Académique Documents
Professionnel Documents
Culture Documents
Roman Wölfel
Martin Pfeffer
Evaluation of sampling technique and transport
Sandra Essbauer
Sylke Nerkelun
media for the diagnostics of adenoviral
Gerhard Dobler eye infections
Adenovirus sampling and transport
Results
5.3 ∞a 6 ∞a 4 ∞a 6 ∞a 6
4.3 4.0 ± 0.01 6 3.9 ± 0.06 3 4.0 ± 0.02 6 4.0 ± 0.02 6
3.3 2.9 ± 0.08 6 2.9 ± 0.11 4 2.9 ± 0.16 6 3.0 ± 0.08 6
FFU focus-forming units, SEM standard error of the mean
a
More than 300 focus-forming units per well
cant (P >0.9). We did not see any comparable cell damage
in the diagnostic process of specimens from locally
anaesthetised patients. Likewise, by using DNA extraction
and qPCR protocols described above, we did not find any
significant influence of oxyprocain on real-time detection
of HAdV-DNA (data not shown), indicating that neither the
local anaesthetic nor the conservation material inhibited
detection of HAdV by qPCR.
study was to determine if local anaesthesia of the eye Results of this study on HAdV infectivity in centrifu-
should be recommended for HAdV specimen collection. gation indicate a significant difference between the tested
Because HAdV replicates mainly within the nucleus [3], it VTS for virus preservation. In contrast to specific virus
is most important to collect as many conjunctival cells as
possible. We showed that topical application of an
anaesthetic is an appropriate way of collecting material
for diagnostics, which does not interfere with downstream
techniques such as qPCR, an important screening method,
or virus isolation by cell culture. The cytotoxic-like effects
that we observed in cell culture at the highest concentration
of local anaesthetics are most likely not relevant for the
investigation of clinical samples. The appropriate amount
of anaesthetic eye drops (50 μl) corresponds to the
complete dose normally administered to a patient’s eye.
Therefore, the amount of local anaesthetics in the specimen
will always be much lower.
References
1. Aoki K, Tagawa Y (2002) A twenty- 4. Darougar S, Walpita P, Thaker U, 8. Ford E, Nelson KE, and Warren D
one year surveillance of adenoviral Viswalingam N, Wishart MS (1984) (1987) Epidemiology of epidemic
conjunctivitis in Sapporo, Japan. Int Rapid culture test for adenovirus iso- keratoconjunctivitis. Epidemiol Rev
Ophthalmol Clin 42:49–54 lation. Br J Ophthalmol 68:405–408 9:244–261
2. Azevedo AM, Durigon EL, Okasima V, 5. Dawson C, Jawetz E, Hanna L, Winn 9. Gardner PS, McQuillen J (1980) Ap-
Queiroz DA, de Moraes-Vasconcelos WE, and Thompson C (1960) A family plication of immunofluorescence. In:
D, Duarte AJ, Grumach AS (2003) outbreak of adenovirus 8 infection Rapid virus diagnosis, 2nd edn.
Detection of influenza, parainfluenza, (epidemic keratoconjunctivitis). Am Butterworth, London, pp 92–109
adenovirus and respiratory syncytial J Hyg 72:279–283 10. Heim A, Ebnet C, Harste G, Pring-
virus during asthma attacks in children 6. Dunn JJ, Billetdeaux E, Skodack-Jones Akerblom P (2003) Rapid and quanti-
older than 2 years old. Allergol L, and Carroll KC (2003) Evaluation of tative detection of human adenovirus
Immunopathol (Madr ) 31:311–317 three Copan viral transport systems for DNA by real-time PCR. J Med Virol
3. Boniuk M, Phillips CA, Hines MJ, and the recovery of cultivatable, clinical 70:228–239
Friedman JB (1966) Adenovirus infec- virus isolates. Diagn Microbiol Infect 11. Huntoon CJ, House RF, Jr., and Smith
tions of the conjunctiva and cornea. Dis 45:191–197 TF (1981) Recovery of viruses from
Trans Am Acad Ophthalmol 7. Fitch CP, Rapoza PA, Owens S, three transport media incorporated into
Otolaryngol 70:1016–1026 Murillo-Lopez F, Johnson RA, Quinn culturettes. Arch Pathol Lab Med
TC, Pepose JS, Taylor HR (1989) 105:436–437
Epidemiology and diagnosis of acute
conjunctivitis at an inner-city hospital.
Ophthalmology 96:1215–1220
12. Jawetz E, Thygeson P, Hanna L, 16. Kowalski RP and Gordon YJ (1989) 21. Richmond S, Burman R, Crosdale E,
Nicholas A, Kimura SJ (1957) The Comparison of direct rapid tests for the Cropper L, Longson D, Enoch BE,
etiology of epidemic keratoconjuncti- detection of adenovirus antigen in Dodd CL (1984) A large outbreak of
vitis. Am J Ophthalmol 43:79–83 routine conjunctival specimens. keratoconjunctivitis due to adenovirus
13. Jensen C, Johnson FB (1994) Com- Ophthalmology 96:1106–1109 type 8. J Hyg (Lond) 93:285–291
parison of various transport media for 17. Lehtomaki K, Julkunen I, Sandelin K, 22. Vastine DW, Schwartz HS,
viability maintenance of herpes simplex Salonen J, Virtanen M, Ranki M, Hovi Yamashiroya HM, Smith RF, and Guth
virus, respiratory syncytial virus, and T (1986) Rapid diagnosis of respiratory SB (1977) Cytologic diagnosis of
adenovirus. Diagn Microbiol Infect Dis adenovirus infections in young adult adenoviral epidemic keratoconjunctivi-
19:137–142 men. J Clin Microbiol 24:108–111 tis by direct immunofluorescence.
14. Johnson FB (1990) Transport of viral 18. Mellman-Rubin TL, Kowalski RP, Invest Ophthalmol Vis Sci 16:195–200
specimens. Clin Microbiol Rev Uhrin M, Gordon YJ (1995) Incidence 23. Warren D, Nelson KE, Farrar JA,
3:120–131 of adenoviral and chlamydial coinfec- Hurwitz E, Hierholzer J, Ford E,
15. Kinchington PR, Turse SE, Kowalski tion in acute follicular conjunctivitis. Anderson LJ (1989) A large outbreak
RP, and Gordon YJ (1994) Use of Am J Ophthalmol 119:652–654 of epidemic keratoconjunctivitis:
polymerase chain amplification reac- 19. Pereira HG, Allison AC, Balfour B problems in controlling nosocomial
tion for the detection of adenoviruses in (1959) Multiplication of adenovirus spread. J Infect Dis 160:938–943
ocular swab specimens. Invest type 5 studied by infectivity titrations 24. Wolfel R, Pfeffer M, Dobler G, and
Ophthalmol Vis Sci 35:4126–4134 and by the fluorescent antibody tech- Finke EJ (2004) [Virological investi-
nique. Virology 7:300–314 gation of Keratoconjunctivitis epidem-
20. Radstrom P, Knutsson R, Wolffs P, ica]. Wehrmed Wehrpharm 2:36–41
Lovenklev M, and Lofstrom C (2004)
Pre-PCR processing: strategies to gen-
erate PCR-compatible samples. Mol
Biotechnol 26:133–146