RESUMEN

Garviecin LG34 produced by Lactococcus garvieae LG34 exhibits wide-spectrum antibacterial activity against both Gram-positive and Gram-negative bacteria. This work aimed at clarifying the antibacterial mode of action of garviecin LG34 against Gram-negative bacterium Salmonella typhimurium. To determine the concentration for the bacteriocin antimicrobial mode experiments, the minimum inhibitory concentration of garviecin LG34 against S. typhimurium CICC21484 was determined as 0.25 mg/ml. Garviecin LG34 decreased the viable count of S. typhimurium CICC21484 and its antibacterial activity was the dose and time dependant. Garviecin LG34 led to the dissipation of transmembrane potential, the rise in the extracellular conductivity, UV-absorbing material at 260 nm, and LDH level of S. typhimurium CICC21484. Scanning electron micrographs results shown that garviecin LG34 cause dramatic deformation and fragmentation including the flagellum shedding, pores formation in surface, and even completely breakage of S. typhimurium cell. Moreover, garviecin LG34 decreased the intracellular ATP level. The results of this study demonstrated that garviecin LG34 can destroy cell structure, increase membrane permeability of S. typhimurium, thereby might be used as biopreservative for treating food borne and salmonellosis resulting from Gram-negative bacterium S. typhimurium.

Asunto(s)

Antibacterianos , Pruebas de Sensibilidad Microbiana , Salmonella typhimurium , Salmonella typhimurium/efectos de los fármacos , Salmonella typhimurium/crecimiento & desarrollo , Antibacterianos/farmacología , Antibacterianos/química , Bacteriocinas/farmacología , Lactococcus/efectos de los fármacos , Adenosina Trifosfato/metabolismo , Potenciales de la Membrana/efectos de los fármacos

RESUMEN

The wild-type Lactococcus lactis strain LAC460 produces two bacteriocin-like phage lysins, LysL and LysP. This study aimed to produce and secrete LysL in various heterologous hosts and an in vitro cell-free expression system for further functional studies. Initially, the lysL gene from L. lactis LAC460 was cloned into Lactococcus cremoris NZ9000 and L. lactis N8 strains, with and without the usp45 signal sequence (SSusp45), under a nisin-inducible promoter. Active LysL was primarily produced intracellularly in recombinant L. lactis N8, with some secretion into the supernatant. Recombinant L. cremoris NZ9000 lysed upon nisin induction, indicating successful lysL expression. However, fusion with Usp45 signal peptide (SPUsp45-LysL) weakened LysL activity, likely due to incomplete signal peptide cleavage during secretion. Active LysL was also produced in vitro, and analysed in SDS-PAGE, giving a 42-kDa band. However, the yield of LysL protein was still low when produced from recombinant lactococci or by in vitro expression system. Therefore, His-tagged LysL was produced in Escherichia coli BL21(DE3). Western blot confirmed the intracellular production of about 44-kDa His-tagged LysL in E. coli. His-tagged active LysL was then purified by Ni-NTA affinity chromatography yielding sufficient 4.34 mg of protein to be used in future functional studies.

Asunto(s)

Bacteriocinas , Lactococcus lactis , Lactococcus lactis/genética , Lactococcus lactis/metabolismo , Lactococcus lactis/virología , Bacteriocinas/genética , Bacteriocinas/metabolismo , Bacteriocinas/biosíntesis , Clonación Molecular , Nisina/farmacología , Nisina/genética , Nisina/metabolismo , Señales de Clasificación de Proteína/genética , Expresión Génica , Lactococcus/genética , Lactococcus/metabolismo , Lactococcus/virología , Bacteriófagos/genética

RESUMEN

Until the late 2000s, lactococci substantially contributed to the discovery of various plasmid-borne phage defence systems, rendering these bacteria an excellent antiphage discovery resource. Recently, there has been a resurgence of interest in identifying novel antiphage systems in lactic acid bacteria owing to recent reports of so-called 'defence islands' in diverse bacterial genera. Here, 321 plasmid sequences from 53 lactococcal strains were scrutinized for the presence of antiphage systems. Systematic evaluation of 198 candidates facilitated the discovery of seven not previously described antiphage systems, as well as five systems, of which homologues had been described in other bacteria. All described systems confer resistance against the most prevalent lactococcal phages, and act post phage DNA injection, while all except one behave like abortive infection systems. Structure and domain predictions provided insights into their mechanism of action and allow grouping of several genetically distinct systems. Although rare within our plasmid collection, homologues of the seven novel systems appear to be widespread among bacteria. This study highlights plasmids as a rich repository of as yet undiscovered antiphage systems.

