Diagnosing Herpetic Genital Ulcers in a Resource-Limited Setting- What Works?

Aims: Globally, viral agents, especially herpes simplex virus (HSV), have overtaken the bacterial causes of genital ulcers. Very few laboratories in India, perform culture techniques and polymerase chain reaction (PCR) for diagnosis of genital ulcers. This study aimed to establish the utility of existing tests, which are cheaper and need less technical expertise, when compared to newer tests such as PCR.Study Design: This cross sectional study was carried out to determine the aetiology of genital ulcers, with emphasis on diagnosis of herpetic ulcers, using newer and more accurate methods of diagnosis and evaluating their performance by comparing against viral culture as gold standard test.Place and Duration of the Study: The study was carried out over a period of one year in the Apex Regional Sexually Transmitted Diseases (STD) Centre at Safdarjung Hospital, New Delhi and the Department of Microbiology, AIIMS, New Delhi.Methodology: Fifty three patients with genital ulcers were included in the study. Specimens from ulcers were taken for various tests, including Giemsa stain, ELISA for HSV-1 & 2, PCR and Viral culture for HSV.Results: HSV was identified in 31 of 53 cases (58.5%), including 03 cases of HSV-1, and 28 cases of HSV-2. Sensitivity and specificity of PCR was 90.0% and 84.85%, respectively. Viral culture positivity was 37.7%.Conclusion: Genital herpes is associated with an increased risk of Human Immunodeficiency Virus (HIV) acquisition, and clinical manifestations are diverse; hence a presumptive diagnosis should be confirmed by reliable laboratory tests. Nucleic acid amplification tests (NAAT) are the most sensitive methods for direct detection of HSV. The extensive validation of these tests allows for their application in routine laboratory settings with consistency and greater diagnostic accuracy. When standardised and used, PCR is a highly reproducible, rapid and labour efficient method for HSV detection.

Alteration in sample preparation to increase the yield of multiplex Polymerase Chain Reaction assay for diagnosis of genital ulcer disease.

Purpose: Genital Ulcer Disease (GUD) is common sexually transmitted infection (STI). Multiple studies have shown that GUDs are strongly associated with the transmission and the acquisition of HIV infection. An accurate diagnosis of common etiology of GUD namely Herpes, syphilis and Chancroid is possible using Multiplex PCR (M-PCR). However, frequent presence of Polymerase Chain Reaction inhibitors in the ulcer swab specimen limits the performance of the assay. In order to overcome this problem, alternative specimen preparation method was used. Materials and Methods: To determine the common etiology, GUD specimens obtained under an STI operations research study were tested with M-PCR after the samples were prepared using Roche Amplicor specimen preparation kit. PCR inhibiting samples were identified from that, which showed negative results. These samples were subjected to phenol-chloroform extraction and ethanol precipitation before the conduct of M-PCR on them. Results: Of the 237 GUD specimens tested, in 145 etiologies could be detected, whereas 92 samples were found negative. Further spiking with one of the target DNA, 128 of the negative samples were found to contain the inhibitors. These 126 samples were then subjected to phenol chloroform extraction and ethanol precipitation followed by M-PCR. Using this method for sample preparation, etiology could be determined in 46 (23%) additional samples. This success rate of altered sample preparation method has been lower than that has reported. Conclusion: The results indicate that sample preparation using phenol chloroform extraction and ethanol precipitation, prior to M-PCR helps to eliminate the inhibitors and increase the yield of the assay. However, being a laborious procedure, it may be used for samples giving negative results after the screening by Roche Amplicor specimen preparation kit.

A Syndromic Approach to the Management of Genital Ulcers

Genital ulcer disease is a common presentation of sexually transmitted infections (STIs) and can cause significant morbidity in patients. Syphilis (chancre of primary syphilis) and genital herpes are the two most significant ulcerative STIs and are occasionally indistinguishable clinically. Infectious syphilis is on the rise in Singapore particularly in men who have sex with men (MSM); whilst genital herpes although showing a decreasing incidence in recent years still presents with more than 500 notifications a year as first episode genital herpes. Due to the breaks in the mucosa of the skin caused by the ulcers, the risk of acquisition as well as transmission of HIV infection are both increased making timely treatment of genital ulcers of utmost importance. Although uncommon in Singapore, chancroid, LGV and granuloma inguinale remain endemic in some parts of Asia and there should be a high index of suspicion for patients who have returned from these regions presenting with ulcerative lesions. All patients who present with genital ulcers should have their blood tested for syphilis and HIV as well as a swab for culture or PCR testing taken from the ulcer to exclude genital herpes.