Safety of potable reuse can be enhanced by improved water quality monitoring techniques for assessing water treatment processes. This study evaluated the efficacy of online bacterial counting for ...continuous monitoring of reverse osmosis (RO) membranes to remove bacteria using real-time bacteriological commercial counters and an on-site pilot-scale RO system. Prior to on-site assessments, the online bacterial counting was verified by comparing the measurement of fluorescent particles in water with flow cytometry. During a seven day pilot test of RO treatment at a water reclamation plant, online bacterial counts in RO permeate were monitored below 15 counts/mL; whereas the bacterial counts in RO feed water were approximately 2500 to 10,000 counts/mL. Removal rates of bacterial counts ranged from 2.6 to 3.1-log (average = 2.9-log) by continuously monitoring bacterial removal. This is greater than a 2-log reduction frequently determined using other water quality surrogates (i.e., electrical conductivity). Overall, the continuous monitoring of bacteria in RO feed and permeate can be implemented without the addition of chemicals to provide near real-time bacterial counts to measure their reduction after RO treatment. This can be developed for continuous performance monitoring of the RO process, providing greater assurance of microbial water quality after RO treatment.
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•Efficacy of online bacterial counting for monitoring RO system integrity was assessed.•Real-time bacteriological commercial counters were used for this study's assessment.•Online bacterial counting was verified by measuring surrogates.•Bacterial counts in RO feed and permeate were monitored online at the pilot scale.•A range of 2.6 to 3.1-log reduction in bacteria was determined by online monitoring.
Fluorescence methods are widely used for the study of marine and freshwater phytoplankton communities. However, the identification of different microalgae populations by the analysis of ...autofluorescence signals remains a challenge. Addressing the issue, we developed a novel approach using the flexibility of spectral flow cytometry analysis (SFC) and generating a matrix of virtual filters (VF) which allowed thorough examination of autofluorescence spectra. Using this matrix, different spectral emission regions of algae species were analyzed, and five major algal taxa were discriminated. These results were further applied for tracing particular microalgae taxa in the complex mixtures of laboratory and environmental algal populations. An integrated analysis of single algal events combined with unique spectral emission fingerprints and light scattering parameters of microalgae can be used to differentiate major microalgal taxa. We propose a protocol for the quantitative assessment of heterogenous phytoplankton communities at the single-cell level and monitoring of phytoplankton bloom detection using a virtual filtering approach on a spectral flow cytometer (SFC-VF).
Photosynthetic organisms maintain optimum levels of photosynthetic pigments in response to environmental changes to adapt to the conditions. The identification of cyanobacteria strains that alleviate ...bleaching has revealed genes that regulate levels of phycobilisome, the main light-harvesting complex. In contrast, the mechanisms of pigment degradation in algae remain unclear, as no nonbleaching strains have previously been isolated. To address this issue, this study attempted to isolate nonbleaching strains of the unicellular red alga
after exposure to nitrogen (N)-depletion based on autofluorescence information. After four weeks under N-depletion, 13 cells from 500,000 cells with almost identical pre- and post-depletion chlorophyll a (Chl a) and/or phycocyanin autofluorescence intensities were identified. These nonbleaching candidate strains were sorted
a cell sorter, isolated on solid medium, and their post-N-depletion Chl a and phycocyanin levels were analyzed. Chl a levels of these nonbleaching candidate strains were lower at 1-4 weeks of N-depletion similar to the control strains, however, their phycocyanin levels were unchanged. Thus, we successfully isolated nonbleaching
strains in which phycocyanin was not degraded under N-depletion,
autofluorescence spectroscopy and cell sorting. This versatile method will help to elucidate the mechanisms regulating pigments in microalgae.
Low-dimensional (<10 nm) semiconductor quantum dots (QDs) have received great attention for potential use in biomedical applications (diagnosis and therapy) for which larger nanoparticles (>10 nm) ...are not suitable. Here, we demonstrate a green, biogenic synthesis route for making CdS QDs with 2–5 nm particle size using tea leaf extract (Camellia sinensis) as a toxic-free particle stabilizing agent. We explored the biological activity of these CdS QDs in different applications, namely, (a) antibacterial activity, (b) bioimaging, and (c) apoptosis of lung cancer cells. The antibacterial activity of the CdS QDs was studied against different types of bacteria growth, showing that CdS QDs effectively inhibit the bacterial growth and exhibit cytotoxicity toward A549 cancer cells when compared to a control (no QD treatment). We compared this cytotoxicity effect on A549 cancer cells with a standard drug, cisplatin, showing comparable results. Additionally, these CdS QDs produce high-contrast fluorescence images of A549 cancer cells indicating a strong interaction with the cancer cell. To further understand the role of CdS QDs in bioimaging and cytotoxicity effect in A549 cells, fluorescence emission and flow cytometry analyses were performed. The fluorescence emission of CdS QDs was recorded with λexc = 410 nm, showing concentration-dependent fluorescence emission centered at 670 nm. From the flow cytometry analysis, it is confirmed that the CdS QDs are arresting the A549 cell growth at the S phase of cell cycle, inhibiting further growth of lung cancer cell. The multifunctional advantages of C. sinensis extract-mediated green CdS QDs will be of widespread interest in implementing in vivo-based bioimaging and therapeutic cancer treatment applications.
