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In the present study, and for the waste valorization, Moringa oleifera seeds-removed ripened pods (SRRP) were used for papersheet production and for the extraction of bioactive compounds. Fibers were characterized by SEM–EDX patterns, while the phytoconstituents in ethanol extract was analyzed by HPLC. The inhibition percentage of fungal mycelial growth (IFMG) of the treated Melia azedarach wood with M. oleifera SRRP extract at the concentrations of 10,000, 20,000, and 30,000 µg/mL against the growth of Rhizoctonia solani and Fusarium culmorum was calculated and compared with fluconazole (25 µg). The produced papersheet was treated with the ethanol extract (4000, 2000, and 1000 µg/mL) and assayed for its antibacterial activity against Agrobacterium tumefaciens, Erwinia amylovora, and Pectobacterium atrosepticum by measuring the inhibition zones and minimum inhibitory concentrations (MICs). According to chemical analysis of M. oleifera SRRP, benzene:alcohol extractives, holocellulose, lignin, and ash contents were 7.56, 64.94, 25.66 and 1.53%, respectively, while for the produced unbleached pulp, the screen pulp yield and the Kappa number were 39% and 25, respectively. The produced papersheet showed tensile index, tear index, burst index, and double fold number values of 58.8 N m/g, 3.38 mN m2/g, 3.86 kPa m2/g, and 10.66, respectively. SEM examination showed that the average fiber diameter was 16.39 µm, and the mass average of for elemental composition of C and O by EDX were, 44.21%, and 55.79%, respectively. The main phytoconstituents in the extract (mg/100 g extract) by HPLC were vanillic acid (5053.49), benzoic acid (262.98), naringenin (133.02), chlorogenic acid (66.16), and myricetin (56.27). After 14 days of incubation, M. oleifera SRRP extract-wood treated showed good IFMG against R. solani (36.88%) and F. culmorum (51.66%) compared to fluconazole, where it observed 42.96% and 53.70%, respectively. Moderate to significant antibacterial activity was found, where the minimum inhibitory concentration (MIC) values were 500, 650, and 250 µg/mL against the growth of A. tumefaciens, E. amylovora, and P. atrosepticum respectively, which were lower than the positive control used (Tobramycin 10 µg/disc). In conclusion, M. oleifera SRRP showed promising properties as a raw material for pulp and paper production as well as for the extraction of bioactive compounds.

The Golden trumpet Handroanthus chrysotrichus is a tree that presents beneficial health properties against various diseases. Thus, this study aims to verify the toxicity of H. chrysotrichus bark extract, observing the effects of exposure to this extract in mice. For this, mice were separated in groups: saline (sterile solution .9%); H. chrysotrichus crude extract (HCCE) 10; HCCE 50, and HCCE 100 mg. kg−1 (p.o.). We analyzed HCCE effects on acute (single exposure) and subchronic protocol (14 days exposure). After both exposures, acute, and subchronic, we collected samples from blood, brain, liver, and kidney tissues for biochemical evaluation. In addition, after subchronic exposure, we performed behavioral tests. Acute exposure caused an increase of lipid peroxidation in liver tissue. Moreover, we observed a significant carbonyl increase in liver and brain tissues from HCCE 50 mg. kg−1. Kidneys presented carbonyl increase in mice treated with the highest concentration. Besides, creatinine increased in the group of the acute exposure at HCCE 100 mg. kg−1. Total leukocyte count decreased in all concentrations tested. Sub-chronic exposure at HCCE 100 mg. kg−1 caused a decrease in the number of crossing and an increase in its self-grooming frequency in the open field test. In this exposure, the brain and liver had a significant increase in carbonyl levels in all concentrations. We concluded that H. chrysotrichus cause behavioral and biochemical alterations in mice. HCCE primary targets seem to be the liver, kidneys, and white cells.

Thalidomide causes teratogenic effects in several animal species and in humans. Accordingly, the World Health Organization banned thalidomide when mothers who took thalidomide during pregnancy delivered abnormal fetuses. After four decades, thalidomide underwent drug “re-purposing” since its antiangiogenic and immunomodulatory effects were therapeutic for multiple myeloma. There are no reports of thalidomide’s effects on prokaryotes, but it showed teratogenic effects in Arabidopsis thaliana, an ancestor of the plant kingdom. This proof of concept study clearly shows that thalidomide caused a significant and reproducible decrease in germination rate, nitric oxide (NO) production, and chlorophyll content of fennel plantlets. Thalidomide also induced the formation of abnormal fennel plantlets with stunting, wrinkling, and curling of fennel shoots and leaves. Notably, quantitative analysis showed that thalidomide caused a 50% increase in the formation of abnormal fennel plantlets and that these negative effects of thalidomide showed a 2.50- to 4-fold decrease when fennel seeds were co-incubated with an NO donor (Spermine NoNoate) or a stable cGMP analog 8-bromo Guanosine 3′,5′-cyclic monophosphate (8-Bromo-cGMP). This study is important because it confirms that thalidomide’s negative effects on fennel seed germination and growth are mediated by attenuation of NO and disruption of NO signaling. This reproducible model of thalidomide-induced, NO-dependent damage in a plant system can be used to further investigate the molecular mechanisms of thalidomide action in plants. Importantly, this study establishes a link between the evolution of development of higher plants and mammals.

Cervical cancer (CC) is a type of pelvic malignant tumor that severely threatens women's health. Current evidence suggests that IER5, as a potential radiosensitizer, promotes irradiation-induced apoptosis in CC tissues in patients undergoing chemoradiotherapy. IER5 has been shown to be involved in the G2/M-phase transition. In the present study, we used Cdc25B as the breakthrough point to explore the underlying mechanism of IER5 in the cell cycle regulation of radiation-damaged HeLa cells. IER5 was evidently upregulated after irradiation, but Cdc25B was significantly downregulated. In monoclonal IER5-silenced HeLa cells, irradiation-induced downregulation of Cdc25B was attenuated. The effect of irradiation on Cdc25B promoter activity was determined by dual-luciferase reporter assays. The response elements on the Cdc25B promoter related to irradiation were predicted by JASPAR. These conserved sequences were mutated individually or in combination by splicing-by-overlap extension PCR, and their function was confirmed by dual-luciferase reporter assays. The enrichment efficiency of transcription factors after irradiation was determined by chromatin immunoprecipitation (ChIP) assay. Both Sp1/Sp3 and NF-YB binding sites were involved in irradiation-mediated regulation of Cdc25B. IER5 was involved in irradiation-mediated regulation of Cdc25B through the NF-YB binding site. Furthermore, ChIP assays showed that IER5 bound to the Cdc25B promoter, and the binding of IER5 to the Cdc25B promoter region in irradiation-induced HeLa cells induced the release of the coactivator p300 through interaction with NF-YB. Taken together, these findings indicate that IER5 is the transcriptional repressor that accelerates the downregulation of Cdc25B expression after irradiation.

Malaria and leishmaniasis are the public health problems in southern Iran. The main activity of vector control is indoor residual spraying using pyrethroids, using different insecticides as larviciding and impregnated bednets. The aim of study was to evaluate the biological assays of bendiocarb wettable powder (WP) at different surfaces of wall. The residual effect of bendiocarb WP80 at 400 milligram/meter square (mg/m2) was evaluated on various local surfaces of rooms such as mud and plaster as well as thatch roofs and wooden. World Health Organization standard cones using contact bioassays were carried out using laboratory reared sugar-fed, 48–72 h old females of Anopheles stephensi. Contact bioassays were carried out on sprayed surfaces for 150 days. Contact bioassay on surfaces treated with bendiocarb WP80 at different surfaces was estimated about 2 months. Fumigant tests of bendiocarb WP80 at 400 mg/m2 revealed 50–93.83% mortality with 1 month persistency. The results showed that carbamate insecticide could be used as rotation with pyrethroids for malaria vector control. Monitoring and evaluation of environmental toxicology of pesticides is important for decision making for choosing appropriate pesticides for disease vector control.

Infections caused by resistant microorganisms are a complex global public health challenge, and the way to combat the increase of resistance is the development of more modern and faster techniques for resistance detection. This study aimed to evaluate the transport of inactivated bacteria impregnated in a filter paper disk to detect carbapenem resistance genes by multiplex real-time PCR (qPCR) using high-resolution melting (HRM). A total of 88 isolates of 10 different species of Enterobacterales harboring well-characterized carbapenem resistance genes were evaluated. A full 10-µL loop of fresh growth of bacteria were impregnated in a filter paper disk, which was left at room temperature for 2 days in order to simulate the time spent in transportation. Bacterial inactivation was performed with 70% ethanol at 15 min. Afterwards, the DNA was extracted from the paper disks for further analysis by qPCR HRM. The time of 15 min in 70% ethanol was enough to inactivate all the isolates tested. It was possible to correctly identify the presence of the carbapenem resistance gene by HRM qPCR in 87 isolates (98.87%) that were transported in the filter paper disks. Our results indicated that it is possible to use filter paper to transport inactivated bacteria and to identify carbapenem resistance genes by qPCR HRM. This alternative tends to facilitate the access to this technology by many laboratories which do not have the qPCR equipment.

