Pseudomonas aphyarum sp. nov., Pseudomonas fontis sp. nov., Pseudomonas idahonensis sp. nov. and Pseudomonas rubra sp. nov., isolated from in, and around, a rainbow trout farm.

During a large-scale bacterial culturing effort of biofilms in the vicinity of a rainbow trout aquaculture facility in Idaho, USA, 10 isolates were identified as having pathogen-inhibiting activity and were characterized further. These isolates were shown to be Gram-negative, rod-shaped bacteria belonging to the genus Pseudomonas. Whole-genome comparisons and multi-locus sequence analysis using four housekeeping genes (16S rRNA, gyrA, rpoB and rpoD) showed that these 10 isolates clustered into four distinct species groups. These comparisons also indicated that these isolates were below the established species cutoffs for the genus Pseudomonas. Further phenotypic characterization using API 20NE, API ZYM and Biolog GENIII assays and chemotaxonomic analysis of cellular fatty acids were carried out. Based on the genomic, physiological and chemotaxonomic properties of these isolates, we concluded that these strains composed four novel species of the genus Pseudomonas. The proposed names are as follows: Pseudomonas aphyarum sp. nov. consisting of strains ID233, ID386T and ID387 with ID386T (=DSM 114641T=ATCC TSD-305T) as the type strain; Pseudomonas rubra sp. nov. consisting of strains ID291T, ID609 and ID1025 with ID291T (=DSM 114640T=ATCC TSD-303T) as the type strain; Pseudomonas idahonensis sp. nov. consisting of strains ID357T and ID1048 with ID357T (=DSM 114609T=ATCC TSD-304T) as the type strain; and Pseudomonas fontis sp. nov. consisting of strains ID656T and ID681 with ID656T (=DSM 114610T=ATCC TSD-306T) as the type strain.

Systematic Detection of Large-Scale Multigene Horizontal Transfer in Prokaryotes.

Horizontal gene transfer (HGT) is central to prokaryotic evolution. However, little is known about the "scale" of individual HGT events. In this work, we introduce the first computational framework to help answer the following fundamental question: How often does more than one gene get horizontally transferred in a single HGT event? Our method, called HoMer, uses phylogenetic reconciliation to infer single-gene HGT events across a given set of species/strains, employs several techniques to account for inference error and uncertainty, combines that information with gene order information from extant genomes, and uses statistical analysis to identify candidate horizontal multigene transfers (HMGTs) in both extant and ancestral species/strains. HoMer is highly scalable and can be easily used to infer HMGTs across hundreds of genomes. We apply HoMer to a genome-scale data set of over 22,000 gene families from 103 Aeromonas genomes and identify a large number of plausible HMGTs of various scales at both small and large phylogenetic distances. Analysis of these HMGTs reveals interesting relationships between gene function, phylogenetic distance, and frequency of multigene transfer. Among other insights, we find that 1) the observed relative frequency of HMGT increases as divergence between genomes increases, 2) HMGTs often have conserved gene functions, and 3) rare genes are frequently acquired through HMGT. We also analyze in detail HMGTs involving the zonula occludens toxin and type III secretion systems. By enabling the systematic inference of HMGTs on a large scale, HoMer will facilitate a more accurate and more complete understanding of HGT and microbial evolution.

Detecting Flavobacterial Fish Pathogens in the Environment via High-Throughput Community Analysis.

