Polymerase Chain Reaction to Detect Infectious Laryngotracheitis Virus in Conjunctival Swabs from Experimentally Infected Chickens

Abstract

The polymerase chain reaction (PCR) was standardized and assessed as a potential diagnostic test for infectious laryngotracheitis using conjunctival swabs collected from experimentally infected chickens. Polymerase chain reaction primers based on the sequence of a 1.1-kb BamHI restriction enzyme fragment of the Ontario 1598 (Ont 1598) strain of infectious laryngotracheitis virus were selected and 300 fg of purified viral DNA were detected by ethidium bromide staining of agarose gels or 30 fg were detected by DNA hybridization. At least five different strains (Ont 1598, ATCC N-71851, LT-IVAX, Laryngo-Vac, and a local strain 322) were amplified whereas other avian pathogens and uninfected cell cultures tested negative. Swabs collected from experimentally infected chickens were analyzed by both PCR and virus isolation on various days postinfection. A comparison of virus isolation to PCR indicated that, in the mid-postinfection phase, PCR and virus isolation appeared to be comparable with a kappa value of greater than 0.8. The polymerase chain reaction was shown to be the better test later in infection, when clinical recovery had occurred.

Molecular detection and characterization of infectious laryngotracheitis virus (Gallid herpesvirus-1) from clinical samples of commercial poultry flocks in India

“…Thus, the PCR assay would be a useful tool to confirm the ILTV. PCR has been found to be more sensitive than virus isolation from clinical samples [33], especially when other contaminant viruses such as adenoviruses are present [1]. All the eight Indian ILTV sequences were closely related with CEO vaccine strains of Italy, USA, Brazil and China.…”

Survey of infectious laryngotracheitis outbreak in layer hens and differential diagnosis with other respiratory pathogens

“…Later, PCR yields higher numbers of positive samples as compared to virus isolation (Alexander & Nagy, 1997). In some cases, the failure to isolate ILTV may be due to low virus concentrations present in carrier birds Williams et al, 1994).…”

“…In 13 of the 24 positive farms, no clinical signs were observed during sample collection. Clinical signs usually appear between 6 and 12 days PI, but the PCR technique can detect DNA virus already on the third day PI (Alexander & Nagy 1997). Consequently, it is possible that samples were collected in the early stages of infection, before to the appearance of clinical signs.…”

“…Polymerase chain reaction (PCR) has been used successfully to detect ILTV DNA from the trachea and extratracheal sites, such as the conjunctiva and the trigeminal ganglia Alexander et al, 1998;Alexander & Nagy, 1997;Williams et al, 1992).…”

Herpesvirus-like respiratory infection in African penguins Spheniscus demersus admitted to a rehabilitation centre

“…Samples were subjected to basic Chelex extraction for DNA isolation. A PCR targeting a 1.1 kb BamHI restriction enzyme fragment of GaHV-1 was performed following the protocol provided by Alexander & Nagy (1997); positive and negative controls were used in each batch as well as the amplification of a housekeeping gene to monitor for sample quality and the presence of PCR inhibitors. Nineteen of these samples were also subjected to a PCR test targeting the DNA polymerase gene of large DNA viruses (LDVs) following the protocols provided by Hanson et al (2006).…”