Bacillus Mycoides Research Articles

Antimicrobial Chlorinated Carbazole Alkaloids from the Sponge‐Associated Actinomycete Streptomyces diacarniLHW51701

Main observation and conclusionFour new chlorinated carbazole alkaloids, chlocarbazomycins (CCBs) A—D, were isolated from sponge associated bacterium Streptomyces diacarni LHW51701. Their structures were elucidated via spectroscopic techniques (including HRESIMS, 1D & 2D NMR), and X‐ray crystallographic analysis. Structurally, the 4‐chloro coupled with 3‐methoxyl substituents in the tricycle nucleus of CCBs A—D and the N‐methoxyl group of CCB‐D are rare in naturally occurred tricyclic carbazole alkaloids. Moreover, CCBs A—D do not bear the typical C1 para‐alkyl and C2 meta‐methyl side chains of bacterial tricycle carbazoles, suggesting a novel mechanism of aromatic ring formation in biosynthesis. The biological evaluation showed that CCB‐C possessed inhibitory activities against pathogenic microorganisms including methicillin‐resistant Staphylococcus aureus (MRSA), Mycobacterium smegmatis, Bacillus mycoides, and Candida albicans.

Bio-Organic Fertilizer: A Green Technology to Reduce Synthetic N and P Fertilizer for Rice Production.

Decomposed organic materials, in combination with plant growth-promoting bacteria (PGPB), are environmentally friendly and reduce synthetic fertilizer use in rice production. A bio-organic fertilizer (BoF) was prepared using kitchen waste (79%), chita-dhan (unfilled rice grain) biochar (15%), rock phosphate (5%), and a consortium of 10 PGPB (1%) to supplement 30% nitrogen and to replace triple superphosphate (TSP) fertilizer in rice production with an improvement of soil health. PGPB were local isolates and identified using 16S ribosomal RNA partial gene sequences as Bacillus mycoides, Proteus sp., Bacillus cereus, Bacillus subtilis, Bacillus pumilus, Paenibacillus polymyxa, and Paenibacillus spp. Isolates could fix N2 by 0.7–1.4 g kg–1, solubilize 0.1–1.2 g kg–1 phosphate, and produce 0.1–40 g kg–1 indoleacetic acid. The performance of BoF was evaluated by 16 field experiments and 18 farmers’ field demonstration trials during the year 2017–2020 in different parts of Bangladesh. Performances of BoF were evaluated based on control (T1), full synthetic fertilizer dose of N, P, and K (T2), BoF (2 t ha–1) + 70% N as urea + 100% K as muriate of potash (T3), 70% N as urea + 100% P as TSP + 100% K as muriate of potash (T4), and 2 t ha–1 BoF (T5) treatments. At the research station, average grain yield improved by 10–13% in T3 compared with T2 treatment. Depending on seasons, higher agronomic N use efficiency (19–30%), physiological N use efficiency (8–18%), partial factor productivity (PFP)N (114–150%), recovery efficiency (RE)N (3–31%), N harvest index (HIN) (14–24%), agronomic P use efficiency (22–25%), partial factor productivity of P (9–12%), AREP (15–23%), and HIP (3–6%) were obtained in T3 compared with T2 treatment. Research results were reflected in farmers’ field, and significant (P < 0.05) higher plant height, tiller, panicle, grain yield, partial factor productivity of N and P were obtained in the same treatment. Application of BoF improved soil organic carbon by 6–13%, along with an increased number of PGPB as compared with full synthetic fertilizer dose. In conclusion, tested BoF can be considered as a green technology to reduce 30% synthetic N and 100% TSP requirements in rice production with improved soil health.

