Endophytic Bacillus subtilis P10 from Prunus cerasifera as a biocontrol agent against tomato Verticillium wilt

Endophytic bacteria serve key roles in the maintenance of plant health and growth. Few studies to date, however, have explored the antagonistic and plant growth-promoting (PGP) properties of Prunus cerasifera endophytes. To that end, we isolated endophytic bacteria from P. cerasifera tissue samples and used a dual culture plate assay to screen these microbes for antagonistic activity against Verticillium dahliae, Botryosphaeria dothidea, Fusarium oxysporum, F. graminearum, and F. moniliforme. Of the 36 strains of isolated bacteria, four (strains P1, P10, P16, and P20) exhibited antagonistic effects against all five model pathogens, and the P10 strain exhibited the strongest antagonistic to five pathogens. This P10 strain was then characterized in-depth via phenotypic assessments, physiological analyses, and 16s rDNA sequencing, revealing it to be a strain of Bacillus subtilis. Application of a P10 cell suspension (1×108 CFU/mL) significantly enhanced the seed germination and seedling growth of tomato in a greenhouse setting. This P10 strain further significantly suppressed tomato Verticillium wilt with much lower disease incidence and disease index scores being observed following P10 treatment relative to untreated plants in pot-based experiments. Tomato plants that had been treated with strain P10 also enhanced defense-related enzymes, peroxidase, superoxide dismutase, and catalase activity upon V. dahliae challenge relative to plants that had not been treated with this endophytic bacterium. The results revealed that the P10 bacterial strain has potential value as a biocontrol agent for use in the prevention of tomato Verticillium wilt.

Fast-Tracking Isolation, Identification and Characterization of New Microalgae for Nutraceutical and Feed Applications

Microalgae contain high-value biochemical compounds including fatty acids (FA), protein and carotenoids, and are promising bioresources to enhance nutrition of food and animal feed. Important requirements for commercial strains are rapid growth and high productivities of desirable compounds. As these traits are believed to be found in aquatic environments with fluctuating conditions, we collected microalgae from marine and freshwater environments that are subjected to eutrophication and/or tidal fluctuations. Using this directed approach, 40 monoalgal cultures were isolated and 25 identified through 18S rDNA sequencing and morphological characterization. Based on their high growth rates (0.28–0.60 day−1) and biomass productivities (0.25–0.44 g L−1day−1) in commercial fertilizer under standardized conditions, six new strains were selected. Scenedesmus sp. GW63 produced quality FA-rich biomass with high omega-3 polyunsaturated FA (28.5% of total FA (TFA)) contents, especially α-linolenic acid (ALA; 20.0% of TFA) with a very low n-6/n-3 ratio (0.4), and high FA productivity (32.6 mg L−1 day−1). A high protein productivity (34.5 mg L−1 day−1) made Desmodesmus sp. UQL1_26 (33.4% of dry weight (DW)) attractive as potential protein-rich feed and nutrition supplement. Monoraphidium convolutum GW5 displayed valuable carotenoid production (0.8% DW) with high carotenoid accumulation capability (0.8 mg L−1 day−1). This research provides a pathway for fast-tracking the selection of high-performing local microalgae from different environments for nutraceuticals, functional foods and animal feed applications.

The Nasal Microbiome of Predicting Bronchopulmonary Dysplasia in Preterm Infants

Bronchopulmonary dysplasia (BPD) is chronic lung disease of prematurity and associated with substantial long-term disabilities. To characterize and compare the nasal swabs microbiome of early stage in premature infants and determine whether microbial diversity or composition in the airway associated with BPD disease. We performed a prospective observational cohort design. Preterm neonates less than 32 weeks of gestation were recruited from NICU, Children's Hospital, Zhejiang University School of Medicine from 2019 to 2020. Sterile foam swabs were collected from anterior nares at 1 and 3 weeks of postnatal age. We used PCR amplification and 16S rDNA sequencing. Neonatal demographic data including gestational age, birth weight, medication administration history were recorded. A total of 98 nasal swabs samples were collected from 54 preterm infants, 13 developed BPD infants and 41 control infants were finally involved in the study. Birth weights ranged from 700 to 2,050 g. Gestational age ranged from 25 2/7to 31 6/7. We found increased in the expression of Prevotella, Marinomonas, Enterobacteriaceae, Weissella, Selenomonas, Oribacterium, Nubsella and Antricoccus in BPD group at two time points. Prevotella was correlated with the severity of BPD (Spearman r=0.361, P=0.000). Given possible roles for noninvasive upper airway microbiota in BPD pathobiology, the nasal microbiome in BPD is a compelling area of research to continue to expand.

