16S Ribosomal Gene Research Articles

ABC-12: Exploring the microbiome in patients (pts) with advanced biliary tract cancer (BTC) in a first-line study of durvalumab in combination with cisplatin/gemcitabine (cis/gem).

TPS4183 Background: Durvalumab/cis/gem improved overall survival (OS) in pts with advanced BTC versus placebo/cis/gem (Oh et al. NEJM Evid 2022). Disruption of the microbiota may impair tumour response to immunotherapy and chemotherapy and a better understanding of its role in the efficacy of these therapeutics in advanced BTC is required. Methods: This is a multi-centre, single arm trial exploring the microbiome in pts receiving durvalumab 1500 mg intravenously (IV) Q3w, in combination with cis 25 mg/m2, gem 1000 mg/m2 (Days 1 and 8, Q3w) up to 8 cycles, followed by durvalumab 1500 mg as monotherapy Q4w, until progression or intolerable toxicity. Pts with an ECOG performance status of ≤1 and histologically-proven BTC, including cholangiocarcinoma and gallbladder carcinoma, who have had no prior systemic chemotherapy for locally advanced or metastatic disease are eligible. Pts must provide a saliva and stool sample prior to commencement of durvalumab/cis/gem and at 18 weeks, or at progression (if earlier than 18 weeks). Taxonomic profiling via 16S Ribosomal ribonucleic acid gene sequencing will examine the differences in the diversity and composition of the pt gut microbiome. Pts must also have availability of a tumour biopsy. This study plans to recruit 70 pts from 10 UK centres (over 12 months). The primary objective is to determine the difference in baseline alpha diversity between “responders” (partial or complete response) and “non-responders” at 18 weeks (RECIST 1.1) in patients treated with durvalumab/cis/gem. Secondary objectives include investigation of the association between microbiome parameters and objective response rate, tumour control (partial + complete response + stable disease), progression-free and OS, and to investigate the interaction between treatment effect and microbiome parameters on clinical outcomes. The tumour biopsy will be used for research into the tumour microbiome and/or factors that may influence response to chemotherapy/immunotherapy, including, but not limited to tumour mutation burden, programmed cell death 1/programmed death-ligand 1 status, and microsatellite instability status. Clinical trial information: ISRCTN11210442 .

Comprehensive study of the intratumoral microbiome in early- vs. late-onset colorectal cancer: Final analysis of COSMO CRC.

3530 Background: Although colorectal cancer (CRC) incidence has declined overall, CRC in individuals under age 45 (early-onset CRC, EOCRC) has risen dramatically. Obesity and diabetes may partially explain this epidemiologic shift; however, many patients (pts) with EOCRC are neither obese nor diabetic. Certain bacteria disrupt colonic luminal integrity and promote inflammation, leading to oncogenic mutations in colonic epithelial cells. Fusobacterium nucleatum ( F. nuc) promotes CRC by suppressing immune response within the tumor microenvironment, activating the β-catenin pathway, and causing chemoresistance due to autophagy. The intratumoral microbiome (MB) in pts with EOCRC may differ from pts with late-onset CRC (LOCRC). Methods: We compared the intratumoral MB in pts with CRC diagnosed before age 45 (EOCRC) and after age 65 (LOCRC) in a prospective/retrospective study between 2017 and 2022. Primary and metastatic tumors were included. DNA was extracted from tumors and analyzed using 16S ribosomal gene sequencing. We compared the frequency of F. nuc and other bacterial and fungal DNA in tumors EOCRC vs. LOCRC pts. Next-generation tumor sequencing and diet questionnaire data were available for some pts. Results: Tumors from 36 EOCRC pts (median age 38 years) and 27 LOCRC pts (median age 72 years) were analyzed. In total, 917 unique bacterial and fungal species were detected. F. nuc was found in 30.6% of EOCRC and 29.6% of LOCRC (p = 0.94). Cladosporium sp. was seen more commonly in EOCRC (30.6% vs. 11%, p = 0.04), whereas Pseudomonas luteola (2.8% vs. 22.2%), Ralstonia sp. (22.2% vs. 48.1%), and Moraxella osloensis (19.4% vs. 44.4%) were seen more commonly in LOCRC (p < 0.05). Clostridium perfringens (11.1%), Escherichia coli (11.1%), Leptotrichia hofstadii (11.1%) , Mycosphaerella sp. (11.1%), Neodevriesia modesta (11.1%), Penicillium sp. (11.1%), and Leptosphaeria sp. (11.1%) were seen exclusively in LOCRC (p < 0.05). There was no significant difference in median MB diversity in EOCRC vs. LOCRC (43 vs. 45 organisms per pt). Median follow up from time of diagnosis was 39.6 months (mo). Twenty-three EOCRC pts (64%) and 19 LOCRC pts (70%) are alive. Median overall survival (mOS) was 75.5 mo (95 CI: 40 - NA) in EOCRC and 60 mo (95% CI: 50.5 - NA) in LOCRC. There was no significant difference in OS in LOCRC vs. EOCRC (Log-rank test p = 0.85). Pts with F. nuc positive tumors (N = 17) had a mOS of 75.5 mo (95 CI: 23 – NA), whereas pts with F. nuc negative tumors (N = 46) had a mOS of 60 mo (95% CI: 50.5 – NA). There was no significant difference in OS in F. nuc positive vs. F. nuc negative groups (Log-rank test p = 0.87). Conclusions: There are significant differences in the intratumoral microbiome in EOCRC and LOCRC. These findings warrant larger, prospective studies to elucidate the role the intratumoral microbiome plays in the carcinogenesis, the tumor immune microenvironment, and responsiveness to specific therapies.

