Fermented Goat’s Milk Contributes to the Recovery of Iron Deficiency Anemia via Modulation of the Gut Microbiome

Iron deficiency anemia (IDA) is a global public health concern affecting 1.6 billion people worldwide. The administration of iron supplements during the treatment of IDA adversely affects the intestinal barrier function and the composition and functionality of the intestinal microbiome, both of which are already altered during IDA. For this reason, it is of great interest to develop nutritional strategies aimed at alleviating these gut alterations associated with IDA and its treatment. In this sense, fermented goat’s milk (FGM) was studied due to its nutritional quality. Our findings showed that in anemic animals the consumption of a FGM-based diet, compared to a standard diet, had positive modulatory effects on the intestinal microbiome. FGM-based diet restored intestinal dysbiosis, the intestinal barrier functionality, and bacterial translocation, contributing to a more efficient recovery of IDA. Therefore, FGM is a useful nutritional tool to ease intestinal alterations occurring during IDA and during its treatment

Characterization of the pentamidine resistant strain <i>Tb</i>R25.

<p>(a) Relative quantification (RQ) of <i>TbAT1/P2</i> expression in wild type AnTat 1.1 and <i>Tb</i>R25 strains estimated by qRT-PCR. (b) IC₅₀ value for diminazene aceturate in the same strains. Error bars indicate S.D. from 3 replicates. (c) Schematic illustration of the <i>AQP2/AQP3</i> locus showing the heterozygote character of <i>Tb</i>R25 strain with a chimeric gene in one allele and the complete deletion of <i>AQP2</i> gene and the intergenic region in the other (deletion from position 3441867 to 3443663 in chromosome 10, -Tb927_10_v5).</p

Sensitive profile of the pentamidine resistant strain <i>Tb</i>R25.

<p>(a) IC₅₀ analysis. Free pentamidine (grey column); pentamidine-chNPs (red column) and NbAn33-pentamidine-chNPs (blue column). Errors bars indicate SEM from 3–9 independent experiments. Fold reductions are indicated in the graph. (b) Therapeutic effect in <i>Tb</i>R25 acute infection mouse model. Survival (Kaplan-Meier plot) of female C57BL/6J mice infected with <i>Tb</i>R25 (inoculum 2.5 x 10⁶ parasites).</p

Endocytosis of NbAn33-chNPs.

<p>(a) Bloodstream trypanosomes observed by fluorescence microscopy after incubation with NbAn33-chNPs-Alexa Fluor 594 (red) and tomato lectin-FITC (TL, green) as described in Materials and Methods. Samples were taken after 2 minutes (bottom panel) and 10 minutes (top panel) of incubation. DNA is stained with DAPI (blue). Regions of colocalization appear yellow in merged images. (b) Parasite viability after incubation with NbAn33-pentamidine-chNPs at 37° and 4°C for 2 h. Cell death was estimated by propidium iodide staining and FACS analysis at three time points. Error bars represent the S.D. from three independent experiments. Statistical significance was *, p<0.05; ***, p<0.001.</p

Circulation kinetics of fluorescent chitosan nanoparticles in mouse model.

<p>Percentage of the initial dose in peripheral blood when injected via intravenous (i.v.) and intraperitoneal (i.p.) vs. time. Error bars represent the S.D. from 5 mice.</p

Sensitive profile of <i>T</i>. <i>brucei</i> bloodstream forms.

<p>(a) IC₅₀ analysis. Pentamidine (grey column); pentamidine-loaded PEGylated chitosan nanoparticles (pentamidine-chNPs, red column) and nanobody-coated pentamidine-loaded PEGylated chitosan nanoparticles (NbAn33-pentamidine-chNPs, blue column). Errors bars indicate S.D. from 3–9 independent experiments. Statistical significance was ***, p<0.001. (b) Therapeutic effect in <i>T</i>. <i>brucei</i> acute infection mouse model. Survival (Kaplan-Meier plot) of female C57BL/6J mice infected with <i>T</i>. <i>brucei</i> AnT1.1 (1 x 10⁴ parasites). The treatment started once the parasites were detected in blood, at the 3rd day after inoculation and consisted in a daily dose in four consecutive days. Treatment with pentamidine, pentamidine-chNPs, NbAn33-pentamidine-chNPs, NbAn33-chNPs empty (nanobody-coated non pentamidine-loaded PEGylated chitosan nanoparticles) and vehicle (physiological saline solution). (c) Parasitemia in <i>T</i>. <i>brucei</i> acute infection mouse model. Treatment with vehicle (physiological saline solution), NbAn33-chNPs empty (nanobody-coated PEGylated-chitosan nanoparticles), free pentamidine, pentamidine-chNPs (pentamidine-loaded PEGylated chitosan nanoparticles), NbAn33-pentamidine-chNPs (nanobody-coated pentamidine-loaded PEGylated chitosan nanoparticles).</p