Antifungal, antiaflatoxigenic, and cytotoxic properties of bioactive secondary metabolites derived from Bacillus species.

Aflatoxins (AFs) are hazardous carcinogens and mutagens produced by some molds, particularly Aspergillus spp. Therefore, the purpose of this study was to isolate and identify endophytic bacteria, extract and characterize their bioactive metabolites, and evaluate their antifungal, antiaflatoxigenic, and cytotoxic efficacy against brine shrimp (Artemia salina) and hepatocellular carcinoma (HepG2). Among the 36 bacterial strains isolated, ten bacterial isolates showed high antifungal activity, and thus were identified using biochemical parameters and MALDI-TOF MS. Bioactive metabolites were extracted from two bacterial isolates, and studied for their antifungal activity. The bioactive metabolites (No. 4, and 5) extracted from Bacillus cereus DSM 31T DSM, exhibited strong antifungal capabilities, and generated volatile organic compounds (VOCs) and polyphenols. The major VOCs were butanoic acid, 2-methyl, and 9,12-Octadecadienoic acid (Z,Z) in extracts No. 4, and 5 respectively. Cinnamic acid and 3,4-dihydroxybenzoic acid were the most abundant phenolic acids in extracts No. 4, and 5 respectively. These bioactive metabolites had antifungal efficiency against A. flavus and caused morphological alterations in fungal conidiophores and conidiospores. Data also indicated that both extracts No. 4, and 5 reduced AFB1 production by 99.98%. On assessing the toxicity of bioactive metabolites on A. salina the IC50 recorded 275 and 300 µg/mL, for extracts No. 4, and 5 respectively. Meanwhile, the effect of these extracts on HepG2 revealed that the IC50 of extract No. 5 recorded 79.4 µg/mL, whereas No. 4 showed no cytotoxic activity. It could be concluded that bioactive metabolites derived from Bacillus species showed antifungal and anti-aflatoxigenic activities, indicating their potential use in food safety.

Extraction and characterization of bioactive secondary metabolites from lactic acid bacteria and evaluating their antifungal and antiaflatoxigenic activity.

Aflatoxins are toxic carcinogens and mutagens formed by some moulds, specifically Aspergillus spp. Therefore, this study aimed to extract and identify bioactive secondary metabolites from Lactobacillus species, to evaluate their efficacy in reducing fungal growth and aflatoxin production and to investigate their toxicity. The bioactive secondary metabolites of Lactobacillus species showed variable degrees of antifungal activity, whereas L. rhamnosus ethyl acetate extract No. 5 exhibited the highest antifungal activity and, thus, was selected for further identification studies. Data revealed that L. rhamnosus ethyl acetate extract No. 5 produced various organic acids, volatile organic compounds and polyphenols, displayed antifungal activity against A. flavus, and triggered morphological changes in fungal conidiophores and conidiospores. L. rhamnosus ethyl acetate extract No. 5 at a 9 mg/mL concentration reduced AFB1 production by 99.98%. When the effect of L. rhamnosus ethyl acetate extract No. 5 on brine shrimp mortality was studied, the extract attained a 100% mortality at a concentration of 400 µg/mL, with an IC50 of 230 µg/mL. Meanwhile, a mouse bioassay was performed to assess the toxicity of L. rhamnosus ethyl acetate extract No. 5, whereas there were no harmful effects or symptoms in mice injected with L. rhamnosus ethyl acetate extract at concentrations of 1, 3, 5, 7, and 9 mg/kg body weight.

Potential synergistic effect of Alhagi graecorum ethanolic extract with two conventional food preservatives against some foodborne pathogens.

The present study aims to screen the anti-bacterial activity and synergistic interaction of A. graecorum Boiss. ethanolic extract with two food preservatives against five strains of foodborne bacteria. Disk diffusion and minimum inhibitory concentration were used for anti-bacterial assay, checkerboard assay and time-kill curve were used for the combination studies. HPLC analysis and molecular docking study were performed to corroborate the in vitro results. The ethanolic extract showed anti-bacterial activity against all tested bacterial strains with inhibition zones from 7.5 to 9.3 mm and MIC values ranged between 1.2 and 1.8 mg mL-1. The combination of the ethanolic extract with Na-benzoate or Na-propionate resulted in synergistic and additive interactions against the tested bacteria with fractional inhibitory concentration index (FICI) ranges 0.31-0.63 and no antagonism was shown. Time-kill curve assay showed that the synergistic and additive combinations have inhibitory effects on the tested strains. The ethanolic extract combination with Na-benzoate or Na-propionate can be used for development new sources of food preservatives. Testing new different natural plant extracts with food preservatives will help develop new anti-bacterial agents.