Asunto(s)

Bacteriófagos , Lactococcus , Plásmidos , Plásmidos/genética , Bacteriófagos/genética , Lactococcus/genética , Lactococcus/virología

RESUMEN

The biosynthetic machinery for cell wall polysaccharide (CWPS) formation in Lactococcus lactis and Lactococcus cremoris is encoded by the cwps locus. The CWPS of lactococci typically consists of a neutral rhamnan component, which is embedded in the peptidoglycan, and to which a surface-exposed side chain oligosaccharide or polysaccharide pellicle (PSP) component is attached. The rhamnan component has been shown for several lactococcal strains to consist of a repeating rhamnose trisaccharide subunit, while the side chain is diverse in glycan content, polymeric status and glycosidic linkage architecture. The observed structural diversity of the CWPS side chain among lactococcal strains is reflected in the genetic diversity within the variable 3' region of the corresponding cwps loci. To date, four distinct cwps genotypes (A, B, C, D) have been identified, while eight subtypes (C1 through to C8) have been recognized among C-genotype strains. In the present study, we report the identification of three novel subtypes of the lactococcal cwps C genotypes, named C9, C10 and C11. The CWPS of four isolates representing C7, C9, C10 and C11 genotypes were analysed using 2D NMR to reveal their unique CWPS structures. Through this analysis, the structure of one novel rhamnan, three distinct PSPs and three exopolysaccharides were elucidated. Results obtained in this study provide further insights into the complex nature and fascinating diversity of lactococcal CWPSs. This highlights the need for a holistic view of cell wall-associated glycan structures which may contribute to robustness of certain strains against infecting bacteriophages. This has clear implications for the fermented food industry that relies on the consistent application of lactococcal strains in mesophilic production systems.

Asunto(s)

Pared Celular , Genotipo , Lactococcus , Leche , Pared Celular/química , Lactococcus/genética , Lactococcus/aislamiento & purificación , Lactococcus/clasificación , Animales , Leche/microbiología , Polisacáridos Bacterianos/química , Polisacáridos Bacterianos/metabolismo , Alimentos Fermentados/microbiología , Polisacáridos/metabolismo , Microbiología de Alimentos

RESUMEN

Dairy industries apply selected lactococcal strains and mixed cultures to produce diverse fermented products with distinctive flavor and texture properties. Innovation of the starter culture functionality in cheese applications embraces natural biodiversity of the Lactococcus species to identify novel strains with alternative flavor or texture forming capacities and/or increased processing robustness and phage resistance. Mobile genetic elements (MGE), like integrative conjugative elements (ICEs) play an important role in shaping the biodiversity of bacteria. Besides the genes involved in the conjugation of ICEs from donor to recipient strains, these elements also harbor cargo genes that encode a wide range of functions. The definition of such cargo genes can only be achieved by accurate identification of the ICE boundaries (delimiting). Here, we delimited 25 ICEs in lactococcal genome sequences with low contig numbers using insertion-sites flanking single-copy core-genome genes as markers for each of the distinct ICE-integrases we identified previously within the conserved ICE-core genes. For ICEs in strains for which genome information with large numbers of contigs is available, we exemplify that CRISPR-Cas9 driven ICE-curing, followed by resequencing, allows accurate delimitation and cargo definition of ICEs. Finally, we compare and contrast the cargo gene repertoire of the 26 delimited lactococcal ICEs, identifying high plasticity among the cargo of lactococccal ICEs and a range of encoded functions that is of apparent industrial interest, including restriction modification, abortive infection, and stress adaptation genes.

Asunto(s)

Genoma Bacteriano , Lactococcus/genética , Secuencias Repetitivas Esparcidas/genética , Sistemas CRISPR-Cas , Conjugación Genética

RESUMEN

To understand the relationship between changes in aroma and bacteria in pigeon breast meat (PBM) during preservation, bacterial communities and volatile compounds in PBM were analyzed using high-throughput sequencing and gas chromatography-ion mobility spectrometry. Analyses of total viable bacteria counts revealed that modified atmospheric packaging (MAP) and electron beam irradiation (EBI) could be used to extend the shelf-life of PBM to 10 d and 15 d, respectively. Furthermore, Lactococcus spp. and Psychrobacter spp. were the dominant bacterial genera of the MAP and EBI groups, respectively. The results of the study revealed 91 volatile organic compounds, one of which, butanal, was the most intense volatile organic compound while being an important source of aroma differences between the physical preservation techniques. Alpha-terpinolene, acetoin-M, gamma-butyrolactone, 1-hexanol-M, and 2,6-dimethyl-4-heptanone may be markers of PBM spoilage. During preservation, the MA group (treatment with 50 % CO2 + 50 % N2) demonstrated greater stabilization of PBM aroma. A Spearman correlation analysis showed that Lactococcus spp., Psychrobacter spp., and Pseudomonas spp. were the dominant bacterial genera of PBM during preservation and were closely related to an increase in the intensity of anisole, 2-methyl-3-furanthiol, and 5-methyl-2-furanmethanol, respectively. Lactococcus spp. and Psychrobacter spp. play crucial roles in the sensory degradation of PBM. In this study, we analyzed the changes in bacterial genera and volatile organic compounds of PBM under different physical preservation techniques to identify a suitable method for preserving PBM and evaluating its freshness.

