Megaterium Isolate Research Articles

Evaluation of Endospore-Forming Bacteria for Suppression of Postharvest Decay of Apple Fruit

The use of microbial biocontrol agents for control of postharvest disease has been the subject of intensive research over the past three decades resulting in commercialization of several biocontrol products. The objective of this research was to test endospore-forming bacteria collected from apple leaves for suppression of bitter rot and blue mold on apple. Bacteria were collected from abandoned, low-input, organic, and conventionally managed orchards in Pennsylvania and were screened for their ability to produce endospores, hydrolyze chitin, reduce pathogen growth in vitro, and suppress postharvest disease in vivo. Several isolates reduced bitter rot lesion size on 'Rome Beauty' from 40-89% compared to untreated controls. Bacillus megaterium isolates, A3-6 and Ae-1, resulted in the greatest suppression of bitter rot lesion size. One isolate, A3-2, suppressed blue mold lesion size. Scanning electron microscopy of inoculated apple wounds suggests parasitism as a mode of action explains the suppression of bitter rot lesion size by isolate A3-6. Of the top seventeen isolates exhibiting biocontrol potential, 70% were collected from abandoned or unmanaged locations. This research demonstrates abandoned apple orchards can be a source of new biocontrol agents for control of postharvest diseases of apple.

THE EFFECT OF DIFFERENT MICROBIAL FERTILIZER DOSES ON YIELD AND YIELD COMPONENTS IN CORIANDER (Coriandrum sativum L.)

Microbial fertilizers containing Bacillus subtilis and Bacillus megaterium isolates are one of the important organic fertilizers, which strengthen the immune system of plants against harmful effects and promote the plant growth by fixing nitrogen from the atmosphere, to obtain better quality products. This study was conducted to determine the effects of different liquid microbial fertilizer doses (0, 500, 1000, 1500, 2000, and 2500 ml 100 L–1) on the yield and yield components of the coriander (Coriandrum sativum L.) grown in Bayburt in 2018 and 2019. The experiment was set up as three replicates in randomized complete block design. The results indicated that there was a considerable variation in the yield and yield components of coriander, and the yield significantly increased as the dose of microbial fertilizer was increased. The maximum essential oil content and seed yield were recorded for the plots grown under the F1 and F3 treatment. It was found that the maximum seed yield was obtained in the F3 treatment (839.0 kg ha–1), and the seed yield decreased in the F4 (754.1 kg ha–1) and F5 (759.9 kg ha–1) treatments. Although the increase in the microbial fertilizer dose increases seed yield, using the fertilizer in optimum dose can be more profitable. Therefore, this study suggests that the fertilizer dose “F3” can improve the productivity of the coriander grown in the areas treated with microbial fertilizer.

Characterisation of Pb-resistant plant growth-promoting rhizobacteria (PGPR) from Scirpus grossus

Abstract Microorganisms that have the potential to improve plant productivity and health are known as plant growth-promoting rhizobacteria. Rhizobacteria play a major role in phytoremediation process by increasing the phytoremediation efficiency, thus the need to identify the superior rhizosphere bacteria has been gaining serious attention. In the current study, Scirpus grossus were exposed to a series of lead-contaminated sand (50, 100, 150, 200 and 300 mg/L) in plastic crates in greenhouse enviroment. Four weeks after exposure, the plants could sustain with toxicity of lead contamination by as much as 300 mg/L. Twenty two colonies of rhizobacteria were isolated from plant roots of Scirpus grossus. The rhizobacteria were characterised based on cell and colony observation. Identification of rhizobacteria was carried out based on analysis of 16S rDNA sequences and were identified as Aeromonas taiwanensis isolate 5E, Bacillus sp. Isolate 7G, Bacillus cereus isolate 8H and 3C , Bacillus velezensis isolate 9I, Bacillus proteolyticus isolate 4D, Bacillus stratosphericus isolate 14N, Bacillus megaterium isolate 11K , Pseudomonas sp. Isolate 12L , Enterobacter cloacae isolate 13M and isolate 16P, Bacillus aerius isolate 15O and Lysinibacillus sp. isolate 10J. The highly lead-resistant rhizobacteria were identified as Bacillus proteolyticus isolate 4D, Bacillus velezensis isolate 9I and Lysinibacillus sp. isolate 10J , which can tolerate against high concentration of lead (300 mg/L). The results showed that these highly lead-resistant rhizobacteria have potential to produce plant growth-promoting traits and have a ability to help S. grossus growth and productivity.

