Root Rot Management in Common Bean (Phaseolus vulgaris L.) Through Integrated Biocontrol Strategies using Metabolites from Trichoderma harzianum, Serratia marcescens, and Vermicompost Tea.

Common bean (Phaseolus vulgaris L.) is an essential food staple and source of income for small-holder farmers across Africa. However, yields are greatly threatened by fungal diseases like root rot induced by Rhizoctonia solani. This study aimed to evaluate an integrated approach utilizing vermicompost tea (VCT) and antagonistic microbes for effective and sustainable management of R. solani root rot in common beans. Fourteen fungal strains were first isolated from infected common bean plants collected across three Egyptian governorates, with R. solani being the most virulent isolate with 50% dominance. Subsequently, the antagonistic potential of vermicompost tea (VCT), Serratia sp., and Trichoderma sp. was assessed against this destructive pathogen. Combinations of 10% VCT and the biocontrol agent isolates displayed potent inhibition of R. solani growth in vitro, prompting in planta testing. Under greenhouse conditions, integrated applications of 5 or 10% VCT with Serratia marcescens, Trichoderma harzianum, or effective microorganisms (EM1) afforded up to 95% protection against pre- and post-emergence damping-off induced by R. solani in common bean cv. Giza 6. Similarly, under field conditions, combining VCT with EM1 (VCT + EM1) or Trichoderma harzianum (VCT + Trichoderma harzianum) substantially suppressed disease severity by 65.6% and 64.34%, respectively, relative to untreated plants. These treatments also elicited defense enzyme activity and distinctly improved growth parameters including 136.68% and 132.49% increases in pod weight per plant over control plants. GC-MS profiling of Trichoderma harzianum, Serratia marcescens, and vermicompost tea (VCT) extracts revealed unique compounds dominated by cyclic pregnane, fatty acid methyl esters, linoleic acid derivatives, and free fatty acids like oleic, palmitic, and stearic acids with confirmed biocontrol and plant growth-promoting activities. The results verify VCT-mediated delivery of synergistic microbial consortia as a sustainable platform for integrated management of debilitating soil-borne diseases, enhancing productivity and incomes for smallholder bean farmers through regeneration of soil health. Further large-scale validation can pave the adoption of this climate-resilient approach for securing food and nutrition security.

Larvicidal potential, antimicrobial properties and molecular docking analysis of Egyptian Mint (Mentha rotundifolia) against Culex pipiens L. (Diptera: Culicidae) and Midgut-borne Staphylococcus aureus.

Mosquitoes prefer stagnant areas near hospitals to live and easily spread pathogenic bacteria. Our current study aims to isolate multidrug-resistant (MDR) Staphylococcus aureus isolates from midguts of Mosquito Culex pipiens and study the potential of mint as a biocontrol strategy against C. pipiens larvae and their midgut-borne S. aureus. Samples of the third and fourth larval instars of C. pipiens were collected from water ponds around three Cairo hospitals. Ciprofloxacin, gentamycin and tetracycline, as well as various concentrations of mint leaf extract (MLE) were tested for antibiotic susceptibility. Sixty-five isolates were obtained and showed antibiotic resistance to tetracycline, gentamycin, ciprofloxacin, and undiluted MLE with resistant percentages (%) of 27.69, 30.76, 17.46, and 23.08%, respectively. Undiluted MLE inhibited 61.53% of the multidrug S. aureus isolates, whereas it couldn't inhibit any of these isolates at dilutions less than 50 µg/mL. The MIC of MLE was ≤ 700 µg/mL, while the MIC of the antibiotics ranged from 0.25 to 5.0 µg/mL for the three antibiotics. The most inhibited S. aureus isolate was identified by 16SrRNA sequencing approach and registered in GenBank as S. aureus MICBURN with gene accession number OQ766965. MLE killed all larval stages after 72 h of exposure, with mortality (%) reaching 93.33 and 100% causing external hair loss, breakage of the outer cuticle epithelial layer of the abdomen, and larvae shrinkage. Histopathology of treated larvae showed destruction of all midgut cells and organelles. Gas chromatography (GC) of MLE revealed that menthol extract (35.92%) was the largest active ingredient, followed by menthone (19.85%), D-Carvone (15.46%), Pulegone (5.0579%). Docking analysis confirmed that alpha guanine and cadinol had the highest binding affinity to both predicted active sites of Culex pipiens acetylcholinesterase. As a result, alpha-guanine and cadinol might have a role as acetylcholinesterase inhibitors.

Optimizing alpha-amylase from Bacillus amyloliquefaciens on bread waste for effective industrial wastewater treatment and textile desizing through response surface methodology.

