Abstract
Background: Enterobacter cloacae subsp. dissolvens bacterium is a known commensal of the gut microflora of Phlebotomus papatasi, the main vector for zoonotic cutaneous Leishmaniasis, and nominated for paratransgenesis in sand flies. In this study, we evaluated dynamics and fitness costs of engineered E. cloacae for its potential to serve as a 'Trojan Horse' in P. papatasi. Methods: The engineered strain of E. cloacae transformed with a constantly active expressed red fluorescent protein plus defensin (EC-DR) plasmid and was fed to sand fly colonies via larval food to larvae. A wild type the bacterium (EC-WT) and intact food were used as controls. Fitness characters as well as dynamics of the EC-DR at various development stages of sand fly larvae were tested by plating homogenized specimens and counting fluorescent expressing colonies on the Tet-BHI agar medium. Results: Enterobacter cloacaeDR producing red fluorescent protein could be isolated from the larvae gut after 36 days when the bacteria were added once in larval pots. The EC-DR with multiple applications had no negative effect on emergence time of instar II larvae, pupae, and adults but increased slightly mortality rate of P. papatasi larvae. The experiment also confirmed lack or weak trans-stadial transmission of E. cloacae DR in P. papatasi. It has minimal fitness cost on P. papatasifeeding behavior and survival. Conclusion: Results of this study showed that E. cloacae DR is suitable for paratransgenesis of P. papatasi at only adult stage because it did not transmit transstadially.
Highlights
Leishmaniasis is one of the ten major tropical diseases that the World Health Organization (WHO) has been recommending and supporting research on their various aspects
At larval stage, when the engineered bacteria was added to larval pot only one time in the first instar larvae, except for two positive larval specimens with a few colonies, all of the samples including different larval stages (I-IV), pupa and newly emerged adults were negative for E. cloacae D-R
When the bacteria were added every 3.5 days to pots, E. cloacae-RD was found in the guts of P. papatasi larvae (Figures 1 and 2), it inadequately transmitted transstadially from larvae to adult stage
Summary
Leishmaniasis is one of the ten major tropical diseases that the World Health Organization (WHO) has been recommending and supporting research on their various aspects. The leishmaniases are complex diseases of (sub) tropical regions of the world caused by different Leishmania species of Trypanosomatidae and spread by sand flies. The cutaneous forms of the disease include the cutaneous (CL), diffuse (DCL), and mucocutaneous (MCL) leishmaniasis, but infections remain asymptomatic in many cases [1,2,3]. Leishmaniasis is endemic in 98 countries, and a total of 350 million people are at risk of developing the disease, including 21 countries in the new world and 79 countries in the ancient world. A total of 12 million people suffer from this disease, and occur between 1 and 2.1 million new cases of leishmaniasis in the world [4]. We evaluated dynamics and fitness costs of engineered E. cloacae for its potential to serve as a 'Trojan Horse' in P. papatasi
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