Mapping immunological and host receptor binding determinants of SARS-CoV spike protein utilizing the Qubevirus platform.

The motifs involved in tropism and immunological interactions of SARS-CoV spike (S) protein were investigated utilizing the Qubevirus platform. We showed that separately, 14 overlapping peptide fragments representing the S protein (F1-14 of 100 residues each) could be inserted into the C terminus of A1 on recombinant Qubevirus without affecting its viability. Additionally, recombinant phage expression resulted in the surface exposure of different engineered fragments in an accessible manner. The F6 from S425-525 was found to contain the binding determinant of the recombinant human angiotensin-converting enzyme 2, with the shortest active binding motif situated between residues S437-492. Upstream, another fragment, F7, containing an overlapping portion of F6 would not bind to recombinant human angiotensin-converting enzyme 2, confirming that a contiguous stretch of residues could adopt the appropriate structural orientation of F6 as an insertion within the Qubevirus. The F6 (S441-460) and other inserts, including F7/F8 (S601-620) and F10 (S781-800), were demonstrated to contain important immunological determinants through recognition and binding of S protein specific (anti-S) antibodies. An engineered chimeric insert bearing the fusion of all three anti-S reactive epitopes improved substantially the recognition and binding to their cognate antibodies. These results provide insights into humoral immune relevant epitopes and tropism characteristics of the S protein with implications for the development of subunit vaccines or other biologics against SARS-CoV.

Protective effect of Bacillus species associated with Rumex dentatus against postharvest soil borne disease in potato tubers and GC-MS metabolite profile.

Potato is constantly exposed to various kinds of phytopathogens which cause diseases during the developmental stage and post-harvest storage. This investigation was designed to assay the anti-phytopathogen activity of bacterial endophytes and their suppressive effects on rot disease in potato. The study also aimed to screen isolates for their plant growth-promoting traits and establish GC-MS-based metabolite profile of the potent isolate. Endophytes were isolated from Rumex dentatus and identified based on 16S rRNA gene. They were screened in dual culture assay against fungal phytopathogens and the potent isolate was tested for its capability to suppress Fusarium rot disease in potato tubers. The mechanism of action of endophytes on the phytopathogens was assessed using scanning electron microcopy. Isolates were also screened in vitro to assay their capability to produce phytohormones, hydrolytic enzymes, and to solubilize phosphates. Endophytic isolates produced proteases with a diameter of halo zone ranging from 7 to 32 mm. Bacillus sp. KL5 exhibited the highest production of indole acetic acid (IAA) with the amount of 104.28 µg/mL and was the most potent antagonist of Fusarium oxysporum and Verticillium dahliae with an inhibitory percentage of 61.53 and 100%, respectively. It showed a reduction of potato rot disease severity by more than 50%. GC-MS of active fractions of KL5 showed the presence of dibutylphthalate and 2,4-di-tert-butylphenol as major metabolites. From this study, it is evident that endophytic Bacillus species from R. dentatus are potent antagonists of F. oxysporum and V. dahliae. Bacillus sp. KL5 is a potent inhibitor of pathogenic F. oxysporum in potato tubers and can be developed as a biocontrol agent.

Plant growth promoting potential of butyl isobutyl phthalate and Streptomyces sp. from Rumex dentatus on rice.

