With the introduction of synthetic antibiotics, many lives including humans and animals have been saved against bacterial infection. An increasing level of antibiotics use, however, raises serious problems of multi-drug resistance and transferring of resistance genes across different environments and countries. Advances in high-throughput sequencing technology and efficient bioinformatics methods allow us to perform a large-scale screening and analysis of resistomes in the human and environmental microbiomes. Recent studies on human microbiomes have revealed a diverse distribution of resistance genes and their transferring activities in the communities. This review discusses recent progresses in metagenomic approaches to identify resistance genes in the human microbiome, including genomic sequence search and functional metagenomics methods. Using Rifampicin ADP-ribosyltransferase as an example, an integrative approach that analyzes the sequences and three-dimensional structures of the proteins derived from resistance genes is also introduced.
ADP Ribose Transferases ; Animals ; Anti-Bacterial Agents ; Bacterial Infections ; Computational Biology ; Drug Resistance, Multiple ; Humans* ; Mass Screening ; Metagenome ; Metagenomics* ; Microbiota ; Rifampin

No abstract available.
Humans* ; Microbiota*

Evidence indicates that Helicobacter pylori is the causative agent of chronic gastritis and perhaps gastric malignancy. Extracellular vesicles (EVs) play an important role in the evolutional process of malignancy due to their genetic material cargo. We aimed to evaluate the clinical significance and biological mechanism of H. pylori EVs on the pathogenesis of gastric malignancy. We performed 16S rDNA-based metagenomic analysis of gastric juices either from endoscopic or surgical patients. From each sample of gastric juices, the bacteria and EVs were isolated. We evaluated the role of H. pylori EVs on the development of gastric inflammation in vitro and in vivo. IVIS spectrum and confocal microscopy were used to examine the distribution of EVs. The metagenomic analyses of the bacteria and EVs showed that Helicobacter and Streptococcus are the two major bacterial genera, and they were significantly increased in abundance in gastric cancer (GC) patients. H. pylori EVs are spherical and contain CagA and VacA. They can induce the production of tumor necrosis factor-α, interleukin (IL)-6 and IL-1β by macrophages, and IL-8 by gastric epithelial cells. Also, EVs induce the expression of interferon gamma, IL-17 and EV-specific immunoglobulin Gs in vivo in mice. EVs were shown to infiltrate and remain in the mouse stomach for an extended time. H. pylori EVs, which are abundant in the gastric juices of GC patients, can induce inflammation and possibly cancer in the stomach, mainly via the production of inflammatory mediators from gastric epithelial cells after selective uptake by the cells.
Adenocarcinoma* ; Animals ; Bacteria ; Epithelial Cells* ; Extracellular Vesicles* ; Gastric Juice* ; Gastritis ; Helicobacter pylori ; Helicobacter* ; Humans ; Immunoglobulin G ; In Vitro Techniques ; Inflammation* ; Interferons ; Interleukin-17 ; Interleukin-8 ; Interleukins ; Macrophages ; Metagenomics ; Mice ; Microscopy, Confocal ; Necrosis ; Stomach ; Stomach Neoplasms ; Streptococcus

Recent evidence has indicated that bacteria-derived extracellular vesicles (EVs) are important for host–microbe communication. The aims of the present study were to evaluate whether bacteria-derived EVs are excreted via the urinary tract and to compare the composition of bacteria-derived EVs in the urine of pregnant and non-pregnant women. Seventy-three non-pregnant and seventy-four pregnant women were enrolled from Dankook University and Ewha Womans University hospitals. DNA was extracted from urine EVs after EV isolation using the differential centrifugation method. 16S ribosomal RNA (16S rRNA) gene sequencing was performed using high-throughput 454 pyrosequencing after amplification of the V1–V3 region of the 16S rDNA. The composition of 13 taxa differed significantly between the pregnant and non-pregnant women. At the genus level, Bacillus spp. EVs were more significantly enriched in the urine of the pregnant women than in that of the non-pregnant women (45.61% vs 0.12%, respectively). However, Pseudomonas spp. EVs were more dominant in non-pregnant women than in pregnant women (13.2% vs 4.09%, respectively). Regarding the compositional difference between pregnant women with normal and preterm delivery, EVs derived from Ureaplasma spp. and the family Veillonellaceae (including Megasphaera spp.) were more abundant in the urine of preterm-delivered women than in that of women with normal deliveries. Taken together, these data showed that Bacillus spp. EVs predominate in the urine of pregnant women, whereas Pseudomonas spp. EVs predominate in the urine of non-pregnant women; this suggests that Bacillus spp. EVs might have an important role in the maintenance of pregnancy.
Bacillus ; Centrifugation ; DNA ; DNA, Ribosomal ; Extracellular Vesicles* ; Female ; Hospitals, University ; Humans ; Megasphaera ; Metagenomics* ; Methods ; Pregnancy ; Pregnant Women ; Pseudomonas ; RNA, Ribosomal, 16S ; Ureaplasma ; Urinary Tract ; Veillonellaceae

