In silico biotechnological potential of Bacillus sp. strain MHSD_37 bacterial endophyte.

BACKGROUND: Endophytic bacteria possess a range of unique characteristics that enable them to successfully interact with their host and survive in adverse environments. This study employed in silico analysis to identify genes, from Bacillus sp. strain MHSD_37, with potential biotechnological applications. RESULTS: The strain presented several endophytic lifestyle genes which encode for motility, quorum sensing, stress response, desiccation tolerance and root colonisation. The presence of plant growth promoting genes such as those involved in nitrogen fixation, nitrate assimilation, siderophores synthesis, seed germination and promotion of root nodule symbionts, was detected. Strain MHSD_37 also possessed genes involved in insect virulence and evasion of defence system. The genome analysis also identified the presence of genes involved in heavy metal tolerance, xenobiotic resistance, and the synthesis of siderophores involved in heavy metal tolerance. Furthermore, LC-MS analysis of the excretome identified secondary metabolites with biological activities such as anti-cancer, antimicrobial and applications as surfactants. CONCLUSIONS: Strain MHSD_37 thereby demonstrated potential biotechnological application in bioremediation, biofertilisation and biocontrol. Moreover, the strain presented genes encoding products with potential novel application in bio-nanotechnology and pharmaceuticals.

Bioremediation of Pb contaminated water using a novel Bacillus sp. strain MHSD_36 isolated from Solanum nigrum.

The Pb bioremediation mechanism of a multi-metal resistant endophytic bacteria Bacillus sp. strain MHSD_36, isolated from Solanum nigrum, was characterised. The strain tested positive for the presence of plant growth promoters such as indoleacetic acid, 1-aminocyclopropane-1-carboxylate deaminase, siderophores, and phosphate solubilization. The experimental data illustrated that exopolysaccharides and cell hydrophobicity played a role in Pb uptake. The data further showed that the cell wall biosorbed a significant amount (71%) of the total Pb (equivalent to 4 mg/L) removed from contaminated water, compared to the cell membrane (11%). As much as 11% of the Pb was recovered from the cytoplasmic fraction, demonstrating the ability of the strain to control the influx of toxic heavy metals into the cell and minimize their negative impacts. Pb biosorption was significantly influenced by the pH and the initial concentration of the toxic ions. Furthermore, the presence of siderophores and biosurfactants, when the strain was growing under Pb stress, was detected through liquid chromatography mass spectrometry. The strain demonstrated a multi-component based Pb biosorption mechanism and thus, has a great potential for application in heavy metal bioremediation.

Draft Genome Sequence of Pantoea sp. Strain MHSD4, a Bacterial Endophyte With Bioremediation Potential.

Pantoea sp. strain MHSD4 is a bacterial endophyte isolated from the leaves of the medicinal plant Pellaea calomelanos. Here, we report on strain MHSD4 draft whole genome sequence and annotation. The draft genome size of Pantoea sp. strain MHSD4 is 4 647 677 bp with a G+C content of 54.2% and 41 contigs. The National Center for Biotechnology Information Prokaryotic Genome Annotation Pipeline tool predicted a total of 4395 genes inclusive of 4235 protein-coding genes, 87 total RNA genes, 14 non-coding (nc) RNAs and 70 tRNAs, and 73 pseudogenes. Biosynthesis pathways for naphthalene and anthracene degradation were identified. Putative genes involved in bioremediation such as copA, copD, cueO, cueR, glnGm, and trxC were identified. Putative genes involved in copper homeostasis and tolerance were identified which may suggest that Pantoea sp. strain MHSD4 has biotechnological potential for bioremediation of heavy metals.

In Silico Analysis of Novel Bacterial Metabolites with Anticancer Activities.

Resistance to anticancer therapeutics is a major global concern. Thus, new anticancer agents should be aimed against novel protein targets to effectively mitigate the increased resistance. This study evaluated the potential of secondary metabolites from a bacterial endophyte, as new anticancer agents, against a novel protein target, fibroblast growth factor. In silico genomic characterization of the Bacillus sp. strain MHSD_37 was used to identify potential genes involved in encoding secondary metabolites with biological activity. The strain was also exposed to stress and liquid chromatography-mass spectrometry used for the identification and annotation of secondary metabolites of oligopeptide class with anticancer activity. Selected metabolites were evaluated for their anticancer activity through molecular docking and Absorption, Distribution, Metabolism, Excretion and Toxicity (ADMET) properties analysis. Phylogenetic analysis revealed that strain MHSD_37 shared close evolutionary relationships with Bacillus at the species level, with no identified relationships at the sub-species level. Both in silico genomic characterization and spectrometry analysis identified secondary metabolites with potential anticancer activity. Molecular docking analysis illustrated that the metabolites formed complexes with the target protein, fibroblast growth factor, which were stabilized by hydrogen bonds. Moreover, the ADMET analysis showed that the metabolites passed the toxicity test for use as a potential drug. Thereby, Bacillus sp. strain MHSD_37 is a potential novel strain with oligopeptide metabolites that can be used as new anticancer agents against novel protein targets.

