VALIDATION OF A FAST AND SIMPLE DIAGNOSTIC KIT FOR HLB CAUSAL AGENT DETECTION

STOLOWICS FG; L. LAROCCA; WERBAJH S; JUAN PEDRO AGOSTINI; JONATHAN REDES; CAROLINA CARRILLO; VOJNOV A

Congreso; REUNIÓN ANUAL DE SOCIEDADES DE BIOCIENCIA 2019; 2019

The Huanglongbing (HLB), is the mostdevastating disease in citrus worldwide, due to the damage itcauses, the difficulty of diagnosis and the speed of its expansion.The causal pathogen, Candidatus Liberibacter spp (Ca. L), istransmitted by the insect Diaphorina citri. The species Ca. Lasiaticus has been detected in Argentina, and the vectordistributed at least in nine provinces. Since 2010, the ArgentinianNational Service for Health and Agro-Food Quality (SENASA), hasimplemented a National HLB Prevention Program to safeguardproductivity in this important sector. Nowadays, HLB diagnosis isperformed by PCR, nested PCR, real time PCR or somecombination of them, requiring purified genomic DNA,sophisticated equipment and qualified human resources. The aimof this study was to evaluate the performance of a sensible, fastand simple diagnosis test based on specific DNA isothermalamplification of Ca. L asiaticus by comparison with PCR andqPCR, considered the Gold standard methods to HLB diagnosis.We applied the test in a group of samples whose true diseasestatus was defined by the mentioned gold standard techniques.Analyzing the results by a 2 x 2 contingency table, wedetermined Sensitivity and Specificity of the test, and thepositive and negative predictive values (PPV and NPV). In a firsttest, 30 DNA samples were analyzed and compared with qPCRtechnique with a concordance in 28 samples (PPV and NPV of100 and 88.88 %, respectively). Adjusting test parameters ofreading out, 23 new samples consisting on midribs and purifiedgenomic DNA from uninfected or Ca. L. asiaticus infected plantlines were analyzed in a blind assay comparing with PCR/nestedPCR applied by the Molecular Laboratory of the EEA Montecarlo(INTA), obtaining 100% of concordance (PPV and NPV, both100%). The results obtained in the present study demonstrateda high quality of our diagnostic test, with low cost, making it avalid and useful tool to support the diagnosis of HLB disease.