Nature’s Pharmacy under Siege: Investigating Antibiotic Resistance Pattern in Endophytic Bacteria of Medicinal Plants

Antibiotic resistance poses a significant global health threat, necessitating a thorough understanding of its prevalence in various ecological contexts. Medicinal plants, renowned for their therapeutic properties, host endophytic bacteria that produce bioactive compounds. Understanding antibiotic resistance dynamics in these bacteria is vital for human health and antibiotic efficacy preservation. In this study, we investigated antibiotic resistance profiles in endophytic bacteria from five medicinal plants: Thankuni, Neem, Aparajita, Joba, and Snake plant. We isolated and characterized 113 endophytic bacteria, with varying resistance patterns observed against multiple antibiotics. Notably, 53 strains were multidrug-resistant (MDR), with 14 exhibiting extensive drug resistance (XDR). Thankuni-associated bacteria displayed 44% MDR and 11% XDR, while Neem-associated bacteria showed higher resistance (60% MDR, 13% XDR). Aparajita-associated bacteria had lower resistance (22% MDR, 6% XDR), whereas Joba-associated bacteria exhibited substantial resistance (54% MDR, 14% XDR). Snake plant-associated bacteria showed 7% MDR and 4% XDR. Genus-specific distribution revealed Bacillus (47%), Staphylococcus (21%), and Klebsiella (11%) as major contributors to MDR. Our findings highlight diverse drug resistance patterns among plant-associated bacteria and underscore the complexity of antibiotic resistance dynamics in diverse plant environments. Identification of XDR strains emphasizes the severity of the antibiotic resistance problem, warranting further investigation into contributing factors.

Construction and application of a three-dimensional vascular variation-based nephrometry scoring system for completely endophytic renal tumors

Background:Completely endophytic renal tumors(CERT)pose significant challenges due to their anatomical complexity and loss of visual clues about tumor location.A facile scoring model based on three-dimensional(3D)reconstructed images will assist in better assessing tumor location and vascular variations.Methods:In this retrospective study,80 patients diagnosed with CERT were included.Forty cases underwent preoperative assessment using 3D reconstructed imaging(3D-Cohort),while the remaining 40 cases were assessed using two-dimensional imaging(2D-Cohort).Vascular variations were evaluated by ascertaining the presence of renal arteries>1,prehilar branching arteries,and arteries anterior to veins.The proposed scoring system,termed RAL,encompassed three critical components:(R)adius(maximal tumor diameter in cm),(A)rtery(occurrence of arterial variations),and(L)ocation relative to the polar line.Comparison of the RAL scoring system was made with established nephrometry scoring systems.Results:A total of 48(60%)patients exhibited at least one vascular variation.In the 2D-Cohort,patients with vascular variations experienced significantly prolonged operation time,increased bleeding volume,and extended warm ischemia time compared with those without vascular variations.Conversely,the presence of vascular vari-ations did not significantly affect operative parameters in the 3D-Cohort.Furthermore,the 2D-Cohort demon-strated a notable decline in both short-and long-term estimated glomerular filtration rate(eGFR)changes com-pared with the 3D-Cohort,a trend consistent across patients with warm ischemia time≥25 min and those with vascular variations.Notably,the 2D-Cohort exhibited a larger margin of normal renal tissue compared with the 3D-Cohort.Elevated RAL scores correlated with larger tumor size,prolonged operation time,extended warm is-chemia time,and substantial postoperative eGFR decrease.The RAL scoring system displayed superior predictive capabilities in assessing postoperative eGFR changes compared with conventional nephrometry scoring systems.Conclusions:Our proposed 3D vascular variation-based nephrometry scoring system offers heightened proficiency in preoperative assessment,precise prediction of surgical complexity,and more accurate evaluation of postoper-ative renal function in CERT patients.