Value-Added Utilization of Citrus Peels in Improving Functional Properties and Probiotic Viability of Acidophilus-bifidus-thermophilus (ABT)-Type Synbiotic Yoghurt during Cold Storage.

Citrus peel, a fruit-processing waste, is a substantial source of naturally occurring health-promoting compounds, including polyphenols, and has great potential as a dietary supplement for enhancing the functional properties of food. The present work aimed to investigate the effects of sour orange (SO), sweet orange (SWO), and lemon (LO) peels on the typical physiochemical, antioxidant, antibacterial, and probiotic properties of synbiotic yoghurt fermented by acidophilus-bifidus-thermophilus (ABT)-type cultures during cold storage (0−28 days). High-performance liquid chromatography-diode array detection (HPLC-DAD) analysis showed that the total phenolic content in the SO peel were more than 2-fold higher than that in the SWO and LO peel. The predominant phenolic compounds were myricetin (2.10 mg/g dry weight) and o-coumaric acid (1.13 mg/g) in SO peel, benzoic acid (0.81 mg/g) and naringin (0.72 mg/g) in SWO peel, and benzoic acid (0.76 mg/g) and quercetin (0.36 mg/g) in LO peel. Only 0.5% (w/w) of citrus peel addition did not reduce the overall acceptance of ABT synbiotic yoghurt but led to increased acidity and decreased moisture during cold storage (14 and 28 days). Additionally, compared to control samples without citrus peel addition, supplementation with citrus peels improved the antioxidant property of the ABT synbiotic yoghurt. ABT milks with SO and SWO peel addition had significantly stronger DPPH radical scavenging activities than that with LO peel addition (p < 0.05). Antibacterial analysis of ABT synbiotic yoghurt with citrus peel addition showed that the diameters of inhibition zones against S. aureus, B. subtilis, and E. coli increased by 0.6−1.9 mm relative to the control groups, suggesting the enhancement of antibacterial activities by citrus peels. The viabilities of probiotic starter cultures (L. acidophilus, S. thermophilus, and Bifidobacterial sp.) were also enhanced by the incorporation of citrus peels in synbiotic yoghurt during cold storage. Hence, our results suggest that citrus peels, especially SO and SWO peels, could be recommended as a promising multifunctional additive for the development of probiotic and synbiotic yoghurt with enhanced antioxidant and antibacterial properties, as well as probiotic viability.

Prevalence and Predictors of Persistent Symptoms After Clearance of SARS-CoV-2 Infection: A Multicenter Study from Egypt.

Background and Aim: Little is known about the persistence of symptoms after clearance of SARS-CoV-2 infection. Our study aimed to assess persistent symptoms in COVID-19 patients after clearance of SARS-CoV-2 infection. Methodology: A multi-center survey was conducted on first wave COVID-19 patients with confirmed SARS-CoV-2 infection. Sociodemographic and clinical characteristics, including presenting symptoms and persistent symptoms after viral clearance and possible factors contributing to persistence of such symptoms, were collected using an online multicomponent questionnaire. Descriptive and inferential statistical analysis was performed to detect the most persisting symptoms and factors contributing to their persistence. Results: Overall, 538 patients were enrolled. Mean age was 41.17 (±SD 14.84), 54.1% were males, and 18.6% were smokers. Hypertension and diabetes were the most reported co-morbidities. Mild symptoms were reported in 61.3% of patients, 51.3% were admitted to hospital and 6.5% were admitted to the intensive care unit. Our study identified 49 types of persisting symptoms. Fatigue (59.1%), sense of fever (46.5%), anorexia (24.3%) and diarrhea (24.3%) were the most commonly reported persisting symptoms followed by loss of taste and smell (22.3%), headache (21.4%), cough (20.8) and dyspnea (21%). The use of hydroxychloroquine, azithromycin and multivitamins were significantly associated with persistence of symptoms (OR = 8.03, 8.89 and 10.12, respectively). Conclusion: Our study revealed that in COVID-19 recovered patients, many patients reported persistence of at least one symptom, particularly fatigue and sense of fever. Follow-up of patients after discharge from hospital is recommended until complete resolution of symptoms.

Chemical profiling, in vitro antimicrobial and antioxidant activities of pomegranate, orange and banana peel-extracts against pathogenic microorganisms.