Diseases caused by the fish pathogens Flavobacterium columnare and Flavobacterium psychrophilum are major contributors of preventable losses in the aquaculture industry. The persistent and difficult-to-control infections caused by these bacteria make timely intervention and prophylactic elimination of pathogen reservoirs important measures to combat these disease-causing agents. In this study, we present two independent assays for detecting these pathogens in a range of environmental samples. Natural water samples were inoculated with F. columnare and F. psychrophilum over 5 orders of magnitude, and pathogen levels were detected using Illumina MiSeq sequencing and droplet digital PCR. Both detection methods accurately identified pathogen-positive samples and showed good agreement in quantifying each pathogen. Additionally, the real-world application of these approaches was demonstrated using environmental samples collected at a rainbow trout (Oncorhynchus mykiss) aquaculture facility. These results show that both methods can serve as useful tools for surveillance efforts in aquaculture facilities, where the early detection of these flavobacterial pathogens may direct preventative measures to reduce disease occurrence. IMPORTANCE Early detection of a deadly disease outbreak in a population can be the difference between mass mortality or mitigated effects. In the present study, we evaluated and compared two molecular techniques for detecting economically impactful aquaculture pathogens. We demonstrate that one of these techniques, 16S rRNA gene sequencing using Illumina MiSeq technology, provides the ability to accurately detect two freshwater fish pathogens, F. columnare and F. psychrophilum, while simultaneously profiling the native microbial community. The second technique, droplet digital PCR, is commonly used for pathogen detection, and the results obtained using the assays we designed with this method served to validate those obtained using the MiSeq method. These two methods offer distinct advantages. The MiSeq method pairs pathogen detection and microbial community profiling to answer immediate and long-term fish health concerns, while the droplet digital PCR method provides fast and highly sensitive detection that is useful for surveillance and rapid clinical responses.

Gut Microbiota-Derived Short-Chain Fatty Acids Promote Poststroke Recovery in Aged Mice.

RATIONALE: The elderly experience profound systemic responses after stroke, which contribute to higher mortality and more severe long-term disability. Recent studies have revealed that stroke outcomes can be influenced by the composition of gut microbiome. However, the potential benefits of manipulating the gut microbiome after injury is unknown. OBJECTIVE: To determine if restoring youthful gut microbiota after stroke aids in recovery in aged subjects, we altered the gut microbiome through young fecal transplant gavage in aged mice after experimental stroke. Further, the effect of direct enrichment of selective bacteria producing short-chain fatty acids (SCFAs) was tested as a more targeted and refined microbiome therapy. METHODS AND RESULTS: Aged male mice (18-20 months) were subjected to ischemic stroke by middle cerebral artery occlusion. We performed fecal transplant gavage 3 days after middle cerebral artery occlusion using young donor biome (2-3 months) or aged biome (18-20 months). At day 14 after stroke, aged stroke mice receiving young fecal transplant gavage had less behavioral impairment, and reduced brain and gut inflammation. Based on data from microbial sequencing and metabolomics analysis demonstrating that young fecal transplants contained much higher SCFA levels and related bacterial strains, we selected 4 SCFA-producers (Bifidobacterium longum, Clostridium symbiosum, Faecalibacterium prausnitzii, and Lactobacillus fermentum) for transplantation. These SCFA-producers alleviated poststroke neurological deficits and inflammation, and elevated gut, brain and plasma SCFA concentrations in aged stroke mice. CONCLUSIONS: This is the first study suggesting that the poor stroke recovery in aged mice can be reversed via poststroke bacteriotherapy following the replenishment of youthful gut microbiome via modulation of immunologic, microbial, and metabolomic profiles in the host.

Macrobdella decora: Old World Leech Gut Microbial Community Structure Conserved in a New World Leech.

Leeches are found in terrestrial, aquatic, and marine habitats on all continents. Sanguivorous leeches have been used in medicine for millennia. Modern scientific uses include studies of neurons, anticoagulants, and gut microbial symbioses. Hirudo verbana, the European medicinal leech, maintains a gut community dominated by two bacterial symbionts, Aeromonas veronii and Mucinivorans hirudinis, which sometimes account for as much as 97% of the total crop microbiota. The highly simplified gut anatomy and microbiome of H. verbana make it an excellent model organism for studying gut microbial dynamics. The North American medicinal leech, Macrobdella decora, is a hirudinid leech native to Canada and the northern United States. In this study, we show that M. decora symbiont communities are very similar to those in H. verbana. We performed an extensive study using field-caught M. decora and purchased H. verbana from two suppliers. Deep sequencing of the V4 region of the 16S rRNA gene allowed us to determine that the core microbiome of M. decora consists of Bacteroides, Aeromonas, Proteocatella, and Butyricicoccus. The analysis revealed that the compositions of the gut microbiomes of the two leech species were significantly different at all taxonomic levels. The R2 value was highest at the genus and amplicon sequence variant (ASV) levels and much lower at the phylum, class, and order levels. The gut and bladder microbial communities were distinct. We propose that M. decora is an alternative to H. verbana for studies of wild-caught animals and provide evidence for the conservation of digestive-tract and bladder symbionts in annelid models.IMPORTANCE Building evidence implicates the gut microbiome in critical animal functions such as regulating digestion, nutrition, immune regulation, and development. Simplified, phylogenetically diverse models for hypothesis testing are necessary because of the difficulty of assigning causative relationships in complex gut microbiomes. Previous research used Hirudo verbana as a tractable animal model of digestive-tract symbioses. Our data show that Macrobdella decora may work just as well without the drawback of being an endangered organism and with the added advantage of easy access to field-caught specimens. The similarity of the microbial community structures of species from two different continents reveals the highly conserved nature of the microbial symbionts in sanguivorous leeches.

