BACKGROUND: MicroRNAs (miRNAs), small RNA molecules of approximately 22 nucleotides, have been shown to be up-or downregulated in specific cell types and disease states. These molecules have become recognized as one of the major regulatory gatekeepers of coding genes in the human genome.CONTENT: We review the structure, nomenclature, mechanism of action, technologies used for miRNA detection, and associations of miRNAs with human cancer. miRNAs are produced in a tissue-specific manner, and changes in miRNA within a tissue type can be correlated with disease status. miRNAs appear to regulate mRNA translation and degradation via mechanisms that are dependent on the degree of complementarity between the miRNA and mRNA molecules. miRNAs can be detected via several methods, such as microarrays, bead-based arrays, and quantitative real-time PCR. The tissue concentrations of specific miRNAs have been associated with tumor invasiveness, metastatic potential, and other clinical characteristics for several types of cancers, including chronic lymphocytic leukemia, and breast, colorectal, hepatic, lung, pancreatic, and prostate cancers.

Epitheliocystis has been associated with heavy mortality and reduced growth of survivors in farmed Atlantic salmon (Salmo salar) (22). Ultrastructural studies of the epitheliocystis agent found in Atlantic salmon have revealed it to be an intracellular gram-negative coccoid bacterium with distinct developmental stages typical of bacteria of the order Chlamydiales (22). Epitheliocystis has been described in other salmonid hosts, e.g., juvenile steelhead trout (Oncorhynchus mykiss) (28) and cultured lake trout (Salvelinus namaycush) (3), as well as in a number of nonsalmonid species, including bluegill (Lepomis macrochirus) (16), striped bass (Morone saxatilis) (32), white perch (Morone americanus) (32), sea bream (Sparus aurata) (25), grey mullet (Liza ramada) (25), and cultured white sturgeon (Acipenser transmontanus) (14). Morphological studies of epitheliocystis agents in sea bream (S. aurata) have provided evidence for two distinct chlamydia-like developmental cycles associated with proliferative and nonproliferative host reactions (5). Although transmission electron microscopic examinations of intracellular inclusions have demonstrated that the agents of epitheliocystis in both salmonid and nonsalmonid hosts are gram-negative bacteria with developmental stages typical of members of the order Chlamydiales (5,14,22,28,32), the genetic relatedness of these bacteria has yet to be determined.Sequence data from the rRNA operon have revised phylogenetic relationships between Chlamydia species and chlamydia-like bacteria (CLB) (7,8,9). Reclassifying chlamydial species on the basis of 16S rRNA gene sequence identity, 16S and 23S ribosomal DNA (rDNA) sequences, and phenotypic characterization is considered by some to be the best means of taxonomically categorizing chlamydiae (24). Based on this approach, species within the family Chlamydiaceae have 16S rRNA gene sequences that are Ͼ90% identical (26), whereas chlamydia-like bacteria, defined as obligate intracellular bacteria having reticulate (RBs) and elementary bodies (EBs) characteristic of chlamydia, have been shown to have Ͼ80% 16S rRNA

BACKGROUND MicroRNAs (miRNAs) are RNA molecules that are involved in the regulation of many cellular processes, including those related to human cancers. The aim of this study was to determine, as a proof of principle, whether specific candidate miRNAs could be detected in fine-needle aspirate (FNA) biopsies of pancreatic ductal adenocarcinoma (PDAC) and could accurately differentiate malignant from benign pancreatic tissues. METHODS We used TaqMan® assays to quantify miRNA levels in FNA samples collected in RNARetain (n = 16) and compared the results with a training set consisting of frozen macrodissected pancreatic samples (n = 20). RESULTS Quantitative reverse-transcription PCR analysis confirmed that miRNA levels are affected in PDAC FNAs and correlate well with the changes observed in the training set of frozen pancreatic samples. Analysis of the amounts produced for a few specific miRNAs enabled identification of PDAC samples. The combination of miR-196a and miR-217 biomarkers further improved the ability to distinguish between healthy tissue, PDAC, and chronic pancreatitis in the training set (P = 8.2 × 10−10), as well as segregate PDAC FNA samples from other FNA samples (P = 1.1 × 10−5). Furthermore, we showed that miR-196a production is likely specific to PDAC cells and that its incidence paralleled the progression of PDAC. CONCLUSIONS To the best of our knowledge, this study is the first to evaluate the diagnostic potential of miRNAs in a clinical setting and has shown that miRNA analysis of pancreatic FNA biopsy samples can aid in the pathologic evaluation of suspicious cases and may provide a new strategy for improving the diagnosis of pancreatic diseases.

Metagenomic sequencing for infectious disease diagnostics is an important tool that holds promise for use in the clinical laboratory. Challenges for implementation so far include high cost, the length of time to results, and the need for technical and bioinformatics expertise. However, the recent technological innovation of nanopore sequencing from Oxford Nanopore Technologies (ONT) has the potential to address these challenges. ONT sequencing is an attractive platform for clinical laboratories to adopt due to its low cost, rapid turnaround time, and user-friendly bioinformatics pipelines. However, this method still faces the problem of base-calling accuracy compared to other platforms. This review highlights the general challenges of pathogen detection in clinical specimens by metagenomic sequencing, the advantages and disadvantages of the ONT platform, and how research to date supports the potential future use of nanopore sequencing in infectious disease diagnostics.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) has the poorest overall prognosis among gastrointestinal cancers; however, curative resection in early-stage PDAC greatly improves survival rates, indicating the importance of early detection. Because abnormal microRNA production is commonly detected in cancer, we investigated noninvasive precursor pancreatic intraepithelial neoplasia (PanIN) lesions for microRNA production as a potential early biomarker of PDAC.