The effect of crop species on DNase-producing bacteria in two soils

PurposeExtracellular deoxyribonucleases (exDNases) from microbial origin contribute substantially to the restriction of extracellular DNA (exDNA) in the soil. Hence, it is imperative to understand the diversity of bacterial species capable of performing this important soil function and how crop species influence their dynamics in the soil. The present study investigates the occurrence of DNase-producing bacteria (DPB) in leachate samples obtained from soils in which the crop species of alfalfa (Medicago sativa L.), canola (Brassica napus L.), soybean (Glycine max [L.] Merr.) and wheat (Triticum aestivum L.) were raised in a growth room.MethodsSelective media containing methyl green indicator was used to screen for DPB from leachate samples, whereas the 16S rRNA sequence analysis was employed to identify the isolates.ResultsThe proportion of culturable DPB ranged between 5.72 and 40.01%; however, we did observe specific crop effects that shifted throughout the growing period. In general, higher proportions of exDNase producers were observed when the soils had lower nutrient levels. On using the 16S rRNA to classify the DPB isolates, most isolates were found to be members of the Bacillus genera, while other groups included Chryseobacterium, Fictibacillus, Flavobacterium, Microbacterium, Nubsella, Pseudomonas, Psychrobacillus, Rheinheimera, Serratia and Stenotrophomonas. Five candidate exDNase/nuclease-encoding proteins were also identified from Bacillus mycoides genomes using online databases.ConclusionResults from this study showed that crop species, growth stage and soil properties were important factors shaping the populations of DPB in leachate samples; however, soil properties seemed to have a greater influence on the trends observed on these bacterial populations. It may be possible to target soil indigenous bacteria that produce exDNases through management to decrease potential unintended effects of transgenes originating from genetically modified organisms (GMOs) or other introduced nucleic acid sequences in the environment.

Isolation and Identification Bacillus Bacteria in Tilapia (Oreochromis niloticus) Using the Vitek-2 Compact

Tilapia, Oreochromis niloticus, is fish commodity that plays an important role in the tropic and subtropics country with their economic value for the community and tolerance for salinity. The high demand for tilapia resulted in the need to increase their production through the stocking density method. The purpose of this research is isolation and identification of pathogenic bacteria in tilapia fish using a Vitek-2 compact. The method used is sampling, perform bacterial isolation using TSB and TSA media using streak plate technique with OSE needle loop and cotton swap, identification of bacteria through gram staining, bacterial pathogenitas test, and using the Vitek-2 compact. The results of this study revealed three species of Bacillus thuringiensis, Bacillus mycoides and Bacillus cereus. All bacteria detected in the tilapia cultures were known as putative pathogens in Fish.

The effect of pH and incubation time on rude protease enzymes activity of Bacillus mycoides from anchovy isolates (Stolephorus sp.)

Proteases are several enzymes that are in important positions in the enzyme industry. eyebrow hydrolysis of peptide bonds on protein. Bacillus mycoides bacteria can grow in Nutrient Broth medium, these bacteria are included in facultative anaerobic bacteria. Bacillus mycoides are resistant to penicillin, positive to form Beta-hemolysis, positively catalyze hydrogen without oxidase, positively reduce nitrates and reduce methylene This study aims to determine the effect of pH and time on the protease enzyme activity of Bacillus mycoides from salted anchovy isolates. This research was conducted using a descriptive method with the treatment of pH concentration which has 5 levels, namely pH 5, 6, 7, 8 and 9 which are grouped based on the incubation time which has 4 levels, namely 6 hours, 12 hours, 18 hours and 24 hours and each was repeated 2 times. The results showed that pH and time affected the protease enzyme activity of Bacillus mycoides from salted anchovy isolate. Different pH treatments gave significantly different effects (F count> F5%) and the optimum activity was obtained at pH 5 of 5.243 U/ml. The different incubation time treatments had a significantly different effect (F count> F5%) and the optimum activity at 24 hours was 5.154 U/ml.

Mass spectrometry-based identification of bacteria isolated from industrially contaminated site in Salamanca (Mexico) and evaluation of their potential for DDT degradation.