Multi-Omics Analysis Reveals Changes in the Intestinal Microbiome, Transcriptome, and Methylome in a Rat Model of Chronic Non-bacterial Prostatitis: Indications for the Existence of the Gut-Prostate Axis

Chronic non-bacterial prostatitis (CNP) is one of the most prevalent diseases in human males worldwide. In 2005, the prostate-gut axis was first proposed to indicate the close relationship between the prostate and the intestine. This study investigated CNP-induced changes of the gut microbiota, gene expression and DNA methylation in a rat model by using multi-omics analysis. Firstly, 16S rDNA sequencing presented an altered structure of the microbiota in cecum of CNP rats. Then, transcriptomic analysis revealed that the expression of 185 genes in intestinal epithelium was significantly changed by CNP. These changes can participate in the immune system, digestive system, metabolic process, etc. Finally, methylC-capture sequencing (MCC-Seq) found 73,232 differentially methylated sites (DMSs) in the DNA of intestinal epithelium between control and CNP rats. A combined analysis of methylomics and transcriptomics suggested an epigenetic mechanism for CNP-induced differential expression genes correlated with intestinal barrier function, immunity, metabolism, enteric infectious disease, etc. More importantly, the transcriptomic, methylomic and gut microbial changes were highly correlated with multiple processes including intestinal immunity, metabolism and epithelial barrier function. In this study, disrupted homeostasis in the gut microbiota, gene expression and DNA methylation were reported in CNP, which supports the existence of the gut-prostate axis.

First Report of Fecal Microflora of Wild Bar-Headed Goose in Tibet Plateau

The bar-headed goose (Anser indicus) has two black spots on its head. It is considered an important bird in China. It breeds in plateau lakes, especially saltwater lakes, and swamp areas. However, the intestinal flora of wild bar-headed geese in the Tibet Autonomous Region is currently not known. In this study, 16S rDNA sequencing was performed on the intestinal microbes of wild bar-headed geese. A total of 513,505 reads of raw data were obtained, and the results analyzed the average number of 128,376 ± 2,392 reads per sample. The microbiota of all samples consists of 10 main bacterial phyla, including Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, Cyanobacteria, Patescibacteria, Deferribacteres, Planctomy-cetes, Fusobacteria, and Tenericutes. The results indicated that Firmicutes (67.34%) was the predominant phylum, followed by Proteobacteria (29.03%) and Cyanobacteria (1.97%). In our research, we identified the intestinal flora of the wild bar-headed goose, which provides valuable information for further research on the gene function of the bar-headed goose and the intestinal flora of wild animals. These findings are also useful and valuable for genetic and high-altitude research in the Tibet Autonomous Region.

Metagenomic analysis decodes the fungal diversity of bio-dynamic preparations

Aim: Biodynamic farming system involves use of 8 different biodynamic preparations (BD 500-BD 507). Multi functionality of any ecosystem is due to its microbial diversity and community composition of microbes. So the present study was aimed to determine the total fungal population viz. unculturable ones, metagenomic analysis was done. Methodology: In the present study, 18S rDNA sequencing of V3-V4 amplicon regions was performed to identify and characterize fungal diversity, which existed in these preparations. Results: Alpha diversity was found to be maximum in BD506 with 868 OTU (operational taxanomic units) and minimum in BD507 with 254 OTU. At phylum level, the most abundant phylum was Ascomycota as recorded in 7 BD preparations with exception in the BD 500 (Unassigned). At genus level highest percentage of OTU abundance was observed for unassigned genus in all BD preparations, except Mortierella in BD 500 and BD 502; Microascus in BD 501 and BD504; Gymnoascus in BD503, Scedosporium in BD 505, Mucor in BD 506 and Hyphopichia in BD 507. On the basis of species diversity, BD502, 503 and 506 showed high percentage of OTU abundance for Mucor racemosus, while Mortierella oligospora was abundant in BD500, Dipodascus geotrichum in BD 501, Kernia pachypleura in BD504, Petriella setifera in BD505 and Hyphopichia burtonii in BD 507. Interpretation: This indicated a unqiue class of fungus predominating each type of BD preparation. Furthermore, a large proportion of unassigned fungi at phylum and genus level were detected in metagenome analysis which might have specific roles in contributing for their overall effectiveness of each kind of BD preparations.

Experimental Evidence for Manure-Borne Bacteria Invasion in Soil During a Coalescent Event: Influence of the Antibiotic Sulfamethazine

The fertilization of agricultural soil by organic amendment that may contain antibiotics, like manure, can transfer bacterial pathogens and antibiotic-resistant bacteria to soil communities. However, the invasion by manure-borne bacteria in amended soil remains poorly understood, being hardly observed. Here, we assessed the invasions of manure-borne bacteria during a coalescence event between manure and soil, in different soils and in the presence or absence of antibiotics. To this end, microcosms of four different soils were amended or not with manure at an agronomical dose and/or exposed or not to the antibiotic sulfamethazine (SMZ). After one month of incubation, the diversity, structure and composition of bacterial communities of the soils were assessed by 16S rDNA sequencing. The invasion of manure-borne bacteria was still perceptible one month after the soil amendment. The results obtained with the soil already amended in situ with manure six months prior to the experiment suggest that some of the bacterial invaders were established in the community over the long term. Even if differences were observed between soils, the invasion was mainly attributable to some of the most abundant OTUs of manure (mainly Firmicutes). SMZ exposure had a limited influence on soil microorganisms. It was significant in only one soil, where it enhanced the invasion potential of some manure-borne invaders.

