57 research outputs found
Peptide and amino acid metabolism is controlled by an OmpR-family response regulator in Lactobacillus casei
A Lactobacillus casei BL23 strain defective in an OmpR-family response regulator encoded by LCABL_18980 (PrcR, RR11), showed enhanced proteolytic activity caused by overexpression of the gene encoding the proteinase PrtP. Transcriptomic analysis revealed that, in addition to prtP expression, PrcR regulates genes encoding peptide and amino acid transporters, intracellular peptidases and amino acid biosynthetic pathways, among others. Binding of PrcR to twelve promoter regions of both upregulated and downregulated genes, including its own promoter, was demonstrated by electrophoretic mobility shift assays showing that PrcR can act as a transcriptional repressor or activator. Phosphorylation of PrcR increased its DNA binding activity and this effect was abolished after replacement of the phosphorylatable residue Asp-52 by alanine. Comparison of the transcript levels in cells grown in the presence or absence of tryptone in the growth medium revealed that PrcR activity responded to the presence of a complex amino acid source in the growth medium. We conclude that the PrcR plays a major role in the control of the peptide and amino acid metabolism in L. casei BL23. Orthologous prcR genes are present in most members of the Lactobacillaceae and Leuconostocaceae families. We hypothesize that they play a similar role in these bacterial groups.This work was financed by funds from the former Spanish Ministry of Science and Innovation (AGL2007-60975 and AGL2010-15679) and Generalitat Valenciana (ACOMP2012/137). A. Revilla-Guarinos was the recipient of a research fellowship from the Instituto Danone.Peer reviewe
Retinoic-Acid-Induced Downregulation of the 67 KDa Laminin Receptor Correlates with Reduced Biological Aggressiveness of Human Neuroblastoma Cells
16 pages (217-232), 6 figuresThe authors are indebted to Dr. S. Menard (Milan, Italy) for the gift of the antibody against 37LRP. C. M. R. L. was supported by Programa de Personal Técnico de Apoyo (PTA-2003-02-00207; Ministry of Education and Science, Spain). This work was supported by grants from the Spanish former Ministry of Education and Science and Ministry of Science and Innovation (SAF2003-00311, SAF2006–00647 and SAF2007–60780) and Generalitat Valenciana (GRUPOS 03/15 and ACOMP 09/212) (to D. B.) , and Instituto de Salud Carlos III (RD20-102 to S. N.).Peer reviewe
Intestinal Explant Cultures from Gilthead Seabream (Sparus aurata, L.) Allowed the Determination of Mucosal Sensitivity to Bacterial Pathogens and the Impact of a Plant Protein Diet
[EN] The interaction between diet and intestinal health has been widely discussed, although in vivo approaches have reported limitations. The intestine explant culture system developed provides an advantage since it reduces the number of experimental fish and increases the time of incubation compared to similar methods, becoming a valuable tool in the study of the interactions between pathogenic bacteria, rearing conditions, or dietary components and fish gut immune response. The objective of this study was to determine the influence of the total substitution of fish meal by plants on the immune intestinal status of seabream using an ex vivo bacterial challenge. For this aim, two