Project Grant R01AI160433

CHARACTERISATION OF DRUG RESISTANCE IN FIELD-COLLECTED SCHISTOSOMES - LARGE-SCALE TREATMENT PROGRAMS WITH PRAZIQUANTEL (PZQ) MONOTHERAPY ARE THE MAINSTAY OF EFFORTS TO CONTROL AND ELIMINATE SCHISTOSOMIASIS. THE POTENTIAL FOR DRUG RESISTANCE IN SCHISTOSOMES IS A CRITICAL AREA OF RESEARCH AS WE SCALE UP TREATMENT COVERAGE. IN THE LABORATORY, PZQ RESISTANCE (PZQ-R) IS EASILY SELECTED, RESULTS IN A >368-FOLD DIFFERENCE IN DRUG RESPONSE, AND GENETIC ANALYSES UNAMBIGUOUSLY IDENTIFY A REGION OF CHR. 3 CONTAINING A TRANSIENT RECEPTOR POTENTIAL (TRP) CHANNEL. IN WESTERN KENYA, APPROXIMATELY 30% OF SCHISTOSOME-INFECTED PATIENTS ARE EGG-POSITIVE FOLLOWING TREATMENT WITH PZQ, AND IN A RECENT STUDY (SCORE) WE OBSERVED SEVERAL "HOTSPOT" VILLAGES THAT FAILED TO LOWER INFECTION INTENSITIES AND PREVALENCE, DESPITE MULTIPLE ANNUAL MASS TREATMENTS WITH PZQ. ONE HYPOTHESIS IS THAT PZQ-R CONTRIBUTES TO THE OBSERVED FAILURE OF MASS PZQ TREATMENT TO LOWER INFECTION. IN AIM 1 WE WILL ASK WHETHER THE PERSISTENCE OF INFECTION IN "HOTSPOT" VILLAGES CAN BE EXPLAINED BY RESISTANCE. WE HAVE ESTABLISHED A NOVEL PLATFORM FOR TESTING FOR THE EXTENT OF PZQ-R IN INDIVIDUAL ADULT SCHISTOSOMES FROM A FIELD SETTING. TO DO THIS WE HAVE ESTABLISHED A LARGE SNAIL BREEDING COLONY, AND HAMSTER BREEDING FACILITY. WE CAN THEREFORE GENERATE LARGE POPULATIONS OF FIELD-DERIVED ADULT WORMS, BY (I) HARVESTING S. MANSONI EGGS FROM INFECTED PATIENTS, (II) INFECTING SNAILS WITH MIRACIDIA, (III) INFECTING HAMSTERS WITH RELEASED CERCARIAE, AND (IV) PERFUSING ADULT WORMS FROM HAMSTERS. WE WILL SCREEN THE DRUG RESPONSE OF INDIVIDUAL S. MANSONI WORMS MAINTAINED ON 96-WELL PLATES AND EXPOSED TO PZQ USING A SIMPLE L-LACTATE ASSAY. USING THIS APPROACH, WE WILL DIRECTLY COMPARE RESISTANCE STATUS OF PARASITES FROM HOTSPOT AND NON-HOTSPOT VILLAGES. WE WILL ALSO COMPARE POOLS OF PZQ-R AND PZQ-S PARASITES ISOLATED FROM THE FIELD TO DETERMINE THE GENOME REGIONS THAT UNDERLIE THE DIFFERENCES OBSERVED, AND TO TEST THE HYPOTHESIS THAT GENETIC VARIATION IN THE TRP CHANNEL ON CHR. 3 UNDERLIES PZQ RESISTANCE IN THE FIELD. IN AIM 2 WE WILL EXPLOIT OUR UNIQUE PLATFORM TO EXAMINE GENETIC VARIATION FOR RESISTANCE TO NEW SCHISTOSOME DRUGS. A SUITE OF COMPOUNDS SHOWING STRONG ACTIVITY AGAINST SCHISTOSOMES IS NOW AVAILABLE. RECENT FIELD STUDIES HAVE SHOWN HIGH LEVELS OF NATURALLY OCCURRING RESISTANCE TO OXAMNIQUINE IN EAST AFRICA WHERE THIS DRUG HAS BEEN MINIMALLY USED, AND COMPARABLE WORK WITH THE FREE-LIVING NEMATODE C. ELEGANS SHOWS NATURALLY OCCURRING RESISTANCE TO A RANGE OF ANTHELMINTIC COMPOUNDS. WE WILL CRITICALLY TEST THE HYPOTHESIS THAT "STANDING VARIATION" FOR RESISTANCE IS COMMON AGAINST SCHISTOSOME DRUGS UNDER DEVELOPMENT. WE WILL THEN COMPARE INDIVIDUAL WORMS AT THE EXTREMES OF THE DRUG RESPONSE SPECTRUM TO IDENTIFY THE GENETIC BASIS OF RESISTANCE. THE KNOWLEDGE GAINED WILL BE CRITICAL TO ON- GOING SCHISTOSOMIASIS ELIMINATION EFFORTS THROUGH MASS PZQ TREATMENT AND WILL PROVIDE VALUABLE INFORMATION FOR THE DEVELOPMENT OF NEW ANTI-SCHISTOSOME DRUGS. THE PROJECT WILL ALSO STIMULATE DEVELOPMENT OF RESEARCH CAPACITY IN GENOME SEQUENCING, COMPUTATIONAL BIOLOGY AND ANTHELMINTHIC PHARMACOLOGY IN KENYA.