-
Sensors (Basel, Switzerland) Oct 2023Colony-Forming Unit (CFU) counting is a complex problem without a universal solution in biomedical and food safety domains. A multitude of sophisticated heuristics and...
Colony-Forming Unit (CFU) counting is a complex problem without a universal solution in biomedical and food safety domains. A multitude of sophisticated heuristics and segmentation-driven approaches have been proposed by researchers. However, U-Net remains the most frequently cited and used deep learning method in these domains. The latter approach provides a segmentation output map and requires an additional counting procedure to calculate unique segmented regions and detect microbial colonies. However, due to pixel-based targets, it tends to generate irrelevant artifacts or errant pixels, leading to inaccurate and mixed post-processing results. In response to these challenges, this paper proposes a novel hybrid counting approach, incorporating a multi-loss U-Net reformulation and a post-processing Petri dish localization algorithm. Firstly, a unique innovation lies in the multi-loss U-Net reformulation. An additional loss term is introduced in the bottleneck U-Net layer, focusing on the delivery of an auxiliary signal that indicates where to look for distinct CFUs. Secondly, the novel localization algorithm automatically incorporates an agar plate and its bezel into the CFU counting techniques. Finally, the proposition is further enhanced by the integration of a fully automated solution, which comprises a specially designed uniform Petri dish illumination system and a counting web application. The latter application directly receives images from the camera, processes them, and sends the segmentation results to the user. This feature provides an opportunity to correct the CFU counts, offering a feedback loop that contributes to the continued development of the deep learning model. Through extensive experimentation, the authors of this paper have found that all probed multi-loss U-Net architectures incorporated into the proposed hybrid approach consistently outperformed their single-loss counterparts, as well as other comparable models such as self-normalized density maps and YOLOv6, by at least 1% to 3% in mean absolute and symmetric mean absolute percentage errors. Further significant improvements were also reported through the means of the novel localization algorithm. This reaffirms the effectiveness of the proposed hybrid solution in addressing contemporary challenges of precise in vitro CFU counting.
PubMed: 37837169
DOI: 10.3390/s23198337 -
Asian Pacific Journal of Allergy and... Dec 2023Cytokine-induced killer (CIK) cells are ex-vivo expanded T cells which present a phenotype of both T and Natural Killer cell properties.
BACKGROUND
Cytokine-induced killer (CIK) cells are ex-vivo expanded T cells which present a phenotype of both T and Natural Killer cell properties.
OBJECTIVE
To compare the proliferation and functional properties of human CIK cells cultured in three cell culture plasticwares.
METHODS
The number and viability of CIK cells were monitored. The expression of surface markers (CD3 and CD56), TH1 cytokines (IFN-γ and TNF-α), and cytolytic granules (granzyme B and perforin) were determined by flow cytometry.
RESULTS
The number of CIK cells cultured in a static bag was highest compared to those in a petri dish and gas-permeable flask. However, CIK cells cultured in all plasticwares similarity expressed surface marker, TH1 cytokines, and cytolytic granules.
CONCLUSIONS
Considering safety, efficacy, and cost, a static bag is the best plasticware for culturing CIK cells.
Topics: Humans; Cytokine-Induced Killer Cells; Cells, Cultured; Interferon-gamma; Cytokines; Cell Culture Techniques
PubMed: 33274953
DOI: 10.12932/AP-140720-0913 -
Scientific Reports Jul 2020Plant growth promoting rhizobacteria (PGPR) are a functionally diverse group of microbes having immense potential as biostimulants and stress alleviators. Their...