Gut microbiota indole-3-propionic acid mediates neuroprotective effect of probiotic consumption in healthy elderly: A randomized, double-blind, placebo-controlled, multicenter trial and in vitro study

The beneficial effects of probiotic consumption on age-related decline in cerebral function have been previously reported in the literature; however, the mechanistic link between gut and brain interactions has not yet been fully elucidated. Therefore, this study aimed to identify the role of gut microbiota-derived metabolites in gut-brain interactions via blood metabolomic profiling analysis in clinical trials and invitro mechanistic studies. A randomized, double-blind, placebo-controlled, multicenter clinical trial was conducted in 63 healthy elderly individuals (≥65 years of age). Participants were administered either placebo (placebo group, N=31) or probiotic capsules (Bifidobacterium bifidum BGN4 and Bifidobacterium longum BORI; probiotics group, N=32) for 12 weeks. Global and targeted metabolomic profiling analyses of their blood samples were then performed using 1H nuclear magnetic resonance and liquid chromatography-mass spectrometry methods, both at baseline and at the end of the trial. Gut microbial analysis was conducted using the 16S ribosomal ribonucleic acid gene sequencing method. Subsequently, microglial BV2 cells were treated invitro with indole-3-propionic acid (IPA) following lipopolysaccharide stimulation, and neuronal SH-SY5Y cells were treated with conditioned media from the BV2 cells. Finally, the levels of pro-inflammatory cytokines in BV2 cells and neurotrophins in SH-SY5Y cells were quantified using a real-time polymerase chain reaction or enzyme-linked immunosorbent assay. The metabolomic profiling analyses showed that probiotic consumption significantly altered the levels of metabolites involved in tryptophan metabolism (P<0.01). Among these metabolites, gut microbiota-produced IPA had a 1.91-fold increase in the probiotics group (P<0.05) and showed a significant relation to gut bacterial profiles (P<0.01). Elevated IPA levels were also positively associated with the level of serum brain-derived neurotropic factor (BDNF) in the probiotics group (r=0.28, P<0.05), showing an inverse trend compared to the placebo group. In addition, invitro treatment with IPA (5μM) significantly reduced the concentration of proinflammatory TNF-α in activated microglia (P<0.05), and neuronal cells cultured with conditioned media from IPA-treated microglia showed a significant increase in BDNF and nerve growth factor production (P<0.05). These results show that gut microbiota-produced IPA plays a role in protecting the microglia from inflammation, thus promoting neuronal function. Therefore, this suggests that IPA is a significant mediator linking the interaction between the gut and the brain in the elderly with probiotic supplementation.

STENOPHYSA MARMORATA (MOLLUSCA: PHYSIDADE) AS AN INTERMEDIATE HOST OF A SNAKE TREMATODE OF THE FAMILY RENIFERIDAE IN BRAZIL.

Species of the family Reniferidae are trematodes found in the oral cavity and esophagus of snakes from Nearctic and Neotropical regions. Although Renifer heterocoelium has been reported in different snake species from South America, the snails involved in its transmission remain unknown. In this study, a xiphidiocercaria emerged from the physid snail Stenophysa marmorata from Brazil and was subjected to morphological and molecular study. The general morphology, including the shape of the stylet and arrangement of penetration glands, resembles that described for reniferid trematodes from North America. Phylogenetic analysis on the basis of nuclear sequences (28S ribosomal deoxyribonucleic acid gene [1,072 base pairs {bp}] and internal transcribed spacer region [ITS, 1,036 bp]) supports identifying this larva as a member of the family Reniferidae, very possibly a species of the genus Renifer. In the 28S analysis, low molecular divergences were found to Renifer aniarum (1.4%) and Renifer kansensis (0.6%), but also concerning other 2 reniferid species, i.e., Dasymetra nicolli (1.4%) and Lechriorchis tygarti (1.0%). Regarding ITS, the divergences between this Brazilian cercaria and R. aniarum or L. tygarti were 1.9% and 8.5%, respectively. In the case of the mitochondrial marker cytochrome oxidase subunit 1 (797 bp), our Reniferidae gen. sp. differs 8.6-9.6% from Paralechriorchis syntomentera, the only reniferid with sequences available for comparison. We discuss the probable conspecificity of the larval stages here reported with R. heterocoelium, the reniferid species reported in South America.

Single Copies of the 5S rRNA Inserted into 45S rDNA Intergenic Spacers in the Genomes of Nototheniidae (Perciformes, Actinopterygii).

In the vast majority of Animalia genomes, the 5S rRNA gene repeats are located on chromosomes outside of the 45S rDNA arrays of the nucleolar organiser (NOR). We analysed the genomic databases available and found that a 5S rDNA sequence is inserted into the intergenic spacer (IGS) between the 45S rDNA repeats in ten species of the family Nototheniidae (Perciformes, Actinopterigii). We call this sequence the NOR-5S rRNA gene. Along with Testudines and Crocodilia, this is the second case of a close association between four rRNA genes within one repetitive unit in deuterostomes. In both cases, NOR-5S is oriented opposite the 45S rDNA. None of the three nucleotide substitutions compared to the canonical 5S rRNA gene influenced the 5S rRNA secondary structure. In transcriptomes of the Patagonian toothfish, we only found NOR-5S rRNA reads in ovaries and early embryos, but not in testis or somatic tissues of adults. Thus, we consider the NOR-5S gene to be a maternal-type 5S rRNA template. The colocalization of the 5S and 45S ribosomal genes appears to be essential for the equimolar production of all four rRNAs in the species that show rDNA amplification during oogenesis. Most likely, the integration of 5S and NOR rRNA genes occurred prior to Nototheniidae lineage diversification.

Integrative taxonomy in the genus Rhabdias stiles et Hassall, 1905 from anuran in Brazil, description of two new species and phylogenetic analyses.

About 20 valid species of the genus Rhabdias are known in the Neotropical region. The present study aimed to describe two new species of Rhabdias parasitizing the lungs of Leptodactylus macrosternum and Leptodactylus podicipinus from Brazil. Distinctive characteristics between these species are numerous and based on body size, size of the buccal capsule, shape and size of the oesophagus, and position of the vulva. Molecular data based on ribosomal genes 28S and ITS region and mitochondrial COI of the two species are presented. Molecular analysis and comparison of the partial mitochondrial COI sequence of Rhabdias matogrossensis n. sp. and Rhabdias guaianensis n. sp. revealed a genetic divergence between these new species and the sequences of Rhabdias spp. previously deposited in GenBank. In the phylogenetic analysis, R. matogrossensis n. sp. was grouped with R. breviensis species complex, and R. guaianensis n. sp. was grouped as a sister group of R. cf. stenochepala. This study contributes to improving the diversity of known species of Rhabdias described in Brazilian anurans.

Improvement and Validation of a Genomic DNA Extraction Method for Human Breastmilk.