Asunto(s)

Columbidae , Microbiología de Alimentos , Psychrobacter , Compuestos Orgánicos Volátiles , Compuestos Orgánicos Volátiles/análisis , Animales , Columbidae/microbiología , Psychrobacter/metabolismo , Odorantes/análisis , Conservación de Alimentos/métodos , Bacterias/clasificación , Carne/microbiología , Carne/análisis , Embalaje de Alimentos/métodos , Lactococcus , Cromatografía de Gases y Espectrometría de Masas , Aldehídos/análisis , Microbiota

RESUMEN

Lactococcus garvieae has recently been identified and listed as one of the causative agents of hyperacute hemorrhagic sepsis in fish. In intensive recirculating aquaculture systems where there are high fish densities and minimal water changes, not only will it be conducive to the growth of bacteria, but Cryptocaryon irritans as a marine protozoan fish parasite is also prone to appear. This study reports the disease status of Trachinotus ovatus in an aquaculture area in Yangjiang City, Guangdong Province. Through the diagnosis of clinical symptoms of the diseased fish, identification of specific primers, 16s rRNA sequences phylogenetic tree analysis, physiological and biochemical identification, and observation of histopathological sections, the result of the experiment is that the mass death of T. ovatus is caused by a mixture of L. garvieae and C. irritants infections. Subsequently, regression infection experiments were performed to verify Koch's law. It was confirmed that the pathogen had strong virulence to T. ovatus. This is the first time that the co-infection of L. garvieae and C. irritans to T. ovatus was found in South China. The research results of this experiment have certain enlightenment significance for the epidemic trend of fish diseases in relevant sea areas.

Asunto(s)

Enfermedades de los Peces , Lactococcus , Filogenia , Animales , Lactococcus/genética , Lactococcus/aislamiento & purificación , Lactococcus/clasificación , Enfermedades de los Peces/microbiología , Enfermedades de los Peces/parasitología , China , Cilióforos/genética , Cilióforos/clasificación , Cilióforos/aislamiento & purificación , Acuicultura , ARN Ribosómico 16S/genética , Coinfección/microbiología , Coinfección/parasitología , Infecciones por Cilióforos/parasitología , Infecciones por Cilióforos/veterinaria , Peces/parasitología , Peces/microbiología , Infecciones por Bacterias Grampositivas/microbiología , Infecciones por Bacterias Grampositivas/veterinaria

RESUMEN

Lactococcus lactis is a lactic acid bacterium of major importance for food fermentation and biotechnological applications. The ability to manipulate its genome quickly and easily through competence for DNA transformation would accelerate its general use as a platform for a variety of applications. Natural transformation in this species requires the activation of the master regulator ComX. However, the growth conditions that lead to spontaneous transformation, as well as the regulators that control ComX production, are unknown. Here, we identified the carbon source, nitrogen supply, and pH as key factors controlling competence development in this species. Notably, we showed that these conditions are sensed by three global regulators (i.e., CcpA, CodY, and CovR), which repress comX transcription directly. Furthermore, our systematic inactivation of known signaling systems suggests that classical pheromone-sensing regulators are not involved. Finally, we revealed that the ComX-degrading MecA-ClpCP machinery plays a predominant role based on the identification of a single amino-acid substitution in the adaptor protein MecA of a highly transformable strain. Contrasting with closely-related streptococci, the master competence regulator in L. lactis is regulated both proximally by general sensors and distantly by the Clp degradation machinery. This study not only highlights the diversity of regulatory networks for competence control in Gram-positive bacteria, but it also paves the way for the use of natural transformation as a tool to manipulate this biotechnologically important bacterium.

