Deciphering the metabolism of Lactobacillus delbrueckii subsp. delbrueckii during soy juice fermentation using phenotypic and transcriptional analysis.

In the context of sustainable diet, the development of soy-based yogurt fermented with lactic acid bacteria is an attractive alternative to dairy yogurts. To decipher the metabolism of Lactobacillus delbrueckii subsp. delbrueckii during soy juice (SJ) fermentation, the whole genome of the strain CIRM-BIA865 (Ld865) was sequenced and annotated. Then Ld865 was used to ferment SJ. Samples were analyzed throughout fermentation for their cell number, carbohydrate, organic acid, free amino acid, and volatile compound contents. Despite acidification, the number of Ld865 cells did not rise, and microscopic observations revealed the elongation of cells from 3.6 µm (inoculation) to 36.9 µm (end of fermentation). This elongation was observed in SJ but not in laboratory-rich medium MRS. Using transcriptomic analysis, we showed that the biosynthesis genes of peptidoglycan and membrane lipids were stably expressed, in line with the cell elongation observed, whereas no genes implicated in cell division were upregulated. Among the main sugars available in SJ (sucrose, raffinose, and stachyose), Ld865 only used sucrose. The transcriptomic analysis showed that Ld865 implemented the two transport systems that it contains to import sucrose: a PTS system and an ABC transporter. To fulfill its nitrogen needs, Ld865 probably first consumed the free amino acids of the SJ and then implemented different oligopeptide transporters and proteolytic/peptidase enzymes. In conclusion, this study showed that Ld865 enables fast acidification of SJ, despite the absence of cell division, leads to a product rich in free amino acids, and also leads to the production of aromatic compounds of interest. IMPORTANCE: To reduce the environmental and health concerns related to food, an alternative diet is recommended, containing 50% of plant-based proteins. Soy juice, which is protein rich, is a relevant alternative to animal milk, for the production of yogurt-like products. However, soy "beany" and "green" off-flavors limit the consumption of such products. The lactic acid bacteria (LAB) used for fermentation can help to improve the organoleptic properties of soy products. But metabolic data concerning LAB adapted to soy juice are lacking. The aim of this study was, thus, to decipher the metabolism of Lactobacillus delbrueckii subsp. delbrueckii during fermentation of a soy juice, based on a multidisciplinary approach. This result will contribute to give tracks for a relevant selection of starter. Indeed, the improvement of the organoleptic properties of these types of products could help to promote plant-based proteins in our diet.

Positive Interactions between Lactic Acid Bacteria Promoted by Nitrogen-Based Nutritional Dependencies.

Nutritional dependencies, especially those regarding nitrogen sources, govern numerous microbial positive interactions. As for lactic acid bacteria (LAB), responsible for the sanitary, organoleptic, and health properties of most fermented products, such positive interactions have previously been studied between yogurt bacteria. However, they have never been exploited to create artificial cocultures of LAB that would not necessarily coexist naturally, i.e., from different origins. The objective of this study was to promote LAB positive interactions, based on nitrogen dependencies in cocultures, and to investigate how these interactions affect some functional outputs, e.g., acidification rates, carbohydrate consumption, and volatile-compound production. The strategy was to exploit both proteolytic activities and amino acid auxotrophies of LAB. A chemically defined medium was thus developed to specifically allow the growth of six strains used, three proteolytic and three nonproteolytic. Each of the proteolytic strains, Enterococcus faecalis CIRM-BIA2412, Lactococcus lactis NCDO2125, and CIRM-BIA244, was cocultured with each one of the nonproteolytic LAB strains, L. lactis NCDO2111 and Lactiplantibacillus plantarum CIRM-BIA465 and CIRM-BIA1524. Bacterial growth was monitored using compartmented chambers to compare growth in mono- and cocultures. Acidification, carbohydrate consumption, and volatile-compound production were evaluated in direct cocultures. Each proteolytic strain induced different types of interactions: strongly positive interactions, weakly positive interactions, and no interactions were seen with E. faecalis CIRM-BIA2412, L. lactis NCDO2125, and L. lactis CIRM-BIA244, respectively. Strong interactions were associated with higher concentrations of tryptophan, valine, phenylalanine, leucine, isoleucine, and peptides. They led to higher acidification rates, lower pH, higher raffinose utilization, and higher concentrations of five volatile compounds. IMPORTANCE Interactions of lactic acid bacteria (LAB) are often studied in association with yeasts or propionibacteria in various fermented food products, and the mechanisms underlying their interactions are being quite well characterized. Concerning interactions between LAB, they have mainly been investigated to test antagonistic interactions. Understanding how they can positively interact could be useful in multiple food-related fields: production of fermented food products with enhanced functional properties or fermentation of new food matrices. This study investigated the exploitation of the proteolytic activity of LAB strains to promote positive interactions between proteolytic and nonproteolytic strains. The results suggest that proteolytic LAB do not equally stimulate nonproteolytic LAB and that the stronger the interactions between LAB are, the more functional outputs we can expect. Thus, this study gives insight into how to create new associations of LAB strains and to guarantee their positive interactions.