Plant growth promoting rhizobacteria (PGPR) are a functionally diverse group of microbes having immense potential as biostimulants and stress alleviators. Their exploitation in agro-ecosystems as an eco-friendly and cost-effective alternative to traditional chemical inputs may positively affect agricultural productivity and environmental sustainability. The present study describes selected rhizobacteria, from a range of origins, having plant growth promoting potential under controlled conditions. A total of 98 isolates (ectophytic or endophytic) from various crop and uncultivated plants were screened, out of which four endophytes (n, L, K and Y) from Phalaris arundinacea, Solanum dulcamara, Scorzoneroides autumnalis, and Glycine max, respectively, were selected in vitro for their vegetative growth stimulating effects on Arabidopsis thaliana Col-0 seedlings with regard to leaf surface area and shoot fresh weight. A 16S rRNA gene sequencing analysis of the strains indicated that these isolates belong to the genera Pseudomonas, Bacillus, Mucilaginibacter and Rhizobium. Strains were then further tested for their effects on abiotic stress alleviation under both Petri-plate and pot conditions. Results from Petri-dish assay indicated strains L, K and Y alleviated salt stress in Arabidopsis seedlings, while strains K and Y conferred increases in fresh weight and leaf area under osmotic stress. Results from subsequent in vivo trials indicated all the isolates, especially strains L, K and Y, distinctly increased A. thaliana growth under both normal and high salinity conditions, as compared to control plants. The activity of antioxidant enzymes (ascorbate peroxidase, catalase and peroxidase), proline content and total antioxidative capacity also differed in the inoculated A. thaliana plants. Furthermore, a study on spatial distribution of the four strains, using either conventional Petri-plate counts or GFP-tagged bacteria, indicated that all four strains were able to colonize the endosphere of A. thaliana root tissue. Thus, the study revealed that the four selected rhizobacteria are good candidates to be explored as plant growth stimulators, which also possess salt stress mitigating property, partially by regulating osmolytes and antioxidant enzymes. Moreover, the study is the first report of Scorzoneroides autumnalis (fall dandelion) and Solanum dulcamara (bittersweet) associated endophytes with PGP effects.
Topics: Arabidopsis; Arabidopsis Proteins; Bacteria; Endophytes; Gene Expression Regulation, Plant; Osmotic Pressure; Phylogeny; Plant Leaves; Plant Shoots; RNA, Ribosomal, 16S; Salt Stress; Soil Microbiology
PubMed: 32728116
DOI: 10.1038/s41598-020-69713-5 -
Current Opinion in Chemical Biology Feb 2020Genetically encoded fluorescent protein-based kinase biosensors are a central tool for illumination of the kinome. The adaptability and versatility of biosensors have... (Review)
Review
Genetically encoded fluorescent protein-based kinase biosensors are a central tool for illumination of the kinome. The adaptability and versatility of biosensors have allowed for spatiotemporal observation of real-time kinase activity in living cells and organisms. In this review, we highlight various types of kinase biosensors, along with their burgeoning applications in complex biological systems. Specifically, we focus on kinase activity reporters used in neuronal systems and whole animal settings. Genetically encoded kinase biosensors are key for elucidation of the spatiotemporal regulation of protein kinases, with broader applications beyond the Petri dish.
Topics: Animals; Biosensing Techniques; Fluorescence Resonance Energy Transfer; Fluorescent Dyes; Humans; Luminescent Proteins; Phosphotransferases
PubMed: 31911398
DOI: 10.1016/j.cbpa.2019.11.005 -
Environmental Health Insights 2022Indoor air quality determines the well-being of occupants. It has been linked to sick building syndrome and building-related diseases which lead to many socio-economic...
Indoor air quality determines the well-being of occupants. It has been linked to sick building syndrome and building-related diseases which lead to many socio-economic problems including reduced productivity and impaired learning. Indoor air quality problem is more serious for prisoners, due to their confinement and exposure condition. However, it has not been studied in our study setting. Thus, this study aimed to determine the indoor air microbial quality and associated factors in Jimma town prison administration, Southwestern Ethiopia. A cross-sectional study design was employed in August 2021. Data on the general condition of the prison rooms and occupancy were collected by trained data collectors using an observational checklist. The microbial sample was collected using a sterilized Petri dish. A total of 19 triplicate air samples were collected using Mannitol salt agar and Sabouroad dextrose agar media for the growth of and fungi respectively. Data were analyzed using SPSS version 23 and presented using tables and a graph. The effect of predictor variables on the microbial load was also analyzed by using linear regression. The finding of this study revealed that the microbial load of indoor air at Jimma town prison administration ranged from 891 to 15 439 and 315 to 3067 CFU/m³ for and fungi respectively. Both and the fungal load of the indoor environment were positively affected by the temperature of the room. Whereas, the floor space per inmate affects the concentration of alone. Almost all rooms of the prison administration had microbial load beyond the acceptable limit. Higher temperature, less floor space per inmate, bad floor cleanness conditions, inadequate ventilation, and dampness were contributing factors to the high load of and fungus. Thus, additional rooms are required to reduce overcrowding and keep room temperature.
PubMed: 36003416
DOI: 10.1177/11786302221118842 -
Plant Disease Mar 2021The panicle branch, which is the key node for transport of photosynthesis products from source to sink, is vulnerable to many diseases caused by fungal pathogens, such...