Asunto(s)

Proteínas Bacterianas , Regulación Bacteriana de la Expresión Génica , Redes Reguladoras de Genes , Lactococcus lactis , Lactococcus lactis/genética , Lactococcus lactis/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Transformación Bacteriana/genética , Lactococcus/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Competencia de la Transformación por ADN/genética

RESUMEN

Co-infection of Lactococcus garvieae and Aeromonas hydrophila, has been confirmed from diseased Nile Tilapia (Oreochromis niloticus), Chithralada strain cultured in a freshwater rearing pond of Alappuzha district of Kerala, India. The aetiological agents behind the disease outbreak were bacteriologically proven and confirmed by 16SrRNA sequencing and phylogenetic analysis. PCR detection of the virulent genes, showed existence of adhesin and hemolysin in L. garvieae and aerolysin in A. hydrophila strain obtained. To fulfil Koch's postulates, challenge experiments were conducted and median lethal dose (LD50) of L. garvieae and A. hydrophila was calculated as 1 × 105.91 CFU per mL and 1 × 105.2 CFU per mL respectively. Histopathologically, eyes, spleen, and kidney were the predominantly infected organs by L. garvieae and A. hydrophila. Out of the 13 antibiotics tested to check antibiotic susceptibility, L. garvieae showed resistance to almost 7 antibiotics tested, with a resistance to Ciprofloxacin while A. hydrophila was found resistant to Streptomycin and Erythromycin. Understanding the complex interaction between Gram-positive and Gram-negative bacteria in the disease process and pathogenesis in fish host will contribute to efficient treatment strategies. As a preliminary investigation into this complex interaction, the present study is aimed at phenotypic and genotypic characterization, pathogenicity evaluation, and antibiotic susceptibility of the co-infecting pathogens in a diseased sample of freshwater-farmed Nile tilapia.

Asunto(s)

Aeromonas hydrophila , Antibacterianos , Cíclidos , Coinfección , Enfermedades de los Peces , Infecciones por Bacterias Gramnegativas , Lactococcus , Filogenia , Animales , Aeromonas hydrophila/genética , Aeromonas hydrophila/aislamiento & purificación , Aeromonas hydrophila/patogenicidad , Aeromonas hydrophila/clasificación , Aeromonas hydrophila/efectos de los fármacos , Cíclidos/microbiología , India , Enfermedades de los Peces/microbiología , Lactococcus/genética , Lactococcus/aislamiento & purificación , Lactococcus/clasificación , Lactococcus/patogenicidad , Infecciones por Bacterias Gramnegativas/microbiología , Infecciones por Bacterias Gramnegativas/veterinaria , Coinfección/microbiología , Coinfección/veterinaria , Antibacterianos/farmacología , Pruebas de Sensibilidad Microbiana , Infecciones por Bacterias Grampositivas/microbiología , Infecciones por Bacterias Grampositivas/veterinaria , ARN Ribosómico 16S/genética , Acuicultura , Proteínas Hemolisinas/genética , Proteínas Hemolisinas/metabolismo

RESUMEN

Lactococcus (Lc.) paracarnosus and the phylogenetically closely related Lc. carnosus species are common members of the microbiota in meat stored under modified atmosphere and at low temperature. The effect of these strains on meat spoilage is controversially discussed. While some strains are known to cause spoilage, others are being studied for their potential to suppress the growth of spoilage and pathogenic bacteria. In this study, Lc. paracarnosus DSM 111017T was selected based on a previous study for its ability to suppress the growth of meat spoilers, including Brochothrix thermosphacta. The mechanism by which this bioprotective strain inhibits competing bacteria and how it contributes to spoilage are not yet known. To answer these two questions, we investigated the effect of four different headspace gas mixtures (simulated air (21 % O2/79 % N2); HiOx-MAP (70 % O2/30 % CO2); nonOx-MAP (70 % N2/ 30 % CO2); simulated vacuum (100 % N2) and the presence of Brochothrix (B.) thermosphacta TMW 2.2101 on the growth and transcriptional response of Lc. paracarnosus DSM 111017T when cultured on a meat simulation agar surface at 4 °C. Analysis of genes specifically upregulated by the gas mixtures used revealed metabolic pathways that may lead to different levels of spoilage metabolites production. We propose that under elevated oxygen levels, Lc. paracarnosus preferentially converts pyruvate from glucose and glycerol to uncharged acetoin/diacetyl instead of lactate to counteract acid stress. Due to the potential production of a buttery off-flavour, the strain may not be suitable as a protective culture in meat packaged under high­oxygen conditions. 70 % N2/ 30 % CO2, simulated vacuum- and the presence of Lc. paracarnosus inhibited the growth of B. thermosphacta TMW 2.2101. However, B. thermosphacta did not affect gene regulation of metabolic pathways in Lc. paracarnosus, and genes previously predicted to be involved in B. thermosphacta growth suppression were not regulated at the transcriptional level. In conclusion, the study indicates that the gas mixture used in packaging significantly affects the metabolism and spoilage potential of Lc. paracarnosus and its ability to inhibit B. thermosphacta growth.