Rice (Oryza sativa L.) is one of the most important staple foods consumed in many countries of the world. It is mostly consumed in developing countries where different chemical fertilizers are used to improve the productivity of the crop plant. In the present study, endophytic actinomycetes isolated from Rumex dentatus were identified morphologically and by scanning electron microscopy. Butyl isobutyl phthalate (BIBP) was isolated from the root endophyte Streptomyces sp. JR9 using column chromatography and HPLC methods. The compound was tested for its effect on rice seed germination. BIBP, extracts, and isolates were evaluated for their plant growth effect on rice in a growth chamber. Isolates were also screened in vitro for phosphate solubilization activity and enzyme production. Indole-3-acetic acid (IAA) and BIBP produced in extracts were quantified and detected using high performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) methods, respectively. BIBP was found to increase the germination of rice seeds by 6 to 12% in treated samples and displayed potent effect at lowest concentration (0.437 µM). Both the compound and the extract depicted significant increases in almost all growth parameters at lowest concentration of 0.125 µg/mL and 62.5 µg/mL, respectively. BIBP also increased significantly shoot length, fresh root, fresh shoot, and dried shoot weight at high concentrations and was more potent than the standard phytohormone IAA. HPLC quantification showed 7.952 µg/mg and 0.371 µg/mg of IAA in extracts of Streptomyces sp. JR9 and the stem endophyte Streptomyces sp. KS3, respectively. IAA containing extract of JR9 increased significantly most growth parameters at lowest concentration (125 µg/mL). The extract of KS3 depicted significant increases in almost all growth parameters at high concentration (500 µg/mL). Our investigation showed that streptomycetes isolated from R. dentatus and BIBP are potent growth promoting agents and can be used in agriculture as bio-fertilizer to improve the growth and productivity of rice. KEY POINTS: • Butyl isobutyl phthalate (BIBP) isolated from endophytic Streptomyces sp. JR9 is a potent rice seed germination activator and promotes significantly the growth of rice • Isolated endophytes showed the ability to produce enzymes and phytohormone IAA • Isolates enhanced significantly the growth of rice.

Antimicrobial and Cytotoxic Potential of Helminthosporin from Rumex abyssiniscus Jacq. Discovered as a Novel Source of Syringic Acid and Bis(2-ethyloctyl) Phthalate.

Rumex abyssinicus Jacq. is a perennial medicinal herb widely used in traditional medicine to treat many diseases. Phytochemicals of the plant were isolated using column chromatography and thin layer chromatography techniques. Extract, fractions and pure compounds were screened for antimicrobial activity against sensitive and multi-drug resistant microbes and their cytotoxicity was performed on different cancer cell lines. The mechanism of action of purified helminthosporin as well as the potent fraction containing a mixture of two compounds was assessed. Fraction R7C3 was the most potent antibacterial with the lowest MIC value of 0.12 µg/mL. Helminthosporin was the most potent compound with the lowest MIC value of 1.95 µg/mL. The compound was more potent than the antibiotic chloramphenicol against multi-drug resistant (MDR) bacteria with MIC equal to 16 µg/mL. The fraction and helminthosporin were shown to destroy the cell wall of the yeast and bacteria, and DNA fragmentation effect on the genome of Candida albicans and Bacillus cereus. Helminthosporin was the most cytotoxic compound with IC50 ˂ 10 µM. Fraction R7C3 showed the most potent cytotoxic effects on all cancer cell lines, with IC50 ranging from ˂1 to 4.35 ng/mL. Our study is the first report on the mechanism of action of helminthosporin, a potent candidate in the development of new drugs against multi-resistant bacteria and cancer cells. In addition, this study uncovered Rumex abyssinicus as a new source of syringic acid and bis(2-ethyloctyl) phthalate.

Antiviral potential of anthraquinones from Polygonaceae, Rubiaceae and Asphodelaceae: Potent candidates in the treatment of SARS-COVID-19, A comprehensive review.

Medicinal plants are being used as an alternative source of health management to cure various human ailments. The healing role is attributed to the hidden dynamic groups of various phytoconstituents, most of which have been recorded from plants and their derivatives. Nowadays, medicinal plants have gained more attention due to their pharmacological and industrial potential. Aromatic compounds are one of the dynamic groups of secondary metabolites (SM) naturally present in plants; and anthraquinones of this group are found to be attractive due to their high bioactivity and low toxicity. They have been reported to exhibit anticancer, antimicrobial, immune-suppressive, antioxidant, antipyretic, diuretic and anti-inflammatory activities. Anthraquinones have been also shown to exhibit potent antiviral effects against different species of viruses. Though, it has been reported that a medicinal plant with antiviral activity against one viral infection may be used to combat other types of viral infections. Therefore, in this review, we explored and highlighted the antiviral properties of anthraquinones of Polygonaceae, Rubiaceae and Asphodelaceae families. Anthraquinones from these plant families have been reported for their effects on human respiratory syncytial virus and influenza virus. They are hence presumed to have antiviral potential against SARS-CoV as well. Thus, anthraquinones are potential candidates that need to be screened thoroughly and developed as drugs to combat COVID-19. The information documented in this review could therefore serve as a starting point in developing novel drugs that may help to curb the SARS-COVID-19 pandemic.

Comprehensive Review of Endophytic Flora from African Medicinal Plants.

