PCR analysis for assessment of bacterial bioburden in orthokeratology lens cases.PURPOSE
 To develop a PCR gel analysis method for assessing the bacterial bioburden in orthokeratology contact lens (OK) case fluid determined by culture.


 PCR was used to assess the bacterial bioburden (colony-forming units per milliliter) of OK after removal and soaking in the storage case for 6 h.  The signal intensity of the PCR bands was analyzed after grayscale image transformation.  The difference (cPCR-d) and ratio (cPCR-r) between a PCR signal and its background were used as two standardized indices of PCR signals.  The association between the two indices of the PCR signals and the bacterial bioburden determined by culture were analyzed with Pearson's correlation coefficient (r) and receiver operating characteristic (ROC) plots.


 Both cPCR-d and cPCR-r showed strong correlations with the bacterial bioburden (r>0.7, p<0.0001).  ROC analysis enabled good determination of the cutoff values for the two PCR indices with acceptable sensitivity and specificity (78-89%) to assess the degree of bacterial contamination.


 PCR analysis provides an alternative and rapid method for assessing the bacterial bioburden of OK lens cases, and these results should serve as a warning to OK wearers to follow appropriate lens care procedures to prevent infection.
PCR
To develop a PCR gel analysis method for assessing the bacterial bioburden in orthokeratology contact lens (OK) case fluid determined by culture.



PCR gel analysis method
The signal intensity of the PCR bands was analyzed after grayscale image transformation. 
PCR bands
The signal intensity of the PCR bands was analyzed after grayscale image transformation. 
grayscale image transformation
The association between the two indices of the PCR signals and the bacterial bioburden determined by culture were analyzed with Pearson's correlation coefficient (r) and receiver operating characteristic (ROC) plots.



Pearson's correlation coefficient (r)
The association between the two indices of the PCR signals and the bacterial bioburden determined by culture were analyzed with Pearson's correlation coefficient (r) and receiver operating characteristic (ROC) plots.



receiver operating characteristic (ROC) plots