The human milk microbiota (HMM) of healthy women can vary substantially, as demonstrated by recent advances in DNA sequencing technology. However, the method used to extract genomic DNA (gDNA) from these samples may impact the observed variations and potentially bias the microbiological reconstruction. Therefore, it is important to use a DNA extraction method that is able to effectively isolate gDNA from a diverse range of microorganisms. In this study, we improved and compared a DNA extraction method for gDNA isolation from human milk (HM) samples to commercial and standard protocols. We evaluated the extracted gDNA using spectrophotometric measurements, gel electrophoresis, and PCR amplifications to assess its quantity, quality, and amplifiability. Additionally, we tested the improved method's ability to isolate amplifiable gDNA from fungi, Gram-positive and Gram-negative bacteria to validate its potential for reconstructing microbiological profiles. The improved DNA extraction method resulted in a higher quality and quantity of the extracted gDNA compared to the commercial and standard protocols and allowed for polymerase chain reaction (PCR) amplification of the V3-V4 regions of the 16S ribosomal gene in all the samples and the ITS-1 region of the fungal 18S ribosomal gene in 95% of the samples. These results suggest that the improved DNA extraction method demonstrates better performance for gDNA extraction from complex samples such as HM.

Impact of High Salt-Intake on a Natural Gut Ecosystem in Wildling Mice.

The mammalian holobiont harbors a complex and interdependent mutualistic gut bacterial community. Shifts in the composition of this bacterial consortium are known to be a key element in host health, immunity and disease. Among many others, dietary habits are impactful drivers for a potential disruption of the bacteria-host mutualistic interaction. In this context, we previously demonstrated that a high-salt diet (HSD) leads to a dysbiotic condition of murine gut microbiota, characterized by a decrease or depletion of well-known health-promoting gut bacteria. However, due to a controlled and sanitized environment, conventional laboratory mice (CLM) possess a less diverse gut microbiota compared to wild mice, leading to poor translational outcome for gut microbiome studies, since a reduced gut microbiota diversity could fail to depict the complex interdependent networks of the microbiome. Here, we evaluated the HSD effect on gut microbiota in CLM in comparison to wildling mice, which harbor a natural gut ecosystem more closely mimicking the situation in humans. Mice were treated with either control food or HSD and gut microbiota were profiled using amplicon-based methods targeting the 16S ribosomal gene. In line with previous findings, our results revealed that HSD induced significant loss of alpha diversity and extensive modulation of gut microbiota composition in CLM, characterized by the decrease in potentially beneficial bacteria from Firmicutes phylum such as the genera Lactobacillus, Roseburia, Tuzzerella, Anaerovorax and increase in Akkermansia and Parasutterella. However, HSD-treated wildling mice did not show the same changes in terms of alpha diversity and loss of Firmicutes bacteria as CLM, and more generally, wildlings exhibited only minor shifts in the gut microbiota composition upon HSD. In line with this, 16S-based functional analysis suggested only major shifts of gut microbiota ecological functions in CLM compared to wildling mice upon HSD. Our findings indicate that richer and wild-derived gut microbiota is more resistant to dietary interventions such as HSD, compared to gut microbiota of CLM, which may have important implications for future translational microbiome research.

Does Methane Contribute to Growth of Invertebrate Communities in Drinking Water?

Methane is a potential source of carbon in drinking water. Typically, it is removed at waterworks during an initial treatment step such as aeration or stripping. Remaining methane may be converted by methane-oxidizing bacteria to organic carbon, which is then available for heterotrophic growth and may ultimately contribute to invertebrate growth. We investigated the presence of invertebrates at a waterworks with incomplete methane removal and at a waterworks without methane. Microscopy and analyses of 16S and 18S ribosomal genes were conducted on filter sand from full-scale biological rapid sand filters. Primary filters with methane were dominated by methane- and ammonia-oxidizing bacteria. Upper layers of secondary filters were dominated by heterotrophic bacteria, while the deepest layer contained 92% eukaryote DNA. Rotifers, nematodes, platyhelminths and annelids constituted 22% of the DNA in the secondary filters. Filters with methane contained higher shares of invertebrates (13%) than the filter without methane (7%). Furthermore, pilot studies were conducted to estimate suitable levels of methane when implementing methane removal technologies. Methane concentrations of 0.24 mg/L caused rapid visible growth. Vacuum stripping and nitrogen addition removed methane to 0.018–0.03 mg/L and prevented growth of methane-oxidizing bacteria.