Asunto(s)

Brochothrix , Técnicas de Cocultivo , Lactococcus , Transcriptoma , Brochothrix/crecimiento & desarrollo , Brochothrix/genética , Brochothrix/metabolismo , Brochothrix/efectos de los fármacos , Lactococcus/metabolismo , Lactococcus/genética , Lactococcus/crecimiento & desarrollo , Microbiología de Alimentos , Vacio , Gases/farmacología , Gases/metabolismo , Oxígeno/metabolismo , Oxígeno/farmacología , Carne/microbiología , Regulación Bacteriana de la Expresión Génica , Dióxido de Carbono/metabolismo , Dióxido de Carbono/farmacología

RESUMEN

BACKGROUND: Although microorganisms are the main cause of spoilage in prepared beef steaks, very few deep spoilage mechanisms have been reported so far. Aiming to unravel the mechanisms during 12 days of storage at 4 °C affecting the quality of prepared beef steak, the present study investigated the changes in microbial dynamic community using a combined high-throughput sequencing combined and bioinformatics. In addition, gas chromatography-mass spectrometry combined with multivariate statistical analysis was utilized to identify marker candidates for prepared steaks. Furthermore, cloud platform analysis was applied to determine prepared beef steak spoilage, including the relationship between microbiological and physicochemical indicators and volatile compounds. RESULTS: The results showed that the dominant groups of Pseudomonas, Brochothrix thermosphacta, Lactobacillus and Lactococcus caused the spoilage of prepared beef steak, which are strongly associated with significant changes in physicochemical properties and volatile organic compounds (furan-2-pentyl-, pentanal, 1-octanol, 1-nonanol and dimethyl sulfide). Metabolic pathways were proposed, among which lipid metabolism and amino acid metabolism were most abundant. CONCLUSION: The present study is helpful with respect to further understanding the relationship between spoilage microorganisms and the quality of prepared beef steak, and provides a reference for investigating the spoilage mechanism of dominant spoilage bacteria and how to extend the shelf life of meat products. © 2024 Society of Chemical Industry.

Asunto(s)

Bacterias , Biología Computacional , Compuestos Orgánicos Volátiles , Bovinos , Animales , Compuestos Orgánicos Volátiles/química , Compuestos Orgánicos Volátiles/metabolismo , Compuestos Orgánicos Volátiles/análisis , Bacterias/genética , Bacterias/clasificación , Bacterias/aislamiento & purificación , Bacterias/metabolismo , Cromatografía de Gases y Espectrometría de Masas , Microbiología de Alimentos , Almacenamiento de Alimentos , Pseudomonas/crecimiento & desarrollo , Pseudomonas/metabolismo , Lactobacillus/metabolismo , Refrigeración , Brochothrix/metabolismo , Brochothrix/crecimiento & desarrollo , Brochothrix/aislamiento & purificación , Lactococcus , Carne Roja/microbiología , Carne Roja/análisis

RESUMEN

AIMS: This study aimed to prospect and isolate lactic acid bacteria (LAB) from an artisanal cheese production environment, to assess their safety, and to explore their bacteriocinogenic potential against Listeria monocytogenes. METHODS AND RESULTS: Samples were collected from surfaces of an artisanal-cheese production facility and after rep-PCR and 16S rRNA sequencing analysis, selected strains were identified as to be belonging to Lactococcus garvieae (1 strain) and Enterococcus faecium (14 isolates, grouped into three clusters) associated with different environments (worktables, cheese mold, ripening wooden shelves). All of them presented bacteriocinogenic potential against L. monocytogenes ATCC 7644 and were confirmed as safe (γ-hemolytic, not presenting antibiotic resistance, no mucus degradation properties, and no proteolytic or gelatinase enzyme activity). Additionally, cell growth, acidification and bacteriocins production kinetics, bacteriocin stability in relation to different temperatures, pH, and chemicals were evaluated. According to performed PCR analysis all studied strains generated positive evidence for the presence of entA and entP genes (for production of enterocins A and enterocins P, respectively). However, pediocin PA-1 associated gene was recorded only in DNA obtained from E. faecium ST02JL and Lc. garvieae ST04JL. CONCLUSIONS: It is worth considering the application of these safe LAB or their bacteriocins in situ as an alternative means of controlling L. monocytogenes in cheese production environments, either alone or in combination with other antimicrobials.