Food waste is a major issue, with one-third of food wasted yearly. This study aimed to produce sustainably the industrial enzyme alpha-amylase using discarded bread waste. Brown (BBW) and white bread waste (WBW) were tested as growth substrates using solid-state and submerged fermentation. The biosynthesized α- amylase is applied to treat starch-heavy industrial wastewater and for textile desizing. Bacillus amyloliquificiens showed the highest starch hydrolysis and enzyme activity on solid and liquid media. α-amylase production by B. amyloliquificiens was optimized via a one-factor-at-a-time evaluation of production parameters. Optimal production occurred by submerged fermentation of BBW inoculated with 2% B. amyloliquificiens at 37 °C and 200rpm for 24 h, reaching 695.2 U/mL α- amylase. The crude enzyme was immobilized on calcium alginate beads with 96.6% efficiency and kept 88.5% activity after 20 reuses, enhancing stability. A Box-Behnken design (BOX) assessed variable interactions. Response surface methodology (RSM) generated a quadratic model and analysis of variance (ANOVA analysis) fitting experimental starch hydrolysis data. Optimal conditions were pH 9, 45 °C, 70% starch, and 27.5 U/mL enzyme incubated for 15 min of contact time, with a high R2 of 0.83. ANOVA confirmed the enzyme's alkaliphilic and thermophilic nature. Using enzyme concentrations ranging from 10.9 to 695.1 U/mL, the enzyme desized textiles in 15 min at pH 9.0 and 45 °C with 96.3% efficiency. Overall, the optimized α- amylase from bread waste has industrial potential for sustainable starch processing.

Biosynthesis and characterization of silver nanoparticles from Punica granatum (pomegranate) peel waste and its application to inhibit foodborne pathogens.

Polyphenolics have been predicted to effectively develop antimicrobial agents for the food industry as food additives and promote human health. This study aims to synthesize pomegranate peel extract (PPE) with silver nanoparticles (AgNPs) against eight foodborne pathogens. Multispectroscopic analysis of UV-vis spectroscopy, Zeta potential, Fourier transform infrared (FTIR) and scanning electron microscopy (SEM) analysis were used to characterize the interaction between PPE and AgNPs. Eight foodborne pathogenic strains (six bacterial and two fungal strains) Bacillus subtilis ATCC 6633, Enterococcus faecalis ATCC 29212, Escherichia coli ATCC 8379, Klebsiella pneumoniae ATCC 00607, Salmonella typhi DSM 17058, Shigella sonnei DSM 5570, Aspergillus flavus ATCC 9643, and Rhizopus oryzae ATCC 96382 were used to test the inhibitory potential of PPW-AgNPs. The reaction colour of PPE-AgNPs from yellow to brown indicated that the nanoparticles were successfully formed. The UV absorption of PPE-AgNPs was detected at 440 nm of 0.9 SPR. SEM image of PPE-AgNPs exhibited spherical shapes with a zeta potential of - 20.1 mV. PPE-AgNPs showed high antimicrobial activity against all tested strains. The highest inhibition activity of PPE-AgNPs was recorded for the B. subtilis strain followed by K. pneumonia, while the highest resistance was noticed for R. oryzae. The components of pomegranate peel were analyzed using gas chromatography-mass spectrometry (GC-MS). The major constituents of pomegranate peel is phenol (51.1%), followed by Isocitronellol (19.41%) and 1-Propanol, 2-(2-hydroxypropyl)- (16.05%). PPE is key in the simple, eco-friendly green synthesis of extracellular stable AgNPs as an alternative source for harmful chemical disinfectants.

Larvicidal potential, toxicological assessment, and molecular docking studies of four Egyptian bacterial strains against Culex pipiens L. (Diptera: Culicidae).

