Cryopreservation of Infant Gut Microbiota with Natural Cryoprotectants.

A growing body of evidence has demonstrated the importance of the gut microbiome in human health. In general, fecal microbial samples are used to study the mechanisms of relevant diseases. In this context, it is worth mentioning that an optimized cryopreservation method is urgently needed to successfully perform clinical diagnosis, therapy, and scientific investigations of the gut microbiome without affecting its viability and biological activity. In this study, we aimed to test the relative cryopreservation efficiency of different nontoxic natural cryoprotectants using infant fecal and meconium samples. First, we selected two facultative and two obligate anaerobic bacteria as the experimental gut microbial strain to compare these cryoprotectants' toxicity and concentration-dependent bacteria viabilities after cryopreservation, then the viabilities and bacterial diversity of mixed facultative and obligate anaerobic bacteria. Finally, we explored the effects of optimized cryoprotectants for meconium and infant feces after cryopreservation using 16S rRNA sequencing analyses. In addition, to better understand the effectiveness of these cryoprotectants, we used different freeze-thaw conditions mimicking real-life situations in the process of distribution. We found that the better choice for the infant fecal sample's cryopreservation was 100 mg/mL trehalose, whereas 200 mg/mL trehalose/betaine was the optimum choice for meconium cryopreservation. We hope that our results will shed light on the importance of natural cryoprotectants toward the long-term and stable viability of invaluable human gut microbial specimens.

Impacts of trehalose and l-proline on the thermodynamic nonequilibrium phase change and thermal properties of normal saline.

Understanding the phase change behavior and thermal properties of cryoprotective agents (CPAs) in biological solutions is essential for enhancing the success of cryopreservation and biobanking. In this study, the phase change behavior and thermal properties of normal saline added with trehalose or l-proline were investigated using differential scanning calorimeter (DSC) and cryomicroscope during freezing and warming. The addition of trehalose or l-proline can eliminate the eutectic formation in normal saline. Trehalose had significantly lower latent heat release than l-proline does at a high concentration of 1 M (P < 0.05), while unfrozen water content of trehalose is significantly lower than that of l-proline at all the concentrations (P < 0.05). It was also found that addition of 0.2 M, 0.3 M and 1 M trehalose can achieve partial vitrification in normal saline and that the glass transition temperature rises along with the increase in concentrations of trehalose. However, no vitrification was observed in normal saline with l-proline at any concentrations. Besides, rates of ice crystal growth in normal saline added with trehalose are slower than those in normal saline with l-proline at the same concentrations. These results suggest that both trehalose and l-proline can act as CPAs by avoiding eutectic formation and inhibiting ice formation in normal saline for cell cryopreservation. It could be useful for CPA selection and designing in the future.

Catalytic Oxidation of NO over MnOx-CeO2 and MnOx-TiO2 Catalysts.

A series of MnOx-CeO2 and MnOx-TiO2 catalysts were prepared by a homogeneous precipitation method and their catalytic activities for the NO oxidation in the absence or presence of SO2 were evaluated. Results show that the optimal molar ratio of Mn/Ce and Mn/Ti are 0.7 and 0.5, respectively. The MnOx-CeO2 catalyst exhibits higher catalytic activity and better resistance to SO2 poisoning than the MnOx-TiO2 catalyst. On the basis of Brunauer-Emmett-Teller (BET), X-ray diffraction (XRD), and scanning transmission electron microscope with mapping (STEM-mapping) analyses, it is seen that the MnOx-CeO2 catalyst possesses higher BET surface area and better dispersion of MnOx over the catalyst than MnOx-TiO2 catalyst. X-ray photoelectron spectroscopy (XPS) measurements reveal that MnOx-CeO2 catalyst provides the abundance of Mn3+ and more surface adsorbed oxygen, and SO2 might be preferentially adsorbed to the surface of CeO2 to form sulfate species, which provides a protection of MnOx active sites from being poisoned. In contrast, MnOx active sites over the MnOx-TiO2 catalyst are easily and quickly sulfated, leading to rapid deactivation of the catalyst for NO oxidation. Furthermore, temperature programmed desorption with NO and O2 (NO + O2-TPD) and in situ diffuse reflectance infrared transform spectroscopy (in situ DRIFTS) characterizations results show that the MnOx-CeO2 catalyst displays much stronger ability to adsorb NOx than the MnOx-TiO2 catalyst, especially after SO2 poisoning.