Identification, technological and safety characterization of Lactobacillus sakei and Lactobacillus curvatus isolated from Argentinean anchovies (Engraulis anchoita)

In this study, the identification and characterization of Lactobacillus previously isolated from fresh anchovies (Engraulis anchoita) are investigated. 16S rDNA partial sequencing assigned all the isolates to belong to the Lactobacillus sakei/curvatus group. Fourteen out of 15 isolates were identified as L. sakei by phenotypic traits: they exhibited catalase activity and fermented melibiose, although only 10 of them hydrolyzed arginine. These results were confirmed by multiplex PCR-based restriction enzyme analysis with HindIII and by restriction fragment length polymorphic (RFLP) analysis of the 16S-23S rDNA intergenic spacer region with TaqI. Among identified isolates, four L. sakei strains and the sole L. curvatus strain showing sensitivity to chloramphenicol, erythromycin and tetracycline and exhibiting high tolerance to NaCl (10-18%) were unable to produce neither dextran nor biogenic amines. Based on technological and safety features, L. sakei SACB704 and L. curvatus SACB03a naturally present in fresh anchovies may be promising strains for the development of a starter culture to accelerate and control the fermentation of salt fermented anchovy-based products.Fil: Belfiore, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Raya, Raul Ricardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Vignolo, Graciela Margarita. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentin

Isolation and characterization of plant growth promoting bacteria isolated from andean soil as potential inoculants of soybean seeds

Argentina is the leading exporter of soybean oil and flour, and the third largest producer of grain. Since, the crop isa matter of great importance to the national economy. Their production depends on the soil as their main resourceto ensure a good productive capacity, so it is necessary to preserve the physical, chemical and biological propertiesof the soil. Although, the indiscriminate use of chemical fertilizers, disturb them. In recent years, there has been atrend towards cleaner production to reduce the use of chemical. One of the alternatives involves biological meansthrough the use of plant growth promoting bacteria. These group of bacteria colonize the rhizosphere of plants andstimulate the plant growth by several mechanisms.The objective of this work was to characterize, identify and evaluate the growth promoting effect of 13 strainsisolated from the Andean vegetation rhizosphere. The bacterial isolates were Enterobacteria, Stenotrophomonas,Pseudomonas, Nocardiodes, Bacillus, Exiguobacterium, Acinetobacter and Lactococcus genera. The results of thebiochemical characterization determined that from the 13 bacterial strains, which produce siderophores, 11possess the catalase enzyme, 10 fixate nitrogen, 12 produce the protease enzyme, 12 solubilize phosphorus, and 11produce indoleacetic acid.The application of different inoculums to the seeds, allowed to obtain plants with longer stem length, moredeveloped roots, larger and more intense coloration leaves than the control plants. The results encourage deeperstudies to achieve the formulation of inoculums to use as a biofertilizer, which would replace chemical fertilizersor reduce their doses.Fil: Belfiore, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Santos, Ana Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Farias, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; Argentin

High-Up: A Remote Reservoir of Microbial Extremophiles in Central Andean Wetlands

