Use of virus suspensions without RNA extraction as RT-PCR templates for detection of Newcastle disease virus

Allantoic fluid (AF) and cell culture supernatant (CCS) obtained from eggs or cells infected with strain I-2 of Newcastle disease virus were processed by different RNA template preparation methods for direct use in reverse transcriptase-polymerase chain reaction (RT-PCR). The objective was to determine the most effective technique for viral RNA extraction with consideration for efficacy, economy and simplicity. Results showed that use of undiluted CCS without RNA extraction or other treatment as template for RT-PCR produced a positive signal whereas direct use of undiluted AF did not. When aliquots of each sample dilution were used, an amplicon was detected from 1:10 dilution of both AF and CCS whereas no PCR products were amplified from both AF and CCS at 1:100 dilution. Both boiled undiluted AF and CCS produced positive signals when were used as templates for RT-PCR. An important contribution of the present study is the evidence that crude CCS, diluted or boiled AF and CCS may be used directly in RT-PCR without further manipulation, and yielded a positive PCR result comparable to those obtained from RNA extracted by silica gel based method.