Comparison of the Fecal Bacteriome of HIV-Positive and HIV-Negative Older Adults.

HIV infection is considered a scenario of accelerated aging. Previous studies have suggested a link between aging, frailty, and gut dysbiosis, but there is a knowledge gap regarding the HIV population. Our objective was to compare the fecal bacteriome of older people with HIV (PWH) and non-HIV controls, and to assess potential links between gut dysbiosis and frailty. A total of 36 fecal samples (24 from PWH and 12 from non-HIV controls) were submitted to a metataxonomic analysis targeting the V3-V4 hypervariable region of the 16S rRNA gene. High-quality reads were assembled and classified into operational taxonomic units. Alpha diversity, assessed using the Shannon index, was higher in the control group than in the HIV group (p < 0.05). The relative abundance of the genus Blautia was higher in the HIV group (p < 0.001). The presence of Blautia was also higher in PWH with depression (p = 0.004), whereas the opposite was observed for the genus Bifidobacterium (p = 0.004). Our study shows shifts in the composition of the PWH bacteriome when compared to that of healthy controls. To our knowledge, this is the first study suggesting a potential link between depression and gut dysbiosis in the HIV population.

Influence of SARS-CoV-2 Status and Aging on the Nasal and Fecal Immunological Profiles of Elderly Individuals Living in Nursing Homes.

In the frame of SARS-CoV-2 infection, studies regarding cytokine profiling of mucosal-related samples are scarce despite being the primary infection sites. The objective of this study was to compare the nasal and fecal inflammatory profiles of elderly individuals living in a nursing home highly affected by COVID-19 (ELD1) with those of elderly individuals living in a nursing home with no cases of SARS-CoV-2 infection (ELD2) and, also, with those of healthy SARS-CoV-2-negative younger adults (YHA). BAFF/TNFSF13B, IL6, IL10 and TNF-α (immunological hallmarks of SARS-CoV-2 infection) were the only immune factors whose concentrations were different in the three groups. Their highest concentrations were achieved in the ELD1 group. Nasal and fecal concentrations of a wide number of pro-inflammatory cytokines were similar in the ELD1 and ELD2 groups but higher than those found in the YHA samples. These results reinforce the hypothesis that immunosenescence and inflammaging rendered the elderly as a highly vulnerable population to a neo-infection, such as COVID-19, which was evidenced during the first pandemic waves.

Immunomodulation of the Vaginal Ecosystem by Ligilactobacillus salivarius CECT 30632 Improves Pregnancy Rates among Women with Infertility of Unknown Origin or Habitual Abortions.

In this study, the probiotic potential of Ligilactobacillus salivarius CECT 30632 was assessed, including properties specifically related with gynecological targets. This strain displayed co-aggregative and antimicrobial activity against a wide spectrum of vaginal pathogens while being respectful with the growth of vaginal lactobacilli. The strain produced a high concentration of lactic acid and displayed α-amylase activity when assayed in vitro. It showed a noticeable survival rate after exposition to conditions similar to those present in the human digestive tract and was adhesive to both vaginal and intestinal cells. Subsequently, their capacity to increase pregnancy rates among women with habitual abortion or infertility of unknown origin was studied. Administration of L. salivarius CECT 30632 (~9 log10 CFU) daily for a maximum of six months to these women was safe and led to a successful pregnancy rate of 67.5% (80% and 55% for women with repetitive abortion and infertile women, respectively). Significant differences in Nugent score, vaginal pH, and vaginal concentrations of lactobacilli, TGF-ß, and VEFG were observed when the samples collected before the intervention were compared with those collected after the treatment among those women who got pregnant. Therefore, this strain can modulate the vaginal ecosystem and lead to better fertility outcomes.

Nutritional composition of green asparagus (Asparagus officinalis L.), edible part and by-products, and assessment of their effect on the growth of human gut-associated bacteria.

Asparagus is considered a healthy food with a high content of bioactive compounds. In this study, the proximate and mineral composition, non-digestible carbohydrates and bioactive compounds of edible spear, spear by-product and root have been evaluated. Their activity on the growth of human gut-associated bacteria has been studied. The results support the high nutritional and functional value of the asparagus, including its by-products, highlighting the potential of the non-edible parts to be used as prebiotics. A remarkable content in xylose, inulin, flavonoids and saponins has been found. It has been shown that the spear by-product can be selectively used to promote the growth of commensal or probiotic lactobacilli and bifidobacteria strains. It has been confirmed that any part of the asparagus has a potential future as a healthy food or as health-promoting ingredients, however more work is required to identify the compounds able to modulate the human gut microbiota.

