TRAPPC6B biallelic variants cause a neurodevelopmental disorder with TRAPP II and trafficking disruptions.

Highly conserved transport protein particle (TRAPP) complexes regulate subcellular trafficking pathways. Accurate protein trafficking has been increasingly recognized to be critically important for normal development, particularly in the nervous system. Variants in most TRAPP complex subunits have been found to lead to neurodevelopmental disorders with diverse but overlapping phenotypes. We expand on limited prior reports on TRAPPC6B with detailed clinical and neuroradiologic assessments, and studies on mechanisms of disease, and new types of variants. We describe 29 additional patients from 18 independent families with biallelic variants in TRAPPC6B. We identified seven homozygous nonsense (n = 12 patients) and eight canonical splice-site variants (n = 17 patients). In addition, we identified one patient with compound heterozygous splice-site/missense variants with a milder phenotype and one patient with homozygous missense variants. Patients displayed non-progressive microcephaly, global developmental delay/intellectual disability, epilepsy and absent expressive language. Movement disorders including stereotypies, spasticity and dystonia were also observed. Brain imaging revealed reductions in cortex, cerebellum and corpus callosum size with frequent white matter hyperintensity. Volumetric measurements indicated globally diminished volume rather than specific regional losses. We identified a reduced rate of trafficking into the Golgi apparatus and Golgi fragmentation in patient-derived fibroblasts that was rescued by wild-type TRAPPC6B. Molecular studies revealed a weakened interaction between mutant TRAPPC6B (c.454C>T, p.Q152*) and its TRAPP binding partner TRAPPC3. Patient-derived fibroblasts from the TRAPPC6B (c.454C>T, p.Q152*) variant displayed reduced levels of TRAPPC6B as well as other TRAPP II complex-specific members (TRAPPC9 and TRAPPC10). Interestingly, the levels of the TRAPPC6B homologue TRAPPC6A were found to be elevated. Moreover, co-immunoprecipitation experiments showed that TRAPPC6A co-precipitates equally with TRAPP II and TRAPP III, while TRAPPC6B co-precipitates significantly more with TRAPP II, suggesting enrichment of the protein in the TRAPP II complex. This implies that variants in TRAPPC6B may preferentially affect TRAPP II functions compared to TRAPP III functions. Finally, we assessed phenotypes in a Drosophila TRAPPC6B-deficiency model. Neuronal TRAPPC6B knockdown impaired locomotion and led to wing posture defects, supporting a role for TRAPPC6B in neuromotor function. Our findings confirm the association of damaging biallelic TRAPPC6B variants with microcephaly, intellectual disability, language impairments, and epilepsy. A subset of patients also exhibited dystonia and/or spasticity with impaired ambulation. These features overlap with disorders arising from pathogenic variants in other TRAPP subunits, particularly components of the TRAPP II complex. These findings suggest that TRAPPC6B is essential for brain development and function, and TRAPP II complex activity may be particularly relevant for mediating this function.

Colonization ability and uniformity of resources and environmental factors determine biological homogenization of soil protists in human land-use systems.

Humans have substantially transformed the global land surface, resulting in the decline in variation in biotic communities across scales, a phenomenon known as "biological homogenization." However, different biota are affected by biological homogenization to varying degrees, but this variation and the underlying mechanisms remain little studied, particularly in soil systems. To address this topic, we used metabarcoding to investigate the biogeography of soil protists and their prey/hosts (prokaryotes, fungi, and meso- and macrofauna) in three human land-use ecosystem types (farmlands, residential areas, and parks) and natural forest ecosystems across subtropical and temperate regions in China. Our results showed that the degree of community homogenization largely differed between taxa and functional groups of soil protists, and was strongly and positively linked to their colonization ability of human land-use systems. Removal analysis showed that the introduction of widespread, generalist taxa (OTUs, operational taxonomic units) rather than the loss of narrow-ranged, specialist OTUs was the major cause of biological homogenization. This increase in generalist OTUs seemingly alleviated the negative impact of land use on specialist taxa, but carried the risk of losing functional diversity. Finally, homogenization of prey/host biota and environmental conditions were also important drivers of biological homogenization in human land-use systems, with their importance being more pronounced in phagotrophic than parasitic and phototrophic protists. Overall, our study showed that the variation in biological homogenization strongly depends on the colonization ability of taxa in human land-use systems, but is also affected by the homogenization of resources and environmental conditions. Importantly, biological homogenization is not the major cause of the decline in the diversity of soil protists, and conservation and study efforts should target at taxa highly sensitive to local extinction, such as parasites.

