8%; sensitivity and specificity were 35.6% and 99.2% respectively. The cPCR gave a prevalence of 25.9%, with a sensitivity of 94.1% and specificity of 81.5%. The cPCR was diagnostically found to be significantly superior over light microscopy technique (X2=45.780, P < 0.001). ConclusioncPCR is superior to light microscopy technique in detecting P. falciparum gametocytes when one considers a successive malaria transmission-blocking intervention.
 Rwanda J Med Health Sci 2023;6(2):175-183

Morphological analyses of sperms causing severe male infertility with light microscopy, motile sperm organellar morphology examination and electron microscopy tecnics.

To evaluate abnormal sperm morphology by three different methods including light microscope, real-time motile sperm organelle morphology assessment at 8050x high magnification and electron microscope and to determine degree of affect on morphologic defects level. Beside the light and electron microscopic techniques, the motile sperm organelle examination can offer information about sperm quality in a simple and quick way and by the IMSI developed through the use of sperm selected by this technique can contribute to the success of in vitro fertilization centers. We believe that sperm selection at high magnifications can increase IVF pregnancy rates in patients with sperm morphology that causes severe male infertility.

#6669 ANALYSIS OF KIDNEY BIOPSIES AND URINE SEDIMENT USING SCANNING ELECTRON MICROSCOPY (SEM) AND ATOMIC FORCE MICROSCOPY (AFM)

Abstract Background and Aims The classification of renal diseases is dependent on the evaluation of kidney biopsies using light microscopy techniques. Ultrastructural details (e.g. podocyte foot processes, amyloid fibrils, Fabri's intracellular inclusions) need transmission electron microscopy (TEM) which has high costs, requires specialized personnel and is time consuming. Therefore, alternative techniques are highly desirable. Here we report our experience with scanning electron microscopy (SEM) and atomic force microscopy (AFM). Method Paraffin-embedded kidney sections (3μm thick) were examined after dewaxing and Jones-methenamine methane staining for type IV collagen fibers. Dried urine sediments were prepared using standard methods for urine cytological analysis. Analysis was then performed using atomic force microscopy (nGauge system) on dried specimens and with Scanning Electron Microscopy (Zeiss) after gold-spattering of the surface. Reference images were obtained by standard microscopy. Results The combination of SEM and thin paraffin sections allows the observation of ultrastructural details in kidney biopsies with a very simple and fast preparation. AFM is even faster than SEM, as it does not require any preparation of the surface of the sample. An important advantage of SEM is the possibility to view the sample at low magnification, which makes it easier to identify the location of the glomeruli. It is thus possible to switch from small magnifications (100x) to very extreme magnifications (10000x). Notably, the interpretation of the images requires knowledge of the artifacts induced by paraffin inclusion. Specifically, structures and cells containing lipids (for example foamy cells) appear empty. Chromatin is also condensed in the nuclei around the periphery of the nucleus and in spike structures whit large empty areas. The recognition of nuclei in SEM and AFM is important for the subsequent interpretation of the images. Podocyte pedicles fusion can be identified relatively easily in SEM and in AFM. Both techniques add relevant ultrastructural details of urine sediment cells, from epithelial cells to isolated tubular cells. Also, casts show many ultrastructural features which are not evident in classical optical microscopy. It is also possible to measure the thickness of the basement membrane with great precision both in SEM and with AFM. The major limitation of this approach is the limited experience of pathologists with these methods and the absence of an atlas of normal and pathological kidney. Conclusion Considering the affordable costs of SEM and AFM instruments and the simple preparation of samples for their analysis, these should be considered a valid alternative to TEM. SEM and AFM are alternative methods, simple and relatively cheap and very fast to obtain ultrastructural details of renal biopsies. We thank ANED and AST for the great support given to research.

The Tripartite Lichen Ricasolia virens: Involvement of Cyanobacteria and Bacteria in Its Morphogenesis.