growth stages of fish were assayed (12 g): phase I (90 days), up to 68 g, and phase II (305 days), up to 250 g. Additionally, in phase II, the effects of long term and short term exposure (15 days) to a plant protein (PP) diet were determined. PP diet altered the mucosal immune homeostasis, the younger fish being more sensitive, and the intestine from fish fed short-term plant diets showed a higher immune response than with long-term feeding. Vibrio alginolyticus (V. alginolyticus) triggered the highest immune and inflammatory response, while COX-2 expression was significantly induced by Photobacterium damselae subsp. Piscicida (P. damselae subsp. Piscicida), showing a positive high correlation between the pro-inflammatory genes encoding interleukin 1 beta (IL1-beta), interleukin 6 (IL-6) and cyclooxygenase 2(COX-2).The research was supported by a grant financed by the Spanish Ministerio de Economia y Competitividad AGL2015-70487-P. and Generalitat Valenciana, IDIFEDER/2020/029 The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. It was additionally granted by Contrato Pre-doctoral para la Formacion de Profesorado Universitario from Subprogramas de Formacion y Movilidad within the Programa Estatal de Promocion del Talento y su Empleabilidad of the Ministerio de Educacion, Cultura y Deporte of Spain.Peñaranda, D.; Bäuerl, C.; Tomas-Vidal, A.; Jover Cerda, M.; Estruch, G.; Pérez Martínez, G.; Martínez-Llorens, S. (2020). Intestinal Explant Cultures from Gilthead Seabream (Sparus aurata, L.) Allowed the Determination of Mucosal Sensitivity to Bacterial Pathogens and the Impact of a Plant Protein Diet. International Journal of Molecular Sciences. 21(20):1-20. https://doi.org/10.3390/ijms21207584S1202120Minghetti, M., Drieschner, C., Bramaz, N., Schug, H., & Schirmer, K. (2017). A fish intestinal epithelial barrier model established from the rainbow trout (Oncorhynchus mykiss) cell line, RTgutGC. Cell Biology and Toxicology, 33(6), 539-555. doi:10.1007/s10565-017-9385-xGómez, G. D., & Balcázar, J. L. (2008). A review on the interactions between gut microbiota and innate immunity of fish: Table 1. FEMS Immunology & Medical Microbiology, 52(2), 145-154. doi:10.1111/j.1574-695x.2007.00343.xJose L Gonzalez Vecino, M. H. (2015). Probiotic and Pathogen Ex-vivo Exposure of Atlantic Salmon (Salmo Salar L.) Intestine from Fish Fed Four Different Protein Sources. Journal of Aquaculture Research & Development, 06(05). doi:10.4172/2155-9546.1000340Nematollahi, A., Decostere, A., Ducatelle, R., Haesebrouck, F., & Pasmans, F. (2005). Development of a gut perfusion model as an alternative to the use of live fish. Laboratory Animals, 39(2), 194-199. doi:10.1258/0023677053739710Lin, Y.-C., & Chen, J.-C. (2001). Acute toxicity of ammonia on Litopenaeus vannamei Boone juveniles at different salinity levels. Journal of Experimental Marine Biology and Ecology, 259(1), 109-119. doi:10.1016/s0022-0981(01)00227-1Nematollahi, A., Pasmans, F., Van den Broeck, W., Ducatelle, R., Haesebrouck, F., & Decostere, A. (2005). Association of Flavobacterium psychrophilum strains with intestinal explants of rainbow trout Oncorhynchus mykiss. Diseases of Aquatic Organisms, 67, 67-72. doi:10.3354/dao067067Harper, G. M., Monfort, M., & Saoud, I. P. (2011). An ex vivo approach to studying the interactions of probiotic Pediococcus acidilactici and Vibrio (Listonella) anguillarum in the anterior intestine of rainbow trout Oncorhynchus mykiss. Journal of Aquaculture Research & Development, s1. doi:10.4172/2155-9546.s1-004Løvmo Martinsen, L., Salma, W., Myklebust, R., Mayhew, T. M., & Ringø, E. (2011). Carnobacterium maltaromaticum vs. Vibrio (Listonella) anguillarum in the midgut of Atlantic cod (Gadus morhua L.): an ex vivo study. Aquaculture Research, 42(12), 1830-1839. doi:10.1111/j.1365-2109.2010.02784.xRen, P., Xu, L., Yang, Y., He, S., Liu, W., Ringø, E., & Zhou, Z. (2013). 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: Effects on performance and nutrient efficiency. Aquaculture Nutrition, 24(4), 1163-1174. doi:10.1111/anu.12655Torrecillas, S., Caballero, M. J., Mompel, D., Montero, D., Zamorano, M. J., Robaina, L., … Izquierdo, M. (2017). Disease resistance and response against Vibrio anguillarum intestinal infection in European seabass ( Dicentrarchus labrax ) fed low fish meal and fish oil diets. Fish & Shellfish Immunology, 67, 302-311. doi:10.1016/j.fsi.2017.06.022Estruch, G., Collado, M. C., Peñaranda, D. S., Tomás Vidal, A., Jover Cerdá, M., Pérez Martínez, G., & Martinez-Llorens, S. (2015). Impact of Fishmeal Replacement in Diets for Gilthead Sea Bream (Sparus aurata) on the Gastrointestinal Microbiota Determined by Pyrosequencing the 16S rRNA Gene. PLOS ONE, 10(8), e0136389. doi:10.1371/journal.pone.0136389Estruch, G., Martínez-Llorens, S., Tomás-Vidal, A., Monge-Ortiz, R., Jover-Cerdá, M., Brown, P. B., & Peñaranda, D. S. (2020). 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SARS-CoV-2 RNA and antibody detection in breast milk from a prospective multicentre study in Spain
This study has been supported by a research grant from Fundacion La Marato-TV3 (MilkCORONA, ref 202106).Objectives To develop and validate a specific protocol
for SARS-CoV-
2 detection in breast milk matrix and to
determine the impact of maternal SARS-CoV-
2 infection
on the presence, concentration and persistence of
specific SARS-CoV-
2 antibodies.
Design and patients This is a prospective, multicentre
longitudinal study (April–December 2020) in 60 mothers
with SARS-CoV-
2 infection and/or who have recovered
from COVID-19. A control group of 13 women before the
pandemic were also included.
Setting Seven health centres from different provinces
in Spain.
Main outcome measures Presence of SARS-CoV-
2
RNA in breast milk, targeting the N1 region of the
nucleocapsid gene and the envelope (E) gene; presence
and levels of SARS-CoV-
2-specific
immunoglobulins
(Igs)—IgA, IgG and IgM—in breast milk samples from
patients with COVID-19.
Results All breast milk samples showed negative
results for presence of SARS-CoV-
2 RNA. We observed
high intraindividual and interindividual variability in
the antibody response to the receptor-binding
domain
of the SARS-CoV-
2 spike protein for each of the three
isotypes IgA, IgM and IgG. Main Protease (MPro)
domain antibodies were also detected in milk. 82.9%
(58 of 70) of milk samples were positive for at least
one of the three antibody isotypes, with 52.9% of these
positive for all three Igs. Positivity rate for IgA was
relatively stable over time (65.2%–87.5%), whereas
it raised continuously for IgG (from 47.8% for the first
10 days to 87.5% from day 41 up to day 206 post-PCR
confirmation).
Conclusions Our study confirms the safety of breast
feeding and highlights the relevance of virus-specific
SARS-CoV-
2 antibody transfer. This study provides crucial
data to support official breastfeeding recommendations
based on scientific evidence.Fundacion La Marato-TV3 20210
SARS-CoV-2 RNA and antibody detection in human milk from a prospective multicenter study in Spain
Objectives To develop and validate a specific protocol for SARS-CoV-2 detection in breast milk matrix and to determine the impact of maternal SARS-CoV-2 infection on the presence, concentration and persistence of specific SARS-CoV-2 antibodies. Design and patients This is a prospective, multicentre longitudinal study (April-December 2020) in 60 mothers with SARS-CoV-2 infection and/or who have recovered from COVID-19. A control group of 13 women before the pandemic were also included. Setting Seven health centres from different provinces in Spain. Main outcome measures Presence of SARS-CoV-2 RNA in breast milk, targeting the N1 region of the nucleocapsid gene and the envelope (E) gene; presence and levels of SARS-CoV-2-specific immunoglobulins (Igs)¿IgA, IgG and IgM¿in breast milk samples from patients with COVID-19. Results All breast milk samples showed negative results for presence of SARS-CoV-2 RNA. We observed high intraindividual and interindividual variability in the antibody response to the receptor-binding domain of the SARS-CoV-2 spike protein for each of the three isotypes IgA, IgM and IgG. Main Protease (MPro) domain antibodies were also detected in milk. 82.9% (58 of 70) of milk samples were positive for at least one of the three antibody isotypes, with 52.9% of these positive for all three Igs. Positivity rate for IgA was relatively stable over time (65.2%-87.5%), whereas it raised continuously for IgG (from 47.8% for the first 10 days to 87.5% from day 41 up to day 206 post-PCR confirmation). Conclusions Our study confirms the safety of breast feeding and highlights the relevance of virus-specific SARS-CoV-2 antibody transfer. This study provides crucial data to support official breastfeeding recommendations based on scientific evidence
Metallomic and Untargeted Metabolomic Signatures of Human Milk from SARS-CoV-2 Positive Mothers
Scope
Lack of information about the impact of maternal severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection on the elemental and metabolomic profile of human milk (HM).
Methods and results
An observational study on HM from mothers with COVID-19 is conducted including a prepandemic control group. Maternal–infant clinical records and symptomatology are recorded. The absolute quantification of elements and untargeted relative metabolomic profiles are determined by inductively coupled plasma mass spectrometry and gas chromatography coupled to mass spectrometry, respectively. Associations of HM SARS-CoV-2 antibodies with elemental and metabolomic profiles are studied. COVID-19 has a significant impact on HM composition. COVID-19 reduces the concentrations of Fe, Cu, Se, Ni, V, and Aluminium (Al) and increases Zn compared to prepandemic control samples. A total of 18 individual metabolites including amino acids, peptides, fatty acids and conjugates, purines and derivatives, alcohols, and polyols are significantly different in HM from SARS-CoV-2 positive mothers. Aminoacyl-tRNA biosynthesis, phenylalanine, tyrosine and tryptophan biosynthesis, phenylalanine, and linoleic acid pathways are significantly altered. Differences are obtained depending on COVID-19 symptomatic and asymptomatic status.
Conclusions
This study provides unique insights about the impact of maternal SARS-CoV-2 infection on the elemental and metabolomic profiles of HM that warrants further research due the potential implications for infant health.This work was supported by the projects PG2018-096608-B-C21(Spanish Ministry of Science and innovation (MCIN). Generación delConocimiento. MCIN/ AEI /10.13039/501100011033/ FEDER “Una man-era de hacer Europa”), UHU-1256905 and UHU-202009 from the FEDERAndalusian operative program (Ministry of Economic Transformation, In-dustry, Knowledge and Universities, Andalusia, Spain), and a researchgrant from Fundación La Marató-TV3 (MilkCORONA, ref 202106). FJSCthanks Junta de Andalucía and University of Huelva for a predoctoralcontract (Ref. SNGJ5-TS-005, Garantía Juvenil). Funding for open accesscharge: Universidad de Huelva / CBUA. The authors are grateful to FEDER(European Community) for financial support through grants UNHU13-1E-1611 and UNHU15-CE-3140. The funding sources had no role in the de-sign and conduct of the study; collection, management, analysis, and in-terpretation of the data; preparation, review, or approval of the manuscript;and decision to submit the manuscript for publicationPeer reviewe
Breastfeeding during the COVID-19 pandemic: analysis of the breastmilk antibodies, neutralization capacity and microbiota profile from infected and vaccinated wome
Resumen del póster presentado a las III Jornadas Científicas PTI+ Salud Global, celebradas en el Centro de Ciencias Humanas y Sociales (CCHS), CSIC (Madrid) del 20 al 22 de noviembre de 2023.[Background] Breastmilk is considered the gold standard in infant nutrition and provides bioactive compounds to the neonate, among them antibodies and microbiota. In the context of the COVID- 19 pandemics, there were great concerns about a possible mother-to-infant transfer of SARS-CoV-2, since limited knowledge about the safety of breastfeeding after natural infection or vaccination, as well as the transfer of protective antibodies and their neutralization capacity, was available. Additionally, there are concerns about potential short- and long-term adverse effects of SARS-CoV-2 infection and vaccine-induced changes to the breastmilk microbiome composition, which contributes in shaping the early-life microbiome.[Methods] This study included 60 mothers which had a confirmed SARS-CoV-2 infection and also, 86 mothers vaccinated with mRNA-based (Comirnaty, mRNA-1273) and adenoviral-vectored vaccines (ChAdOx1 nCoV-19) were recruited and breastmilk samples were collected longitudinally from baseline up to 30 days after the second dose at seven or eight time points (depending on vaccine type). In COVID-19 lactating mothers, the presence of SARS-CoV-2 was assessed by RT-qPCR targeting the N1 region of the nucleocapsid gene and the envelope (E) gene. In both studies, the levels of SARS-CoV-2 RBD-specific IgA, IgM and IgG were determined by ELISA. The neutralization capacity was tested using pseudotyped vesicular stomatitis virus carrying either the Wuhan-Hu-1, Delta, or BA.1 Omicron spike proteins. To assess the microbiome composition, DNA from breastmilk samples was extracted and the V3-V4 region of the 16S rRNA gene was sequenced using the MiSeq system of Illumina.[Results] After SARS-CoV-2 infection, no virus-specific RNA was detected in breastmilk samples. Determination
of antibody levels in mothers with confirmed SARS-CoV-2 infection showed that 82.9% (58 of 70) of milk samples were positive for at least one of the three tested antibody isotypes. Vaccination elicited also a strong induction of SARS-CoV-2-specific antibodies, which was higher in IgG when compared to COVID-19 convalescent women and was strongly increased after the 2nd dose. mRNA-based vaccines induced higher IgG and IgA levels when compared to the adenovirus- vectored vaccine, and women with previous virus exposure increased their IgG antibodies levels after the first dose to a similar level observed in vaccinated women after the second dose. When assessing the neutralization capacity, natural infection resulted in higher neutralizing titers that correlated positively with levels of SARS-CoV-2-specific immunoglobulin A in breastmilk. Breastmilk samples from COVID-19 convalescent mothers infected during the first wave (Wuhan-Hu-1 strain) neutralized less effectively Omicron BA.1 than the Wuhan-Hu-1 variant. In addition, significant differences in the capacity to produce neutralizing antibodies were observed between both mRNA- based vaccines and the adenovirus-vectored ChAdOx1 COVID-19 vaccine. First results of the analysis of the breastmilk microbiome found no significant differences in the mean diversity of species (alpha-diversity) after natural SARS-CoV-2 infection, whereas some specific bacterial groups were increased (e.g. Enterobacteriaceae).[Conclusions] Overall, our results indicate that breastmilk from naturally infected women or those vaccinated with
mRNA-based vaccines contain SARS-CoV-2 neutralizing antibodies that could potentially provide protection to breastfed infants from infection.Peer reviewe
Differences in the expression of cell envelope proteinases (CEP) in two Lactobacillus paracasei probiotic strains
Proteinase PrtP (EC:3.4.21.96) is a cell envelope proteinase (CEP) highly expressed in the probiotic strain Lactobacillus paracasei BL312(VSL#3) that accounts for its anti-inflammatory properties. The main aim of this work is to understand differences in CEP expression between this strain and L. paracasei BL23. Hence, differences in the regulation by amino acid sources of four proteinase related genes (prtP, prsA, prtR1 and prtR2) were determined by RT-qPCR in BL312(VSL#3) and BL23 using as a reference BL368, a BL23 derepressed mutant lacking the response regulator (RR) PrcR. BL312(VSL#3) showed greater expression of prtP (2- to 3-fold) than BL23, and prtP was highly repressed by peptone in both strains. Two other putative CEP genes, prtR1 and prtR2, showed a low expression profile. Interestingly, when the prsA-prtP promoter region from both strains, and deleted mutants, were cloned in vector pT1GR, expression of the gfp and mrfp fluorescent reporters was always repressed in BL23 (high or low peptone) and derepressed in BL368, revealing an interesting mechanism of regulation affecting specifically to this promoter. In conclusion, BL312(VSL#3) has higher expression of prtP and other CEP related genes than BL23, that could respond to a natural deregulation in this strain, possibly independent from the RR PrcR.Grants of the Spanish Ministry of Science and Universities AGL2010‐1567, AGL2015-70487-P, RTI2018-097982-B-I00, and from the Generalitat Valenciana ACOMP2012/137. Support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI).Peer reviewe
Lactobacillus casei extracellular vesicles stimulate EGFR pathway likely due to the presence of proteins P40 and P75 bound to their surface
In the complex interplay of beneficial bacteria with the host, there are few examples of bacterial metabolites and effector molecules that have been consistently identified. Protective effects on the intestinal epithelium have been ascribed to P40 and P75, two well characterized cell wall muramidases, present in the culture supernatant of strains belonging to the taxon Lactobacillus casei/paracasei/rhamnosus. This work reports that Lactobacillus casei BL23 extracellular vesicles (BL23 EVs) have a small size (17–20 nm or 24–32 nm, depending on the method used) and contain lipoteichoic acid (LTA). Interestingly, all detected P40 and most of P75 were associated to EVs and possibly located at their external surface, as shown by proteinase K digestion. Biosensor assays showed that both proteins bind LTA and vesicles, suggesting that they could bind to ligands like LTA present on BL23 EVs. Native BL23 EVs have a moderate proinflammatory effect and they were able to induce phosphorylation of the epidermal growth factor receptor (EGFR), showing an effect similar to purified P40 and P75 and leading to the conclusion that the activity described in the supernatant (postbiotic) of these bacteria would be mainly due to P40 and P75 bound to EVs.Grant of the Spanish Ministry of Science and Universities RTI2018-097982-B-I00 and the CONSOLIDER Proposal nber. 25506, that provided the Biacore T100 instrumentPeer reviewe
Shifts in gut microbiota composition in an APP/PSS1 transgenic mouse model of Alzheimer's disease during lifespan
Alzheimer's disease (AD) is the most common form of dementia and one of the major causes of disability and dependency in older people. Accumulating evidences link gut microbiota with different diseases and its relationship with neurodegenerative diseases is becoming most intriguing. This study was aimed to compare the gut microbiota of transgenic APP/PS1 (TG) mice, a well‐established deterministic mouse model of AD, with their C57BL/6 wild‐type (WT) littermates. Faecal samples were collected from 3‐, 6‐ and 24‐month‐old mice and analysed by pyrosequencing of the V1–V3 region of the bacterial 16S rRNA genes. Bacterial profiles were similar in all young mice (3 months old), and started to diverge so that 6‐month‐old WT and TG mice had different and more diverse microbiota. During ageing, Turicibacteriaceae (typical mice bacterial group) and Rikenellaceae increased in all groups, although total Bacteroidetes remained stable. TG mice were characterized by an increase in Proteobacteria after 6 months, particularly the genus Sutterella (Betaproteobacteria), interestingly also increased in autism disorder. Also, the inflammation related family Erysipelotrichaceae was more abundant in TG mice at 24 months compared to wild‐type control. In summary, AD pathology in mice shifts the gut microbiota towards profiles that share features with autism and inflammatory disorders.Financed by the Spanish Ministerio de Economia y Competitividad AGL2015‐70487‐PPeer reviewe
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