1887

Abstract

A nested PCR assay (TPILC-PCR) was developed to detect and distinguish between assemblages A and B from human faeces by analysis of the triose phosphate isomerase gene (). The assay comprised an initial multiplexed block-based amplification. This was followed by two separate real-time PCR assays specific for assemblages A and B using a LightCycler and SYBR Green I to identify PCR products by melting-point analysis. RFLP analysis was applied to distinguish assemblage A groups I and II. The real-time nested PCR was evaluated using DNA extracted from purified giardial trophozoites, oocysts, whole faeces containing a range of potential pathogens (including ), faecal smears and bacterial suspensions. The assay was specific, sensitive, reproducible and rapid.

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2003-08-01
2024-03-28
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References

  1. Adam R. D. 2000; The Giardia lamblia genome. Int J Parasitol 30:475–484 [CrossRef]
    [Google Scholar]
  2. Amar C, Pedraza-Diaz S, McLauchlin J. 2001; Extraction and genotyping of Cryptosporidium parvum DNA from fecal smears on glass slides stained conventionally for direct microscope examination. J Clin Microbiol 39:401–403 [CrossRef]
    [Google Scholar]
  3. Amar C. F, Dear P. H, Pedraza-Diaz S, Looker N, Linnane E, McLauchlin J. 2002; Sensitive PCR-restriction fragment length polymorphism assay for detection and genotyping of Giardia duodenalis in human feces. J Clin Microbiol 40:446–452 [CrossRef]
    [Google Scholar]
  4. Farthing M. J. G. 1995; Giardia lamblia . In Infections of the Gastrointestinal Tract . pp 1081–1106 Edited by Blaser M. J., Smith P. D., Ravdin J. I., Greenberg H. B., Guerrant R. L. New York: Raven Press;
  5. Hall T. A. 1999; BioEdit: a friendly biological sequence alignment editor and analysis program for Window 95/98/NT. Nucleic Acids Symp Ser 41:95–98
    [Google Scholar]
  6. Homan W. L, Gilsing M, Bentala H, Limper L, van Knapen F. 1998; Characterization of Giardia duodenalis by polymerase-chain-reaction fingerprinting. Parasitol Res 84:707–714 [CrossRef]
    [Google Scholar]
  7. Isaac-Renton J. L. 1991; Laboratory diagnosis of giardiasis. Clin Lab Med 11:811–827
    [Google Scholar]
  8. LeChevallier M. W, Norton W. D, Siegel J. E, Abbaszadegan M. 1995; Evaluation of the immunofluorescence procedure for detection of Giardia cysts and Cryptosporidium oocysts in water. Appl Environ Microbiol 61:690–697
    [Google Scholar]
  9. McLauchlin J, Pedraza-Díaz S, Amar-Hoetzeneder C, Nichols G. L. 1999; Genetic characterization of Cryptosporidium strains from 218 patients with diarrhea diagnosed as having sporadic cryptosporidiosis. J Clin Microbiol 37:3153–3158
    [Google Scholar]
  10. McLauchlin J, Amar C, Pedraza-Díaz S, Nichols G. L. 2000; Molecular epidemiological analysis of Cryptosporidium spp.in the United Kingdom: results of genotyping Cryptosporidium spp. in 1,705 fecal samples from humans and 105 fecal samples from livestock animals. J Clin Microbiol 38:3984–3990
    [Google Scholar]
  11. Patel S, Pedraza-Díaz S, McLauchlin J. 1999; The identification of Cryptosporidium species and Cryptosporidium parvum directly from whole faeces by analysis of a multiplex PCR of the 18S rRNA gene and by PCR/RFLP of the Cryptosporidium outer wall protein (COWP) gene. Int J Parasitol 29:1241–1247 [CrossRef]
    [Google Scholar]
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