Human milk from tandem feeding dyads does not differ in metabolite and metataxonomic features when compared to single nursling dyads under six months of age

Given the long-term advantages of exclusive breastfeeding to infants and their mothers, there is both an individual and public health benefit to its promotion and support. Data on the composition of human milk over the course of a full period of lactation for a single nursling is sparse, but data on human milk composition during tandem feeding (feeding children of different ages from different pregnancies) is almost entirely absent. This leaves an important knowledge gap that potentially endangers the ability of parents to make a fully informed choice on infant feeding. We compared the metataxonomic and metabolite fingerprints of human milk samples from 15 tandem feeding dyads to that collected from ten exclusively breastfeeding single nursling dyads where the nursling is under six months of age. Uniquely, our cohort also included three tandem feeding nursling dyads where each child showed a preferential side for feeding—allowing a direct comparison between human milk compositions for different aged nurslings. Across our analysis of volume, total fat, estimation of total microbial load, metabolite fingerprinting, and metataxonomics, we showed no statistically significant differences between tandem feeding and single nursling dyads. This included comparisons of preferential side nurslings of different ages. Together, our findings support the practice of tandem feeding of nurslings, even when feeding an infant under six months

Metabolic phenotyping and strain characterisation of pseudomonas aeruginosa Isolates from cystic fibrosis patients using rapid evaporative ionisation mass spectrometry

Rapid evaporative ionisation mass spectrometry (REIMS) is a novel technique for the real-time analysis of biological material. It works by conducting an electrical current through a sample, causing it to rapidly heat and evaporate, with the analyte containing vapour channelled to a mass spectrometer. It was used to characterise the metabolome of 45 Pseudomonas aeruginosa (P. aeruginosa) isolates from cystic fibrosis (CF) patients and compared to 80 non-CF P. aeruginosa. Phospholipids gave the highest signal intensity; 17 rhamnolipids and 18 quorum sensing molecules were detected, demonstrating that REIMS has potential for the study of virulence-related metabolites. P. aeruginosa isolates obtained from respiratory samples showed a higher diversity, which was attributed to the chronic nature of most respiratory infections. The analytical sensitivity of REIMS allowed the detection of a metabolome that could be used to classify individual P. aeruginosa isolates after repeated culturing with 81% accuracy, and an average 83% concordance with multilocus sequence typing. This study underpins the capacities of REIMS as a tool with clinical applications, such as metabolic phenotyping of the important CF pathogen P. aeruginosa, and highlights the potential of metabolic fingerprinting for fine scale characterisation at a sub-species level

Properties of film forming dispersions and films based on chitosan containing basil and thyme essential oils

[EN] Film-forming emulsions and films of chitosan containing basil or thyme essential oil, with and without oleic acid were characterised as to the emulsion stability (particle size, z-potential and rheological behaviour) and barrier, mechanical and optical properties of the films. The losses of the essential oil during the film formation were also quantified as well as the antifungal effect of the films against Aspergillus niger, Botrytis cinerea and Rhizopus stolonifer. The retention of essential oil in the films was greatly dependent on the stability of the film-forming emulsion. The addition of oleic acid (OA) to the chitosan-essential oil formulations enhanced the emulsion stability and oil retention in the film, at the same time that it improved the water vapour barrier properties of the film. Lipids reduced the film stretchability but when OA was present in the formulation, this reduction was mitigated, as well as the changes in colour provoked by the essential oils, whereas OA reduced the film transparency. Chitosan films with thyme or basil essential oil did not inhibit the growth of the tested fungi.The authors acknowledge the financial support provided by the Universitat Politecnica de Valencia (UPPTE/2012/183) and the Ministerio de Ciencia e Innovacion (AGL2010-20694). Author A. Perdones thanks the Universitat Politecnica de Valencia for an FPIgrant.Perdones Montero, Á.; Chiralt, A.; Vargas, M. (2016). Properties of film forming dispersions and films based on chitosan containing basil and thyme essential oils. Food Hydrocolloids. 57:271-279. https://doi.org/10.1016/j.foodhyd.2016.02.006S2712795

Effect of chitosan lemon essential oil coatings on storage-keeping quality of strawberry