Longstanding industrial deposits of 1-chloro-4-[2,2,2-trichloro-1-(4-chlorophenyl)ethyl]benzene (DDT) impose environmental threat in Salamanca city, located in central Mexico. Native bacteria from this location were isolated and identified, and their potential utility for DDT biodegradation was examined. Twenty-five isolates were obtained, and cell lysates were analyzed by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) with BiotyperTR; twenty-one organisms were identified at species level, and the other four were assigned to genus. The most abundant species corresponded to Bacillus (44%) and Pseudomonas genera (20%). Eight bacteria could grow in the presence of 200mg/L of DDT. Two-week exposure of Lysinibacillus fusiformis, Bacillus mycoides, Bacillus pumilus, and Bacillus cereusto DDT 50mg/L and 200mg/L, caused percentage pesticide degradation in the range 41-48% and 26-31%, respectively. Other four bacteria presented lower degradation rates. Gas chromatography-mass spectrometry (GC-MS) analysis of the spent media revealed that eight isolates assisted the conversion of DDT, DDD (1,1-dichloro-2,2-bis-(4-chlorophenyl)ethane), and DDE (1,1-dichloro-2,2-bis-(4-chlorophenyl)ethylene) to DDMU (1,1-(2-chloro-1,1-ethenediyl)-bis-(4-chlorobenzene)); however, DDNU (2,2-bis(4-chlorophenyl)ethylene), DBP (4,4'-dichlorobenzophenone(bis(4-chlorophenyl)methanone)) and DBH (bis(4-chlorophenyl)methanol) were found only for L. fusiformis, B. mycoides, B. cereus, B. marisflavi, and B. megaterium. Within the context of DDT biodegradation, the first three were the most promising isolates and further studies will be aimed at setting the experimental conditions for efficient mineralization of DDT congeners.

The influence of disease-inducing agents other than Dermatophilus congolensis on the severity of bovine dermatophilosis

This study was designed to assess the influence of disease-inducing agents other than Dermatophilus congolensis on the incidence of bovine dermatophilosis from cattle farms in Federal University of Agriculture, Abeokuta and Olabisi Onabanjo University, Ogun State and also from Akinyele cattle market in Oyo State. Skin scrapings collected from cattle with typical lesions of dermatophilosis were examined for the presence of bacteria, fungi and ectoparasites. Out of the 57 animals examined with typical skin lesions of dermatophilosis, Dermatophilus congolensis organism was cultured from 23 cases. Other bacterial organisms identified included Staphylococcus aureus (28 cases), Staphylococcus albus (35 cases), Bacillus mycoides (14 cases), Bacillus cereus (1 case), haemolytic Streptococcus (5 cases) Pseudomonas aeruginosa (13 cases), Escherichia coli (16 cases), Proteus spp (4 cases) and Norcardia spp (5 cases). Ticks (mostly Amblyomma variegatum) were seen in all the 57 animals while Demodex bovis, Sarcoptes scabiei var bovis and lice were seen in 15, 4 and 8 cases respectively. It was inferred that secondary bacterial invasions are associated with the pathogenesis and severity of the infection and the presence of ticks and mange mites may serve as the portal of entry for dermatophilosis infection. The control of dermatophilosis should take into account of arthropod vectors and secondary bacterial complications.

Biosynthesized silver nanoparticles by Aspergillus terreus NRRL265 for imparting durable antimicrobial finishing to polyester cotton blended fabrics: Statistical optimization, characterization, and antitumor activity evaluation

In the current study, the conditions that influence the biosynthesis of silver nanoparticles (AgNPs) by A. terreus NRRL265 cell free filtrate (CFF) were first considered through the “one factor per time” ordinary strategy. At that point a response surface methodology (RSM) was built to study and identify the interdependence between the efficient factors that influence the biosynthesis of AgNPs, utilizing the central composite design (CCD) procedure. Sphere shaped AgNPs in a range between 13 and 49 nm were detected via electron microscopy approaches. The spectroscopy of Fourier Transform Infrared (FTIR) suggested the contribution of peptides as a reducing and capping tool. The viability of AgNPs like an antimicrobial factor was set up by means of its capacity to apply a significant antimicrobial action in opposition to Candida albicans, Bacillus mycoides, and Escherichia coli. Human Caucasian breast adenocarcinoma (MCF7) tumor cells showed a direct dose–response relationship with AgNPs to give an IC50 value of 46.7 μg/mL. AgNP deposition on polyester cotton blended fabrics surface was detected as a thin reliable surface layer through checking scanning electron microscopy (SEM). In spite of the presence of little amounts of silver on the treated fabrics as demonstrated by EDX analysis profile, the fabrics treated with AgNPs were capable to restrain the tested microorganism’s growth.