Metataxonomic Analysis of Milk Samples From SARS-CoV-2-Positive and SARS-CoV-2-Negative Women.

Objective: To assess the impact of SARS-CoV-2 viral infection on the metataxonomic profile and its evolution during the first month of lactation. Methods: Milk samples from 37 women with full-term pregnancies and mild SARS-CoV-2 infection and from 63 controls, collected in the first and fifth postpartum weeks, have been analyzed. SARS-CoV-2 RNA was assessed by reverse transcription polymerase chain reaction (RT-PCR) both in cases and controls. After DNA extraction, the V3-V4 hypervariable region of the gene 16S rRNA was amplified and sequenced using the MiSeq system of Illumina. Data were submitted for statistical and bioinformatics analyses after quality control. Results: All the 1st week and 5th week postpartum milk samples were negative for SARS-CoV-2 RNA. Alpha diversity showed no differences between milk samples from the study and control group, and this condition was maintained along the observation time. Analysis of the beta-diversity also indicated that the study and control groups did not show distinct bacterial profiles. Staphyloccus and Streptococcus were the most abundant genera and the only ones that were detected in all the milk samples provided. Disease state (symptomatic or asymptomatic infection) did not affect the metataxonomic profile in breast milk. Conclusion: These results support that in the non-severe SARS-CoV-2 pregnant woman infection the structure of the bacterial population is preserved and does not negatively impact on the human milk microbiota.

Interactions between human milk oligosaccharides, microbiota and immune factors in milk of women with and without mastitis.

Lactational mastitis is an excellent target to study possible interactions between HMOs, immune factors and milk microbiota due to the infectious and inflammatory nature of this condition. In this work, microbiological, immunological and HMO profiles of milk samples from women with (MW) or without (HW) mastitis were compared. Secretor status in women (based on HMO profile) was not associated to mastitis. DFLNH, LNFP II and LSTb concentrations in milk were higher in samples from HW than from MW among Secretor women. Milk from HW was characterized by a low bacterial load (dominated by Staphylococcus epidermidis and streptococci), high prevalence of IL10 and IL13, and low sialylated HMO concentration. In contrast, high levels of staphylococci, streptococci, IFNγ and IL12 characterized milk from MW. A comparison between subacute (SAM) and acute (AM) mastitis cases revealed differences related to the etiological agent (S. epidermidis in SAM; Staphylococcus aureus in AM), milk immunological profile (high content of IL10 and IL13 in SAM and IL2 in AM) and milk HMOs profile (high content of 3FL in SAM and of LNT, LNnT, and LSTc in AM). These results suggest that microbiological, immunological and HMOs profiles of milk are related to mammary health of women.

Administration of Ligilactobacillus salivarius CECT 30632 to elderly during the COVID-19 pandemic: Nasal and fecal metataxonomic analysis and fatty acid profiling.

Elderly was the most affected population during the first COVID-19 and those living in nursing homes represented the most vulnerable group, with high mortality rates, until vaccines became available. In a previous article, we presented an open-label trial showing the beneficial effect of the strain Ligilactobacillus salivarius CECT 30632 (previously known as L. salivarius MP101) on the functional and nutritional status, and on the nasal and fecal inflammatory profiles of elderly residing in a nursing home highly affected by the pandemic. The objective of this post-hoc analysis was to elucidate if there were changes in the nasal and fecal bacteriomes of a subset of these patients as a result of the administration of the strain for 4 months and, also, its impact on their fecal fatty acids profiles. Culture-based methods showed that, while L. salivarius (species level) could not be detected in any of the fecal samples at day 0, L. salivarius CECT 30632 (strain level) was present in all the recruited people at day 120. Paradoxically, the increase in the L. salivarius counts was not reflected in changes in the metataxonomic analysis of the nasal and fecal samples or in changes in the fatty acid profiles in the fecal samples of the recruited people. Overall, our results indicate that L. salivarius CECT 30632 colonized, at least temporarily, the intestinal tract of the recruited elderly and may have contributed to improvements in their functional, nutritional, and immunological status, without changing the general structure of their nasal and fecal bacteriomes when assessed at the genus level. They also suggest the ability of low abundance bacteria to train immunity.

Dietary Habits and Relationship with the Presence of Main and Trace Elements, Bisphenol A, Tetrabromobisphenol A, and the Lipid, Microbiological and Immunological Profiles of Breast Milk.

Breastfeeding is the best way to feed an infant, although it can also be a source of abiotic contaminants such as heavy metals or bisphenol A (BPA). The early life exposure to these compounds can lead to serious toxic effects in both the short and long-term. These substances can reach breast milk through the mother's habits, diet being one of the main routes of exposure. The aim of the present work was to analyse possible associations between the dietary habits of women and the content of major trace elements, BPA, fatty acids and lipids, and the microbiological and immunological profiles of human milk. Possible associations between major trace elements and BPA and the lipid, microbiological and immunological profiles were also analysed. The results of this study support that the microbiological composition of human milk is associated with the dietary habits of the women, and that the consumption of canned drinks is related to the presence of BPA in human milk. Furthermore, some relationships were found between the amount of major trace elements and the microbiological and immunological profile of the milk samples. Finally, the presence of BPA was associated with changes in the immunological profile of human milk.

Administration of Ligilactobacillus salivarius MP101 in an Elderly Nursing Home during the COVID-19 Pandemic: Immunological and Nutritional Impact.

The elderly population living in nursing homes is particularly vulnerable to COVID-19 although individual susceptibility to SARS-CoV-2 infection may be related to the host microbiota. The objective of this work was to investigate the effect of Ligilactobacillus salivarius MP101 on the functional (Barthel index), cognitive (GDS/FAST), and nutritional (MNA) status as well as on the nasal and fecal inflammatory profiles of elderly residents living in a nursing home that is highly affected by COVID-19. A total of 25 residents participated in the trial, which involved the daily ingestion of a dairy product (L. salivarius MP101: 9.3 log10 CFU per unit) for 4 months. Nasal and fecal samples were analyzed for 37 immune factors at recruitment and at the end of the study. After the trial, no change in the GDS/FAST scores were found but, in contrast, the values for the Barthel index and the MNA score improved significantly. The concentrations of some immune factors changed significantly after the trial, including a decrease in the concentrations of BAFF/TNFSF13B, APRIL/TNFSF13, IL8, IL31, osteopontin, sTNF-R1, and sTNF-R2, and an increase in chitinase 3-like 1, IL19, IL35, and pentraxin 3 was also observed. In conclusion, L. salivarius MP101 seems to be a promising strain for improving or maintaining health in this highly vulnerable population.

Immediate Pre-Partum SARS-CoV-2 Status and Immune Profiling of Breastmilk: A Case-Control Study.

Objetive: To address the prevalence of SARS-CoV-2 and the evolutionary profile of immune compounds in breastmilk of positive mothers according to time and disease state. Methods: Forty-five women with term pregnancies with confirmed non-severe SARS-CoV-2 infection (case group), and 96 SARS-CoV-2 negative women in identical conditions (control group) were approached, using consecutive sample. Weekly (1st to 5th week postpartum) reverse transcription polymerase chain reaction (RT-PCR) in nasopharyngeal swabs (cases) and breastmilk (cases and controls) were obtained. Concentration of cytokines, chemokines, and growth factors in breastmilk (cases and controls) were determined at 1st and 5th week post-partum. Results: Thirty-seven (study group) and 45 (control group) women were enrolled. Symptomatic infection occurred in 56.8% of women in the study group (48% fever, 48% anosmia, 43% cough). SARS-CoV-2 RNA was not found in breastmilk samples. Concentrations of cytokines (IFN-γ, IL-1ra, IL-4, IL-6, IL-9, IL-13, and TNF-α) chemokines (eotaxin, IP-10, MIP-1α, and RANTES) and growth factors (FGF, GM-CSF, IL7, and PDGF-BB) were higher in breastmilk of the study compared with the control group at 1st week postpartum. Immune compounds concentrations decreased on time, particularly in the control group milk samples. Time of nasopharyngeal swab to become negative influenced the immune compound concentration pattern. Severity of disease (symptomatic or asymptomatic infection) did not affect the immunological profile in breast milk. Conclusions: This study confirms no viral RNA and a distinct immunological profile in breastmilk according to mother's SARS-CoV-2 status. Additional studies should address whether these findings indicate efficient reaction against SARS-CoV-2 infection, which might be suitable to protect the recipient child.

Application of Ligilactobacillus salivarius CECT5713 to Achieve Term Pregnancies in Women with Repetitive Abortion or Infertility of Unknown Origin by Microbiological and Immunological Modulation of the Vaginal Ecosystem.

In this study, the cervicovaginal environment of women with reproductive failure (repetitive abortion, infertility of unknown origin) was assessed and compared to that of healthy fertile women. Subsequently, the ability of Ligilactobacillus salivarius CECT5713 to increase pregnancy rates in women with reproductive failure was evaluated. Vaginal pH and Nugent score were higher in women with reproductive failure than in fertile women. The opposite was observed regarding the immune factors TGF-ß 1, TFG-ß 2, and VEFG. Lactobacilli were detected at a higher frequency and concentration in fertile women than in women with repetitive abortion or infertility. The metataxonomic study revealed that vaginal samples from fertile women were characterized by the high abundance of Lactobacillus sequences, while DNA from this genus was practically absent in one third of samples from women with reproductive failure. Daily oral administration of L. salivarius CECT5713 (~9 log10 CFU/day) to women with reproductive failure for a maximum of 6 months resulted in an overall successful pregnancy rate of 56%. The probiotic intervention modified key microbiological, biochemical, and immunological parameters in women who got pregnant. In conclusion, L. salivarius CECT5713 has proved to be a good candidate to improve reproductive success in women with reproductive failure.

Metataxonomic and immunological analysis of milk from ewes with or without a history of mastitis.

Mastitis is a highly prevalent condition that has a great impact on milk production and animal welfare, and often requires substantial management efforts. For this reason, it is generally considered an important threat to the dairy industry. Many microbial, host, and environmental factors can protect against, predispose to, or influence the development of mastitis. The objective of this work was to characterize the milk microbiota of Manchega ewes, and to compare samples from animals with and without a history of mastitis. We analyzed milk samples from 36 ewes belonging to 2 different farms (18 ewes from each farm) using culture-dependent and culture-independent techniques. We also analyzed several immune compounds to investigate associations of mastitis with 3 main variables: farm; history of mastitis or no mastitis; and parity number. Both culture-dependent and culture-independent techniques showed that ewe milk harbored a site-specific complex microbiota and microbiome. Staphylococcus epidermidis was the main species driving the difference between farm A (where it was the dominant species) and B (where it was not). In contrast, samples from farm B were characterized by the presence of a wide spectrum of other coagulase-negative staphylococci. Some of these species have already been associated with subclinical intramammary infections in ruminants. Of the 10 immune compounds assayed in this study, 3 were related to a history of mastitis [IL-8, IFN-γ, and IFN-gamma-induced protein 10 (IP-10)]. Increases in IL-8 concentrations in milk seemed to be a feature of subclinical mastitis in sheep, and in this study, this immune factor was detected only in samples from ewes with some episodes of mastitis and from the group with the highest somatic cell count. We also observed a positive correlation between the samples with the highest somatic cell count and IFN-γ and IP-10 levels. Our results suggest that these 3 compounds could be used as biomarkers for the negative selection of mastitis-prone animals, particularly when somatic cell count is very high.

Early-life intake of major trace elements, bisphenol A, tetrabromobisphenol A and fatty acids: Comparing human milk and commercial infant formulas.

In the present study, the presence of a wide spectrum of major and trace elements (As, Ag, Al, Ba, Cd, Co, Cr, Cu, Hg, Mn, Ni, Sr, Sb, Se, Sn, Pb, V, and Zn), fatty acids, as well as some pollutants like free and total BPA and tetrabromobisphenol A (TBBPA), was analysed in human milk (n = 53) and infant formula (n = 50) samples. In addition, the infant exposure to these chemicals was assessed. The content of free BPA and several elements (Al, Ca, Cr, Cu, Fe, K, Mg, Mn, Na, Ni, Sn, Sr, and Zn) was higher (p < 0.01) in infant formula samples. Furthermore, human milk contained levels of BPA and elements that, in almost all cases, were well below their respective EFSA and/or WHO thresholds, being also independent of the maternal characteristics (e.g., age, BMI or breastfeeding period). The fatty acid profiling also revealed major differences between human milk and infant formulas, which should be taken in account in the development of new formulas as well as in specific recommendations for the diet of breastfeeding mothers. Anyway, the results of this study reinforce that breastfeeding should be always the first feeding option in early life.

Pluripotency markers in tissue and cultivated cells in vitro of different regions of human amniotic epithelium.

Studies have described the presence of pluripotent markers in vivo and in vitro in human amnion. However, the amnion can be divided into reflected, placental and umbilical regions that are anatomically and functionally heterogeneous. Here, we evaluated the expression of pluripotency markers in tissue and cultivated cells in vitro of different regions of human amnion. To this end, we determined the presence of the core pluripotency factors OCT-4, NANOG and SOX-2 by immunofluorescence and RT-PCR and also performed transcriptome analysis of the different regions of amnion tissue. We identified the mRNA and protein of the pluripotency factors in the different regions of human amnion tissue. However, the OCT-4 and NANOG immunolocalization was cytoplasmic, whereas SOX-2 immunolocalization was nuclear regardless of the region analyzed. Moreover, we found three subpopulations of cells in the in vitro cultures of reflected and placental amnion: cells with immunostaining only in the nucleus, only in the cytoplasm, or in both compartments. Yet no statistically significant differences were found between the reflected and placental amnion. These results suggest a homogeneous distribution of the pluripotency transcription factors of the different regions of human amnion to isolate stem cells that can be used in regenerative medicine.

Capturing the ephemeral human pluripotent state.

During human development, pluripotency is present only in early stages of development. This ephemeral cell potential can be captured in vitro by obtaining pluripotent stem cells (PSC) with self-renewal properties, the human embryonic stem cells (hESC). However, diverse studies suggest the existence of a plethora of human PSC (hPSC) that can be derived from both embryonic and somatic sources, depending on defined culture conditions, their spatial origin, and the genetic engineering used for reprogramming. This review will focus on hPSC, covering the conventional primed hESC, naïve-like hPSC that resemble the ground-state of development, region-selective PSC, and human induced PSC (hiPSC). We will analyze differences and similarities in their differentiation potential as well as in the molecular circuitry of pluripotency. Finally, we describe the need for human feeder cells to derive and maintain hPSC, because they could emulate the interaction of in vivo pluripotent cells with extraembryonic structures that support development. Developmental Dynamics 245:762-773, 2016. © 2016 Wiley Periodicals, Inc.

Human amniotic epithelial cells as feeder layer to derive and maintain human embryonic stem cells from poor-quality embryos.

Data from the literature suggest that human embryonic stem cell (hESC) lines used in research do not genetically represent all human populations. The derivation of hESC through conventional methods involve the destruction of viable human embryos, as well the use of mouse embryonic fibroblasts as a feeder layer, which has several drawbacks. We obtained the hESC line (Amicqui-1) from poor-quality (PQ) embryos derived and maintained on human amniotic epithelial cells (hAEC). This line displays a battery of markers of pluripotency and we demonstrated the capacity of these cells to produce derivates of the three germ layers.

[Human amniotic epithelium (HAE) as a possible source of stem cells (SC)]. / Las células del epitelio amniótico humano (CEAh) como posible fuente de células troncales (CT).

There have been major recent advances in the field of developmental biology due to the investigation on stem cells (SC). Stem cells are characterized by their capacity of auto-renewal and differentiation to different cellular phenotypes. Based on the developmental stage, they can be classified into two different types: embryonic SCs and adult SCs. It has been widely reported that several problems need to be resolved before their possible clinical applications. As a result, fetal membranes have been suggested as an alternative source of SCs. In the human amniotic epithelium, the presence of markers of pluripotent SC´s has been reported, and its capacity as a feeder layer for expansion of different SC types. Also, fetal membranes are a discarded product after delivery, and thus there are not any ethical issues related to its use. In conclusion, the human amniotic epithelium can be a strong candidate for regenerative medicine.

Markers of Pluripotency in Human Amniotic Epithelial Cells and Their Differentiation to Progenitor of Cortical Neurons.

Human pluripotent stem cells (hPSC) have promise for regenerative medicine due to their auto-renovation and differentiation capacities. Nevertheless, there are several ethical and methodological issues about these cells that have not been resolved. Human amniotic epithelial cells (hAEC) have been proposed as source of pluripotent stem cells. Several groups have studied hAEC but have reported inconsistencies about their pluripotency properties. The aim of the present study was the in vitro characterization of hAEC collected from a Mexican population in order to identify transcription factors involved in the pluripotency circuitry and to determine their epigenetic state. Finally, we evaluated if these cells differentiate to cortical progenitors. We analyzed qualitatively and quantitatively the expression of the transcription factors of pluripotency (OCT4, SOX2, NANOG, KLF4 and REX1) by RT-PCR and RT-qPCR in hAEC. Also, we determined the presence of OCT4, SOX2, NANOG, SSEA3, SSEA4, TRA-1-60, E-cadherin, KLF4, TFE3 as well as the proliferation and epigenetic state by immunocytochemistry of the cells. Finally, hAEC were differentiated towards cortical progenitors using a protocol of two stages. Here we show that hAEC, obtained from a Mexican population and cultured in vitro (P0-P3), maintained the expression of several markers strongly involved in pluripotency maintenance (OCT4, SOX2, NANOG, TFE3, KLF4, SSEA3, SSEA4, TRA-1-60 and E-cadherin). Finally, when hAEC were treated with growth factors and small molecules, they expressed markers characteristic of cortical progenitors (TBR2, OTX2, NeuN and ß-III-tubulin). Our results demonstrated that hAEC express naïve pluripotent markers (KLF4, REX1 and TFE3) as well as the cortical neuron phenotype after differentiation. This highlights the need for further investigation of hAEC as a possible source of hPSC.