Feasibility of enhanced phytoextraction of Zn contaminated soil with Zn mobilizing and plant growth promoting endophytic bacteria

Three bacterial endophytes of Sedum alfredii, VI8L2, II8L4 and VI8R2, were examined for promoting soil Zn bioavailability and Zn accumulation in S. alfredii. Results showed that three strains were re-introduced into S. alfredii rhizosphere soils under Zn stress and resulted in better plant growth, as roots biomass increased from 80% to 525% and shoot biomass from 11% to 47% compared with the uninoculated ones. Strains IVsLz, II8L4 and IVsR2 significantly increased shoot and root Zn concentrations in the ZnCO3 contaminated soil. Inoculation with strain IVsL2 resulted in 44% and 39% higher shoot and root Zn concentrations, while strain IV8R2 significantly decreased shoot Zn concentration in the Zn3（PO4）2 contaminated soils. In the aged contaminated soil, isolates IVsL2, IIsL4 and IVsR2 significantly increased root Zn concentration, but decreased shoot Zn concentration of Sedum alfredii. It suggested that endophytes might be used for enhancing phytoextraction efficiency.

Endophytic Occupation in Nodules of Rhynchosia Plants from Semiarid Regions of Argentina

Beneficial microbes can improve soil health by promoting soil structure,nutrient cycling,and disease suppression.In addition,a wide array of rhizospheric microbes are responsible for producing metabolically active compounds including various types of plant growth regulators.So,microbial biodiversity studies could contribute to the improvement of agricultural practices in deprived areas,such as the Pampean semiarid region.The vast majority of studies conducted on endophytic microorganisms have focused on intensive crop legume species.In contrast,little attention has been paid to microorganisms of native legumes,whose ecology is not directly affected by human action.In this study,endophytic microorganisms isolated from root nodules of a selected native legume of the genus Rhynchosia were characterized.Viable isolates were studied with a focus on their plant growth-pro-moting rhizobacteria(PGPR)properties.Considering the edaphic characteristics of the Pampean semiarid region,the isolates obtained were evaluated for their ability to grow under three salt stress conditions(50,100,and 200 mM NaCl)and four different pH values(6,7,8,and 9).Based on their PGPR activities,the selected strains were phylogenetically grouped using BOX-PCR.The results showed great variability among the isolates in terms of the characteristics studied.Native legumes manifested a wide endophytic variability and remarkable perfor-mance in PGPR activities.We conclude that they could be used as potential bioinoculants for legume cultivation,an excellent alternative to the use of chemical fertilizers that currently pollute the environment.

Studies on the Isolation of Endophytic Bacteria from Tomato Plants and Their Growth-promoting Activities

[Objective] The study aimed to investigate the growth-promoting activities of endophytic bacteria from tomato plants.[Method]The endophytic bacteria isolated from different tissues of tomato plants were analyzed for the effects of their growth-promoting activities on the germination and growth of tomato plants.The bacteria with growth-promoting activity were preliminarily identified.[Result]Totally 59 endophytic bacterial strains were isolated from roots and stems of tomatoes,of which 4 showed significantly growth-promoting activity to germination and growth of tomato.The results suggest that these strains are endowed with the potential capability of growth-promoting.[Conclusion]The endophytic bacteria with growth-promoting activity were found among the isolates from tomato plants.This provided a good foundation for utilization of these bacteria with growth-promoting activity.

Screening for Siderophore-producing Endophytic Bacteria against Fusarium oxysporum

[Objective] With a strain of endophytic antagonistic bacteria with good disease control efficacy in pot as trial material,the paper was to explore the antagonistic mechanism of siderophore against Fusarium oxysporum.[Method] Whether the strain produced siderophore and its fluorescent property was judged by MSA（Modified Sugar-Aspartic acid） plate.The siderophore activity of strains in liquid MSA medium was determined through the absorbance values at particular wavelength.The inhibition effects of it siderophore against F.oxysporum under different concentrations of Fe3＋ were compared.Then the strain was preliminarily identified based on morphological,physiological and biochemical characteristics [Result] The fluorescent siderophore produced by the endophytic bacteria in MSA medium had a strong inhibition effect against F.oxysporum.With the increasing concentration of Fe3＋,the inhibition effect against the pathogen weakened.The preliminary analysis showed these strains belonged to Bacillus.[Conclusion] Bacillus could compete the absorption of Fe3＋ by secreting siderophore to inhibit the growth of F.oxysporum.