Fitness and fur colouration: Testing the camouflage and thermoregulation hypotheses in an Arctic mammal.

Selection for crypsis has been recognized as an important ecological driver of animal colouration, whereas the relative importance of thermoregulation is more contentious with mixed empirical support. A potential thermal advantage of darker individuals has been observed in a wide range of animal species. Arctic animals that exhibit colour polymorphisms and undergo seasonal colour moults are interesting study subjects for testing the two alternative hypotheses: demographic performance of different colour morphs might be differentially affected by snow cover with a cryptic advantage for lighter morphs, or conversely by winter temperature with a thermal advantage for darker morphs. In this study, we explored whether camouflage and thermoregulation might explain differences in reproduction and survival between the white and blue colour morphs of the Arctic fox Vulpes lagopus under natural conditions. Juvenile and adult survival, breeding propensity and litter size were measured for 798 captive-bred and released or wild-born Arctic foxes monitored during an 11-year period (2007-2017) in two subpopulations in south-central Norway. We investigated the proportion of the two colour morphs and compared their demographic performance in relation to spatial variation in duration of snow cover, onset of snow season and winter temperatures. After population re-establishment, a higher proportion of blue individuals was observed among wild-born Arctic foxes compared to the proportion of blue foxes released from the captive population. Our field study provides the first evidence for an effect of colour morph on the reproductive performance of Arctic foxes under natural conditions, with a higher breeding propensity of the blue morph compared to the white one. Performance of the two colour morphs was not differentially affected by the climatic variables, except for juvenile survival. Blue morph juveniles showed a tendency for higher survival under colder winter temperatures but lower survival under warmer temperatures compared to white morph juveniles. Overall, our findings do not consistently support predictions of the camouflage or the thermoregulation hypotheses. The higher success of blue foxes suggests an advantage of the dark morph not directly related to disruptive selection by crypsis or thermoregulation. Our results rather point to physiological adaptations and behavioural traits not necessarily connected to thermoregulation, such as stress response, immune function, sexual behaviour and aggressiveness. Our findings highlight the need to explore the potential role of genetic linkage or pleiotropy in influencing the fitness of white and blue Arctic foxes as well as other species with colour polymorphisms.

Independent domestication events in the blue-cheese fungus Penicillium roqueforti.

Domestication provides an excellent framework for studying adaptive divergence. Using population genomics and phenotypic assays, we reconstructed the domestication history of the blue cheese mould Penicillium roqueforti. We showed that this fungus was domesticated twice independently. The population used in Roquefort originated from an old domestication event associated with weak bottlenecks and exhibited traits beneficial for pre-industrial cheese production (slower growth in cheese and greater spore production on bread, the traditional multiplication medium). The other cheese population originated more recently from the selection of a single clonal lineage, was associated with all types of blue cheese worldwide except Roquefort, and displayed phenotypes more suited for industrial cheese production (high lipolytic activity, efficient cheese cavity colonization ability and salt tolerance). We detected genomic regions affected by recent positive selection and putative horizontal gene transfers. This study sheds light on the processes of rapid adaptation and raises questions about genetic resource conservation.