Influence of cultivation conditions on IAA - producing activity of endophytic bacterial strains isolated from morning-glory (IPOMOEA PES-CAPRAE(L.) R.Br.)

This work presents the experimental study results of the influence of the culture medium on the ability to IAA synthesis of three endophytic strains TH10R, TH11T, and TH13T from roots of Ipomoea pes-caprae. Three investigated strains give the highest IAA concentration after 96 h of cultivation. A significant increase in IAA biosynthesis was obtained by cultivating the TH10R strain in a medium containing lactose or starch as a carbon source and NH4Cl or KNO3 as a nitrogen source. The TH11T strain produces the maximum amount of IAA, using glucose or xylose and KNO3 or NH4NO3 as carbon and nitrogen sources, respectively. Sucrose is a suitable carbon source for the TH13T strain; on a sucrose-containing medium, the TH13T strain produces the highest IAA amount. The most active strain is TH10R, identified as Bacillus mycoides and named Bacillus mycoides TH10R.

MogR Is a Ubiquitous Transcriptional Repressor Affecting Motility, Biofilm Formation and Virulence in Bacillus thuringiensis.

Flagellar motility is considered an important virulence factor in different pathogenic bacteria. In Listeria monocytogenes the transcriptional repressor MogR regulates motility in a temperature-dependent manner, directly repressing flagellar- and chemotaxis genes. The only other bacteria known to carry a mogR homolog are members of the Bacillus cereus group, which includes motile species such as B. cereus and Bacillus thuringiensis as well as the non-motile species Bacillus anthracis, Bacillus mycoides and Bacillus pseudomycoides. Furthermore, the main motility locus in B. cereus group bacteria, carrying the genes for flagellar synthesis, appears to be more closely related to L. monocytogenes than to Bacillus subtilis, which belongs to a separate phylogenetic group of Bacilli and does not carry a mogR ortholog. Here, we show that in B. thuringiensis, MogR overexpression results in non-motile cells devoid of flagella. Global gene expression profiling showed that 110 genes were differentially regulated by MogR overexpression, including flagellar motility genes, but also genes associated with virulence, stress response and biofilm lifestyle. Accordingly, phenotypic assays showed that MogR also affects cytotoxicity and biofilm formation in B. thuringiensis. Overexpression of a MogR variant mutated in two amino acids within the putative DNA binding domain restored phenotypes to those of an empty vector control. In accordance, introduction of these mutations resulted in complete loss in MogR binding to its candidate flagellar locus target site in vitro. In contrast to L. monocytogenes, MogR appears to be regulated in a growth-phase dependent and temperature-independent manner in B. thuringiensis 407. Interestingly, mogR was found to be conserved also in non-motile B. cereus group species such as B. mycoides and B. pseudomycoides, which both carry major gene deletions in the flagellar motility locus and where in B. pseudomycoides mogR is the only gene retained. Furthermore, mogR is expressed in non-motile B. anthracis. Altogether this provides indications of an expanded set of functions for MogR in B. cereus group species, beyond motility regulation. In conclusion, MogR constitutes a novel B. thuringiensis pleiotropic transcriptional regulator, acting as a repressor of motility genes, and affecting the expression of a variety of additional genes involved in biofilm formation and virulence.

Antimicrobial Activity of Soil Clostridium Enriched Conditioned Media Against Bacillus mycoides, Bacillus cereus, and Pseudomonas aeruginosa.

