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1.
PLoS One ; 19(4): e0298905, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38578734

RESUMO

Nematodes are keystone actors of soil, freshwater and marine ecosystems, but the complexity of morphological identification has limited broad-scale monitoring of nematode biodiversity. DNA metabarcoding is increasingly used to assess nematode diversity but requires universal primers with high taxonomic coverage and high taxonomic resolution. Several primers have been proposed for the metabarcoding of nematode diversity, many of which target the 18S rRNA gene. In silico analyses have a great potential to assess key parameters of primers, including taxonomic coverage, resolution and specificity. Based on a recently-available reference database, we tested in silico the performance of fourteen commonly used and one newly optimized primer for nematode metabarcoding. Most primers showed very good coverage, amplifying most of the sequences in the reference database, while four markers showed limited coverage. All primers showed good taxonomic resolution. Resolution was particularly good if the aim was the identification of higher-level taxa, such as genera or families. Overall, species-level resolution was higher for primers amplifying long fragments. None of the primers was highly specific for nematodes as, despite some variation, they all amplified a large number of other eukaryotes. Differences in performance across primers highlight the complexity of the choice of markers appropriate for the metabarcoding of nematodes, which depends on a trade-off between taxonomic resolution and the length of amplified fragments. Our in silico analyses provide new insights for the identification of the most appropriate primers, depending on the study goals and the origin of DNA samples. This represents an essential step to design and optimize metabarcoding studies assessing nematode diversity.


Assuntos
Ecossistema , Nematoides , Humanos , Animais , DNA Ribossômico/genética , Código de Barras de DNA Taxonômico , Nematoides/genética , RNA Ribossômico 18S/genética , Biodiversidade
2.
Int J Mol Sci ; 25(7)2024 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-38612702

RESUMO

Cystic fibrosis (CF) is an inherited genetic disorder which manifests primarily in airway disease. Recent advances in molecular technologies have unearthed the diverse polymicrobial nature of the CF airway. Numerous studies have characterised the genus-level composition of this airway community using targeted 16S rDNA sequencing. Here, we employed whole-genome shotgun metagenomics to provide a more comprehensive understanding of the early CF airway microbiome. We collected 48 sputum samples from 11 adolescents and children with CF over a 12-month period and performed shotgun metagenomics on the Illumina NextSeq platform. We carried out functional and taxonomic analysis of the lung microbiome at the species and strain levels. Correlations between microbial diversity measures and independent demographic and clinical variables were performed. Shotgun metagenomics detected a greater diversity of bacteria than culture-based methods. A large proportion of the top 25 most-dominant species were anaerobes. Samples dominated by Staphylococcus aureus and Prevotella melaninogenica had significantly higher microbiome diversity, while no CF pathogen was associated with reduced microbial diversity. There was a diverse resistome present in all samples in this study, with 57.8% agreement between shotgun metagenomics and culture-based methods for detection of resistance. Pathogenic sequence types (STs) of S. aureus, Pseudomonas aeruginosa, Haemophilus influenzae and Stenotrophomonas maltophilia were observed to persist in young CF patients, while STs of S. aureus were both persistent and shared between patients. This study provides new insight into the temporal changes in strain level composition of the microbiome and the landscape of the resistome in young people with CF. Shotgun metagenomics could provide a very useful one-stop assay for detecting pathogens, emergence of resistance and conversion to persistent colonisation in early CF disease.


Assuntos
Fibrose Cística , Microbiota , Criança , Humanos , Adolescente , Staphylococcus aureus , Bioensaio , DNA Ribossômico , Microbiota/genética
3.
J Helminthol ; 98: e32, 2024 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-38618914

RESUMO

Two new species of the genus Sectonema found in northern Iran are characterized, including morphological descriptions and molecular (18S-, 28S-rDNA) analyses. Sectonema tehranense sp. nov. is distinguished by its 7.22 - 8.53 mm long body, lip region offset by constriction and 24 - 31 µm wide with perioral lobes and abundant setae- or cilia-like projections covering the oral field, mural tooth 15.5 - 17 µm long at its ventral side, neck 1091 - 1478 µm long, pharyngeal expansion occupying 61 - 71% of the total neck length, female genital system diovarian, uterus simple and 3.9 - 4.2 times the corresponding body diameter long, transverse vulva (V = 49 - 59), tail short and rounded (44 - 65 µm, c = 99 - 162, c' = 0.6 - 0.8), spicules 111 - 127 µm long, and 7 - 10 spaced ventromedian supplements with hiatus. Sectonema noshahrense sp. nov. displays a 4.07 - 4.73 mm long body, lip region offset by constriction and 23 - 25 µm wide with perioral lobes and abundant setae- or cilia-like projections covering the oral field, odontostyle 14 - 14.5 µm long, neck 722 - 822 µm long, pharyngeal expansion occupying 66 - 68% of the total neck length, female genital system diovarian, uterus simple and 2.4 - 2.7 times the corresponding body diameter long, transverse vulva (V = 54 - 55), tail convex conoid (39 - 47 µm, c = 91 - 111, c' = 0.8 - 0.9), spicules 82 µm long, and seven spaced ventromedian supplements with hiatus. Molecular analyses confirm a maximally supported (Epacrolaimus + Metaporcelaimus + Sectonema) clade and a tentative biogeographical pattern, with sequences of Indolamayan taxa forming a clade separated from those of Palearctic ones. Parallel or convergent evolution processes might be involved in the phylogeny of the species currently classified under Sectonema. This genus is certainly more heterogeneous than previously assumed.