Identification of the bacteria and their metabolic activities associated with the microbial spoilage of custard cream desserts.

The famous French dessert "ile flottante" consists of a sweet egg white foam floating on a vanilla custard cream, which contains highly nutritive raw materials, including milk, sugar and egg. Spoilage issues are therefore a key concern for the manufacturers. This study explored the bacterial diversity of 64 spoiled custard cream desserts manufactured by 2 French companies. B. cereus group bacteria, coagulase negative Staphylococcus, Enterococcus and Leuconostoc spp. were isolated from spoiled products. Thirty-one bacterial isolates representative of the main spoilage species were tested for their spoilage abilities. Significant growth and pH decrease were observed regardless of species. While off-odours were detected with B. cereus group and staphylococci, yoghurt odours were detected with Enterococcus spp. and Leuconostoc spp. B. cereus group bacteria produced various esters and several compounds derived from amino acid and sugar metabolism. Most Staphylococci produced phenolic compounds. Enterococcus spp. and Leuconostoc spp. isolates produced high levels of compounds derived from sugar metabolism. Each type of spoilage bacteria was associated with a specific volatile profile and lactic acid was identified as a potential marker of spoilage of custard cream-based desserts. These findings provide valuable information for manufacturers to improve food spoilage detection and prevention of chilled desserts made with milk and egg.

Diversity of the metabolic profiles of a broad range of lactic acid bacteria in soy juice fermentation.

This study explores the ability of lactic acid bacteria (LAB) to ferment soy juice. The ability of 276 LAB strains from 25 species to ferment the principal soy carbohydrates, sucrose, raffinose or stachyose was tested in synthetic media and a soy juice. Fermented soy juices (FSJs) were characterized for their odor. Selected FSJs were characterized by targeted metabolomics. All Streptococcus, 83% of Leuconostoc and Lactobacillus and 41% of Lactococcus strains were sucrose-positive, while only 36% of all the LAB strains tested were raffinose-positive and 6% stachyose-positive. Nearly all (97%) the sucrose-positive strains fermented soy juice, indicating that an ability to use sucrose is a good criterion to select strains for soy juice fermentation. Among the most efficient acidifying strains, 46 FSJs had an odor deemed to be acceptable. FSJ composition was dependent on both species and strains: 17/46 strains deglycosylated soy juice isoflavones, the 27 S. thermophilus strains converted a mean 4.4 ± 0.1 g/L of sucrose into 3.0 ± 0.1 g/L of lactic acid versus 5.2 ± 0.1 g/L into 2.2 ± 0.1 g/L for the 18 Lactobacillus and one Lactococcus strains. This study highlights the diversity of the metabolic profiles of LAB strains in soy juice fermentation.

The Candidate Blood-stage Malaria Vaccine P27A Induces a Robust Humoral Response in a Fast Track to the Field Phase 1 Trial in Exposed and Nonexposed Volunteers.

Background: P27A is an unstructured 104mer synthetic peptide from Plasmodium falciparum trophozoite exported protein 1 (TEX1), the target of human antibodies inhibiting parasite growth. The present project aimed at evaluating the safety and immunogenicity of P27A peptide vaccine in malaria-nonexposed European and malaria-exposed African adults. Methods: This study was designed as a staggered, fast-track, randomized, antigen and adjuvant dose-finding, multicenter phase 1a/1b trial, conducted in Switzerland and Tanzania. P27A antigen (10 or 50 µg), adjuvanted with Alhydrogel or glucopyranosil lipid adjuvant stable emulsion (GLA-SE; 2.5 or 5 µg), or control rabies vaccine (Verorab) were administered intramuscularly to 16 malaria-nonexposed and 40 malaria-exposed subjects on days 0, 28, and 56. Local and systemic adverse events (AEs) as well as humoral and cellular immune responses were assessed after each injection and during the 34-week follow-up. Results: Most AEs were mild to moderate and resolved completely within 48 hours. Systemic AEs were more frequent in the formulation with alum as compared to GLA-SE, whereas local AEs were more frequent after GLA-SE. No serious AEs occurred. Supported by a mixed Th1/Th2 cell-mediated immunity, P27A induced a marked specific antibody response able to recognize TEX1 in infected erythrocytes and to inhibit parasite growth through an antibody-dependent cellular inhibition mechanism. Incidence of AEs and antibody responses were significantly lower in malaria-exposed Tanzanian subjects than in nonexposed European subjects. Conclusions: The candidate vaccine P27A was safe and induced a particularly robust immunogenic response in combination with GLA-SE. This formulation should be considered for future efficacy trials. Clinical Trials Registration: NCT01949909, PACTR201310000683408.

