A guidebook of spatial transcriptomic technologies, data resources and analysis approaches.

Advances in transcriptomic technologies have deepened our understanding of the cellular gene expression programs of multicellular organisms and provided a theoretical basis for disease diagnosis and therapy. However, both bulk and single-cell RNA sequencing approaches lose the spatial context of cells within the tissue microenvironment, and the development of spatial transcriptomics has made overall bias-free access to both transcriptional information and spatial information possible. Here, we elaborate development of spatial transcriptomic technologies to help researchers select the best-suited technology for their goals and integrate the vast amounts of data to facilitate data accessibility and availability. Then, we marshal various computational approaches to analyze spatial transcriptomic data for various purposes and describe the spatial multimodal omics and its potential for application in tumor tissue. Finally, we provide a detailed discussion and outlook of the spatial transcriptomic technologies, data resources and analysis approaches to guide current and future research on spatial transcriptomics.

Electron transfer from Geobacter sulfurreducens to mixed methanogens improved methane production with feedstock gases of H2 and CO2.

In order to solve problems of poor utilization of H2 and CO2 in biomethane conversion with mixed methanogens due to multi-channel competition and nondirectional electron transfer, Geobacter sulfurreducens were cocultured with mixed methanogens to promote oriented metabolic pathway of H2 and CO2 to produce CH4. When inoculation volume ratio of G. sulfurreducens to mixed methanogens was 2:4, CH4 yield increased to 0.24 mL/ml H2 (close to the maximum theoretical yield of 0.25 mL/ml H2) and conversion efficiency of H2 to CH4 increased from 72 to 96%. Electrochemical detection and three-dimensional fluorescence spectra showed that the co-culture system had an increased metabolic capacity and spectral intensity of fulvic acid-like compounds was enhanced, which mediated direct interspecific electron transfer to produce CH4. The 16S rRNA gene sequencing showed that relative abundance of G. sulfurreducens and Methanoculleus increased, indicating an established syntrophic relationship between G. sulfurreducens and Methanoculleus.

Microbial electrochemical degradation of lipids for promoting methane production in anaerobic digestion.

In order to solve problems of low methane production from lipids in anaerobic digestion, microbial electrochemical degradation was proposed to promote methane yield of glycerol trioleate (a typical lipid component of food waste). The beta-oxidation of lipids was strengthened with an applied voltage to promote electron transfer and anaerobic digestion. SEM images showed that a lot of spherical and rod-shaped microbes adhered to electrode surfaces. Cyclic voltammetry showed that electron transfer rate constant at 0.8 V was 14.4-fold that at 0 V. Three-dimensional fluorescence spectroscopy showed that small organic degraded molecules were used more efficiently in anaerobic digestion. The methane yield of glycerol trioleate increased to 791.6 mL/g-TVS (at 0.8 V), while methane production peak rate increased to 26.8 mL/g-TVS/d with a shortened peak time to 24th day. The overall energy conversion efficiency in methane production increased from 53.6 to 60.1% due to microbial electrochemical degradation of lipids.

A sodium percarbonate/ultraviolet system generated free radicals for degrading capsaicin to alleviate inhibition of methane production during anaerobic digestion of lipids and food waste.

To alleviate inhibition of anaerobic digestion caused by capsaicin, which is easily soluble in the lipid components of food waste (FW), an advanced oxidation process with sodium percarbonate/ultraviolet (SPC/UV) was used to generate free radicals for degrading capsaicin and recovering methane production. Free radical sweeping showed that the free radicals OH, O2- and CO3- worked together to degrade capsaicin. Gas chromatography-mass spectrometry showed that capsaicin likely had four degradation pathways via conversion into benzoquinone, and finally into carbon dioxide and water. The degradation rate of capsaicin in lipids increased from 62.2% to 96.0% when the SPC concentration increased from 2 mmol/L to 32 mmol/L (UV intensity = 20.66 mW/cm2). The degradation rate increased from 70.9% to 94.6% when the UV intensity increased from 20.66 mW/cm2 to 46.86 mW/cm2 (SPC concentration = 4 mmol/L). The subsequent products after capsaicin degradation were subjected to anaerobic digestion either directly or by adding FW. The reduced intracellular oxidative kinases of anaerobic digestion microorganisms recovered the CH4 yield from 27.2 mL/g-total volatile solids (TVS) with capsaicin to 311.2 mL/g-TVS with degraded capsaicin, which was 40.7% that of the control group (765.3 mL/g-TVS without capsaicin). After adding FW, the CH4 yield of SPC/UV degradation effluent was 504.1 mL/g-TVS, which was 82.6% that of the control group (610.4 mL/g-TVS).

Inhibition of N-Vanillylnonanamide in anaerobic digestion of lipids in food waste: Microorganisms damage and blocked electron transfer.