Isolation and Identification of a Taxol-producing Endophytic Fungus Identified from Taxus media

[Objective] The aim was to isolate and identify a taxol-producing endophytic fungus from Taxus media. [Method] 32 strains of endophytic fungi were identified form the inner bark of T. media,and their fermentation products were detected by high performance liquid chromatography (HPLC). [Result] Through the screening,a strain of taxol-producing endophytic fungi M57 was obtained,which could produce 45-50 μg/L of taxol,and M57 was defined as Rhizopus sp. through morphological observation and 18S rDNA sequence analysis. [Conclusion] The finding of Rhizopus sp. M57 provided a promising strain for producing taxol with taxol-producing fungi fermentation process.

Determination of Growth-promoting and Antagonistic Action of Endophytic Bacteria Strains Itb57 and Itb295 of Tobacco

[ Objective ] The paper was to study the growth-promoting and antagonistic action of endophytic bacteria strains Itb57 and Itb295 of tobacco to explore their functions in biological control. [ Method] The growth-promoting effects of bacterial suspension ~ff endophytic bacteria Itb57 and Itb295 on tobacco seedling un- der different treatment modes were studied using potting method in greenhouse. The antagonistic action of bacterial suspension of endophytic bacteria Itb57 and Itb295 on Phytophthora nicotianae, Alternaria alternata and Botrytis cinerea were measured by duel culture method. [ Result] Bacterial suspensions of enduphytic bacteria Itb57 and Itb295 had certain growth-promoting effects on tobacco seedling, which could significantly increase the fresh weight and dry weight in aerial part; the growth-promoting effect of soaking ＋ spraying and irrigating treatment was the best. Itb57 strain had good antagonistic action against P. nicotianae. A. alterna- ta and B. cinerea, while Itb295 strain only had good antifungal effect against P. nicotianae. [ Conclusion] The results provided basis for the study and application of tobacco endophytic bacteria strains Itb57 and Itb295 in biocontrol of tobacco diseases.

Impacts of Irrigation with Reclaimed Water on Endophytic Bacteria in Chinese Cabbage in Urumqi City

[Objective] This study aimed to investigate the feasibility of agricultural irri- gation with reclaimed water in Urumqi City. [Method] Chinese cabbages were used as experimental materials and irrigated with control water, 50% reclaimed water and 100% reclaimed water, and then the number of endophytic bacteria in Chinese cab- bages was measured. [Result] Using 50% reclaimed water, the fresh weight of Chi- nese cabbage was improved by 68.94%; however, with the deepening internalization of exogenous microorganism internalization, the total number of endophytic bacteria and coliform flock in Chinese cabbages treated with 50% reclaimed water was sig- nificantly higher than the other two treatment groups （P〈0.05）. [Conclusion] Re- claimed water enhances both the weight of Chinese cabbage and the number of endophytic bacteria, which is unsuitable for irrigation of edible crops such as Chi- nese cabbage. Strengthening disinfection during the process of sewerage treatment could solve this problem.

Isolation and Screening of Black Spot-Resistant Endophytic Fungi Strains from Rosa chinensis

[Objective] This study aimed to isolate and screen black-spot-resistant en- dophytic fungi strains. [Method] Various species of healthy Rosa chinensis were col- lected from Xi＇an, Xianyang, Baoji and Weinan City in Shaanxi Province. Endophytic fungi was isolated from stems and leaves, and purified by 2-3 times of inoculation to screen endophytic fungi antagonistic to Marssonina rosae with modified punching method. [Result] Samples collected from Xianyang exhibited the highest colonization rate and isolation rate; endophytic fungi strains isolated from stems presented the highest colonization rate and isolation rate compared with leaves. A total of 67 en- dophytic fungi strains were isolated from Rosa chinensis, including 3 black spot-re- sistant strains which were all derived from Baoji. [Conclusion] This study laid the foundation for further screening candidate strains of biocontrol fungi and environ- ment-friendly fungal biological control.