Development of antifungal ingredients for dairy products: From in vitro screening to pilot scale application.

Biopreservation represents a complementary approach to traditional hurdle technologies for reducing microbial contaminants (pathogens and spoilers) in food. In the dairy industry that is concerned by fungal spoilage, biopreservation can also be an alternative to preservatives currently used (e.g. natamycin, potassium sorbate). The aim of this study was to develop antifungal fermentates derived from two dairy substrates using a sequential approach including an in vitro screening followed by an in situ validation. The in vitro screening of the antifungal activity of fermentates derivating from 430 lactic acid bacteria (LAB) (23 species), 70 propionibacteria (4 species) and 198 fungi (87 species) was performed against four major spoilage fungi (Penicillium commune, Mucor racemosus, Galactomyces geotrichum and Yarrowia lipolytica) using a cheese-mimicking model. The most active fermentates were obtained from Lactobacillus brevis, Lactobacillus buchneri, Lactobacillus casei/paracasei and Lactobacillus plantarum among the tested LAB, Propionibacterium jensenii among propionibacteria, and Mucor lanceolatus among the tested fungi. Then, for the 11 most active fermentates, culture conditions were optimized by varying incubation time and temperature in order to enhance their antifungal activity. Finally, the antifungal activity of 3 fermentates of interest obtained from Lactobacillus rhamnosus CIRM-BIA1952, P. jensenii CIRM-BIA1774 and M. lanceolatus UBOCC-A-109193 were evaluated in real dairy products (sour cream and semi-hard cheese) at a pilot-scale using challenge and durability tests. In parallel, the impact of these ingredients on organoleptic properties of the obtained products was also assessed. In semi-hard cheese, application of the selected fermentates on the cheese surface delayed the growth of spoilage molds for up to 21 days, without any effect on organoleptic properties, P. jensenii CIRM-BIA1774 fermentate being the most active. In sour cream, incorporation of the latter fermentate at 2 or 5% yielded a high antifungal activity but was detrimental to the product organoleptic properties. Determination of the concentration limit, compatible with product acceptability, showed that incorporation of this fermentate at 0.4% prevented growth of fungal contaminants in durability tests but had a more limited effect against M. racemosus and P. commune in challenge tests. To our knowledge, this is the first time that the workflow followed in this study, from in vitro screening using dairy matrix to scale-up in cheese and sour cream, is applied for production of natural ingredients relying on a large microbial diversity in terms of species and strains. This approach allowed obtaining several antifungal fermentates which are promising candidates for dairy products biopreservation.

Mesencephalic dopamine neurons interfacing the shell of nucleus accumbens and the dorsolateral striatum in the rat.

