The mass use of deltamethrin collars to control and prevent canine visceral leishmaniasis: A field effectiveness study in a highly endemic area.

BACKGROUND: Visceral leishmaniasis (VL) is a zoonosis of great importance. Limitations in current VL control measures compromise efficacy, indicating the need to implement new strategies. The aim of this study was to evaluate the effectiveness of the mass use of deltamethrin-impregnated collars in dogs as a public health measure to control and prevent canine visceral leishmaniasis (CVL). METHODOLOGY: An interventional study was implemented in two endemic areas in the district of Monte Gordo (Bahia-Brazil): an intervention area, in which VL seronegative dogs were collared, and a control area in which only conventional CVL control measures were applied. At baseline, seropositive dogs were removed and seronegative dogs were included. Dogs were then reevaluated every 7-8 months for almost two years. At each time point, dogs in the intervention area that remained seronegative received new collars and newly identified seronegative dogs were included and collared. The local zoonosis control authorities were notified of any dogs that tested seropositive in both areas, which were subsequently marked for euthanasia as mandated by the Brazilian Ministry of Health. PRINCIPAL FINDINGS: In the first serological survey, seroprevalence was similar in both areas. At the second evaluation, significant reductions in seroprevalence were seen in both areas, while seroprevalence in the intervention area reduced to 6.0% during the final evaluation versus an increase of 11.0% in the control area. This significant increase and the estimated relative risk (RR = 0.55) indicated protection against CVL in the intervention area. Although CVL incidence did not differ significantly between the areas, an increased tendency was observed in the control area, which could be due to low seroconversion rates throughout the study or a high loss to follow-up. CONCLUSIONS/SIGNIFICANCE: Although our evaluation of the effectiveness of deltamethrin-impregnated collars as a community-wide public health control measure was inconclusive, this measure likely provides protection over time. In endemic areas of Brazil, this strategy represents an operational challenge for local zoonosis control authorities, indicating the need for adjustments, including improved collar design.

A ready-to-use duplex qPCR to detect Leishmania infantum DNA in naturally infected dogs.

Canine visceral leishmaniasis (CVL) is a systemic disease caused by Leishmania infantum. A precise CVL diagnosis would allow for a faster and more specific treatment. Quantitative PCR (qPCR) is a sensitive and specific technique that can diagnose CVL and also monitor parasite load in the animal during the course of the infection or treatment. The aim of this study was to develop a ready-to-use (gelified and freezer-free) duplex qPCR for the identification of infected animals. We combined a new qPCR protocol that detects the canine 18S rRNA gene with an existing protocol for L. infantum kDNA detection, creating a duplex qPCR. This duplex method was then developed into a ready-to-use format. The performance of the duplex and singleplex reactions were compared in the traditional format (liquid and freezer-stored). Furthermore, the duplex qPCR performance was compared between the ready-to-use and traditional formats. The singleplex and new duplex qPCR exhibited the same detection limit in the traditional format (0.1 parasites/reaction). The ready-to-use format showed a detection limit of 1 parasite/reaction without affecting the reaction efficiency. The performance of the new qPCR protocol in the two formats was assessed using canine tissue samples from 82 dogs in an endemic CVL area that were previously characterized by standard serological and parasitological protocols. Splenic aspirates provided a higher rate of positivity (92.9%) followed by skin (50%) and blood (35.7%). The reported detection limits were observed for all tissues studied. Our results show that the amplification of L. infantum kDNA and canine DNA in a single tube, using either the traditional or ready-to-use format, exhibited the same diagnostic performance as amplification of the parasite kDNA alone. The detection of the host gene strengthens the qPCR results by confirming the presence and quality of DNA in the samples and the absence of polymerase inhibitors. The ready-to-use duplex qPCR format has many advantages. By joining two qPCR protocols into one, more results can be obtained in the same amount of time with reduced costs and embedded quality control. Reagents are preloaded and stored on the plate, reducing the operator's hands-on time to set up a reaction, as well as decreasing manipulation steps, which reduces the risk of mistakes or contamination. Thus, the ready-to-use duplex format turns qPCR into a robust, easy-to-use tool, which could help increase the availability of qPCR for CVL diagnosis.