Lessons from Digestive-Tract Symbioses Between Bacteria and Invertebrates.

In most animals, digestive tracts harbor the greatest number of bacteria in the animal that contribute to its health: by aiding in the digestion of nutrients, provisioning essential nutrients and protecting against colonization by pathogens. Invertebrates have been used to enhance our understanding of metabolic processes and microbe-host interactions owing to experimental advantages. This review describes how advances in DNA sequencing technologies have dramatically altered how researchers investigate microbe-host interactions, including 16S rRNA gene surveys, metagenome experiments, and metatranscriptome studies. Advantages and challenges of each of these approaches are described herein. Hypotheses generated through omics studies can be directly tested using site-directed mutagenesis, and findings from transposon studies and site-directed experiments are presented. Finally, unique structural aspects of invertebrate digestive tracts that contribute to symbiont specificity are presented. The combination of omics approaches with genetics and microscopy allows researchers to move beyond correlations to identify conserved mechanisms of microbe-host interactions.

Effects of Host Species Identity and Diet on the Biodiversity of the Microbiota in Puerto Rican Bats.

Microbiota perform vital functions for their mammalian hosts, making them potential drivers of host evolution. Understanding effects of environmental factors and host characteristics on the composition and biodiversity of the microbiota may provide novel insights into the origin and maintenance of these symbiotic relationships. Our goals were to (1) characterize biodiversity of oral and rectal microbiota in bats from Puerto Rico; and (2) determine the effects of geographic location and host characteristics on that biodiversity. We collected bats and their microbiota from three sites, and used four metrics (species richness, Shannon diversity, Camargo evenness, Berger-Parker dominance) to characterize biodiversity. We quantified the relative importance of site, host sex, host species-identity, and host foraging-guild on biodiversity of the microbiota. Microbe biodiversity was highly variable among conspecifics. Geographical location exhibited consistent effects, whereas host sex did not. Within each host guild, host species exhibited consistent differences in biodiversity of oral microbiota and of rectal microbiota. Oral microbe biodiversity was indistinguishable between guilds, whereas rectal microbe biodiversity was significantly greater in carnivores than in herbivores. The high intraspecific and spatial variation in microbe biodiversity necessitate a relatively large number of samples to statistically isolate the effects of environmental or host characteristics on the microbiota. Species-specific biodiversity of oral microbiota suggests these communities are structured by direct interactions with the host immune system via epithelial receptors. In contrast, the number of microbial taxa that a host gut supports may be driven by host diet-diversity or composition.

Evaluation of strategies for the assembly of diverse bacterial genomes using MinION long-read sequencing.