Genetic Diversity of Antagonistic Endophytic Bacteria Isolated from Sophora alopecuroide

[ Objective ] The paper was to study and analyze the genetic diversity of 48 endophytic bacteria isolated from Sophora alopecuroide with strong antagonis- tic effect against Fusarium oxysporum and Verticillium dahliae, and carry out sequence measurement and phylogenetic analysis on 10 representative strains. [ Meth- od] Using total DNA ERIC-PCR and 16S rRNA sequencing analysis method, a total of 48 endophytic bacteria were carried out genetic diversity and phylogenetic analysis. [Result] ERIC-PCR fingerprint map showed that 48 endophytic bacteria were divided into 6 ERIC groups and 2 strains with independent groups at Wat- son distance of 0.31. Representative strains were selected from each group to determine 16S rRNA gene for phylogenetic analysis, and the results showed that these strains belonged to Bacillus atrophaeus, Bacillus subtilis, Bacillus cereus, Bacillus megaterium, Bacillus pumilus and Serratia marcescens, respectively. [ Conclu- sion ] The genetic diversity of 48 strains of bacteria was obvious. Determination of 48 strains of bacteria on diversity and phylogenetic status could lay foundation for study on mechanism of F. oxysporum and V. dahliae, which could also provide new strain sources for biological control of diseases in cotton production.

Separation and Purification of Antioxidant Endophytic Fungi from Stevia Leaves

[Objective] The aim was to separate and purify antioxidant endophytic fun- gi from Stevia leaves. [Method] Stevia leaves were used as materials for endophytic fungi screening and purification. Then, liquid culture was conducted to test fermented supernatant abilities of antioxidant, DPPH free radical clearing and chemilumines- cence inhibition. [Result] Forty two endophytic fungi were separated from F01 to F42. F41 was proved to be the strongest in antioxidant aspect; F37 was the strongest in DPPH clearing; F02, F07, F20, F25, F37, and F416 stood out in a comprehensive comparison of the above aspects. Flow injection chemiluminescenct method was used to test the above six fermentation liquors, and the results showed that antioxidant abilities of the six were quite strong for all of chemiluminescence rates were over 70%. Through preliminary morphological identification, bacterial colony was found white or yellowy and only F02 was of dissepiment. [Conclusion] Fungi with high-antioxidation activities were separated, which paved the way for future research.

Preliminary Studies on Antifungal Activity of Xanthium sibiricum and the Endophytic Fungi

[Objective] The study was aimed to provide theoretical foundation for developing a new fungicide with high activity,low toxicity and low residua.[Method] Based on the plant growth rate and inhibition zone methods,the antifungal activity of extracts of every organ of Xanthium sibiricum,the fermentation liquid of endophytic fungi MC4-23,and the extract of mycelium were tested with six kinds of pathogenic fungi and three kinds of pathogenic bacteria respectively.[Result] The extract of every organ of Xanthium sibiricum,the fermentation liquid of endophyic fungi MC4-23,and the extract of mycelium all had antifungal activities.The acetone extracts of Xanthium sibiricum had stronger antifungal activities than the ethanol extracts of Xanthium sibiricum.The extracts of leaf have the strongest antifungal activities against fungi and bacteria.The inhibition rates of the six pathogenic fungi were all above 47.8%,and the inhibition diameters of the three pathogenic bacteria were all above 0.5 centimeter.The fermentation liquid of endophytic fungi also had strong activities against the fungi and bacteria.The inhibition rates of the six pathogenic fungi were all above 68.1%.The extracts of root of Xanthium sibiricum and the mycelium of endophytic fungi were all had antifungal activities.[Conclusion] Xanthium sibiricum and endophyic fungi MC4-23 had great value of development and utilization.