The panicle branch, which is the key node for transport of photosynthesis products from source to sink, is vulnerable to many diseases caused by fungal pathogens, such as Magnaporthe oryzae, Cochliobolus miyabeanus. Among these diseases, rice blast is the most important one which causes devastating losses in many regions. In 2019 and 2020, panicle branch rot of rice with a symptom which could be mistaken with rice blast was observed in a paddy field, where is not traditional epidemical region of rice blast, in Fuyang, Zhejiang province. In 2020, similar symptom was also observed in Hubei and Anhui Province. In a paddy field in Fuyang, the symptom appeared on more than 30% investigated panicles. Diseased panicle exhibited brown to black lesions on primary or secondary branches as well as pedicels, however the grain and the neck of spike could not be infected which is the most obvious difference with rice blast. Obviously, the disease can't destroy the entire function of branch and blank grain was rarely observed, so its damage is not comparable with neck blast. Normally, it caused incomplete grain filing commonly leading to 5% - 25% grain weight loss. During the booting stage of rice, local solar irradiation time and temperature were fewer and lower than common years which may be responsible for losses caused by this disease. After surface sterilized, lesion parts cut from infected branches from 25 panicle samples were cultured on 2% water agar at 28℃ for 24-28 h, and fungi were isolated and purified by mycelial tip transferring. Among 31 isolates, 26 showed similar cultural characters. The wool-like mycelia were luxuriant and grew rapidly on PDA spreading the whole 9 cm petri dish in less than a week at 28 ℃. The mycelia were white to ashen at beginning and gradually turned black from center of the plate after 5 days culture at 28 ℃. Hyphae were smooth, branched, septate, hyaline or pale brown. Conidia were single-celled, black, spherical to subspherical, and 10.2 to 14.6 × 12.2 to 15.7 μm (n=50) in dimension and born on tip of hyaline and ampulliform conidiophores. The fungus showed similar morphological characteristics with Nigrospora oryzae (1). ITS sequences of 6 representative strains of the fungus were amplified, sequenced with primer pair, ITS1/ITS4 (2), and submitted into GenBank with an accession number, MW228165. Phylogenetic analysis was conducted with sequences of reference strains (3). The result showed that the fungus obtained in this study was fallen into the same group with N.oryzae. In view of above both morphological and molecular analysis, the strains were finally identified as N. oryzae. Pathogenicity tests were conducted in triplicate with rice panicles in initial heading stage. Fifty panicles were wounded on branches with needles and inoculated by spreading the conidia suspension (10μl, 1 × 106 conidia ml-1) on the wounds. The panicles used as control were treated in same way with 10μl of sterile water. The inoculated and control plants were kept in dark, 25 ℃ and relative humidity of more than 85% for 24 h in culture chamber. Symptoms appeared on 44 of 50 inoculated panicles which were basically similar with those observed in paddy field, while negative controls remained symptomless. The fungi re-isolated from inoculated panicles were also confirmed as N. oryzae by both morphological and molecular analysis. To the best of our knowledge, this is the first report of N. oryzae causing panicle branch rot disease on Oryza sativa (rice). This disease not only cause yield losses and lower milling quality, but could also be mistaken as rice blast incurring unnecessary fungicides spray.
PubMed: 33728954
DOI: 10.1094/PDIS-11-20-2423-PDN -
Sensors (Basel, Switzerland) Aug 2021Automatic tracking of () in standard Petri dishes is challenging due to high-resolution image requirements when fully monitoring a Petri dish, but mainly due to...
Automatic tracking of () in standard Petri dishes is challenging due to high-resolution image requirements when fully monitoring a Petri dish, but mainly due to potential losses of individual worm identity caused by aggregation of worms, overlaps and body contact. To date, trackers only automate tests for individual worm behaviors, canceling data when body contact occurs. However, essays automating contact behaviors still require solutions to this problem. In this work, we propose a solution to this difficulty using computer vision techniques. On the one hand, a skeletonization method is applied to extract skeletons in overlap and contact situations. On the other hand, new optimization methods are proposed to solve the identity problem during these situations. Experiments were performed with 70 tracks and 3779 poses (skeletons) of . Several cost functions with different criteria have been evaluated, and the best results gave an accuracy of 99.42% in overlapping with other worms and noise on the plate using the modified skeleton algorithm and 98.73% precision using the classical skeleton algorithm.