BACKGROUND: The use of natural preservatives became of great interest; good examples of these natural preservation agents are plant peels. The use of plant peels has dual benefits; first is their antimicrobial activity against food-borne pathogens, while the second is minimizing agro-industrial wastes. RESULTS: The evaluation of the antimicrobial potential of both methanolic and ethanolic extracts of three fruit peels (orange, pomegranate, and banana), against 4 Gram-positive (G+), 3 Gram-negative bacteria (G-), and 2 fungal strains revealed that both pomegranate peel extracts exhibited significantly higher inhibitory effect on all tested G+ bacteria. Methanolic extract of pomegranate peel gave higher activity than the ethanolic one against G+ and G- bacteria except for S. typhimurium. Against A. flavus and A. niger, both pomegranate and orange extracts showed activity ranging between 65 and 100% more than the positive control. The ethanolic extracts of all tested peels showed a considerable capacity of antioxidant compounds compared to the methanolic extracts. The highest antioxidant capacity was found for ethanolic and methanolic extracts of pomegranate, 66.870 and 56.262 mg/ml, respectively. Generally, the concentration of total phenolic compounds was higher than that of total flavonoids followed by tannins. The highest readings of all tested constituents were reported for pomegranate extracts followed by orange and then banana. The total phenolic content, total flavonoids, and tannins were proportional to antioxidant values. GC-MS of pomegranate peel extracts identified 23 compounds in the methanolic extract versus 31 compounds in the ethanolic one. These components were identified based on their retention times and mass spectral fragmentation pattern. 5-hydroxymethylfufural (HMF) represented the major component in both methanolic and ethanolic extracts with peak area percentage of 65.78% and 48.43%, respectively. CONCLUSIONS: The results showed negative effect of methanolic and ethanolic extracts of pomegranate on G+ and G- bacteria and two fungal pathogenic strains. The phytochemical analysis regarded these results to the high content of phenols, flavonoids, and tannins. GC-MS chromatogram identified many compounds known to be effective as antioxidants and antibacterial and antifungal agents. These indications show that pomegranate peel may be a superior natural food-preserver, but further studies about the suitable formulation, dosage, and possible side-effects are still needed.

Barriers for resuming endoscopy service in the context of COVID-19 pandemic: A multicenter survey from Egypt.

BACKGROUND: The current coronavirus disease 2019 (COVID-19) pandemic has affected routine endoscopy service across the gastroenterology community. This led to the suspension of service provision for elective cases. AIM: To assess the potential barriers for resuming the endoscopy service in Egypt. METHODS: A national online survey, four domains, was disseminated over a period of 4 wk in August 2020. The primary outcome of the survey was to determine the impact of the COVID-19 pandemic on the endoscopy service and barriers to the full resumption of a disabled center(s). RESULTS: A hundred and thirteen Egyptian endoscopy centers participated in the survey. The waiting list was increased by ≥ 50% in 44.9% of areas with clusters of COVID-19 cases (n = 49) and in 35.5% of areas with sporadic cases (n = 62). Thirty nine (34.8%) centers suffered from staff shortage, which was considered a barrier against service resumption by 86.4% of centers in per-protocol analysis. In multivariate analysis, the burden of cases in the unit locality, staff shortage/recovery and the availability of separate designated rooms for COVID-19 cases could markedly affect the resumption of endoscopy practice (P = 0.029, < 0.001 and 0.02, respectively) and Odd's ratio (0.15, 1.8 and 0.16, respectively). CONCLUSION: The COVID-19 pandemic has led to restrictions in endoscopic volumes. The staff shortage/recovery and the availability of COVID-19 designed rooms are the most important barriers against recovery. Increasing working hours and dividing endoscopy staff into teams may help to overcome the current situation.

Recycling of agro-wastes for Ganoderma lucidum mushroom production and Ganoderma post mushroom substrate as soil amendment.

The in vitro growth of Ganoderma mycelia on six agro-wastes namely, broad bean stalks (BBS), cotton stalk (CS), maize straw (MS), rice straw (RS), sugarcane bagasse (SCB) and wheat straw (WS) supplemented with wheat bran (WB) or corn gluten (CG) was evaluated. Among the substrates used, CS appeared best followed by SCB and RS. WB showed best supplementation for mycelial growth. CO2 emission values exhibited accurate measurements to decide the suitability of such agro-waste for growth rather than visual observations. CS+RS+SCB+WB in combination proven its superiority for in vitro growth and active spawn development substrate. In mushroom house, this particular formula proved its superiority and was on par with recommended EG formula; it gave the highest yield (195.16 g Kg-1), biological efficiency (19.52%), protein (16.69%), polysaccharides (3.613%) and minerals (3433 mg/100 g). Spawn running period was the shortest in treatments inoculated with agro-waste-based spawns. With 40% biochar, days required to the complete mycelium colonization and fructification were 10.60 and 23.00, respectively. At 10% biochar, highest yields (238.40 g Kg-1), biological efficiencies (23.84%), protein (19.58%) and minerals (4092 mg/100 g) were obtained. The higher the biochar level, the higher the reduction in emitted CO2, the loss in C and the increase in N of Ganoderma post mushroom substrates (GPMSs). Under greenhouse conditions, almost all the tested GPMSs, at 0.125 or 0.25%, encouraged the reproduction of reniform nematodes and improved plant growth criteria.

