Prevalence of Lentilacobacillus hilgardii over Lactiplantibacillus plantarum in Low-Temperature Spontaneous Malolactic Fermentation of a Patagonian Pinot Noir

The spontaneous malolactic fermentation (MLF) in a centenary winery from Patagonia, Argentina, is conducted by predominantly mesophilic Oenococcus oeni and Lactiplantibacillus plantarum. In this region, MLF takes place from 14 to 4 °C, leading to heat cellars incurring in higher costs and non-sustainable practices. Previously, psychrotrophic strains of O. oeni had been obtained from a Patagonian wine. The goal of this work was to identify the Lactobacillaceae microbiota related to low-temperature MLF and assess their contribution. Nine sychrotrophicc Lentilactibacillus hilgardii strains were identified by sequencing the 16S rRNA gene, and the strains typified by RAPD-PCR. All strains consumed L-malic acid at 4 and 10 °C in sterile wine. The selected UNQLh1.1 strain revealed implantation capacity and L-malic acid consumption at 4 and 10 °C in the presence of the native microbial consortium. Furthermore, the histidine decarboxylase (hdc) gene was not detected in any of the Len. hilgardii strains. The prevalence of Len. hilgardii under low-temperature conditions represents a novelty compared to previous findings of LAB diversity in the MLF of Patagonian wines. The native Patagonian psychrotrophic Len. hilgardii strains are a new player in fermentations conducted at low temperatures with the potential to be used as a sustainable MLF starter.Fil: Manera, Camila. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Rivas, Gabriel Alejandro. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Flores, Naiquen Elizabeth. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Brizuela, Natalia Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Caballero, Adriana Carmen. Universidad Nacional del Comahue; Argentina. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas; ArgentinaFil: Semorile, Liliana Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Valdes la Hens, Danay. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Complete Genome Sequencing of Lactobacillus plantarum UNQLp 11 Isolated from a Patagonian Pinot Noir Wine

Lactobacillus plantarum UNQLp 11 strain was isolated from a Patagonian Pinot noir wine at the oldest commercial winery (110 years old) in General Roca, North Patagonia, Argentina, and has demonstrated its ability to survive during winemaking processes and successfully carry out malolactic fermentation. This work aimed to obtain the whole assembled genome of the UNQLp 11 strain, analysing its architecture and the possible functions of the predicted genes from the oenological properties of this strain. The genome size is 3 534 932 bp, with a mean GC content of 44.2%, 3 412 CDS, 80 transposons and 148 tandem repeats. A comparison between the genome size and gene content of 14 Lb. plantarum strains from different origins was performed, and UNQLp 11 exhibited the largest size. The in silico genome-wide analysis allowed us to confirm the existence of genes encoding enzymes involved in the synthesis of several metabolites of oenological interest, in addition to bacteriocins and exopolysaccharides. Furthermore, it is possible to speculate on this strain’s adaptation to different environments, as it is able to use diverse substrates for its growth. All these features suggest the potential of UNQLp 11 to be a good starter culture for malolactic fermentation.Fil: Iglesias, Nestor Gabriel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Instituto de Microbiología Básica y Aplicada; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Brizuela, Natalia Soledad. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Tymczyszyn, Emma Elizabeth. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Hollmann, Axel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet Noa Sur. Centro de Investigación en Biofísica Aplicada y Alimentos. - Universidad Nacional de Santiago del Estero. Centro de Investigación en Biofísica Aplicada y Alimentos; ArgentinaFil: Valdes la Hens, Danay. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Semorile, Liliana Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Bravo Ferrada, Barbara Mercedes. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Relative expression of stress-related genes during acclimation at low temperature of psychrotrophic Oenococcus oeni strains from Patagonian wine