Diversity and Activity of Soil N2O-Reducing Bacteria Shaped by Urbanization.

Nitrous oxide (N2O) is a potent greenhouse gas with various production pathways. N2O reductase (N2OR) is the primary N2O sink, but the distribution of its gene clades, typically nosZI and atypically nosZII, along urbanization gradients remains poorly understood. Here we sampled soils from forests, parks, and farmland across eight provinces in eastern China, using high-throughput sequencing to distinguish between two N2O-reducing bacteria clades. A deterministic process mainly determined assemblies of the nosZI communities. Homogeneous selection drove nosZI deterministic processes, and both homogeneous and heterogeneous selection influenced nosZII. This suggests nosZII is more sensitive to environmental changes than nosZI, with significant changes in community structure over time or space. Ecosystems with stronger anthropogenic disturbance, such as urban areas, provide diverse ecological niches for N2O-reducing bacteria (especially nosZII) to adapt to environmental fluctuations. Structural equation modeling (SEM) and correlation analyses revealed that pH significantly influences the community composition of both N2O-reducing bacteria clades. This study underscores urbanization's impact on N2O-reducing bacteria in urban soils, highlighting the importance of nosZII and survival strategies. It offers novel insights into the role of atypical denitrifiers among N2O-reducing bacteria, underscoring their potential ecological importance in mitigating N2O emissions from urban soils.

Lipid Metabolites as Potential Regulators of the Antibiotic Resistome in Tetramorium caespitum.

Antibiotic resistance genes (ARGs) are ancient but have become a modern critical threat to health. Gut microbiota, a dynamic reservoir for ARGs, transfer resistance between individuals. Surveillance of the antibiotic resistome in the gut during different host growth phases is critical to understanding the dynamics of the resistome in this ecosystem. Herein, we disentangled the ARG profiles and the dynamic mechanism of ARGs in the egg and adult phases of Tetramorium caespitum. Experimental results showed a remarkable difference in both gut microbiota and gut resistome with the development of T. caespitum. Meta-based metagenomic results of gut microbiota indicated the generalizability of gut antibiotic resistome dynamics during host development. By using Raman spectroscopy and metabolomics, the metabolic phenotype and metabolites indicated that the biotic phase significantly changed lipid metabolism as T. caespitum aged. Lipid metabolites were demonstrated as the main factor driving the enrichment of ARGs in T. caespitum. Cuminaldehyde, the antibacterial lipid metabolite that displayed a remarkable increase in the adult phase, was demonstrated to strongly induce ARG abundance. Our findings show that the gut resistome is host developmental stage-dependent and likely modulated by metabolites, offering novel insights into possible steps to reduce ARG dissemination in the soil food chain.

[Detection model of atrial fibrillation based on multi-branch and multi-scale convolutional networks].

Atrial fibrillation (AF) is a life-threatening heart condition, and its early detection and treatment have garnered significant attention from physicians in recent years. Traditional methods of detecting AF heavily rely on doctor's diagnosis based on electrocardiograms (ECGs), but prolonged analysis of ECG signals is very time-consuming. This paper designs an AF detection model based on the Inception module, constructing multi-branch detection channels to process raw ECG signals, gradient signals, and frequency signals during AF. The model efficiently extracted QRS complex and RR interval features using gradient signals, extracted P-wave and f-wave features using frequency signals, and used raw signals to supplement missing information. The multi-scale convolutional kernels in the Inception module provided various receptive fields and performed comprehensive analysis of the multi-branch results, enabling early AF detection. Compared to current machine learning algorithms that use only RR interval and heart rate variability features, the proposed algorithm additionally employed frequency features, making fuller use of the information within the signals. For deep learning methods using raw and frequency signals, this paper introduced an enhanced method for the QRS complex, allowing the network to extract features more effectively. By using a multi-branch input mode, the model comprehensively considered irregular RR intervals and P-wave and f-wave features in AF. Testing on the MIT-BIH AF database showed that the inter-patient detection accuracy was 96.89%, sensitivity was 97.72%, and specificity was 95.88%. The proposed model demonstrates excellent performance and can achieve automatic AF detection.

