Molecular and Nanotechnologic Approaches to Etiologic Diagnosis of Infectious Syndromes

Sankar, Sathish; Ramamurthy, Mageshbabu; Nandagopal, Balaji; Srikanth, Padma; Venkatraman, Ganesh; Sridharan, Gopalan

doi:10.1007/bf03256405

Cited by 6 publications

(3 citation statements)

References 53 publications

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“…Early detection would help provide appropriate patient care, prevent outbreaks, and determine treatment outcome [1]. In syndromic diagnosis, where different pathogens present with similar symptoms, the use of highly sensitive and specific diagnostic methods would help in understanding the real spectrum of a particular disease in the specific geographic population [2]. Pathogens such as Salmonella Typhi, Salmonella Paratyphi A and Burkholderia pseudomallei are the important causes of acute febrile illness especially in tropical and subtropical countries [3,4].…”

Section: Correspondence Dr Satish Sankarmentioning

confidence: 99%

A novel multiplex real-time PCR for the detection of Salmonella Typhi, Salmonella Paratyphi A and Burkholderia pseudomallei in clinical samples

Saravanan¹,

Rajendiran²,

Sankar³

et al. 2019

APJHS

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Enteric fever, caused by infection with Salmonella Typhi or Paratyphi A, B, or C (typhoidal Salmonella) and melioidosis are among the most common bacterial causes of acute febrile illness in tropical and subtropical countries. The diseases are widely spread even in developed countries mainly affecting travellers returning from endemic areas. Melioidosis is an increasingly recognized fatal septicemic infection also mimicking tuberculosis. These pathogens are largely underreported due to lack of sensitive and specific diagnostic tools. In this study, an inhouse multiplex real-time PCR assay was developed for the simultaneous detection of Salmonella Typhi, S. Paratyphi A and B. pseudomallei using specific primers. The inhouse developed assay was evaluated on buffy coat and serum samples collected from patients with acute febrile illness at four different centres. The assay had a detection limit of less than 1 genome copy for S. Typhi and S. Paratyphi A and 18 genome copies per 10 µl of PCR input for B. pseudomallei. Among the 1101 samples tested by multiplex real-time PCR, one (0.09%) sample was positive for S. Typhi. One sample was positive for blood culture identified as S. Typhi but negative by real-time PCR. The samples were negative for S. Paratyphi A and B. pseudomallei. The multiplex real-time assay would be highly useful as a diagnostic aid for the syndromic approach and comprehensive diagnosis of enteric fever and melioidosis. The assay could improve the overall diagnostic capability and be a useful tool during outbreak investigations.

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Section: Correspondence Dr Satish Sankarmentioning

confidence: 99%

A novel multiplex real-time PCR for the detection of Salmonella Typhi, Salmonella Paratyphi A and Burkholderia pseudomallei in clinical samples

Saravanan¹,

Rajendiran²,

Sankar³

et al. 2019

APJHS

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“…In addition, multiplex realtime PCR assays on the same automated instrument to simultaneously detect and screen for a variety of pathogens are currently under development and are expected in time to replace stand-alone assays for specific pathogens. 4 Despite these exciting possibilities, the cost of such multiplex instrumentation is a significant limitation for many smaller clinical laboratories. This review focuses on the individual real-time PCR assays that are currently widely available in the diagnosis of infectious disease emergencies.…”

Section: Real-time Pcrmentioning

confidence: 99%

Role of Molecular Diagnostics in the Management of Infectious Disease Emergencies

Krishna

Cunnion

2012

Medical Clinics of North America

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In the setting of infectious disease emergencies, rapid and accurate identification of the causative agent is critical to optimizing antimicrobial therapy in a timely manner. It is clearly evident that the age of molecular diagnostics is now upon us, with real-time PCR becoming the standard of diagnosis for many infectious disease emergencies in either monoplex or multiplex format. Other molecular techniques such as whole or partial genome sequencing, microarrays, broad-range PCR, restriction fragment length polymorphisms, and molecular typing are also being used. However, for most small clinical laboratories, implementation of these advanced molecular techniques is not feasible owing to the high cost of instrumentation and reagents. If these tests are not available in-house, samples can be sent to national reference laboratories (eg, Mayo Medical Laboratories and Quest Diagnostics) for real-time PCR assays that can be completed in 1 day. It is anticipated that over time commercial real-time PCR tests and instrumentation will become more standardized and affordable, allowing individual laboratories to conduct tests locally, thus further reducing turnaround time. Although real-time PCR has been proved to expand our diagnostic capability, it must be stressed that such molecular methodology constitutes only an additional tool in the diagnosis of infectious diseases in emergency situations. Phenotypic methodologies (staining, cultures, biochemical tests, and serology) still play a critical role in identifying, confirming, and providing antibiotic susceptibility testing for many microbial pathogens. As multiplex assays become increasingly available, there will be even greater temptation for taking a “shotgun” approach to diagnostic testing. These new technologies will not substitute for a proper history and physical examination leading to a thoughtful differential diagnosis. None the less, these new molecular tests increase the capability of the diagnostician to rapidly identify the microbiological etiology of an infection. An added advantage of rapid diagnostic tests often not emphasized is the capability to rule out certain diagnoses for which unnecessary antimicrobial therapy may otherwise be instituted and/or continued.

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“…Early and rapid diagnosis of the infecting agent would enable prompt and appropriate therapy. Many molecular techniques have been evaluated for rapid diagnosis of infectious syndromes, and include real-time multiplex PCR, DNA microarray, loop-mediated isothermal amplification, and other similar assays [48].…”

Section: Introductionmentioning

confidence: 99%

Application of viromics: a new approach to the understanding of viral infections in humans

et al. 2017

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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.

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Molecular and Nanotechnologic Approaches to Etiologic Diagnosis of Infectious Syndromes

Cited by 6 publications

References 53 publications

A novel multiplex real-time PCR for the detection of Salmonella Typhi, Salmonella Paratyphi A and Burkholderia pseudomallei in clinical samples

A novel multiplex real-time PCR for the detection of Salmonella Typhi, Salmonella Paratyphi A and Burkholderia pseudomallei in clinical samples

Role of Molecular Diagnostics in the Management of Infectious Disease Emergencies

Application of viromics: a new approach to the understanding of viral infections in humans

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