Maternal serum amyloid A level as a novel marker of primary unexplained recurrent early pregnancy loss.

OBJECTIVE: To assess maternal serum amyloid A (SAA) levels among women with primary unexplained recurrent early pregnancy loss (REPL). METHODS: A prospective study was conducted among women with missed spontaneous abortion in the first trimester at Ain Shams University Maternity Hospital, Cairo, Egypt, between January 21 and December 25, 2014. Women with at least two consecutive primary unexplained REPLs and no previous live births were enrolled. A control group was formed of women with no history of REPL who had at least one previous uneventful pregnancy with no adverse outcomes. Serum samples were collected to measure SAA levels. The main outcome was the association between SAA and primary unexplained REPL. RESULTS: Each group contained 96 participants. Median SAA level was significantly higher among women with REPL (50.0 µg/mL, interquartile range 26.0-69.0) than among women in the control group (11.6 µg/mL, interquartile range 6.2-15.5; P<0.001). The SAA level was an independent indicator of primary unexplained REPL, after adjusting for maternal age and gestational age (odds ratio 1.12, 95% confidence interval 1.06-1.19; P<0.001). CONCLUSION: Elevated SAA levels found among women with primary unexplained REPL could represent a novel biomarker for this complication of pregnancy.

Isolation and characterization of multifunctional Streptomyces species with antimicrobial, nematicidal and phytohormone activities from marine environments in Egypt.

Different strategies have been employed for selective isolation of Streptomycetes from 20 marine samples varied in their biological nature. The recovery of Streptomycetes isolates (112) was influenced preferentially by different strategies; sediment samples were the best source of potential candidate Streptomycetes. All isolates exhibited antimicrobial activities with variable spectrum; the most promising isolates (31) were phenotypically characterized and identified as Streptomyces sp.; these isolates exhibited variable capacity for secretion of numerous hydrolytic enzymes such as catalase, protease, amylase, lipase, lecithinase, asparaginase, chitinase and pectinase. All the strains resisted both penicillin and streptomycin, 29 were sensitive to neomycin; the majority of strains (25) showed multiple antibiotic resistance index greater than 0.2; 23, 22 and 13 degraded the shrimp shell, chicken feather and corn cob, respectively, producing bioactive substance(s) which indicates their diversity and their ecological role in the marine ecosystem. At least 28 strains exhibited nematicidal activity in vitro and in vivo against root-knot nematode and supported plant growth. In vitro, the assessed Streptomyces species exhibited the ability to produce gibberellic acid, indole acetic acid, abscisic acid, kinetin and benzyladenine. Except for indole acetic acid, this is the first report concerning the ability of marine Streptomyces to produce such phytohormones and the use of shrimp shell waste as a mono component medium for production of phytohormones. The study is efficacious in selecting effective biodiverse strains of marine Streptomyces that may work under diverse agro-ecological conditions as a useful element in plant nutrition and as biocontrol agents involved in integrated management programs.

Identification of mosquito larvicidal bacterial strains isolated from north Sinai in Egypt.

In the present study, two of the most toxic bacterial strains of Bacillus sphaericus against mosquito were identified with the most recent genetic techniques. The PCR product profiles indicated the presence of genes encoding Bin A, Bin B and Mtx1 in all analyzed strains; they are consistent with protein profiles. The preliminary bioinformatics analysis of the binary toxin genes sequence revealed that the open reading frames had high similarities when matched with nucleotides sequence in the database of other B. sphaericus strains. The biological activity of B. sphaericus strains varied according to growing medium, and cultivation time. The highest yield of viable counts, spores and larvicidal protein were attained after 5 days. Poly (P) medium achieved the highest yield of growth, sporulation, protein and larvicidal activity for all tested strains compared to the other tested media. The larvicidal protein produced by local strains (B. sphaericus EMCC 1931 and EMCC 1932) in P medium was more lethal against the 3rd instar larvae of Culex pipiens than that of reference strains (B. sphaericus 1593 and B. sphaericus 2297). The obtained results revealed that P medium was the most effective medium and will be used in future work in order to optimize large scale production of biocide by the locally isolated Bacillus sphaericus strains.