In the present study, we evaluated the transcriptional response of four stress-related genes in three Oenococcus oeni strains after acclimation at two different temperatures. Gene expression was analyzed at time zero and after 48 h acclimation at 18 and 21 °C. After the acclimation period cells were inoculated into sterile Pinot noir wine and MLF was followed for 25 days to investigate if different acclimation temperatures could influence cell survival and MLF performance. l-malic acid consumption, population survival, and transcriptional behavior were different upon the acclimation temperature. rmlB and hsp20 genes presented a considerable increase in their expression level when strains were acclimated at 18 °C particularly in the psychrotrophic strains UNQOe19 and UNQOe4 isolated from Patagonian Pinot noir wine in comparison with the control strain (ATCC 27310). The increase in rmlB and hsp20 expression could account for the better survival of these strains in Pinot noir in comparison with the control strain. In addition, Patagonian populations acclimated at 18 °C were able to consume a higher percentage of l-malic acid in comparison with cells acclimated at 21 °C. Our results suggest that gene expression analysis of cells acclimated at sub-optimal temperatures could benefit the selection of psychrotrophic strains aimed as starter cultures.Fil: Olguin, Nair Temis. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Valdes la Hens, Danay. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Delfederico, Lucrecia. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Semorile, Liliana Carmen. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentin

Prevalence of Lactobacillus plantarum and Oenococcus oeni during spontaneous malolactic fermentation in Patagonian red wines revealed by polymerase chain reaction-denaturing gradient gel electrophoresis with two targeted genes

Background and Aims:Information regarding the composition and dynamics of microbial communities through-out winemaking is always valuable to control the process and contributes to improving the quality of wine. The aimsof this work were to analyse the diversity of lactic acid bacteria (LAB) species present during the spontaneousmalolactic fermentation (MLF) of Patagonian red wines, and to isolate and identify native LAB species.Methods and Results:Samples of Merlot and Pinot Noir wines were collected during three vintages at onecommercial cellar. Through polymerase chain reaction (PCR)-rpoB/denaturing gradient gel electrophoresis (DGGE),it was possible to identify 15 LAB species and through PCR-16S rRNAV3 region/DGGE, 11 LAB species, duringspontaneous MLF of these wines, mostly,Oenococcus oeniandLactobacillus plantarum. These two species were alsolargely recovered by culture and analysed by the random amplified polymorphic DNA–PCR technique. Isolates ofthese two LAB species exhibited considerable genotypic heterogeneity.Conclusions:The prevalence ofO. oeniandLb. plantarumin all samples suggests that both species are involved inleading the spontaneous MLF of these Patagonian wines. The analysis of the LAB microbiota in wine by PCR-DGGEusing two gene regions enabled addition to the number of species that can usually be detected during MLF.Significance of the Study:The study offered a more complete knowledge of the LAB community present in winesin a cellar in the Argentine Patagonia during the spontaneous MLF. Our findings are particularly relevant, becausethey contribute to the possibility of designing an MLF starter composed of native strains ofLb. plantarumandO. oeniFil: Valdes la Hens, Danay. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Bravo Ferrada, Barbara Mercedes. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Delfederico, Lucrecia. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Caballero, Adriana Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico CONICET - Patagonia Norte. Instituto de Investigación y Desarrollo en Ingeniería de Procesos, Biotecnología y Energías Alternativas. IDEPA - Subsede San Antonio Oeste; Argentina. Universidad Nacional del Comahue; ArgentinaFil: Semorile, Liliana Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentin

Growth and consumption of l-malic acid in wine-like medium by acclimated and non-acclimated cultures of Patagonian Oenococcus oeni strains

Five Oenococcus oeni strains, selected from spontaneous malolactic fermentation (MLF) of Patagonic Pinot noir wine, were assessed for their use as MLF starter cultures. After the individual evaluation of tolerance to some stress conditions, usually found in wine (pH, ethanol, SO2, and lysozyme), the behavior of the strains was analyzed in MLO broth with 14 % ethanol and pH 3.5 in order to test for the synergistic effect of high ethanol level and low pH and, finally, in a wine-like medium. Although the five strains were able to grow in MLO broth under low pH and/or high ethanol, they must be acclimated to grow in a wine-like medium. Additionally, glycosidase and tannase activities were evaluated, showing differences among the strains. The potential of the strains to ferment citrate was tested and two of the five strains showed the ability to metabolize this substrate. We did not detect the presence of genes encoding histidine, tyrosine descarboxylase, and putrescine carbamoyltransferase. All the strains tested exhibited good growth capacity and ability to consume l-malic acid in a wine-like medium after cell acclimation, and each of them showed a particular enzyme profile, which might confer different organoleptic properties to the wine.Fil: Bravo Ferrada, Barbara Mercedes. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Hollmann, Axel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro de Investigaciones y Transferencia de Santiago del Estero. Universidad Nacional de Santiago del Estero. Centro de Investigaciones y Transferencia de Santiago del Estero; ArgentinaFil: Brizuela, Natalia Soledad. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Valdes la Hens, Danay. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Tymczyszyn, Emma Elizabeth. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Semorile, Liliana Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentin

Lactobacillus plantarum as a malolactic starter culture in winemaking: A new (old) player?

Malolactic fermentation (MLF) is a process in winemaking responsible for the conversion of l-malic acid to l-lactic acid and CO2, which reduces the total acidity, improves the biological stability, and modifies the aroma profile of wine. MLF takes place during or after alcoholic fermentation and is carried out by one or more species of lactic acid bacteria (LAB), which are either present in grapes and cellars or inoculated with malolactic starters during the winemaking process. Although the main bacterium among LAB used in commercial starter cultures for MLF has traditionally been Oenococcus oeni, in the last decade, Lactobacillus plantarum has also been reported as a malolactic starter, and many works have shown that this species can survive and even grow under harsh conditions of wine (i.e., high ethanol content and low pH values). Furthermore, it has been proved that some strains of L. plantarum are able to conduct MLF just as efficiently as O. oeni. In addition, L. plantarum exhibits a more diverse enzymatic profile than O. oeni, which could play an important role in the modification of the wine aroma profile. This enzymatic diversity allows obtaining several starter cultures composed of different L. plantarum biotypes, which could result in distinctive wines. In this context, this review focuses on showing the relevance of L. plantarum as a MLF starter culture in winemaking.How to cite: Brizuela NS, Tymczyszyn E, Semorile LC, et al. Lactobacillus plantarum as a malolactic starter culture in winemaking: a new (old) player? Electron J Biotechnol 2019;38. https://doi.org/10.1016/j.ejbt.2018.12.002. Keywords: Alcoholic fermentation, Aroma, Grapes, Lactic acid bacteria, Lactobacillus plantarum, Malolactic fermentation, Malolactic starter culture, Wine, Winemakin

Comparative vinification assays with selected Patagonian strains of Oenococcus oeni and Lactobacillus plantarum

The performance of Patagonian Lactobacillus plantarum and Oenococcus oeni strains as malolactic starter cultures was compared. Two autochthonous strains of each species were selected, based on the presence of aroma-related genes, and inoculated in sterile wine of high ethanol content. The effects of initial inoculum size and acclimation treatment on the efficiency of malolactic fermentation (MLF) were analyzed for each strain. O. oeni strains were able to successfully conduct the MLF only when the inoculum concentration was higher than 1.108 CFU/mL and cells were acclimated in sublethal ethanol concentrations. The increase of ethanol concentration in the acclimation medium also improved the kinetics of malic acid consumption. Successful MLF with L. plantarum strains required lower inocula and no acclimation treatment. In addition, these strains showed a better profile of aroma-related genes than O. oeni. L. plantarum strains appeared to be more efficient than O. oeni strains as candidates for malolactic starter cultures to be used in Patagonian red wines.Fil: Brizuela, Natalia Soledad. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Bravo Ferrada, Barbara Mercedes. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Valdes la Hens, Danay. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Hollmann, Axel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro de Investigaciones y Transferencia de Santiago del Estero. Universidad Nacional de Santiago del Estero. Centro de Investigaciones y Transferencia de Santiago del Estero; ArgentinaFil: Delfederico, Lucrecia. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Caballero, Adriana Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Instituto de Investigación y Desarrollo en Ingeniería de Procesos, Biotecnología y Energías Alternativas. Universidad Nacional del Comahue. Instituto de Investigación y Desarrollo en Ingeniería de Procesos, Biotecnología y Energías Alternativas; ArgentinaFil: Tymczyszyn, Emma Elizabeth. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Semorile, Liliana Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentin