Antibiotic resistance is rising in important bacterial pathogens. Phage therapy (PT), the use of bacterial viruses infecting the pathogen in a species-specific way, is a potential alternative. T4-like coliphages or a commercial Russian coliphage product or placebo was orally given over 4 days to Bangladeshi children hospitalized with acute bacterial diarrhea. Safety of oral phage was assessed clinically and by functional tests; coliphage and Escherichia coli titers and enteropathogens were determined in stool and quantitative diarrhea parameters (stool output, stool frequency) were measured. Stool microbiota was studied by 16S rRNA gene sequencing; the genomes of four fecal Streptococcus isolates were sequenced. No adverse events attributable to oral phage application were observed (primary safety outcome). Fecal coliphage was increased in treated over control children, but the titers did not show substantial intestinal phage replication (secondary microbiology outcome). 60% of the children suffered from a microbiologically proven E. coli diarrhea; the most frequent diagnosis was ETEC infections. Bacterial co-pathogens were also detected. Half of the patients contained phage-susceptible E. coli colonies in the stool. E. coli represented less than 5% of fecal bacteria. Stool ETEC titers showed only a short-lived peak and were otherwise close to the replication threshold determined for T4 phage in vitro. An interim analysis after the enrollment of 120 patients showed no amelioration in quantitative diarrhea parameter by PT over standard care (tertiary clinical outcome). Stool microbiota was characterized by an overgrowth with Streptococcus belonging to the Streptococcus gallolyticus and Streptococcus salivarius species groups, their abundance correlated with quantitative diarrhea outcome, but genome sequencing did not identify virulence genes. Oral coliphages showed a safe gut transit in children, but failed to achieve intestinal amplification and to improve diarrhea outcome, possibly due to insufficient phage coverage and too low E. coli pathogen titers requiring higher oral phage doses. More knowledge is needed on in vivo phage-bacterium interaction and the role of E. coli in childhood diarrhea for successful PT. The study was supported by a grant from Nestlé Nutrition and Nestlé Health Science. The trial was registered with Identifier NCT00937274 at ClinicalTrials.gov.

We investigated the amplification and purification of phage preparations with respect to titer, contamination level, stability, and technical affordability. Using various production systems (wave bags, stirred-tank reactors, and Erlenmeyer flasks), we obtained peak titers of 10 ⁹ to 10 ¹⁰ PFU/ml for T4-like coliphages. Phage lysates could be sterilized through 0.22-μm membrane filters without titer loss. Phages concentrated by differential centrifugation were not contaminated with cellular debris or bacterial proteins, as assessed by electron microscopy and mass spectrometry, respectively. Titer losses occurred by high-speed pelleting of phages but could be decreased by sedimentation through a sucrose cushion. Alternative phage concentration methods are prolonged medium-speed centrifugation, strong anion-exchange chromatography, and ultrafiltration, but the latter still allowed elevated lipopolysaccharide contamination. T4-like phages could not be pasteurized but maintained their infectivity titer in the cold chain. In the presence of 10 mM magnesium ions, phages showed no loss of titer over 1 month at 30°C.

Summary: Eighty‐nine T 4‐like phages from our phage collection were tested against four collections of childhood diarrhoea‐associated E scherichia coli isolates representing different geographical origins ( M exico versus B angladesh), serotypes (69 O , 27 H serotypes), pathotypes ( ETEC , EPEC , EIEC , EAEC , VTEC , S higella ), epidemiological settings (community and hospitalized diarrhoea) and years of isolation. With a cocktail consisting of 3 to 14 T 4‐like phages, we achieved 54% to 69% coverage against predominantly EPEC isolates from M exico, 30% to 53% against mostly ETEC isolates from a prospective survey in B angladesh, 24% to 61% against a mixture of pathotypes isolated from hospitalized children in B angladesh, and 60% coverage against S higella isolates. In comparison a commercial R ussian phage cocktail containing a complex mixture of many different genera of coliphages showed 19%, 33%, 50% and 90% coverage, respectively, against the four above‐mentioned collections. Few O serotype‐specific phages and no broad‐host range phages were detected in our T 4‐like phage collection. Interference phenomena between the phage isolates were observed when constituting larger phage cocktails. Since the coverage of a given T 4‐like phage cocktail differed with geographical area and epidemiological setting, a phage composition adapted to a local situation is needed for phage therapy approaches against E . coli pathogens.

Often blamed for bringing green aromas and astringency to wines, the use of stems is also empirically known to improve the aromatic complexity and freshness of some wines. Although applied in different wine-growing regions, stems use remains mainly experimental at a cellar level. Few studies have specifically focused on the compounds extracted from stems during fermentation and maceration and their potential impact on the must and wine matrices. We identified current knowledge on stem chemical composition and inventoried the compounds likely to be released during maceration to consider their theoretical impact. In addition, we investigated existing studies that examined the impact of either single stems or whole clusters on the wine quality. Many parameters influence stems’ effect on the wine, especially grape variety, stem state, how stems are incorporated, when they are added, and contact duration. Other rarely considered factors may also have an impact, including vintage and ripening conditions, which could affect the lignification of the stem.

This article synthetizes the observations from five experiments and brings practical results about the effects of the timing and intensity of defoliation on white and red Vitis vinifera cultivars. – Pinot noir, Gamay, Merlot, Chasselas and Doral. Intensive defoliation (removal of 6 basal leaves + 6 lateral shoots per shoot) was completed at three developmental stages of the grapevine, i.e., preflowering, flowering and bunch closure. Preflowering defoliation had tremendous consequences on vine agronomic performance, mainly to the detriment of berry set; the yield was highly affected by preflowering defoliation (approximately −35 % in comparison with no defoliation). The intensity of defoliation allowed its impact on the yield to be modulated. The intensity of defoliation also had a positive impact against millerandage, sunburn symptoms and Botrytis cinerea development. The berry skin thickness doubled, and the polyphenol concentration significantly increased with preflowering defoliation. Due to preflowering defoliation, the red wines were often preferred for their color and mouthfeel. However, this practice had a negligible impact on white wine composition. Regardless, preflowering defoliation did not have a negative impact on wine parameters. In the context of this study, preflowering defoliation seems to be an interesting technique to reduce vigor and control the high production potential. Preflowering defoliation also represents a prophylactic solution to reduce both chemical applications and cluster-thinning costs.

To date, there are no specific guidelines for the use of bioacidifying yeasts in winemaking. In this work, we aimed to characterize an oenological strain of Lachancea thermotolerans (Lt), a non-Saccharomyces lactic acid-producing yeast, and to test different sequential inoculation conditions with Saccharomyces cerevisiae (Sc). The results of bench scale vinifications showed that both the strategy of inoculating Sc 12 h after Lt and the mixing of Lt and Sc during fermentation delivered an acceptable increase in lactic acid (2 g/L) and a decrease in pH (about 0.15 units). Therefore, both strategies were implemented in winery experiments. Our results at the cellar scale showed no increase in acidity, which was likely due to the presence of indigenous yeasts. Overall, our experience shows the difficulty of translating laboratory protocols into cellar experiments and calls for further research into new strategies for implementing acidifying yeasts.