BACKGROUND: Short-read sequencing technologies have made microbial genome sequencing cheap and accessible. However, closing genomes is often costly and assembling short reads from genomes that are repetitive and/or have extreme %GC content remains challenging. Long-read, single-molecule sequencing technologies such as the Oxford Nanopore MinION have the potential to overcome these difficulties, although the best approach for harnessing their potential remains poorly evaluated. RESULTS: We sequenced nine bacterial genomes spanning a wide range of GC contents using Illumina MiSeq and Oxford Nanopore MinION sequencing technologies to determine the advantages of each approach, both individually and combined. Assemblies using only MiSeq reads were highly accurate but lacked contiguity, a deficiency that was partially overcome by adding MinION reads to these assemblies. Even more contiguous genome assemblies were generated by using MinION reads for initial assembly, but these assemblies were more error-prone and required further polishing. This was especially pronounced when Illumina libraries were biased, as was the case for our strains with both high and low GC content. Increased genome contiguity dramatically improved the annotation of insertion sequences and secondary metabolite biosynthetic gene clusters, likely because long-reads can disambiguate these highly repetitive but biologically important genomic regions. CONCLUSIONS: Genome assembly using short-reads is challenged by repetitive sequences and extreme GC contents. Our results indicate that these difficulties can be largely overcome by using single-molecule, long-read sequencing technologies such as the Oxford Nanopore MinION. Using MinION reads for assembly followed by polishing with Illumina reads generated the most contiguous genomes with sufficient accuracy to enable the accurate annotation of important but difficult to sequence genomic features such as insertion sequences and secondary metabolite biosynthetic gene clusters. The combination of Oxford Nanopore and Illumina sequencing can therefore cost-effectively advance studies of microbial evolution and genome-driven drug discovery.

Age-related changes in the gut microbiota influence systemic inflammation and stroke outcome.

OBJECTIVE: Chronic systemic inflammation contributes to the pathogenesis of many age-related diseases. Although not well understood, alterations in the gut microbiota, or dysbiosis, may be responsible for age-related inflammation. METHODS: Using stroke as a disease model, we tested the hypothesis that a youthful microbiota, when established in aged mice, produces positive outcomes following ischemic stroke. Conversely, an aged microbiota, when established in young mice, produces negative outcomes after stroke. Young and aged male mice had either a young or an aged microbiota established by fecal transplant gavage (FTG). Mice were subjected to ischemic stroke (middle cerebral artery occlusion; MCAO) or sham surgery. During the subsequent weeks, mice underwent behavioral testing and fecal samples were collected for 16S ribosomal RNA analysis of bacterial content. RESULTS: We found that the microbiota is altered after experimental stroke in young mice and resembles the biome of uninjured aged mice. In aged mice, the ratio of Firmicutes to Bacteroidetes (F:B), two main bacterial phyla in gut microbiota, increased ∼9-fold (p < 0.001) compared to young. This increased F:B ratio in aged mice is indicative of dysbiosis. Altering the microbiota in young by fecal gavage to resemble that of aged mice (∼6-fold increase in F:B ratio, p < 0.001) increased mortality following MCAO, decreased performance in behavioral testing, and increased cytokine levels. Conversely, altering the microbiota in aged to resemble that of young (∼9-fold decrease in F:B ratio, p < 0.001) increased survival and improved recovery following MCAO. INTERPRETATION: Aged biome increased the levels of systemic proinflammatory cytokines. We conclude that the gut microbiota can be modified to positively impact outcomes from age-related diseases. Ann Neurol 2018;83:23-36.

Flagellar regulation mediated by the Rcs pathway is required for virulence in the fish pathogen Yersinia ruckeri.

The flagellum is a complex surface structure necessary for a number of activities including motility, chemotaxis, biofilm formation and host attachment. Flagellin, the primary structural protein making up the flagellum, is an abundant and potent activator of innate and adaptive immunity and therefore expression of flagellin during infection could be deleterious to the infection process due to flagellin-mediated host recognition. Here, we use quantitative RT-PCR to demonstrate that expression of the flagellin locus fliC is repressed during the course of infection and subsequently up-regulated upon host mortality in a motile strain of Yersinia ruckeri. The kinetics of fliC repression during the infection process is relatively slow as full repression occurs 7-days after the initiation of infection and after approximately 3-logs of bacterial growth in vivo. These results suggests that Y. ruckeri possesses a regulatory system capable of sensing host and modulating the expression of motility in response. Examination of the master flagellar operon (flhDC) promoter region for evidence of transcriptional regulation and regulatory binding sites revealed potential interaction with the Rcs pathway through an Rcs(A)B Box. Deletion of rcsB (ΔrcsB) by marker-exchange mutagenesis resulted in overproduction of flagellin and unregulated motility, showing that the Rcs pathway negatively regulates biosynthesis of the flagellar apparatus. Experimental challenge with ΔrcsB and ΔrcsBΔfliC1ΔfliC2 mutants revealed that mutation of the Rcs pathway results in virulence attenuation which is dependent on presence of the flagellin gene. These results suggest that the inappropriate expression of flagellin during infection triggers host recognition and thus immune stimulation resulting in attenuation of virulence. In addition, RNAseq analyses of the ΔrcsB mutant strain verified the role of this gene as a negative regulator of the flagellar motility system and identified several additional genes regulated by the Rcs pathway.