Morphological Changes Caused by Bacillus megaterium on Adult Emergence of Fall Webworm’s Pupa, Hyphantria cunea (Drury) (Lepidoptera: Erebidae)

Insecticidal effects of Bacillus megaterium isolates maintained from bacterial oozed pupae were studied on pupae and 2nd instar larvae of Hyphantria cunea. The bacterial isolates were identified according to morphological, physiological, biochemical and sequence analyses. This study reported that Bacillus megaterium isolates, HMA5 and BM1, caused different abnormal wings shapes and hair loss on thorax and abdomen of Hyphantria cunea. In addition, the HMA5 and BM1 treated pupae showed significantly reduced ability of adult emergence rates. The percentages of the adult emergence from pupae were 1% and 2% for HMA5 and BM1, respectively.

Colonization and biodegradation of the cross-linked potassium polyacrylate component of water absorbing geocomposite by soil microorganisms

The colonization and biodegradation of the superabsorbent polymer component of water absorbing geocomposite by microorganisms occurring in the soil were investigated using a pot experiment. The polymer used in this study was technical cross-linked potassium polyacrylate (tKPA). It was placed inside a plastic skeleton structure covered with a polyester fabric sheath and incubated in non-sterile Haplic Luvisol soil sown with grass for nine months. After this time, we noticed a 77.88% reduction of 4.25 g of the tKPA initial dry mass. The highest number of soil microbes that colonized and utilized tKPA as the sole carbon and energy source was found after the first month of incubation. It ranged from 4.02 log10 CFU g−1 of water absorbed by tKPA for fungi and protists to 6.99 log10 CFU for bacteria; during the next eight months, the number decreased to 3.05 and 5.63 log10 CFU, respectively.Bacillus megaterium isolate 37SBG and the Acremonium sclerotigenum – Acremonium egyptiacum complex isolate 25SFG shown to possess the ability to degrade tKPA were selected among the microorganisms that colonized the superabsorbent polymer. After 60 days of incubation of these microorganisms in mineral media supplemented with tKPA as the sole nutrient source, 280.50 mg of the initial polymer dry mass declined by 30.28% and 20.50% for 37SBG and 25SFG, respectively. In addition, after 30 days of growth in a medium mimicking the composition of grass root exudates that was additionally enriched with ultra pure cross-linked potassium polyacrylate (upKPA), 17.21% of the 267 mg initial dry mass of upKPA was degraded by B. megaterium 37SBG. Enzymatic activity similar to lignin peroxidase appears to be connected with this process.The tKPA geocomposite component was also effectively colonized by phytopathogens from the genus Phytophthora. Their numbers ranged from 0.70 to 2.26 log10 CFU g−1 of water absorbed by tKPA after 5 and 12 months, respectively, of the geocomposite incubation in the soil. In addition, some colonized tKPA microorganisms were able to limit the multiplication of the pathogen. The mycelial growth of Phytophthora cactorum and Rhizoctonia solani was inhibited in the presence of the A. sclerotigenum–A. egyptiacum complex isolate 25SFG.We concluded that the colonization of tKPA by both microbes with the ability to biodegrade it and phytopathogenic oomycetes could limit the soil application of geocomposite as a plant growth support. The biodegradation of tKPA by soil bacteria, fungi, and protists seems to also be important for the reduction of the amount of polymer residues deposited in the soil. In addition, this study expands our knowledge about antagonistic interactions between polymer degrading microbes and soil-borne pathogens.

Isolation and characterization of Bacillus megaterium isolates from dead pentatomids and their insecticidal activity to Palomena prasina nymphs

Bacillus megaterium isolates, were demonstrated to be efficient biocontrol agents against the green shield bug ( Palomena prasina L., Heteroptera : Pentatomidae ). Firstly hazelnut orchards were surveyed and four B. megaterium isolates were obtained from P. prasina . The morphological, physiological and biochemical characteristics of B. megaterium isolates were determined according to the standardized methodology. Additionally 16S rRNA gene sequence analyse was performed to determine the isolates. Analysis of the 16S rRNA gene confirmed that isolates Sa-1, Sa-5, SAk-2 and SAkc-19 are B. megaterium , with 100% sequence homology to the type strains of B. megaterium . Effectiveness of these isolates was tested against P. prasina nymphs in laboratory, at 25±1°C and 70±5 RH. Isolates were bioassayed against nymphs and dead individuals were counted daily, following a 12 days lasting treatment. Lethal time (LT 50 and LT 90 ) values of B. megaterium isolates were calculated. LT 50 values of the two most active isolates were 1.91 and 11.18 days. The highest mortality rate was 98% obtained with the treatment of SAkc-2 isolates at concentrations of 10 8 cfu ml -1 on the 12 th day of post treatment.

