The gut microbiota is currently recognized as an important factor regulating the homeostasis of the gastrointestinal tract and influencing the energetic metabolism of the host as well as its immune ...and central nervous systems. Determining the gut microbiota composition of healthy subjects is therefore necessary to establish a baseline allowing the detection of microbiota alterations in pathologic conditions. Accordingly, the aim of this study was to characterize the gut microbiota of healthy Chilean subjects using 16S rRNA gene sequencing. Fecal samples were collected from 41 young, asymptomatic, normal weight volunteers (age: 25 ± 4 years; ♀:48.8%; BMI: 22.5 ± 1.6 kg/m
) with low levels of plasma (IL6 and hsCRP) and colonic (fecal calprotectin) inflammatory markers. The V3-V4 region of the 16S rRNA gene of bacterial DNA was amplified and sequenced using MiSeq Illumina system. 109,180 ± 13,148 sequences/sample were obtained, with an α-diversity of 3.86 ± 0.37. The dominant phyla were Firmicutes (43.6 ± 9.2%) and Bacteroidetes (41.6 ± 13.1%), followed by Verrucomicrobia (8.5 ± 10.4%), Proteobacteria (2.8 ± 4.8%), Actinobacteria (1.8 ± 3.9%) and Euryarchaeota (1.4 ± 2.7%). The core microbiota representing the genera present in all the subjects included
,
,
(phylum Bacteroidetes),
,
,
,
,
,
,
,
,
,
,
(phylum Firmicutes),
(phylum Verrucomicrobia), and
(phylum Actinobacteria). Butyrate-producing genera including
,
,
, and
were detected. The family Methanobacteriaceae was reported in 83% of the subjects and
, the most representative sulfate-reducing genus, in 76%. The microbiota of the Chilean individuals significantly differed from those of Papua New Guinea and the Matses ethnic group and was closer to that of the Argentinians and sub-populations from the United States. Interestingly, the microbiota of the Chilean subjects stands out for its richness in Verrucomicrobia; the mucus-degrading bacterium
is the only identified member of this phylum. This is an important finding considering that this microorganism has been recently proposed as a hallmark of healthy gut due to its anti-inflammatory and immunostimulant properties and its ability to improve gut barrier function, insulin sensitivity and endotoxinemia. These results constitute an important baseline that will facilitate the characterization of dysbiosis in the main diseases affecting the Chilean population.
This study was done to determine the percentage of Salmonella spp. in camels from three provinces (Karbala, Al-Najaf and AL-Muthana) in Iraq with different age and both sexes. Total of 250 fecal ...samples from 250 camels were collected. Diagnostic study depended upon the morphological and cultural properties of the isolates on some selective media like Xylose lysine deoxycholate (XLD) and Salmonella Shigella (SS) agars which were used in addition to different biochemical tests and molecular assay by PCR for detection of virulence gene invasion A (invA) with Phylogenetic study. The clinical signs appearing on animals infected with Salmonella were greenish diarrhea, loss of appetite with mild systemic reaction. Bacteriological and molecular tests revealed isolation of five Salmonella isolates with invA gene. Two of these isolates were sequenced. The results showed that the first strain S. enterica subspecies typhimurium (LC730846) converged with a group of global strains with one node, as it converged with the global strain that held the clade (MK017934.1 and MT460418.1). While the second local strain S. enterica serovar enteritidis (LC730849) appeared with a new node and it is not affiliated with any association with the world S. enterica strains. It is concluded that the presence of Salmonella spp. in camel needs monitoring in order to minimize the risks of infection exposed the human beings.
This study was done to determine the percentage of Salmonella spp. in camels from three provinces (Karbala, Al-Najaf and AL-Muthana) in Iraq with different age and both sexes. Total of 250 fecal samples from 250 camels were collected. Diagnostic study depended upon the morphological and cultural properties of the isolates on some selective media like Xylose lysine deoxycholate (XLD) and Salmonella Shigella (SS) agars which were used in addition to different biochemical tests and molecular assay by PCR for detection of virulence gene invasion A (invA) with Phylogenetic study. The clinical signs appearing on animals infected with Salmonella were greenish diarrhea, loss of appetite with mild systemic reaction. Bacteriological and molecular tests revealed isolation of five Salmonella isolates with invA gene. Two of these isolates were sequenced. The results showed that the first strain S. enterica subspecies typhimurium (LC730846) converged with a group of global strains with one node, as it converged with the global strain that held the clade (MK017934.1 and MT460418.1). While the second local strain S. enterica serovar enteritidis (LC730849) appeared with a new node and it is not affiliated with any association with the world S. enterica strains. It is concluded that the presence of Salmonella spp. in camel needs monitoring in order to minimize the risks of infection exposed the human beings.