Assuntos
Helmintos , Nematoides , Feminino , Animais , Irã (Geográfico) , Citoesqueleto , DNA Ribossômico/genética , Nematoides/genética
4.
J Orthop Surg Res ; 19(1): 236, 2024 Apr 12.
Artigo em Inglês | MEDLINE | ID: mdl-38609952

RESUMO

OBJECTIVE: Osteonecrosis of the femoral head (ONFH) is a severe disease that primarily affects the middle-aged population, imposing a significant economic and social burden. Recent research has linked the progression of non-traumatic osteonecrosis of the femoral head (NONFH) to the composition of the gut microbiota. Steroids and alcohol are considered major contributing factors. However, the relationship between NONFH caused by two etiologies and the microbiota remains unclear. In this study, we examined the gut microbiota and fecal metabolic phenotypes of two groups of patients, and analyzed potential differences in the pathogenic mechanisms from both the microbial and metabolic perspectives. METHODS: Utilizing fecal samples from 68 NONFH patients (32 steroid-induced, 36 alcohol-induced), high-throughput 16 S rDNA sequencing and liquid chromatography with tandem mass spectrometry (LC-MS/MS) metabolomics analyses were conducted. Univariate and multivariate analyses were applied to the omics data, employing linear discriminant analysis effect size to identify potential biomarkers. Additionally, functional annotation of differential metabolites and associated pathways was performed using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Subsequently, Spearman correlation analysis was employed to assess the potential correlations between differential gut microbiota and metabolites. RESULTS: High-throughput 16 S rDNA sequencing revealed significant gut microbial differences. At the genus level, the alcohol group had higher Lactobacillus and Roseburia, while the steroid group had more Megasphaera and Akkermansia. LC-MS/MS metabolomic analysis indicates significant differences in fecal metabolites between steroid- and alcohol-induced ONFH patients. Alcohol-induced ONFH (AONFH) showed elevated levels of L-Lysine and Oxoglutaric acid, while steroid-induced ONFH(SONFH) had increased Gluconic acid and Phosphoric acid. KEGG annotation revealed 10 pathways with metabolite differences between AONFH and SONFH patients. Correlation analysis revealed the association between differential gut flora and differential metabolites. CONCLUSIONS: Our results suggest that hormones and alcohol can induce changes in the gut microbiota, leading to alterations in fecal metabolites. These changes, driven by different pathways, contribute to the progression of the disease. The study opens new research directions for understanding the pathogenic mechanisms of hormone- or alcohol-induced NONFH, suggesting that differentiated preventive and therapeutic approaches may be needed for NONFH caused by different triggers.


Assuntos
Microbioma Gastrointestinal , Pessoa de Meia-Idade , Humanos , Cabeça do Fêmur , Cromatografia Líquida , Espectrometria de Massas em Tandem , Etanol , Esteroides/efeitos adversos , DNA Ribossômico
5.
Environ Microbiol ; 26(3): e16615, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38501240

RESUMO

Microbial communities are commonly characterised through the metabarcoding of environmental DNA. This DNA originates from both viable (including dormant and active) and dead organisms, leading to recent efforts to distinguish between these states. In this study, we further these approaches by distinguishing not only between viable and dead cells but also between dormant and actively growing cells. This is achieved by sequencing both rRNA and rDNA, in conjunction with propidium monoazide cross-linked rDNA, to partition the active, dormant and relic fractions in environmental samples. We apply this method to characterise the diversity and assemblage structure of these fractions of microeukaryotes in intertidal sediments during a wet-dry-rewet incubation cycle. Our findings indicate that a significant proportion of microeukaryotic phylotypes detected in the total rDNA pools originate from dormant and relic microeukaryotes in the sediments, both in terms of richness (dormant, 13 ± 2%; relic, 47 ± 5%) and read abundance (dormant, 20 ± 7%; relic, 14 ± 5%). The richness and sequence proportion of dormant microeukaryotes notably increase during the transition from wet to dry conditions. Statistical analyses suggest that the dynamics of diversity and assemblage structure across different activity fractions are influenced by various environmental drivers. Our strategy offers a versatile approach that can be adapted to characterise other microbes in a wide range of environments.


Assuntos
Microbiota , Microbiota/genética , DNA Ribossômico/genética
6.
BMC Vet Res ; 20(1): 104, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38491459

RESUMO

BACKGROUND: members of the genus Sarcocystis are intracellular obligate protozoan parasites classified within the phylum Apicomplexa and have an obligate heteroxenous life cycle involving two hosts. A more comprehensive understanding of the prevalence and geographic range of different Sarcocystis species in marine ecosystems is needed globally and nationally. Hence, the objective of this study was to document the incidence of Sarcocystis infection in sharks within the aquarium ecosystem of Egypt and to identify the species through the characterization of the SSU rDNA gene. METHODS: All organs of the mako shark specimen underwent macroscopic screening to detect the existence of a Sarcocystis cyst. Ten cysts were collected from the intestine and processed separately to extract the genomic DNA. The polymerase chain reaction (PCR) was accomplished by amplifying a specific 18S ribosomal RNA (rRNA) gene fragment. Subsequently, the resulting amplicons were subjected to purification and sequencing processes. RESULTS: Macroscopic examination of the mako shark intestinal wall sample revealed the presence of Sarcocystis cysts of various sizes and shapes, and sequencing of the amplicons from Sarcocystis DNA revealed a 100% nucleotide identity with the sequence of Sarcocystis tenella recorded from sheep in Iran; The mako shark sequence has been deposited in the GeneBank with the accession number OQ721979. This study presents the first scientific evidence demonstrating the presence of the Sarcocystis parasite in sharks, thereby documenting this specific marine species as a novel intermediate host in the Sarcocystis life cycle. CONCLUSIONS: This is the first identification of Sarcocystis infection in sharks, and we anticipate it will be an essential study for future screenings and establishing effective management measures for this disease in aquatic ecosystems.


Assuntos
Sarcocystis , Tubarões , Animais , Ovinos/genética , Sarcocystis/genética , Ecossistema , Tubarões/genética , Filogenia , Oceano Índico , DNA Ribossômico , Estágios do Ciclo de Vida
7.
mSystems ; 9(3): e0000824, 2024 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-38426796

RESUMO

The crucial function of circulating microbial DNA (cmDNA) in peripheral blood is gaining recognition because of its importance in normal physiology and immunity in healthy individuals. Evidence suggests that cmDNA in peripheral blood is derived from highly abundant, translocating gut microbes. However, the associations with and differences between cmDNA in peripheral blood and the gut microbiome remain unclear. We collected blood, urine, and fecal samples from volunteers to compare their microbial information via 16S rDNA sequencing. The results revealed that, compared with gut microbial DNA, cmDNA in peripheral blood was associated with reduced diversity and a distinct microbiota composition. The cmDNA in the blood reflects the biochemical processes of microorganisms, including synthesis, energy conversion, degradation, and adaptability, surpassing that of fecal samples. Interestingly, cmDNA in blood showed a limited presence of DNA from anaerobes and gram-positive bacteria, which contrast with the trend observed in fecal samples. Furthermore, analysis of cmDNA revealed traits associated with mobile elements and potential pathologies, among others, which were minimal in stool samples. Notably, cmDNA analysis indicated similarities between the microbial functions and phenotypes in blood and urine samples, although greater diversity was observed in urine samples. Source Tracker analysis suggests that gut microbes might not be the main source of blood cmDNA, or a selective mechanism allows only certain microbial DNA into the bloodstream. In conclusion, our study highlights the composition and potential functions associated with cmDNA in peripheral blood, emphasizing its selective presence; however, further research is required to elucidate the mechanisms involved.IMPORTANCEOur research provides novel insights into the unique characteristics and potential functional implications of circulating microbial DNA (cmDNA) in peripheral blood. Unlike other studies that analyzed sequencing data from fecal or blood microbiota in different study cohorts, our comparative analysis of cmDNA from blood, urine, and fecal samples from the same group of volunteers revealed a distinct blood-specific cmDNA composition. We discovered a decreased diversity of microbial DNA in blood samples compared to fecal samples as well as an increased presence of biochemical processes microbial DNA in blood. Notably, we add to the existing knowledge by documenting a reduced abundance of anaerobes and gram-positive bacteria in blood compared to fecal samples according to the analysis of cmDNA and gut microbial DNA, respectively. This observation suggested that a potential selective barrier or screening mechanism might filter microbial DNA molecules, indicating potential selectivity in the translocation process which contrasts with the traditional view that cmDNA primarily originates from random translocation from the gut and other regions. By highlighting these differences, our findings prompt a reconsideration of the origin and role of cmDNA in blood circulation and suggest that selective processes involving more complex biological mechanisms may be involved.


