Xenodiagnosis to evaluate the infectiousness of humans to sandflies in an area endemic for visceral leishmaniasis in Bihar, India: a transmission-dynamics study

Abstract

Summary Background Visceral leishmaniasis, also known on the Indian subcontinent as kala-azar, is a fatal form of leishmaniasis caused by the protozoan parasite Leishmania donovani and transmitted by the bites of the vector sandfly Phlebotomus argentipes . To achieve and sustain elimination of visceral leishmaniasis, the transmission potential of individuals exposed to L donovani from across the infection spectrum needs to be elucidated. The aim of this study was to evaluate the relative infectiousness to the sandfly vector of patients with visceral leishmaniasis or post-kala-azar dermal leishmaniasis, before and after treatment, and individuals with asymptomatic infection. Methods In this prospective xenodiagnosis study done in Muzaffarpur district of Bihar, India, we included patients with clinically confirmed active visceral leishmaniasis or post-kala-azar dermal leishmaniasis who presented to the Kala-Azar Medical Research Center. These participants received treatment for L donovani infection. We also included asymptomatic individuals identified through a serosurvey of 17 254 people living in 26 high-transmission clusters. Eligible participants were aged 12–64 years, were HIV negative, and had clinically or serologically confirmed L donovani infection. During xenodiagnosis, the forearms or lower legs of participants were exposed to 30–35 female P argentipes sandflies for 30 min. Blood-engorged flies were held in an environmental cabinet at 28°C and 85% humidity for 60–72 h, after which flies were dissected and evaluated for L donovani infection by microscopy and quantitative PCR (qPCR). The primary endpoint was the proportion of participants with visceral leishmaniasis or post-kala-azar dermal leishmaniasis, before and after treatment, as well as asymptomatic individuals, who were infectious to sandflies, with a participant considered infectious if promastigotes were observed in one or more individual flies by microscopy, or if one or more of the pools of flies tested positive by qPCR. Findings Between July 12, 2016, and March 19, 2019, we recruited 287 individuals, including 77 with active visceral leishmaniasis, 26 with post-kala-azar dermal leishmaniasis, and 184 with asymptomatic infection. Of the patients with active visceral leishmaniasis, 42 (55%) were deemed infectious to sandflies by microscopy and 60 (78%) by qPCR before treatment. No patient with visceral leishmaniasis was found to be infectious by microscopy at 30 days after treatment, although six (8%) were still positive by qPCR. Before treatment, 11 (42%) of 26 patients with post-kala-azar dermal leishmaniasis were deemed infectious to sandflies by microscopy and 23 (88%) by qPCR. Of 23 patients who were available for xenodiagnosis after treatment, one remained infectious ...

Towards a Sustainable Vector-Control Strategy in the Post Kala-Azar Elimination Era

“…PDKL, in particular, is considered the major interepidemic reservoir because such patients can have skin lesions for years without being systemically ill (Zijlstra et al, 2017). Recent studies of kalaazar and PKDL patients confirm historical data on their infectiousness (Molina et al, 2017;Mondal et al, 2019;Singh et al, 2021). Overall, 55.3% of enrolled PKDL patients had a positive direct xenodiagnosis result after sand flies fed on patients' lesions; the proportion was highest for those with nodular lesions but macular PKDL patients also infected sand flies at a substantial rate (Mondal et al, 2019).…”

“…The model assumes that, in addition to VL and PKDL patients, asymptomatic carriers play a role in VL transmission, though with a much lower rate of infectiousness (Le Rutte et al, 2017). Studies that provide quantitative data needed to establish the potential of asymptomatic individuals as reservoirs for parasite transmission to vector populations are in progress (Singh et al, 2014;Tiwary et al, 2017;Singh et al, 2021). Non-human reservoir hosts have been sought but never proven to play a role in leishmanial transmission in the ISC (Singh et al, 2020).…”

Assessing L. donovani Skin Parasite Load: A Proof of Concept Study of a Microbiopsy Device in an Indian Setting

“…rK39 ELISA was performed as described previously, using the same cut-off titer of ≥ 14 percentage point positivity (pp) to define a positive test as identified in the paper of Hasker et al (mean value for a healthy non-endemic control plus three standard deviations) (Hasker et al, 2013). Whole Blood Analysis (WBA) was performed as described elsewhere (Singh et al, 2021). In short, antigen-specific IFN-g levels produced in response to soluble Leishmania antigen (SLA) stimulation were determined by subtracting background levels measured in the non-stimulated samples.…”

“…In short, antigen-specific IFN-g levels produced in response to soluble Leishmania antigen (SLA) stimulation were determined by subtracting background levels measured in the non-stimulated samples. The result was considered positive when the IFN-g concentration in the antigen wells was 52.28 pg/ml or higher (Singh et al, 2021).…”

“…While the potential of diseased individuals to transmit Leishmania parasites to sand flies is firmly established (Swaminath et al, 1942;Costa et al, 2000;Vergel et al, 2006;Molina et al, 2017;Mondal et al, 2019), human xenodiagnosis studies have never been able to illustrate transmission from asymptomatic individuals -although it needs to be noted that the number of studies on this topic is limited (Costa et al, 2000;Singh et al, 2021). There are, however, some signs pointing in this direction, such as the presence of live L. donovani amastigotes in the blood of asymptomatically infected individuals in a VL endemic Indian village (Sharma et al, 2000).…”