Bacillus dicomae sp. nov., a new member of the Bacillus cereus group isolated from medicinal plant Dicoma anomala.

A Gram-stain-positive, endospore-forming endophytic bacterial strain designated MHSD28T was isolated from surface-sterilized leaves of Dicoma anomala collected from Eisleben, Botlokwa, Limpopo Province, South Africa. The phenotypic and phylogenetic characteristics of strain MHSD28T were consistent with those of members within the Bacillus cereus group. Comparative analysis between this strain and its relatives confirmed that it belongs to this group and forms a monophyletic branch. The digital DNA-DNA hybridization values between strain MHSD28T and its relatives were lower than the 70 % threshold for species delineation. To further determine its phylogenetic position, multi-locus sequence analysis (MLSA) based on five concatenated housekeeping gene (gyrB, atpD, DnaK, rpoB and rpoD) sequences, phenotypic analysis, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) biotyper identification, fatty acid and polar lipid profile analyses were carried out. Phenotypic characterization, MLSA, whole genome sequence based analyses and MALDI-TOF results placed strain MHSD28T within the B. cereus group. The major fatty acids were iso-C15 : 0 and summed feature 3 and the main polar lipids were diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. The respiratory quinone was menaquinone-7. The cell-wall peptidoglycan structure included meso-diaminopimelic acid. Considering the above results, strain MHSD28T represents a novel species of the B. cereus group, for which the name Bacillus dicomae sp. nov. is proposed. The type strain is MHSD28T (=BD 2262T=LMG 32287T=CECT 30671T).

Genetic Engineering of Escherichia coli BL21 (DE3) with a codon-optimized insecticidal toxin complex gene tccZ.

A toxin complex consists of a high-molecular-weight group of toxins that exhibits insecticidal activity against insect pests. These toxins are a promising alternative to Bacillus thuringiensis (Bt) toxins that have been extensively utilized in insect pest control. Herein, a codon-optimized insecticidal gene (tccZ) (381 bp) identified in Pantoea ananatis strain MHSD5 (a bacterial endophyte previously isolated from Pellaea calomelanos) was ligated into the pET SUMO expression vector and expressed in Escherichia coli BL21 (DE3). We report the success of cloning the tccZ gene into the pET SUMO vector and ultimately the transformation into E. coli BL21 (DE3) competent cells. However, despite conducting a time course of expression as well as isopropyl ß-d-1-thiogalactopyranoside (IPTG) dosage optimization to determine optimal conditions for expression, TccZ protein expression could not be detected on Stain-Free and Coomassie-stained SDS-PAGE gels.

Exploring indigenous freshwater chlorophytes in integrated biophotovoltaic system for simultaneous wastewater treatment, heavy metal biosorption, CO2 biofixation and biodiesel generation.

The study explored the combined photosynthetic activities of two green microalgal species, Tetradesmus obliquus and Tetradesmus reginae, on an integrated biophotovoltaic (BPV) platform for simultaneous wastewater treatment, toxic metal biosorption, carbon biofixation, bioelectricity generation and biodiesel production. The experimental setup comprised of a dual-chambered BPV with copper anode surrounded by T. obliquus in BG11 media, and copper cathode with T. reginae in municipal wastewater separated by Nafion 117 membrane. The study reported a maximum power density of 0.344 Wm-2 at a cell potential of 0.415 V with external resistance of 1000 Ω and 0.3268 V maximum open-circuit voltage. The wastewater electrical conductivity and pH increased from 583 ± 22 to 2035 ± 29.31 mS/cm and 7.403 ± 0.174 to 8.263 ± 0.055 respectively, signifying increased photosynthetic and electrochemical activities. Residual nitrogen, phosphorus, chemical oxygen demand, arsenic, cadmium, chromium and lead removal efficiencies by T. reginae were 100%, 80.68%, 71.91%, 47.6%, 88.82%, 71.24% and 92.96%, respectively. T. reginae accumulated maximum biomass of 0.605 ± 0.033 g/L with a CO2 biosequestration rate of 0.166 ± 0.010 gCO2/L/day and 42.40 ± 1.166% lipid content. Methyl palmitate, methyl undecanoate and 13-octadecenoic acid with relative abundances of 37.24%, 24.80% and 12.02%, respectively were confirmed.