Recent studies have evaluated alternatives to the use of live animals in colony health monitoring. Currently, an alternative method that is suitable for all rack types and that has been verified to detect the infectious agents most commonly excluded from mouse colonies is unavailable. We compared the use of filter paper placed on the inside floor of mouse cages to the traditional use of sentinel mice in the detection of several prevalent murine pathogens including mouse hepatitis virus (MHV), murine norovirus (MNV), minute virus of mice (MVM), mouse parvovirus (MPV), Theiler murine encephalomyelitis virus (TMEV), Helicobacter spp., Syphacia obvelata, and Aspiculuris tetraptera. Experimental groups comprised 7 cages containing either 2 pieces of filter paper on the cage floor or 2 ICR sentinel mice. Soiled bedding from pet-store mice was transferred to the experimental cages weekly for 8 wk. At 1 and 2 mo after bedding transfer, the filter papers were evaluated by PCR and sentinel mice were tested by serology and fecal PCR. Filter papers detected all pathogens as effectively (MHV, MNV, MPV, MVM, TMEV S. obvelata, and A. tetraptera) or more effectively (Helicobacter spp.) than sentinel mice at both time points. Filter papers more readily detected pathogens with a high copy number per RT-PCR analysis than a low copy number. Helicobacter spp. were not detected by sentinel mice at either time point. These results indicate that the use of filter paper placed on the interior floor of empty mouse cages and exposed to soiled bedding is efficient in detecting bacteria, endoparasites, and most of the common mouse viruses included in an animal health monitoring program.

Benzene metabolites (HQ and BQ) are toxic compounds and their presence in human cause alteration in cellular respiration and kidney damage. In the current study, Saccharomyces cerevisiae has been used as a model organism and acute exposure of hydroquinone (HQ) decreased cell growth and increased reactive oxygen species (ROS). The expression of apoptosis regulatory genes (YCA1, NUC1, YSP1 and AIF1) were increased with HQ exposure in the wild-type cells. HQ exposure in the wild-type cells altered both the phospholipid and neutral lipid levels. Phosphatidylcholine is a vital membrane lipid that has a vital role in membrane biogenesis and was increased significantly with HQ. The neutral lipid results were supported with lipid droplets data and mRNA expression study. The phospholipid knockouts (Kennedy pathway) accumulated neutral lipids via the CDP-DAG (cytidine-diphosphate-diacylglycerol) pathway genes both in the presence and absence of HQ.

Chlorpyrifos (CPF) is an organophosphate insecticide commonly used to treat fruit and vegetable crops. CPF can cause severe adverse effects on body organs including the liver and central nervous system. This study investigated the CPF-induced inflammation in mice and explored the role of intestinal flora changes in liver inflammation. Adult C57BL/6 male mice were exposed to a CPF of 0.01-, 0.1-, 1- and 10-mg/kg bodyweight for 12 weeks. The mice in experimental group given CPF solution dissolved in corn oil vehicle by gavage, was administered by intraoral gavage for 5 days per week for 12 weeks. Histopathological examination and inflammatory factor detection were performed on mice liver tissue. Faeces were used for 16S ribosomal RNA high-throughput sequencing to explore the impact of CPF on intestinal flora structure and diversity. The results showed that 1- and 10-mg/kg CPF caused different degrees of liver focal inflammation. The structure of intestinal flora changed significantly in mice including the decreased beneficial bacteria (Akkermansia, Prevotella and Butyricimonas) and increased pathogenic bacteria (Helicobacter and Desulfovibrio). Meanwhile, the results of Q-RT-PCR showed that there was more total bacterial DNA in the liver tissue of the mice treated with 10-mg/kg groups. In conclusion, the imbalance of intestinal flora, the decreased abundance of beneficial bacteria and the increased abundance of pathogenic bacteria, as well as the increase of total bacterial DNA in the liver tissues, maybe associated with the liver focal inflammation induced by CPF.

Changes in ecosystems after the removal of stress provide a rich source of information for conservation science. We used a long-term regional data set on the performance of white sucker (Catostomus commersonii) collected before and after the closure of a pulp mill to explore recovery in fish. Physiological indicators, including liver enzymes and plasma steroids, showed some compelling changes after the closure of the mill consistent with reduced exposure to pulp mill effluent but did not unequivocally demonstrate recovery. However, persistent signals in these fish may indicate effects of impoundment or discharge of sewage. We also used quantile regression with environmental covariates and bootstrap iteration to determine if systematic variation remained in relative body weight, liver weight, and gonad weight. In fish formerly exposed to pulp mill effluent, we found evidence of improvements (male gonad weight and liver weight of males and females), degradation (gonad weight of females), and no change (body weight). Although the observed patterns may be associated with closure of the mill, some differences were also found at regional locations, suggesting roles of additional stressors and challenging the clear association of change at the Mattagami River exposure site with the closure of the mill. However, fish captured at this location show responses consistent with regional locations, suggesting no residual impacts and highlights the challenges of identifying changes in fish even after large and known interventions. Environ Toxicol Chem 2021;40:162-176. © 2020 SETAC.

In this study, Rhododendron luteum and Rhododendron ponticum were evaluated as raw material for pulp and paper production. 12 different sodium borohydride (NaBH4) added cooking trials were performed for each sample and kraft method was used for pulp production. Pulp properties, such as yield, kappa number and viscosity, and physical properties, such as breaking length and burst index, were determined for each trial. Besides, the effects of active alkali and NaBH4 on the pulp and paper properties were also examined. Optimum cooking conditions were obtained by using 18 % active alkali for NaBH4-free cooking experiments and 0.5 % NaBH4 and 18 % active alkali for NaBH4-added cooking experiments. In NaBH4-added pulping condition, the screened yield, kappa number and viscosity of R. luteum were found to be 43.4 %, 40.1 and 949 cm3/g1, respectively. The respective values for R. ponticum were 41.9 %, 44.5 and 885 cm3/g1. The screened yields of R. luteum and R. ponticum increased by about 2.8 % and 5.3 %, respectively, with 5 % addition of NaBH4 compared to NaBH4-free cooking experiments. Furthermore, with the addition of NaBH4, the kappa numbers decreased while the viscosity increased. The physical properties of the produced papers were also improved by using NaBH4 in cooking liquor. According to the obtained results, it was found that R. luteum and R. ponticum species can be evaluated for pulp and paper production., U ovom je radu istražena mogućnost uporabe biljaka Rhododendron luteum i Rhododendron ponticum kao sirovine za proizvodnju celuloze i papira. Za svaki uzorak provedeno je 12 različitih ispitivanja kuhanja s natrijevim borhidridom (NaBH4), a celuloza je proizvedena kraft postupkom. Za svako ispitivanje određena su svojstva celuloze poput prinosa, kappa broja i viskoznosti, te fizička svojstva kao što su duljina lomljenja i indeks pucanja papira. Osim toga, ispitani su učinci aktivne lužine i NaBH4 na svojstva celuloze i papira. Optimalni uvjeti kuhanja postignuti su upotrebom 18 % aktivne lužine za eksperimentalno kuhanje bez NaBH4 i upotrebom 0,5 % NaBH4 i 18 % aktivne lužine za eksperimentalno kuhanje s dodatkom NaBH4. U proizvodnji celuloze iz biljke R. luteum s dodatkom NaBH4 utvrđeno je da prinos prosijavanja iznosi 43,4 %, da je kappa broj 40,1, a viskoznost 949 cm3/g, dok su vrijednosti za celulozu dobivenu iz biljke R. ponticum redom 41,9 %, 44,5 i 885 cm3/g. Prinos prosijavanja biljaka R. luteum i R. ponticum uz dodatak 5 % NaBH4 povećao se oko 2,8 % i 5,3 % u usporedbi s eksperimentalnim kuhanjem bez dodatka NaBH4. Nadalje, uz dodatak NaBH4 smanjuju se kappa brojevi, a viskoznosti se povećavaju. Fizička svojstva proizvedenih papira također se poboljšavaju dodavanjem NaBH4 tekućini za kuhanje. Iz dobivenih je rezultata utvrđeno da se biljke R. luteum i R. ponticum mogu upotrebljavati u proizvodnji celuloze i papira.