Asunto(s)

Bacteriocinas , Queso , Enterococcus faecium , Microbiología de Alimentos , Lactococcus , Listeria monocytogenes , Queso/microbiología , Enterococcus faecium/genética , Enterococcus faecium/aislamiento & purificación , Enterococcus faecium/metabolismo , Lactococcus/genética , Lactococcus/aislamiento & purificación , Bacteriocinas/farmacología , Brasil , Listeria monocytogenes/genética , Listeria monocytogenes/efectos de los fármacos , ARN Ribosómico 16S/genética , Antibacterianos/farmacología

RESUMEN

In this study, the feasibility of promoting the lactic acid (LA) fermentation of food waste (FW) with iron tailings (ITs) addition was explored. The best LA yield was 0.91 g LA/g total sugar when 1 % ITs were added into the system. The mechanisms for promoting LA production were acidification alleviation effects and reduction equivalent supply of ITs. Furthermore, the addition of ITs promoted carbohydrate hydrolysis, and the carbohydrates digestibility reached 88.85 % in the 1 % ITs group. The ITs also affected the microbial communities, Lactococcus gradually replaced Streptococcus as the dominant genus, and results suggested that Lactococcus had a positive correlation with LA production and carbohydrate digestibility. Finally, the complex LAB in FW had significant effects on heavy metal removal from ITs, and the removal efficiency Cr, As, Pb, Cd, and Hg can reach 50.84 %, 26.72 %, 59.65 %, 49.75 % and 78.87 % in the 1 % ITs group, respectively.

Asunto(s)

Fermentación , Hierro , Ácido Láctico , Hierro/metabolismo , Ácido Láctico/metabolismo , Ácido Láctico/biosíntesis , Metales Pesados , Alimentos , Residuos , Hidrólisis , Concentración de Iones de Hidrógeno , Lactococcus/metabolismo , Alimento Perdido y Desperdiciado

RESUMEN

Background: Lactococcus species are used to ferment milk to yogurt, cheese, and other products. The gram-positive coccus causes diseases in amphibia and fish and is a rare human pathogen. Patients and Methods: A 51-year-old male underwent laparoscopic cholecystectomy for acute and chronic calculous cholecystitis. Lactococcus lactis was isolated from pus from his gallbladder empyema. Results: Our institutional database was searched for other cases of Lactococcus spp. infections and four patients (2 males, 2 females; aged 51, 64, 78, and 80 years) were identified during a four-year period. The three other patients had positive blood cultures associated with pneumonia, toxic megacolon, and severe gastroenteritis. All isolates were monocultures with Lactococcus lactis (2), Lactococcus garvieae (1) and Lactococcus raffinolactis (1). Two patients died related to their sepsis. We report the second case of cholecystitis involving Lactococcus. Conclusions: Lactococcus is a very rare pathogen mainly causing blood stream infections but needs to be considered to cause serious surgical infections in humans.

Asunto(s)

Colecistitis Aguda , Infecciones por Bacterias Grampositivas , Lactococcus lactis , Lactococcus , Humanos , Masculino , Persona de Mediana Edad , Colecistectomía Laparoscópica , Colecistitis Aguda/microbiología , Colecistitis Aguda/cirugía , Infecciones por Bacterias Grampositivas/microbiología , Infecciones por Bacterias Grampositivas/diagnóstico , Lactococcus/aislamiento & purificación , Lactococcus lactis/aislamiento & purificación

RESUMEN

Aquaculturists use polyploid fish to maximize production albeit with some unintended consequences including compromised behaviors and physiological function. Given benefits of probiotic therapies (e.g., improved immune response, growth, and metabolism), we explored probiotic supplementation (mixture of Bifidobacterium, Lactobacillus, and Lactococcus), to overcome drawbacks. We first examined fish gut bacterial community composition using 16S metabarcoding (via principal coordinate analyses and PERMANOVA) and determined probiotics significantly impacted gut bacteria composition (p = 0.001). Secondly, we examined how a genomic disruptor (triploidy) and diet supplements (probiotics) impact gene transcription and behavioral profiles of hatchery-reared Chinook salmon (Oncorhynchus tshawytscha). Juveniles from four treatment groups (diploid-regular feed, diploid-probiotic feed, triploid-regular feed, and triploid-probiotic feed; n = 360) underwent behavioral assays to test activity, exploration, neophobia, predator evasion, aggression/sociality, behavioral sensitivity, and flexibility. In these fish, transcriptional profiles for genes associated with neural functions (neurogenesis/synaptic plasticity) and biomarkers for stress response and development (growth/appetite) were (i) examined across treatments and (ii) used to describe behavioral phenotypes via principal component analyses and general linear mixed models. Triploids exhibited a more active behavioral profile (p = 0.002), and those on a regular diet had greater Neuropeptide Y transcription (p = 0.02). A growth gene (early growth response protein 1, p = 0.02) and long-term neural development genes (neurogenic differentiation factor, p = 0.003 and synaptysomal-associated protein 25-a, p = 0.005) impacted activity and reactionary profiles, respectively. Overall, our probiotic treatment did not compensate for triploidy. Our research highlights novel applications of behavioral transcriptomics for identifying candidate genes and dynamic, mechanistic associations with complex behavioral repertoires.

