Detection and discrimination of herpes simplex viruses, Haemophilus ducreyi, Treponema pallidum, and Calymmatobacterium (Klebsiella) granulomatis from genital ulcers.

BACKGROUND: Genital ulcer disease (GUD) is commonly caused by pathogens for which suitable therapies exist, but clinical and laboratory diagnoses may be problematic. This collaborative project was undertaken to address the need for a rapid, economical, and sensitive approach to the detection and diagnosis of GUD using noninvasive techniques to sample genital ulcers. METHODS: The genital ulcer disease multiplex polymerase chain reaction (GUMP) was developed as an inhouse nucleic acid amplification technique targeting serious causes of GUD, namely, herpes simplex viruses (HSVs), H. ducreyi, Treponema pallidum, and Klebsiella species. In addition, the GUMP assay included an endogenous internal control. Amplification products from GUMP were detected by enzyme linked amplicon hybridization assay (ELAHA). RESULTS: GUMP-ELAHA was sensitive and specific in detecting a target microbe in 34.3% of specimens, including 1 detection of HSV-1, three detections of HSV-2, and 18 detections of T. pallidum. No H. ducreyi has been detected in Australia since 1998, and none was detected here. No Calymmatobacterium (Klebsiella) granulomatis was detected in the study, but there were 3 detections during ongoing diagnostic use of GUMP-ELAHA in 2004 and 2005. The presence of C. granulomatis was confirmed by restriction enzyme digestion and nucleotide sequencing of the 16S rRNA gene for phylogenetic analysis. CONCLUSIONS: GUMP-ELAHA permitted comprehensive detection of common and rare causes of GUD and incorporated noninvasive sampling techniques. Data obtained by using GUMP-ELAHA will aid specific treatment of GUD and better define the prevalence of each microbe among at-risk populations with a view to the eradication of chancroid and donovanosis in Australia.

Non-healing genital herpes mimicking donovanosis in an immunocompetent man.

Although atypical presentations of herpetic infection in immunocompetent individuals are common, they very rarely have the extensive, chronic and verrucous appearances seen in the immunocompromised host. We report a case of genital herpes manifesting as painless chronic non-healing genital ulcers with exuberant granulation tissue in an immunocompetent man. Owing to this morphology, the ulcers were initially mistaken for donovanosis. To the best of our knowledge, such a presentation of genital herpes in an immunocompetent individual has not been described previously.

Granuloma inguinale (donovanosis): an unusual cause of otitis media and mastoiditis in children.

Granuloma inguinale (donovanosis) is seen predominantly in adults (it rarely occurs in children) and mainly affects genital skin and mucosa. Infection occurs at other skin and mucosal sites, and hematogenous dissemination to bone also has been described. The infection responds dramatically to appropriate antibiotic treatment. We present two cases of granuloma inguinale occurring in children (8 months and 5 months of age) causing mastoiditis and external ear discharges. A temporal lobe abscess also developed in the 8-month-old child. Subsequent computed tomography scans showed marked improvement in the brain lesion after treatment. The second child had a polypoid mass in the middle ear that on biopsy showed the features of granuloma inguinale. The mother of this child had biopsy-proven granuloma inguinale of the uterine cervix. These cases indicate that granuloma inguinale can be transmitted during vaginal delivery, and careful cleansing of neonates born to infected mothers is recommended.

Growth and cultural characteristics of Calymmatobacterium granulomatis--the aetiological agent of granuloma inguinale (Donovanosis).