Mosquito control in Egypt depends on applying chemical synthetic pesticides that impact negatively on human health and the environment as well as the development of antibiotic and chemical resistance. This study aims to control the 3rd and 4th instars of Culex pipiens larvae using four bacterial strains. According to Phenotypic and molecular identification, the four isolates were identified as Bacillus subtilis MICUL D2023, Serratia marcescens MICUL A2023, Streptomyces albus LARVICID, and Pseudomonas fluorescens MICUL B2023. All strains were deposited in GenBank under accession numbers OQ764791, OQ729954, OQ726575, and OQ891356, respectively. Larvicidal activity of all microbial strain metabolites against a field strain of C. pipiens explored low LC50 results and reached its lowest values on the 3rd day with values of 6.40%, 38.4%, and 46.33% for P. fluorescens, S. albus, and S. marcescens, respectively. In addition, metabolites of P. fluorescence were more toxic than those of S. albus, followed by S. marcescens. B. subtilis shows no larvicidal effect on both field and lab mosquito strains. Microscopic alterations of 3rd and 4th instars showed toxic effects on different body parts (thorax, midgut, and anal gills), including losing external hairs, abdominal breakage, and larvae shrinkage, as well as different histological malformations in the digestive tract, midgut, and cortex. GC-MS analysis detected 51, 30, and 32 different active compounds from S. albus, S. marcescens, and P. fluorescens, respectively. GC detected 1, 2-BENZEA2:A52NEDICARBOXYLIC ACID, 2-Cyclohexene-1-carboxylic-acid-5-2-butenyl-methyl ester, and 3 octadecahydro2R3S4Z9Z-11R-12S from S. albus, S. marcesens, and P. fluorescens, respectively. Total protein, Total carbohydrate, and Acetylcholine esterase activity indicated significantly low levels on the 3rd day. All strain metabolites were safe against HSF cell lines. The docking results confirmed the role of the produced metabolites as larvicidal agents and Acetylcholine esterase inhibition. Such a problem need more studies on applying more and more natural pesticides.

Utilization of biosynthesized silver nanoparticles from Agaricus bisporus extract for food safety application: synthesis, characterization, antimicrobial efficacy, and toxicological assessment.

The emergence of antimicrobial resistance in foodborne bacterial pathogens has raised significant concerns in the food industry. This study explores the antimicrobial potential of biosynthesized silver nanoparticles (AgNPs) derived from Agaricus bisporus (Mushroom) against foodborne bacterial pathogens. The biosynthesized AgNPs were characterized using various techniques, including UV-visible spectroscopy, X-ray diffraction, Fourier transform infrared spectroscopy, high-resolution scanning electron microscopy with energy dispersive X-ray spectroscopy, dynamic light scattering, and zeta potential analysis. The antibacterial activity of the AgNPs was tested against a panel of foodborne bacterial strains, and their cytotoxicity was evaluated on normal human skin fibroblasts. Among the tested strains, Pseudomonas aeruginosa ATCC 27853 showed the highest sensitivity with an inhibition zone diameter (IZD) of 48 mm, while Klebsiella quasipneumoniae ATTC 700603 and Bacillus cereus ATCC 11778 displayed the highest resistance with IZDs of 20 mm. The silver cations released by AgNPs demonstrated strong bactericidal effects against both Gram-positive (G + ve) and Gram-negative (G - ve) bacteria, as evidenced by the minimum inhibitory concentration/minimum bactericidal concentration (MBC/MIC) ratio. Moreover, cytotoxicity testing on normal human skin fibroblasts (HSF) indicated that AgNPs derived from the mushroom extract were safe, with a cell viability of 98.2%. Therefore, AgNPs hold promise as an alternative means to inhibit biofilm formation in the food industry sector.

Enhancing durability and sustainable preservation of Egyptian stone monuments using metabolites produced by Streptomyces exfoliatus.

Despite their threatens for Egyptian stone monuments, A few studies focused on using biocontrol agents against deteriorative fungi and bacteria instead of using chemical assays that leave residuals leading to human toxicity and environmental pollution. This work aims to isolate and identify fungal and bacterial isolates that showed deteriorative activities from stone monuments in Temple of Hathor, Luxor, Egypt, as well as determine the inhibitory activity of metabolites produced by Streptomyces exfoliatus SAMAH 2021 against the identified deteriorative fungal and bacterial strains. Moreover, studying the spectral analysis, toxicological assessment of metabolites produced by S. exfoliatus SAMAH 2021 against health human cell fibroblast, and colorimetric measurements on the selected stone monuments. Ten samples were collected from Temple of Hathor, Luxor, Egypt. Three fungal isolates and one bacterial isolate were obtained and identified as A. niger isolate Hathor 2, C. fioriniae strain Hathor 3, P. chrysogenum strain HATHOR 1, and L. sphaericus strain Hathor 4, respectively. Inhibitory potential of the metabolites in all concentrations used (100-25%) against the recommended antibiotics (Tetracycline 10 µg/ml and Doxycycline (30 µg/ml) showed an inhibitory effect toward all tested deteriorative pathogens with a minimum inhibition concentration (MIC) of 25%. Cytotoxicity test confirmed that microbial filtrate as the antimicrobial agent was safe for healthy human skin fibroblast with IC50 of < 100% and cell viability of 97%. Gas chromatography analysis recorded the existence of thirteen antimicrobial agents, Cis-vaccenic acid; 1,2-Benzenedicarboxylic acid; ç-Butyl-ç-butyrolactone and other compounds. Colorimetric measurements confirmed no color or surface change for the limestone-treated pieces. The use of the metabolite of microbial species antimicrobial as a biocontrol agent raises contemporary issues concerning the bio-protection of the Egyptian monuments to reduce chemical formulas that are toxic to humans and pollute the environment. Such serious problems need further investigation for all kinds of monuments.