Assuntos
Microbioma Gastrointestinal , Microbiota , Humanos , Fezes/química , Microbioma Gastrointestinal/genética , DNA Ribossômico/análise , Análise de Sequência de DNA
8.
Bioinformatics ; 40(3)2024 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-38441320

RESUMO

MOTIVATION: The ribosomal DNA (rDNA) arrays are highly repetitive and homogenous regions which exist in all life. Due to their repetitiveness, current assembly methods do not fully assemble the rDNA arrays in humans and many other eukaryotes, and so variation within the rDNA arrays cannot be effectively studied. RESULTS: Here, we present the tool ribotin to assemble full length rDNA copies, or morphs. Ribotin uses a combination of highly accurate long reads and extremely long nanopore reads to resolve the variation between rDNA morphs. We show that ribotin successfully recovers the most abundant morphs in human and nonhuman genomes. We also find that genome wide consensus sequences of the rDNA arrays frequently produce a mosaic sequence that does not exist in the genome. AVAILABILITY AND IMPLEMENTATION: Ribotin is available on https://github.com/maickrau/ribotin and as a package on bioconda.


Assuntos
Genoma , Software , Humanos , DNA Ribossômico/genética , Análise de Sequência de DNA/métodos , Eucariotos/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos
9.
Behav Brain Res ; 464: 114930, 2024 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-38432300

RESUMO

Depression is a serious disease that has a significant impact on social functioning. However, the exact causes of depression are still not fully understood. Therefore, it is necessary to explore new pathways leading to depression. In this study, we used 16 S rDNA to examine changes in gut microbiota and predict related pathways in depression-like mice. Additionally, we employed liquid chromatography-mass spectrometry (LC-MS) to identify changes in amino acids and gas chromatography-mass spectrometry (GC-MS) to identify changes in short-chain fatty acids (SCFAs) in fecal samples. We conducted Pearson/Spearman correlation analysis to investigate the associations between changes in amino acids/SCFAs and behavioral outcomes. The 16 S rDNA sequencing revealed significant alterations in gut microbiota at the phylum and genus levels in mice subjected to chronic unpredictable mild stress (CUMS). The relative abundances of Bacteroidetes, Proteobacteria, Bacteroides, and Alloprevotella were increased, while Firmicutes, Verrucomicrobia, Actinobacteria, Lactobacillus, Akkermansia, Lachnospirillum, and Enterobacter were decreased in the CUMS mice. We used PICRUSt software to annotate the kyoto encyclopedia of genes and genomes (KEGG) pathway function related to depression-like behavior in mice. Our analysis identified sixty functional pathways associated with the gut microbiota of mice exhibiting depression-like behavior. In the amino acid concentration analysis, we observed decreased levels of hydroxyproline and tryptophan, and increased levels of alanine in CUMS mice. In the SCFAs concentration assay, we found decreased levels of butyric acid and valeric acid, and increased levels of acetic acid in CUMS mice. Some of these changes were significantly correlated with depressive-like behaviors. Our study contributes to the understanding of the mechanism of the gut-brain axis in the occurrence and development of depression.


Assuntos
Aminoácidos , Microbioma Gastrointestinal , Animais , Camundongos , Ácidos Graxos Voláteis , Ácido Butírico , Bacteroidetes , DNA Ribossômico , Depressão
10.
Biol Pharm Bull ; 47(3): 680-691, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38522942

RESUMO

Cholelithiasis, commonly known as gallstones, represents a prevalent hepatobiliary disorder. This study aimed to elucidate the therapeutic role and mechanism of Danyankang capsulein treating cholelithiasis induced by a high-fat diet in C57BL/6 mice. The therapeutical potential of Danyankang was assessed through biochemical analyses, histopathological examinations, protein detection, and 16S rDNA sequencing. A high-fat diet resulted in cholelithiasis manifestation in mice, with discernable abnormal serum biochemical indices and disrupted biliary cholesterol homeostasis. Danyankang treatment notably ameliorated liver inflammation symptoms and rectified serum and liver biochemical abnormalities. Concurrently, it addressed biliary imbalances. Elevated expressions of toll-like receptor 4 (TLR4), nuclear factor-kappaB (NF-κB)/pNF-κB, HMGCR, CYP7A1, and CYP8B1 observed at the inception of cholelithiasis, were notably reduced upon Danyankang administration. Furthermore, 16S rDNA analysis revealed a decline in species number and diversity of the intestinal flora in cholelithiasis-treated mice, while the decline was reversed with Danyankang treatment. Danyankang capsules reduced the abundance of Verrucomicrobiota and increased the abundance of Actinobacteriota and Proteobacteria. In conclusion, the present study demonstrates that Danyankang exerts potent therapeutic efficacy against high-fat diet-induced cholelithiasis. This beneficial outcome is potentially linked to the inhibition of the TLR4/pNF-κB and SHP/CYP7A1/CYP8B1 signaling pathways, as well as the enhancement of intestinal flora species abundance.