Objective To report PCR results and vaccination status of rabbits with rabbit haemorrhagic disease following an investigation into sudden or unexpected death. Materials and methods PCR testing for RHDV2 and RHDV1 was performed on rabbit liver samples at two laboratories. Laboratory A reported results as positive or negative; Laboratory B reported results quantitatively as RNA copies per mg liver, categorised as negative, inconclusive or positive. The vaccination status of rabbits with both histopathological features of rabbit haemorrhagic disease and positive PCR test results were collated. Results PCR results matched histopathological findings in 188 of 195 (96%) cases. Seven individuals showed equivocal results, all of which had histopathological features of RHD but three tested PCR-negative and four results conflicted between laboratories. RHDV2 was the serotype detected in all PCR-positive cases. Histological features of rabbit haemorrhagic disease and PCR test results were positive in 125 rabbits; 51 unvaccinated, 56 in-date with Nobivac Myxo-RHD and 13 vaccinated against RHDV2 - although nine of these were vaccinated within 10 days of death. Clinical significance PCR testing complements histopathology in cases of sudden death in rabbits by confirming the diagnosis and identifying virus serotype, but there can be false negatives. Although RHDV2 is currently prevalent in UK pet rabbits, vaccination against both RHDV1 and RHDV2 is recommended. Failures of RHDV2 vaccine are infrequent.

Objectives To report clinical features, gross post mortem and histopathological findings from an investigation into sudden or unexpected death in rabbits that was undertaken during an outbreak of rabbit haemorrhagic disease. Materials and methods Using a standard protocol, veterinarians were invited to submit case histories and results of their post mortem examination of pet rabbits that died unexpectedly. Histopathological examination of heart, lungs, liver, spleen and kidney samples was collated with macroscopic appearance and clinical details. Results Hepatocellular necrosis, characteristic of rabbit haemorrhagic disease, was observed in 185 of 300 (62%) submissions, often accompanied by glomerular thrombosis and changes in other organs. Evidence of rabbit haemorrhagic disease was not apparent on histopathology in 113 of 300 (38%) rabbits. Gross post mortem examination by veterinary practitioners did not always reflect reported histopathological changes. No macroscopic abnormalities were seen in 78/185 (42%) of rabbit haemorrhagic disease cases. Rapid death and death of other rabbits in the household were common features of rabbit haemorrhagic disease. Ante mortem clinical signs included anorexia, collapse, lethargy, seizures, icterus, bleeding from the mouth, dyspnoea, hypothermia, pyrexia, bradycardia or poor blood clotting. Clinical importance Rabbit haemorrhagic disease can be suspected from a history of sudden death, especially if multiple rabbits are affected. There is not always macroscopic evidence of the disease but histopathology is useful to support or refute a diagnosis of rabbit haemorrhagic disease and provide information about other causes of death.

Polygalacturonase (PG) activity in plants can serve as an important index for plant disease. However, the conventional method to detect PG activity is a complex process and requires a skilled technician and expensive analytical equipment. In this study, a paper-based colorimetric sensor was developed based on the principle of the ruthenium red (RR) dye method for easy and simple measurement of PG activity. The proposed paper-based sensor has a three-layer structure for detection of PG activity in samples. The sensor sensitivity was enhanced by optimizing the pH of the sodium acetate buffer used in polygalacturonic acid (PGA)-RR complex formation and the reaction temperature for PG and the PGA-RR complex. Further, for quantitative analysis of PG activity, Delta RGB analysis was conducted to detect color changes in the sensing window of the sensor. Results presented that the linear measurement range of the paper sensor was 0.02-0.1 unit with the limit of detection of 0.02 unit, which showed a similar detection range, but a lower detection limit, compared to the spectrophotometry. Furthermore, PG activity based on culture condition was measured using samples from Sclerotium cepivorum to verify the potential application of the developed paper-based sensor in the field. The measured activity showed no statistically significant difference from the values obtained from the spectrophotometry at 95% confidence level. Therefore, the paper-based colorimetric sensor can be used to predict plant diseases in Allium crops during the stage of pathogen invasion, potentially contributing to the improvement of crop production.

A bleached kraft pulp mill discharging effluent to the Mattagami River in northern Ontario, Canada, closed after almost 90 yr of operation. During its operation, effluent from the mill influenced biota in the downstream areas. To assess shifts in the reliance of biota from mill-derived nutrients, the isotopic composition (δ13 C and δ15 N) of white sucker (Catostomus commersoni) muscle and whole mayflies (Hexagenia sp.) were compared before (1990s) and after the pulp mill's closure (2012-2014). To better understand other potential sources of spatial and temporal change, samples from 3 other tributaries in the basin with dams, ongoing pulp mill operations, sites receiving sewage, and at several reference sites were collected and compared. Irrespective of time period, biota collected at sites downstream of both dams and active pulp mills tended to have elevated δ13 C values, but variable changes in δ15 N (negligible in most cases) when compared with upstream samples. The isotopic composition of mayflies varied at reference sites over time, with decreasing values of δ13 C and δ15 N (mayflies only) with increasing depth, and there was evidence of lower δ13 C in fish after the pulp mill closure. Overall, these results suggest the importance of long-term, regional-scale measurements for documenting the effects of stressors on nutrient use by aquatic species. Environ Toxicol Chem 2020;39:1207-1218. © 2020 SETAC.

espanolObjetivo. Los patogenos se pueden transmitir a los billetes debido a los habitos antihigienicos personales. El objetivo del estudio fue buscar los posibles patogenos en los billetes que circulan en el mercado y tambien observar su resistencia antibacteriana asi como sus diversos factores de virulencia utilizando metodos genotipicos y fenotipicos. Material y metodos. Se recogieron al azar un total de 150 muestras de billetes entre agosto de 2017 y marzo de 2018. Se utilizaron los sistemas VITEK para la identificacion y las pruebas de sensibilidad a los antimicrobianos, respectivamente. Los genes de resistencia a los antimicrobianos (mecA, van, betalactamasas de espectro ampliado [BLEA] y carbapenemasas) y los genes de virulencia estafilococica (SE, pvl y tsst -1) se determinaron mediante PCR a tiempo real. Resultados. Se detecto la presencia de cepas de Staphylococcus aureus, Staphylococcus coagulasa negativos (SCN), Enterococcus spp, bacterias gramnegativas, bacterias gramnegativas no fermentativas y Candida spp en un 48%, 54,7%, 56%, 21,3%, 18,7% y 4% de los billetes, respectivamente. Se observo la presencia de S. aureus resistente a meticilina, Enterococcus resistentes a vancomicina y gramnegativos productores de BLEA en un 46,8%, 1,3% y 28,7%, respectivamente. Los genes Pvl, tsst-1 y SE se encontraron en un 2,8/4,9%; 1,4/1,2% y 100/87,8% de las cepas de S. aureus/SCN, respectivamente. El gen sea fue el gen enterotoxigenico mas frecuente. Los genes blaTEM, blaSHV, blaCTX-M-2, blaCTX-M-1, blaKPC, y blaOXA-48 se encontraron 55,8%, 46,5%, 41,2%, 18,6%, 18,6%, y 18,6%, respectivamente en cepas gramnegativas. Conclusion. Estos resultados son muy importantes para resaltar el estado higienico de los billetes. De este modo, los billetes pueden contribuir a la propagacion de patogenos y de la resistencia a los antimicrobianos. Por lo tanto, es posible que debamos comenzar a utilizar productos alternativos a los billetes. EnglishObjetives. Pathogens can be transmitted to banknotes due to the personal unhygienic habits. The aim of study was to find the possible pathogens on the banknotes circulating in the market and also to present their antibacterial resistance and their various virulence factors using genotypic and phenotypic methods. Material and methods. A total of 150 samples of banknotes were randomly collected between August 2017 and March 2018. VITEK systems were used for identification and antimicrobial susceptibility testing respectively. Antimicrobial resistance genes (mecA, van, extended-spectrum β-lactamase [ESBL] and carbapenemases) and staphyloccoccal virulence genes (staphyloccoccal enterotoxins [SEs], pvl, and tsst-1) were determined using with real-time PCR. Results. Staphylococcus aureus, coagulase-negative staphylococci (CoNS), Enterococcus spp., Gram-negative enteric bacteria, non-fermentative Gram-negative bacteria and Candida spp. were detected 48%, 54.7%, 56%, 21.3%, 18.7%, and 4%, respectively. Methicillin-resistant S. aureus, vancomycin-resistant enterococci and ESBL producing Gram-negative were found 46.8%, 1.3%, and 28.7%, respectively. Pvl, tsst-1, and SEs genes were found in a 2.8/4.9%, 1.4/1.2%, and 100/ 87.8% of the S. aureus/CoNS strains, respectively. The sea gene was found the most common enterotoxigenic gene. blaTEM, blaSHV, blaCTX-M-2, blaCTX-M-1, blaKPC, and blaOXA-48 were found 55.8%, 46.5%, 41.2%, 18.6%, 18.6%, and 18.6%, respectively in Gram-negative strains. Conclusion. These results is very important to highlight hygienic status of paper currencies. This can be considered as an indication that banknotes may contribute to the spread of pathogens and antimicrobial resistance. Therefore, we may need to start using alternative products instead of banknotes.

Cytochrome P450 (CYP450), and in particular CYP3A4, is the most abundantly expressed CYP450 isozyme implicated in many drug-drug and medicinal plant-drug interactions. Therefore, incorporation of CYP3A4 enzyme screening at an early stage of drug discovery is preferable in order to avoid enzymatic interactions. Here we present for the first time a paper-based CYP3A4 immobilized sol-gel-derived a platform using resorufin benzyl ether as a fluorogenic enzyme substrate used to investigate enzyme activity. The fluorescence intensity of the product can be simply quantified by using a handheld digital microscope and an image analysis software. The limit of quantitation was 0.35 μM with good precision (RSDs 0.05), while offering simplicity and lower cost. Kinetic parameters of the immobilized CYP3A4 in sol-gel coated paper were calculated from the Lineweaver-Burk plot, including Michaelis constant (Km) and maximum velocity (Vmax), which were 2.71 ± 0.35 μM and 0.43 ± 0.05 μM/min, respectively. Moreover, a functional test of these devices was conducted by assessments of known CYP3A4 inhibitors (i.e. ketoconazole, itraconazole) and inducers (i.e. phenytoin, carbamazepine). To further demonstrate the broad range of uses, the devices were utilized to assay plant extracts i.e. Areca catechu seeds, Camellia sinensis leaves, Eclipta prostrata aerial part, providing results in good agreement with previous studies. Furthermore, the sol-gel immobilized enzyme stored at 4 °C can increase storage stability, offering the activity of 86.3 ± 0.4% after 3-weeks storage, equivalent to the activity of the free enzyme solution after 1-week storage. The developed paper-based devices offer versatility, portability and low-cost.

Ethnopharmacobotanical information reports that Parkinsonia aculeata infusion is used to control diabetes-related complications and dyslipidemia. However, few studies are reported on the safe use of this species. The aim of this study is to evaluate the acute toxicity, embryotoxicity and cytotoxicity of a polar fraction obtained from hydroethanolic extract of P. aculeata (PfrHEPA). For the acute toxicity test, we considered the Up and Down method which the guidelines are described by the Organization for Economic Cooperation and Development (OECD N°425). The animals were treated with PfrHEPA (2000 mg/kg) or with distilled water (10 ml/kg) by gavage and observed from Day 1 to14. For embryotoxicity assay, zebrafish embryos were exposed to PfrHEPA (100 mg/L) and toxicity parameters were observed during four consecutive days. The cytotoxicity of PfrHEPA (5, 10, 25, 50, 75 and 100 μg/ml, respectively) was performed on normal cell lines (mesenchymal stem cells, African green monkey renal cells and mouse pre-adipocytes 3 T3-L1 using the MTT salt reduction assay. In the acute toxicity test, no mortality was observed in mice treated with PfrHEPA (2000 mg/kg), as well as behavioral changes, histopathological abnormalities and hematological and biochemical variables. In the embryotoxicity test, no abnormal changes related to the toxicological parameters were observed in the period of 96 h. Regarding the cytotoxicity assay, PfrHEPA showed no cytotoxic effect on the normal cell lines tested, with an IC50 value > 100 μg/ml. These results suggest the safe use of P. aculeata, however, more trials are needed for PfrHEPA to be presented as new safe therapeutic proposal for the control of metabolic disorders.

East Coast fever is caused by Theileria parva, and poses serious concerns for dairy farmers owing to massive economic losses. In the current study, we compared three methods (DNA extraction kits, FTA-NaOH and FTA-TENT) of DNA extraction to identify the most economical and reliable method. A survey for T. parva prevalence was conducted in dairy cattle in Mbarara, Uganda. Cytochrome C oxidase subunit I (COI) and T. parva-p104 genes were amplified to compare the methods. FTA-NaOH-based polymerase chain reaction (PCR) yielded the best detection rate for both COI gene and p104 gene. Prevalence of T. parva was 45.0% and 83.3% at animal and farm-level, respectively. FTA-NaOH based-PCR is simple, highly sensitive and cost-effective tool for T. parva diagnosis in resource constrained settings.

The increasing prevalence of antibiotic-resistant bacteria needs rapid identification and efficient destruction routes. This study proposes testing paper derived from electrospun fibrous mats and aggregation-induced emission (AIE) probes for trace sensing and simultaneous destruction of antibiotic-resistant E. coli. Aptamers are conjugated on fibers for selective capture of E. coli, and the capture capability can be regenerated via rinsing with salt solution. Hydroxyl tetraphenylethene (TPE) is linked with two cephalosporin molecules to construct TPE-Cep probes, and the fluorescence emission is turned on specifically in the presence of β-lactamase, which is a critical marker for screening resistant bacteria. Fibrous mats are lit up only in the presence of antibiotic-resistant bacteria, and the fluorescence intensity changes could be statistically fitted into an equation for quantitative analysis. Fibrous strips display apparent color changes from blue to green for a visual readout of bacterial levels, and the limit of detection (LOD) is much lower than those of previous paper substrates. In addition, the TPE-Cep probes could produce reactive oxygen species (ROS) under room light illumination to kill the captured bacteria. Thus, the integration of aptamer-grafted electrospun fibers and functional AIE probes provides potential for selective capture, trace imaging and photodynamic destruction of antibiotic-resistant bacteria.

Loop-mediated isothermal amplification or LAMP has been identified to be an efficient technology for point-of-care diagnostics. Paper-based LAMP technique has tremendous potential in replacing the existing tube-based technology as the manufacturing cost of a paper-based device is comparatively lower and easy-to-use. LAMP-based paper diagnostic device for Mycobacterium tuberculosis (MTB) detection is of extreme importance as it will help in early and rapid diagnosis of the affected patients. The fabrication of these devices requires assessment of design parameters on the extent of LAMP amplification reaction. Hence, CFD studies would be extremely beneficial from the design perspective. The current work presents an insight into the CFD simulations for LAMP amplification reaction on a porous paper membrane (nitrocellulose membrane). The convection-diffusion-reaction model is solved on a COMSOL Multiphysics 5.0 platform. Studies on effect of pore size, aspect ratio and initial DNA concentration on the extent of DNA amplification reaction have been carried out. The current paper-based technique is effective in detecting a minimum of 5 copies of DNA contrasting the previous semi-quantitative technique which demonstrated the detection of minimum 98 copies. Overall, the simulation results displayed almost 96% enhancement in the DNA amplification rate on paper membrane. Graphical abstract Graphical abstract for the computational study of DNA amplification reaction via LAMP technique on a porous paper membrane.

The gut microbiota is recognized as having major impact in health and disease. Sample storage is an important aspect to obtain reliable results. Mostly recommended is immediate freezing, however, this is not always feasible. Faecal occult blood test (FOBT) papers are an appealing solution in such situations, and most studies find these to be applicable, showing no major changes within 7 days storage at room temperature (RT). As fieldwork often requires RT storage for longer periods, evaluation of this is warranted. We performed 16S rRNA gene sequencing of 19 paired faecal samples immediately frozen or kept five weeks and five months at RT on FOBT papers. Alpha-diversity evaluation revealed no effect of FOBT storage, and evaluation of beta-diversity showed that host explained 65% of community variation, while storage method explained 5%. Evaluation of community dispersion and the Firmicutes/Bacteroidetes ratio revealed a larger effect of storage time for fresh-frozen samples. Single taxa evaluation (order-to-genus level) showed significant alterations of four (of 37) genera after five weeks and five genera after five months. When comparing the two timepoints, alterations were only detectable for fresh-frozen samples. Our findings reveal that long term storage on FOBT papers is an applicable approach for microbiota research.

Pseudoalteromonas (BB2-AT2) is a ubiquitous marine heterotroph, often associated with labile organic carbon sources in the ocean (e.g. phytoplankton blooms and sinking particles). Heterotrophs hydrolyze exported photosynthetic materials, components of the biological carbon pump, with the use of diverse metalloenzymes containing zinc (Zn), manganese (Mn), cobalt (Co), and nickel (Ni). Studies on the metal requirements and cytosolic utilization of metals for marine heterotrophs are scarce, despite their relevance to global carbon cycling. Here, we characterized the Zn, Mn, Co, and Ni metallome of BB2-AT2. We found that the Zn metallome is complex and cytosolic Zn is associated with numerous proteins for transcription (47.2% of the metallome, obtained from singular value decomposition of the metalloproteomic data), translation (33.5%), proteolysis (12.8%), and alkaline phosphatase activity (6.4%). Numerous proteolytic enzymes also appear to be putatively associated with Mn, and to a lesser extent, Co. Putative identification of the Ni-associated proteins, phosphoglucomutase and a protein in the cupin superfamily, provides new insights for Ni utilization in marine heterotrophs. BB2-AT2 relies on numerous transition metals for proteolytic and phosphatase activities, inferring an adaptative potential to metal limitation. Our field observations of increased alkaline phosphatase activity upon addition of Zn in field incubations suggest that such metal limitation operates in sinking particulate material collected from sediment traps. Taken together, this study improves our understanding of the Zn, Mn, Co, and Ni metallome of marine heterotrophic bacteria and provides novel and mechanistic frameworks for understanding the influence of nutrient limitation on biogeochemical cycling.