The human gastrointestinal tract harbors a diverse and complex microbiome, which interacts in a variety of ways with the host. There is compelling evidence that gut microbial dysbiosis, defined as an ...alteration of diversity and abundance in intestinal microbes, is an etiological factor in inflammatory bowel disease (IBD). Membrane vesicles (MVs), which are nano-sized particles released by bacteria, have been found to interact with the host and modulate the development and function of the immune system. As a result MVs have been suggested to play a critical role in both health and disease. In this study we developed a method to isolate, characterize and assess the immunoreactivity of heterogeneous populations of MVs from fecal samples (fMVs) of healthy volunteers. We successfully isolated 2*10
9
-2*10
10
particles/ml from 0.5 gram of feces by using a combination of ultrafiltration and size exclusion chromatography (SEC) from 10 fecal samples. Bead-based flowcytometry in combination with tunable resistive pulse sensing (TRPS) provided a reliable method for (semi-)quantitative determination of fMVs originating from both Gram-positive and Gram-negative bacteria, while transmission electron microscopy confirmed the presence of fMVs. Real time 16s PCR on bacterial cell fractions or isolated fMVs DNA of the most common phyla (
Firmicutes, Bacteroidetes, Actinobacteria and Proteobacteria
) revealed differences in the relative abundance between bacteria and the fMVs. Moreover, fMVs evoke the release of TNF-α by THP-1 cells in a dose-dependent matter. Also, a significant positive correlation was found between Actinobacteria/γ-Proteobacteria derived vesicles and the release of TNF-α. It has become increasingly clear that fMVs could provide an additional layer to the definition of homeostasis or dysbiosis of the microbiota. The current study supports their potential involvement in the intestinal homeostasis or inflammatory disorders and provides putative interesting incentives for future research.
•A comprehensive study of peripheral and CSF immune cells in NMDAR encephalitis.•NMDAR encephalitis exhibited higher neutrophil count and MLR in peripheral blood.•CD3 + and CD4 + T cells in CSF of ...NMDAR encephalitis lowered than viral encephalitis.•Neurological status of NMDAR encephalitis was related to immune cell parameters.•Expression patterns of T lymphocyte subsets differed in NMDAR/ Viral encephalitis.
To investigate the immunopathogenic mechanisms of anti-N-methyl-D-aspartate receptor encephalitis (NMDAR-E) by characterizing the changes of immune cells in both peripheral blood (PB) and cerebrospinal fluid (CSF) of patients with NMDAR-E.
Cytology and flow cytometry were used to explore and compare different immunological parameters in PB and CSF of patients with NMDAR-E, viral encephalitis (VE) and healthy volunteers. Moreover, different models were established to assess the possibility of identifying NMDAR-E patients based on PB and CSF parameters.
The neutrophil counts and monocyte-to-lymphocyte ratios (MLR) in PB are higher in NMDAR-E patients than in both VEs and controls (P < 0.001, respectively), while the percentages of CD3 + T, CD4 + T lymphocytes, and the leukocytes count in CSF were lower in NMDAR-Es than in VEs (P < 0.01, respectively). The higher percentages of CD8 + T cells in blood and CSF were both correlated with more severe NMDAR-E (P < 0.05, respectively). The poor neurological status group had significantly higher PB leukocytes but lower CSF leukocyte count (P < 0.05). Longitudinal observations in patients with NMDAR-E showed a decreasing trend of leukocyte count, neutrophils count, neutrophil-to-monocyte ratios (NMR), and neutrophil-to-lymphocyte ratios (NLR) with the gradual recovery of neurological function.
The expression patterns of T lymphocyte subsets were different in patients with NMDAR-E and viral encephalitis. The changing trends of leukocyte and lymphocyte populations in peripheral blood and cerebrospinal fluid may provide clues for the diagnosis of different types of encephalitides, including NMDARE, and can be used as immunological markers to assess and predict the prognosis.