Ricasolia virens is an epiphytic lichen-forming fungus mainly distributed in Western Europe and Macaronesia in well-structured forests with ecological continuity that lack eutrophication. It is considered to be threatened or extinct in many territories in Europe (IUCN). Despite its biological and ecological relevance, studies on this taxon are scarce. The thalli are tripartite, and the mycobiont has a simultaneous symbiotic relationship with cyanobacteria and green microalgae, which represent interesting models to analyse the strategies and adaptations resulting from the interactions of lichen symbionts. The present study was designed to contribute to a better understanding of this taxon, which has shown a clear decline over the last century. The symbionts were identified by molecular analysis. The phycobiont is Symbiochloris reticulata, and the cyanobionts (Nostoc) are embedded in internal cephalodia. Light, transmission electron and low-temperature scanning microscopy techniques were used to investigate the thallus anatomy, ultrastructure of microalgae and ontogeny of pycnidia and cephalodia. The thalli are very similar to its closest relative, Ricasolia quercizans. The cellular ultrastructure of S. reticulata by TEM is provided. Non-photosynthetic bacteria located outside the upper cortex are introduced through migratory channels into the subcortical zone by the splitting of fungal hyphae. Cephalodia were very abundant, but never as external photosymbiodemes.

Pollen morphology of family Thymelaeaceae Juss. in India and its taxonomic implications

We provide pollen morphology and ultrastructure of 26 plant taxa, comprising 14 genera, 21 species, 1 sub-species and 3 varieties, of the Thymelaeaceae family from the Indian sub-continent using light micsoscopic (LM) and scanning electron microscopic (SEM) techniques. The principal aim was to provide new and useful information regarding the extent of the pollen morphological diversity among the Thymelaeaceae taxa and, thus, contribute to a better understanding of the taxonomy of the studied taxa. The pollen grains of the studied taxa are essentially radially symmetrical, apolar, (released as) monad, pantoporate with a typical crotonoid pattern, and spheroidal (mostly) in shape; but widespread variations with respect to the number and dispensation of pores, size, exine (ectoexine and endoexine) thickness, and the shape itself (spheroidal, sub-spheroidal and prolate spheroidal), as well as in the amb (outline) have been also recorded. Pollen identification keys, based on the pollen micro-morphological attributes, were also constructed in order to easily identify the 26 studied Thymelaeceae taxa up to species and/or varieties level (high taxonomic resolution, showing taxonomic significance). The pollen morphological attributes of the 26 studied plant taxa of the Thymelaeaceae family were subjected to Principal Component Analysis (PCA) and Hierarchical Component Analysis (HCA), which suggest four clusters. The cophenetic correlation coefficient also validates the four group of clustering. The study will significantly help in the precise pollen identification in fossil (pollen) records for the reconstruction of vegetation dynamics and contemporary climate change during the Quaternary Period in India. Moreover, the present study also helps in determining the phylogenetic and evolutionary trend on the basis of the presence or absence of exine ornamentation and having more or less apertures among the pollen grains of the 26 studied Thymelaeaceae taxa.

Notes on the female reproductive system of the red‐and‐black froghopper, Cercopis vulnerata Rossi, 1807 (Hemiptera: Cercopidae)‐light and electron microscopy studies

AbstractThe morphology of the female reproductive system in Cercopis vulnerata, an economically important species, was examined with light and scanning electron microscopy techniques. The female reproductive system of C. vulnerata has two ovaries, each containing 14 telotrophic meroistic ovarioles joined by two lateral oviducts that open in a common oviduct. In this type of ovarioles, the trophocytes within the germarium are transferred to the developing oocytes in the vitellarium via the nutritive cords. The oocytes in the vitellarium have three different development stages: previtello, vitello and choriogenesis. The ovarioles open in the lateral oviduct with a thin canal‐shaped pedicel. A pair of lateral oviducts opens distally into the common oviduct. The oviducts are surrounded by a monolayer cubical epithelium with deep folds toward the lumen and a thick muscle layer. A pair of spermatheca and a bursa copulatrix are seen near the common oviduct. Spermatheca has an elongated shape. The bursa copulatrix has an ovoid tube‐like structure. No work has been done on the female reproductive system of Cercopidae to date. This study was conducted to fill this gap and forms part of the female reproductive system morphology of this economically important insect.

Synergistic Action of MCL-1 Inhibitor with BCL-2/BCL-XL or MAPK Pathway Inhibitors Enhances Acute Myeloid Leukemia Cell Apoptosis and Differentiation.