Up-regulation of NF-kB and depletion of glutathione are acceptable mechanisms of Ifosfamide (IFO) renal toxicity. This investigation elucidates the role of free radicals, inflammatory and apoptotic markers in ifosfamide-induced rentoxicity and the protection of Annona species planted in Egypt (squamosa, cherimolia), and the hybrid between them (atemoya). Annona seeds extracts were prepared and phytochemical analyses were assessed. Rats were injected with saline or with IFO alone or in combination with Annona squamosa or Annona cherimolia or Annona atemoya orally. Biochemical, gene expression, histological and immune-histological examinations were performed. The results demonstrated that IFO elevated creatinine, sodium, magnesium and urea, along with depleted serum potassium and albumin levels. IFO caused a significant reduction in renal GSH, significant increases in renal MDA, and NOx, and up-regulated iNOS. In addition, IFO treatment showed increase mRNA and protein expression of NF-kB, while down-regulated mRNA and protein expression of Bcl-2, and strong immunohistological expression of caspase-3 as well as BAX in kidney tissues. Different Annona seed extracts significantly enhanced the sharpness of renal injury and improved oxidant responses induced by IFO. Annona extracts down-regulated iNOS, NF-kB expressions, as well as, down-regulated caspase-3 and BAX immunohistological expressions, and up-regulated Bcl-2 expression. A. squamosa showed a more pronounced protective effect. Histopathological examination was in accordance with biochemical results. Recent results suggest that Annona species ameliorated the renal toxicity of ifosfamide by down-regulation of NOx, iNOS, NF-kB, BAX, caspase-3 and MDA in addition to up-regulation of GSH and Bcl-2.

Bacillus cereus is one of the most common foodborne pathogens found in various kinds of staple foods such as rice and wheat. A rapid and accurate detection method for this pathogen is highly desirable for the sustainable production of relevant food products. While several classical and molecular-based detection methods are available for the identification of B. cereus, they suffered one or more limitations such as the requirement for a tedious and time-consuming process, less than ideal specificity, and the lack of portability. Herein, we developed the first paper-based sensing device that exhibits high species specificity with sufficiently low limit of detection for the visual detection of specific DNA sequences of B. cereus. The success is attributed to the strategic planning of fabrication in various dimensions including thorough bioinformatics search for highly specific genes, the use of the pyrrolidinyl peptide nucleic acid (PNA) probe whose selectivity advantage is well documented, and an effective PNA immobilization and DNA-binding visualization method with an internal cross-checking system for validating the results. Testing in rice matrices indicates that the sensor is capable of detecting and distinguishing B. cereus from other bacterial species. Hence, this paper-based sensor has potential to be adopted as a practical means to detect B. cereus in food industries.

This study was aimed to evaluate toxicity in repeated doses for 28 days, reproductive toxicity and cytotoxicity of a polar fraction obtained from the hydroethanolic extract of Parkinsonia aculeata (PfrHEPA) in experimental models. To perform the toxicity test in repeated doses for 28 days, male and female Wistar rats were treated via orogastric for 28 days with PfrHEPA (35, 70 or 140 mg/kg) according to the guidelines established by the Organisation for Economic Co-operation and Development (OECD) number 407 (1995). For assessment, the impact of PfrHEPA on the reproductive output various parameters were measured, including maternal weight, no. of pregnant females, female fertility index (%), gestation lengthtime, implantation sites, litter size and placental index of test animals. The cytotoxicity of PfrHEPA was performed on the tumor lines NCI-H292 (human lung carcinoma), HL-60 (human promyelocytic leukemia) and HCT-116 (colorectal cancer). In the repeated dose toxicity test for 28 days, no mortality was observed in the male and female rats treated with PfrHEPA as well as morphological changes and biochemical and hematological parameters. In the reproductive toxicity test, no abnormalities were observed related to the toxicological parameters in both mothers and offspring. Regarding the cytotoxicity assay, the PfrHEPA fraction did not demonstrate significant cytotoxic effect on the cell lines analyzed. The present results suggest the use of PfrHEPA is safe and well tolerated in rats. Further studies are planned to identify and purify the active compounds for subsequent in vivo evaluation.

The present work reports the biobleaching effect on OPEFB pulp upon utilisation of extracellular xylano-pectinolytic enzymes simultaneously yielded from Bacillus amyloliquefaciens ADI2. The impacts of different doses, retention times, pH, and temperatures required for the pulp biobleaching process were delineated accordingly. Here, the OPEFB pulp was subjected to pre-treatment with xylano-pectinolytic enzymes generated from the same alkalo-thermotolerant isolate that yielded those of higher quality. Remarkable enhanced outcomes were observed across varying pulp attributes: for example, enzyme-treated pulp treated to chemical bleaching sequence generated improved brightness of 11.25%. This resulted in 11.25% of less chlorine or chemical consumption required for obtaining pulp with optical attributes identical to those generated via typical chemical bleaching processes. Ultimately, the reduced consumption of chlorine would minimise the organochlorine compounds found in an effluent, resulting in a lowered environmental effect of paper-making processes overall as a consequence. This will undoubtedly facilitate such environmentally-friendly technology incorporation in the paper pulp industry of today.

Significance: Real-time monitoring of the heart rate and blood flow is crucial for studying cardiovascular dysfunction, which leads to cardiovascular diseases. Aim: This study aims at in-depth understanding of high-speed cardiovascular dynamics in a zebrafish embryo model for various biomedical applications via frequency-comb-referenced quantitative phase imaging (FCR-QPI). Approach: Quantitative phase imaging (QPI) has emerged as a powerful technique in the field of biomedicine but has not been actively applied to the monitoring of circulatory/cardiovascular parameters, due to dynamic speckles and low frame rates. We demonstrate FCR-QPI to measure heart rate and blood flow in a zebrafish embryo. FCR-QPI utilizes a high-speed photodetector instead of a conventional camera, so it enables real-time monitoring of individual red blood cell (RBC) flow. Results: The average velocity of zebrafish’s RBCs was measured from 192.5 to 608.8 μm∕s at 24 to 28 hour-post-fertilization (hpf). In addition, the number of RBCs in a pulsatile blood flow was revealed to 16 cells/pulse at 48 hpf. The heart rates corresponded to 94 and 142 beats-per-minute at 24 and 48 hpf. Conclusions: This approach will newly enable in-depth understanding of the cardiovascular dynamics in the zebrafish model and possible usage for drug discovery applications in biomedicine. National Research Foundation (NRF) Published version Funding: National Research Foundation of the Republic of Korea (NRF-2012R1A3A1050386, NRF-2020R1A2C2102338, NRF-2020R1A2C210233811, and NRF-2021R1A4A1031660); KAIST UP Program; Singapore National Research Foundation (NRF-NRFF2015-02).

AIMS: The utilization of micro‐organisms in pulp and paper industries has proved biobleaching technology as an environmentally friendly alternative to the conventional approach. In this paper, the effect of actinobacterial fermentation broth on pulp biobleaching has been investigated. METHODS AND RESULTS: Actinobacterial colonies were isolated from lignocellulose‐rich soil samples and screened for xylanase production and bleaching activity. The most efficient isolate in bleaching activity showed 100% similarity to Streptomyces rutgersensis. Pulp treatment with 5‐day fermentation broth of this strain showed up to 7% increase in brightness (30°C for 6 h, pH (5–7)) compared to untreated (control) pulp. Also, after 60 min biotreatment, significant reduction (12·5%) in consumption of bleaching chemicals was achieved to obtain final brightness of 55%. CONCLUSION: Actinobacterial fermentation broth can be considered as a rich source of effective biobleaching agents which may be considered as environmental friendly and cost‐effective technique in comparison with traditional method. SIGNIFICANCE AND IMPACT OF THE STUDY: Our findings showed ability of S. rutgersensis UTMC 2445 in bleaching chemomechanical paper pulp. Also, two strains of Saccharothrix, a rare actinobacterium, with biobleaching activity were introduced. In the proposed method, there is no need to use purified enzymes, and biobleaching process can be done using the fermentation broth.

