Minimum detection limit of an in-house nested-PCR assay for herpes simplex virus and varicella zoster virus

Rodrigues, Diógenes; de‐Paris, Fernanda; Paiva, Rodrigo Minuto

doi:10.1590/0037-8682-1520-2013

Cited by 10 publications

(6 citation statements)

References 11 publications

(15 reference statements)

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“…Taking into account a median number of ten IS6110 copies per bacteria, the analytical sensitivity of the PCR can be estimated around 250 copies of MTB/mL. The sensitivity of the assay for HSV DNA was comparable to others HSV nucleic acid tests and probably sufficient to detect most of the HSV CNS infections regarding viral load generally observed in CSF . However, a better sensitivity for HSV detection, able to detect <500‐200 HSV DNA copies/mL, would be preferable to detect some of the specimens with low HSV DNA concentration .…”

Section: Discussionmentioning

confidence: 91%

Combined testing for herpes simplex virus and Mycobacterium tuberculosis DNA in cerebrospinal fluid of patients with aseptic meningitis in Burkina Faso, West Africa

Zida

Kolia-Diafouka

Kania

et al. 2018

Clinical Laboratory Analysis

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Background Little is known about the involvement of herpes simplex virus (HSV) or Mycobacterium tuberculosis (MTB) as potentially curable causes of central nervous system (CNS) infections in sub‐Saharan Africa. Objective In this study, we developed a PCR assay dedicated to simultaneous testing of HSV1/HSV2 and MTB in Burkina Faso, a country where HSV is neglected as a cause of CNS infection and where TB prevalence is high. Methods A consensus HSV1/HSV2 set of primers and probe were designed and combined to primers and probe targeting the IS6110 repetitive insertion sequence of MTB. Analytical performances of the assay were evaluated on reference materials. Cerebrospinal fluid (CSF) collected from subjects with aseptic meningitis was tested for HSV1/HSV2 and MTB DNA. Results The UL29 gene was chosen as a highly conserved region targeted by the HSV1/HSV2 nucleic acid test. The lower limits of detection were estimated to be 2.45 copies/µL for HSV1, 1.72 copies/µL for HSV2, and 2.54 IS6110 copies per µL for MTB. The PCR was used in 202 CSF collected from subjects suspected of aseptic meningitis. Five samples (2.46%) tested positive, including two children positive for HSV1 (0.99%) and three adults tested positive for MTB (1.47%). Conclusion Using an in‐house real‐time PCR assay, we showed that both HSV and MTB are etiologic pathogens contributing to aseptic meningitis in Burkina Faso. This molecular test may have clinical utility for early diagnosis for those treatable CNS infections.

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Section: Discussionmentioning

confidence: 91%

Combined testing for herpes simplex virus and Mycobacterium tuberculosis DNA in cerebrospinal fluid of patients with aseptic meningitis in Burkina Faso, West Africa

Zida

Kolia-Diafouka

Kania

et al. 2018

Clinical Laboratory Analysis

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“…These Viruses are related with a variety of clinical signs and symptoms of several organs, including central nervous system (CNS) diseases, skin lesion, eye and keratoconjunctivitis. In immunocompromised patients, the viruses may also trigger severe clinical appearance (6). Herpes viruses cause various ocular diseases which are considered as one of the main causes of eye infection etiology and blindness worldwide.…”

Section: Virus (Vzv) In Eye Infectionmentioning

confidence: 99%

“…Many molecular biology laboratories use in-house PCR assaystechniques specifically developed by laboratories for the diagnosis of infectious diseases. However, the lack of standardization and the poor reproducibility of in-house techniques have been argued as limitations against their routine use as diagnostic methods (6). PCR has been shown in recent studies to be a rapid and reliable method for detecting HSV infection (10).…”

Section: Virus (Vzv) In Eye Infectionmentioning

confidence: 99%

Prevalence of Herpes Simplex Virus type-1, 2 and Varicella Zoster Virus (VZV) in Eye Infection