Granuloma inguinale is a chronic destructive granulomatous disease of the genitalia. The clinical diagnosis is often unreliable and the definitive diagnosis is based on the visualisation of 'Donovan bodies' in tissue smears or biopsy specimens. The organism implicated in its aetiology, Calymmatobacterium granulomatis, was reported to have been cultured > 30 years ago, but little is known about the organism because of its fastidious nature and the difficulty in culturing it. Twenty-two biopsy specimens from female patients with clinical and laboratory-confirmed granuloma inguinale were treated with amikacin 10 mg/L and inoculated in a monocyte co-culture system with peripheral blood mononuclear cells (PBMC) from a single donor and autologous sera. The method was subsequently modified by pretreatment of specimens with vancomycin 5 mg/L and metronidazole 10 mg/L in addition to amikacin 10 mg/L for the purpose of decontamination, pooled blood donor PBMC and by the use of heat-inactivated fetal calf serum instead of autologous serum for culture. This modified method was used to culture additional biopsy specimens and genital ulcer scrapings from female and male patients, respectively. All monocyte co-cultures were examined by a rapid Giemsa (RapiDiff) stain and by an indirect immunofluorescence test with immune sera. Representative cultures were examined by transmission electron microscopy. C. granulomatis was successfully isolated in pure culture by the monocyte co-culture system from four biopsy specimens and 14 genital ulcer scrapings. The cultured organisms were visible both intra- and extra-cellularly and were extremely pleomorphic, with characteristic single and biopolar condensation. The numbers of the organisms increased after each passage. All positive cultures showed bright fluorescence when tested with immune sera. Transmission electron microscopy of the cultured bacteria demonstrated a typical gram-negative cell wall consisting of an outer membrane, middle electron opaque layer and an inner plasma membrane. The capsule was thick and electron dense. Numerous electron dense granules were present within the cytoplasm.

[Epidemiology and pathogenesis of donovanosis]. / Reflexões sobre a epidemiologia e patogenia da donovanose.

Thirty three cases of donovanosis have been observed at the Dermatology Department of Cayenne's Hospital between 1970 and 1982, only in black people. The high frequency of rectal lesions of passive homosexual patients and the vagina contamination in women without sexual intercourse postulate for the intestinal reservoir of the bacteria, and the possibility of auto-inoculation. There is probably a cellular immune deficit. Donovanosis is often associated with other diseases, and in our practice in French Guyana, with leprosy. The skin of the genitalia is the most frequently infected, the mucosa generally resistant. The intracellular life of the "Donovan bodies" "Calymmatobacterium granulomatosis"--in the cytoplasm of large mononuclear cells seems an argument for an anergic disease.

European guideline for the management of donovanosis, 2010.

Donovanosis is a rare sexually transmitted infection now mainly seen in sporadic cases in Papua New Guinea, South Africa, India, Brazil and Australia. The causative organism is Calymmatobacterium granulomatis though a proposal has been put forward that the organism be reclassified as Klebsiella granulomatis comb. nov. The incubation period is approximately 50 days with genital papules developing into ulcers that increase in size. Four types of lesions are described - ulcerogranulomatous, hypertrophic, necrotic and sclerotic. The diagnosis is usually confirmed by microscopic identification of characteristic Donovan bodies on stained tissue smears. More recently, polymerase chain reaction (PCR) methods have been developed. The recommended treatment is azithromycin 1 g weekly until complete healing is achieved.

A colorimetric detection system for Calymmatobacterium granulomatis.

OBJECTIVE: To incorporate the first polymerase chain reaction (PCR) assay for Calymmatobacterium granulomatis into a colorimetric detection system for use in routine diagnostic laboratories. METHODS: A capture oligonucleotide specific for the Klebsiella phoE gene was covalently linked to tosyl activated magnetic beads. Biotinylated phoE PCR products obtained from 14 positive specimens from patients with donovanosis and isolates of Klebsiella pneumoniae, K rhinoscleromatis, and K ozaenae were cleaved with HaeIII for the purpose of differentiation, captured by the prepared beads, and subjected to standard EIA detection methodology. Eight samples from unrelated genital conditions underwent the same procedure. It was anticipated from the sequence data that the biotinylated fragment would be cleaved from the capture oligonucleotide target region in the three Klebsiella phoE products (that is, a negative colorimetric result) while the entire fragment of interest would remain intact in the positive C granulomatis phoE products (that is, a positive colorimetric result). RESULTS: All 14 positive specimens from patients with donovanosis gave strong colorimetric readings with this detection system. Isolates of K pneumoniae, K rhinoscleromatis, K ozaenae, and the eight specimens from unrelated genital conditions were negative. CONCLUSION: The successful development of a colorimetric detection system for C granulomatis incorporating two levels of specificity enables the molecular diagnosis of this condition to be undertaken by routine diagnostic laboratories. This should have an important role in the Australian government's campaign to eradicate donovanosis by 2003 though the test still needs to undergo trials and be validated using a larger number of samples from geographically diverse parts of the world in order to ascertain the generalisability of the methodology.