[EN] Film-forming dispersions (FFD) were prepared with 1% high molecular weight chitosan and 3% lemon essential oil and were submitted to two different homogenization treatments. The particle size and viscosity of the FFD were evaluated and stand-alone coatings were characterized in terms of water vapour permeability (WVP) and antimicrobial activity. The FFD were applied to cold-stored strawberries, cv. Camarosa, and the physicochemical properties, fungal decay and respiration rate of strawberries were determined throughout cold storage at 5 degrees C. The use of microfluidization to prepare chitosan-based FFD led to a significant reduction in the particle size and apparent viscosity of the dispersions, with no significant effect on the decrease in the WVP of the stand-alone coatings. Chitosan coatings did not show a significant effect in terms of the acidity, pH and soluble solid content of strawberries throughout storage. In contrast, coatings slowed down the respiration rate of samples when lemon essential oil was added to the FFD. Adding lemon essential oil enhanced the chitosan antifungal activity both in in vitro tests and during cold storage in strawberries inoculated with a spore suspension of Botrytis cinerea.The authors acknowledge the financial support provided by Conselleria de Empresa, Universidad y Ciencia (project GV/2010/082), Universitat Politecnica de Valencia (project PAID-06-09-2834) and Ministerio de Economia y Competitividad (project AGL2010-20694). Author A. Perdones is grateful to Universitat Politecnia de Valencia for a FPI grant.Perdones Montero, A.; Sánchez González, L.; Chiralt, A.; Vargas, M. (2012). Effect of chitosan lemon essential oil coatings on storage-keeping quality of strawberry. Postharvest Biology and Technology. 70:32-41. doi:10.1016/j.postharvbio.2012.04.002S32417

Effect of chitosan lemon essential oil coatings on volatile profile of strawberries during storage

[EN] Chitosan coatings containing lemon essential oils were described as effective at controlling fruit fungal decay at 20 C during 7 days. In this work, GC MS was used to characterise the volatile compounds of strawberries during cold storage in order to analyse the influence of fruit coatings with chitosan, containing or not containing lemon essential oil, on the volatile profile of the fruits. The coatings affected the metabolic pathways and volatile profile of the fruits. Pure chitosan promoted the formation of esters and dimethyl furfural in very short time after coating, while coatings containing lemon essential oil incorporated terpenes (limonene, c-terpinene, p-cymene and alpha-citral) to the fruit volatiles and enhanced the fermentative process, modifying the typical fruit aroma composition. No effect of chitosan coatings was sensorially perceived, the changes induced by lemon essential oil were notably appreciated.The authors acknowledge the financial support provided by Ministerio de Economia y Competividad (AGL2013-42989-R). Angela Perdones also thanks the Universidad Politecnica de Valencia for the FPI Grant.Perdones Montero, Á.; Escriche Roberto, MI.; Chiralt, A.; Vargas, M. (2016). Effect of chitosan lemon essential oil coatings on volatile profile of strawberries during storage. Food Chemistry. 197:979-986. https://doi.org/10.1016/j.foodchem.2015.11.054S97998619

Rapid Evaporative Ionisation Mass Spectrometry (REIMS) provides accurate direct from culture species identification within the genus Candida

Members of the genus Candida, such as C. albicans and C. parapsilosis, are important human pathogens. Other members of this genus, previously believed to carry minimal disease risk, are increasingly recognised as important human pathogens, particularly because of variations in susceptibilities to widely used anti-fungal agents. Thus, rapid and accurate identification of clinical Candida isolates is fundamental in ensuring timely and effective treatments are delivered. Rapid Evaporative Ionisation Mass Spectrometry (REIMS) has previously been shown to provide a high-throughput platform for the rapid and accurate identification of bacterial and fungal isolates. In comparison to commercially available matrix assisted laser desorption ionisation time of flight mass spectrometry (MALDI-ToF), REIMS based methods require no preparative steps nor time-consuming cell extractions. Here, we report on the ability of REIMS-based analysis to rapidly and accurately identify 153 clinical Candida isolates to species level. Both handheld bipolar REIMS and high-throughput REIMS platforms showed high levels of species classification accuracy, with 96% and 100% of isolates classified correctly to species level respectively. In addition, significantly different (FDR corrected P value < 0.05) lipids within the 600 to 1000 m/z mass range were identified, which could act as species-specific biomarkers in complex microbial communities

Human milk from tandem feeding dyads does not differ in metabolite and metataxonomic features when compared to single nursling dyads under six months of age

Given the long-term advantages of exclusive breastfeeding to infants and their mothers, there is both an individual and public health benefit to its promotion and support. Data on the composition of human milk over the course of a full period of lactation for a single nursling is sparse, but data on human milk composition during tandem feeding (feeding children of different ages from different pregnancies) is almost entirely absent. This leaves an important knowledge gap that potentially endangers the ability of parents to make a fully informed choice on infant feeding. We compared the metataxonomic and metabolite fingerprints of human milk samples from 15 tandem feeding dyads to that collected from ten exclusively breastfeeding single nursling dyads where the nursling is under six months of age. Uniquely, our cohort also included three tandem feeding nursling dyads where each child showed a preferential side for feeding—allowing a direct comparison between human milk compositions for different aged nurslings. Across our analysis of volume, total fat, estimation of total microbial load, metabolite fingerprinting, and metataxonomics, we showed no statistically significant differences between tandem feeding and single nursling dyads. This included comparisons of preferential side nurslings of different ages. Together, our findings support the practice of tandem feeding of nurslings, even when feeding an infant under six months