Bacillus megaterium strains derived from water and soil exhibit differential responses to the herbicide mesotrione.

The intense use of herbicides for weed control in agriculture causes selection pressure on soil microbiota and water ecosystems, possibly resulting in changes to microbial processes, such as biogeochemical cycles. These xenobiotics may increase the production of reactive oxygen species and consequently affect the survival of microorganisms, which need to develop strategies to adapt to these conditions and maintain their ecological functionality. This study analyzed the adaptive responses of bacterial isolates belonging to the same species, originating from two different environments (water and soil), and subjected to selection pressure by herbicides. The effects of herbicide Callisto and its active ingredient, mesotrione, induced different adaptation strategies on the cellular, enzymatic, and structural systems of two Bacillus megaterium isolates obtained from these environments. The lipid saturation patterns observed may have affected membrane permeability in response to this herbicide. Moreover, this may have led to different levels of responses involving superoxide dismutase and catalase activities, and enzyme polymorphisms. Due to these response systems, the strain isolated from water exhibited higher growth rates than did the soil strain, in evaluations made in oligotrophic culture media, which would be more like that found in semi-pristine aquatic environments. The influence of the intracellular oxidizing environments, which changed the mode of degradation of mesotrione in our experimental model and produced different metabolites, can also be observed in soil and water at sites related to agriculture. Since the different metabolites may present different levels of toxicity, we suggest that this fact should be considered in studies on the fate of agrochemicals in different environments.

Efficacy of native antagonistic bacterial isolates in biological control of crown gall disease in Egypt

In vitro analyzing the antagonistic activity of seventy native bacterial isolates towards plant tumorigenic Agrobacterium tumefaciens resulted in a selection of eight potential biocontrol agents. These isolates were screened for their antagonistic effect in vitro as well as their efficacy in reducing gall formation in planta. They were identified using Biolog microplates system as Bacillus megaterium, Paenibacillus polymyxa, Pseudomonas fragi (two isolates), Pseudomonas viridilivd, Pseudomonas asplenii, Curtobacterium flaccumfaciens and Curtobacterum sp.All antagonists tested exhibited considerable inhibitory activity in vitro and significantly reduced incidence and size of galls in rose shoots, kalanchoe leaves and squash fruits with variable degrees on the tested hosts. C. flaccumfaciens reduced the incidence of crown gall up to 100% in the case of rose shoots and kalanchoe leaves whereas the same antagonist reduced galling of squash fruits to 75%. Likewise, P. asplenii, P. viridilivd and P. polymyxa reduced the incidence of crown gall up to 100% in the case of kalanchoe leaves and squash fruits, whereas they reduced galling of rose shoots to 66.7%, 55.6% and 44.5% respectively. In the same manner, the two isolates of P. fragi reduced galling up to 100% in squash fruits, while it was 88.9% in rose shoots and kalanchoe leaves. Interestingly, B. megaterium isolate completely suppressed the gall development in rose shoots, whereas the gall incidence was 100% in kalanchoe leaves and 25% in squash fruits. Bacterial isolates characterized in this study may be considered as potential sources of novel bioactive metabolites as well as promising candidates to develop new biocontrol agents for controlling crown gall disease.

