Prevalence of chloroquine resistance in urban versus rural communities near Kumasi, Ghana

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The fight against malaria, despite recent strides, is still not fully won. This fact can be attributed to ineffective diagnostic tools for rapid and accurate diagnosis, use of older drugs against which the parasite has developed resistance, and failure to rigorously practice prophylactic measures. Chloroquine was once the preferred drug to treat malaria in Ghana; about 8 years ago the WHO recommended a switch to artemisinin-based combination therapy (ACTs). Thus, sensitivity to chloroquine might be expected to return. One hypothesis is that the sensitivity of Plasmodium falciparum to chloroquine might be higher in rural areas with less access to chloroquine, as compared to urban centers with easy access to it as well as other treatment options that work on the same mechanism. An alternative hypothesis is that chloroquine circulates more widely in the rural sector among licensed chemical sellers (LCS) and informal vendor networks. This study aimed: A) to determine the accuracy of routine thick smear diagnosis under usual working conditions as compared with nested PCR and B) to ascertain the prevalence of chloroquine resistance in central, urban Kumasi (KATH, 100 samples), peri-urban Kumasi (KNUST-Hospital, 108 samples) and a more rural area location (Nkwantakese and Pampatea, about 20 km north of Kumasi, 203 samples). A total of 403 thick smears and blood spots with an initial malaria diagnosis made by trained microscopists were collected in the three locations from December, 2012 to January, 2013. Genomic DNA was extracted from the spots and a segment of the Plasmodium falciparum chloroquine resistance transporter gene (pfcrt) was amplified. The amplicons were digested with the restriction enzyme Xap1 to determine the presence of the K76T mutation in pfcrt gene of malaria parasites. Out of 403 samples, a total of 27 (7%) were diagnosed positive by microscopy as compared to 39 (10%) by PCR, with 376 (93%) and 364 (91%) identified negative by microscopy and PCR, respectively. Only 13 smear positive samples were confirmed by PCR and 26 smears were falsely diagnosed negative by microscopy. Although there were far too few positive malaria tests for statistical significance, chloroquine resistance in the P. falciparum population did not decrease as one moved from the urban centres (KATH and KNUST-Hospital) towards the more rural setting (Nkwantakese and Pampatea). A resistance of 87% (20 out 23) was observed for the urban and peri-urban centres together, whereas, a higher prevalence of 94% (15 out of 16) was noted in the rural areas. This preliminary result suggests that chloroquine use might actually be higher in the villages than the urban area. More work is required to validate this very preliminary trend.
This thesis is presented to the Department of Biochemistry & Biotechnology in partial fulfillment for the requirement for the Award of Master of Science in Biotechnology.