Rod-cone dystrophy in an adult with GNB1-related disorder: An expansion of the phenotype and natural history.

GNB1-related disorder is characterized by intellectual disability, abnormal tone, and other variable neurologic and systemic features. GNB1 encodes the ß1 subunit of the heterotrimeric G-protein, a complex with a key role in signal transduction. Consistent with its particularly high expression in rod photoreceptors, Gß1 forms a subunit of retinal transducin (Gαtß1γ1 ), which mediates phototransduction. In mice, GNB1 haploinsufficiency has been associated with retinal dystrophy. In humans, however, although vision and eye movement abnormalities are common in individuals with GNB1-related disorder, rod-cone dystrophy is not yet an established feature of this condition. We expand the phenotype of GNB1-related disorder with the first confirmed report of rod-cone dystrophy in an affected individual, and contribute to a further understanding of the natural history of this condition in a mildly affected 45-year-old adult.

Temporal dynamics of soil bacterial network regulate soil resistomes.

Soil bacteria are diverse and form complicated ecological networks through various microbial interactions, which play important roles in soil multi-functionality. However, the seasonal effects on the bacterial network, especially the relationship between bacterial network topological features and soil resistomes remains underexplored, which impedes our ability to unveil the mechanisms of the temporal-dynamics of antibiotic resistance genes (ARGs). Here, a field investigation was conducted across four seasons at the watershed scale. We observed significant seasonal variation in bacterial networks, with lower complexity and stability in autumn, and a wider bacterial community niche in summer. Similar to bacterial communities, the co-occurrence networks among ARGs also shift with seasonal change, particularly with respect to the topological features of the node degree, which on average was higher in summer than in the other seasons. Furthermore, the nodes with higher betweenness, stress, degree, and closeness centrality in the bacterial network showed strong relationships with the 10 major classes of ARGs. These findings highlighted the changes in the topological properties of bacterial networks that could further alter antibiotic resistance in soil. Together, our results reveal the temporal dynamics of bacterial ecological networks at the watershed scale, and provide new insights into antibiotic resistance management under environmental changes.

Effects of Low Temperature on Pedicel Abscission and Auxin Synthesis Key Genes of Tomato.

Cold stress usually causes the abscission of floral organs and a decline in fruit setting rate, seriously reducing tomato yield. Auxin is one of the key hormones that affects the abscission of plant floral organs; the YUCCA (YUC) family is a key gene in the auxin biosynthesis pathway, but there are few research reports on the abscission of tomato flower organs. This experiment found that, under low temperature stress, the expression of auxin synthesis genes increased in stamens but decreased in pistils. Low temperature treatment decreased pollen vigor and pollen germination rate. Low night temperature reduced the tomato fruit setting rate and led to parthenocarpy, and the treatment effect was most obvious in the early stage of tomato pollen development. The abscission rate of tomato pTRV-Slfzy3 and pTRV-Slfzy5 silenced plants was higher than that of the control, which is the key auxin synthesis gene affecting the abscission rate. The expression of Solyc07g043580 was down-regulated after low night temperature treatment. Solyc07g043580 encodes the bHLH-type transcription factor SlPIF4. It has been reported that PIF4 regulates the expression of auxin synthesis and synthesis genes, and is a key protein in the interaction between low temperature stress and light in regulating plant development.

SCN2A-related epilepsy of infancy with migrating focal seizures: report of a variant with apparent gain- and loss-of-function effects.