The rise of antimicrobial resistant bacteria has fast-tracked the exploration for novel antimicrobial compounds. Reports on antimicrobial producing soil anaerobes such as Clostridium spp. are very limited. In the present study, the antimicrobial activity of soil Clostridium enriched conditioned/spent media (CMs) against Bacillus mycoides, Bacillus cereus and Pseudomonas aeruginosa was assessed by turbidimetric growth inhibition assay. Our results highlighted the antimicrobial potential of soil Clostridium enriched conditioned media against pathogenic and spoilage bacteria. Farm 4 soil conditioned medium (F4SCM) demonstrated a greater growth inhibition activity against all three tested microorganisms in comparison to other soil conditioned media. Non-targeted metabolite profiling of all soil conditioned media revealed distinctive polar and intermediate-polar metabolites in F4SCM, consistent with its strong antimicrobial property. Moreover, 539 significantly abundant metabolites including some unique features were detected in F4SCM suggesting its substantial and specialized chemical diversity. This study putatively identified seven significantly high metabolites in F4SCM; 3-hydroxyphenylacetic acid, γ-aminobutyric acid, creatine, tryptamine, and 2-hydroxyisocaproic acid. Tryptamine and 2-hydroxyisocaproic acid were previously reported to have antimicrobial properties. The present study shows that soil Clostridium spp. are a promising group of bacteria producing metabolites with antimicrobial activity and provides future prospects for clostridial antimicrobial discovery within their metabolic diversity.

Phenylacetic Acid and Methylphenyl Acetate From the Biocontrol Bacterium Bacillus mycoides BM02 Suppress Spore Germination in Fusarium oxysporum f. sp. lycopersici

Bacillus mycoides strain BM02 originally isolated from the tomato rhizosphere was found to have beneficial functions in tomato by promoting plant growth and reducing the severity of Fusarium wilt caused by Fusarium oxysporum f. sp. lycopersici (Fol). Cytological experiments demonstrated B. mycoides BM02 reduced Fol invasion by reducing spore attachment and increasing hyphal deformation in hydroponics-grown tomato root tissues. Two volatile antifungal compounds, phenylacetic acid (PAA) and methylphenyl acetate (MPA), were identified from the culture filtrates of B. mycoides BM02 by GC-MS analysis. Chemically synthesized PAA, and to a lower extent MPA, suppressed spore germination but have no effects on the hyphal growth of Fol. Our results indicated that the biocontrol agent B. mycoides BM02 produced an array of bioactive compounds including PAA and MPA to suppress plant diseases caused by Fol and other pathogenic microorganisms.

Divergence in environmental adaptation between terrestrial clades of the Bacillus cereus group.

The Bacillus cereus group encompasses beneficial and harmful species in diverse niches and has a much debated taxonomy. Investigating whether selection has led to ecological divergence between phylogenetic clades can help understand the basis of speciation, and has implications for predicting biological safety across this group. Using three most terrestrial species in this group (B. cereus, Bacillus thuringiensis and Bacillus mycoides) we charactererized ecological specialization in terms of resource use, thermal adaptation and fitness in different environmental conditions and tested whether taxonomic species or phylogenetic clade best explained phenotypic variation. All isolates grew vigorously in protein rich media and insect cadavers, but exploitation of soil or plant derived nutrients was similarly weak for all. For B. thuringiensis and B. mycoides, clade and taxonomic species were important predictors of relative fitness in insect infections. Fully psychrotolerant isolates could outcompete B. thuringiensis in insects at low temperature, although psychrotolerance predicted growth in artificial media better than clade. In contrast to predictions, isolates in the Bacillus anthracis clade had sub-optimal growth at 37°C. The common ecological niche in these terrestrial B. cereus species is the ability to exploit protein rich resources such as cadavers. However, selection has led to different phylogenetic groups developing different strategies for accessing this resource. Thus, clades, as well as traditional taxonomic phenotypes, predict biologically important traits.

Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria.