Abstract
The microorganisms that inhabit the human gastrointestinal tract comprise a complex ecosystem with functions that significantly contribute to our systemic metabolism and have an impact on ...health and disease. In line with its importance, the human gastrointestinal microbiota has been extensively studied. Despite the fact that a significant part of the intestinal microorganisms has not yet been cultured, presently over 1000 different microbial species that can reside in the human gastrointestinal tract have been identified. This review provides a systematic overview and detailed references of the total of 1057 intestinal species of Eukarya (92), Archaea (8) and Bacteria (957), based on the phylogenetic framework of their small subunit ribosomal RNA gene sequences. Moreover, it unifies knowledge about the prevalence, abundance, stability, physiology, genetics and the association with human health of these gastrointestinal microorganisms, which is currently scattered over a vast amount of literature published in the last 150 years. This detailed physiological and genetic information is expected to be instrumental in advancing our knowledge of the gastrointestinal microbiota. Moreover, it opens avenues for future comparative and functional metagenomic and other high-throughput approaches that need a systematic and physiological basis to have an impact.
This historical review summarizes the over 1000 species associated with the human gastrointestinal tract – it provides a systematic presentation of all currently recognized gastrointestinal tract Bacteria, Archaea and Eukarya coupled with the reports of their prevalence, abundance, their stability in the ecosystem, physiology, genetics, and association with human health.
The early intestinal microbiota exerts important stimuli for immune development, and a reduced microbial exposure as well as caesarean section (CS) has been associated with the development of ...allergic disease. Here we address how microbiota development in infants is affected by mode of delivery, and relate differences in colonisation patterns to the maturation of a balanced Th1/Th2 immune response.
The postnatal intestinal colonisation pattern was investigated in 24 infants, born vaginally (15) or by CS (nine). The intestinal microbiota were characterised using pyrosequencing of 16S rRNA genes at 1 week and 1, 3, 6, 12 and 24 months after birth. Venous blood levels of Th1- and Th2-associated chemokines were measured at 6, 12 and 24 months.
Infants born through CS had lower total microbiota diversity during the first 2 years of life. CS delivered infants also had a lower abundance and diversity of the Bacteroidetes phylum and were less often colonised with the Bacteroidetes phylum. Infants born through CS had significantly lower levels of the Th1-associated chemokines CXCL10 and CXCL11 in blood.
CS was associated with a lower total microbial diversity, delayed colonisation of the Bacteroidetes phylum and reduced Th1 responses during the first 2 years of life.
With the gut microbiome being implicated in many diseases and even offering the potential for future therapies, its study has been gaining interest over the years. However, to gain more in-depth ...insight and understanding of the impact of the microbiome on our health, it is critical to have accurate microbial data that is reproducible and comparable across studies. Variation in experimental techniques, particularly DNA extraction and sequencing methods, may result in variability and the inability to compare results. In this chapter, we describe in detail our microbiome analysis methodology focusing on stool samples.
Gut dysbiosis is heavily involved in the development of various human diseases. There are thousands of publications per year for investigating the role of gut microbiota in diseases. However, ...emerging evidence has indicated the frequent data inconsistency between different studies, which is largely overlooked. There are many factors that can cause data variation and inconsistency during the process of microbiota study, in particular, sample storage conditions and sequencing process. Here, we systemically evaluated the impacts of six fecal sample storage conditions (three non-commercial storage protocols, -80°C, -80°C with 70% ethanol (ET_-80°C), 4°C with 70% ethanol (ET_4°C), and three commercial storage reagents, OMNIgeneGUT OMR-200 (GT) and MGIEasy (MGIE) at room temperature, and Longsee at 4°C (LS) on gut microbiome profile based on 16S rRNA gene sequencing. In addition, we also investigated the impacts of storage periods (1 and 2 weeks, or 6 months) and sequencing platform on microbiome profile. The efficacy of storage conditions was evaluated by DNA yield and quality, α and β diversity, relative abundance of the dominant and functional bacteria associated with short-chain fatty acid (SCFA) production, and BAs metabolism. Our current study suggested that -80°C was acceptable for fecal sample storage, and the addition of 70% ethanol had some benefits in maintaining the microbial community structure. Meanwhile, we found that samples in ET_4°C and GT reagents were comparable, both of them introduced some biases in α or β diversity, and the relative abundance of functional bacteria. Samples stored in MGIE reagent resulted in the least variation, whereas the most obvious variations were introduced by LS reagents. In addition, our results indicated that variations caused by storage condition were larger than that of storage time and sequencing platform. Collectively, our study provided a multi-dimensional evaluation on the impacts of storage conditions, storage time periods, and sequencing platform on gut microbial profile.
Escherichia coli is a common cause of acute diarrhea mainly in young children and, less frequently, in elderly or immunosuppressed patients. Many types of E. coli are part of the normal enteric ...flora, but can cause urinary tract or nervous system infections.
To study the prevalence of the main types and serogroups of diarrheagenic E. coli among hospitalized children with enteric infections.