Early-life antibiotics attenuate regulatory T cell generation and increase the severity of murine house dust mite-induced asthma.

INTRODUCTION: Early-life exposure to antibiotics (ABX) has been linked to increases in asthma severity and prevalence in both children and laboratory animals. We explored the immunologic mechanisms behind this association using a mouse model of house dust mite (HDM)-induced asthma and early-life ABX exposure. METHODS: Mice were exposed to three short courses of ABX following weaning and experimental asthma was thereafter induced. Airway cell counts and differentials; serum immunoglobulin E (IgE); pulmonary function; lung histopathology; pulmonary regulatory T cells (Tregs); and the fecal microbiome were characterized following ABX exposure and induction of experimental asthma. RESULTS: Asthma severity was increased in mice exposed to ABX, including: airway eosinophilia, airway hyper-reactivity, serum HDM-specific IgE, and lung histopathology. ABX treatment led to sharp reduction in fecal microbiome diversity, including the loss of pro-regulatory organisms such as Lachnospira. Pulmonary Tregs were reduced with ABX treatment, and this reduction was directly proportional to diminished microbiome diversity. CONCLUSION: Intermittent exposure to ABX early in life worsened the severity of experimental asthma and reduced pulmonary Tregs; the latter change correlated with decreased microbiome diversity. These data may suggest targets for immunologic or probiotic therapy to counteract the harmful effects of childhood ABX.

Influence of Feeding Type on Gut Microbiome Development in Hospitalized Preterm Infants.

BACKGROUND: Premature infants have a high risk for dysbiosis of the gut microbiome. Mother's own milk (MOM) has been found to favorably alter gut microbiome composition in infants born at term. Evidence about the influence of feeding type on gut microbial colonization of preterm infants is limited. OBJECTIVE: The purpose of this study was to explore the effect of feeding types on gut microbial colonization of preterm infants in the neonatal intensive care unit. METHODS: Thirty-three stable preterm infants were recruited at birth and followed up for the first 30 days of life. Daily feeding information was used to classify infants into six groups (MOM, human donor milk [HDM], Formula, MOM + HDM, MOM + Formula, and HDM + Formula) during postnatal days 0-10, 11-20, and 21-30. Stool samples were collected daily. DNA extracted from stool was used to sequence the 16S rRNA gene. Exploratory data analysis was conducted with a focus on temporal changes of microbial patterns and diversities among infants from different feeding cohorts. Prediction of gut microbial diversity from feeding type was estimated using linear mixed models. RESULTS: Preterm infants fed MOM (at least 70% of the total diet) had highest abundance of Clostridiales, Lactobacillales, and Bacillales compared to infants in other feeding groups, whereas infants fed primarily HDM or formula had a high abundance of Enterobacteriales compared to infants fed MOM. After controlling for gender, postnatal age, weight, and birth gestational age, the diversity of gut microbiome increased over time and was constantly higher in infants fed MOM relative to infants with other feeding types (p < .01). DISCUSSION: MOM benefits gut microbiome development of preterm infants, including balanced microbial community pattern and increased microbial diversity in early life.

Mucinivorans hirudinis gen. nov., sp. nov., an anaerobic, mucin-degrading bacterium isolated from the digestive tract of the medicinal leech Hirudo verbana.