BackgroundBacterial biofilms have become a major threat to human health. The objective of this study was to isolate amylase-producing bacteria from soil to determine the overall inhibition of certain pathogenic bacterial biofilms.MethodsWe used serial dilution and the streaking method to obtain a total of 75 positive amylase isolates. The starch-agar plate method was used to screen the amylolytic activities of these isolates, and we used morphological and biochemical methods to characterize the isolates. Optimal conditions for amylase production and purification using Sephadex G-200 and SDS-PAGE were monitored. We screened these isolates’ antagonistic activities and the purified amylase against pathogenic and multi-drug-resistant human bacteria using the agar disk diffusion method. Some standard antibiotics were controlled according to their degree of sensitivity. Finally, we used spectrophotometric methods to screen the antibiofilm 24 and 48 h after application of filtering and purifying enzymes in order to determine its efficacy at human pathogenic bacteria.ResultsThe isolated Bacillus species were Bacillus megaterium (26.7%), Bacillus subtilis (16%), Bacillus cereus (13.3%), Bacillus thuringiesis (10.7%), Bacillus lentus (10.7%), Bacillus mycoides (5.3%), Bacillus alvei (5.3%), Bacillus polymyxa (4%), Bacillus circulans (4%), and Micrococcus roseus (4%). Interestingly, all isolates showed a high antagonism to target pathogens. B. alevi had the highest recorded activity (48 mm) and B. polymyxa had the lowest recorded activity (12 mm) against Staphylococcus aureus (MRSA) and Escherichia coli, respectively. On the other hand, we detected no antibacterial activity for purified amylase. The supernatant of the isolated amylase-producing bacteria and its purified amylase showed significant inhibition for biofilm: 93.7% and 78.8%, respectively. This suggests that supernatant and purified amylase may be effective for clinical and environmental biofilm control.DiscussionOur results showed that soil bacterial isolates such as Bacillus sp. supernatant and its purified amylase are good antibiofilm tools that can inhibit multidrug-resistant former strains. They could be beneficial for pharmaceutical use. While purified amylase was effective as an antibiofilm, the isolated supernatant showed better results.

Antibiotic activity and resistance of lactic acid bacteria and other antagonistic bacteriocin-producing microorganisms

Introduction. Increased resistance of microorganisms to traditional antibiotics has created a practical need for isolating and synthesizing new antibiotics. We aimed to study the antibiotic activity and resistance of bacteriocins produced by lactic acid bacteria and other microorganisms.&#x0D; Study objects and methods. We studied the isolates of the following microorganism strains: Bacillus subtilis, Penicillium glabrum, Penicillium lagena, Pseudomonas koreenis, Penicillium ochrochloron, Leuconostoc lactis, Lactobacillus plantarum, Leuconostoc mesenteroides, Pediococcus acidilactici, Leuconostoc mesenteroides, Pediococcus pentosaceus, Lactobacillus casei, Lactobacillus fermentum, Bacteroides hypermegas, Bacteroides ruminicola, Pediococcus damnosus, Bacteroides paurosaccharolyticus, Halobacillus profundi, Geobacillus stearothermophilus, and Bacillus caldotenax. Pathogenic test strains included Escherichia coli, Salmonella enterica, Staphylococcus aureus, Pseudomonas aeruginosa, Bacillus mycoides, Alcaligenes faecalis, and Proteus vulgaris. The titer of microorganisms was determined by optical density measurements at 595 nm. &#x0D; Results and discussion. We found that eleven microorganisms out of twenty showed high antimicrobial activity against all test strains of pathogenic and opportunistic microorganisms. All the Bacteroides strains exhibited little antimicrobial activity against Gramnegative test strains, while Halobacillus profundi had an inhibitory effect on Gram-positive species only. The Penicillium strains also displayed a slight antimicrobial effect on pathogenic test strains.&#x0D; Conclusion. The antibiotic resistance of the studied lactic acid bacteria and other bacteriocin-producing microorganisms allows for their use in the production of pharmaceutical antibiotic drugs.

