Fig 1.
Steps for designing and conducting the larvae-parasites interaction experiment.
A) Confining the larval range of motion within a restricted area using physical barriers to increase the likelihood of encountering the parasite; B) Direct interaction between larvae and parasites in a controlled environment for a specified time period (24 hours) under standard laboratory conditions; C) Determining the number of viable cells in study groups using the MTT assay, where purple color indicates formazan crystals produced by metabolically active viable cells.
Fig 2.
Pre-oral (A-C) and oral (D-E) digestion of Leishmania major by Lucilia sericata larvae.
A: Deformation of L. major induced by larval excretion/secretion products; B: lysis of deformed parasites; C: comprehensive lysis of parasites within the interactive environment; D: micro-dissection of larvae exposed to the parasite (middle: third instar spiracles; right: salivary glands connected to the digestive canal); E: complete alimentary canal; F: gut tissues of the larvae stained with Giemsa, showing the presence of some bacteria but no visible L. major.
Fig 3.
Comparison of the average survival percentages of parasites across different study groups at consecutive 30-minute intervals to determine the optimal duration for larval-parasite interactions.
A one-way analysis of variance (ANOVA) test, corrected using a Mixed Model ANOVA, was applied at a 95% confidence level to assess variations among the groups (Larvae/Parasites/Liver, Larvae/Parasites/Spleen, and Larvae/Parasites/RPMI).
Fig 4.
Comparison of the mean percentage of parasite survival across different study groups to identify the appropriate larval-parasite interaction environment.
A one-way analysis of variance (ANOVA) test, corrected using Brown-Forsythe, was conducted at a 95% confidence level. The analysis compared three interactive groups (larvae/parasite/liver, larva/parasite/spleen, larva/parasite/RPMI) to the control group (parasite only).
Fig 5.
Comparison of the mean percentage of viable parasites between the intervention group (L. sericata and L. major in RPMI 1640 supplemented with 10% serum) and the control group (L. major in RPMI 1640 supplemented with 10% serum) outside the larval body, following a four-hour interaction.
The viability of parasites was assessed using the trypan blue exclusion method.
Fig 6.
Comparison of the metabolically active cells among the treatment group (L. sericata and L. major in RPMI 1640 medium supplemented with 10% serum), negative control group (L. sericata in RPMI 1640 medium with 10% serum), and positive control group (L. major in RPMI 1640 medium with 10% serum) outside the larval body, following a four-hour interaction.
The cell activities were assessed using the MTT assay.
Fig 7.
Species-specific nested PCR of Leishmania major, using the ITS2 and 5.8S gene.
Lanes: 1, positive control (~213 bp); M, 100-bp ladder (Fermentas, USA); 2, positive control (~166 bp); 3, negative control (larval body); 4 and 5 (larvae exposed to L. major).
Table 1.
A preliminary guideline for implementing maggot debridement therapy in patients with zoonotic cutaneous leishmaniasis, outlining patient selection criteria, larval preparation, application methods, monitoring procedures, and integration with conventional treatments, while specifying variations based on lesion type, size, depth, and patient health profile.