Assuntos
Colelitíase , Microbioma Gastrointestinal , Camundongos , Animais , Dieta Hiperlipídica/efeitos adversos , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo , Esteroide 12-alfa-Hidroxilase , Camundongos Endogâmicos C57BL , Fígado/metabolismo , NF-kappa B/metabolismo , Colelitíase/tratamento farmacológico , Colelitíase/patologia , DNA Ribossômico
11.
Sci Rep ; 14(1): 6442, 2024 03 18.
Artigo em Inglês | MEDLINE | ID: mdl-38499675

RESUMO

Two gene regions commonly used to characterise the diversity of eukaryotic communities using metabarcoding are the 18S ribosomal DNA V4 and V9 gene regions. We assessed the effectiveness of these two regions for characterising diverisity of coastal eukaryotic microalgae communities (EMCs) from tropical and temperate sites. We binned amplicon sequence variants (ASVs) into the high level taxonomic groups: dinoflagellates, pennate diatoms, radial centric diatoms, polar centric diatoms, chlorophytes, haptophytes and 'other microalgae'. When V4 and V9 generated ASV abundances were compared, the V9 region generated a higher number of raw reads, captured more diversity from all high level taxonomic groups and was more closely aligned with the community composition determined using light microscopy. The V4 region did resolve more ASVs to a deeper taxonomic resolution within the dinoflagellates, but did not effectively resolve other major taxonomic divisions. When characterising these communities via metabarcoding, the use of multiple gene regions is recommended, but the V9 gene region can be used in isolation to provide high-level community biodiversity to reflect relative abundances within groups. This approach reduces the cost of sequencing multiple gene regions whilst still providing important baseline ecosystem function information.


Assuntos
Diatomáceas , Dinoflagelados , Microalgas , Ecossistema , Microalgas/genética , Biodiversidade , Diatomáceas/genética , DNA Ribossômico/genética , Dinoflagelados/genética , RNA Ribossômico 18S/genética , Filogenia
12.
PLoS One ; 19(3): e0298814, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38507310

RESUMO

Here, we describe a novel water mold species, Saprolegnia velencensis sp. n. from Lake Velence, in Hungary. Two strains (SAP239 and SAP241) were isolated from lake water, and characterized using morphological and molecular markers. In addition, phylogenetic analyses based on ITS-rDNA regions and on the RNA polymerase II B subunit (RPB2) gene complemented the study. The ITS-rDNA of the two strains was 100% identical, showed the highest similarity to that of S. ferax (with 94.4% identity), and they formed a separate cluster in both the ITS-rDNA and RPB2-based maximum likelihood phylogenetic trees with high bootstrap support. Although mature oogonia and antheridia were not seen under in vitro conditions, the S. velencensis sp. n. could be clearly distinguished from its closest relative, S. ferax, by the length and width of sporangia, as the new species had shorter and narrower sporangia (163.33±70.07 and 36.69±8.27 µm, respectively) than those of S. ferax. The two species also differed in the size of the secondary cysts (11.63±1.77 µm), which were slightly smaller in S. ferax. Our results showed that S. velencensis sp. n. could not be identified with any of the previously described water mold species, justifying its description as a new species.


Assuntos
Saprolegnia , Saprolegnia/genética , Hungria , Lagos , Filogenia , Fungos/genética , DNA Ribossômico/genética , Água
13.
Plant Cell Rep ; 43(4): 95, 2024 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-38472393

RESUMO

KEY MESSAGE: Both bacterial and fungal endophytes exhibited one or more plant growth-promoting (PGP) traits. Among these strains, the Paenibacillus peoriae SYbr421 strain demonstrated the greatest activity in the direct biotransformation of tuber powder from D. nipponica into diosgenin. Endophytes play crucial roles in shaping active metabolites within plants, significantly influencing both the quality and yield of host plants. Dioscorea nipponica Makino accumulates abundant steroidal saponins, which can be hydrolyzed to produce diosgenin. However, our understanding of the associated endophytes and their contributions to plant growth and diosgenin production is limited. The present study aimed to assess the PGP ability and potential of diosgenin biotransformation by endophytes isolates associated with D. nipponica for the efficient improvement of plant growth and development of a clean and effective approach for producing the valuable drug diosgenin. Eighteen bacterial endophytes were classified into six genera through sequencing and phylogenetic analysis of the 16S rDNA gene. Similarly, 12 fungal endophytes were categorized into 5 genera based on sequencing and phylogenetic analysis of the ITS rDNA gene. Pure culture experiments revealed that 30 isolated endophytic strains exhibited one or more PGP traits, such as nitrogen fixation, phosphate solubilization, siderophore synthesis, and IAA production. One strain of endophytic bacteria, P. peoriae SYbr421, effectively directly biotransformed the saponin components in D. nipponica. Moreover, a high yield of diosgenin (3.50%) was obtained at an inoculum size of 4% after 6 days of fermentation. Thus, SYbr421 could be used for a cleaner and more eco-friendly diosgenin production process. In addition, based on the assessment of growth-promoting isolates and seed germination results, the strains SYbr421, SYfr1321, and SYfl221 were selected for greenhouse experiments. The results revealed that the inoculation of these promising isolates significantly increased the plant height and fresh weight of the leaves and roots compared to the control plants. These findings underscore the importance of preparing PGP bioinoculants from selected isolates as an additional option for sustainable diosgenin production.


Assuntos
Dioscorea , Diosgenina , Endófitos/genética , Endófitos/metabolismo , Dioscorea/genética , Dioscorea/microbiologia , Diosgenina/metabolismo , Filogenia , Raízes de Plantas , DNA Ribossômico/metabolismo
14.
Mar Pollut Bull ; 201: 116198, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38428045

RESUMO

Metabarcoding analysis is an effective technique for monitoring the domoic acid-producing Pseudo-nitzschia species in marine environments, uncovering high-levels of molecular diversity. However, such efforts may result in the overinterpretation of Pseudo-nitzschia species diversity, as molecular diversity not only encompasses interspecies and intraspecies diversities but also exhibits extensive intragenomic variations (IGVs). In this study, we analyzed the V4 region of the 18S rDNA of 30 strains of Pseudo-nitzschia multistriata collected from the coasts of China. The results showed that each P. multistriata strain harbored about a hundred of unique 18S rDNA V4 sequence varieties, of which each represented by a unique amplicon sequence variant (ASV). This study demonstrated the extensive degree of IGVs in P. multistriata strains, suggesting that IGVs may also present in other Pseudo-nitzschia species and other phytoplankton species. Understanding the scope and levels of IGVs is crucial for accurately interpreting the results of metabarcoding analysis.


Assuntos
Diatomáceas , Diatomáceas/genética , DNA Ribossômico , Fitoplâncton/genética , Sequenciamento de Nucleotídeos em Larga Escala , China
15.
PeerJ ; 12: e16627, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38500531

RESUMO

Background: Dung beetles provide many important ecosystem services, including dung decomposition, pathogen control, soil aeration, and secondary seed dispersal. Yet, the biology of most dung beetles remains unknown. Natural diets are poorly studied, partly because previous research has focused on choice or attraction experiments using few, easily accessible dung types from zoo animals, farm animals, or humans. This way, many links within natural food webs have certainly been missed. In this work, we aimed to establish a protocol to analyze the natural diets of dung beetles using DNA gut barcoding. Methods: First, the feasibility of gut-content DNA extraction and amplification of 12s rDNA from six different mammal dung types was tested in the laboratory. We then applied the method to beetles caught in pitfall traps in Ecuador and Germany by using 12s rDNA primers. For a subset of the dung beetles caught in the Ecuador sampling, we also used 16s rDNA primers to see if these would improve the number of species we could identify. We predicted the likelihood of amplifying DNA using gut fullness, DNA concentration, PCR primer, collection method, and beetle species as predictor variables in a dominance analysis. Based on the gut barcodes, we generated a dung beetle-mammal network for both field sites (Ecuador and Germany) and analyzed the levels of network specificity. Results: We successfully amplified mammal DNA from dung beetle gut contents for 128 specimens, which included such prominent species as Panthera onca (jaguar) and Puma concolor (puma). The overall success rate of DNA amplification was 53%. The best predictors for amplification success were gut fullness and DNA concentration, suggesting the success rate can be increased by focusing on beetles with a full gut. The mammal dung-dung beetle networks differed from purely random network models and showed a moderate degree of network specialization (H2': Ecuador = 0.49; Germany = 0.41). Conclusion: We here present a reliable method of extracting and amplifying gut-content DNA from dung beetles. Identifying mammal dung via DNA reference libraries, we created mammal dung-dung beetle trophic networks. This has benefits over previous methods because we inventoried the natural mammal dung resources of dung beetles instead of using artificial mammal baits. Our results revealed higher levels of specialization than expected and more rodent DNA than expected in Germany, suggesting that the presented method provides more detailed insights into mammal dung-dung beetle networks. In addition, the method could have applications for mammal monitoring in many ecosystems.