Many people in different African countries are suffering from different diseases many of which result in serious life threat and public health problems with high risk of infection and mortality. Due to less accessibility and high cost of modern drugs, people of this continent often depend on traditional medicine using medicinal plants to manage the diseases. Africa has large tropical rain forests, which are very rich in medicinal plants. Many of them have been scientifically proven for their medicinal values. These medicinal plants which constitute a large repertoire of endophytes have not been significantly explored for the isolation of these microorganisms and their bioactive secondary metabolites. This review summarizes the research on endophytes isolated from medicinal plants of Africa, their pharmacological potential and some of their biotechnological aspects. Novel compounds reported from endophytes from Africa with their biological activities have also been reviewed. Information documented in this review might serve as starting point for future researches on endophytes in different African countries.

Evolutionary Qß Phage Displayed Nanotag Library and Peptides for Biosensing.

We selected a novel biotin-binding peptide for sensing biotin, biotinylated proteins, and nucleotides. From a 15-mer library displayed on the RNA coliphage Qß, a 15-amino acid long peptide (HGHGWQIPVWPWGQG) hereby referred to as a nanotag was identified to selectively bind biotin. The target selection was achieved through panning with elution by infection. The selected peptide was tested as a transducer for an immunogenic epitope of the foot-and-mouth disease virus (FMDV) on Qß phage platform separated by a linker. The biotin-tag showed no significant influence on the affinity of the epitope to its cognate antibody (SD6). The nanotag-bound biotin selectively fused either to the C- or N-terminus of the epitope. The epitope would not bind or recognize SD6 while positioned at the N-terminus of the nanotag. Additionally, the biotin competed linearly with the SD6 antibody in a competitive ELISA. Competition assays using the selected recombinant phage itself as a probe or transducer enable the operationalization of this technology as a biosensor toolkit to sense and quantify SD6 analyte. Herein, the published Strep II nanotag (DVEWLDERVPLVET) was used as a control and has similar functionalities to our proposed novel biotin-tag thereby providing a new platform for developing devices for diagnostic purposes.

Mapping immunological and host receptor binding determinants of SARS-CoV spike protein utilizing the Qubevirus platform.

The motifs involved in tropism and immunological interactions of SARS-CoV spike (S) protein were investigated utilizing the Qubevirus platform. We showed that separately, 14 overlapping peptide fragments representing the S protein (F1-14 of 100 residues each) could be inserted into the C-terminus of A1 on recombinant Qubevirus without affecting its viability. Additionally, recombinant phage expression resulted in the surface exposure of different engineered fragments in an accessible manner. The F6 from S 425-525 , was found to contain the binding determinant of the recombinant human angiotensin converting enzyme 2 (rhACE2), with the shortest active binding motif situated between residues S 437-492 . Upstream, another fragment, F7, containing an overlapping portion of F6 would not bind to rhACE2, confirming not just only that residues were linear but equally also the appropriate structural orientation of F6 upon the Qubevirus. The F6 (S 441-460 ) and other inserts, including F7/F8 (S 601-620 ) and F10 (S 781-800 ), were demonstrated to contain important immunological determinants through recognition and binding of S protein specific (anti-S) antibodies. An engineered chimeric insert bearing the fusion of all three anti-S reactive epitopes, improved substantially the recognition and binding to their cognate antibodies. These results provide insights into humoral immune relevant epitopes and tropism characteristics of the S protein with implications for the development of subunit vaccines or other biologics against SARS-CoV. Significance: Mapping epitopes within the receptor binding domains of viruses which are essential for viral tropism is critical for developing antiviral agents and subunit vaccines. In this study we have engineered the surface of Qubevirus to display a peptide library derived from the SARS-CoV S protein. In biopanning with S protein antibodies, we have identified three peptide fragments (EP1, EP2 and EP3) which reacted selectively with antibodies specific to the S protein. We demonstrated that all recombinant phage displayed peptide fragments both individually and as chimera exposed important immunological epitopes to their cognate antibodies. A peptide fragment F6 situated at S 425-525 , was found containing the binding determinant of the recombinant human angiotensin converting enzyme 2 (rhACE2), with the shortest active binding motif situated between residues S 437-492 . The platform is rapidly to identify epitopes and receptor binding sites within viral receptors found in target host cell. Thus, this platform holds great significance.