The Antibacterial, Antitumor Activities, and Bioactive Constituents' Identification of Alectra sessiliflora Bacterial Endophytes.

Due to increased antimicrobial resistance against current drugs, new alternatives are sought. Endophytic bacteria associated with medicinal plants are recognized as valuable sources of novel secondary metabolites possessing antimicrobial, antitumor, insecticidal, and antiviral activities. In this study, five bacterial endophytes were isolated and identified from the medicinal plant, Alectra sessiliflora, and their antibacterial and antitumor activities were investigated. In addition, the crude extracts of the endophytes were analyzed using gas chromatography (GC) coupled with time-of-flight mass spectrometry (TOF-MS). The identified bacterial endophytes belong to three genera viz Lysinibacillus, Peribacillus, and Bacillus, with the latter as the dominant genus with three species. Ethyl acetate extracts from the endophytes were used for antimicrobial activity against eleven pathogenic strains through minimum inhibitory concentration (MIC). The antitumor activity against the Hela cervical, Hek 293 kidney, and A549 lung carcinoma cells was determined by the MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxy-phenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay. Lysinibacillus sp. strain AS_1 exhibited broad antibacterial activity against the pathogenic strains with MIC values ranging from 4 to 8 mg/ml, while Bacillus sp. strain AS_3 displayed MIC of 0.25 mg/ml. Crude extracts of Lysinibacillus sp. strain AS_1, Peribacillus sp. strain AS_2, and Bacillus sp. strain AS_3 showed growth inhibition of more than 90% against all the cancer cell lines at a concentration of 1,000 µg/ml. Untargeted secondary metabolite profiling of the crude extracts revealed the presence of compounds with reported biological activity, such as antimicrobial, antioxidant, anti-inflammatory, antitumor, and antidiabetic properties. This study reported for the first time, bacterial endophytes associated with A. sessiliflora with antibacterial and antitumor activities.

Data on metagenomic profiles of bacterial endophyte communities associated with Dicoma anomala.

Plants harbor varied communities of bacterial endophytes which play a crucial role in plant health and growth. Dicoma anomala is a medicinal plant that is known for its excellent ethnomedicinal uses which include treatment of coughs, fever, ulcers, and dysentery. This data in Brief article provides information on the diversity of bacterial endophytes associated with a medicinal plant, Dicoma anomala targeting the 16S rRNA gene using Illumina sequencing technology during three different seasons. Plant samples were collected in Eisleben, Limpopo province, South Africa, in the months of April, June, August and October 2018. The dataset revealed that the leaf samples collected in August had the highest species diversity as indicated by the Shannon index (4.25), Chao1 (1456.01), abundance-based coverage estimator (ACE) (1492.07) and the Simpson indices of diversity (0.05) irrespective of the species. The order of the bacterial endophyte's richness in D. anomala was April> October> June> August, from lowest to highest. The taxonomic composition analysis showed that most endophytic bacteria were composed of Proteobacteria, Actinobacteria, Firmicutes, Bacteroidetes and Chloroflexi. Some endophytic bacteria were found to be tissue specific. Sequences of Cutibacterium, Acinetobacter and Methylobacterium were prevalent in the leaves, whereas Amycolatopsis and Bradyrhizobium were the dominant genera in the root samples.

Antimicrobial Susceptibility Profiles among Pseudomonas aeruginosa Isolated from Professional SCUBA Divers with Otitis Externa, Swimming Pools and the Ocean at a Diving Operation in South Africa.

SCUBA divers are predisposed to otitis externa caused by Pseudomonas aeruginosa, which is becoming increasingly multi-drug resistant (MDR). The present work assessed the antibiotic resistance profiles of P. aeruginosa obtained from SCUBA divers and their environment in Sodwana Bay, South Africa. Bacterial isolates from a total of 137 random water and ear swab samples were identified using biochemical and molecular methods. P. aeruginosa strains were further evaluated for antibiotic susceptibility using the Kirby-Bauer assay. Double disk synergy test (DDST) to confirm metallo-ß-lactamase (MBL) production and PCR amplification of specific antibiotic resistance genes was performed. All (100%) 22 P. aeruginosa isolates recovered were resistant to 6 of the ß-lactams tested including imipenem but exhibited susceptibility to trimethoprim-sulfamethoxazole. MBL production was observed in 77% of isolates while the most prevalent extended-spectrum ß-lactamase (ESBL) genes present included blaAmpC (86.9%) followed by blaTEM (82.6%). Sulfonamide resistance was largely encoded by sul1 (63.6%) and sul2 (77.3%) genes with a high abundance of class 1 integrons (77.3%) of which 18.2% carried both Intl1 and Intl2. P. aeruginosa found in Sodwana Bay exhibits multi-drug resistance (MDRce) to several pharmaceutically important drugs with the potential to transfer antibiotic resistance to other bacteria if the judicious use of antibiotics for their treatment is not practiced.