English

  The causal agents of bacterial blight in lupine (Lupinis termis) were isolated from leaves displaying symptoms in Minia governorate, Egypt. The pathogens were characterized by biochemical and physiological tests, and identified as Bacillus megaterium. Tissue extracts prepared from experimentally diseased shoot systems showed great pectolytic and cellulolytic activities while healthy tissue extracts of both organs showed slight activities of the enzymes. In varietal response test, four lupine cultivars that is, Australian, Balady, Giza 1 and Giza 2 were tested for their susceptibility to all tested B. megaterium isolates and the cultivar Balady was most sensitive to all isolates. However, isolates B2 and B6 were more virulent than the others. Two isolates of B. megaterium isolates that caused blight on lupine plants were subsequently tested on other foliar plants representatives of 17 plant species. Both B. megaterium isolates could not infect any of the tested plants but produced small necrotic spots on faba bean leaves. These results indicate the host specificity of this bacterium towards its host plant and accordingly justify the suggested forma specials lupini to be given to the lupine bacterial pathogen B. megaterium. According to the literature review, this is the first report on the occurrence of B. megaterium as a causal agent of leaf blight of lupine plants in Egypt.   Key words: Bacillus megaterium, lupine, pectolytic and cellulolytic activities, cultivars.

Assessment of Application Timing of Bacillus spp. to Suppress Pre- and Postharvest Diseases of Apple.

Four isolates of Bacillus spp. were tested in a 2-year field study for biological control of pre- and postharvest diseases of apple. For the preharvest test, bacteria were applied to 'Golden Delicious' and 'Rome Beauty' trees in May or May and June. Foliar apple scab severity was assessed weekly. After harvest, fruit were wounded and then either left untreated or given a postharvest application of the bacteria. Wounded fruit were then inoculated with the bitter rot pathogen and lesion size was measured over 8 days. Bacillus megaterium isolate A3-6, B. mycoides isolate A1-1, and B. cereus isolate FLS-5 applied in May or May and June significantly reduced fruit and foliar apple scab severity in both years. A postharvest application of the bacteria (alone or in combination with a preharvest application) resulted in the greatest suppression of bitter rot on both cultivars (P < 0.04). The May + June + postharvest application of isolate A3-6 resulted in the greatest suppression of bitter rot, with an average of 45 and 95% reduction in lesion size compared with nontreated controls on 'Golden Delicious' and 'Rome Beauty' fruit, respectively. Results from this study indicate that preharvest applications of the bacteria were able to reduce foliar and fruit scab and an additional postharvest application was effective in reducing bitter rot severity.

Histamine contents of salted seafood products in Taiwan and isolation of halotolerant histamine-forming bacteria

Fifty-seven salted seafood products sold in the fishing village stores in Taiwan, including salted fish product, salted mollusc product and salted shrimp product, were tested to determine the occurrence of histamine and histamine-forming bacteria. Although the average content of each of nine different biogenic amines in all samples was less than 5.0mg/100g, 10.5% (6/57) of tested samples had the histamine content greater than the 5.0mg/100g allowable limit suggested by the US Food and Drug Administration. One histamine-producing bacterial strain, capable of producing 78.5ppm of histamine in trypticase soy broth supplemented with 1.0% l-histidine (TSBH), was identified as Bacillus megaterium. The B. megaterium isolate was a halotolerant bacterium which grew well to an elevated NaCl concentration of 15% in TSBH medium. Besides, it had a consistent ability to produce >300ppm of histamine at 10% NaCl concentration in TSBH medium after 72h.

Integrated Management of Damping-off, Root and/or Stem Rot Diseases of Chickpea and Efficacy of the Suggested Formula