The Central Andes region displays unexplored ecosystems of shallow lakes and salt flats at mean altitudes of 3700 m. Being isolated and hostile, these so-called “High-Altitude Andean Lakes” (HAAL) are pristine and have been exposed to little human influence. HAAL proved to be a rich source of microbes showing interesting adaptations to life in extreme settings (poly-extremophiles) such as alkalinity, high concentrations of arsenic and dissolved salts, intense dryness, large daily ambient thermal amplitude, and extreme solar radiation levels. This work reviews HAAL microbiodiversity, taking into account different microbial niches, such as plankton, benthos, microbial mats and microbialites. The modern stromatolites and other microbialites discovered recently at HAAL are highlighted, as they provide unique modern—though quite imperfect—analogs of environments proxy for an earlier time in Earth's history (volcanic setting and profuse hydrothermal activity, low atmospheric O2 pressure, thin ozone layer and high UV exposure). Likewise, we stress the importance of HAAL microbes as model poly-extremophiles in the study of the molecular mechanisms underlying their resistance ability against UV and toxic or deleterious chemicals using genome mining and functional genomics. In future research directions, it will be necessary to exploit the full potential of HAAL poly-extremophiles in terms of their biotechnological applications. Current projects heading this way have yielded detailed molecular information and functional proof on novel extremoenzymes: i.e., DNA repair enzymes and arsenic efflux pumps for which medical and bioremediation applications, respectively, are envisaged. But still, much effort is required to unravel novel functions for this and other molecules that dwell in a unique biological treasure despite its being hidden high up, in the remote Andes.Fil: Albarracín, Virginia Helena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Planta Piloto de Procesos Industriales Microbiologicos; ArgentinaFil: Kurth, Daniel German. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Planta Piloto de Procesos Industriales Microbiologicos; ArgentinaFil: Ordoñez, Omar Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Planta Piloto de Procesos Industriales Microbiologicos; ArgentinaFil: Belfiore, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Planta Piloto de Procesos Industriales Microbiologicos; ArgentinaFil: Luccini, Eduardo Alfredo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentina. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Facultad de Química e Ingeniería-Rosario; ArgentinaFil: Salum, Graciela Marisa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Física de Rosario. Universidad Nacional de Rosario. Instituto de Física de Rosario; ArgentinaFil: Piacentini, Ruben Dario Narciso. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Física de Rosario. Universidad Nacional de Rosario. Instituto de Física de Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Exactas, Ingeniería y Agrimensura; Argentina. Universidad Tecnológica Nacional. Facultad Regional Concepción del Uruguay; ArgentinaFil: Farias, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Planta Piloto de Procesos Industriales Microbiologicos; Argentin

Extremophilic bacteria restrict the growth of Macrophomina phaseolina by combined secretion of polyamines and lytic enzymes

[EN] Extremophilic microorganisms were screened as biocontrol agents against two strains of Macrophomina phaseolina (Mp02 and 06). Stenotrophomonas sp. AG3 and Exiguobacterium sp. S58 exhibited a potential in vitro antifungal effect on Mp02 growth, corresponding to 52.2% and 40.7% inhibition, respectively. This effect was confirmed by scanning electron microscopy, where images revealed marked morphological alterations in fungus hyphae. The bacteria were found to secrete lytic enzymes and polyamines. Exiguobacterium sp. S56a was the only strain able to reduce the growth of the two strains of M. phaseolina through their supernatant. Antifungal supernatant activity was correlated with the ability of bacteria to synthesize and excrete putrescine, and the exogenous application of this polyamine to the medium phenocopied the bacterial antifungal effects. We propose that the combined secretion of putrescine, spermidine, and lytic enzymes by extremophilic microorganism predispose these microorganisms to reduce the disease severity occasioned by M. phaseolina in soybean seedlings.The authors acknowledge the generous financial support by the PICT V Bicentenario 2010 1788 Project (FONCyT, Argentina). This work was performed in the context of a project called ¿Análisis de Adaptación al Cambio Climático en Humedales Andinos¿. ID: 6188775¿8-LP13. Ministerio del Medio Ambiente, Región de Antofagasta. We are also grateful to Lic. C. Pérez Brandan for providing us with the M. phaseolina strains used in this study.Santos, AP.; Nieva Muratore, L.; Sole-Gil, A.; Farías, ME.; Ferrando Monleón, AR.; Blazquez Rodriguez, MA.; Belfiore, C. (2021). Extremophilic bacteria restrict the growth of Macrophomina phaseolina by combined secretion of polyamines and lytic enzymes. Plant Biotechnology Reports. 32:1-9. https://doi.org/10.1016/j.btre.2021.e00674S193

Extremophile cyanobacteria: in vitro production of protective compounds against UVB radiation