Benthic Cyanobacterial Diversity and Antagonistic Interactions in Abrolhos Bank: Allelopathy, Susceptibility to Herbivory, and Toxicity

Benthic cyanobacterial mats (BCMs) are conspicuous components of coral reef communities, where they play key ecological roles as primary producers among others. BCMs often bloom and might outcompete neighboring benthic organisms, including reef-building corals. We investigated the cyanobacterial species composition of three BCMs morphotypes from the marginal reef complex of Abrolhos Bank (Southeastern Brazil). Also, we assessed their allelopathic effects on coral zooxanthellae, their susceptibility to herbivory by fish, and their toxicity to brine shrimp nauplii. Morphology and 16S rDNA sequencing unveiled the cyanobacteria Moorena bouillonii, Okeania erythroflocculosa, Adonisia turfae, Leptolyngbya sp., and Halomicronema sp. as components of BCMs from Abrolhos. BCMs cell-free filtrates and extracts exerted an allelopathic effect by reducing the growth of the ex hospite Symbiodinium sp. in culture. BCMs-only treatments remained untouched in field susceptibility assays in contrast to macroalgae only and mixed BCMs-macroalgae treatments that had the macroalgae fully removed by reef fish. Crude aqueous extracts from BCMs were toxic to brine shrimps in acute assays. Besides unveiling the diversity of BCMs consortia in Abrolhos, our results cast some light on their allelopathy, antiherbivory, and toxicity properties. These antagonistic interactions might promote adverse cascading effects during benthic cyanobacteria blooms and in gradual shifts to BCMs-dominated states.

New Biocalcifying Marine Bacterial Strains Isolated from Calcareous Deposits and Immediate Surroundings

Marine bacterial biomineralisation by CaCO3 precipitation provides natural limestone structures, like beachrocks and stromatolites. Calcareous deposits can also be abiotically formed in seawater at the surface of steel grids under cathodic polarisation. In this work, we showed that this mineral-rich alkaline environment harbours bacteria belonging to different genera able to induce CaCO3 precipitation. We previously isolated 14 biocalcifying marine bacteria from electrochemically formed calcareous deposits and their immediate environment. By microscopy and µ-Raman spectroscopy, these bacterial strains were shown to produce calcite-type CaCO3. Identification by 16S rDNA sequencing provided between 98.5 and 100% identity with genera Pseudoalteromonas, Pseudidiomarina, Epibacterium, Virgibacillus, Planococcus, and Bhargavaea. All 14 strains produced carbonic anhydrase, and six were urease positive. Both proteins are major enzymes involved in the biocalcification process. However, this does not preclude that one or more other metabolisms could also be involved in the process. In the presence of urea, Virgibacillus halodenitrificans CD6 exhibited the most efficient precipitation of CaCO3. However, the urease pathway has the disadvantage of producing ammonia, a toxic molecule. We showed herein that different marine bacteria could induce CaCO3 precipitation without urea. These bacteria could then be used for eco-friendly applications, e.g., the formation of bio-cements to strengthen dikes and delay coastal erosion.

Plant growth promoting potential of multifarious endophytic Pseudomonas lini strain isolated from Cicer arietinum L.

Chickpea (Cicer arietinum), one of the major pulse crops in India, endured extreme reduction in production due to various abiotic and biotic stresses. Endophytic bacteria residing in the nodules and roots of chickpea plants enable host in combating these stresses. Twenty one endophytic bacteria isolated from nodules and roots of chickpea were screened for multiple plant growth promoting traits like ammonia, organic acid, siderophore, hydrogen cyanide (HCN) and phytohormone indole acetic acid (IAA) production. Out of these, 86% isolates produced ammonia, around 50% isolates produced organic acid, HCN and siderophore, 29% isolates produced ACC (1-aminocyclopropane-1-carboxylic acid) deaminase, while only 14% isolates solubilized phosphate. Interestingly, all the isolates were able to produce IAA ranging from 11.6–85.2 μg/ml, isolate CPJN 13 being the maximum IAA producer (85.5±2.33 μg/ml). Isolate CPJN13 was selected for IAA optimization studies. The yield of IAA increased up to 4 fold i.e. 331±4.96 μg/ml at optimized conditions. IAA production was also confirmed by TLC and HPLC analysis of crude IAA extract. The application of CPJN13 on chickpea seedlings resulted in significant increase in plant growth parameters. The 16S rDNA sequencing of CPJN13 revealed its similarity with Pseudomonas lini strain and submitted to NCBI with accession number MF574502. To best of our knowledge, this is the first report of the presence of P. lini as endophyte in chickpea nodules. The results of this study imply that the endophytic P. lini has a potential role to enhance the plant growth.