Microbial Degradation, Spectral analysis and Toxicological Assessment of Malachite Green Dye by Streptomyces exfoliatus.

Malachite green (MG) dye is a common environmental pollutant that threatens human health and the integrity of the Earth's ecosystem. The aim of this study was to investigate the potential biodegradation of MG dye by actinomycetes species isolated from planted soil near an industrial water effluent in Cairo, Egypt. The Streptomyces isolate St 45 was selected according to its high efficiency for laccase production. It was identified as S. exfoliatus based on phenotype and 16S rRNA molecular analysis and was deposited in the NCBI GenBank with the gene accession number OL720220. Its growth kinetics were studied during an incubation time of 144 h, during which the growth rate was 0.4232 (µ/h), the duplication time (td) was 1.64 d, and multiplication rate (MR) was 0.61 h, with an MG decolorization value of 96% after 120 h of incubation at 25 °C. Eleven physical and nutritional factors (mannitol, frying oil waste, MgSO4, NH4NO3, NH4Cl, dye concentration, pH, agitation, temperature, inoculum size, and incubation time) were screened for significance in the biodegradation of MG by S. exfoliatus using PBD. Out of the eleven factors screened in PBD, five (dye concentration, frying oil waste, MgSO4, inoculum size, and pH) were shown to be significant in the decolorization process. Central composite design (CCD) was applied to optimize the biodegradation of MG. Maximum decolorization was attained using the following optimal conditions: food oil waste, 7.5 mL/L; MgSO4, 0.35 g/L; dye concentration, 0.04 g/L; pH, 4.0; and inoculum size, 12.5%. The products from the degradation of MG by S. exfoliatus were characterized using high-performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS). The results revealed the presence of several compounds, including leuco-malachite green, di(tert-butyl)(2-phenylethoxy) silane, 1,3-benzenedicarboxylic acid, bis(2-ethylhexyl) ester, 1,4-benzenedicarboxylic acid, bis(2-ethylhexyl) ester, 1,2-benzenedicarboxylic acid, di-n-octyl phthalate, and 1,2-benzenedicarboxylic acid, dioctyl ester. Moreover, the phytotoxicity, microbial toxicity, and cytotoxicity tests confirmed that the byproducts of MG degradation were not toxic to plants, microbes, or human cells. The results of this work implicate S. exfoliatus as a novel strain for MG biodegradation in different environments.

Midguts of Culex pipiens L. (Diptera: Culicidae) as a potential source of raw milk contamination with pathogens.

Despite their importance, only few studies focused on the mosquitoes borne microbial diseases, especially bacterial and fungal diseases, their vectorial capacity toward microorganisms, and their important role in raw milk contamination with pathogens in some unsanitary dairy farms. In Egypt, where Culex pipiens is the historical main vector of lymphatic filariasis, only few studies discussed the isolation of pathogens from the midguts of different stages of C. pipiens. This study aims to isolate and identify the pathogenic symbiotic microorganisms inside the midgut of adult female C. pipiens as well as investigate its ability to transmit their midgut pathogens to raw milk. A total of 750 field strain C. pipiens larvae of the second and third larval instars were collected from ponds water around the livestock farms in Mariotteya, Giza, Egypt, for microbial pathogen isolation and identification. All collected larval instars were transported to the laboratory at the Research and Training Center on Vectors of Diseases (RTC), Ain Shams University, where they were maintained for further studies. Six groups of C. pipiens were tested for the incidence of various pathogenic microorganisms in their midguts and their possibility to contaminate commercial sterilized milk. Traditional PCR assays and sequencing method detected and identified 16srRNA genes of the predominant hemolytic isolates from milk and midguts of female C. pipiens. The phylogenetic analyses of the obtained isolates were performed based on NCBI data. Three strains of Bacillus anthracis strain CPMESA 2021, Staphylococcus warneri strain CPSAME 2021, and Bacillus cereus strain CPSEMA 2021, which represent most food pathogens, were found in the midguts of C. pipiens and were submitted to the GenBank database with the accession numbers OK585071, OK576651, and OK585052, respectively. The isolation of these strains from mosquitoes raises contemporary issues concerning milk safety, such as bacterial isolates, the degree of the vectorial capacity of mosquitoes, milk production and processing conditions, and human pathogenicity. Such serious issues need further investigation.

