Tuberculosis is an under-recognized yet catastrophic health problem, particularly in developing countries. The HIV pandemic has served to increase the number of susceptible individuals, and multidrug-resistance and poor socioeconomic conditions also augment the prevalence and the consequences of the disease. To control the disease and its spread, it is vital that tuberculosis diagnostics are accurate and rapid. Whereas microscopy and culture have several limitations (low sensitivity is a problem for the former, while the latter has a delayed turnaround time), PCR-based techniques targeting regions of the Mycobacterium tuberculosis genome such as IS6110 have proved to be useful. The purpose of this review is to assess the use of PCR-RFLP, nested PCR and real-time PCR protocols and the choice of target regions for the detection of M. tuberculosis. Real-time PCR for the detection of M. tuberculosis target genes in clinical specimens has contributed to improving diagnosis and epidemiologic surveillance in the past decade. However, targeting one genome sequence such as IS6110 may not by itself be sufficiently sensitive to reach 100% diagnosis, especially in the case of pulmonary tuberculosis. Additional testing for target genome sequences such as hsp65 seems encouraging. An interesting approach would be a multiplex real-time PCR targeting both IS6110 and hsp65 to achieve comprehensive and specific molecular diagnosis. This technology needs development and adequate field testing before it becomes the acceptable gold standard for diagnosis.

Hantavirus infections are now recognized to be a global problem. The hantaviruses include several genotypic variants of the virus with different distributions in varying geographical regions. The virus genotypes seem to segregate in association with certain manifestations specific for each syndrome. They primarily include HFRS, HCPS, febrile illness with or without mild involvement of renal diseases. In the course of our study on hantavirus etiology of febrile illnesses, we recovered a hantavirus strain identified by nPCR. This has been sequenced to be Hantaan-like virus (partial S segment). The current manuscript is focused on understanding the N protein coded by S segment in terms of variation of amino acid sequences of the virus genotypes associated with HFRS. The diagnosis of this infection is achieved by PCR testing of serum/plasma or demonstration of IgM/IgG in serum. The limitations of PCR are temporal often not positive after 7 days of onset of infection. IgM detection is possible around this period and up to 21 days. IgG detection is less definitive in acute infections. Here, we report characterization of the sequence diversity of HFRS strains, 3D structure of Hantaan N protein, and B-cell epitopes on this molecule. We predicted a 20 amino acid sequence length peptide by using BepiPred online server in IEDB analysis resource program. We suggest this peptide may be used for development of geographic region-specific immunoassays like EIAs for antibody detection, monoclonal antibody development, and immunoblots (line immunoassay). J. Cell. Biochem. 118: 1182-1188, 2017. © 2016 Wiley Periodicals, Inc.

Hantaviruses are emerging viral pathogens that causes hantavirus cardiopulmonary syndrome (HCPS) in the Americas, a severe, sometimes fatal, respiratory disease in humans with a case fatality rate of ≥50%. IgM and IgG-based serological detection methods are the most common approaches used for laboratory diagnosis of hantaviruses. Such emerging viral pathogens emphasizes the need for improved rapid diagnostic devices and vaccines incorporating pan-specific epitopes of genotypes. We predicted linear B-cell epitopes for hantaviruses that are specific to genotypes causing HCPS in humans using in silico prediction servers. We modeled the Andes and Sin Nombre hantavirus nucleocapsid protein to locate the identified epitopes. Based on the mean percent prediction probability score, epitope IMASKSVGS/TAEEKLKKKSAF was identified as the best candidate B-cell epitope specific for hantaviruses causing HCPS. Promiscuous epitopes were identified in the C-terminal of the protein. Our study for the first time has reported pan-specific B-cell epitopes for developing immunoassays in the detection of antibodies to hantaviruses causing HCPS. Identification of epitopes with pan-specific recognition of all genotypes causing HCPS could be valuable for the development of immunodiagnositic tools toward pan-detection of hantavirus antibodies in ELISA. J. Cell. Biochem. 118: 2320-2324, 2017. © 2017 Wiley Periodicals, Inc.

Our results suggest that the real-time PCR assay was more sensitive compared with the conventional PCR. The advantage of real-time PCR is that it can be performed much faster than conventional PCR. Real-time PCR is less time-consuming, less labour-intensive and also reduces the chance of contamination as there is no post-amplification procedure. In the entire study population, the major viruses detected using real-time PCR were EBV (34%), HSV-2 (10.8%) and VZV (6.8%).

This review is focused at exploring the strengths of modern technology driven data compiled in the areas of virus gene sequencing, virus protein structures and their implication to viral diagnosis and therapy. The information for virome analysis (viromics) is generated by the study of viral genomes (entire nucleotide sequence) and viral genes (coding for protein). Presently, the study of viral infectious diseases in terms of etiopathogenesis and development of newer therapeutics is undergoing rapid changes. Currently, viromics relies on deep sequencing, next generation sequencing (NGS) data and public domain databases like GenBank and unique virus specific databases. Two commonly used NGS platforms: Illumina and Ion Torrent, recommend maximum fragment lengths of about 300 and 400 nucleotides for analysis respectively. Direct detection of viruses in clinical samples is now evolving using these methods. Presently, there are a considerable number of good treatment options for HBV/HIV/HCV. These viruses however show development of drug resistance. The drug susceptibility regions of the genomes are sequenced and the prediction of drug resistance is now possible from 3 public domains available on the web. This has been made possible through advances in the technology with the advent of high throughput sequencing and meta-analysis through sophisticated and easy to use software and the use of high speed computers for bioinformatics. More recently NGS technology has been improved with single-molecule real-time sequencing. Here complete long reads can be obtained with less error overcoming a limitation of the NGS which is inherently prone to software anomalies that arise in the hands of personnel without adequate training. The development in understanding the viruses in terms of their genome, pathobiology, transcriptomics and molecular epidemiology constitutes viromics. It could be stated that these developments will bring about radical changes and advancement especially in the field of antiviral therapy and diagnostic virology.