Flow cytometers are instruments used for the rapid quantitative analysis of cell suspension. Traditional flow cytometry uses multi-channel filters to detect fluorescence, whereas full-spectrum ...fluorescence based on dispersion detection is a more effective and accurate method. The application of various dispersion schemes in flow cytometry spectroscopy has been studied. From the perspective of modern detectors and demand for the miniaturization of flow cytometry, prism dispersion exhibits higher and more uniform light energy utilization, meaning that it is a more suitable dispersion method for small flow cytometers, such as microfluidic flow cytometers. Prism dispersion designs include the size, number, and placement of prisms. By deducing the formula of the final position of light passing through the prism and combining it with the formula of transmittance, the design criteria of the top angle and the incident angle of the prism in pursuit of the optimum transmittance and dispersion index can be obtained. Considering the case of multiple prisms, under the premise of pursuing a smaller size, the optimal design criteria for dispersion system composed of multiple prisms can be obtained. The design of prism dispersion fluorescence detection was demonstrated with a microfluidic flow cytometer, and the effectiveness of the design results was verified by microsphere experiments and practical biological experiments. This design criterion developed in this study is generally applicable to spectral flow cytometers.
Eriodictyol is an important flavonoid and is commonly present across the plant kingdom. Flavonoids have been reported to show incredible pharmacological potential. However, the anticancer activity of ...the important flavonoid eriodictyol has not been well reported. In the present study we determined its anticancer potential against the human lung cancer cell line A549.
The initial cytotoxicity induced by eriodictyol was measured by MTT assay. Flow cytometry was used to study the effects of eriodictyol on apoptosis, cell cycle phase distribution and mitochondrial membrane potential loss. The comet assay was used to measure DNA damage induced by eriodictyol in cancer cells while the western blot assay indicated effects of the compound on Bax/Blc-2 and PI3K/AKT/m-TOR proteins.
The results showed that eriodictyol has an IC
value of 50 μM against human lung cancer cells as compared to the IC
of 95 µM against non-cancerous FR2 cells. The molecule exerted its anticancer activity through induction of apoptosis by regulating the Bcl-2/Bax signalling pathway. It caused cell cycle arrest of human lung cancer A549 cells at G2/M phase. Eriodictyol was also found to cause a reduction of the mitochondrial membrane potential in a dose-dependent manner. Additionally, eriodictyol effectively inhibited the mTOR/PI3K/Akt signalling pathway in a dose-dependent manner.
Based on the above findings, we conclude that eriodictyol exerts its anticancer activity through induction of mitochondrial apoptosis and G2/M cell cycle arrest and inhibition of the TOR/PI3K/Akt cascade, indicating that it may have potential as a lead compound in the treatment of lung cancer, provided further in depth studies are done.
The beef industry continues to be interested in reliable rapid detection technologies for shiga toxin-producing Escherichia coli (STEC). Current rapid technologies require several hours of ...pre-enrichment and additional time on the rapid technology instrument. A flow cytometer-based system (RAPID-B®) has been shown to improve the turn-around for results with a more rapid pre-enrichment requiring only 6.5 h pre-enrichment for a 25 g and 8.5 h for a 375 g sample, followed by an additional 30 min time to achieve final results using the screening technology. The purpose of this study was to validate the RAPID-B® technology for non-O157 STEC detection as compared to the USDA-FSIS reference method which utilizes the BAX® system. A total of 180 STEC isolates from various sources and 20 non-STEC strains were used to evaluate specificity and sensitivity using the RAPID-B® flow cytometer. Also, three different weights (25, 325 and 375 g) of beef trim and ground beef samples were spiked with each STEC to verify detection sensitivity of BAX® system and RAPID-B® flow cytometer. For both methods, samples were confirmed by culturing using the USDA-FSIS reference method regardless of the screening result. The RAPID-B® flow cytometer showed that 180 isolates were all positive and the 20 non-STEC strains were all negative. For spiked beef samples, overall detection sensitivity was the same for both the BAX® system and RAPID-B® flow cytometer. When detection sensitivity was based on sample weight, there was no differences in 25 and 375 g samples between RAPID-B® flow cytometer and USDA-FSIS reference method. The RAPID-B® system yielded the same sensitivity as the reference method with a decrease of over 10 h of pre-enrichment time and 3 h of rapid screening detection time. In conclusion, the RAPID-B® flow cytometer based on whole cell detection generated similar results as BAX® system therefore the RAPID-B® flow cytometer system could be a valuable rapid method for the detection of non-O157 STEC in beef products.
•We compared the non-O157 STEC detection sensitivity between BAX® system and RAPID-B® flow cytometer.•The RAPID-B® flow cytometer exhibited similar results as BAX® system.•The RAPID-B® flow cytometer is a rapid detection method for non-O157 STEC in beef products.
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