SCN2A encodes a voltage-gated sodium channel (NaV1.2) expressed throughout the central nervous system in predominantly excitatory neurons. Pathogenic variants in SCN2A are associated with epilepsy and neurodevelopmental disorders. Genotype-phenotype correlations have been described, with loss-of-function variants typically being associated with neurodevelopmental delay and later-onset seizures, whereas gain-of-function variants more often result in early infantile-onset epilepsy. However, the true electrophysiological effects of most disease-causing SCN2A variants have yet to be characterized. We report an infant who presented with migrating focal seizures in the neonatal period. She was found to have a mosaic c.2635G>A, p.Gly879Arg variant in SCN2A. Voltage-clamp studies of the variant expressed on adult and neonatal NaV1.2 isoforms demonstrated a mixed gain and loss of function, with predominantly a loss-of-function effect with reduced cell surface expression and current density. Additional small electrophysiological alterations included a decrease in the voltage dependence of activation and an increase in the voltage dependence of inactivation. This finding of a predominantly loss-of-function effect was unexpected, as the infant's early epilepsy onset would have suggested a predominantly gain-of-function effect. This case illustrates that our understanding of genotype-phenotype correlations is still limited and highlights the complexity of the underlying electrophysiological effects of SCN2A variants.NEW & NOTEWORTHY Voltage-gated sodium channels play an important role in the central nervous system, mutations in which have been reported to be responsible for epilepsy. We report here an infant presenting with epilepsy of infancy with migrating focal seizures (EIMFS) in the neonatal period with a mosaic c.2635G>A, resulting in a p.Gly879Arg missense mutation on the SCN2A gene encoding NaV1.2 sodium channels. Biophysical characterization of this variant revealed a mixture of gain- and loss-of-function effects.

Microbial Multitrophic Communities Drive the Variation of Antibiotic Resistome in the Gut of Soil Woodlice (Crustacea: Isopoda).

Multitrophic communities inhabit in soil faunal gut, including bacteria, fungi, and protists, which have been considered a hidden reservoir for antibiotic resistance genes (ARGs). However, there is a dearth of research focusing on the relationships between ARGs and multitrophic communities in the gut of soil faunas. Here, we studied the contribution of multitrophic communities to variations of ARGs in the soil woodlouse gut. The results revealed diverse and abundant ARGs in the woodlouse gut. Network analysis further exhibited strong connections between key ecological module members and ARGs, suggesting that multitrophic communities in the keystone ecological cluster may play a pivotal role in the variation of ARGs in the woodlouse gut. Moreover, long-term application of sewage sludge significantly altered the woodlice gut resistome and interkingdom communities. The variation portioning analysis indicated that the fungal community has a greater contribution to variations of ARGs than bacterial and protistan communities in the woodlice gut after long-term application of sewage sludge. Together, our results showed that changes in gut microbiota associated with agricultural practices (e.g., sewage sludge application) can largely alter the gut interkingdom network in ecologically relevant soil animals, with implications for antibiotic resistance, which advances our understanding of the microecological drivers of ARGs in terrestrial ecosystem.

Does Plant Identity Affect the Dispersal of Resistomes Above and Below Ground?

Resistomes are ubiquitous in natural environments. Previous studies have shown that both the plant phyllosphere and soil-borne nematodes were reservoirs of above- and below-ground resistomes, respectively. However, the influence of plant identity on soil, nematode, and phyllosphere resistomes remains unclear. Here, a microcosm experiment was used to explore the characteristics of bacterial communities and resistomes in soil, nematode, and phyllosphere associated with six different plant identities (Lactuca sativa, Cichorium endivia, Allium fistulosum, Coriandrum sativum, Raphanus sativus, and Mesembryanthemum crystallinum). A total of 222 antibiotic resistance genes (ARGs) and 7 mobile genetic elements (MGEs) were detected by high-throughput quantitative PCR from all samples. Plant identity not only significantly affected the diversity of resistomes in soil, nematode, and phyllosphere but also influenced the abundance of resistomes in nematodes. Shared bacteria and resistomes indicated a possible pathway of resistomes transfer through the soil-nematode-phyllosphere system. Structural equation models revealed that plant identity had no direct effect on phyllosphere ARGs, but altered indirectly through complex above- and below-ground interactions (soil-plant-nematode trophic transfer). Results also showed that bacteria and MGEs were key factors driving the above- and below-ground flow of resistomes. The study extends our knowledge about the top-down and bottom-up dispersal patterns of resistomes.

Denitrification and N2O Emission in Estuarine Sediments in Response to Ocean Acidification: From Process to Mechanism.