Over a period of 5 years, 1,160 hospitalized children with acute diarrhea syndrome were studied. Fecal samples underwent culturing, biochemical and phenotypic identification.
Among the studied patients, 112/1,160 children (9.7%) had diarrhea caused by E. coli, and only 4 of the isolates were lactose-negative. The most common was diarrhea caused by ETEC – 65/112 (58.0%), followed by EPEC – 38/112 (33.9%), and in third place – EHES 9/112 (8.0%). We did not isolate EIEC types. Depending on the group of E. coli, we observed some differences in the clinical presentation and specifics in the distribution of patients by age.
The study shows that this causative agent is common among Bulgarian children with diarrhea. Unfortunately, in Bulgaria the microbiological network is still not able to adequately respond to the challenges of the extended serodiagnosis for detection of diarrheagenic E. coli, which is performed in Western Europe and North America.
The tigecycline resistance gene tet(X4) has been widely reported in animals and animal products in some Asian countries including China in recent years but only sporadically detected in human. In ...this study, we investigated the prevalence and genetic features of tet(X4)-positive clinical E. coli strains. A total of 462 fecal samples were collected from patients in four hospitals located in four provinces in China in 2023. Nine tet(X4)-positive E. coli strains were isolated and subjected to characterization of their genetic and phenotypic features by performing antimicrobial susceptibility test, whole-genome sequencing, bioinformatic and phylogenetic analysis. The majority of the test strains were found to exhibit resistance to multiple antimicrobial agents including tigecycline but remained susceptible to colistin and meropenem. A total of seven different sequence types (STs) and an unknown ST type were identified among the nine tet(X4)-positive strains. Notably, the tet(X4) gene in six out of these nine tet(X4)-positive E. coli strains was located in a IncFIA-HI1A-HI1B hybrid plasmid, which was an tet(X4)-bearing epidemic plasmid responsible for dissemination of the tet(X4) gene in China. Furthermore, the tet(X4) gene in four out of nine tet(X4)-positive E. coli isolates could be successfully transferred to E. coli EC600 through conjugation. In conclusion, this study characterized the epidemic tet(X4)-bearing plasmids and tet(X4)-associated genetic environment in clinical E. coli strains, suggested the importance of continuous surveillance of such tet(X4)-bearing plasmids to control the increasingly widespread dissemination of tigecycline-resistant pathogens in clinical settings in China.
BACKGROUNDColorectal cancer (CRC), the third most common cause of death in both males and females worldwide, shows a positive response to therapy and usually a better prognosis when detected at an ...early stage. However, the survival rate declines when the diagnosis is late and the tumor spreads to other organs. Currently, the measures widely used in the clinic are fecal occult blood test and evaluation of serum tumor markers, but the lack of sensitivity and specificity of these markers restricts their use for CRC diagnosis. Due to its high sensitivity and precision, colonoscopy is currently the gold-standard screening technique for CRC, but it is a costly and invasive procedure. Therefore, the implementation of custom-made methodologies including those with minimal invasiveness, protection, and reproducibility is highly desirable. With regard to other screening methods, the screening of fecal samples has several benefits, and metabolomics is a successful method to classify the metabolite shift in living systems as a reaction to pathophysiological influences, genetic modifications, and environmental factors. AIMTo characterize the variation groups and potentially recognize some diagnostic markers, we compared with healthy controls (HCs) the fecal nuclear magnetic resonance (NMR) metabolomic profiles of patients with CRC or adenomatous polyposis (AP). METHODSProton nuclear magnetic resonance spectroscopy was used in combination with multivariate and univariate statistical approaches, to define the fecal metabolic profiles of 32 CRC patients, 16 AP patients, and 38 HCs well matched in age, sex, and body mass index. RESULTSNMR metabolomic analyses revealed that fecal sample profiles differed among CRC patients, AP patients, and HCs, and some discriminatory metabolites including acetate, butyrate, propionate, 3-hydroxyphenylacetic acid, valine, tyrosine and leucine were identified. CONCLUSIONIn conclusion, we are confident that our data can be a forerunner for future studies on CRC management, especially the diagnosis and evaluation of the effectiveness of treatments.
Human gut microbiota can be studied through the characterization of microorganisms present in feces. Metaproteomics has arisen as a good approach to investigate this vast community. However, the ...processing of fecal samples in order to obtain the largest number of proteins from gut microbiota to be subsequently analyzed by means of metaproteomics is a challenge. Here we describe a protocol to approach this task. It includes two main steps: the first step of humectation and dispersion of the feces, followed by the separation of microorganisms from other fecal components such as roughage and food debris, and the second step in which microbial cells are broken up and microbiota proteins recovered for MS analysis. Detailed procedures for sample preparation, protein extraction, trypsin digestion, and mass spectrometry analysis for gut microbiota samples are provided.
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