Acute myeloid leukemia (AML) is a hematological malignancy characterized by excessive proliferation of abnormal myeloid precursors accompanied by a differentiation block and inhibition of apoptosis. Increased expression of an anti-apoptotic MCL-1 protein was shown to be critical for the sustained survival and expansion of AML cells. Therefore, herein, we examined the pro-apoptotic and pro-differentiating effects of S63845, a specific inhibitor of MCL-1, in a single-agent treatment and in combination with BCL-2/BCL-XL inhibitor, ABT-737, in two AML cell lines: HL-60 and ML-1. Additionally, we determined whether inhibition of the MAPK pathway had an impact on the sensitivity of AML cells to S63845. To assess AML cells' apoptosis and differentiation, in vitro studies were performed using PrestoBlue assay, Coulter electrical impedance method, flow cytometry, light microscopy and Western blot techniques. S63845 caused a concentration-dependent decrease in the viability of HL-60 and ML-1 cells and increased the percentage of apoptotic cells. Combined treatment with S63845 and ABT-737 or MAPK pathway inhibitor enhanced apoptosis but also induced differentiation of tested cells, as well as altering the expression of the MCL-1 protein. Taken together, our data provide the rationale for further studies regarding the use of MCL-1 inhibitor in combination with other pro-survival protein inhibitors.

Confocal interferometric scattering microscopy reveals 3D nanoscopic structure and dynamics in live cells

Bright-field light microscopy and related phase-sensitive techniques play an important role in life sciences because they provide facile and label-free insights into biological specimens. However, lack of three-dimensional imaging and low sensitivity to nanoscopic features hamper their application in many high-end quantitative studies. Here, we demonstrate that interferometric scattering (iSCAT) microscopy operated in the confocal mode provides unique label-free solutions for live-cell studies. We reveal the nanometric topography of the nuclear envelope, quantify the dynamics of the endoplasmic reticulum, detect single microtubules, and map nanoscopic diffusion of clathrin-coated pits undergoing endocytosis. Furthermore, we introduce the combination of confocal and wide-field iSCAT modalities for simultaneous imaging of cellular structures and high-speed tracking of nanoscopic entities such as single SARS-CoV-2 virions. We benchmark our findings against simultaneously acquired fluorescence images. Confocal iSCAT can be readily implemented as an additional contrast mechanism in existing laser scanning microscopes. The method is ideally suited for live studies on primary cells that face labeling challenges and for very long measurements beyond photobleaching times.

Pollen morphology of the endemic genera of the Madeira archipelago, Portugal

This study presents the first palynological characterisation of the five endemic plant genera of the Madeira archipelago: Chamaemeles Lindl, Melanoselinum Hoffm., Monizia Lowe, Musschia Dumort and Sinapidendron Lowe. Pollen grain morphology of ten endemic species was studied using light and scanning electron microscopy techniques. The size and shape of pollen grains, the polar axis, the equatorial diameter, and the exine ornamentation were measured and described. We found that the pollen grains of the five endemic genera are all medium-size monads. The close relative apiaceous Melanoselinum and Monizia differ in polar (P) and equatorial (E) diameter size and exine ornamentation while Sinapidendron species show differences in P, E, and P/E ratios. The pollen grains of the two Musschia species are very similar to each other, but differ in morphology and ornamentation from the Macaronesian endemic bellflowers Azorina vidalii and Canarina canariensis. This study unveiled differences between the endemic taxa and with their close related species, thus providing support to previous taxonomic findings.

New species of Pavlovophyceae (Haptophyta) and revision of the genera Exanthemachrysis, Rebecca and Pavlova

The justification of the 4 genera that currently compose the class Pavlovophyceae is based on a low number of species and a relative paucity of available, traceable and referenced cultures. Previous integrative phylogeny work revealed strains that can refine and strengthen our knowledge of the genera in the class. The application of multiple light and electron microscopy techniques allowed us to prioritize the cytomorphological characters (pyrenoid, thylakoid, stigma, knob-scales, life stage / life cycle) used for the taxonomy of these algae and to describe two new species: Exanthemachrysis fresneliae Véron sp. nov. and Rebecca billardiae Véron sp. nov. Consequently, revisions of the two genera Exanthemachrysis Lepailleur emend. Véron and Rebecca Green emend. Véron were made. In addition, the genus Pavlova Butcher emend Véron is revised in the light of these characters. Particular emphasis is placed on the life stages and habitat of the species.

Pollen morphological investigation of selected species of family Asteraceae from Pakistan by using light and scanning electron microscopy.