Off-colony screening of biosynthetic libraries by rapid laser-enabled mass spectrometry

Leveraging advances in DNA synthesis and molecular cloning techniques, synthetic biology increasingly makes use of large construct libraries to explore large design spaces. For biosynthetic pathway engineering the ability to screen these libraries for a variety of metabolites of interest is essential. If the metabolite of interest or the metabolic phenotype is not easily measurable, screening soon becomes a major bottleneck involving time-consuming culturing, sample preparation, and extraction. To address this, we demonstrate the use of automated Laser-Assisted Rapid Evaporative Ionisation Mass Spectrometry (LA-REIMS) - a form of ambient laser desorption ionisation mass spectrometry - to perform rapid mass spectrometry analysis direct from agar plate yeast colonies without sample preparation or extraction. We use LA-REIMS to assess production levels of violacein and betulinic acid directly from yeast colonies at a rate of 6 colonies per minute. We then demonstrate the throughput enabled by LA-REIMS by screening over 450 yeast colonies in under 4 hours, while simultaneously generating recoverable glycerol stocks of each colony in real-time. This showcases LA-REIMS as a pre-screening tool to complement downstream quantification methods such as LCMS. Through pre-screening several hundred colonies with LA-REIMS, we successfully isolate and verify a strain with a 2.5-fold improvement in betulinic acid production. Finally, we show that LA-REIMS can detect 20 out of a panel of 27 diverse biological molecules, demonstrating the broad applicability of LA-REIMS to metabolite detection. The rapid and automated nature of LA-REIMS makes this a valuable new technology to complement existing screening technologies currently employed in academic and industrial workflows

Utilisation of Ambient Laser Desorption Ionisation Mass Spectrometry (ALDI-MS) improves lipid-based microbial species level identification

The accurate and timely identification of the causative organism of infection is important in ensuring the optimum treatment regimen is prescribed for a patient. Rapid evaporative ionisation mass spectrometry (REIMS), using electrical diathermy for the thermal disruption of a sample, has been shown to provide fast and accurate identification of microorganisms directly from culture. However, this method requires contact to be made between the REIMS probe and microbial biomass; resulting in the necessity to clean or replace the probes between analyses. Here, optimisation and utilisation of ambient laser desorption ionisation (ALDI) for improved speciation accuracy and analytical throughput is shown. Optimisation was completed on 15 isolates of Escherichia coli, showing 5 W in pulsatile mode produced the highest signal-to-noise ratio. These parameters were used in the analysis of 150 clinical isolates from ten microbial species, resulting in a speciation accuracy of 99.4% - higher than all previously reported REIMS modalities. Comparison of spectral data showed high levels of similarity between previously published electrical diathermy REIMS data. ALDI does not require contact to be made with the sample during analysis, meaning analytical throughput can be substantially improved, and further, increases the range of sample types which can be analysed in potential direct-from-sample pathogen detection

Metabonomics and the gut microbiome associated with primary response to Anti-TNF therapy in Crohn’s Disease

Background and Aims Anti-tumour necrosis factor [anti-TNF] therapy is indicated for treatment of moderate to severe inflammatory bowel disease [IBD], but has a primary non-response rate of around 30%. We aim to use metabonomic and metataxonomic profiling to identify predictive biomarkers of anti-TNF response in Crohn’s disease. Methods Patients with luminal Crohn’s disease, commencing anti-TNF therapy, were recruited with urine, faeces, and serum samples being collected at baseline and 3-monthly. Primary response was defined according to a combination of clinical and objective markers of inflammation. Samples were measured using three UPLC-MS assays: lipid, bile acid, and Hydrophillic Interaction Liquid Chromatography [HILIC] profiling with 16S rRNA gene sequencing of faeces. Results Samples were collected from 76 Crohn’s disease patients who were anti-TNF naïve and from 13 healthy controls. There were 11 responders, 37 non-responders, and 28 partial responders in anti-TNF-treated Crohn’s patients. Histidine and cysteine were identified as biomarkers of response from polar metabolite profiling [HILIC] of serum and urine. Lipid profiling of serum and faeces found phosphocholines, ceramides, sphingomyelins, and triglycerides, and bile acid profiling identified primary bile acids to be associated with non-response to anti-TNF therapy, with higher levels of phase 2 conjugates in non-responders. Receiver operating curves for treatment response demonstrated 0.94 +/ -0.10 [faecal lipid], 0.81 +/- 0.17 [faecal bile acid], and 0.74 +/- 0.15 [serum bile acid] predictive ability for anti-TNF response in Crohn’s disease. Conclusions This prospective, longitudinal cohort study of metabonomic and 16S rRNA gene sequencing analysis demonstrates that a range of metabolic biomarkers involving lipid, bile acid, and amino acid pathways may contribute to prediction of response to anti-TNF therapy in Crohn’s disease