Genome In Silico and In Vitro Analysis of the Probiotic Properties of a Bacterial Endophyte, Bacillus Paranthracis Strain MHSD3.

Spore-forming Bacillus species are gaining interest in human health recently, due to their ability to withstand the harsh environment of the gastrointestinal tract. The present study explores probiotic features of Bacillus paranthracis strain MHSD3 through genomic analysis and in vitro probiotic assays. The draft genome of strain MHSD3 contained genes associated with tolerance to gastrointestinal stress and adhesion. Cluster genes responsible for the synthesis of antimicrobial non-ribosomal peptide synthetases, bacteriocins, and linear azole-containing peptides were identified. Additionally, strain MHSD3 was able to survive in an acidic environment, had the tolerance to bile salt, and exhibited the capability to tolerate gastric juices. Moreover, the isolate was found to possess strong cell surface traits such as high auto-aggregation and hydrophobicity indices of 79 and 54%, respectively. Gas chromatography-mass spectrometry analysis showed that the strain produced secondary metabolites such as amino acids, phenolic compounds, and organic acid, known to exert health-promoting properties, including the improvement of gastrointestinal tract health.

Photorhabdus sp. ETL Antimicrobial Properties and Characterization of Its Secondary Metabolites by Gas Chromatography-Mass Spectrometry.

Entomopathogenic nematodes (EPNs) are known to be highly pathogenic to insect pests, due to their associated symbiotic bacteria, which produce virulence factors, exo-enzymes and other harmful secondary metabolites to conquer, kill, and degrade their insect hosts. However, these properties are not fully characterized. This study reports on the antimicrobial activities of Photorhabdus sp. strain ETL, symbiotically associated to an insect pathogenic nematode, Heterorhabditis zealandica, against human pathogenic bacteria and toxigenic fungi, as well as the non-targeted profiling of its secondary metabolites (SMs) using gas chromatography coupled to high-resolution time-of-flight mass spectrometry. Fatty acids including 3-eicosene, (E)-; 5-eicosene, (E)-; eicosene; 9-octadecenamide; undecanoic acid with shown antimicrobial activities were detected. This provided more insight on the composition and bioactivities of SMs produced by the Photorhabdus sp.

Genome assembly and annotation of Photorhabdus heterorhabditis strain ETL reveals genetic features involved in pathogenicity with its associated entomopathogenic nematode and anti-host effectors with biocontrol potential applications.

The genome sequences of entomopathogenic nematode (EPN) bacteria and their functional analyses can lead to the genetic engineering of the bacteria for use as biocontrol agents. The bacterial symbiont Photorhabdus heterorhabditis strain ETL isolated from an insect pathogenic nematode, Heterorhabditis zealandica strain ETL, collected in the northernmost region of South Africa was studied to reveal information that can be useful in the design of improvement strategies for both effective and liquid production method of EPN-based pesticides. The strain ETL genome was found closely related to the type strain genome of P. australis DSM 17,609 (~60 to 99.9% CDSs similarity), but closely related to the not yet genome-sequenced type strain, P. heterorhabditis. It has a genome size of 4,866,148 bp and G + C content of 42.4% similar to other Photorhabdus. It contains 4,351 protein coding genes (CDSs) of which, at least 84% are shared with the de facto type strain P. luminescens subsp. laumondii TTO1, and has 318 unknown CDSs and the genome has a higher degree of plasticity allowing it to adapt to different environmental conditions, and to be virulent against various insects; observed through genes acquired through horizontal gene transfer mechanisms, clustered regularly interspaced short palindromic repeats, non-determined polyketide- and non-ribosomal peptide- synthase gene clusters, and many genes associated with uncharacterized proteins; which also justify the strain ETL's genes differences (quantity and quality) compared to P. luminescens subsp. laumondii TTO1. The protein coding sequences contained genes with both bio-engineering and EPNs mass production importance, of which numerous are uncharacterized.