PURPOSE: Atopic dermatitis (AD) is an inflammatory skin disease, significantly affecting the quality of life. Using AD as a model system, we tested a successive identification of AD-associated microbes, followed by a culture-independent serum detection of the identified microbe. METHODS: A total of 43 genomic DNA preparations from washing fluid of the cubital fossa of 6 healthy controls, skin lesions of 27 AD patients, 10 of which later received treatment (post-treatment), were subjected to high-throughput pyrosequencing on a Roche 454 GS-FLX platform. RESULTS: Microbial diversity was decreased in AD, and was restored following treatment. AD was characterized by the domination of Staphylococcus, Pseudomonas, and Streptococcus, whereas Alcaligenaceae (f), Sediminibacterium, and Lactococcus were characteristic of healthy skin. An enzyme-linked immunosorbent assay (ELISA) showed that serum could be used as a source for the detection of Staphylococcus aureus extracellular vesicles (EVs). S. aureus EV-specific immunoglobulin G (IgG) and immunoglobulin E (IgE) were quantified in the serum. CONCLUSIONS: A metagenomic analysis together with a serum detection of pathogen-specific EVs provides a model for successive identification and diagnosis of pathogens of AD.
Alcaligenaceae ; Dermatitis, Atopic* ; Diagnosis ; DNA ; Enzyme-Linked Immunosorbent Assay ; Extracellular Vesicles ; Humans ; Immunoglobulin E ; Immunoglobulin G ; Immunoglobulins ; Lactococcus ; Metagenomics* ; Pseudomonas ; Quality of Life ; Skin ; Skin Diseases ; Staphylococcus ; Staphylococcus aureus ; Streptococcus

Background: Cholecystitis is an important risk factor for gallbladder cancer, but the bile microbiome and its association with gallbladder disease has not been investigated fully.We aimed to analyze the bile microbiome in normal conditions, chronic cholecystitis, and gallbladder cancer, and to identify candidate bacteria that play an important role in gallbladder carcinogenesis. Methods: We performed metagenome sequencing on bile samples of 10 healthy individuals, 10 patients with chronic cholecystitis, and 5 patients with gallbladder cancer, and compared the clinical, radiological, and pathological characteristics of the participants. Results: No significant bacterial signal was identified in the normal bile. The predominant dysbiotic bacteria in both chronic cholecystitis and gallbladder cancer were those belonging to the Enterobacteriaceae family. Klebsiella increased significantly in the order of normal, chronic cholecystitis, and gallbladder cancer. Patients with chronic cholecystitis and dysbiotic microbiome patterns had larger gallstones and showed marked epithelial atypia, which are considered as precancerous conditions. Conclusion We investigated the bile microbiome in normal, chronic cholecystitis, and gallbladder cancer. We suggest possible roles of Enterobacteriaceae, including Klebsiella, in gallbladder carcinogenesis. Our findings reveal a possible link between a dysbiotic bile microbiome and the development of chronic calculous cholecystitis and gallbladder cancer.

PURPOSE: The microbial environment is an important factor that contributes to the pathogenesis of atopic dermatitis (AD). Recently, it was revealed that not only bacteria itself but also extracellular vesicles (EVs) secreted from bacteria affect the allergic inflammation process. However, almost all research carried out so far was related to local microorganisms, not the systemic microbial distribution. We aimed to compare the bacterial EV composition between AD patients and healthy subjects and to experimentally find out the beneficial effect of some bacterial EV composition METHODS: Twenty-seven AD patients and 6 healthy control subjects were enrolled. After urine and serum were obtained, EVs were prepared from samples. Metagenomic analysis of 16s ribosomal DNA extracted from the EVs was performed, and bacteria showing the greatest difference between controls and patients were identified. In vitro and in vivo therapeutic effects of significant bacterial EV were evaluated with keratinocytes and with Staphylococcus aureus-induced mouse AD models, respectively. RESULTS: The proportions of Lactococcus, Leuconostoc and Lactobacillus EVs were significantly higher and those of Alicyclobacillus and Propionibacterium were lower in the control group than in the AD patient group. Therefore, lactic acid bacteria were considered to be important ones that contribute to the difference between the patient and control groups. In vitro, interleukin (IL)-6 from keratinocytes and macrophages decreased and cell viability was restored with Lactobacillus plantarum-derived EV treatment prior to S. aureus EV treatment. In S. aureus-induced mouse AD models, L. plantarum-derived EV administration reduced epidermal thickening and the IL-4 level. CONCLUSIONS: We suggested the protective role of lactic acid bacteria in AD based on metagenomic analysis. Experimental findings further suggest that L. plantarum-derived EV could help prevent skin inflammation.
Alicyclobacillus ; Animals ; Bacteria ; Cell Survival ; Dermatitis, Atopic* ; DNA, Ribosomal ; Extracellular Vesicles* ; Healthy Volunteers ; Humans ; In Vitro Techniques ; Inflammation ; Interleukin-4 ; Interleukins ; Keratinocytes ; Lactic Acid ; Lactobacillus* ; Lactococcus ; Leuconostoc ; Macrophages ; Metagenomics ; Mice ; Microbiota ; Probiotics ; Propionibacterium ; Skin ; Staphylococcus* ; Therapeutic Uses

Background: Cholecystitis is an important risk factor for gallbladder cancer, but the bile microbiome and its association with gallbladder disease has not been investigated fully.We aimed to analyze the bile microbiome in normal conditions, chronic cholecystitis, and gallbladder cancer, and to identify candidate bacteria that play an important role in gallbladder carcinogenesis. Methods: We performed metagenome sequencing on bile samples of 10 healthy individuals, 10 patients with chronic cholecystitis, and 5 patients with gallbladder cancer, and compared the clinical, radiological, and pathological characteristics of the participants. Results: No significant bacterial signal was identified in the normal bile. The predominant dysbiotic bacteria in both chronic cholecystitis and gallbladder cancer were those belonging to the Enterobacteriaceae family. Klebsiella increased significantly in the order of normal, chronic cholecystitis, and gallbladder cancer. Patients with chronic cholecystitis and dysbiotic microbiome patterns had larger gallstones and showed marked epithelial atypia, which are considered as precancerous conditions. Conclusion We investigated the bile microbiome in normal, chronic cholecystitis, and gallbladder cancer. We suggest possible roles of Enterobacteriaceae, including Klebsiella, in gallbladder carcinogenesis. Our findings reveal a possible link between a dysbiotic bile microbiome and the development of chronic calculous cholecystitis and gallbladder cancer.