Three anaerobic bacterial strains were isolated from the digestive tract of the medicinal leech Hirudo verbana, using mucin as the primary carbon and energy source. These strains, designated M3(T), M4 and M6, were Gram-stain-negative, non-spore-forming and non-motile. Cells were elongated bacilli approximately 2.4 µm long and 0.6 µm wide. Growth only occurred anaerobically under mesophilic and neutral pH conditions. All three strains could utilize multiple simple and complex sugars as carbon sources, with glucose fermented to acid by-products. The DNA G+C contents of strains M3(T), M4 and M6 were 44.9, 44.8 and 44.8 mol%, respectively. The major cellular fatty acid of strain M3(T) was iso-C15 : 0. Phylogenetic analysis of full-length 16S rRNA gene sequences revealed that the three strains shared >99 % similarity with each other and represent a new lineage within the family Rikenellaceae of the order Bacteroidales, phylum Bacteroidetes. The most closely related bacteria to strain M3(T) based on 16S rRNA gene sequences were Rikenella microfusus DSM 15922(T) (87.3 % similarity) and Alistipes finegoldii AHN 2437(T) (87.4 %). On the basis of phenotypic, genotypic and physiological evidence, strains M3(T), M4 and M6 are proposed as representing a novel species of a new genus within the family Rikenellaceae, for which the name Mucinivorans hirudinis gen. nov., sp. nov. is proposed. The type strain of Mucinivorans hirudinis is M3(T) ( = ATCC BAA-2553(T) = DSM 27344(T)).

Effect of organic loading on the microbiota in a temperature-phased anaerobic digestion (TPAD) system co-digesting dairy manure and waste whey.

Temperature-phased anaerobic digestion (TPAD) has gained increasing attention because it provides the flexibility to operate digesters under conditions that enhance overall digester performance. However, research on impact of organic overloading rate (OLR) to microbiota of TPAD systems was limited. In this study, we investigated the composition and successions of the microbiota in both the thermophilic and the mesophilic digesters of a laboratory-scale TPAD system co-digesting dairy manure and waste whey before and during organic overloading. The thermophilic and the mesophilic digesters were operated at 50 and 35 °C, respectively, with a hydraulic retention time (HRT) of 10 days for each digester. High OLR (dairy manure with 5 % total solid and waste whey of ≥60.4 g chemical oxygen demand (COD)/l/day) resulted in decrease in pH and in biogas production and accumulation of volatile fatty acids (VFAs) in the thermophilic digester, while the mesophilic digester remained unchanged except a transient increase in biogas production. Both denaturant gradient gel electrophoresis (DGGE) and Illumina sequencing of 16S ribosomal RNA (rRNA) gene amplicons showed dramatic change in microbiota composition and profound successions of both bacterial and methanogenic communities. During the overloading, Thermotogae was replaced by Proteobacteria, while Methanobrevibacter and archaeon classified as WCHD3-02 grew in predominance at the expense of Methanoculleus in the thermophilic digester, whereas Methanosarcina dominated the methanogenic community, while Methanobacterium and Methanobrevibacter became less predominant in the mesophilic digester. Canonical correspondence analysis (CCA) revealed that digester temperature and pH were the most influential environmental factors that explained much of the variations of the microbiota in this TPAD system when it was overloaded.

Comparison of the microbial communities in solid-state anaerobic digestion (SS-AD) reactors operated at mesophilic and thermophilic temperatures.

