Browsing by Autor "Audisio, M C"

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Antagonistic effects of Bacillus subtilis subsp. subtilis and B. amyloliquefaciens against Macrophomina phaseolina: SEM study of fungal changes and UV-MALDI-TOF MS analysis of their bioactive compounds.
(2016) Torres, M J; Brandan, C Pérez; Petroselli, G; Erra-Balsells, R; Audisio, M C
The antifungal effect of Bacillus subtilis subsp. subtilis PGPMori7 and Bacillus amyloliquefaciens PGPBacCA1 was evaluated against Macrophomina phaseolina (Tassi) Goid. Cell suspension (CS), cell-free supernatant (CFS) and the lipopeptide fraction (LF) of PGPMori7 and PGPBacCA1 were screened against three different M. phaseolina strains. CS exhibited the highest inhibitory effect (around 50%) when compared to those of CFS and LF, regardless of the fungal strain studied. The synthesis of lipopeptides was studied by UV-MALDI TOF. Chemical analysis of Bacillus metabolite synthesis revealed that surfactin and iturin were mainly produced in liquid medium. Potential fengycin was also co-produced when both Bacillus were cultivated in solid medium. In co-culture assays, the bacterial colony-fungal mycelium interface at the inhibition zone was evaluated by both scanning electron microscopy (SEM) and UV-MALDI TOF, the former to determine the structural changes on M. phaseolina cells and the latter to identify the main bioactive molecules involved in the inhibitory effect. PGPBacCA1 produced surfactin, iturin and fengycin in the inhibition zone while PGPMori7 only produced these metabolites within its colony and not in the narrow inhibition zone. Interestingly, SEM revealed that PGPBacCA1 induced damage in M. phaseolina sclerotia, generating a fungicidal effect as no growth was observed when normal growth conditions were reestablished. In turn, PGPMori7 inhibited the growth of the Macrophomina mycelium without fungal injury, resulting only in a fungistatic activity. From these results, it was determined that the two bacilli significantly inhibited the growth of an important phytopathogenic fungus by at least two different mechanisms: lipopeptide synthesis and competition among microorganisms.
Bacillus subtilis subsp. subtilis CBMDC3f with antimicrobial activity against Gram-positive foodborne pathogenic bacteria: UV-MALDI-TOF MS analysis of its bioactive compounds.
(2015) Torres, M J; Petroselli, G; Daz, M; Erra-Balsells, R; Audisio, M C
In this work a new Bacillus sp. strain, isolated from honey, was characterized phylogenetically. Its antibacterial activity against three relevant foodborne pathogenic bacteria was studied; the main bioactive metabolites were analyzed using ultraviolet matrix assisted laser desorption-ionization mass spectrometry (UV-MALDI MS). Bacillus CBMDC3f was phylogenetically characterized as Bacillus subtilis subsp. subtilis after rRNA analysis of the 16S subunit and the gyrA gene (access codes Genbank JX120508 and JX120516, respectively). Its antibacterial potential was evaluated against Listeria monocytogenes (9 strains), B. cereus (3 strains) and Staphylococcus aureus ATCC29213. Its cell suspension and cell-free supernatant (CFS) exerted significant anti-Listeria and anti-S. aureus activities, while the lipopeptides fraction (LF) also showed anti-B. cereus effect. The UV-MALDI-MS analysis revealed surfactin, iturin and fengycin in the CFS, whereas surfactin predominated in the LF. The CFS from CBMDC3f contained surfactin, iturin and fengycin with four, two and four homologues per family, respectively, whereas four surfactin, one iturin and one fengycin homologues were identified in the LF. For some surfactin homologues, their UV-MALDI-TOF/TOF (MS/MS; Laser Induced Decomposition method, LID) spectra were also obtained. Mass spectrometry analysis contributed with relevant information about the type of lipopeptides that Bacillus strains can synthesize. From our results, surfactin would be the main metabolite responsible for the antibacterial effect.
Beneficial Effects of Bacillus subtilis subsp. subtilis Mori2, a Honey-Associated Strain, on Honeybee Colony Performance.
(2012) Sabaté, D C; Cruz, M S; Benítez-Ahrendts, M R; Audisio, M C
A Bacillus spp. strain isolated from a honey sample in Morillos (Salta, Argentina) was phylogenetically characterized as B. subtilis subsp. subtilis Mori2. The strain was administered to bee colonies as a monoculture in one litre of sugarcane syrup (125 g/L) at a final concentration of 10(5) spores/mL to evaluate the bee colony performance. The treated colony was monitored, and any changes were compared with the control hives. All conditions were identical (weather, nourishment and supervision), except for the Bacillus spore supplement. The new nourishment, which was administered monthly from May to December 2010, was accepted by the bees and consumed within ca. 24-48 h. Photograph records and statistic analyses revealed significant differences in the open and operculated brood areas between the treated and control groups. The status of the colony improved after the second administration of the Bacillus spores until the end of the experiment. A higher number of bees were counted in the treated groups (26% more than the control) with respect to the initial number. Furthermore, at the time of harvest, honey storage in the treated hives was 17% higher than in the control hives. In addition, spore counts of both Nosema sp. and Varroa sp. foretica in treated hives were lower than in the control hives. These results with experimental hives would indicate that B. subtilis subsp. subtilis Mori2 favoured the performance of bees; firstly, because the micro-organism stimulated the queen's egg laying, translating into a higher number of bees and consequently more honey. Secondly, because it reduced the prevalence of two important bee diseases worldwide: nosemosis and varroosis.