Driven by a bright prospect for rapid, portable and cost-effective point-of-care testing, an assembled Pasteur pipette device to integrate nucleic acid extraction, amplification and detection was developed to detect B. cereus in a sample-to-answer format. Denaturation Bubble-mediated Strand Exchange Amplification (SEA) was chosen to perform isothermal amplification because it requires only a pair of primers and one Bst DNA polymerase. The established SEA can detect as low as 1.0 × 10−13 M genomic DNA of B. cereus, which was comparable with the previously reported method for B. cereus detection. The assembled Pasteur pipette allows sample-to-answer diagnostic in a simple, low-cost, portable, and disposable format. The inherent function of Pasteur pipette enables direct liquid handling without the need of extra pipettes, syringes or pumps. Visual readout was achieved by using a pH sensitive dye, further simplifying result judgment process. The detection limit for B. cereus is 1.0 × 104 CFU/mL in pure cultures, while the detection limit in artificially contaminated milk is 1.0 × 105 CFU/mL without enrichment and 1.0 × 100 CFU/mL following 12 h enrichment. Considering that typical cell counts in food samples associated to food poisoning are 1.0 × 105 to 1.0 × 108 CFU per gram/milliliter B. cereus, our Pasteur pipette is enough sensitive for answer-to-sample diagnosis of B. cereus even directly from foods without enrichment. The whole diagnostic procedure could be completed within 50 min, dramatically decreasing the detection time. In a word, the assembled Pasteur pipette device, combined with a homemade metal bath, possesses great potential for sample-to-answer application in resource-limited settings.

Objectives To assess the prevalence of canine parainfluenza virus, canine adenovirus type 2, canine distemper virus, canine respiratory coronavirus and influenza virus A infections in: (1) privately-owned or, (2) kennelled dogs showing signs consistent with canine infectious respiratory disease and, (3) clinically healthy dogs in Vienna, Austria. Materials and methods Prospectively, nasal and tonsillar swabs from 214 dogs affected with infectious respiratory disease, and 50 healthy control dogs were tested for nucleic acids specific to the various viral infections. Concurrent bronchoalveolar lavage fluid from 31 dogs with chronic respiratory disease was investigated for the same viral pathogens. Additionally, anti-canine respiratory coronavirus antibody concentrations were measured in paired blood samples from 30 acutely diseased dogs. Results Canine respiratory coronavirus (7.5%) and canine parainfluenza virus (6.5%) were the most commonly detected viruses in samples from the upper airways of dogs with respiratory infections. Serological results showed a significant seroconversion in response to coronavirus in 50% of the examined cases. None of the samples was positive for influenza virus A-specific nucleic acid. Canine coronavirus-specific nucleic acid was detected in 4.0% of healthy dogs. Clinical significance Canine coronavirus should be considered as a clinically relevant cause of infectious respiratory disease in crowded dog populations. For sample collection, the nasal mucosa can be recommended as the favoured site. Analysis of paired serum samples aids verification of canine coronavirus infection in respiratory disease.

In this study, an internal circulation-anoxic/aerobic (IC-A/O) process followed by ultrafiltration (UF) and reverse osmosis (RO) system was applied for paper wastewater reclamation. The IC-AO system presented a stable and efficient performance, achieving high removal of chemical oxygen demand (COD), total organic carbon (TOC) and total nitrogen (TN) with methane production rate of 132.8 mL/d. Acute toxicity to Daphnia magna (D. magna) was reduced significantly (83.2%) and the spearman's rank correlation analysis indicated that the toxicity of effluents from each reactor were positively correlated with COD and TOC. Hexadecanoic acid, octadecanoic acid and benzophenone were the main toxic contributors for biological effluent. Microbial community revealed that Anaerolinea was significantly related with organic pollutants. The UF-RO system further removed pollutants and toxicity with the final effluent COD, TOC, ammonium nitrogen (NH4+-N) and TN of 32.6, 18.8, 0.3 and 9.2 mg/L, respectively, which proved that it was feasible for paper wastewater reuse. This study presented an efficient, practical and environmentally competitive system, and paved a foundation for the treatment and reuse of paper wastewater.

Managing food contamination from bacteria has been an ongoing issue in the public health and industrial fields. Enzymatic substrates possessing optical properties, e.g. fluorescence or color manifestation, are widely exploited in pathogenic/non-pathogenic bacteria culture methods. Recently, various chromogenic substrates have been utilized in the development of point-of-care diagnostic tools. Herein, four types of chromogenic substrates were exploited to develop paper-based sensors for major foodborne pathogens. We designed a compact sized three-dimensional paper device with a simple user interface. By inserting functional layers in the middle of multilayers, pre-lysis and pH regulation steps were excluded and the analysis time was subsequently reduced, while only one sample droplet was needed for the whole analysis process. After the enzymatic reactions had proceeded, target-specific colors appeared. When it was combined with enrichment, 101 cfu mL-1 of pathogens were successfully detected in 4-8 hours, while those in milk samples were readily sensed in 12 hours. The proposed bacteria sensor exhibited great advantages of low cost, portability and simple operation, while showing a respectable limit-of-detection as low as 101 cfu mL-1 and below. Significantly, we emphasize that it takes fewer steps than existing methods and provides a reduced analysis time owing to the layer functionalization.

Identification of Cronobacter represent a major challenge for laboratories testing powdered infant formula (PIF). In the present study, two biochemical galleries and three molecular methods have been applied to confirm 276 Cronobacter spp. and non-Cronobacter isolates from different sources. Using the latest database of API 20 E and ID 32 E biochemical miniaturized kits, 53% and 78% of the isolates were identified respectively. From the available results, total accuracy for Cronobacter detection was in 97% (API 20 E) and 99% (ID 32 E). The three molecular methods were based on rRNA based lateral flow, Real Time PCR combined with either a hybridization or hydrolysis probe. For all three methods total accuracy was more than 99%. A pilot trial using Next Generation Sequencing (NGS) correctly identified 58 out of 66 isolates (88%) in DNA mixtures. The results indicate that the commercially available approaches such as ID 32 E, rRNA based lateral flow and Real Time PCR are all suitable for Cronobacter identification at the genus level. The NGS method may become a suitable alternative in the future, provided that the sequence database is improved.

In recent years, increasing efforts have focused on production of organic-inorganic compound fertilizers using ammonium sulfite pulping waste liquor. However, their ecological effects on soil have not been studied. In this study, earthworm Eisenia fetida was exposed to various doses (0, 0.13, 0.26 and 0.52 kg/m2) for different time (7, 14, 21, and 28 d) to evaluate the effects of fertilizers made from pulping waste liquor, including reactive oxygen species (ROS) generation, antioxidant enzymes activities, glutathione S-transferase enzyme (GST) activity, malondialdehyde (MDA) content and DNA damage. Results showed that there were significant increase of ROS and MDA levels after 14 d, inducing production of antioxidant enzymes (superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT)) as well as GST. Before 14 d, excess ROS and MDA caused damage to the DNA of earthworms, leading to gradual increase of Olive tail moment (OTM) in the comet assay. With the exposure time extended to 28 d, owing to the combined effects of elimination of free radicals by antioxidant enzymes and detoxification enzymes as well as self-repairing function of cells, ROS and MDA levels declined slightly and OTM gradually decreased. In summary, this study indicated that there was a toxicological effect on earthworms when fertilizers made from pulping waste liquor were applied to soil, which needs more attention.

Alcoholism has been linked to problems with male reproductive function. The combined effects of alcohol, cannabis, and tobacco were compared in this study. A total of 35 rats were assigned randomly into seven groups A–G: animals in A were administered distilled water. Animals in B–G were either administered alcohol orally (30 ml 40% alcohol) or exposed to smoke from ignited tobacco (exposure to smoke from 0.7 g tobacco for 5 min) or cannabis (exposure to smoke from 0.7 g tobacco and cannabis for 5 min): B (orally administered alcohol), C (exposed to the smoke from tobacco), D (exposed to smoke from cannabis), E (treated with alcohol and exposed to smoke from tobacco), F (treated with alcohol and exposed to smoke from cannabis), G (treated with alcohol and exposed to smokes from tobacco and cannabis). Assays were carried on the testicular homogenate after a 14-day treatment. There was a significant increase in activity of alkaline phosphatase (P ≤ 0.05), concentrations of cholesterol, glutathione reductase, and malondialdehyde in treated rats by the co-administration of alcohol with cannabis and tobacco compared with the control group. The combined treatment also caused degeneration and morphological distortions of testicular cells. The biochemical and histoarchitectural change was due to oxidative damage attributable to the synergistic effects. The high binding energy of tetrahydrocannabinol ligand to prostate acid phosphatase may be a prediction that the ligand can have an inhibitory effect on the function of enzymes in the prostate.