To study the inhibited degradation metabolism and anaerobic digestion of typical lipids in food waste, an artificially produced capsaicin, N-Vanillylnonanamide, a typical soluble component in waste lipids, was added to a glycerol trioleate anaerobic digestion system. The microorganisms damage and blocked electron transfer caused by N-Vanillylnonanamide during anaerobic digestion were further clarified. Scanning electron microscopy and transmission electron microscopy images demonstrated that N-Vanillylnonanamide (≥4 wt%) structurally damaged microorganisms via cell membrane breakage, which impair their function. N-Vanillylnonanamide inhibited the activities of the key enzyme CoA, AK, F420, and CoM, which are relevant for both degradation metabolism and anaerobic digestion. 16S rRNA analysis showed that dominant bacterial and archaeal communities markedly decreased after anaerobic digestion of glycerol trioleate with N-Vanillylnonanamide (≥4 wt%). For example, the proportion of Methanosarcina decreased from 30 % to 6 %. Current-voltage curves indicated that the electron transfer rate in the community of microorganisms decreased by 99 % from 4.67 × 10-2 to 5.66 × 10-4 s-1 in response to N-Vanillylnonanamide (40 wt%). The methane yield during anaerobic digestion of glycerol trioleate decreased by 84.0 % from 780.21-142.10 mL/g-total volatile solids with N-Vanillylnonanamide (40 wt%).

Nanoscale zero-valent iron improved lactic acid degradation to produce methane through anaerobic digestion.

Serious inhibition of methane production in an anaerobic digestion (AD) system can be caused by propionic acid, which is derived from lactic acid degradation. Nanoscale zero-valent iron (nZVI) was used in this study to improve conversion of propionic acid into acetic acid, thereby promoting methane production. The methane yield was markedly enhanced when nZVI concentration increased from 0 to 2 g/L; however, it decreased when nZVI concentration further increased to 8 g/L. At an nZVI concentration of 2 g/L, the methane yield increased by 37% from 398.5 to 546.4 mL CH4/g TVS. The abundance of Candidatus Cloacamonas in the bacterial community increased from 2.17% to 3.78%, which facilitated conversion of propionic acid into acetic acid. Meanwhile, the abundances of Methanomassiliicoccus and Methanosarcina in archaeal community increased, which was beneficial to methane production. Cyclic voltammetry showed that the electron transfer coefficient in the AD system increased from 0.029 to 0.034 s-1.

Improving CH4 production and energy conversion from CO2 and H2 feedstock gases with mixed methanogenic community over Fe nanoparticles.

To achieve methanogenic community optimization and improve the conversion efficiency of CO2 to CH4, Fe nanoparticles were used to promote the Methanothermobacter abundance in methanogens, which significantly increased the conversion efficiency of CO2 and H2 feedstock gases to CH4 product. High-throughput 16S rRNA gene sequencing analysis revealed that Methanothermobacter abundance markedly increased from 7 to 16% when the Fe nanoparticles concentration increased from 0 to 1.5 g/LR (the working volume in the bioreactor). Therefore, the CH4 yield significantly promoted from 0.105 to 0.186 L/LR. However, when the Fe nanoparticles concentration was further increased to 2 g/LR, methanogenesis was inhibited due to toxic effects. The electron transfer constant kapp of anaerobic sludge increased by 32.8-fold to 5.77 × 10-2 s-1 when the Fe nanoparticles concentration increased from 0 to 1.5 g/LR, which significantly promoted carbon conversion efficiency from 52.9 to 92.9% and energy conversion efficiency from 46.3 to 76.9%.

Hydrothermal alkali pretreatment contributes to fermentative methane production of a typical lipid from food waste through co-production of hydrogen with methane.

In order to relieve the suppression problems of methanogenesis with microorganisms surrounded by undegraded lipids in food waste, hydrothermal alkali pretreatment was utilized to degrade lipids for promoted methane production through the co-production process of hydrogen with methane. GC-MS results demonstrated that oleic acids and hexadecanoic acids derived from degraded glycerol trioleate increased (from 43.29% to 58.22%, and from 1.06% to 8.25%, respectively) when the pretreatment temperature was increased from 160 °C to 220 °C. SEM, TEM and FTIR analyses showed that the pre-treatment at 220 °C effectively degraded 87.56% of glycerol trioleate and drastically relieved the covering of methanogens by non-degraded lipids. The methane yield and the production peak rate of glycerol trioleate also increased (from 636.85 to 877.47 mL CH4/g-total volatile solid (VS), and from 32.60 to 51.22 mL CH4/g-VS/d, respectively), which led to an increased energy conversion efficiency from 48.05% to 66.21% through the co-production of hydrogen with methane.

Investigating hydrothermal pretreatment of food waste for two-stage fermentative hydrogen and methane co-production.

The growing amount of food waste (FW) in China poses great pressure on the environment. Complex solid organics limit the hydrolysis of FW, hence impairing anaerobic digestion. This study employed hydrothermal pretreatment (HTP) to facilitate the solubilization of FW. When HTP temperature increased from 100 to 200°C, soluble carbohydrate content first increased to a peak at 140°C and then decreased, whereas total carbohydrate content was negatively correlated with increasing temperature due to the enhanced degradation and Maillard reactions. Protein solubilization was dramatically promoted after HTP, whereas protein degradation was negligibly enhanced. The hydrogen and methane yields from hydrothermally pretreated FW under the optimum condition (140°C, 20min) through two-stage fermentation were 43.0 and 511.6mL/g volatile solids, respectively, resulting in an energy conversion efficiency (ECE) of 78.6%. The ECE of pretreated FW was higher than that of untreated FW by 31.7%.