Global estuarine ecosystems are experiencing severe nitrogen pollution and ocean acidification (OA) simultaneously. Sedimentary denitrification is an important way of reactive nitrogen removal but at the same time leads to the emission of large amounts of nitrous oxide (N2O), a potent greenhouse gas. It is known that OA in estuarine regions could impact denitrification and N2O production; however, the underlying mechanism is still underexplored. Here, sediment incubation and pure culture experiments were conducted to explore the OA impacts on microbial denitrification and the associated N2O emissions in estuarine sediments. Under neutral (in situ) conditions, fungal N2O emission dominated in the sediment, while the bacterial and fungal sources had a similar role under acidification. This indicated that acidification decreased the sedimentary fungal denitrification and likely inhibited the activity of fungal denitrifiers. To explore molecular mechanisms, a denitrifying fungal strain of Penicillium janthinellum was isolated from the sediments. By using deuterium-labeled single-cell Raman spectroscopy and isobaric tags for relative and absolute quantitation proteomics, we found that acidification inhibited electron transfers in P. janthinellum and downregulated expressions of the proteins related to energy production and conservation. Two collaborative pathways of energy generation in the P. janthinellum were further revealed, that is, aerobic oxidative phosphorylation and TCA cycle and anoxic pyruvate fermentation. This indicated a distinct energy supply strategy from bacterial denitrification. Our study provides insights into fungi-mediated nitrogen cycle in acidifying aquatic ecosystems.

Fates of Antibiotic Resistance Genes in the Gut Microbiome from Different Soil Fauna under Long-Term Fertilization.

Applying organic fertilizers has been well documented to facilitate the dissemination of antibiotic resistance genes (ARGs) in soil ecosystems. However, the role of soil fauna in this process has been seldom addressed, which hampers our ability to predict the fate of and to manage the spread of ARGs. Here, using high-throughput quantitative polymerase chain reaction (HT-qPCR), we examined the effect of long-term (5-, 8-, and 10-year) fertilization treatments (control, inorganic fertilizers, and mixed fertilizers) on the transfer of ARGs between soil, nematodes, and earthworms. We found distinct fates for ARGs in the nematodes and earthworms, with the former having higher enriched levels of ARGs than the latter. Fertilization impacted the number and abundance of ARGs in soil, and fertilization duration altered the composition of ARGs. Shared ARGs among soil, nematodes, and earthworm guts supported by a fast expectation-maximization microbial source tracking analysis demonstrated the trophic transfer potential of ARGs through this short soil food chain. The transfer of ARGs was reduced by fertilization duration, which was mainly ascribed to the reduction of ARGs in the earthworm gut microbiota. This study identified the transfer of ARGs in the soil-nematode-earthworm food chain as a potential mechanism for a wider dissemination of ARGs in the soil ecosystem.

How can fertilization regimes and durations shape earthworm gut microbiota in a long-term field experiment?

The positive roles of earthworms on soil functionality has been extensively documented. The capacity of the earthworm gut microbiota on decomposition and nutrient cycling under long-term fertilization in field conditions has rarely been studied. Here, we report the structural, taxonomic, and functional responses of Eisenia foetida and Pheretima guillelmi gut microbiota to different fertilization regimes and durations using 16S rRNA gene-based Illumina sequencing and high-throughput quantitative PCR techniques. Our results revealed that the core gut microbiota, especially the fermentative bacteria were mainly sourced from the soil, but strongly stimulated with species-specificity, potential benefits for the host and soil health. The functional compositions of gut microbiota were altered by fertilization with fertilization duration being more influential than fertilization regimes. Moreover, the combination of organic and inorganic fertilization with the longer duration resulted in a higher richness and connectivity in the gut microbiota, and also their functional potential related to carbon (C), nitrogen, and phosphorus cycling, particularly the labile C decomposition, denitrification, and phosphate mobilization. We also found that long-term inorganic fertilization increased the abundance of pathogenic bacteria in the P. guillelmi gut. This study demonstrates that understanding earthworm gut microbiota can provide insights into how agricultural practices can potentially alter soil ecosystem functions through the interactions between soil and earthworm gut microbiotas.

A diagnostic approach to syndromic retinal dystrophies with intellectual disability.

Inherited retinal dystrophies are a group of monogenic disorders that, as a whole, contribute significantly to the burden of ocular disease in both pediatric and adult patients. In their syndromic forms, retinal dystrophies can be observed in association with intellectual disability, frequently alongside other systemic manifestations. There are now over 80 genes implicated in syndromic retinal dystrophies with intellectual disability. Identifying and accurately characterizing these disorders allows the clinician to narrow the differential diagnosis, evaluate for relevant associated features, arrive at a timely and accurate diagnosis, and address both sight-threatening ocular manifestations and morbidity-causing systemic manifestations. The co-occurrence of retinal dystrophy and intellectual disability in an individual can be challenging to investigate, diagnose, and counsel given the considerable phenotypic and genotypic heterogeneity that exists within this broad group of disorders. We performed a review of the current literature and propose an algorithm to facilitate the evaluation, and clinical and mechanistic classification, of these individuals.