Valorization of Agro-Industrial By-products: Use of Rice Husk as a Source of Microorganisms to Denitrification of Water

Rice husk, which is an agricultural waste, provides a feasible alternative for the growth and propagation of denitrifying microorganisms. Nitrate and nitrite were removed using Immobilized Microorganisms (MOIM) or Microorganisms in Solution (MOSO). Microorganisms present in the rice husk biomass responsible for denitrification were identified as Pseudomonas, and other microorganisms have also been identified, as Oerskovia spp. Enterococcus sp. Bacillus mycoides and Escherichia coli. The influence of pH, temperature, C/N ratio and carbon source on biological denitrification were investigated. MOIM and MOSO consortium had optimal denitrifying performance at 25-30 °C and in pH 7-8. MOSO has average denitrification efficiency larger than MOIM. The MOIM denitrification efficiency was more sensitive to pH changes than the MOSO. Ethanol and sodium acetate were carbon sources for the denitrifying process. The efficiency of nitrate and nitrite removal using MOSO and ethanol or acetate with 1:1, 1:2, 1:3 and 1:4 C/N ratios were equivalents and above 97.00%. The denitrifying process presented was robust and it presented nitrate removal close to 100% during 10 cycles.

Use of radiofrequency electromagnetic radiation to remove biofilms from canvasses

A method based on the use of electromagnetic radiation (EMR) in the radiofrequency band was experimented to eliminate heterotrophic organisms from biofilms encrusting linen canvasses used for paintings. Artificial biofilms were used for exposure to EMR. Biofilms were grown on fragments of canvasses sampled from the backside of a painting. The fragments, after sterilization, were inoculated with several heterotrophic microorganisms previously isolated from other canvasses used for paintings and identified by molecular analyses as the bacteria Bacillus mycoides and Bacillus subtilis, and the microfungi Aspergillus versicolor, Cladosporium cladosporioides, Cladosporium oxysporum and Fusarium oxysporum. The newly formed biofilms resulted to contain the same microorganisms used for the inocula. They were exposed to EMR obtained by an amplitude-modulating radiofrequency sine wave with a train of rectangular pulses with 200 ns repetition time and 10% duty cycle. The exposure protocol consisted of three applications of two hours each, every other day. Denaturing gradient gel electrophoresis analyses carried out on the EMR-treated biofilms showed that all the microorganisms disappeared after the third application. The amplitude of the adopted EMR was not high enough to cause any significant increase in the temperature of the substrates. Therefore, the methodology showed to be non-invasive for the artefacts and its exposure levels not hazardous for operators and the environment compared to other techniques adopted in the field of conservation of cultural heritage. Among the latter techniques, the most common one is based on the use of biocides that, although effective, present problems in all the above-described areas.

Bio stimulation of indigenous microorganisms with Gomeya: a bioremediation technique

The burden of heavy metals in the environment can be reduced using organic amendment stimulated bacterial remediation. This study employed cattle manure slurry stimulated bacterial inoculum to treat heavy metals-contaminated soil. Samples of contaminated soil and cattle manure were collected from the area surrounding a steel rolling mill and a commercial animal pen respectively. Bacteria were isolated using pour plate technique; identified using various biochemical tests and screened for resistance to heavy metal salts by incorporating heavy metal salts into agar plates. The contaminated soil and manure slurry were analysed for heavy metals and then sterilised separately. Five kilogram of the sterilised contaminated soil was weighed and mixed with 100g of sterilized cow dung slurry and aseptically packed into plastic nursery bags. Bacterial samples showing high tolerance to heavy metal salts were introduced into the bags singly and as a consortium for bioremediation exercise. Thirty-six bacterial isolates were obtained from the contaminated soil. Chemical analysis revealed that the soil was heavily contaminated especially with lead and chromium with concentrations of 1505.1-2333.6 and 1526.0-1678.7 mg/kg, respectively. Alcaligenes faecalis, Pseudomonas azotoformans and Bacillus mycoides exhibited high tolerance to heavy metals salt and were selected for bioremediation. Post bioremediation analysis of the soil samples revealed a reduction in the concentration of heavy metals concentration with major reduction in the concentration of chromium in groups treated with P. azotoformans. Biostimulation of microorganisms with organic amendment effectively remediated heavy metals contaminated soil and can be employed in the treatment of such contaminated environments.