Assuntos
Besouros , Ecossistema , Animais , Besouros/genética , DNA Ribossômico , Fezes , Mamíferos
16.
Parasitol Res ; 123(3): 158, 2024 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-38460006

RESUMO

Cryptosporidium spp., Enterocytozoon bieneusi and Encephalitozoon spp. are the most common protistan parasites of vertebrates. The results show that pigeon populations in Central Europe are parasitised by different species of Cryptosporidium and genotypes of microsporidia of the genera Enterocytozoon and Encephalitozoon. A total of 634 and 306 faecal samples of captive and feral pigeons (Columba livia f. domestica) from 44 locations in the Czech Republic, Slovakia and Poland were analysed for the presence of parasites by microscopy and PCR/sequence analysis of small subunit ribosomal RNA (18S rDNA), 60 kDa glycoprotein (gp60) and internal transcribed spacer (ITS) of SSU rDNA. Phylogenetic analyses revealed the presence of C. meleagridis, C. baileyi, C. parvum, C. andersoni, C. muris, C. galli and C. ornithophilus, E. hellem genotype 1A and 2B, E. cuniculi genotype I and II and E. bieneusi genotype Peru 6, CHN-F1, D, Peru 8, Type IV, ZY37, E, CHN4, SCF2 and WR4. Captive pigeons were significantly more frequently parasitised with screened parasite than feral pigeons. Cryptosporidium meleagridis IIIa and a new subtype IIIl have been described, the oocysts of which are not infectious to immunodeficient mice, whereas chickens are susceptible. This investigation demonstrates that pigeons can be hosts to numerous species, genotypes and subtypes of the studied parasites. Consequently, they represent a potential source of infection for both livestock and humans.


Assuntos
Criptosporidiose , Cryptosporidium , Encephalitozoon , Enterocytozoon , Microsporidiose , Humanos , Animais , Camundongos , Columbidae , Enterocytozoon/genética , Cryptosporidium/genética , Encephalitozoon/genética , Criptosporidiose/epidemiologia , Criptosporidiose/parasitologia , Microsporidiose/epidemiologia , Microsporidiose/veterinária , Microsporidiose/parasitologia , Filogenia , Galinhas , Europa (Continente)/epidemiologia , DNA Ribossômico , Variação Genética , Genótipo , Fezes/parasitologia
17.
Parasites Hosts Dis ; 62(1): 85-97, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38443773

RESUMO

This study aimed to describe the morphological and molecular characteristics of Paralecithodendrium longiforme (Digenea: Lecithodendriidae) adults and cercariae isolated in Thailand. Adult flukes were isolated from the Chinese pipistrelle bat (Hypsugo sp.), and cercariae were detected in the viviparid snail (Filopaludina martensi martensi) from Chiang Mai province. The morphological characteristics were observed and described using conventional methods, and the molecular characteristics with internal transcribed spacer 2 (ITS2) and 28S rDNA gene sequences. The adult flukes were fusiform, 0.84-0.98 mm in length, and 0.37-0.49 mm in width, and were distinguishable from other species by the presence of longitudinal uterine coils. The cercariae were nonvirgulate xiphidiocercariae, with the oral sucker bigger than the acetabulum, the tail without fin fold, a body size of 117.5-138.3 × 48.3-52.2 µm, and a tail size of 100.7-103.7 × 15.0-18.9 µm. Molecular studies revealed that the adults and cercariae shared 99.3% (ITS2) and 99.6% (28S rDNA) homology with each other. They were phylogenetically close to P. longiforme with an identity of 94.5% for ITS2 and 98.7% for 28S rDNA. This study provides new information on the natural definitive host and first intermediate host of P. longiforme in Thailand. The discovery of its cercarial stage in Filopaludina snails highlights the importance of monitoring the associated second intermediate host and prevention and control of this potentially zoonotic trematode.


Assuntos
Quirópteros , Trematódeos , Animais , Tailândia , Trematódeos/genética , Cercárias/genética , DNA Ribossômico/genética , Caramujos/genética , China
18.
Methods Mol Biol ; 2756: 247-255, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38427297

RESUMO

DGGE (denaturing gradient gel electrophoresis) is a nucleic acid separation technique applied to the evaluation of microbial biodiversity. This technique is quite rapid and cheap compared to other types of analysis. Here we describe the comparison of nematode communities inhabiting different ecosystems. After an ecologically representative sampling collection and the nematode extraction from soil, nematodes are centrifuged in Eppendorf tubes to facilitate DNA extraction. DNA from the whole community of each type of soil is extracted, amplified with primers for 18 S rDNA and used in DGGE analysis. The profiles of DGGE can be analyzed with appropriate software, and biodiversity indices can be estimated.