Pathogenic effects of inhibition of mTORC1/STAT3 axis facilitates Staphylococcus aureus-induced pyroptosis in human macrophages.

BACKGROUND: Pyroptosis is a recently identified pathway of caspase-mediated cell death in response to microbes, lipopolysaccharide, or chemotherapy in certain types of cells. However, the mechanism of how pyroptosis is regulated is not well-established. METHODS: Herein, the intracellular bacteria were detected by staining and laser confocal microscopy and TEM. Live/dead cell imaging assay was used to examine macrophage death. Western blot and immunohistochemical staining were used to examine the protein changes. IFA was used to identify typical budding vesicles of pyroptosis and the STAT3 nuclear localization. SEM was used to observe the morphological characteristics of pyroptosis. ELISA was used to detect the level of inflammatory cytokines. Pyroptosis was filmed in macrophages by LSCM. RESULTS: S. aureus was internalized by human macrophages. Intracellular S. aureus induced macrophage death. S. aureus invasion increased the expression of NLRP3, Caspase1 (Casp-1 p20) and the accumulation of GSDMD-NT, GSDMD-NT pore structures, and the release of IL-1ß and IL-18 in macrophages. Macrophages pyroptosis induced by S. aureus can be abrogated by blockage of S. aureus phagocytosis. The pyroptosic effect by S. aureus infection was promoted by either rapamycin or Stattic, a specific inhibitor for mTORC1 or STAT3. Inhibition of mTORC1 or STAT3 induced pyroptosis. mTORC1 regulated the pyroptosic gene expression through governing the nuclear localization of STAT3. mTORC1/STAT3 axis may play a regulatory role in pyroptosis within macrophages. CONCLUSIONS: S. aureus infection induces human macrophage pyroptosis, inhibition of mTORC1/STAT3 axis facilitates S. aureus-induced pyroptosis. mTORC1 and STAT3 are associated with pyroptosis. Our findings demonstrate a regulatory function of the mTORC1/STAT3 axis in macrophage pyroptosis, constituting a novel mechanism by which pyroptosis is regulated in macrophages. Video Abstract Macrophages were infected with S. aureus for 3 h (MOI 25:1), and pyroptosis was filmed in macrophages by laser confocal microscopy. A representative field was recorded. Arrow indicates lysing dead cell.

Trophic Transfer of Antibiotic Resistance Genes in a Soil Detritus Food Chain.

The presence and spread of antibiotic resistance genes (ARGs) are causing substantial global public concern; however, the dispersal of ARGs in the food chain is poorly understood. Here, we experimented with a soil collembolan ( Folsomia candida)-predatory mite ( Hypoaspis aculeifer) model food chain to study trophic transfer of ARGs in a manure-contaminated soil ecosystem. Our results showed that manure amendment of soil could significantly increase ARGs in the soil collembolan microbiome. With the ARGs in the prey collembolan microbiome increasing, an increase in ARGs in the predatory mite microbiome was also observed, especially for three high abundant ARGs ( blaSHV, fosX and aph6ia). Three unique ARGs were transferred into the microbiome of the predatory mite from manure amended soil via the prey collembolan ( aac(6' )-lb(akaaacA4), yidY_mdtL and tolC). Manure amendment altered the composition and structure and reduced the diversity of the microbiomes of the prey collembolan and the predatory mite. We further reveal that bacterial communities and mobile genetic elements were two important drivers for the trophic transfer of ARGs, not just for ARGs distribution in the samples. These findings suggest that the importance of food chain transmission of ARGs for the dispersal of resistance genes in soil ecosystems may be underestimated.

Mobile Incubator for Iron(III) Reduction in the Gut of the Soil-Feeding Earthworm Pheretima guillelmi and Interaction with Denitrification.