A light and scanning electron microscopy technique was conducted to investigate the palynological features of 19 species belonging to 15 genera of the family Asteraceae. The species under investigation yielded pollen in a range of shapes, including spheroidal, prolate, and subprolate. Trizoncolporate, Tricolporate, and Tetracolporate are the three types of pollen apertures observed in examined species. Except for Gazania rigens, which has reticulate ornamentation under SEM, the exine pattern in all studied species is echinate. The majority of the species had isopolar polarity, whereas some members were apolar and heteropolar. The quantitative parameters that are polar to equatorial diameter, P/E ratio, length of colpi, width of colpi, length of the spine, width of spine, and exine thickness was measured by using light microscopy. The Coreopsis tinctoria had the lowest mean polar 19.75 μm to mean equatorial diameter 18.25 μm while the Silybum marianum had the highest polar 44.7μm to equatorial 48.2μm value. The value for colpi length/width was highest in Cirsium arvensis and lowest in C. tinctoria, measuring 9.7/13.2μm and 2.7/4.7μm, respectively. The spine length varied from 0.5μm in Sonchus arvensis to 5.5μm in Calendula officinalis. Verbesina encelioides had the highest exine thickness measurement (3.3μm), whereas S. arvensis had the lowest (0.3μm). The pollen surface of Tagetes erectus, has the highest number (65) whereas the lowest spines number (20) is found in S. arvensis. A taxonomic key based on pollen traits is provided for quick identification of the species. Significant implications for the systematics of the Asteraceae family can be drawn from the reported pollen's quantitative and qualitative data. RESEARCH HIGHLIGHTS: SEM and LM technique was used to study the palynological features of pollen grains. Quantitative and qualitative pollen features of 19 species of family Asteraceae are described. Morphological traits of pollen grains are important for the systematics of family Asteraceae.

Peroxisomes Are Highly Abundant and Heterogeneous in Human Parotid Glands

The parotid gland is one of the major salivary glands producing a serous secretion, and it plays an essential role in the digestive and immune systems. Knowledge of peroxisomes in the human parotid gland is minimal; furthermore, the peroxisomal compartment and its enzyme composition in the different cell types of the human parotid gland have never been subjected to a detailed investigation. Therefore, we performed a comprehensive analysis of peroxisomes in the human parotid gland's striated duct and acinar cells. We combined biochemical techniques with various light and electron microscopy techniques to determine the localization of parotid secretory proteins and different peroxisomal marker proteins in parotid gland tissue. Moreover, we analyzed the mRNA of numerous gene encoding proteins localized in peroxisomes using real-time quantitative PCR. The results confirm the presence of peroxisomes in all striated duct and acinar cells of the human parotid gland. Immunofluorescence analyses for various peroxisomal proteins showed a higher abundance and more intense staining in striated duct cells compared to acinar cells. Moreover, human parotid glands comprise high quantities of catalase and other antioxidative enzymes in discrete subcellular regions, suggesting their role in protection against oxidative stress. This study provides the first thorough description of parotid peroxisomes in different parotid cell types of healthy human tissue.

Placentation in the Mexican scincid lizard Plestiodon brevirostris (Squamata: Scincidae).

Viviparity is the reproductive pattern in which females gestate eggs within their reproductive tract to complete their development and give birth to live offspring. Within extant sauropsids, only the Squamata (e.g., snakes, lizards, and amphisbaenians) evolved viviparity, representing 20% of the existing species. The genus Plestiodon is represented by 43 species and is one of the most widely distributed genera of the Scincidae in Mexico. The goal of this research has been to determine the placental morphology and ontogeny during gestation in the lizard Plestiodon brevirostris. Specimens were dissected to obtain the embryonic chambers and the embryos were categorized to carry out the correlation between the development stage and the placenta development. The embryonic chambers were processed using the conventional histological technique for light microscopy. The identified embryonic stages were 4, 29, 34, 36, and 39. A thin eggshell surrounds the egg in early developmental stages; however, this structure is already absent in the embryonic hemisphere during the developmental stage 29. The results revealed that P. brevirostris is a lecithotrophic species, but a close maternal-fetal relationship is established by tissue apposition. Ontogenically, the placental types that form in the embryonic hemisphere are the chorioplacenta, choriovitelline placenta, and chorioallantoic placenta; whereas the omphaloplacenta is formed in the abembryonic hemisphere. The structure of the chorioallantoic placenta in P. brevirostris suggests that it may play a role during gas exchange between the mother and the embryo, due to the characteristics of the epithelia that comprise it. The structure of embryonic and maternal epithelia of the omphaloplacenta suggests a role in the absorption of the eggshell during gestation and possibly in the transport or diffusion of some nutrients. In general, it is evident that ontogeny and placental characteristics of P. brevirostris match those of other species of viviparous lecithotrophic scincids.