The Andean Altiplano-Puna is a sedimentary volcanic plateau at an average altitude of 4000 m located between latitudes 13°and 27° south. Solar irradiance is 165% higher than that at the level of the sea with instant flow of UV-B that reaches 17 Wm2,low nutrient concentration particularly phosphorous; presence of heavy metals and arsenic and broad fluctuation of thetemperature of the air, ranging from 20 °C during the day to -10 ° C at night. Even though these conditions are highly limiting,previous results from our laboratory have described the microbial diversity of different lakes, mats and crust of the altiplanoand revealed an unexpectedly diverse microbial community, including several genera of cyanobacteria. Cyanobacteria isolatedfrom extreme conditions like Andean microbial mats and crust could produce mycosporine-like amino acids (MAAs) as amitigation strategy to reduce the damaging effect of ultraviolet radiation. In order to probe the production of MAAs from thesecyanobacteria, the analysis of the methanolic extract by spectrophotometry was used as a rapid method to know the presenceor absence of these compounds. Different strains of cyanobacteria were placed in quartz tubes and exposed to UVB radiationfor 4 and 24 hours. Then the culture was centrifuged at 10000 rpm during 10 min, and 0,15 g of biomass were placed in tubeswith 15 ml of methanol during 24 h in dark. Methanolic extract was analyzed in a spectrophotometer by scanning from 250nm to 750 nm. Also, the methanolic extract was analyzed with high performance liquid chromatographic (HPLC), using WatersAlliance 2695e - Waters PDA 2998 Detector - Empower 2 Software, Column: Gemini C18-5u-4.6 x 250 mm and the mobilephase: 0.1% acetic acid in methanol 2.5%. The cyanobacteria that showed presence of MAAs were cultivated in differentconditions in order to determine the most convenient: condition 1: light 24 h, without shaking and room temperature; condition2: light:dark 12:12 h, without shaking and 28°C; condition 3: light:dark 12:12 h, without shaking but bubbling air into thereactor and 28° C. The strain GTAR 001, Anabaena sp., showed the most significant peak of absorbance at 334 nm when wasexposed for 24 h to UVB. The MAAs reported for cyanobacteria have peaks between 310 and 360 nm and the peak of 334 nmcorresponds to Shinorine. On the other hand, the strain GTAR 001 produced significantly more biomass with condition 3 andin a shorter period. These preliminary results agree with the MAAs reported for Anabaena sp. isolated from rice paddy fieldand hyper saline pond/marine habitat. It will be necessary to analyze this compound with liquid chromatography coupled withtandem mass spectrometry for the final determination.Fil: Lencina, María Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Farias, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Belfiore, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaLVII Annual Meeting of the Argentine Society for Biochemistry and Molecular Biology Research; XVI Annual Meeting of the Argentinean Society for General MicrobiologyCiudad Autónoma de Buenos AriesArgentinaAsociación Civil de Microbiología GeneralSociedad Argentina de Investigación en Bioquímica y Biología Molecula

Combination of bioprotective cultures with EDTA to reduce Escherichia coli O157:H7 in frozen ground-beef patties