The effects of antibiotics and illness on gut microbial composition in the fawn-footed mosaic-tailed rat (Melomys cervinipes).

The gut microbiota are critical for maintaining the health and physiological function of individuals. However, illness and treatment with antibiotics can disrupt bacterial community composition, the consequences of which are largely unknown in wild animals. In this study, we described and quantified the changes in bacterial community composition in response to illness and treatment with antibiotics in a native Australian rodent, the fawn-footed mosaic-tailed rat (Melomys cervinipes). We collected faecal samples during an undiagnosed illness outbreak in a captive colony of animals, and again at least one year later, and quantified the microbiome at each time point using 16s ribosomal rRNA gene sequencing. Gut bacterial composition was quantified at different taxonomic levels, up to family. Gut bacterial composition changed between time periods, indicating that illness, treatment with antibiotics, or a combination affects bacterial communities. While some bacterial groups increased in abundance, others decreased, suggesting differential effects and possible co-adapted and synergistic interactions. Our findings provide a greater understanding of the dynamic nature of the gut microbiome of a native Australian rodent species and provides insights into the management and ethical well-being of animals kept under captive conditions.

Insights on the particle-attached riverine archaeal community shifts linked to seasons and to multipollution during a Mediterranean extreme storm event.

Even if Archaea deliver important ecosystem services and are major players in global biogeochemical cycles, they remain poorly understood in freshwater ecosystems. To our knowledge, no studies specifically address the direct impact of xenobiotics on the riverine archaeome. Using environmental DNA metabarcoding of the 16S ribosomal gene, we previously demonstrated bacterial communities significant shifts linked to pollutant mixtures during an extreme flood in a typical Mediterranean coastal watercourse. Here, using the same methodology, we sought to determine whether archaeal community shifts coincided with the delivery of environmental stressors during the same flood. Further, we wanted to determine how archaea taxa compared at different seasons. In contrast to the bacteriome, the archaeome showed a specific community in summer compared to winter and autumn. We also identified a significant relationship between in situ archaeome shifts and changes in physicochemical parameters along the flood, but a less marked link to those parameters correlated to river hydrodynamics than bacteria. New urban-specific archaeal taxa significantly related to multiple stressors were identified. Through statistical modeling of both domains, our results demonstrate that Archaea, seldom considered as bioindicators of water quality, have the potential to improve monitoring methods of watersheds.

Genome skimming elucidates the evolutionary history of Octopoda

Phylogenies for Octopoda have, until now, been based on morphological characters or a few genes. Here we provide the complete mitogenomes and the nuclear 18S and 28S ribosomal genes of twenty Octopoda specimens, comprising 18 species of Cirrata and Incirrata, representing 13 genera and all five putative families of Cirrata (Cirroctopodidae, Cirroteuthidae, Grimpoteuthidae, Opisthoteuthidae and Stauroteuthidae) and six families of Incirrata (Amphitretidae, Argonautidae, Bathypolypodidae, Eledonidae, Enteroctopodidae, and Megaleledonidae) which were assembled using genome skimming. Phylogenetic trees were built using Maximum Likelihood and Bayesian Inference with several alignment matrices. All mitochondrial genomes had the ‘typical’ genome composition and gene order previously reported for octopodiforms, except Bathypolypus ergasticus, which appears to lack ND5, two tRNA genes that flank ND5 and two other tRNA genes. Argonautoidea was revealed as sister to Octopodidae by the mitochondrial protein-coding gene dataset, however, it was recovered as sister to all other incirrate octopods with strong support in an analysis using nuclear rRNA genes. Within Cirrata, our study supports two existing classifications suggesting neither is likely in conflict with the true evolutionary history of the suborder. Genome skimming is useful in the analysis of phylogenetic relationships within Octopoda; inclusion of both mitochondrial and nuclear data may be key.

Comparative analyses of intratumoral microbiome with diet in patients with early- vs late-onset colorectal cancer.