Parallel corticostriatonigral circuits have been proposed that separately process motor, cognitive, and emotional-motivational information. Functional integration requires that interactions exist between neurons participating in these circuits. This makes it imperative to study the complex anatomical substrate underlying corticostriatonigral circuits. It has previously been proposed that dopaminergic neurons in the ventral mesencephalon may play a role in this circuit interaction. Therefore, we studied in rats convergence of basal ganglia circuits by depositing an anterograde neuroanatomical tracer into the ventral striatum together with a retrograde fluorescent tracer ipsilaterally in the dorsolateral striatum. In the mesencephalon, using confocal microscopy, we looked for possible appositions of anterogradely labeled fibers and retrogradely labeled neurons, "enhancing" the latter via intracellular injection of Lucifer Yellow. Tyrosine hydroxylase (TH) immunofluorescence served to identify dopaminergic neurons. In neurophysiological experiments, we combined orthodromic stimulation in the medial ventral striatum with recording from ventral mesencephalic neurons characterized by antidromic stimulation from the dorsal striatum. We observed terminal fields of anterogradely labeled fibers that overlap populations of retrogradely labeled nigrostriatal cell bodies in the substantia nigra pars compacta and lateral ventral tegmental area (VTA), with numerous close appositions between boutons of anterogradely labeled fibers and nigrostriatal, TH-immunopositive neurons. Neurophysiological stimulation in the medial ventral striatum caused inhibition of dopaminergic nigrostriatal neurons projecting to the ventrolateral striatal territory. Responding nigrostriatal neurons were located in the medial substantia nigra and adjacent VTA. Our results strongly suggest a functional link between ventromedial, emotional-motivational striatum, and the sensorimotor dorsal striatum via dopaminergic nigrostriatal neurons.

Strains of the Lactobacillus casei group show diverse abilities for the production of flavor compounds in 2 model systems.

Cheese flavor development is directly connected to the metabolic activity of microorganisms used during its manufacture, and the selection of metabolically diverse strains represents a potential tool for the production of cheese with novel and distinct flavor characteristics. Strains of Lactobacillus have been proven to promote the development of important cheese flavor compounds. As cheese production and ripening are long-lasting and expensive, model systems have been developed with the purpose of rapidly screening lactic acid bacteria for their flavor potential. The biodiversity of 10 strains of the Lactobacillus casei group was evaluated in 2 model systems and their volatile profiles were determined by gas chromatography-mass spectrometry. In model system 1, which represented a mixture of free AA, inoculated cells did not grow. In total, 66 compounds considered as flavor contributors were successfully identified, most of which were aldehydes, acids, and alcohols produced via AA metabolism by selected strains. Three strains (DPC2071, DPC3990, and DPC4206) had the most diverse metabolic capacities in model system 1. In model system 2, which was based on processed cheese curd, inoculated cells increased in numbers over incubation time. A total of 47 compounds were identified, and they originated not only from proteolysis, but also from glycolytic and lipolytic processes. Tested strains produced ketones, acids, and esters. Although strains produced different abundances of volatiles, diversity was less evident in model system 2, and only one strain (DPC4206) was distinguished from the others. Strains identified as the most dissimilar in both of the model systems could be more useful for cheese flavor diversification.

Adaptation of Propionibacterium freudenreichii to long-term survival under gradual nutritional shortage.

BACKGROUND: Propionibacterium freudenreichii is an Actinobacterium widely used in the dairy industry as a ripening culture for Swiss-type cheeses, for vitamin B12 production and some strains display probiotic properties. It is reportedly a hardy bacterium, able to survive the cheese-making process and digestive stresses. RESULTS: During this study, P. freudenreichii CIRM-BIA 138 (alias ITG P9), which has a generation time of five hours in Yeast Extract Lactate medium at 30 °C under microaerophilic conditions, was incubated for 11 days (9 days after entry into stationary phase) in a culture medium, without any adjunct during the incubation. The carbon and free amino acids sources available in the medium, and the organic acids produced by the strain, were monitored throughout growth and survival. Although lactate (the preferred carbon source for P. freudenreichii) was exhausted three days after inoculation, the strain sustained a high population level of 9.3 log10 CFU/mL. Its physiological adaptation was investigated by RNA-seq analysis and revealed a complete disruption of metabolism at the entry into stationary phase as compared to exponential phase. CONCLUSIONS: P. freudenreichii adapts its metabolism during entry into stationary phase by down-regulating oxidative phosphorylation, glycolysis, and the Wood-Werkman cycle by exploiting new nitrogen (glutamate, glycine, alanine) sources, by down-regulating the transcription, translation and secretion of protein. Utilization of polyphosphates was suggested.