The effectiveness of bacteriocin-producing Lactobacillus curvatus CRL705 and Lactococcus lactis CRL1109 in combination with Na2EDTA on frozen ground-beef patties contaminated with Escherichia coli O157:H7, was investigated under temperature abuse conditions (5 C during 9 days). The presence of the bioprotective cultures (ca. 107 CFU/g) and chelator (48 mM) resulted in one log CFU/g reduction for E. coliLactobacillus curvatus CRL705 and Lactococcus lactis CRL1109 in combination with Na2EDTA on frozen ground-beef patties contaminated with Escherichia coli O157:H7, was investigated under temperature abuse conditions (5 C during 9 days). The presence of the bioprotective cultures (ca. 107 CFU/g) and chelator (48 mM) resulted in one log CFU/g reduction for E. coli2EDTA on frozen ground-beef patties contaminated with Escherichia coli O157:H7, was investigated under temperature abuse conditions (5 C during 9 days). The presence of the bioprotective cultures (ca. 107 CFU/g) and chelator (48 mM) resulted in one log CFU/g reduction for E. coliC during 9 days). The presence of the bioprotective cultures (ca. 107 CFU/g) and chelator (48 mM) resulted in one log CFU/g reduction for E. coli7 CFU/g) and chelator (48 mM) resulted in one log CFU/g reduction for E. coli strain, compared to the control on day 0. Similarly, a significant decline for indigenous coliforms in ground-beef patties was also observed in the presence of bacteriocinogenic strains and chelator. However, in the absence of Na2EDTA, neither E. coli nor coliforms were inhibited by the bioprotective cultures, the pathogen reaching similar counts than control samples (5.22 and 3.60 log CFU/g, respectively) at 9 days. When the growth of bacteriocinogenic strains on patties was evaluated, they were able to increase their population producing bacteriocins after 48 h up to the end of incubation period while a near neutral pH in the presence of Na2EDTA was detected. Non substantial effect on of ground-beef patties color was produced in the presence of bioprotective cultures, while a darker color developed in those added with the chelator. The simultaneous treatment with bioprotective cultures and Na2EDTA may be of value for the control of E. coli O157:H7 in temperature abused ground-beef patties.ficant decline for indigenous coliforms in ground-beef patties was also observed in the presence of bacteriocinogenic strains and chelator. However, in the absence of Na2EDTA, neither E. coli nor coliforms were inhibited by the bioprotective cultures, the pathogen reaching similar counts than control samples (5.22 and 3.60 log CFU/g, respectively) at 9 days. When the growth of bacteriocinogenic strains on patties was evaluated, they were able to increase their population producing bacteriocins after 48 h up to the end of incubation period while a near neutral pH in the presence of Na2EDTA was detected. Non substantial effect on of ground-beef patties color was produced in the presence of bioprotective cultures, while a darker color developed in those added with the chelator. The simultaneous treatment with bioprotective cultures and Na2EDTA may be of value for the control of E. coli O157:H7 in temperature abused ground-beef patties.2EDTA, neither E. coli nor coliforms were inhibited by the bioprotective cultures, the pathogen reaching similar counts than control samples (5.22 and 3.60 log CFU/g, respectively) at 9 days. When the growth of bacteriocinogenic strains on patties was evaluated, they were able to increase their population producing bacteriocins after 48 h up to the end of incubation period while a near neutral pH in the presence of Na2EDTA was detected. Non substantial effect on of ground-beef patties color was produced in the presence of bioprotective cultures, while a darker color developed in those added with the chelator. The simultaneous treatment with bioprotective cultures and Na2EDTA may be of value for the control of E. coli O157:H7 in temperature abused ground-beef patties.2EDTA was detected. Non substantial effect on of ground-beef patties color was produced in the presence of bioprotective cultures, while a darker color developed in those added with the chelator. The simultaneous treatment with bioprotective cultures and Na2EDTA may be of value for the control of E. coli O157:H7 in temperature abused ground-beef patties.2EDTA may be of value for the control of E. coli O157:H7 in temperature abused ground-beef patties.E. coli O157:H7 in temperature abused ground-beef patties.Fil: Castellano, Patricia Haydee. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Tucumán. Centro de Referencia Para Lactobacilos (i); ArgentinaFil: Belfiore, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Tucumán. Centro de Referencia para Lactobacilos (i); ArgentinaFil: Vignolo, Graciela Margarita. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Tucumán. Centro de Referencia para Lactobacilos (i); Argentin

Temperature, soluble solids and pH effect on Alicyclobacillus acidoterrestris viability in lemon juice concentrate

