Haemophilus ducreyi as a cause of skin ulcers in children from a yaws-endemic area of Papua New Guinea: a prospective cohort study

Background Skin infections with ulceration are a major health problem in countries of the south Pacifi c region. Yaws, caused by Treponema pallidum subspecies pertenue and diagnosed by the presence of skin ulcers and a reactive syphilis serology, is one major cause, but this infection can be confused clinically with ulcers due to other causative agents. We investigated T pallidum pertenue and another bacterium known to cause skin infections in the Pacifi c islands— Haemophilus ducreyi—as causes of skin ulceration in a yaws-endemic region. Additionally, we identifi ed specifi c signs and symptoms associated with these causative agents of cutaneous ulcers and compared these fi ndings with laboratory-based diagnoses. Methods We did a prospective cohort study of fi ve yaws-endemic villages (total population 3117 people) during a yaws elimination campaign in Papua New Guinea in April, 2013. We enrolled all consenting patients with chronic moist or exudative skin ulcers. We undertook a detailed dermatological assessment, syphilis serology, and PCR on lesional swabs to detect the presence of T pallidum pertenue and H ducreyi. Patients with PCR-confi rmed bacterial infections were included in a comparative analysis of demographics and clinical features. Findings Full outcome data were available for 90 people with skin ulcers. Of these patients, 17 (19%) had negative results in all molecular tests and were therefore excluded from the comparative analyses. A bacterial cause was identifi ed in 73 (81%) participants—either H ducreyi (n=42), T pallidum pertenue (yaws; n=19), or coinfection with both organisms (dual infection; n=12). The demographic characteristics of the patients infected with yaws and with H ducreyi were similar. Skin lesions in patients with yaws and in those with dual infection were larger than those in patients infected with H ducreyi (p=0·071). The lesions in patients with yaws and dual infection were more circular in shape (79% and 67%) than in those infected with H ducreyi (21%; p<0·0001); more likely to have central granulating tissue (90% and 67% vs 14%; p<0·0001); and more likely to have indurated edges (74% and 83% vs 31%; p=0·0003). The prevalence of reactive combined serology (positive T pallidum haemagglutination test and rapid plasmin reagin titre of ≥1:8) was higher in cases of yaws (63%) and dual infections (92%) than in H ducreyi infections (29%; p<0·0001). Interpretation In this yaws-endemic community, H ducreyi is an important and previously unrecognised cause of chronic skin ulceration. Reactive syphilis serology caused by latent yaws can occur in ulcers with the presence of H ducreyi alone. The introduction of PCR for ulcer surveillance could improve the accuracy of diagnosis in countries with yaws eradication campaigns

Re-emergence of yaws after single mass azithromycin treatment followed by targeted treatment: a longitudinal study

Background: Yaws is a substantial cause of chronic disfiguring ulcers in children in at least 14 countries in the tropics. WHO's newly adopted strategy for yaws eradication uses a single round of mass azithromycin treatment followed by targeted treatment programmes, and data from pilot studies have shown a short-term significant reduction of yaws. We assessed the long-term efficacy of the WHO strategy for yaws eradication. Methods: Between April 15, 2013, and Oct 24, 2016, we did a longitudinal study on a Papua New Guinea island (Lihir; 16 092 population) in which yaws was endemic. In the initial study, the participants were followed for 12 months; in this extended follow-up study, clinical, serological, and PCR surveys were continued every 6 months for 42 months. We used genotyping and travel history to identify importation events. Active yaws confirmed by PCR specific for Treponema pallidum was the primary outcome indicator. The study is registered with ClinicalTrials.gov, number NCT01955252. Findings: Mass azithromycin treatment (coverage rate of 84%) followed by targeted treatment programmes reduced the prevalence of active yaws from 1·8% to a minimum of 0·1% at 18 months (difference from baseline −1·7%, 95% CI, −1·9 to −1·4; p<0·0001), but the infection began to re-emerge after 24 months with a significant increase to 0·4% at 42 months (difference from 18 months 0·3%, 95% CI 0·1 to 0·4; p<0·0001). At each timepoint after baseline, more than 70% of the total community burden of yaws was found in individuals who had not had the mass treatment or as new infections in non-travelling residents. At months 36 and 42, five cases of active yaws, all from the same village, showed clinical failure following azithromycin treatment, with PCR-detected mutations in the 23S ribosomal RNA genes conferring resistance to azithromycin. A sustained decrease in the prevalence of high-titre latent yaws from 13·7% to <1·5% in asymptomatic children aged 1–5 years old and of genetic diversity of yaws strains from 0·139 to less than 0·046 between months 24 and 42 indicated a reduction in transmission of infection. Interpretation: The implementation of the WHO strategy did not, in the long-term, achieve elimination in a high-endemic community mainly due to the individuals who were absent at the time of mass treatment in whom yaws reactivated; repeated mass treatment might be necessary to eliminate yaws. To our knowledge, this is the first report of the emergence of azithromycin-resistant T p pertenue and spread within one village. Communities' surveillance should be strengthened to detect any possible treatment failure and biological markers of resistance