Leaf and stem anatomical characterization of Euphorbia hirta L., a tolerant species to glyphosate

Euphorbia hirta L. is a weed species that is tolerant to the most used herbicide in agriculture, glyphosate. The anatomical characteristics of plants influence the processes of absorption and translocation of herbicides. The objective of this work was to characterize the anatomy of the aerial vegetative axis (leaves and stem) of E. hirta, to support the establishment of strategies for better control of this species with herbicides. The plants were grown in a greenhouse under controlled conditions. When they reached sizes between 8 and 12 cm, samples of stems and leaves were collected, fixed in FAA 50, and stored in 70% ethanol. Subsequently, the samples were processed following usual light microscopy techniques. In cross-section, the stem of E. hirta has a circular shape. The leaf epidermis is uniseriate composed of isodiametric cells of compact arrangement and with the presence of multicellular trichomes and anthocyanin. As for the morphometric parameters evaluated, the young leaves have a lower thickness in the abaxial epidermis. Based on the anatomical characteristics observed in E. hirta, the main barriers that can act in the absorption of herbicides are the high hairiness and the high content of anthocyanin in the epidermal cells.

HIV-1 transmission: modelling and direct visualization in the third dimension.

Identifying initial events of mucosal entry of human immunodeficiency virus type-1 (HIV-1) in laboratory-based, physiologically relevant and high-throughput contexts may aid in designing effective strategies to block local transmission and spread of HIV-1. Several paradigms have been posited for how HIV-1 crosses mucosal barriers to establish infection based on two dimensional (2D) culture-based or animal-based models. Nevertheless, despite these models stemming from 2D culture and animal studies, monolayers of cells poorly replicate the complex niche that influences viral entry at mucosal surfaces, whereas animal models often inadequately reproduce human disease pathophysiology and are prohibitively expensive. Organoids, having never been directly utilized in HIV-1 transmission investigations, may offer a compromise between 2D culture and animal models as they provide a platform that mimics the biophysical and biochemical niche of mucosal tissues. Importantly, observation of events downstream of viral inoculation is potentially accessible to researchers via an array of microscopy techniques. Because of the potential insights organoids may provide in this context, we offer this review to highlight key physiological factors of HIV-1 transmission at common mucosal sites and a discussion to highlight how many of these factors can be recapitulated in organoids, their current limitations and what questions can initially be addressed, particularly using a selective inclusion of quantitative light microscopy techniques. Harnessing organoids for direct observation of HIV-1 entry at mucosal sites may uncover potential therapeutic targets which prevent the establishment of HIV-1 infection.

"Ectomosphere": Insects and Microorganism Interactions.

This study focuses on interacting with insects and their ectosymbiont (lato sensu) microorganisms for environmentally safe plant production and protection. Some cases help compare ectosymbiont microorganisms that are insect-borne, -driven, or -spread relevant to endosymbionts' behaviour. Ectosymbiotic bacteria can interact with insects by allowing them to improve the value of their pabula. In addition, some bacteria are essential for creating ecological niches that can host the development of pests. Insect-borne plant pathogens include bacteria, viruses, and fungi. These pathogens interact with their vectors to enhance reciprocal fitness. Knowing vector-phoront interaction could considerably increase chances for outbreak management, notably when sustained by quarantine vector ectosymbiont pathogens, such as the actual Xylella fastidiosa Mediterranean invasion episode. Insect pathogenic viruses have a close evolutionary relationship with their hosts, also being highly specific and obligate parasites. Sixteen virus families have been reported to infect insects and may be involved in the biological control of specific pests, including some economic weevils. Insects and fungi are among the most widespread organisms in nature and interact with each other, establishing symbiotic relationships ranging from mutualism to antagonism. The associations can influence the extent to which interacting organisms can exert their effects on plants and the proper management practices. Sustainable pest management also relies on entomopathogenic fungi; research on these species starts from their isolation from insect carcasses, followed by identification using conventional light or electron microscopy techniques. Thanks to the development of omics sciences, it is possible to identify entomopathogenic fungi with evolutionary histories that are less-shared with the target insect and can be proposed as pest antagonists. Many interesting omics can help detect the presence of entomopathogens in different natural matrices, such as soil or plants. The same techniques will help localize ectosymbionts, localization of recesses, or specialized morphological adaptation, greatly supporting the robust interpretation of the symbiont role. The manipulation and modulation of ectosymbionts could be a more promising way to counteract pests and borne pathogens, mitigating the impact of formulates and reducing food insecurity due to the lesser impact of direct damage and diseases. The promise has a preventive intent for more manageable and broader implications for pests, comparing what we can obtain using simpler, less-specific techniques and a less comprehensive approach to Integrated Pest Management (IPM).