title: The in vitro effects of proxymetacaine, fluorescein, and fusidic acid on real-time PCR assays used for the diagnosis of Feline herpesvirus 1 and Chlamydophila felis infections.
abstract: OBJECTIVE
  To investigate the possible inhibition of qPCR assays used for the diagnosis of ocular infections in cats by proxymetacaine, fluorescein, and fusidic acid, which are commonly used in veterinary ophthalmology.
  
  
  METHODS
  Fluorescein, proxymetacaine, and fusidic acid were tested for possible inhibition of a triplex qPCR assay designed to detect Chlamydophila felis, Feline herpesvirus 1 (FHV-1), and the feline 28S ribosomal DNA (28S rDNA) gene by comparing threshold cycle (C(t) ) values of samples with and without the three products. A second experiment was carried out to measure the effects of various dilutions of fusidic acid.
  
  
  RESULTS
  No statistically significant differences were detected between the C. felis, FHV-1, and 28S rDNA C(t) values with and without proxymetacaine or fluorescein. However, there was a statistically significant increase in FHV-1 (P < 0.01), C. felis (P < 0.01), and 28S rDNA (P < 0.05) C(t) values when fusidic acid was used. When dilutions of fusidic acid were tested, the results revealed that only the 1:2 dilution caused a statistically significant increase (P < 0.01) in the FHV-1 Ct values.
  
  
  CONCLUSIONS
  Proxymetacaine and fluorescein did not interfere with our qPCR assays for the detection of C. felis and FHV-1. The presence of fusidic acid caused a small inhibitory effect of doubtful clinical significance. In vivo studies are required to establish the clinical relevance of this study and to confirm our findings.
authors.affiliation: School of Veterinary Sciences, Small Animal Hospital, University of Bristol, Bristol, UK. SergiSegarra.vet@gmail.com
authors.forename: Sergi
authors.initials: S
authors.lastname: Segarra
authors.forename: Kostas
authors.initials: K
authors.lastname: Papasouliotis
authors.forename: Chris
authors.initials: C
authors.lastname: Helps
mesh_terms: D018138:Alphaherpesvirinae; D000779:Anesthetics, Local / Q000494:pharmacology*; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002371:Cat Diseases / Q000175:diagnosis* / Q000382:microbiology / Q000821:virology; D002415:Cats; D023501:Chlamydophila; D023521:Chlamydophila Infections / Q000175:diagnosis / Q000662:veterinary*; D015818:Eye Infections, Bacterial / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D015828:Eye Infections, Viral / Q000175:diagnosis / Q000662:veterinary; D005188:False Negative Reactions; D019793:Fluorescein / Q000494:pharmacology*; D005456:Fluorescent Dyes / Q000494:pharmacology*; D005672:Fusidic Acid / Q000494:pharmacology*; D006566:Herpesviridae Infections / Q000175:diagnosis / Q000662:veterinary; D066298:In Vitro Techniques; D011430:Propoxycaine / Q000494:pharmacology*; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary*
chemical_list: D000779:Anesthetics, Local; D000900:Anti-Bacterial Agents; D005456:Fluorescent Dyes; D005672:Fusidic Acid; C005717:proxymetacaine; D011430:Propoxycaine; D019793:Fluorescein