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Analytical Chemistry Jul 2019Single cell RNA sequencing is a technology that provides the capability of analyzing the transcriptome of a single cell from a population. So far, single cell RNA...
Single cell RNA sequencing is a technology that provides the capability of analyzing the transcriptome of a single cell from a population. So far, single cell RNA sequencing has been focused mostly on human cells due to the larger starting amount of RNA template for subsequent amplification. One of the major challenges of applying single cell RNA sequencing to microbial cells is to amplify the femtograms of the RNA template to obtain sufficient material for downstream sequencing with minimal contamination. To achieve this goal, efforts have been focused on multiround RNA amplification, but would introduce additional contamination and bias. In this work, we for the first time coupled a microfluidic platform with multiple displacement amplification technology to perform single cell whole transcriptome amplification and sequencing of Porphyromonas somerae, a microbe of interest in endometrial cancer, as a proof-of-concept demonstration of using single cell RNA sequencing tool to unveil gene expression heterogeneity in single microbial cells. Our results show that the bacterial single-cell gene expression regulation is distinct across different cells, supporting widespread heterogeneity.
Topics: Equipment Design; Gene Expression Profiling; Gene Expression Regulation, Bacterial; Microfluidic Analytical Techniques; Nucleic Acid Amplification Techniques; Porphyromonas; Single-Cell Analysis; Transcriptome
PubMed: 31188565
DOI: 10.1021/acs.analchem.8b04773 -
Journal of Clinical Microbiology Mar 2015A yet-undescribed bacterial species, tentatively named "Porphyromonas katsikii," was isolated from individuals of a small goat herd with pyogranulomatous pneumonia...
A yet-undescribed bacterial species, tentatively named "Porphyromonas katsikii," was isolated from individuals of a small goat herd with pyogranulomatous pneumonia during an outbreak of acute respiratory disease. The isolated bacteria grew in the form of black-pigmented colonies after 14 days of incubation under anaerobic conditions at 37°C on a tryptic soy blood agar medium. The bacteria were identified as a yet-undescribed Porphyromonas species by determination of the nucleotide sequence of the rrs 16S rRNA gene, and this species was tentatively named Porphyromonas katsikii. PCR amplification with specific primers for this yet-undescribed species revealed the presence of P. katsikii in the lung tissue of all affected animals, while no PCR signals were evidenced from the lungs of healthy goats or from goats with pasteurellosis caused by Mannheimia haemolytica. These data indicate P. katsikii as the causative agent of acute respiratory distress. P. katsikii is phylogenetically related to Porphyromonas somerae and Porphyromonas levii, which cause pathologies in humans and animals, respectively. P. katsikii was not detected by PCR from samples of the gingival pockets or of the faces of healthy goats.
Topics: Anaerobiosis; Animals; Bacteriological Techniques; Cluster Analysis; DNA, Bacterial; DNA, Ribosomal; Disease Outbreaks; Goat Diseases; Goats; Lung; Molecular Sequence Data; Phylogeny; Pigments, Biological; Pneumonia, Bacterial; Porphyromonas; RNA, Ribosomal, 16S; Sequence Analysis, DNA; Temperature
PubMed: 25540395
DOI: 10.1128/JCM.02682-14 -
Journal of Clinical Microbiology Sep 2005Porphyromonas levii is an anaerobic, pigmented gram-negative bacillus originally isolated from bovine rumen. We describe 58 human clinical strains of P. levii-like...
Porphyromonas levii is an anaerobic, pigmented gram-negative bacillus originally isolated from bovine rumen. We describe 58 human clinical strains of P. levii-like organisms, isolated from various human clinical specimens that are phenotypically similar to the type strain of P. levii, a rumen isolate (ATCC 29147). Our biochemical, comparative 16S rRNA sequence analyses, and DNAlpha-DNA relatedness studies indicate that the human P. levii-like organisms are similar to each other but genetically different from the P. levii type strain isolated from bovine rumen. We therefore propose the name Porphyromonas somerae to encompass the human P. levii-like organisms. P. somerae was predominantly isolated from patients with chronic skin and soft tissue or bone infections, especially in the lower extremities.
Topics: Animals; Bacterial Typing Techniques; Bacteroidaceae Infections; DNA, Bacterial; DNA, Ribosomal; Humans; Molecular Sequence Data; Nucleic Acid Hybridization; Phenotype; Porphyromonas; RNA, Ribosomal, 16S; Sequence Analysis, DNA
PubMed: 16145091
DOI: 10.1128/JCM.43.9.4455-4459.2005