Diets of soil-feeding earthworms contain abundant nitrate and iron(III) oxides, which are potential electron acceptors for mineralization of organic compounds. The earthworm gut provides an ideal habitat for ingested iron(III)-reducing microorganisms. However, little is known about iron(III) reduction and its interaction with other processes in the guts of earthworms. Here, we determined the dynamics of iron(III) and revealed its interaction with the turnover of organic acids and nitrate in the gut of the earthworm Pheretima guillelmi. Samples from gut contents combined with anoxic incubation were used for chemical analysis and 16S rRNA based Illumina sequencing. Chemical analysis showed that higher ratios of iron(II)/iron(III), nitrite/nitrate, and more abundant organic acids were contained in the in vivo gut of the earthworm P. guillelmi than those in the in situ soil. A higher rate of iron(III) reduction was detected in treatments of microcosmic incubation with gut contents (IG gut) than that with soil (IG soil), and nitrate reduction occurred earlier than iron(III) reduction in both treatments. Potential iron(III) reducers were dominated by fermentative genera Clostridium, Bacillus, and Desulfotomaculum in the treatment of IG gut, while they were dominated by dissimilatory iron(III)-reducing genera Geobacter in the treatment of IG soil. The iron(III)-reducing microbial community shared several genera with denitrifers in the treatment of IG gut, revealing a close link between iron(III) reduction and denitrification in the gut of earthworms. Collectively, our findings demonstrated that iron(III) reduction occurred along the gut and provided novel insights into the great contribution of earthworm gut microbiota on Fe and the associated C and N cycling in soil environments.

RNA Stable Isotope Probing of Potential Feammox Population in Paddy Soil.

Anaerobic ammonium oxidation coupled to iron reduction (Feammox) is a recently discovered pathway contributing to nitrogen loss in various ecosystems such as paddy soils and sediments. However, little is known about the microbes driving Feammox in an agricultural ecosystem. Here, we demonstrated the occurrence of Feammox in paddy soils of Southern China using a 15N isotopic tracing technique, and examined the microbial communities associated with Feammox using RNA based stable isotope probing (RNA-SIP) combined with Illumina sequencing. Feammox was detected in all collected soils with direct N2 production as the dominant Feammox pathway. It was estimated that approximately 6.91% of the applied nitrogen fertilizers were lost through Feammox in the paddy soils. RNA-SIP results showed that the composition of enriched active microbial communities were dependent on soil properties, especially the soil pH and grain size. Geobacter were enriched in most soils across various properties. The abundance of enriched GOUTA19 were significantly higher in soils with low pH than those in soils with medium pH and high pH, and the relative abundance of active Nitrososphaeraceae and Pseudomonas only increased in soils with medium and high pH during 4-day of incubation. These results suggested Feammox is a ubiquitous and important process for N loss. Geobacter, GOUTA19, Nitrososphaeraceae and Pseudomonas were active during the incubation that favored Feammox and the growth of Feammox microbes, suggesting these microbes were potentially associated with Feammox in natural agricultural soils.

Adsorbed Sulfamethoxazole Exacerbates the Effects of Polystyrene (∼2 µm) on Gut Microbiota and the Antibiotic Resistome of a Soil Collembolan.

Microplastics pollution in the environment is now receiving worldwide attention; however, the effects of copollution of antibiotics and microplastics on the gut microbiome of globally distributed and functionally important nontarget soil animals remain poorly understood. We studied a model collembolan (Folsomia candida) and found that the ingestion of microplastics (polystyrene, 2-2.9 µm) substantially altered the gut microbiome, antibiotic resistance gene (ARG) profile, and the isotopic fractionation in the soil collembolan tissue. Importantly, collembolans exposed to polystyrene microplastics loaded with sulfamethoxazole (MA) presented a distinctive gut microbiome, ARG profile, and isotopic fractionation compared to those exposed to polystyrene alone (MH). We observed that the abundance of ARGs and mobile genetic elements (MGEs) in the MA-treated collembolan guts was significantly higher than in the MH and the control treatments. There were also strong interactions between the gut microbiome and ARGs in the collembolan guts. We further found that bacterial ß-diversity correlated significantly with the δ13C and δ15N values in collembolan body tissues. Together, our results indicate that changes in isotopic fractionation and ARG profiles in the collembolan were induced by the changes in gut microbiota and suggest that microplastics from diverse sources may have profound influences on soil fauna and soil food webs.