Omidian

Sheikhi-Shooshtari

Fazeli³

2015

Iran J Virol

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Background and Aims: Herpes simplex virus (HSV) is a common cause of corneal disease and is the leading infectious cause of corneal blindness among developed nations. This study is aimed to provide an estimation of the incidence of Herpes Simplex Virustype-1, 2(HSV1, 2) and Varicella Zoster Virus (VZV) in tears and swab from eye infection by polymerase chain reaction (PCR) in eye disease in a suspected community. Materials and Methods: Fifty subjects without signs of ocular herpetic disease enrolled in the study. Serum samples from all subjects were tested for HSV IgG antibodies by enzymelinked immunosorbent assay (ELISA). Subjects were instructed to collect tear samples by touching the inner surface of the lower eyelid with an individually wrapped, sterile cotton swab and to place the swab immediately into labeled sterile tubes. Swabs were kept at 4°C until processed. Nucleic acid was extracted from the samples and PCR-amplified for HSV DNA. Results: Among 50 samples, 3 samples were refused because internal controls were negative. HSV infection was established in 10% (5 out of 50) of all patients. The prevalence of HSV infection in patients with no clinical suspicion of herpetic keratitis was 6%. Histopathologic evaluation revealed that among samples with positive PCR results, 100% had evidence of inflammation, 55% had giant cells, 39% had necrosis, 59% had vascularization, 67 % had ulcer and 100% of them had inclusion bodies. Conclusions: Because some of the patients with no clinical suspicion of herpes infection were found positive, we suggest that HSV to be considered as one of the underlying etiologies in any patient with corneal scar and keratitis. Therefore, performing further diagnostic methods, including PCR and histopathology, is mandatory to clearly understand the infection.

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“…For HSV-1, the sensitivity range was 96.7-100 % (Van Der Pol et al 2012). High analytical sensitivity of the HSV and VZV detection was achieved also by the method of a duplex nested-PCR assay that allowed the detection of 5 copies/μL for HSV and 10 copies/μL for VZV (Rodrigues et al 2013). VZV diagnosis is confirmed by the presence of VZV DNA or anti-VZV IgG, or both in cerebrospinal fluid (CSF) (Gilden et al 1994).…”

Section: Alphaherpesvirus Genomicsmentioning

confidence: 99%

Analytical methods in herpesvirus genomics

Blaškovičová

Labuda

2014

Acta Chimica Slovaca

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Genomics is a branch of bioanalytical chemistry characterized as the study of the genome structure and function. Genome represents the complete set of chromosomal and extrachromosomal genes of an organism, a cell, an organelle or a virus. There are at least five from eight species of herpesviruses commonly widespread among humans, Herpes simplex virus type 1 and 2, Varicella zoster virus, Epstein-Barr virus and Cytomegalovirus. Human gammaherpesviruses can cause serious diseases including B-cell lymphoma and Kaposi’s sarcoma. Diagnostics and study of the herpesviruses is directly dependent on the development of modern analytical methods able to detect and determine the presence and evolution of herpesviral particles/ genomes. Diagnostics and genomic characterization of human herpesvirus species is based on bioanalytical methods such as polymerase chain reaction (PCR), DNA sequencing, gel electrophoresis, blotting and others. The progress in analytical approaches in the herpesvirus genomics is reviewed in this article.

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Minimum detection limit of an in-house nested-PCR assay for herpes simplex virus and varicella zoster virus

Cited by 10 publications

References 11 publications

Combined testing for herpes simplex virus and Mycobacterium tuberculosis DNA in cerebrospinal fluid of patients with aseptic meningitis in Burkina Faso, West Africa

Combined testing for herpes simplex virus and Mycobacterium tuberculosis DNA in cerebrospinal fluid of patients with aseptic meningitis in Burkina Faso, West Africa

Prevalence of Herpes Simplex Virus type-1, 2 and Varicella Zoster Virus (VZV) in Eye Infection

Analytical methods in herpesvirus genomics

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