Identification of Taxus cuspidata Sieb.et Zucc endophytic fungi─ new recorded-genus-species of China and the metabolite

A total of 94 isolates of endophytic fungi were isolated from the bark of 200-yr.-old Taxus cuspidata Sieb. et Zucc. in the primeval forest of the Changbai Mountain Natural Reserve, and 19 species of endophytic fungi were identified. Among the taxa 2 new species and 1 new varied species as well as the 6 known species of China had been described in a previous article published on Journal of Forestry Research, Vol. 14 (4): 290-294 (2003). This paper made a detail description on the morpho-logical characteristics of the 9 new recorded-genus-species and 1 new record species with illustrations, including Staphy-lotrichum coccosporum Meyer et Nicot, Botryodema lateritium Papendoorf & Upadhyay, Oedocephalum glomerulosum (Rul-liard) Sacc., Trichosporonoides oedocephalis Haskins & Spencer, Beniowskia sphaeroidea (Kal Chbrenner et Caeke) Mason, Thermomyces verrucosus Pugh, Blakeman & Morgan-jones, Gilmaniella humicola Brron , Arthrinium sphaerosperma (Per.:Fr.) Ellis, Ambrosiella brunnea (Verrall) Batra, and Cytosporina notha Diedz (1 new record species). And the habitats, hosts, distribu-tions and fruiting periods were recorded for the new recorded-genus-species. The determination result of thin layer chroma-tography (TLC) showed that the liquid fermentation metabolites of the strains could not react positively with taxoids developer - the vanillic aldehyde and dilute bismuth potassium iodide.

Isolation of the Endophytic Fungi of Paris polyphylla var. yunnanensis and Their Effects on the Embryo Development of P. polyphylla var. yunnanensis Seeds

[Objective] This study aimed to isolate the endophytic fungi of Paris polyphylla var. yunnanensis and investigate their effects on the embryo development of P. polyphylla var. yunnanensis seeds. [Method] The endophytic fungi of P. polyphylla were isolated and identified morphologically, and their effects on the embryo development of P. polyphylla var. yunnanensis seeds were studied by using paraffin sectioning and microphotography. [Result] Nine endophytic fungi, i.e. P. polyphylla var. yunnanensis endophytic fungi PPYEF-1, PPYEF-2, PPYEF-3, PPYEF-4, PPYEF-5, PPYEF-6, PPYEF-7, PPYEF-8 and PPYEF-9 belonging to seven genera in five families, three orders were isolated from the rhizomes. Except PPYEF-4 (Cladosporium sp.), other fungi could promote the embryo development of the P. polyphylla var. yunnanensis seeds, mostly reaching the extremely significant or significant level. PPYEF-9 (Trichoderma sp.) resulted in the highest embryo length and embryo-emerging ratio. [Conclusion] This paper could provide a reference for the application of the endophytic fungi of P. polyphylla var. yunnanensis in the dormancy-breaking of P. polyphylla var. yunnanensis seeds.

Effect of Endophytic Fungi AL12 on the Metabolites Distribution in Organs of Atraetylodes lancea

[Objective] To examine the effect of endophytic fungi ALl2 （Gilmaniella sp.） on metabolites distribution in organs of Atractylodes lancea. [Method] Endophytic fungi ALl2 was inoculated on Atraetylodes lancea plantlets in tissue culture, and the distribution of cellulose, hemicellulose, lignin, soluble sugar in leaves and roots of the inoculated group were detected. The weight of leaves and roots were compared. Gas Chromatography was used to analyze the volatile oil components. [Result] Compared with the control group, the average fresh weight and dry weight of leaves and roots of A. lancea which had been symbiosed with ALl2 increased significantly. The content of lignin and soluble sugar increased in the leaves of the inoculated group, and the content of cellulose, hemicellulose, lignin, soluble sugar and volatile oil also increased in roots. [Conclusion] The results indicate that symbiosed with ALl2 is benefit for the development of A. lancea roots and can promote the transfer and accumulation of the medicinal components to the roots.