QUIRKY interacts with STRUBBELIG and PAL OF QUIRKY to regulate cell growth anisotropy during Arabidopsis gynoecium development.

Organ morphogenesis largely relies on cell division and elongation, which need to be both coordinated between cells and orchestrated with cytoskeleton dynamics. However, components that bridge the biological signals and the effectors that define cell shape remain poorly described. We have addressed this issue through the functional characterisation of QUIRKY (QKY), previously isolated as being involved in the STRUBBELIG (SUB) genetic pathway that controls cell-cell communication and organ morphogenesis in Arabidopsis. QKY encodes a protein containing multiple C2 domains and transmembrane regions, and SUB encodes an atypical LRR-receptor-like kinase. We show that twisting of the gynoecium observed in qky results from the abnormal division pattern and anisotropic growth of clustered cells arranged sporadically along the gynoecium. Moreover, the cortical microtubule (CMT) network of these cells is disorganised. A cross to botero, a katanin mutant in which the normal orientation of CMTs and anisotropic cell expansion are impaired, strongly reduces silique deviation, reinforcing the hypothesis of a role for QKY in CMT-mediated cell growth anisotropy. We also show that QKY is localised at the plasma membrane and functions in a multiprotein complex that includes SUB and PAL OF QUIRKY (POQ), a previously uncharacterised PB1-domain-containing protein that localises both at the plasma membrane and in intracellular compartments. Our data indicate that QKY and its interactors play central roles linking together cell-cell communication and cellular growth.

Spatial Distribution of Lactococcus lactis Colonies Modulates the Production of Major Metabolites during the Ripening of a Model Cheese.

In cheese, lactic acid bacteria are immobilized at the coagulation step and grow as colonies. The spatial distribution of bacterial colonies is characterized by the size and number of colonies for a given bacterial population within cheese. Our objective was to demonstrate that different spatial distributions, which lead to differences in the exchange surface between the colonies and the cheese matrix, can influence the ripening process. The strategy was to generate cheeses with the same growth and acidification of a Lactococcus lactis strain with two different spatial distributions, big and small colonies, to monitor the production of the major ripening metabolites, including sugars, organic acids, peptides, free amino acids, and volatile metabolites, over 1 month of ripening. The monitored metabolites were qualitatively the same for both cheeses, but many of them were more abundant in the small-colony cheeses than in the big-colony cheeses over 1 month of ripening. Therefore, the results obtained showed that two different spatial distributions of L. lactis modulated the ripening time course by generating moderate but significant differences in the rates of production or consumption for many of the metabolites commonly monitored throughout ripening. The present work further explores the immobilization of bacteria as colonies within cheese and highlights the consequences of this immobilization on cheese ripening.

Lactobacillus and Leuconostoc volatilomes in cheese conditions.

New strains are desirable to diversify flavour of fermented dairy products. The objective of this study was to evaluate the potential of Leuconostoc spp. and Lactobacillus spp. in the production of aroma compounds by metabolic fingerprints of volatiles. Eighteen strains, including five Lactobacillus species (Lactobacillus fermentum, Lactobacillus helveticus, Lactobacillus paracasei, Lactobacillus rhamnosus, Lactobacillus sakei) and three Leuconostoc species (Leuconostoc citreum, Leuconostoc lactis, and Leuconostoc mesenteroides) were incubated for 5 weeks in a curd-based slurry medium under conditions mimicking cheese ripening. Populations were enumerated and volatile compounds were analysed by headspace trap gas chromatography-mass spectrometry (GC-MS). A metabolomics approach followed by multivariate statistical analysis was applied for data processing and analysis. In total, 12 alcohols, 10 aldehydes, 7 esters, 11 ketones, 5 acids and 2 sulphur compounds were identified. Very large differences in concentration of volatile compounds between the highest producing strains and the control medium were observed in particular for diacetyl, 2-butanol, ethyl acetate, 3-methylbutanol, 3-methylbutanoic acid and 2-methylbutanoic acid. Some of the characterized strains demonstrated an interesting aromatizing potential to be used as adjunct culture.