Effectiveness of single-dose azithromycin to treat latent yaws: a longitudinal comparative cohort study

BACKGROUND: Treatment of latent yaws is a crucial component of the WHO yaws eradication strategy to prevent relapse and the resulting transmission to uninfected children. We assessed the effectiveness of single-dose azithromycin to treat patients with latent yaws. METHODS: This population-based cohort study included children (age <20 years) living on Lihir Island, Papua New Guinea, with high-titre (rapid plasma reagin titre >/=1:8) latent or active yaws, between April, 2013, and May, 2015. Latent yaws was defined as lack of suspicious skin lesions or presence of ulcers negative for Treponema pallidum subsp pertenue on PCR, and active yaws was defined as ulcers positive for T pertenue on PCR. All children received one oral dose of 30 mg/kg azithromycin. The primary endpoint was serological cure, defined as a two-dilution decrease in rapid plasma reagin titre by 24 months after treatment. Treatment of latent yaws was taken to be non-inferior to that of active yaws if the lower limit of the two-sided 95% CI for the difference in rates was higher than or equal to -10%. This study is registered with ClinicalTrials.gov, number NCT01955252. FINDINGS: Of 311 participants enrolled, 273 (88%; 165 with latent yaws and 108 with active yaws) completed follow-up. The primary endpoint was achieved in 151 (92%) participants with latent yaws and 101 (94%) with active yaws (risk difference -2.0%, 95% CI -8.3 to 4.3), meeting the prespecified criteria for non-inferiority. INTERPRETATION: On the basis of decline in serological titre, oral single-dose azithromycin was effective in participants with latent yaws. This finding supports the WHO strategy for the eradication of yaws based on mass administration of the entire endemic community irrespective of clinical status. FUNDING: Newcrest Mining Limited and ISDIN laboratories

Evaluation of Azithromycin Resistance in Treponema pallidum Specimens From Madagascar

Abstract:Treponema pallidum resistance to azithromycin has been documented in the US, Canada, and Ireland. We found no evidence of resistance to azithromycin in specimens from 141 patients with syphilitic lesions in Madagascar suggesting resistance is geographically isolated and supporting use of azithromycin as alternative treatment for early syphilis in Madagascar

Streptococcus pyogenes Is Associated with Idiopathic Cutaneous Ulcers in Children on a Yaws-Endemic Island

Exudative cutaneous ulcers (CU) in yaws-endemic areas are associated with Treponema pallidum subsp. pertenue (TP) and Haemophilus ducreyi (HD), but one-third of CU cases are idiopathic (IU). Using mass drug administration (MDA) of azithromycin, a yaws eradication campaign on Lihir Island in Papua New Guinea reduced but failed to eradicate yaws; IU rates remained constant throughout the campaign. To identify potential etiologies of IU, we obtained swabs of CU lesions (n = 279) and of the skin of asymptomatic controls (AC; n = 233) from the Lihir Island cohort and characterized their microbiomes using a metagenomics approach. CU bacterial communities were less diverse than those of the AC. Using real-time multiplex PCR with pathogen-specific primers, we separated CU specimens into HD-positive (HD+), TP+, HD+TP+, and IU groups. Each CU subgroup formed a distinct bacterial community, defined by the species detected and/or the relative abundances of species within each group. Streptococcus pyogenes was the most abundant organism in IU (22.65%) and was enriched in IU compared to other ulcer groups. Follow-up samples (n = 31) were obtained from nonhealed ulcers; the average relative abundance of S. pyogenes was 30.11% in not improved ulcers and 0.88% in improved ulcers, suggesting that S. pyogenes in the not improved ulcers may be azithromycin resistant. Catonella morbi was enriched in IU that lacked S. pyogenes As some S. pyogenes and TP strains are macrolide resistant, penicillin may be the drug of choice for CU azithromycin treatment failures. Our study will aid in the design of diagnostic tests and selective therapies for CU.IMPORTANCE Cutaneous ulcers (CU) affect approximately 100,000 children in the tropics each year. While two-thirds of CU are caused by Treponema pallidum subspecies pertenue and Haemophilus ducreyi, the cause(s) of the remaining one-third is unknown. Given the failure of mass drug administration of azithromycin to eradicate CU, the World Health Organization recently proposed an integrated disease management strategy to control CU. Success of this strategy requires determining the unknown cause(s) of CU. By using 16S rRNA gene sequencing of swabs obtained from CU and the skin of asymptomatic children, we identified another possible cause of skin ulcers, Streptococcus pyogenes Although S. pyogenes is known to cause impetigo and cellulitis, this is the first report implicating the organism as a causal agent of CU. Inclusion of S. pyogenes into the integrated disease management plan will improve diagnostic testing and treatment of this painful and debilitating disease of children and strengthen elimination efforts