Alicyclobacillus acidoterrestris is a thermoacidophilic, non-pathogenic, spore-forming bacterium detected in spoiled commercial pasteurized fruit juice. Apple, white grape and tomato are particularly susceptible. A. acidoterrestris spores are resistant to lemon juice pasteurization (2 min at 82°C), and they can germinate and grow causing spoilage. This contamination is characterized by a medicinal or disinfectant smell attributed to guaiacol (o-dihydroxybenzene) production and other taint chemicals. The aim of this work was to study the influence of temperature (82, 86, 92 and 95 °C), total soluble solids (SS) (6.20, 9.8, 50 and 68°Brix) and pH (2.28, 2.45, 2.80, 3.25, 3.5) on decimal reduction time (D) of the A. acidoterrestris in clarified and non-clarified concentrated lemon juice. Once D-value was determined, the resistance of A. acidoterrestris at the assayed temperatures was confirmed. SS and pH influence spore viability, because spore resistance increases with higher SS (50°Brix 22 min 82 °C–68°Brix 28 min 82 °C) and pH values (pH 2.28, 17 min–pH 4.00, 22 min). Bacterial growth was lower in clarified lemon juice, 26 min at 82 °C, than in non-clarified lemon juice, 51 min at 82 °C. Temperature was the parameter that had the greatest influence on the D value.Fil: Belfiore, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; Argentina. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Biotecnología; ArgentinaFil: Maldonado, Maria Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; Argentina. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Biotecnología; ArgentinaFil: Navarro, Antonio Roberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán; Argentina. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Biotecnología; Argentin

Proteomic Approach of Adaptive Response to Arsenic Stress in Exiguobacterium sp. S17. An Extremophile Strain Isolate from High Altitude Andean Lake Stromatolite

North West of Argentina is particularly rich in wetlands located in the Puna in a range of altitude of 3,600 to 4,600 m above sea level. Most of these high-altitude Andean lakes are inhospitable areas suffering from extreme conditions such as high content of toxic elements particularly Arsenic. Exiguobacterium sp. S17, isolated from stromatolite in Laguna Socompa, showed tolerance to high arsenic concentration. The assays revealed that this strain was able to growth in presence of 10mM and 150mM of As(III) and As(V) respectively. To understand the mechanisms implicated in Exiguobacterium sp. S17 tolerance arsenic to proteomic approach was conducted. Comparative proteomic analyses of S17 arsenic exposed and unexposed to reveal that 25 spots were differentially expressed. Identification of proteins was performed by MALDI-TOF/MS, and the results obtained revealed that proteins involved in energy-metabolism, stress, transport, and in proteins synthesis were expressed under arsenic stress. To our knowledge, this work represents the first proteomic study of arsenic tolerance in an Exiguobacterium strain.Fil: Belfiore, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Ordoñez, Omar Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Farias, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; Argentin

Characterization of Lactobacillus curvatus G, a lactocin 705 and antilisteria-deficient mutant

Lactobacillus curvatus CRL705 produces two bacteriocins, Lac705 and AL705, and harbors two plasmids, pRC18 and pRC12. The gene determinants of lactocin Lac705, genes #705/#705, imm705, abc705 and acc705, are encoded in pRC18, a plasmid of 18600 bp whose DNA sequence has been determined. In this study, the characterization of a Lac705- and AL705- deficient mutant, named strain G, is described. Strain G has been isolated after transforming L. curvatus CRL705 with plasmid pTV01::Tn917 and growing the transformants at high temperature (39C). Confirmation that strain G derived from CRL705 was determined by API50 analysis and by RAPD reactions, using primers RAPD1, RAPD2, REP1RI, REP2I and REPV. Agarose gel electrophoresis showed that plasmids pRC12 and pRC18 were absent in strain G. However, the presence pRC18 in strain G, and therefore of the lactocin Lac705 operon, was confirmed by PCR reactions using a set of three pRC18-specific primers (primers #1/#2, #3/#4 and #5/#6). The abundance and size of the amplified PCR products suggested that plasmid pRC18 is present in strain G in a low copy number and that the structural genes of lactocin Lac705 (705/705) are rearranged, which could explain the phenotype observed.Fil: Olaya Passarell, María José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Belfiore, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Raya, Raul Ricardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaIII Simposio Internacional de Bacterias Lácticas; II Encuentro de la Red Argentina de Bacterias LácticasTucumanArgentinaConsejo Nacional de Investigaciones Científicas y Técnicas. Centro de Referencia para LactobacilosMinisterio de Salud PúblicaAgencia Nacional de Promoción Científica y Tecnológic