The Antibacterial Activity of Crude Extracts of Secondary Metabolites from Bacterial Endophytes Associated with Dicoma anomala.

Endophytic bacteria isolated from medicinal plants are recognized valuable sources of novel bioactive compounds with various activities such as antimicrobial, anticancer, and antiviral. In this study, eleven bacterial endophytes were isolated from surface sterilized roots and leave tissues, of medicinal plant Dicoma anomala. The bacterial endophytes were identified by sequencing the 16S rRNA gene, and belong to five genera viz Bacillus, Staphylococcus, Stenotrophomonas, Enterobacter, and Pantoea. The dominant genera were Bacillus with five strains, Staphylococcus with two strains, and Stenotrophomonas with two strains. The crude extracts of seven selected bacterial endophytes indicated antimicrobial activity against five pathogenic strains Escherichia coli (ATCC 25922), Bacillus cereus (ATCC 10876), Staphylococcus aureus (NCTC 6571), Pseudomonas aeruginosa (ATCC 27853), and Klebsiella oxytoca (ATCC 13182), with significant inhibition concentration ranging from 0.312 mg/ml to 0.625 mg/ml. Finally, based on the data analysis of the crude extracts of the endophytes, we identified bioactive secondary metabolites with reported biological activities such as antimicrobial, anti-inflammatory, and antioxidant properties with biotechnological applications in medicine, agriculture, and other industries. This study reported for the first time bacterial endophytes associated with D. anomala, with antimicrobial activity against bacterial pathogens.

Draft Genome Sequence of Pseudarthrobacter phenanthrenivorans Strain MHSD1, a Bacterial Endophyte Isolated From the Medicinal Plant Pellaea calomelanos.

Pseudarthrobacter phenanthrenivorans strain MHSD1 is a bacterial endophyte isolated from sterilized leaves of Pellaea calomelanos, a medicinal plant capable of growing in arid environments. Here, we report the draft genome sequence and annotation of this bacterial endophyte. The draft genome sequence of P. phenanthrenivorans strain MHSD1 has 4 450 468 bp with a G + C content of 65.30%. The National Center for Biotechnology Information Prokaryotic Genome Annotation Pipeline identified a total of 4004 protein-coding genes, 56 genes coding for RNAs, and 82 pseudogenes. Biosynthesis pathways for various phytohormones such as auxin, salicylic acid, ethylene, cytokinin, jasmonic acid, abscisic acid, and gibberellins were identified. Putative genes involved in various characteristics of bacterial endophyte lifestyle such as transport, motility, adhesion, membrane proteins, secretion and delivery systems, plant cell wall modification, and detoxification were identified. Phylogenomic analysis showed P. phenanthrenivorans strain MHSD1 to be a subspecies of P. phenanthrenivorans Sphe3.

Data on draft genome sequence of Bacillus sp. strain MHSD28, a bacterial endophyte isolated from Dicoma anomala.

Here, we present the draft genome sequence of Bacillus sp. strain MHSD28 which was sequenced, and assembled with a total length of 5,571,729 bp. The genome has 43 contigs, the largest contig with 1,785,042 bp, N50 of 1,474,247 bp, G + C% content of 35.23%. The strain was isolated from surface sterilized leaves of Dicoma anomala, obtained in Limpopo province, South Africa. The genome has 5792 total genes which include 5701 protein coding sequences (CDS), 192 pseudogenes, 7 rRNA genes with 3 operons (5S, 16S and 23S), 79 tRNA genes and 5 noncoding RNA (ncRNA) genes. This whole genome shotgun project has been deposited in DDBJ/ENA/GenBank under accession number VHIV00000000. The version described in this paper is version VHIV01000000.

Draft genome sequence of Pantoea ananatis strain MHSD5 isolated from surface sterilized leaves of medicinal plant, Pellaea calomelanos obtained in South Africa.

Pantoea ananatis strain MHSD5 is a bacterial endophyte isolated from the surface sterilized leaves of Pellaea calomelanos, which is a medicinal plant obtained in Limpopo province of South Africa. We present here the draft genome sequence and annotation of P. ananatis strain MHSD5. The genome assembly was 4.6 Mb in size with an N50 of 550,557 bp. A total of 4,350 putative protein coding sequence genes were predicted with PGAAP. This is the first draft genome of a bacterial endophyte symbiotically associated with P. calomelanos. This Whole Genome Shotgun project has been deposited at DDBJ/ENA/GenBank under the accession PUEK00000000. The version described in this paper is version PUEK01000000.