Simplified PCR for detection of Haemophilus ducreyi and diagnosis of chancroid

Abstract

A simplified PCR was developed for detection of Haemophilus ducreyi in samples from chancroid patients. The strategy included a straightforward chloroform extraction sample preparation method, a one-tube nested PCR to minimize contamination risks, and a colorimetric method for detection of products. Primers were designed from published nucleotide sequences of the 16S rRNA gene of H. ducreyi, with longer outer primers for annealing at a higher temperature and shorter inner primers labelled with biotin and digoxigenin for binding with avidin and colorimetric detection. The PCR technique detected all 35 strains of H. ducreyi tested, from four different geographical regions, and was negative for other, related strains of bacteria and for the common contaminating bacteria tested. Of 25 samples from H. ducreyi culture-positive chancroid patients, 24 were PCR positive and 1 produced a weak reaction. Of 83 samples from clinical cases of chancroid in the Republic of South Africa, 69 were PCR positive. The sensitivity of PCR compared with that of clinical diagnosis was 83%. All 50 negative control samples were negative. Encouraging results were also obtained with a consecutive series of 25 genital ulcer patients in Tanzania, of whom 9 were PCR positive. The adaptations of this simplified PCR strategy, at the sensitivity and specificity levels obtained, mean it will be useful for detection of H. ducreyi in areas where the organism is endemic, particularly where testing by culture is difficult or impossible.

Using a gel‐free PCR‐ELISA for the molecular identification of wheat genotypes carrying wheat–rye translocations

“…4). As pointed out by West et al. (1995), the use of a high copy number target sequence as the test target (the repetitive RIS‐1 element; Moore et al.…”

“…The aim of this work was to develop a robust and simple PCR‐ELISA gel‐free assay to speed the identification of 1RS‐carrying wheat genotypes in breeding programmes, based on the direct amplicon capture approach reported by West et al. (1995).…”

“…1RS rye chromosome detection by PCR‐ELISA: PCR conditions were the same as above except that forward and reverse primers were 5′‐end labelled with digoxigenin (DIG) and biotin, respectively (Biosource International, Camarillo, CA, USA) following West et al. (1995).…”

Herpes Simplex Virus (HSV) Infection in Men with Genital Ulcer Disease (GUD) - as Observed in Dhaka Medical College Hospital

“…Detection of H. ducreyi by PCR technique: a dacron tipped swab was subsequently rubbed against the ulcer base and stored in a sterile 1 molar phosphate buffered saline solution (PBS; 50mM sodium phosphate, 0.15M sodium chloride; pH 7.5), but without sodium chenodeoxycholate 9 and stored at -700c until shipped to the laboratory. Three different primers were used for the detection of the H. ducreyi genome [10][11][12] . Only sample giving a positive result with three primers were considered as being positive.…”

Development of a Serological Test for Haemophilus ducreyi for Seroprevalence Studies

“…H. ducreyi is a fastidious, slow-growing gram-negative rod, which is known for its inability to synthesize heme. Recent reviews of chancroid and H. ducreyi are available (1,21,37). The incubation period for chancroid is between 4 and 7 days, when a small inflammatory papule or pustule containing polymorphonuclear leukocytes may be seen.…”