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Mutualistic bacteria have different forms of interaction with the host. In contrast to the invasion of pathogenic bacteria, naturally occurring internalization of commensal bacteria has not been studied in depth. Three in vitro methods, gentamicin protection, flow cytometry and confocal laser scanning microscopy, have been implemented to accurately assess the internalization of two lactobacillus strains—Lacticaseibacillus paracasei BL23 and Lacticaseibacillus rhamnosus GG—in Caco-2 and T84 intestinal epithelial cells (IECs) under a variety of physiological conditions and with specific inhibitors. First and most interesting, internalization occurred at a variable rate that depends on the bacterial strain and IEC line, and the most efficient was BL23 internalization by T84 and, second, efficient internalization required active IEC proliferation, as it improved naturally at the early confluence stages and by stimulation with epidermal growth factor (EGF). IFN-γ is bound to innate immune responses and autolysis; this cytokine had a significant effect on internalization, as shown by flow cytometry, but increased internalization was not perceived in all conditions, possibly because it was also stimulating autolysis and, as a consequence, the viability of bacteria after uptake could be affected. Bacterial uptake required actin polymerization, as shown by cytochalasin D inhibition, and it was partially bound to clathrin and caveolin dependent endocytosis. It also showed partial inhibition by ML7 indicating the involvement of cholesterol lipid rafts and myosin light chain kinase (MLCK) activation, at least in the LGG uptake by Caco-2. Most interestingly, bacteria remained viable inside the IEC for as long as 72 h without damaging the epithelial cells, and paracellular transcytosis was observed. These results stressed the fact that internalization of commensal and mutualistic bacteria is a natural, nonpathogenic process that may be relevant in crosstalk processes between the intestinal populations and the host, and future studies could determine its connection to processes such as commensal tolerance, resilience of microbial populations or transorganic bacterial migration.

Genetic studies in rare species are important to determine their patterns of genetic diversity among populations and implement conservation plans aimed to reduce extinction risks. Pachyphytum caesium is an endemic species with extremely small populations in central Mexico. This work analyzes the diversity and genetic structure of Pachyphytum cesium (Crassulaceae) to change the conservation status and propose conservation strategies. Twelve dominant ISSR markers were used to describe the genetic diversity of all known populations. Additionally, we carried out two simulations to explore how the loss of individuals or the local populations extinction affect the genetics parameters of the species. The average results indicated moderate levels of genetic diversity (number of alleles = 89.7 ± 3.9, expected heterozygosity = 0.212 ± 0.0, and percentage of polymorphic loci = 56.1 ± 3.9), parameters that decreased significantly when simulations were performed in P. caesium. Additionally, a genetic structure of the populations was found with low gene flow (Nm = 0.92). Genetic parameters are negatively affected by the loss of individuals and the local extinction of populations. Based on our results, we propose to reclassify the conservation status of the species in danger of extinction, hence a series of conservation strategies are provided to prevent its disappearance.

Cliff-dwelling plant species are highly specialized and adapted to a vulnerable, fragmented, and are mostly endemic, narrowly-distributed and threatened. As a contribution to the conservation efforts of endemic cliff-dwelling species, this study provides an overview of the effects of habitat loss on the abundance and distribution of Pachyphytum caesium (Crassulaceae) due to human disturbances. To achieve this objective, we first conducted a retrospective analysis from 2003–2013 to assess the effects of land use change on the abundance of P. caesium. Secondly, we estimate the abundance and distribution of P. caesium throughout the study area, as well as analyze the effect of rock-climbing activities on the density and population structure of P. caesium. The results suggest differences in population abundance among sites is due to the adverse effects of habitat loss. P. caesium presents a very restricted distribution with small and fragmented populations. In addition, guava agriculture has a significant impact on the chemical soil properties of the hillsides, causing a significant effect on the occurrence of P. caesium, while sport activities remove both the soil and the plants from the cliffs. According to the results, P. caesium is classified as a plant species with extremely small populations (PSESP), and it is highly vulnerable to habitat disturbance. Its conservation is thus a priority to ensure its permanence.