Background Tungiasis is a neglected tropical skin disease caused by the sand flea Tunga penetrans. Female fleas penetrate the skin, particularly at the feet, and cause severe inflammation. This study aimed to characterize disease burden in two highly affected regions in Kenya, to test the use of thermography to detect tungiasis-associated inflammation and to create a new two-level classification of disease severity suitable for mapping, targeting, and monitoring interventions. Methods From February 2020 to April 2021, 3532 pupils age 8–14 years were quasi-randomly selected in 35 public primary schools and examined for tungiasis and associated symptoms. Of the infected pupils, 266 were quasi-randomly selected and their households visited, where an additional 1138 family members were examined. Inflammation was assessed using infra-red thermography. A Clinical score was created combining the number of locations on the feet with acute and chronic symptoms and infra-red hotspots. Results The overall prevalence of tungiasis among all the school pupils who were randomly selected during survey rounds 1 and 3 was 9.3% [95% confidence interval (CI): 8.4–10.3]. Based on mixed effects logistic models, the odds of infection with tungiasis among school pupils was three times higher in Kwale (coastal Kenya) than in Siaya [western Kenya; adjusted odds ratio (aOR) = 0.36, 95% CI: 0.18–0.74]; three times higher in males than in females (aOR = 3.0, 95% CI: 2.32–3.91) and three times lower among pupils sleeping in a house with a concrete floor (aOR = 0.32, 95% CI: 0.24–0.44). The odds of finding an infected person among the household population during surveys before the COVID-19 pandemic was a third (aOR = 0.32, 95% CI: 0.19–0.53) of that when schools were closed due to COVID-19 restrictions and approximately half (aOR = 0.44, 95% CI: 0.29–0.68) in surveys done after school re-opening (round 3). Infection intensity was positively correlated with inflammation as measured by thermography (Spearman’s rho = 0.68, P < 0.001) and with the clinical score (rho = 0.86, P < 0.001). Based on the two-level classification, severe cases were associated with a threefold higher level of pain (OR = 2.99, 95% CI: 2.02–4.43) and itching (OR = 3.31, 95% CI: 2.24–4.89) than mild cases. Conclusions Thermography was a valuable addition for assessing morbidity and the proposed two-level classification of disease severity clearly separated patients with mild and severe impacts. The burden of tungiasis was considerably higher in households surveyed during COVID-19 restrictions suggesting underlying risks are found in the home environment more than in school.

Tungiasis is a neglected tropical disease caused by skin-penetrating female Tunga penetrans fleas. Although tungiasis causes severe health problems, its ecology is poorly understood and morphological descriptions of the larvae are unavailable. To identify T. penetrans immature stages and sites where they develop, diagnostic PCRs are required. However, flea larvae feed on soil organic matter rich in PCR inhibitors. Here, three DNA preparation methods, including a soil DNA kit that removes inhibitors, a simple ammonium acetate precipitation approach (AmAcet) and a crude lysate of larvae (CL), were combined with amplification by the highly processive FIREPol® Taq or the inhibitor-resistant Phusion® polymerase. Independent of the polymerase used, the frequency of successful amplification, Cq values and PCR efficacies for the low-cost CL and AmAcet methods were superior to the commercial kit for amplification of a 278 bp partial internal transcribed spacer-2 (ITS-2) and a 730 bp pan-Siphonaptera cytochrome oxidase II PCR. For the CL method combined with Phusion® polymerase, the costs were approximately 20-fold lower than for the methods based on the soil DNA kit, which is a considerable advantage in resource-poor settings. The ITS-2 PCR did not amplify Ctenocephalides felis genomic or Tunga trimammilata ITS-2 plasmid DNA, meaning it can be used to specifically identify T. penetrans.

Tungiasis is a neglected tropical disease caused by skin-penetrating female Tunga penetrans fleas. Although tungiasis causes severe health problems, its ecology is poorly understood and morphological descriptions of larvae are unavailable. To identify T. penetrans immature stages and sites where they develop, diagnostic PCRs are required. However, flea larvae feed on soil organic matter rich in PCR inhibitors. Here, three DNA preparation methods, a soil DNA kit removing inhibitors, a simple ammonium acetate precipitation approach (AmAcet) and a crude lysate of larvae (CL), were combined with amplification by the highly processive FIREPol&reg; Taq or the inhibitor-resistant Phusion&reg; polymerase. Independent of the polymerase used, frequency of successful amplification, Cq values and PCR efficacies for the low-cost CL and AmAcet methods were superior to the commercial kit for amplification of a 278 bp partial internal transcribed spacer-2 (ITS-2) and a 730 bp pan-Siphonaptera cytochrome oxidase I PCR. For the CL method combined with Phusion&reg; polymerase, costs were approximately 20-fold lower than for methods based on the soil DNA kit, which is a considerable advantage in resource-poor settings. The ITS-2 PCR did not amplify Ctenocephalides felis genomic or Tunga trimammilata ITS-2 plasmid DNA allowing it to be used to specifically identify T. penetrans.

Background: Tungiasis is a common but extremely neglected tropical skin disease caused by the sand flea Tunga penetrans. Female sand fleas penetrate the skin, particularly at the feet, and cause severe inflammation with pain and itching, acute and chronic morbidity. This study aimed to characterize disease burden in two highly affected regions in Kenya, to test the use of simple thermography to detect tungiasis-associated inflammation and to create a new two-level classification of disease severity that may be used for mapping, targeting, and monitoring interventions in future. Methods: From February 2020 to April 2021, 3,532 pupils age 8-14 years were randomly selected in 35 public primary schools and examined for tungiasis and associated pathology. Of the infected pupils, 266 were randomly selected and their households visited. An additional 1,138 family members were examined. Infra-red thermography was used to assess inflammation. An all-pathology score was created combining the number of locations on the feet with acute and chronic pathologies and infra-red hotspots. Results: Based on multilevel mixed effects logistic models, the odds of infection with tungiasis among school pupils was three times as high in Kwale than in Siaya (adjusted odds ratio (AOR)0.28, 95% CI 0.1-0.6); three times higher in males than in females (AOR2.9, 95%CI 2.2-3.7) and three times lower among pupils sleeping in a house with a concrete floor (AOR0.32,95%CI 0.24-0.44). The odds of finding an infected person among the household population during active school terms was approximately a half of that when schools were closed due to COVID-19 (AOR0.44, 95% CI 0.3-0.7). Infection intensity was positively correlated with inflammation (Spearman’s rho =0.68, p<0.001) and all-pathology score (rho 0.86, p<0.001). Based on the two-level classification, severe cases were associated with a threefold higher level of pain (OR2.99, 95% CI 2-4.4) and itching (OR3.31, 95% CI 2.2-4.9) than mild cases. Conclusions: Thermography was a valuable addition for assessing morbidity and the proposed two-level classification of disease severity clearly separated patients with mild and severe impacts. The burden of tungiasis severely increased during COVID-19 restrictions and reinforced that underlying risks are found in the home environment more than in school.