Topics: Algorithms; Animals; Caenorhabditis elegans; Skeleton
PubMed: 34451062
DOI: 10.3390/s21165622 -
Toxins Oct 2021is known to produce relatively large amounts of alkaloids; however, their ecological consequences remain unclear. In this study, we evaluated the allelopathic potential...
is known to produce relatively large amounts of alkaloids; however, their ecological consequences remain unclear. In this study, we evaluated the allelopathic potential of the main alkaloids, including aloperine, matrine, oxymatrine, oxysophocarpine, sophocarpine, sophoridine, as well as their mixture both in distilled HO and in the soil matrix. Our results revealed that all the alkaloids possessed inhibitory activity on four receiver species, i.e., , , and . The strength of the phytotoxicity of the alkaloids was in the following order: sophocarpine > aloperine > mixture > sophoridine > matrine > oxysophocarpine > oxymatrine (in Petri dish assays), and matrine > mixture > sophocarpine > oxymatrine > oxysophocarpine > sophoridine > aloperine (in pot experiments). In addition, the mixture of the alkaloids was found to significantly increase the IAA content, MDA content and POD activity of seedlings, whereas CTK content, ABA content, SOD activity and CAT activity of seedlings decreased markedly. Our results suggest might produce allelopathic alkaloids to improve its competitiveness and thus facilitate the establishment of its dominance; the potential value of these alkaloids as environmentally friendly herbicides is also discussed.
Topics: Alkaloids; Allelopathy; Amaranthus; Lolium; Medicago sativa; Setaria Plant; Sophora
PubMed: 34678999
DOI: 10.3390/toxins13100706 -
Sensors (Basel, Switzerland) Jun 2022The measurement of biological tissues' dielectric properties plays a crucial role in determining the state of health, and recent studies have reported microwave...
The measurement of biological tissues' dielectric properties plays a crucial role in determining the state of health, and recent studies have reported microwave biosensing to be an innovative method with great potential in this field. Research has been conducted from the tissue level to the cellular level but, to date, cellular adhesion has never been considered. In addition, conventional systems for diagnosing tumor aggressiveness, such as a biopsy, are rather expensive and invasive. Here, we propose a novel microwave approach for biosensing adherent cancer cells with different malignancy degrees. A circular patch resonator was designed adjusting its structure to a standard Petri dish and a network analyzer was employed. Then, the resonator was realized and used to test two groups of different cancer cell lines, based on various tumor types and aggressiveness: low- and high-aggressive osteosarcoma cell lines (SaOS-2 and 143B, respectively), and low- and high-aggressive breast cancer cell lines (MCF-7 and MDA-MB-231, respectively). The experimental results showed that the sensitivity of the sensor was high, in particular when measuring the resonant frequency. Finally, the sensor showed a good ability to distinguish low-metastatic and high-metastatic cells, paving the way to the development of more complex measurement systems for noninvasive tissue diagnosis.
Topics: Cell Adhesion; Humans; MCF-7 Cells; Microwaves; Neoplasms
PubMed: 35746165
DOI: 10.3390/s22124383 -
Medical Mycology Journal 2021We postulated that disinfection of viable Trichophyton species in shoes would help reduce the number of patients with tinea pedis in Japan and that this might be...
We postulated that disinfection of viable Trichophyton species in shoes would help reduce the number of patients with tinea pedis in Japan and that this might be accomplished safely using volatile components of essential oils. As vapor of lemongrass (Cymbopogon citratus) oil and citral have strong antimicrobial activities against Trichophyton, we examined the conditions under which lemongrass oil or citral show optimal antimicrobial activity in shoes. First, we investigated whether or not a strong antimicrobial effect could be obtained by combining with terpene aldehydes or aromatic aldehydes. When combined with citral, perillaldehyde showed superior antimicrobial activity to citronellal, cinnamaldehyde, cuminaldehyde, hydroxycitronellal, and vanillin. The combined effects of citral and perillaldehyde against Trichophyton mentagrophytes, Bacillus subtilis, and Candida albicans as volatile components dotted on filter paper placed away from the petri dish inoculated with fungi or bacteria were examined. Citral (2.5 mg/mL) and perillaldehyde (2.5 mg/mL) showed a greater inhibitory effect on growth of C. albicans than either solution alone in the aromatogram (disc diffusion) descent method (fractional inhibitory concentration [FIC] index of 0.58). Citral (2.5 mg/mL) and perillaldehyde (1.25 mg/mL) vapors in a closed box synergistically inhibited growth of B. subtilis and T. mentagrophytes (FIC indexes of 0.5 and 0.38, respectively). These results suggested that this combination would be safe and useful for disinfection of shoes.
Topics: Anti-Infective Agents; Cymbopogon; Humans; Oils, Volatile; Perilla; Trichophyton
PubMed: 34853254
DOI: 10.3314/mmj.21-00011