Asunto(s)

Microbioma Gastrointestinal , Lactococcus , Probióticos , Salmón , Transcriptoma , Triploidía , Animales , Probióticos/farmacología , Probióticos/administración & dosificación , Salmón/genética , Salmón/microbiología , Lactococcus/genética , Lactobacillus/genética , Conducta Animal/efectos de los fármacos

RESUMEN

Hyperuricemia (HUA) is a widespread metabolic disorder. Probiotics have drawn increasing attention as an adjunctive treatment with fewer side effects. However, thus far the effective strains are limited and the mechanisms for their serum uric acid (SUA)-lowering effect are not well understood. Along this line, we conducted the current study using a hyperuricemia mouse model induced by potassium oxonate and adenine. A novel strain of Lactococcus cremoris named D2022 was identified to have significant SUA-lowering capability. Lactococcus cremoris D2022 significantly reduced SUA levels by inhibiting uric acid synthesis and regulating uric acid transportation. It was also found that Lactococcus cremoris D2022 alleviated HUA-induced renal inflammatory injury involving multiple signaling pathways. By focusing on the expression of NLRP3-related inflammatory genes, we found correlations between the expression levels of these genes and free fatty acid receptors (FFARs). In addition, oral administration of Lactococcus cremoris D2022 increased short-chain fatty acids (SCFAs) in cecal samples, which may be one of the mechanisms by which oral probiotics alleviate renal inflammation. Serum untargeted metabolomics showed changes in a variety of serum metabolites associated with purine metabolism and inflammation after oral administration of Lactococcus cremoris D2022, further confirming its systemic bioactivity. Finally, it was proved that Lactococcus cremoris D2022 improved intestinal barrier function. In conclusion, Lactococcus cremoris D2022 can alleviate HUA and HUA-induced nephropathy by increasing the production of SCFAs in the gut and systemic metabolism.

Asunto(s)

Hiperuricemia , Riñón , Lactococcus , Probióticos , Ácido Úrico , Animales , Hiperuricemia/tratamiento farmacológico , Ratones , Probióticos/farmacología , Probióticos/administración & dosificación , Masculino , Ácido Úrico/sangre , Riñón/metabolismo , Microbioma Gastrointestinal/efectos de los fármacos , Inflamación , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL

RESUMEN

The diet during pregnancy, or antenatal diet, influences the offspring's intestinal health. We previously showed that antenatal butyrate supplementation reduces injury in adult murine offspring with dextran sulfate sodium (DSS)-induced colitis. Potential modulators of butyrate levels in the intestine include a high fiber diet or dietary supplementation with probiotics. To test this, we supplemented the diet of pregnant mice with high fiber, or with the probiotic bacteria Lactococcus lactis subspecies cremoris or Lactobacillus rhamnosus GG. We then induced chronic colitis with DSS in their adult offspring. We demonstrate that a high fiber antenatal diet, or supplementation with Lactococcus lactis subspecies cremoris during pregnancy diminished the injury from DSS-induced colitis in offspring. These data are evidence that antenatal dietary interventions impact offspring gut health and define the antenatal diet as a therapeutic modality to enhance offspring intestinal health.

Asunto(s)

Colitis , Microbioma Gastrointestinal , Lactococcus lactis , Lactococcus , Femenino , Embarazo , Animales , Ratones , Lactococcus lactis/genética , Suplementos Dietéticos , Butiratos

RESUMEN

Lactic Acid Bacteria (LAB) are predominantly probiotic microorganisms and the most are Generally Recognized As Safe (GRAS). LAB inhabit in the human gut ecosystem and are largely found in fermented foods and silage. In the last decades, LAB have also has been found in plant microbiota as a new class of microbes with probiotic activity to plants. For this reason, today the scientific interest in the study and isolation of LAB for agronomic application has increased. However, isolation protocols from complex samples such as plant tissues are scarce and inefficient. In this study, we developed a new protocol (CLI, Complex samples LAB Isolation) which yields purified LAB from plants. The sensitivity of CLI protocol was sufficient to isolate representative microorganisms of LAB genera (i.e. Leuconostoc, Lactococcus and Enterococcus). CLI protocol consists on five steps: i) sample preparation and pre-incubation in 1% sterile peptone at 30 °C for 24-48 h; ii) Sample homogenization in vortex by 10 min; iii) sample serial dilution in quarter-strength Ringer solution, iv) incubation in MRS agar plates with 0.2% of sorbic acid, with 1% of CaCO3, O2 < 15%, at pH 5.8 and 37 °C for 48 h.; v) Selection of single colonies with LAB morphology and CaCO3-solubilization halo. Our scientific contribution is that CLI protocol could be used for several complex samples and represents a useful method for further studies involving native LAB.