208 Background: Early-onset colorectal cancer (CRC) is a growing public health problem with an incidence rate increase of 7.9% in individuals aged 20-29, 4.9% in those aged 30-39, and 1.6% in those aged 40-49 during 2004-2016. The etiology of this rise is not well established, but there is emerging evidence that the intra-tumoral microbiome may be implicated. We sought to categorize the interaction between diet and intra-tumoral microbiome in early- vs. late-onset CRC. Methods: Intra-tumoral microbiome data were collected from patients enrolled in a retrospective/prospective study of early ( &lt; 45 years) vs. late-onset (≥ 65 years) CRC. Formalin-fixed paraffin-embedded tumor samples were microdissected for tumor vs. non-tumor, then DNA was extracted and analyzed using 16S ribosomal gene sequencing. The DHQ3 diet questionnaire was used to ascertain diet history for a period of 12 months prior to diagnosis. Twelve bacteria of interest were selected for analysis based on increased prevalence and known potential association with CRC. Diet groups of interest for analysis included high-risk groups: fried foods, red meat, and alcohol. Results: A subset of 3 bacteria were further analyzed, Bacillus sp., Fusobacterium nucleatum, and Moraxella osloensis, based on relation to the group of patients diagnosed with CRC before the age of 45 (N = 8) and after age 65 (N = 12). Bacteria of interest had a similar prevalence in early- (38%, 3/8) and late-onset patients (33%, 4/12). Fifteen of the 20 patients had one or more of the bacteria of interest present. 71% (5/7) of both early- and late-onset patients who had Bacillus present had no alcohol in their diets. 100% (4/4) of the late onset patients with Bacillus present all had the same diet pattern: no tree nuts, high amounts of potato chips, no alcohol, and no red meat. 83% (5/6) of the early onset patients with 1 or more bacteria present had a high amount of potato chips in their diets. 80% (4/5) patients who had none of the bacteria of interest present also had no red meat in their diet. All early- and late-onset patients with Moraxella had a diet that consisted of more high-risk food groups, in particular potato chips. Conclusions: There were no significant differences in intra-tumoral microbiome in patients with early- vs. late-onset CRC. However, there may be an association between certain bacteria and high-risk diets. Results were limited by a small sample size, and further research should be done to better understand the relationship between intra-tumoral microbiome and diet.

Phylogenetic analyses reveal bat communities in Northwestern Mexico harbor a high diversity of novel cryptic ectoparasite species.

Parasites are integral parts of ecosystem function and important drivers of evolutionary processes. Characterizing ectoparasite diversity is fundamental to studies of host-parasite interactions, evolution, and conservation, and also for understanding emerging disease threats for some vector borne pathogens. With more than 1400 species, bats represent the second most speciose mammalian clade, but their ectoparasite fauna are poorly known for most species. We sequenced mitochondrial Cytochrome Oxidase C subunit I and nuclear 18S ribosomal gene fragments, and used Bayesian phylogenetic analyses to characterize ectoparasite taxon identity and diversity for 17 species of parasitized bats sampled along the Baja California peninsula and in Northwestern Mexico. The sequence data revealed multiple novel lineages of bat bugs (Cimicidae), flies (Nycteribiidae and Streblidae), and ticks (Argasidae). Within families, the new linages showed more than 10% sequence divergence, which is consistent with separation at least at the species level. Both families of bat flies showed host specificity, particularly on Myotis species. We also identified new records for the Baja peninsula of one tick (Carios kelleyi), and of five Streblid bat fly species. One Nycteribiid bat fly haplotype from Pallid bat (Antrozous pallidus) hosts was found throughout the peninsula, suggesting potential long distance co-dispersal with hosts. Different bat bug and tick communities were found in the north and south of the peninsula. This study is the first systematic survey of bat ectoparasites in the Baja California peninsula, revealing novel lineages that are highly genetically differentiated from other parts of North America. For some ectoparasite species, haplotype distributions may reflect patterns of bat migration. This work is a first step in characterizing ectoparasite diversity over the Baja California peninsula, and understanding how ecological and evolutionary interactions shape bat ectoparasite communities among host species in different parts of their ranges.

