[Show abstract][Hide abstract] ABSTRACT: We report the draft genome sequence of the Stenotrophomonas maltophilia strain EPM1, found in association with a culture of Giardia duodenalis. The draft genome sequence of S. maltophilia strain EPM1, obtained with Roche 454 GS-FLX Titanium technology, is composed of 19 contigs totaling 4,785,869 bp, with a G+C content of 66.37%.
[Show abstract][Hide abstract] ABSTRACT: PCR assays have been developed mainly to assist investigations into the epidemiology of Giardia duodenalis, the only species in the Giardia genus having zoonotic potential. However, a reliable identification of all species is of practical importance, particularly when water samples and samples from wild animals are investigated. The aim of the present work was to genotype Giardia species and G. duodenalis assemblages using as a target the region spanning the 5.8S gene and the 2 flanking internal transcribed spacers (ITS1 and ITS2) of the ribosomal gene. Primers were designed to match strongly conserved regions in the 3' end of the small subunit and in the 5' end of the large subunit ribosomal genes. The corresponding region (about 310 bp) was amplified from 49 isolates of both human and animal origin, representing all G. duodenalis assemblages as well as G. muris and G. microti. Sequence comparison and phylogenetic analysis showed that G. ardeae, G. muris, G. microti as well as the 7 G. duodenalis assemblages can be easily distinguished. Since the major subgroups within the zoonotic assemblages A and B can be identified by sequence analysis, this assay is also informative for molecular epidemiological studies.
[Show abstract][Hide abstract] ABSTRACT: Of the seven genetic groups, or assemblages, currently recognized in the Giardia duodenalis species complex, only assemblages A and B are associated with human infection, but they also infect other mammals. Recent investigations have suggested the occurrence of genetic exchanges among isolates of G. duodenalis, and the application of assemblage-specific PCR has shown both assemblages A and B in a significant number of human infections. In this work, three real-time quantitative (qPCR) assays were developed to target the G. duodenalis triose phosphate isomerase, glutamate dehydrogenase, and open reading frame C4 sequences. Primers were designed to allow the specific amplification of the DNA of assemblage A or B and to generate products distinguishable by their melting curves or, after qPCR, by their sequences, sizes, or restriction patterns. The assays showed full specificity and detected DNA from a single trophozoite (4 to 8 target copies). We applied these assays, as well as a TaqMan assay that targets the beta-giardin gene, to genomic DNA extracted from 30 human stools and to Giardia cysts purified by immunomagnetic capture from the same samples. Simultaneous detection of both assemblages was observed in a large number of DNAs extracted from stools, and experiments on the cysts purified from the same samples showed that this was essentially attributable to mixed infections, as only one assemblage was detected when dilutions of cysts were tested. In a few cases, detection of both assemblages was observed even when single cysts were tested. This result, which suggests the presence of recombinants, needs to be confirmed using more accurate methods for cyst separation and enumeration. The assays described in this study can be used to detect Giardia cysts infectious to humans in samples from animals and in water and food.