Enhanced Molecular Typing of Treponema pallidum : Geographical Distribution of Strain Types and Association with Neurosyphilis

Strain typing is a tool for determining diversity and epidemiology of infections

Yaws re-emergence and bacterial drug resistance selection after mass administration of azithromycin: a genomic epidemiology investigation.

BACKGROUND: In a longitudinal study assessing the WHO strategy for yaws eradication using mass azithromycin treatment, we observed resurgence of yaws cases with dominance of a single JG8 sequence type and emergence of azithromycin-resistant Treponema pallidum subspecies pertenue (T p pertenue). Here, we analyse genomic changes in the bacterial population using samples collected during the study. METHODS: We did whole bacterial genome sequencing directly on DNA extracted from 37 skin lesion swabs collected from patients on Lihir Island, Papua New Guinea, between April 1, 2013, and Nov 1, 2016. We produced phylogenies and correlated these with spatiotemporal information to investigate the source of new cases and the emergence of five macrolide-resistant cases. We used deep amplicon sequencing of surveillance samples to assess the presence of minority macrolide-resistant populations. FINDINGS: We recovered 20 whole T p pertenue genomes, and phylogenetic analysis showed that the re-emerging JG8 sequence type was composed of three bacterial sublineages characterised by distinct spatiotemporal patterns. Of five patients with resistant T p pertenue, all epidemiologically linked, we recovered genomes from three and found no variants. Deep sequencing showed that before treatment, the index patient had fixed macrolide-sensitive T p pertenue, whereas the post-treatment sample had a fixed resistant genotype, as did three of four contact cases. INTERPRETATION: In this study, re-emergence of yaws cases was polyphyletic, indicating multiple epidemiological sources. However, given the genomic and epidemiological linkage of resistant cases and the rarity of resistance alleles in the general population, azithromycin resistance is likely to have evolved only once in this study, followed by onward dissemination. FUNDING: Wellcome and Provincial Deputation of Barcelona

Treponema pallidum subsp. pallidum DNA and RNA in Semen of a Syphilis Patient Without Genital or Anal Lesions

Treponema pallidum subsp. pallidum DNA and RNA were detected in a semen specimen of a syphilis patient with no genital or anal sores and no clinically evident orchitis. No nucleic acids were found in a urine sample of the same patient collected immediately before the semen sample. Exposure to the syphilis agent through semen could account for transmission episodes in the absence of direct contact with a syphilitic sore

Development of a Multilocus Sequence Typing (MLST) scheme for Treponema pallidum subsp. pertenue: Application to yaws in Lihir Island, Papua New Guinea.

Yaws is a neglected tropical disease, caused by Treponema pallidum subsp. pertenue. The disease causes chronic lesions, primarily in young children living in remote villages in tropical climates. As part of a global yaws eradication campaign initiated by the World Health Organization, we sought to develop and evaluate a molecular typing method to distinguish different strains of T. pallidum subsp. pertenue for disease control and epidemiological purposes.Published genome sequences of strains of T. pallidum subsp. pertenue and pallidum were compared to identify polymorphic genetic loci among the strains. DNA from a number of existing historical Treponema isolates, as well as a subset of samples from yaws patients collected in Lihir Island, Papua New Guinea, were analyzed using these targets. From these data, three genes (tp0548, tp0136 and tp0326) were ultimately selected to give a high discriminating capability among the T. pallidum subsp. pertenue samples tested. Intragenic regions of these three target genes were then selected to enhance the discriminating capability of the typing scheme using short readily amplifiable loci. This 3-gene multilocus sequence typing (MLST) method was applied to existing historical human yaws strains, the Fribourg-Blanc simian isolate, and DNA from 194 lesion swabs from yaws patients on Lihir Island, Papua New Guinea. Among all samples tested, fourteen molecular types were identified, seven of which were found in patient samples and seven among historical isolates or DNA. Three types (JG8, TD6, and SE7) were predominant on Lihir Island.This MLST approach allows molecular typing and differentiation of yaws strains. This method could be a useful tool to complement epidemiological studies in regions where T. pallidum subsp. pertenue is prevalent with the overall goals of improving our understanding of yaws transmission dynamics and helping the yaws eradication campaign to succeed