Elicitation on Artemisinin Biosynthesis in Artemisia annua Hairy Roots by the Oligosaccharide Extract from the Endophytic Colletotrichum sp. B501

The oligosaccharide elicitor from the mycelial wall of an endophytic Colletotrichum sp. B501 promoted the production of artemisinin in Artemisia annua L. hairy root culture. When hairy roots of 22-day-old cultures (later growth phase) were exposed to the elicitor (20 mg/L) for 4 d, the maximum content of artemisinin reached 1.15 mg/g, a 64.29% increment over the control. The electron X-ray microanalysis disclosed the rapid accumulation of Ca 2+ in the elicited cortical cells of hairy root. The electronic microscope observation revealed the high electron density area in vacuole of elicited cells. During the first day of elicitation the peroxidase activity of hairy roots was improved sharply. Some cellular morphological changes including cell shrinkage, condensation of cytoplasm and nuclear fragmentation, coincident with the appearance of DNA ladders, were observed after the third day of elicitation. It was suggested that the oligosaccharide elicitor triggered the programmed cell death, which may provide the substance or chemical signal for artemisinin biosynthesis.

Chemical constituents of marine mangrove-derived endophytic fungus Alternaria tenuissima EN-192

A chemical investigation of the ethyl acetate extract of the fermentation broth of Alternaria tenuissima EN- 192, an endophytic fungus obtained from the stems of the marine mangrove plant Rhizophora stylosa, resulted in the isolation of nine known secondary metabolites, including four indole-diterpenoids： penijanthine A （1）, paspaline （2）, paspalinine （3）, and penitrem A （4）; three tricycloalternarene derivatives： tricycloalternarene 3a （5）, tricycloalternarene lb （6）, and tricycloalternarene 2b （7）; and two alternariol congeners： djalonensone （8） and alternariol （9）. The chemical structures of these metabolites were characterized through a combination of detailed spectroscopic analyses and their comparison with reports from the literature. The inhibitory activities of each isolated compound against four bacteria were evaluated and compounds 5 and 8 displayed moderate activity against the aquaculture pathogenic bacterium Vibrio anguillarum, with inhibition zone diameters of 8 and 9 mm, respectively, at 100 gg/disk. To the best of our knowledge, this is the first report on the secondary metabolites of mangrove-derived Alternaria tenuissima and also the first report of the isolation ofindole-diterpenoids from fungal genus Alternaria.

Biogenesis of antibacterial silver nanoparticles using the endophytic bacterium Bacillus cereus isolated from Garcinia xanthochymus

Objective:To synthesize the ecofriendly nanoparticles,which is viewed as an alternative to the chemical method which initiated the use of microbes like bacteria and fungi in their synthesis.Methods:The current study uses the endophytic bacterium Bacillus cereus isolated from the Garcinia xanthochymus to synthesize the silver nanoparticles(AgNPs).The AgNPs were synthesized by reduction of silver nitrate solution by the endophytic bacterium after incubation for 3-5 d at room temperature.The synthesis was initially observed by colour change from pale white to brown which was confirmed by UV-Vis spectroscopy.The AgNPs were further characterized using FTIR,SEM-EDX and TEM analyses.Results:The synthesized nanoparticles were found to he spherical with the size in the range of 20-40 nm which showed a slight aggregation.The energy-dispersive spectra of the nanoparticle dispersion confirmed the presence of elemental silver.The AgNPs were found to have antibacterial activity against a few pathogenic bacteria like Escherichia coli,Pseudomonas aeruginosa,Staphylococcus aureus,Salmonella typhi and Klebsiella pneumoniae.Conclusions:The endophytic bacteria identified as Bacillus cereus was able to synthesize silver nanoparticles with potential antibacterial activity.

A New Cytotoxic Sterol Produced by an Endophytic Fungus from Castaniopsis fissa at the South China Sea Coast

A new sterol, ergosta-8(9),22-diene-3,5,6,7-tetraol(3β,5α,6β,7α,22E)(A) together with three known sterols: 3β, 5α,6β-trihydroxyergosta-7,22-diene (B), 3β-hydroxy-5α,8α- epidioxyer-gosta-6,22-diene (C) and ergosterol (D) were isolated from the mycelia of an unidentified endophytic fungus separated from Castaniopsis fissa (chestnut tree). Compound A exhibited potent selective cytotoxicity against Bel-7402, NCI4460 and L-02 cell lines with IC50 values 8.445, 5.03, 13.621 μg/mL, respectively.