Highlights • The alkaline – HC miscanthus fibers lignin content was decreased by 41.54% • Pulp fiber shape characteristics were negatively influenced due to cavitation force. • Results could verify the efficacy of alkaline - HC pretreatment as a delignification method. • The alkaline – HC process displayed similar results with other conventional methods. • Miscanthus grass fibers could potentially substitute hardwood fibers in pulp blends., One way of satisfying increased market demand and simultaneously achieving a reduced environmental load in the industrial paper production is the use of fibrous agricultural residues. The aims of this study were i) to investigate the effect of alkaline – hydrodynamic cavitation (HC) pre-treatments on the delignification of Miscanthus × giganteus stalks (MGS) and ii) establishing the suitability of MGS as feedstock and their exploitation in pulp and paper manufacturing. It was demonstrated that the proposed treatment is an efficient delignification method for the non-wood fiber sources, such as miscanthus. A significant outcome of this work was the observation that HC treatment preserved the fibres lengths and surface quality of raw MGS, but at the same time increased the amount of kinked and curled fibers present in cavitated miscanthus fibers. The average miscanthus fiber length was found to be relatively short at 0.45 (±0.28) mm, while the slenderness ratio, the flexibility coefficient and Runkel ratio values were calculated to be 28.13, 38.16 and 1.62, respectively. The estimated physical properties of MGS pulp hand-sheets were 24.88 (±3.09) N m g−1 as the tensile index, 0.92 (±0.06) kPa m2 g−1 as the burst index and 4.0 (±0.37) mN m2 g−1 as the tear index. Overall the current work demonstrated effective use of hydrodynamic cavitation for improving the processing in pulp and paper manufacturing.

Qing Hao Gan Cao (QHGC), a Chinese medicinal formula containing Artemisia annua and Glycyrrhizae Radix et Rhizoma, has been used to treat sunstroke and as an antiviral agent for more than 800 years. It has not previously been subject to a toxicological safety evaluation in acute and subacute (28 days) studies. Therefore, the acute and subacute toxicity of an aqueous extract of QHGC were evaluated in vivo. For the QHGC preparation, the botanical raw materials were crushed into pieces and mixed in the ratio of 10:1 in distilled water for 12 h, then boiling three times for 2 h each time. The three decoctions were mixed and filtered, then spray-dried with hot air at 160°C for 30 min, and stored at room temperature. For the acute toxicity test, 72.0 g/kg of QHGC extract was administered by gavage to male and female mice. Body weight, general observations, and autopsy results were recorded. No mortality or toxicity signs were observed during the studies. For the subacute toxicity test, 4.0, 8.0, or 16.0 g/kg/day of QHGC extract was administered to rats for 28 days. General observations and mortality, body weight, biochemical and hematological parameters, organ weight, and pathological morphology were analyzed. The acute and subacute toxicity studies did not show significant changes in body weight, general observations, hematology and biochemical parameters, organ weight, and liver, spleen, stomach, duodenum, testis, ovary, lung, heart, and kidney histopathological analyses. The consumption of QHGC aqueous extract can be considered safe within the conditions of this study.

The use of natural substances derived from traditional Chinese medicine and natural plants as safe radiosensitizing adjuvants is a new trend for cancer radiotherapy. Ganoderma lucidum has been used as a traditional Chinese medicine with a history of more than 2000 years. Ganoderic acid T (GAT) is a typical triterpene of G. lucidum, which has strong cytotoxicity to cancer cells, but whether it has radiation sensitization effect has not been explored. In this work, we treated the HeLa cells with different concentrations of GAT before exposure to gamma-ray radiation and investigated its influence on the radiosensitivity. The cell viability, apoptosis rate, necoptosis rate, intracellular ATP level, cell cycle, the amount of H2AX and 53BP1, reactive oxygen species, and mitochondrial membrane potential were examined. Apoptotic, necroptotic, and autophagic biomarker proteins, including caspase 8, cytochrome c, caspase 3, RIPK, MLKL, P62, and LC3, were analyzed. As a result, we confirmed that with treatment of GAT, the gamma-ray radiation induced both apoptosis and necroptosis in HeLa cells, and with increase of GAT, the percentage ratio of necroptosis was increased. The involved pathways and mechanisms were also explored and discussed.

Significance: Infrared (IR) inhibition can selectively block peripheral sensory nerve fibers, a potential treatment for autonomic-dysfunction-related diseases (e.g., neuropathic pain and interstitial cystitis). Lowering the IR inhibition threshold can increase its translational potentials. Aim: Infrared induces inhibition by enhancing potassium channel activation. We hypothesized that the IR dose threshold could be reduced by combining it with isotonic ion replacement. Approach: We tested the IR inhibition threshold on the pleural-abdominal connective of Aplysia californica. Using a customized chamber system, the IR inhibition was applied either in normal saline or in isotonic ion-replaced saline, which could be high glucose saline, high choline saline, or high glucose/high choline saline. Each modified saline was at a subthreshold concentration for inhibiting neural conduction. Results: We showed that isotonically replacing ions in saline with glucose and/or choline can reduce the IR threshold and temperature threshold of neural inhibition. Furthermore, the size selectivity of IR inhibition was preserved when combined with high glucose/high choline saline. Conclusions: The present work of IR inhibition combined with isotonic ion replacement will guide further development of a more effective size-selective IR inhibition modality for future research and translational applications.

We characterised fibres and papers of microfibrillated silk from Bombyx mori produced by mechanical and enzymatic process. Milling increased the specific surface area of fibres from 1.5 to 8.5 m2/g and that enzymatic pre-treatment increased it further to 16.5 m2/g. These fibrils produced a uniform, significantly strong (tenacity 55 Nm/g) and stiff (Young's modulus > 2 GPa) papers. Enzymatic pre-treatment did not reduce molecular weight and tensile strength of papers but significantly improved fibrillation. Silk remained highly crystalline throughout the fibrillation process. Protease biodegradation was more rapid after fibrillation. Biodegradation was impacted by structural change due to enzymatic pre-treatment during the fibrillation. Biodegraded silk had much higher thermal degradation temperature. The unique combination of high strength, slow yet predicable degradation and controllable wicking properties make the materials ideally suited to biomedical and healthcare applications.

In this article, we aimed to investigate the anti-aging activity of Lantana camara ethanolic leaves-extract in Drosophila melanogaster: survival and longevity (life span). L. camara leaves were collected and washed thoroughly of sand particles, air-dried, ground, and extracted by the maceration method using ethanol as a solvent. Phytochemical screening was carried out. 168-hour LC50 was determined by exposing fruit flies to concentrations ranging from 5 to 5000 mg/10 g diet for 7-days. Twenty-eight-day survival and longevity studies were carried out by administering L. camara ethanolic leaves extract at 5, 10, and 20 mg/10 g diet to 1–3 days old fruit flies. Each concentration was replicated four times with 50 fruit flies each. The emergence rate of young fruit flies from eggs laid by fruit flies administered L. camara leaves-extracts were also carried out. The total yield of the extraction was determined to be 18%. Phytochemical analysis revealed the presence of alkaloids, Flavonoids, Phenol, steroids, cardiac glycosides, and carbohydrates. 168-hour LC50 of L. camara was also determined to be 1135 mg/10 g diet. L. camara significantly prolonged (P

Pomegranate (Punica granatum L.) is a fruit used extensively in traditional medicine by ancient and modern cultures. Different parts of the tree and fruit, such as leaf, peel, pericarp, aril, seed, and juice contain considerable amounts of phenolic compounds with high antioxidant activities. To improve its storability, pomegranate juice was microencapsulated by spray drying. The present study evaluated microencapsulated pomegranate juice (MPJ) for toxic effects in Wistar rats and CD-1 mice to determine if MPJ can be considered safe for human consumption and used as a nutraceutical. No deaths or deleterious effects occurred when high doses of 5000 mg/kg were orally administered in rats for 14 days, indicating an absence of subacute toxicity. Similarly, 3000 mg/kg MPJ administered to CD-1 mice for 90 days did not show subchronic toxicity. In fact, MPJ resulted in lowered weight gain in both rats and mice. Cytotoxic and microbiological analyses of MPJ were also performed. MPJ did not cause any cytotoxicity in epithelial cell culture as tested using the Alamar blue assay. Additionally, histopathological analysis of kidney and liver corroborated the absence of toxicity in CD-1 mice. The microbial load of the MPJ was low, and no pathogenic bacteria were present. In conclusion, the results reported here show that high doses of MPJ are apparently innocuous in rats and mice for the 14 and 90 days investigated, respectively. Although preliminary, our results suggest that MPJ may be safe to ingest and may even have beneficial effects in reducing weight gain.