Diagnostic Accuracy of Emergency Physician-Performed Ultrasound for Acute Appendicitis in a Remote Location.

BACKGROUND: Preoperative imaging for suspected acute appendicitis (AA), such as ultrasonography (US), was shown to improve diagnostic accuracy and patient outcomes. Criteria for diagnosis of AA by US are well established and reliable. In previous studies, US assessments were always performed by skilled radiologist physicians. However, a radiologist and computed tomography scanning equipment are not always available in the community hospitals or remote sites of developing countries. OBJECTIVE: Our aim was to assess a diagnostic pathway using clinical evaluation, routine US performed by an emergency physician, and clinical re-evaluation for patients suspected of having AA. METHODS: Patients suspected of having AA admitted to the emergency department in a developing country were prospectively enrolled between November 2010 and January 2011. Clinical and US data were studied. A noncompressible appendix with a diameter ≥6 mm was the main US diagnosis criterion. RESULTS: Among the 104 included patients, surgery was performed on 28. Of the 25 patients with positive US, 22 actually had AA, matching the surgical report. The remaining 76 patients without US appendicitis criteria underwent clinical follow-up and had medical conditions. Sensitivity of US was 88%, specificity was 96%, positive predictive value was 88%, and negative predictive value was 96%. The likelihood ratios for our US assessment highlight the need for a test with enhanced diagnostic accuracy. CONCLUSIONS: A diagnostic strategy using clinical evaluations, routine US performed by emergency physicians, and clinical re-evaluation of patients with acute abdominal pain is appropriate to provide positive results for the diagnosis and treatment of appendicitis in remote locations.

SQUINT promotes stem cell homeostasis and floral meristem termination in Arabidopsis through APETALA2 and CLAVATA signalling.

Plant meristems harbour stem cells, which allow for the continuous production of new organs. Here, an analysis of the role of SQUINT (SQN) in stem cell dynamics in Arabidopsis is reported. A close examination of sqn mutants reveals defects that are very similar to that of weak clavata (clv) mutants, both in the flower meristem (increased number of floral organs, occasional delay in stem cell termination) and in the shoot apical meristem (meristem and central zone enlargement, occasional fasciation). sqn has a very mild effect in a clv mutant background, suggesting that SQN and the CLV genes act in the same genetic pathway. Accordingly, a loss-of-function allele of SQN strongly rescues the meristem abortion phenotype of plants that overexpress CLV3. Altogether, these data suggest that SQN is necessary for proper CLV signalling. SQN was shown to be required for normal accumulation of various miRNAs, including miR172. One of the targets of miR172, APETALA2 (AP2), antagonizes CLV signalling. The ap2-2 mutation strongly suppresses the meristem phenotypes of sqn, indicating that the effect of SQN on stem cell dynamics is largely, but not fully, mediated by the miR172/AP2 tandem. This study refines understanding of the intricate genetic networks that control both stem cell homeostasis and floral stem cell termination, two processes that are critical for the proper development and fertility of the plant.

Relationships between isotopic values and oxidative status: insights from populations of gentoo penguins.

Feeding strategies can affect the balance between the production of reactive oxygen species and antioxidant defences (i.e. oxidative status). This is ecologically relevant, as variation in oxidative status can in turn strongly affect fitness. However, how animals regulate their oxidative status through their feeding behaviour under natural conditions remains poorly understood. Thus, relating the isotopic values of free-ranging animals to their oxidative status may prove useful. Here, we considered three colonies of gentoo penguins (Pygoscelis papua) in which we measured (1) δ(13)C and δ(15)N values, and (2) antioxidant defences and oxidative damage. We found that colonies with the highest δ(13)C and δ(15)N values also had the highest levels of antioxidant defences and oxidative damage, resulting in positive relationships between isotopic values and markers of oxidative status. As a result, colony segregation in terms of isotopic values was reflected by segregation in terms of oxidative markers (although more markedly for oxidative damage than for antioxidant defences). Interestingly, variation in the estimated contribution of krill in the diet of penguins followed an opposite pattern to that observed for markers of oxidative status, providing evidence that inter-population differences in terms of foraging strategies can result in inter-population differences in terms of oxidative status. More studies examining simultaneously oxidative status, isotopic signature, foraging behaviour and food allocation between parents and young are, however, needed to understand better the interplay between the foraging strategies adopted by animals in their natural habitat and their oxidative status.