Bacillus mycoides A1 and Bacillus tequilensis A3 inhibit the growth of a member of the phytopathogen Colletotrichum gloeosporioides species complex in avocado.

Avocado is affected by Colletotrichum gloeosporioides causing anthracnose. Antagonistic microorganisms against C. gloeosporioides represent an alternative for biological control. Accordingly, in the present study, we focused on the isolation and characterization of potential antagonist bacteria against a member of the C. gloeosporioides species complex with respect to their possible future application. Samples of avocado rhizospheric soil were aquired from an orchard located in Ocuituco, Morelos, Mexico, aiming to obtain bacterial isolates with potential antifungal activity. From the soil samples, 136 bacteria were isolated and they were then challenged against a member of the C. gloeosporioides species complex; only three bacterial isolates A1, A2 and A3 significantly diminished mycelial fungal growth by 75%, 70% and 60%, respectively. Two of these isolates were identified by 16S rRNA as Bacillus mycoides (A1 and A2) and the third was identified as Bacillus tequilensis (A3). Bacillus mycoides bacterial cell-free supernatant reduced the mycelial growth of a member of the C. gloeosporioides species complex isolated from avocado by 65%, whereas Bacillus tequilensis A3 supernatant did so by 25% after 3 days post inoculation. Bacillus tequilensis mycoides A1 was a producer of proteases, indolacetic acid and siderophores. Preventive treatment using a cell-free supernatant of B. mycoides A1 diminished the severity of anthracnose disease (41.9%) on avocado fruit. These results reveal the possibility of using B. mycoides A1 as a potential biological control agent. © 2020 Society of Chemical Industry.

Diversity of nitrogen-fixing rhizobacteria associated with sugarcane: a comprehensive study of plant-microbe interactions for growth enhancement in Saccharum spp.

BackgroundNitrogen is an essential element for sugarcane growth and development and is generally applied in the form of urea often much more than at recommended rates, causing serious soil degradation, particularly soil acidification, as well as groundwater and air pollution. In spite of the importance of nitrogen for plant growth, fewer reports are available to understand the application and biological role of N2 fixing bacteria to improve N2 nutrition in the sugarcane plant.ResultsIn this study, a total of 350 different bacterial strains were isolated from rhizospheric soil samples of the sugarcane plants. Out of these, 22 isolates were selected based on plant growth promotion traits, biocontrol, and nitrogenase activity. The presence and activity of the nifH gene and the ability of nitrogen-fixation proved that all 22 selected strains have the ability to fix nitrogen. These strains were used to perform 16S rRNA and rpoB genes for their identification. The resulted amplicons were sequenced and phylogenetic analysis was constructed. Among the screened strains for nitrogen fixation, CY5 (Bacillus megaterium) and CA1 (Bacillus mycoides) were the most prominent. These two strains were examined for functional diversity using Biolog phenotyping, which confirmed the consumption of diverse carbon and nitrogen sources and tolerance to low pH and osmotic stress. The inoculated bacterial strains colonized the sugarcane rhizosphere successfully and were mostly located in root and leaf. The expression of the nifH gene in both sugarcane varieties (GT11 and GXB9) inoculated with CY5 and CA1 was confirmed. The gene expression studies showed enhanced expression of genes of various enzymes such as catalase, phenylalanine-ammonia-lyase, superoxide dismutase, chitinase and glucanase in bacterial-inoculated sugarcane plants.ConclusionThe results showed that a substantial number of Bacillus isolates have N-fixation and biocontrol property against two sugarcane pathogens Sporisorium scitamineum and Ceratocystis paradoxa. The increased activity of genes controlling free radical metabolism may at least in part accounts for the increased tolerance to pathogens. Nitrogen-fixation was confirmed in sugarcane inoculated with B. megaterium and B. mycoides strains using N-balance and 15N2 isotope dilution in different plant parts of sugarcane. This is the first report of Bacillus mycoides as a nitrogen-fixing rhizobacterium in sugarcane.