Characterisation of the cell envelope and adhesive properties of Lactobacillus johnsonii CRL1647, a probiotic from the honeybee gut.
(2025) Novicov-Fanciotti, M; Dentice Maidana, S; Elean, M; Villena, J; Audisio, M C
Lactobacillus johnsonii CRL1647 has been studied due to its beneficial effects on Apis mellifera L. bee colonies. In this work, we analyzed the characteristics of its cell envelope and the relationship of this bacterial structure with adhesion. The study revealed that CRL1647 cells did not harbor S-layer proteins, whereas L. acidophilus ATCC4356, used as a positive control, showed a typical S-layer protein band. L. johnsonii CRL1647 hemagglutinated with sheep erythrocytes. Interestingly, the hemagglutination abilities of L. johnsonii CRL1647 were affected by the treatments with proteinase K and sodium metaperiodate. Transmission electron microscopy (TEM) analysis confirmed that L. acidophilus ATCC4356 has S-layer and revealed that the L. johnsonii CRL1647 cell surface was full of prolongations like 'hairs'. These ultra-structures completely disappeared after the treatment with proteinase K. The CRL1647 strain was able to efficiently adhere to intestinal epithelial cells and was phagocyted by macrophages. Both, adhesion and phagocytosis were significantly diminished when CRL1647 cells were pretreated with proteinase K. From these results, it can be inferred that the principal molecules involved in the adherence of L. johnsonii CRL1647 have a glycoprotein structure, differing them from S-layer proteins.
Effect of Lactobacillus johnsonii CRL1647 on different parameters of honeybee colonies and bacterial populations of the bee gut.
(2015) Audisio, M C; Sabaté, D C; Benítez-Ahrendts, M R
Lactobacillus johnsonii CRL1647, isolated from the intestinal tract of a worker-bee in Salta, Argentina, was delivered to Apis mellifera L. honey bee colonies according to two different administration schedules: 1×10(5) cfu/ml every 15 days (2011) or monthly (2012). The effect of each treatment on the bee-colony performance was monitored by measuring honey production, and the prevalence of varroasis and nosemosis. Worker bees from each assay were randomly captured 3 days after administration and assayed for the following intestinal culturable and defined bacterial populations: total aerobic microorganisms, Bacillus spp. spores, Lactobacillus spp., Enterococcus spp. and enterobacteria. Interestingly, both treatments generated a similar increase in honey production in treated colonies compared to controls: 36.8% (every 15 days) and 36.3% (monthly). Nosema index always exhibited a reduction when lactobacilli were administered; in turn, Varroa incidence was lower when the lactobacilli were administered once a month. Moreover, the administration of L. johnsonii CRL1647 every 15 days produced an increase in the total number of aerobic microorganisms and in bacteria belonging to the genera Lactobacillus and Enterococcus; at the same time, a decrease was observed in the number of total spores at the end of the treatment. The number of enterobacteria was constant and remained below that of control hives at the end of the assay. On the other hand, the delivery of lactobacilli once a month only showed an increase in the number of bacteria belonging to the genus Lactobacillus; meanwhile, viable counts of the remaining microorganisms assayed were reduced. Even though it seems that both treatments were similar, those bee colonies that received L. johnsonii CRL1647 every 15 days became so strong that they swarmed.
Honey yield of different commercial apiaries treated with Lactobacillus salivarius A3iob, a new bee-probiotic strain.
(2018) Fanciotti, M Novicov; Tejerina, M; Benítez-Ahrendts, M R; Audisio, M C
The main objective of this study was to determine the impact of Lactobacillus salivarius A3iob, a honey bee gut-associated strain (GenBank code access KX198010), on honey yield. Independent assays were conducted from May to September 2014 and 2015, in three commercial apiaries: Tilquiza, El Carmen and Yala, all located in north-western Argentina. Local Apis mellifera L. bees were kept in standard Langstroth hives; treated hives were fed once a month with 1×105 cfu/ml viable Lactobacillus cells, administered to the bees through a Doolittle-type feeder in 125 g/l sucrose syrup. Control hives were only given the syrup mixed with MRS sterile broth. The main honey harvest was done in December in all groups and we found that there was an overall increase in honey yield from the treated hives. In 2014, all treated hives produced between 2.3 to 6.5 times more honey than the controls. However, in 2015, higher honey average yields in the treated hives at El Carmen and Yala were obtained, yet not at Tilquiza, because of a slight mishap. They experienced the swarming of several bee colonies due to a higher number of bees without appropriate management, which caused the control group to yield more honey compared to the hives fed with Lactobacillus. Interestingly, at El Carmen, two honey harvests were recorded: one in winter and another in summer (July and December 2015, respectively). This unexpected result arose from the particular flora of the region, mainly Tithonia tubaeformis, which blooms in winter. L. salivarius A3iob cells prove to be a natural alternative that will positively impact the beekeepers' economy by providing a higher honey yield.