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Chapter 15 : Detection of spp. and Specific DNA in Respiratory Secretions by PCR-Enzyme-Linked Immunosorbent Assay and Comparison with Conventional Methods

Authors: Diane S. J. Lindsay1, William H. Abraham2, Alistair W. Brown3, Giles F. S. Edwards4
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Affiliations: 1: Scottish Reference Laboratory, Stobhill Hospital, Glasgow, UK G21 3UW; 2: Scottish Reference Laboratory, Stobhill Hospital, Glasgow, UK G21 3UW; 3: Scottish Reference Laboratory, Stobhill Hospital, Glasgow, UK G21 3UW; 4: Scottish Reference Laboratory, Stobhill Hospital, Glasgow, UK G21 3UW
Content Type: Monograph
Publication Year: 2006

Category: Bacterial Pathogenesis

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Detection of spp. and Specific DNA in Respiratory Secretions by PCR-Enzyme-Linked Immunosorbent Assay and Comparison with Conventional Methods, Page 1 of 2

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Abstract:

This chapter talks about a study in which a PCR-enzyme-linked immunosorbent assay (ELISA) was devised for the detection of and –specific DNA in human respiratory secretions. The PCR-ELISA incorporates a solid-phase probe hybridization step to verify the PCR reaction. Briefly, DNA was extracted from a total of 343 routine respiratory secretions using Nucleospin tissue columns. The PCR-ELISA was performed using reagents from the Roche Diagnostic PCR-ELISA (Dig Detection) kit Cat No. 1636111 following the manufacturers’ instructions. A total of 343 respiratory samples were tested, of which 21 patients were identified as cases of legionellosis by fulfilling one or more of the criteria for a definitive case. With PCR-based assays becoming more sensitive and specific, perhaps the distinction of a definitive case should incorporate more molecular techniques. The 16S RNA PCR-ELISA has the same sensitivity as the 5S RNA PCR/SB technique but can also identify and is more sensitive than either direct fluorescent antibody (DFA) or culture. The lack of suitable samples, i.e., respiratory secretions, should not inhibit investigation, as specific DNA has been found in serum and urine.

Citation: Lindsay D, Abraham W, Brown A, Edwards G. 2006. Detection of spp. and Specific DNA in Respiratory Secretions by PCR-Enzyme-Linked Immunosorbent Assay and Comparison with Conventional Methods, p 55-57. In Cianciotto N, Kwaik Y, Edelstein P, Fields B, Geary D, Harrison T, Joseph C, Ratcliff R, Stout J, Swanson M (ed), . ASM Press, Washington, DC. doi: 10.1128/9781555815660.ch15

Key Concept Ranking

Legionella pneumophila
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Molecular Techniques
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References

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1. Fallon, R. J. 1981. Laboratory diagnosis of Legionnaires’ disease. Assoc. Clin. Pathol. 99: 1– 15.
2. Helbig, J. H.,, S. A. Uldum,, S. Bernander,, C. L. Luck,, G. Wewalka,, B. Abraham,, V. Gaia, and, T. G. Harrison. 2003. Clinical utility of urinary antigen detection for diagnosis of community-acquired, travel-associated and nosocomial Legionnaires’ disease. J. Clin. Microbiol. 41: 838– 840.
3. Lindsay, D. S. J.,, W. H. Abraham,, W. Findlay,, P. Christie,, F. Johnston, and, G. F. S. Edwards. 2004. Laboratory diagnosis of Legionnaires’ disease due to Legionella pneumophila serogroup 1: comparison of phenotypic and genotypic methods. J. Med. Microbiol. 53: 183– 187.
4. McDade, J. E.,, C. C. Shepard, and, D. W. Fraser. 1977. Legionnaires’ disease: isolation of a bacterium and demonstration of its role in respiratory disease. N. Engl. J. Med. 297: 1197– 1203.
5. Murdoch, D. R. 2003. Nucleic acid amplification tests for the diagnosis of pneumonia. Clin. Infect. Dis. 36: 1162– 1170.
6. Sopena, N.,, M. Sabria-Leal,, M. L. Pedro-Botet,, E. Padilla,, J. Dominguez,, J. Morera, and, P. Tudela. 1998. Comparative study of the clinical presentation of Legionella pneumophila and other community-acquired pneumonias. Chest 113: 1195– 1200.
7. van der Zee, A.,, H. Verbakel,, C. de Jong,, R. Pot,, M. Peeters,, J. Schellekens, and, A. Bergmans. 2002. Clinical validation of diagnosis of Legionella infections, p. 189–192. In R. Marre,, Y. Abu Kwaik,, C. Bartlett,, N. P. Cianciotto,, B. S. Fields,, M. Frosch,, J. Hacker, and, P. C. Luck (ed.), Legionella, ASM Press, Washington, DC.

Tables

Detection of Legionella spp. and Legionella pneumophila-Specific DNA in Respiratory Secretions by PCR-Enzyme-Linked Immunosorbent Assay and Comparison with Conventional Methods
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Citation: Lindsay D, Abraham W, Brown A, Edwards G. 2006. Detection of spp. and Specific DNA in Respiratory Secretions by PCR-Enzyme-Linked Immunosorbent Assay and Comparison with Conventional Methods, p 55-57. In Cianciotto N, Kwaik Y, Edelstein P, Fields B, Geary D, Harrison T, Joseph C, Ratcliff R, Stout J, Swanson M (ed), . ASM Press, Washington, DC. doi: 10.1128/9781555815660.ch15
/content/10.1128/9781555815660.ch15.ch15tab01
TABLE 1
DFA, culture and PCR results on the 21 definitive and 5 presumptive cases
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TABLE 1

DFA, culture and PCR results on the 21 definitive and 5 presumptive cases

Citation: Lindsay D, Abraham W, Brown A, Edwards G. 2006. Detection of spp. and Specific DNA in Respiratory Secretions by PCR-Enzyme-Linked Immunosorbent Assay and Comparison with Conventional Methods, p 55-57. In Cianciotto N, Kwaik Y, Edelstein P, Fields B, Geary D, Harrison T, Joseph C, Ratcliff R, Stout J, Swanson M (ed), . ASM Press, Washington, DC. doi: 10.1128/9781555815660.ch15

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