A methodological approach to screen diverse cheese-related bacteria for their ability to produce aroma compounds.

Microorganisms play an important role in the development of cheese flavor. The aim of this study was to develop an approach to facilitate screening of various cheese-related bacteria for their ability to produce aroma compounds. We combined i) curd-based slurry medium incubated under conditions mimicking cheese manufacturing and ripening, ii) powerful method of extraction of volatiles, headspace trap, coupled to gas chromatography-mass spectrometry (HS-trap-GC-MS), and iii) metabolomics-based method of data processing using the XCMS package of R software and multivariate analysis. This approach was applied to eleven species: five lactic acid bacteria (Leuconostoc lactis, Lactobacillus sakei, Lactobacillus paracasei, Lactobacillus fermentum, and Lactobacillus helveticus), four actinobacteria (Brachybacterium articum, Brachybacterium tyrofermentans, Brevibacterium aurantiacum, and Microbacterium gubbeenense), Propionibacterium freudenreichii, and Hafnia alvei. All the strains grew, with maximal populations ranging from 7.4 to 9.2 log (CFU/mL). In total, 52 volatile aroma compounds were identified, of which 49 varied significantly in abundance between bacteria. Principal component analysis of volatile profiles differentiated species by their ability to produce ethyl esters (associated with Brachybacteria), sulfur compounds and branched-chain alcohols (H. alvei), branched-chain acids (H. alvei, P. freudenreichii and L. paracasei), diacetyl and related carbonyl compounds (M. gubbeenense and L. paracasei), among others.

The secreted esterase of Propionibacterium freudenreichii has a major role in cheese lipolysis.

Free fatty acids are important flavor compounds in cheese. Propionibacterium freudenreichii is the main agent of their release through lipolysis in Swiss cheese. Our aim was to identify the esterase(s) involved in lipolysis by P. freudenreichii. We targeted two previously identified esterases: one secreted esterase, PF#279, and one putative cell wall-anchored esterase, PF#774. To evaluate their role in lipolysis, we constructed overexpression and knockout mutants of P. freudenreichii CIRM-BIA1(T) for each corresponding gene. The sequences of both genes were also compared in 21 wild-type strains. All strains were assessed for their lipolytic activity on milk fat. The lipolytic activity observed matched data previously reported in cheese, thus validating the relevance of the method used. The mutants overexpressing PF#279 or PF#774 released four times more fatty acids than the wild-type strain, demonstrating that both enzymes are lipolytic esterases. However, inactivation of the pf279 gene induced a 75% reduction in the lipolytic activity compared to that of the wild-type strain, whereas inactivation of the pf774 gene did not modify the phenotype. Two of the 21 wild-type strains tested did not display any detectable lipolytic activity. Interestingly, these two strains exhibited the same single-nucleotide deletion at the beginning of the pf279 gene sequence, leading to a premature stop codon, whereas they harbored a pf774 gene highly similar to that of the other strains. Taken together, these results clearly demonstrate that PF#279 is the main lipolytic esterase in P. freudenreichii and a key agent of Swiss cheese lipolysis.

Microgradients of pH do not occur around Lactococcus colonies in a model cheese.

Lactococci inoculated into cheese grow as colonies producing lactic acid. The pH microgradients were investigated around colonies in a complex food such as cheese. The results, obtained using a nondestructive technique, demonstrated that pH microgradients did not occur regardless of the acidification kinetics and the size of the colony.