Detection of Mycobacterium tuberculosis with nested polymerase chain reaction analysis in enucleated eye ball in Eales' disease.

Nested polymerase chain reaction (nPCR) was performed on enucleated eyeball for detection of Mycobacterium tuberculosis (M. tb) genome in a patient with Eales' disease. PCR analysis in all previous studies has been done mainly using aqueous, vitreous and epiretinal membranes from these patients. Paraffin wax embedded tissue section of the enucleated eyeball was analyzed by histopathology and nPCR targeting MPB64 gene and IS6110 region of M. tb genome. Lymphocytic infiltration was seen in the vitreous, iris and the retinal tissue. Ziehl Neelsen stain was negative for acid fast bacilli. Caseation necrosis was not seen in any section. Agarose gel electrophoretogram showed positive results with 200 bp specific amplified product targeting MPB64 gene, whereas nPCR targeting IS6110 region was negative. Since biopsy proven M. tb is extremely difficult in ocular tissues due to extensive necrosis, the nPCR technique aided in the diagnosis.

Multilevel Precision-Based Rational Design of Chemical Inhibitors Targeting the Hydrophobic Cleft of Toxoplasma gondii Apical Membrane Antigen 1 (AMA1).

Toxoplasma gondii is an intracellular Apicomplexan parasite and a causative agent of toxoplasmosis in human. It causes encephalitis, uveitis, chorioretinitis, and congenital infection. T. gondii invades the host cell by forming a moving junction (MJ) complex. This complex formation is initiated by intermolecular interactions between the two secretory parasitic proteins-namely, apical membrane antigen 1 (AMA1) and rhoptry neck protein 2 (RON2) and is critically essential for the host invasion process. By this study, we propose two potential leads, NSC95522 and NSC179676 that can efficiently target the AMA1 hydrophobic cleft, which is a hotspot for targeting MJ complex formation. The proposed leads are the result of an exhaustive conformational search-based virtual screen with multilevel precision scoring of the docking affinities. These two compounds surpassed all the precision levels of docking and also the stringent post docking and cumulative molecular dynamics evaluations. Moreover, the backbone flexibility of hotspot residues in the hydrophobic cleft, which has been previously reported to be essential for accommodative binding of RON2 to AMA1, was also highly perturbed by these compounds. Furthermore, binding free energy calculations of these two compounds also revealed a significant affinity to AMA1. Machine learning approaches also predicted these two compounds to possess more relevant activities. Hence, these two leads, NSC95522 and NSC179676, may prove to be potential inhibitors targeting AMA1-RON2 complex formation towards combating toxoplasmosis.

Detection of Mycobacterium tuberculosis by polymerase chain reaction in a case of orbital tuberculosis.

Orbital tuberculosis is quite uncommon. We report a case of orbital tuberculosis in a 3-year-old child from Bangladesh who presented with swelling and discharging sinus in the lower part of the orbit. Histopathology revealed a granulomatous inflammation with caseation necrosis. Polymerase chain reaction (PCR) showed amplification of the Mycobacterium tuberculosis genome. The patient responded to a course of antituberculous treatment. Mycobacterium tuberculosis should be considered in the differential diagnosis of inflammatory orbital disease in the Indian subcontinent where tuberculosis is prevalent.