Exogenous Paclobutrazol Reinforces the Antioxidant and Antimicrobial Properties of Lavender (Lavandula officinalis L.) Oil through Modulating Its Composition of Oxygenated Terpenes.

Plant growth regulators can affect the primary and secondary metabolites of various plant species. However, the effect of paclobutrazol (PBZ) on the composition of lavender oil, especially related to the terpenoid pathway, is still unclear in literatures. In this study, the effect of PBZ as a foliar spray (0.200, 400 and 600 ppm) on the vegetative growth, phytochemical content, and both antioxidant and antimicrobial properties of lavender oil were investigated. The results indicated that all examined PBZ treatments led to a significant (p ≤ 0.05) decrease in growth parameters compared to the untreated plants. Meanwhile, the yield of essential oil was significantly decreased by the treatment of PBZ at 200 ppm compared to the control. In contrast, applied-PBZ significantly enhanced the chlorophyll content and displayed a marked change in the composition of the essential oil. This change included an obvious and significant increase in 3-carene, eucalyptol, γ-terpinene, α-pinocarvone, caryophyllene, ß-vetivenene, ß-santalol, ledol, geranyl isovalerate, farnesol, caryophyllene oxide, and phytol percentage. Generally, the highest significant values were achieved by the treatment of 400 ppm compared to the other treatments. Furthermore, this treatment showed the highest free radical scavenging activity against DPPH (1,1-diphenyl-2-picrylhydrazyl) by 13% over the control. Additionally, to determine the antimicrobial activities of the extracted oil, each treatment was examined against two strains of Gram positive bacteria (S. aureus and B. cereus), two strains of Gram negative bacteria (S. enteritidis and E. coli), and two fungal species (C. albicans and A. niger) represent the yeast modal and filamentous fungus, respectively. The findings demonstrated that all examined species were more sensitive to the oil that was extracted from lavender plants, treated with 400 ppm PBZ, compared to the other concentrations.

Factors Influencing Decolourization and Detoxification of Remazol Brilliant Blue R Dye by Aspergillus flavus.

<b>Background and Objectives:</b> Anthraquinone synthetic dyes are widely used in textile, dyeing and paper painting. The discharge of these dyes into the environment causes detriment. The removal of physiochemical dyes is sometimes unsuccessful and expensive. Biological removal is inexpensive, eco-friendly and may break down organic contaminants. In the current work, a fungal technique was applied to decolorize and detoxify dye. <b>Materials and Methods:</b> Dye decolorizing fungi isolation, selection and identification of the most effective isolate and dye decolorization optimization based on carbon and nitrogen sources. In addition, the product's cytotoxicity and metabolites were tested. The enzyme activities were measured to determine dye decolorization. <b>Results:</b> Decolorization of reactive blue 19 dye by the most effective fungal strain isolate (5BF) isolated from industrial effluents were studied. This isolate was identified as <i>Aspergillus flavus</i> based on phenotypic characteristics and confirmed using 18S rRNA gene sequencing. Thin-layer chromatography indicated that this strain is aflatoxins free. Furthermore, metabolites produced from dye treatment with <i>A. flavus</i> were assessed using gas chromatography-mass spectrometry. Toxicity data revealed that <i>A. flavus</i> metabolites were not toxic to plants. Using a one-factor-at-a-time optimization technique, a maximum decolorization percentage (99%) was obtained after 72 hrs in the presence of mannitol and NH₄NO₃ or NH₄Cl as carbon and nitrogen sources. Two enzymes (laccase and manganese peroxidase) were shown to be active during dye decolorization by <i>A. flavus</i>. <b>Conclusion:</b> The <i>A. flavus</i> strain was shown to be safe when it came to removing dye from a synthetic medium with high efficiency and their metabolites had no negative influence on the environment. As a result, this strain will be used in the future for dye wastewater bioremediation.

First study in Qatar to reveal high Legionella counts in cooling towers.

Legionella spp. is transmitted from water to humans by aerosol-generating devices, including cooling towers (CTs). There have not been published reports about Legionella in these systems in Qatar. Ten CTs in Qatar University were sampled on a monthly basis. Bacteria were recovered from 90 water samples by filtration and concentration. Legionella DNA copy number (CN) was assessed by quantitative RT-PCR. Legionella DNA was detected in 100% of the samples. The bacterial counts ranged from 0.006 to 199.56 CFU/mL, and critical counts were found in 51 (56.7 %) samples. Moreover, 7 (7.8%) samples showed a count of more than 100 CFU/mL. The highest counts were found in the months of May and June. These results suggest that this organism is found in high number in tested CTs, presenting a potential health risk to the local population.