Polymerase chain reaction targeting insertion sequence for the diagnosis of extrapulmonary tuberculosis.

BACKGROUND & OBJECTIVES Diagnosis of extrapulmonary tuberculosis (EPTB) is difficult using conventional diagnostic methods. This study was conducted to evaluate the use of polymerase chain reaction (PCR) in diagnosis of definitive and probable extrapulmonary tuberculosis patients, and to assess the performance of insertion sequence (IS) 6110 based PCR assay as compared to conventional culture by Lowenstein-Jensen (LJ) method for the diagnosis of EPTB. METHODS A total of 178 non repeated clinical specimens were collected from clinically suspected extrapulmonary tuberculosis patients. The specimens included 59 ascitic fluid, 54 pleural fluid, 25 cerebrospinal fluid (CSF), 12 fine needle aspiration (FNA), 8 urine, 7 pus, 6 synovial fluid, 2 skin tissue, one pericardial fluid, one liver abscess, one pancreatic cyst fluid, one omental biopsy and one semen sample. All these clinical samples were subjected to Ziehl-Neelsen staining (ZN) for acid fast bacilli (AFB) and culture on LJ medium. PCR was performed by targeting 123bp fragment of insertion sequence IS6110 of Mycobacterium tuberculosis (MTB). RESULTS Of the 178 specimens, 10 (5.61%) were ZN smear positive for AFB, six (3.37%) were L-J culture positive from 10 AFB smear positive cases and 48 (26.96%) were PCR IS 6110 positive for M. tuberculosis. INTERPRETATION & CONCLUSIONS PCR using IS6110 primer was able to pick up more EPTB patients compared to conventional L-J culture method for detection of M. tuberculosis. False positive PCR IS6110 in three CSF samples may be due to latent TB infection which was limitation in this study

Distribution of Papaya Ring Spot Virus Infecting Papaya in Kerala, India

In the midst of the rising interest of papaya cultivation in Kerala, it is inevitable to study the current status of the Papaya ring spot disease in the state. Knowing the severity and distribution of PRSV isolates, causing ring spot disease, helps in the better formulation of effective management strategies against it. Purposive sampling surveys carried out in 2021-22 across five agro ecological units (AEUs) of Kerala, recorded papaya ring spot disease incidence (PRSD) ranging from 50 to 100 per cent in the cultivated areas with vulnerability index of 33.54 to 98.22. Highest disease incidence was recorded from AEU 8- Southern Laterites: Thiruvananthapuram (100.0%) and highest vulnerability index was recorded from Kalliyoor (98.22). Twenty symptomatic samples tested positive for PRSV in double antibody sandwich- ELISA (DAS-ELISA) using PRSV polyclonal antiserum. RT-PCR using coat protein gene specific primers RKJ 52 and 3 yielded amplicons of size approximately ~890bp in all the samples. Virulence of the Kerala isolates was evaluated based on the symptom expression, disease incidence and vulnerability index, upon mechanical transmission of PRSV on two months old papaya plants (variety- Red Lady). PRSV isolate from Kalliyoor (TVM1) inoculated on papaya plants expressed chlorosis, mottling, malformation of leaves and stunting with 96.80 vulnerability index. Maximum vulnerability index and severe symptoms including stunting in the inoculated plants were observed in isolates from southern and central Kerala, which include Thiruvananthapuram, Alappuzha and Thrissur had the most virulent PRSV isolates compared to north Kerala