[Donovanosis. Apropos of a new case of granuloma inguinale in France]. / La Donovanose. A propos d'un nouveau cas de granulome inguinal en France.

We report a new case of granuloma inguinale in France. A 47 years old Turkish man living in France for eight years had an ulcerovegetative lesion on the corona sulcus and inner aspect of the prepuce for three months. Donovanosis was established by granulation time smears and Giemsa's stain showing Donovan bodies inside large mononuclear cells. Complete resolution occurred after treatment with ampicillin for three weeks and later doxycycline for a month. Granuloma inguinale is rare in France and must not be forgotten. Epidemiologic data, clinical manifestations and therapeutic indications are reviewed.

Genital ulcer disease in women in Durban, South Africa.

OBJECTIVE: To study the microbial aetiology of genital ulcer disease (GUD) in women. DESIGN: Microbial and clinical assessment of genital ulcers in women. SETTING: City Health sexually transmitted diseases clinic, King Edward VIII Hospital, Durban, South Africa. PARTICIPANTS: 100 Zulu women with genital ulceration who had not received antibiotics in the previous two weeks. RESULTS: Syphilis was diagnosed in 40%, genital herpes in 18%, donovanosis (granuloma inguinale) in 16%, chancroid in 14%, lymphogranuloma venereum in 7% and scabies in 2%. No recognised cause was detected in 18%. Secondary syphilis was diagnosed in 21%, primary syphilis in 16% and mixed primary and secondary syphilis in 3%. Multiple infections were detected in 13 women, of whom 12 had syphilis. Bleeding was observed from the ulcers of 59 during swab collection. Three women had HIV-1 antibodies. Neisseria gonorrhoeae was isolated from the ulcers and endocervix of two women and from the endocervix alone in nine. Generalised scabies was diagnosed in 14. CONCLUSIONS: All the major causes of GUD are prevalent in Zulu women in Durban: secondary syphilis was the commonest diagnosis. Donovanosis, which often presents late with large ulcers, and genital herpes are now significant problems. Mixed infections with coexisting syphilis are common. All women in this population with GUD should be treated for syphilis and receive oral antibiotics effective for chancroid and donovanosis.

Genital ulcer disease in men in Durban, South Africa.

OBJECTIVE: To study the microbial aetiology of genital ulcer disease (GUD) in men. DESIGN: Microbiological and clinical assessment of genital ulcers in men. SETTING: City Health sexually transmitted diseases clinic, King Edward VIII Hospital, Durban, South Africa. PARTICIPANTS: 100 Zulu men with genital ulcers who had not received antibiotics in the previous four weeks. RESULTS: Syphilis was diagnosed in 42%, chancroid in 22%, donovanosis (granuloma inguinale) in 11%, genital herpes in 10% and lymphogranuloma venereum (LGV) in 6%. No pathogens were identified in 24%. Mixed infections were detected in 14 men, in whom 13 had syphilis. Five men had HIV-1 antibodies. Neisseria gonorrhoeae was isolated from the ulcers and urethra in seven men and from the urethra alone in five. Scabies was diagnosed clinically in eight. CONCLUSIONS: All the major causes of GUD are prevalent in Zulu men in Durban. Primary syphilis was the commonest and was invariably present in mixed infections. Donovanosis was under-reported and was associated with a long delay before presentation. In this population, genital ulcers other than superficial lesions should be treated with anti-syphilitic therapy and oral antibiotics effective against chancroid and donovanosis.