Assuntos
Ecossistema , Nematoides , Animais , Biodiversidade , DNA Ribossômico/genética , Reação em Cadeia da Polimerase/métodos , Nematoides/genética , Solo , Eletroforese em Gel de Poliacrilamida , DNA Bacteriano/genética , RNA Ribossômico 16S/genética , Microbiologia do Solo
19.
Food Res Int ; 182: 114173, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38519188

RESUMO

Wet-type grinder (WG) is a nanofiber technology used to atomize dietary fiber-rich materials. WG-treated okara (WGO) exhibits high dispersion and viscosity similar to those of viscous soluble dietary fibers. Here, we studied the effect of WGO supplementation on obesity and gut microbiota composition in high-fat diet (HFD)-fed mice. WGO intake suppressed body weight gain and fat accumulation, improved glucose tolerance, lowered cholesterol levels, and prevented HFD-induced decrease in muscle mass. WGO supplementation also led to cecum enlargement, lower pH, and higher butyrate production. The bacterial 16S ribosomal RNA genes (16S rDNA) were sequenced to determine the gut microbiota composition of the fecal samples. Sequencing of bacterial 16S rDNA revealed that WGO treatment increased the abundance of butyrate producer Ruminococcus and reduced the abundances of Rikenellaceae, Streptococcaceae, and Prevotellaceae, which are related to metabolic diseases. Metabolomics analysis of the plasma of WGO- and cellulose-treated mice were conducted using ultra-high-performance liquid chromatography-mass spectrometry. Metabolic pathway analysis revealed that the primary bile acid biosynthesis pathway was significantly positively regulated by WGO intake instead of cellulose. These results demonstrate that WG is useful for improving functional properties of okara to prevent metabolic syndromes, including obesity, diabetes, and dyslipidemia.


Assuntos
Microbioma Gastrointestinal , Camundongos , Animais , Obesidade/tratamento farmacológico , Obesidade/prevenção & controle , Obesidade/metabolismo , Celulose/farmacologia , Butiratos , DNA Ribossômico/farmacologia
20.
J Ethnopharmacol ; 327: 117989, 2024 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-38462026

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Massa Medicata Fermentata, a fermented Chinese medicine, is produced by the fermentation of six traditional Chinese medicines. Liu Shenqu (LSQ) and charred Liu Shenqu (CLSQ) have been used for strengthening the spleen and enhancing digestion for over a thousand years, and CLSQ is commonly used in clinical practice. However, it is unclear whether there is a difference in the spleen strengthening and digestion effects between LSQ and CLSQ, as well as their mechanisms of action. AIM OF STUDY: This study aims to compare the effects of LSQ and CLSQ on the digestive function of functional dyspepsia (FD) rats and reveal their mechanisms of action. MATERIALS AND METHODS: SPF grade SD rats were randomly divided into 6 groups: control group, model group, Liu Shenqu decoction low-dosage (LSQ LD) group, Liu Shenqu decoction high-dosage (LSQ HD) group, charred Liu Shenqu decoction low-dosage (CLSQ LD) group, and charred Liu Shenqu decoction high-dosage (CLSQ HD) group. Rats were injected intraperitoneally with reserpine to create an FD model and then treated by intragastric administration. During this period, record the weight and food intake of the animals. After 18 days of treatment, specimens of the gastric antrum, spleen, and duodenum of rats were taken for pathological staining and immunohistochemical detection of Ghrelin protein expression. Enzyme linked immunosorbent assay (ELISA) was used to determine the concentration of relevant gastrointestinal hormones in serum. The 16 S rDNA sequencing method was used to evaluate the effect of cecal contents on the structure of the gut microbiota in experimental rats. Plasma metabolomics analysis was performed using ultra high performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UPLC-QTOF-MS) to further reveal their mechanism of action. RESULTS: LSQ and CLSQ improved the pathological tissue histological structure of FD rats and increased the levels of MTL and GAS hormones in serum and the levels of ghrelin in the gastric antrum, spleen, and duodenum, while reducing VIP, CCK, and SP hormone levels. The above results showed that the therapeutic efficacy of CLSQ is better than that of LSQ. Futhermore, the mechanism of action of LSQ and CLSQ were revealed. The 16 S rDNA sequencing results showed that both LSQ and CLSQ can improve the composition and diversity of the gut microbiota. And metabolomic analysis demonstrated that 20 metabolites changed after LSQ treatment, and 16 metabolites underwent continuous changes after CLSQ treatment. Further analysis revealed that LSQ mainly intervened in the metabolic pathways of glycerol phospholipid metabolism and arginine and proline metabolism, but CLSQ mainly intervened in the metabolic pathways of ether lipid metabolism, sphingolipid metabolism, and glycerophospholipid metabolism. CONCLUSIONS: Both LSQ and CLSQ can improve functional dyspepsia in FD rats, but CLSQ has a stronger improvement effect on FD. Although their mechanisms of action are all related to regulating gastrointestinal hormone secretion, significantly improving intestinal microbiota disorders, and improving multiple metabolic pathways, but the specific gut microbiota and metabolic pathways they regulate are different.


Assuntos
Medicamentos de Ervas Chinesas , Dispepsia , Microbiota , Ratos , Animais , Grelina/uso terapêutico , Dispepsia/tratamento farmacológico , Ratos Sprague-Dawley , Medicamentos de Ervas Chinesas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Metabolômica/métodos , DNA Ribossômico
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