Dietary supplement of mushrooms promotes SCFA production and moderately associates with IgA production: A pilot clinical study.

Mushrooms are rich in dietary fiber, and fiber intake has been reported to increase the levels of short-chain fatty acids (SCFAs). It has also been reported that SCFAs promote immunoglobulin A (IgA) production, indicating involvement in systemic immunity. The objective of this study was to evaluate the effects of mushroom consumption on the amount of intestinal IgA. We also aimed to comprehensively evaluate the gut microbiota and intestinal metabolome and to conduct an exploratory analysis of their relationship with IgA. Healthy adults (n = 80) were enrolled in a parallel group trial. Participants consumed a diet with mushrooms or a placebo diet once daily for 4 weeks. Gut microbiota profiles were assessed by sequencing the bacterial 16S ribosomal RNA-encoding gene. Intestinal metabolome profiles were analyzed using capillary electrophoresis-time of flight mass spectrometry (CE-TOFMS). Mushroom consumption tended to increase IgA levels at 4 weeks of consumption compared to those in the control group (p = 0.0807; Hedges' g = 0.480). The mushroom group had significantly higher levels of intestinal SCFAs, such as butyrate and propionate, than the control group (p = 0.001 and 0.020; Hedges' g = 0.824 and 0.474, respectively). Correlation analysis between the changes in the amount of intestinal IgA and the baseline features of the intestinal environment showed that the increasing amount of intestinal IgA was positively correlated with the baseline levels of SCFAs (Spearman's R = 0.559 and 0.419 for butyrate and propionate, respectively). Consumption of mushrooms significantly increased the intestinal SCFAs and IgA in some subjects. The increase in intestinal IgA levels was more prominent in subjects with higher SCFA levels at baseline. This finding provides evidence that mushroom alters the intestinal environment, but the intensity of the effect still depends on the baseline intestinal environment. This trial was registered at www.umin.ac.jp as UMIN000043979.

Galacto-Oligosaccharides Increase the Abundance of Beneficial Probiotic Bacteria and Improve Gut Architecture and Goblet Cell Expression in Poorly Performing Piglets, but Not Performance.

Poorly performing piglets receiving commercial milk replacers do not benefit from the naturally occurring probiotic galacto-oligosaccharides otherwise found in sow milk. Study objectives were to investigate the effects of complete milk replacer supplemented with galacto-oligosaccharides on the microbiome, gut architecture and immunomodulatory goblet cell expression of poorly performing piglets that could benefit from milk replacement feeding when separated from sows and housed with fit siblings in environmentally controlled pens. The study is novel in that it is one of the first to investigate the effects of supplementing complete milk replacer with galacto-oligosaccharides in poorly performing piglets. Gastrointestinal tract samples were collected from piglets, and the microbiome composition was assessed by 16s ribosomal ribonucleic acid gene sequencing. Gut architectural features, villus/crypt ratio and enumeration of goblet cells in tissues were assessed by histopathological techniques. The most abundant taxa identified at the genus level were Lactobacillus, Streptococcus, Prevotella, Lactococcus and Leuconostoc. Milk replacer plus galacto-oligosaccharides significantly improved gut architectural features and villus/crypt ratio throughout the gastrointestinal tract, increased the number of goblet cells and revealed a differential abundance of beneficial probiotic bacteria, particularly Lactobacillus and Bifidobacterium. In these respects, galacto-oligosaccharide-supplemented milk replacer may be a useful addition to animal husbandry in poorly performing, non-thriving animals when moved to environmentally controlled pens away from sows and fit siblings, thereby modulating the microbiome and gastrointestinal tract performance.

Microbial communities on eelgrass (Zostera marina) thriving in Tokyo Bay and the possible source of leaf-attached microbes.