The microbiomes involved in liquid anaerobic digestion process have been investigated extensively, but the microbiomes underpinning solid-state anaerobic digestion (SS-AD) are poorly understood. In this study, microbiome composition and temporal succession in batch SS-AD reactors, operated at mesophilic or thermophilic temperatures, were investigated using Illumina sequencing of 16S rRNA gene amplicons. A greater microbial richness and evenness were found in the mesophilic than in the thermophilic SS-AD reactors. Firmicutes accounted for 60 and 82 % of the total Bacteria in the mesophilic and in the thermophilic SS-AD reactors, respectively. The genus Methanothermobacter dominated the Archaea in the thermophilic SS-AD reactors, while Methanoculleus predominated in the mesophilic SS-AD reactors. Interestingly, the data suggest syntrophic acetate oxidation coupled with hydrogenotrophic methanogenesis as an important pathway for biogas production during the thermophilic SS-AD. Canonical correspondence analysis (CCA) showed that temperature was the most influential factor in shaping the microbiomes in the SS-AD reactors. Thermotogae showed strong positive correlation with operation temperature, while Fibrobacteres, Lentisphaerae, Spirochaetes, and Tenericutes were positively correlated with daily biogas yield. This study provided new insight into the microbiome that drives SS-AD process, and the findings may help advance understanding of the microbiome in SS-AD reactors and the design and operation of SS-AD systems.

Early Life Experience and Gut Microbiome: The Brain-Gut-Microbiota Signaling System.

BACKGROUND: Over the past decades, advances in neonatal care have led to substantial increases in survival among preterm infants. With these gains, recent concerns have focused on increases in neurodevelopment morbidity related to the interplay between stressful early life experiences and the immature neuroimmune systems. This interplay between these complex mechanisms is often described as the brain-gut signaling system. The role of the gut microbiome and the brain-gut signaling system have been found to be remarkably related to both short- and long-term stress and health. Recent evidence supports that microbial species, ligands, and/or products within the developing intestine play a key role in early programming of the central nervous system and regulation of the intestinal innate immunity. PURPOSE: The purpose of this state-of-the-science review is to explore the supporting evidence demonstrating the importance of the brain-gut-microbiota axis in regulation of early life experience. We also discuss the role of gut microbiome in modulating stress and pain responses in high-risk infants. A conceptual framework has been developed to illustrate the regulation mechanisms involved in early life experience. CONCLUSIONS: The science in this area is just beginning to be uncovered; having a fundamental understanding of these relationships will be important as new discoveries continue to change our thinking, leading potentially to changes in practice and targeted interventions.

Seasonal and spatial variations in microbial activity at various phylogenetic resolutions at a groundwater - surface water interface.

We investigated the seasonal and spatial variation in activity and density of the metabolically active in situ microbial community (AIMC) at a landfill leachate-impacted groundwater - surface water interface (GSI). A series of AIMC traps were designed and implemented for AIMC sampling and microbial activity and density examinations. Measurements were made not only at the level of bacterial domain but also at the levels of alphaproteobacterial Rhizobiales order and gammaproteobacterial Pseudomonas genus, both of which included a large number of iron-oxidizing bacteria as revealed from previous analysis. Consistently higher microbial activities with less variation in depth were measured in the AIMC traps than in the ambient sediments. Flood disturbance appeared to control AIMC activity distributions at the gradually elevated GSI. The highest AIMC activities were generally obtained from locations closest to the free surface water boundary except during the dry season when microbial activities were similar across the entire GSI. A clone library of AIMC 16S rRNA genes was constructed, and it confirmed the predominant role of the targeted alphaproteobacterial group in AIMC activity and composition. This taxon constituted 2%-14% of all bacteria with similar activity distribution profiles. The Pseudomonas group occupied only 0.1‰-0.5‰ of the total bacterial density, but its activity was 27 times higher than the bacterial average. Of the 16S rRNA sequences in the AIMC clone library, 7.5% were phylogenetically related to putative IOB, supporting the occurrence and persistence of active microbial iron oxidation across the studied iron-rich GSI ecosystem.

Slowing Alzheimer's disease progression through probiotic supplementation.

The lack of affordable and effective therapeutics against cognitive impairment has promoted research toward alternative approaches to the treatment of neurodegeneration. In recent years, a bidirectional pathway that allows the gut to communicate with the central nervous system has been recognized as the gut-brain axis. Alterations in the gut microbiota, a dynamic population of trillions of microorganisms residing in the gastrointestinal tract, have been implicated in a variety of pathological states, including neurodegenerative disorders such as Alzheimer's disease (AD). However, probiotic treatment as an affordable and accessible adjuvant therapy for the correction of dysbiosis in AD has not been thoroughly explored. Here, we sought to correct the dysbiosis in an AD mouse model with probiotic supplementation, with the intent of exploring its effects on disease progression. Transgenic 3xTg-AD mice were fed a control or a probiotic diet (Lactobacillus plantarum KY1032 and Lactobacillus curvatus HY7601) for 12 weeks, with the latter leading to a significant increase in the relative abundance of Bacteroidetes. Cognitive functions were evaluated via Barnes Maze trials and improvements in memory performance were detected in probiotic-fed AD mice. Neural tissue analysis of the entorhinal cortex and hippocampus of 10-month-old 3xTg-AD mice demonstrated that astrocytic and microglial densities were reduced in AD mice supplemented with a probiotic diet, with changes more pronounced in probiotic-fed female mice. In addition, elevated numbers of neurons in the hippocampus of probiotic-fed 3xTg-AD mice suggested neuroprotection induced by probiotic supplementation. Our results suggest that probiotic supplementation could be effective in delaying or mitigating early stages of neurodegeneration in the 3xTg-AD animal model. It is vital to explore new possibilities for palliative care for neurodegeneration, and probiotic supplementation could provide an inexpensive and easily implemented adjuvant clinical treatment for AD.

Draft Genome Sequence of Plesiomonas shigelloides MD22D9, Isolated from the Digestive Tract of Macrobdella decora.

Here, we present a draft genome sequence of Plesiomonas shigelloides MD22D9, isolated from the digestive tract of the North American medicinal leech Macrobdella decora. The gut microbiome of the medicinal leech is hypothesized to be critical for maintaining host fitness. This genome can provide insights into this uncharacterized microbe-host relationship.

Development of a biomarker signature using grating-coupled fluorescence plasmonic microarray for diagnosis of MIS-C.

Multisystem inflammatory syndrome in children (MIS-C) is a rare but serious condition that can develop 4-6 weeks after a school age child becomes infected by SARS-CoV-2. To date, in the United States more than 8,862 cases of MIS-C have been identified and 72 deaths have occurred. This syndrome typically affects children between the ages of 5-13; 57% are Hispanic/Latino/Black/non-Hispanic, 61% of patients are males and 100% have either tested positive for SARS-CoV-2 or had direct contact with someone with COVID-19. Unfortunately, diagnosis of MIS-C is difficult, and delayed diagnosis can lead to cardiogenic shock, intensive care admission, and prolonged hospitalization. There is no validated biomarker for the rapid diagnosis of MIS-C. In this study, we used Grating-coupled Fluorescence Plasmonic (GCFP) microarray technology to develop biomarker signatures in pediatric salvia and serum samples from patients with MIS-C in the United States and Colombia. GCFP measures antibody-antigen interactions at individual regions of interest (ROIs) on a gold-coated diffraction grating sensor chip in a sandwich immunoassay to generate a fluorescent signal based on analyte presence within a sample. Using a microarray printer, we designed a first-generation biosensor chip with the capability of capturing 33 different analytes from 80  µ L of sample (saliva or serum). Here, we show potential biomarker signatures in both saliva and serum samples in six patient cohorts. In saliva samples, we noted occasional analyte outliers on the chip within individual samples and were able to compare those samples to 16S RNA microbiome data. These comparisons indicate differences in relative abundance of oral pathogens within those patients. Microsphere Immunoassay (MIA) of immunoglobulin isotypes was also performed on serum samples and revealed MIS-C patients had several COVID antigen-specific immunoglobulins that were significantly higher than other cohorts, thus identifying potential new targets for the second-generation biosensor chip. MIA also identified additional biomarkers for our second-generation chip, verified biomarker signatures generated on the first-generation chip, and aided in second-generation chip optimization. Interestingly, MIS-C samples from the United States had a more diverse and robust signature than the Colombian samples, which was also illustrated in the MIA cytokine data. These observations identify new MIS-C biomarkers and biomarker signatures for each of the cohorts. Ultimately, these tools may represent a potential diagnostic tool for use in the rapid identification of MIS-C.