Eleven fungal isolates were isolated from naturally infected chickpea roots collected from different locations in New Valley Governorate (Egypt). The isolated fungi were purified and identified as Rhizoctonia solani (5 isolates), Fusarium solani (4 isolates) and Sclerotinia sclerotiorum (2 isolates). The isolated fungi proved their pathogenicity on cv. ‘Giza 3’. Response of chickpea cvs. ‘Giza 1’, ‘Giza 2’, ‘Giza 3’, ‘Giza 4’, ‘Giza 88’, ‘Giza 195’, ‘Giza 531’ to infection by the tested fungi was significantly varied. ‘Giza 1’ was the most resistant one followed by ‘Giza 531’, while the other tested cvs. were highly susceptible. Seven biocontrol agents, namely Bacillus subtilis, B. megaterium, B. cereus, Trichoderma viride, T. harzianum, Aspergillus sp., Penicillium sp. isolated from chickpea rhizosphere, were tested for their antagonistic action against the tested pathogens. B. subtilis isolate BSM1, B. megaterium isolate TVM5, T. viride isolate TVM2 and T. harzianum isolate THM4 were the most antagonistic ones to the tested fungi in vitro, while the other isolates were moderate or weak antagonists. The most antagonistic isolates as well as the commercial biocide Rhizo-N were applied as seed treatment for controlling damping-off, root and/or stem rot diseases caused by the tested fungi under greenhouse conditions. The obtained data showed that all tested antagonistic isolates were able to cause significant reduction of damping-off, root and/or stem rot diseases in chickpea plants. T. viride (isolate TVM2) and B. megaterium (isolate BMM5) proved to be the most effective isolates for controlling the diseases. Under field condition, the obtained data indicated that all the tested antagonistic isolates significantly reduced damping-off, root and/or stem rot. T. viride (isolate TVM2) and B. megaterium (isolate BMM5) recorded the highest reduction of damping-off, root and/or stem rot in all sowing dates. Sowing of treated seeds with bioagents in first of November gave the highest protection against root diseases in chickpea. The reduction in damping-off, root and/or stem rot severity was significantly reflected on the produced seed yield. In this respect, seeds previously treated with T. viride (TVM2) produced the highest seed yield in all sowing dates followed by seed treated with B. megaterium (TVM5). First of November was the best sowing date to reduce these diseases and to increase seed yield/fed. On the other hand, the antagonistic isolates isolated from chickpea rhizosphere, were most active than the commercial biocide Rhizo-N in reducing chickpea root diseases and increase of seed yield in greenhouse and field conditions.

ISOLATION OF BACILLUS MEGATERIUM FROM APHIS POMI (HOMOPTERA: APHIDIDAE) AND ASSESSMENT OF ITS PATHOGENICITY

SUMMARY Bacillus megaterium isolates were recovered from the green apple aphid, Aphis pomi De Geer, collected from apple trees in the Samsun province of Turkey. The lethality of nine B. megaterium isolates was assayed on A. pomi at 28-30°C. The isolates caused between 92% and 100% mortality within one week, with no significant differences amongst them. In contrast, there was a significant difference between all isolates and the control at P<0.05. The highest death rate (100%) was induced by the bacterial isolate No. 23 (Bmin23). After bacterial application on A. pomi, there was a slowing down of movement, cessation of feeding and development of a brown-black discoloration in diseased aphids within three days. Death followed within five days.

Diversity of Bacillus megaterium isolates cultured from honeys

Bacillus megaterium isolates recovered from Argentinean honeys ( n = 48) were compared with isolates recovered from honey from other countries ( n = 4) and with strain NRRL B-939. All the isolates and the NRRL strain were characterized using rep-PCR fingerprinting with primers BOX-, REP- and ERIC-; restriction fragment length polymorphism analysis of a 16S rRNA gene fragment (16S rRNA PCR/RFLP), and morphological and biochemical tests. There was a high degree of diversity, both phenotypic and genotypic among the isolates of B. megaterium. Our results not only show considerable levels of diversity among isolates of B. megaterium analyzed here but also that repetitive sequences such as BOX, ERIC and REP are reliable tools to characterize isolates at the genomic level. Haemolytic activity was shown by 77% of the isolates, as β-haemolysis (40 isolates) or α-haemolysis (one isolate). Within the β-haemolytic group, 10% of isolates produced a discontinuous haemolytic pattern, which is usually correlated with enterotoxin activity in other bacterial species. In addition, coagulase activity was detected in 74% of 53 isolates tested suggesting a pathogenic activity not ordinarily reported as a property of B. megaterium. According to our knowledge, this is the first report of phenotypic and genotypic characterization of B. megaterium isolates from honey.

BACTERIA FOUND ON AMERICAN CHESTNUT BARK AND THEIR POTENTIAL IN BIOCONTROL OF CHESTNUT BLIGHT

Summary The American chestnut (Castanea dentata [Marsh.] Borkh.) became susceptible to a blight fungus, Cryphonectria parasitica [Murr.] Bar, in the 1870s. This was imported on seedings from Asia, and American chestnut had no resistance to it. Since then, all attempts to eradicate or control the disease have met with very limited success or failure. This paper briefly reviews the history of the biocontrol of chestnut blight, from the use of ‘hypovirulent’ strains of C. parasitica to the use of a bacterium of Castanea spp. that are (1) antagonistic to C. parasitica and (2) can survive for an extensive time period on the bark of C. dentata. The study reveals that some Bacillus megaterium isolates from C. dentata bark may have potential in biological control. The results also provide an explanation for the success of mud packs in ‘healing’ cankers.