China
Africa
Explore chapters and articles related to this topic
Dientamoeba fragilis Infection
Published in Dongyou Liu, Handbook of Foodborne Diseases, 2018
Candela Menéndez Fernández-Miranda, Jonathan Fernández Suarez, Noelia Moran Suarez, Javier Fernández Domínguez, María Martínez Sela, Mercedes Rodríguez Pérez, Azucena Rodríguez-Guardado
Real-time PCR is considered as the diagnostic method of choice by most authors.8,27,28,52–54,58,61 Several papers27,28,101,102 have compared PCR with other diagnostic methods such as culture or conventional microscopy, finding that real-time PCR based on the small-subunit ribosomal RNA gene of D. fragilis demonstrates 100% sensitivity and specificity versus 40% and 100%, respectively, by culture and 34.3% and 99%, respectively, by microscopy.
Malabsorption and microsporidia
Published in Ronald R. Watson, NUTRIENTS and FOODS in AIDS, 2017
Five genera of microsporidia are known to cause disease in humans, and include Enterocytozoon bieneusi and Septata intestinalis, which are intestinal pathogens, and Encephalitozoon spp., Nosema spp., and Pleistophora sp. which are rare and nonintestinal and may cause keratitis, myositis, encephalitis, sinusitis, bronchitis, pneumonitis, hepatitis, and nephritis.5–8 Recently, genetic studies of the S. intestinalis rrs gene encoding small subunit ribosomal RNA (srRNA)9 indicates that this protozoan is more closely related to Encephalitozoon and should be reclassified Encephalitozoon intestinalis.
Molecular Mycology and Emerging Fungal Pathogens
Published in Johan A. Maertens, Kieren A. Marr, Diagnosis of Fungal Infections, 2007
The small subunit ribosomal RNA gene, or 18S rDNA in fungi, is one useful molecular chronometer for inferring evolutionary relationships between organisms, though additional targets such as the large subunit rRNA, cytochrome b, beta-tubulin, chitin synthase, and heat shock protein genes have been used. PCR amplification of rDNA from fungi is facilitated by the presence of highly conserved sequences in the 18S rRNA gene. PCR primers targeting these conserved sites can be used to amplify segments of the gene containing more variable and, hence, phylogenetically informative sequences. When these PCR products are sequenced and aligned with known 18S rDNA sequences, phylogenetic trees depicting inferred evolutionary relationships can be generated. One problem with use of a single gene for phylogenetic analysis is that the output reflects the phylogeny of the gene, which may differ from the phylogeny of the organism in the unusual scenario of gene transfer between organisms. The taxonomic position of an organism is bolstered when analysis of several genes in the organism suggests the same phylogeny (4).
Validation and maintaining laboratory developed molecular tests compliant with ISO15189 for diagnosis of intestinal parasitic infections
Published in Expert Review of Molecular Diagnostics, 2022
In the in silico sensitivity the theoretically ability of the assay to detect genetic variants of the target organism is evaluated. In other words it provides insight in the genetic variation within the primer and probe sequences used in the assay. Target sequences are obtained through browsing the GenBank for sequences of the target organism and by BLAST search of the primers and probes. In Figure 1, an alignment of the small subunit ribosomal RNA gene of different Giardia intestinalis genotypes (assemblages) with the primers and probe used in the real-time PCR [8] is given as an example. A 100% homology within all assemblages is shown within the reverse primer and probe sequences. The mismatch shown in the forward primer sequence in Giardia assemblage E (goat, sheep, cow) is expected not to cause loss of sensitivity because the mismatch is seven positions away from the 3ʹ end of the primer sequence which is the starting point for the Taq polymerase. In this example it can be concluded that the in silico sensitivity analysis fulfills the predetermined criterion.
The ‘in vivo lifestyle’ of bile acid 7α-dehydroxylating bacteria: comparative genomics, metatranscriptomic, and bile acid metabolomics analysis of a defined microbial community in gnotobiotic mice
Published in Gut Microbes, 2020
Jason M. Ridlon, Saravanan Devendran, João Mp Alves, Heidi Doden, Patricia G. Wolf, Gabriel V. Pereira, Lindsey Ly, Alyssa Volland, Hajime Takei, Hiroshi Nittono, Tsuyoshi Murai, Takao Kurosawa, George E. Chlipala, Stefan J. Green, Alvaro G. Hernandez, Christopher J. Fields, Christy L. Wright, Genta Kakiyama, Isaac Cann, Purna Kashyap, Vance McCracken, H. Rex Gaskins
Genomic DNA was extracted from cecum samples using a Maxwell® 16 Tissue DNA Purification Kit, with extractions conducted on a Maxwell16 automated extraction system (Promega, Madison, WI). Genomic DNA was PCR amplified with primers 515F-modified and 926R30 targeting the V4-V5 variable region of the microbial small subunit ribosomal RNA gene using a two-stage “targeted amplicon sequencing (TAS)” protocol.31,32 The primers contained 5ʹ common sequence tags (known as common sequence 1 and 2, CS1 and CS2) as described previously.33 The CS1_515F and CS2_926R primer sequences were 5ʹ-ACACTGACGACATGGTTCTACAGTGYCAGCMGCCGCGGTAA-3ʹ and 5ʹ-TACGGTAGCAGAGACTTGGTCTCCGYCAATTYMTTTRAGTTT-3ʹ, respectively, with the underlined regions indicating the common sequence tags. First stage PCR amplifications were performed in 10 µl reactions in 96-well plates, using the MyTaq HS 2X mastermix (Bioline, Taunton, MA). PCR conditions were 95°C for 5 min, followed by 28 cycles of 95°C for 30 sec, 50°C for 60 sec and 72°C for 90 sec.
Metabarcoding and metabolomics offer complementarity in deciphering marine eukaryotic biofouling community shifts
Published in Biofouling, 2018
Jean-François Briand, Xavier Pochon, Susanna A. Wood, Christine Bressy, Cédric Garnier, Karine Réhel, Félix Urvois, Gérald Culioli, Anastasija Zaiko
After immersion for month, 96 panels were scraped and the material isolated for DNA extraction, which was performed on all samples using a Power Soil DNA Isolation kit (Qiagen, Courtaboeuf, France), following the manufacturer’s instructions (Camps et al. 2014). Triplicate DNA extracts were pooled and a two-step tailed PCR amplicon procedure (Pramanik and Li 2002) employed for generating an Illumina MiSeqTM library for the 32 samples. The universal primers Uni18SF (5’-AGGGCAAKYCTGGTGCCAGC-3’) and Uni18SR (5’-GRCGGTATCTRATCGYCTT-3’) (Zhan et al. 2013) were used to amplify the eukaryotic V4 region of the nuclear small subunit ribosomal RNA (18S rRNA) gene. The primers for the first PCR were modified to include IlluminaTM overhang adaptors as described in (Kozich et al. 2013). PCR amplifications were undertaken on an Eppendorf Mastercycler (Eppendorf, Germany) in a total volume of 30 μl using AmpliTaq Gold® 360 PCR Master Mix (Life Technologies, USA), 1 μl of each primer (10 μM, IDT DNA, USA) and 1 μl of template DNA. Reaction cycling conditions were: 94 °C for 3 min, followed by 25 cycles of 94 °C for 45s, 55 °C for 45s, 72 °C for 2 min, and a final extension of 72 °C for 5 min.