Azaspiracids (AZAs) are lipophilic polyether toxins produced by Azadinium and Amphidoma species of marine microalgae. The main dinoflagellate precursors AZA1 and AZA2 are metabolized by shellfish to produce an array of AZA analogues. Many marine toxins undergo fatty acid esterification in shellfish, therefore mussel tissues contaminated with AZAs were screened for intact fatty acid esters of AZAs using liquid chromatography-high resolution mass spectrometry. Acyl esters were primarily observed for AZAs containing hydroxy groups at C-3 with 3-O-palmitoylAZA4 identified as the most abundant acyl ester, while other fatty acid esters including 18:1, 16:1, 17:0, 20:2 and 18:0 acyl esters were detected. The structures of these acyl derivatives were determined through LC-MS/MS experiments, and supported by periodate cleavage reactions and semi-synthesis of palmitate esters of the AZAs. Esters of the hydroxy groups at C-20 or C-21 were not observed in mussel tissue. The relative proportion of the most abundant AZA ester was less than 3% of the sum of the major free AZA analogues. These findings reveal an additional metabolic pathway for AZAs in shellfish., Highlights • Fatty acid esters of azaspiracids were identified in mussels (Mytilus edulis). • Fatty acid esters of azaspiracids with hydroxy groups at C-3 were primarily observed. • Fatty acid esters of regulated azaspiracids (AZA1, 2, −3) were absent. • Structures were determined with LC-HRMS and confirmed by semi-synthesis of palmitate esters and periodate cleavage. • This work reveals an additional metabolic pathway for azaspiracids in shellfish.

Vector competence refers to the ability of a vector to acquire, maintain, and transmit a pathogen. Collecting mosquito saliva in medium-filled capillary tubes has become the standard for approximating arbovirus transmission. However, this method is time-consuming and labor-intensive. Here we compare the capillary tube method to an alternative high-throughput detection method the collection of saliva on paper cards saturated with honey, with (FTA card) and without (filter paper) reagents for the preservation of nucleic acid for Aedes aegypti and Aedes albopictus mosquitoes infected with two emerging genotypes of the chikungunya virus (CHIKV). Model results showed that the Asian genotype CHIKV dissemination in the harvested legs of both Ae. aegypti and Ae. albopictus increased the odds of females having a positive salivary infection and higher salivary viral titers, while for the IOL genotype the same effect was observed only for Ae. aegypti. Of the three tested detection methods, the FTA card was significantly more effective at detecting infected saliva of Ae. aegypti and Ae. albopictus females than the capillary tube and filter paper was as effective as the capillary tube for the Asian genotype. We did not find significant effects of the detection method in detecting higher viral titer for both Asian and IOL genotypes. Our results are discussed in light of the limitations of the different tested detection methods.

Background For microorganisms on a paper surface, the lack of water is one of the most important stress factors. A strain of Bacillus megaterium FDU301 was isolated from plaques on a paper surface using culture medium with polyethylene glycol 200 (PEG200) to simulate an arid condition. Global transcriptomic analysis of B. megaterium FDU301 grown under normal and simulated arid conditions was performed via RNA-seq technology to identify genes involved in arid stress adaptation. Results The transcriptome of B. megaterium FDU301 grown in LB medium under arid (15% PEG200 (w/w)) and normal conditions were compared. A total of 2941 genes were differentially expressed, including 1422 genes upregulated and 1519 genes downregulated under arid conditions. Oxidative stress-responsive regulatory genes perR, fur, and tipA were significantly upregulated, along with DNA protecting protein (dps), and catalase (katE). Genes related to Fe2+ uptake (feoB), sporulation stage II (spoIIB, spoIIE, spoIIGA), small acid-soluble spore protein (sspD), and biosynthesis of compatible solute ectoine (ectB, ectA) were also highly expressed to various degrees. Oxidative phosphorylation-related genes (atpB, atpE, atpF, atpH, atpA, atpG, atpD, atpC) and glycolysis-related genes (pgk, tpiA, frmA) were significantly downregulated. Conclusion This is the first report about transcriptomic analysis of a B. megaterium to explore the mechanism of arid resistance. Major changes in transcription were seen in the arid condition simulated by PEG200 (15%), with the most important one being genes related to oxidative stress. The results showed a complex mechanism for the bacteria to adapt to arid stress.

Lake Van fish (Alburnus tarichi Guldenstadt 1814) is the only fish that can adapt to the extreme conditions (pH 9.8 salinity 0.2% and alkalinity 151.2 meq/L) of Lake Van. In this study, it was aimed to determine the cytotoxic and genotoxic effects of chlorpyrifos (CPF) on Lake Van fish primary gill cell culture. Gill epithelium from Lake Van fish was isolated enzymatically and grown in primary culture on Leibovitz’s L-15 medium. After different doses (0.01, 0.1, 1, and 10 μM) of CPF were applied to the gill cells, the total oxidant status (TOS), total antioxidant status (TAS), malondialdehyde (MDA), and DNA damage levels (8-hydroxyguanine (8-OHdG)) were examined at the end of 24 and 48 h. It was determined that the TOS, MDA, and 8-OHdG levels increased in the cells exposed to high doses (1 and 10 μM) of CPF and the TAS was decreased (P

Objectives The emergence of the 2019 novel coronavirus (SARS-CoV-2) has necessitated evaluation of the potential for SARS-CoV-2 infection in dogs and cats. Using a large data set, we evaluated the frequency of SARS-CoV-2 and other respiratory pathogens in samples submitted for respiratory testing from mid-February to mid-April 2020. Materials and methods A SARS-CoV-2 real-time PCR was developed and validated. A subset of canine and feline samples submitted for respiratory pathogen panel testing to reference laboratories in Asia, Europe, and North America were also tested for SARS-CoV-2. The frequency of respiratory pathogens was compared for the February-April period of 2020 and 2019. Results Samples from 4616 patients were included in the study and 44% of canine and 69% of feline samples were PCR positive with Mycoplasma cynos and Bordetella bronchiseptica and Mycoplasma felis and feline calicivirus, respectively. No SARS-CoV-2 infections were identified. Positive results for respiratory samples were similar between years. Clinical significance The data in this study suggest that during the emergence of the SARS-CoV-2 pandemic in early 2020, respiratory diseases in tested pet cats and dogs were caused by common veterinary pathogens and that SARS-CoV-2 infections in dogs and cats are rare.

In modern times, viruses still threaten people's lives. Among them, norovirus was the main pathogenic factor in the cause of gastroenteritis and foodborne illness, of which the GII.4 and GII.17 genotypes are prevalent in China and most parts of the world. A simple and low-cost platform for rapid and accurate norovirus detection remains a major challenge. After the cell-free system and paper-based chromogenic system were optimized, a rapid and specific norovirus detection method was established based on norovirus-specific sequences in combination with toehold switch elements. The development of a visible color change during detection eliminates the need for any complicated instruments. We validated this strategy and its specificity in differentiating GII.4, GII.17, Zika virus, and human coronavirus HKU1. The results showed that the optimized detection system not only provided a simple and rapid detection method for the sufficient differentiation of the two norovirus genotypes but also showed high specificity and no cross-reactivity with other viruses. Using nucleic acid isothermal amplification, this assay showed a limit of detection of 0.5 pM for the GII.4 genotype and 2.6 fM for the GII.17 genotype in reactions that could be observed directly with the naked eye. Our results suggested that this paper-based colorimetric method could serve as a simple and low-cost visual detection method for pathogens in clinical samples, especially in remote or rural areas.

Aims This study was designed to analyse the prevalence of class 1 and class 2 integron integrase genes among antibiotic-resistant coliform bacteria isolated from paper currency circulating in Pakistan. Methods and results A total of 500 individual currency notes were collected from different food vending sites at Lahore, Pakistan. Bacterial population were identified by biochemical and PCR techniques. Antimicrobial susceptibility testing was performed by disc diffusion assay. The highest bacterial population on currency was found from street vendors and butcher shops. Escherichia coli was found to be the most prevalent coliform bacteria followed by Klebsiella sp. and Enterobacter sp. PCR amplification of antimicrobial resistance gene showed the presence of ampC, blaTEM , blaNDM-1 , qnrA, tet(A) and tet(B) genes among coliform isolates. A total of 47 integron integrase bearing strains of coliform bacteria were analysed. Sequence analysis showed the presence of dfrA1-aadA1, dfrA1, dfrA5, dfrA7, aadA1, aadA4 cassette arrays in class 1 integron and dfrA1-sat2-aadA1 in class 2 integrase genes. Conclusion Circulating currency was heavily contaminated with antimicrobial-resistant coliform bacteria bearing class 1 and class 2 integron integrase genes. Significance and impact of the study This study describes a potential threat of severe bacterial infections due to improper hand hygiene and community sanitation when dealing with the currency notes.