Zostera marina (eelgrass) is classified as one of the marine angiosperms and is widely distributed throughout much of the Northern Hemisphere. The present study investigated the microbial community structure and diversity of Z. marina growing in Futtsu bathing water, Chiba prefecture, Japan. The purpose of this study was to provide new insight into the colonization of eelgrass leaves by microbial communities based on leaf age and to compare these communities to the root-rhizome of Z. marina, and the surrounding microenvironments (suspended particles, seawater, and sediment). The microbial composition of each sample was analyzed using 16S ribosomal gene amplicon sequencing. Each sample type was found to have a unique microbial community structure. Leaf-attached microbes changed in their composition depending on the relative age of the eelgrass leaf. Special attention was given to a potential microbial source of leaf-attached microbes. Microbial communities of marine particles looked more like those of eelgrass leaves than those of water samples. This finding suggests that leaf-attached microbes were derived from suspended particles, which could allow them to go back and forth between eelgrass leaves and the water column.

Gathering morphological and molecular data for identifying two species of trematodes from the neotropical Vaillant's frog, Lithobates vaillanti (Amphibia: Ranidae) in the highlands of Chiapas, south-eastern Mexico.

Mexico possesses a large diversity of amphibians partly due to its complex topography and transitional position between the Nearctic and Neotropical biogeographical regions. However, its helminth parasite fauna has been relatively poorly studied. Specimens of the Vaillant's frog, Lithobates vaillanti (Brocchi) were sampled in the tropical rain forest of Nahá, in the Chiapas Highlands, and examined for parasites. Two trematode species were collected from their hosts; morphologically, specimens were allocated to the genera Langeronia Caballero and Bravo-Hollis, 1949 and Haematoloechus Looss, 1899, respectively. Individuals were sequenced for two molecular markers (the mitochondrial cytochrome c oxidase gene, and the ribosomal gene 28S), and processed for morphological analyses, including scanning electron microscopy. The new evidence was not enough to accomplish the identification at species level of Langeronia sp. due to the lack of sequence data from the type localities of Langeronia parva Christian, 1970 and Langeronia macrocirra Caballero and Bravo-Hollis, 1949. Likewise, the newly generated data were useful to properly identify the adult specimens of lung flukes as Haematoloechus complexus Seely, 1906.

Extension of the known range of the snapping shrimp Alpheus christofferseni Anker, Hurt and Knowlton, 2007 (Caridea: Alpheidae)

Alpheus christofferseni Anker, Hurt and Knowlton, 2007 was described based on four specimens from Atol das Rocas, northeastern Brazil, and one specimen from Bocas del Toro, the Caribbean coast of Panama. Here, we report the collection of two specimens from Pernambuco, Brazil (~ 8°41’S), thus increasing the knowledge on species distribution along the Brazilian coast by approximately five latitudinal degrees south of its type locality (Atol das Rocas, Rio Grande do Norte, ~ 3°51'S). We also report a substantial increase in the known bathymetric distribution from shallow intertidal to approximately 51 m. Notes on morphological variation are provided based on the new material, and the first sequence of the 16S ribosomal subunit gene for the Brazilian coast is provided and compared with congeners.

Stephanostomum minankisi n. sp. (Digenea: Acanthocolpidae) from dusky flounder Syacium papillosum (Paralichthydae) from southern Gulf of Mexico: A new species without spines?

A new species of the genus Stephanostomum is described for the southeastern Gulf of Mexico based on morphological and nucleotide evidence. Stephanostomum minankisi n. sp. infects the intestine of the dusky flounder Syacium papillosum in the Yucatan Continental Shelf, Mexico (Yucatan Peninsula). Sequences of the 28S ribosomal gene were obtained and compared with available sequences of the other species and genera of the families Acanthocolpidae and Brachycladiidae from GenBank. A phylogenetic analysis was conducted, including 39 sequences, 26 of which represented 21 species and six genera of the family Acanthocolpidae. The new species is characterized by the absence of circumoral spines and spines on the tegument. Nonetheless, scanning electron microscopy consistently revealed the pits of 52 circumoral spines distributed in a double row with 26 spines each, and forebody spined. Other distinctive features of this species are testes in contact (sometimes overlapping), the vitellaria running along the body lateral fields to the mid-level of the cirrus-sac, pars prostatica and ejaculatory duct similar in length, and uroproct present. The phylogenetic tree showed that the three species found as parasites of dusky flounder (the new adult species and two in metacercaria stages) were grouped into two different clades. S. minankisi n. sp. was the sister species of Stephanostomum sp. 1 (Bt = 56) and formed a clade with S. tantabiddii, supported by high bootstrap values (100).