Asunto(s)

Lactobacillales , Lactobacillales/aislamiento & purificación , Lactobacillales/clasificación , Plantas/microbiología , Leuconostoc/aislamiento & purificación , Probióticos/aislamiento & purificación , Lactococcus/aislamiento & purificación , Enterococcus/aislamiento & purificación , Ácido Láctico/metabolismo

RESUMEN

Bacterial reverse transcriptases (RTs) are a large and diverse enzyme family. AbiA, AbiK and Abi-P2 are abortive infection system (Abi) RTs that mediate defense against bacteriophages. What sets Abi RTs apart from other RT enzymes is their ability to synthesize long DNA products of random sequences in a template- and primer-independent manner. Structures of AbiK and Abi-P2 representatives have recently been determined, but there are no structural data available for AbiA. Here, we report the crystal structure of Lactococcus AbiA polymerase in complex with a single-stranded polymerization product. AbiA comprises three domains: an RT-like domain, a helical domain that is typical for Abi polymerases, and a higher eukaryotes and prokaryotes nucleotide-binding (HEPN) domain that is common for many antiviral proteins. AbiA forms a dimer that distinguishes it from AbiK and Abi-P2, which form trimers/hexamers. We show the DNA polymerase activity of AbiA in an in vitro assay and demonstrate that it requires the presence of the HEPN domain which is enzymatically inactive. We validate our biochemical and structural results in vivo through bacteriophage infection assays. Finally, our in vivo results suggest that AbiA-mediated phage defense may not rely on AbiA-mediated cell death.

Asunto(s)

Bacteriófagos , Lactococcus , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Bacteriófagos/genética , Cristalografía por Rayos X , Lactococcus/virología , Lactococcus/genética , Modelos Moleculares , Dominios Proteicos , Multimerización de Proteína , ADN Polimerasa Dirigida por ARN/metabolismo , ADN Polimerasa Dirigida por ARN/química , ADN Polimerasa Dirigida por ARN/genética , Relación Estructura-Actividad

RESUMEN

Piscine lactococcosis is a significant threat to cultured and wild fish populations worldwide. The disease typically presents as a per-acute to acute hemorrhagic septicemia causing high morbidity and mortality, recalcitrant to antimicrobial treatment or management interventions. Historically, the disease was attributed to the gram-positive pathogen Lactococcus garvieae. However, recent work has revealed three distinct lactococcosis-causing bacteria (LCB)-L. garvieae, L. petauri, and L. formosensis-which are phenotypically and genetically similar, leading to widespread misidentification. An update on our understanding of lactococcosis and improved methods for identification are urgently needed. To this end, we used representative isolates from each of the three LCB species to compare currently available and recently developed molecular and phenotypic typing assays, including whole-genome sequencing (WGS), end-point and quantitative PCR (qPCR) assays, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), API 20 Strep and Biolog systems, fatty acid methyl ester analysis (FAME), and Sensititre antimicrobial profiling. Apart from WGS, sequencing of the gyrB gene was the only method capable of consistent and accurate identification to the species and strain level. A qPCR assay based on a putative glycosyltransferase gene was also able to distinguish L. petauri from L. garvieae/formosensis. Biochemical tests and MALDI-TOF MS showed some species-specific patterns in sugar and fatty acid metabolism or protein profiles but should be complemented by additional analyses. The LCB demonstrated overlap in host and geographic range, but there were relevant differences in host specificity, regional prevalence, and antimicrobial susceptibility impacting disease treatment and prevention. IMPORTANCE: Lactococcosis affects a broad range of host species, including fish from cold, temperate, and warm freshwater or marine environments, as well as several terrestrial animals, including humans. As such, lactococcosis is a disease of concern for animal and ecosystem health. The disease is endemic in European and Asian aquaculture but is rapidly encroaching on ecologically and economically important fish populations across the Americas. Piscine lactococcosis is difficult to manage, with issues of vaccine escape, ineffective antimicrobial treatment, and the development of carrier fish or biofilms leading to recurrent outbreaks. Our understanding of the disease is also widely outdated. The accepted etiologic agent of lactococcosis is Lactococcus garvieae. However, historical misidentification has masked contributions from two additional species, L. petauri and L. formosensis, which are indistinguishable from L. garvieae by common diagnostic methods. This work is the first comprehensive characterization of all three agents and provides direct recommendations for species-specific diagnosis and management.

Asunto(s)

Enfermedades de los Peces , Infecciones por Bacterias Grampositivas , Lactococcus , Lactococcus/genética , Lactococcus/aislamiento & purificación , Lactococcus/clasificación , Animales , Enfermedades de los Peces/microbiología , Infecciones por Bacterias Grampositivas/microbiología , Infecciones por Bacterias Grampositivas/veterinaria , Peces/microbiología , Secuenciación Completa del Genoma , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción