|Budget Amount *help
¥3,500,000 (Direct Cost : ¥3,500,000)
Fiscal Year 2005 : ¥600,000 (Direct Cost : ¥600,000)
Fiscal Year 2004 : ¥1,300,000 (Direct Cost : ¥1,300,000)
Fiscal Year 2003 : ¥1,600,000 (Direct Cost : ¥1,600,000)
We had reported during the period from 2003 to 2004 that we determined all the base sequences of cytochrome b (cyt b) of the causative Leishmania parasites and concluded that it was possible to identify and classify the causative Leishmania parasites according to the phylogenetic tree designed from the obtained cyt b genes. The purpose of the study in 2005 was to analyze cyt b genes of the causative Leismania parasites infected in the tissue specimens of the patients in Islamic Republic of Pakistan and to investigate the kinds, frequencies, and geographic distributions of the causative parasites as well as their relationship to the clinical symptoms.
During the two-year period from January 2003 to December 2004, we diagnosed 214 patients with Leishmaniasis who showed cutaneous lesions. They consisted of 119 males and 95 females and lived in the vicinities of Quetta in Balochistan Province (1700-1800m above sea level), and Larkana and Sukkur in Sindhi Province (100m above s
ea level). Sixty out of the 214 patients were confirmed to be cutaneous Leishmaniasis. The 60 patients were comprised of 40 males and 20 females whose ages ranged from 10 months to 50 years with the average of 20.7 years. We analyzed the cutaneous symptoms (dry type, wet type, dry and wet type), causative parasites, and geographic distributions of these 60 patients. Following the traditional classification of the clinical symptoms, we described them as dry type, wet type, and dry and wet type which showed both of the two types.
The methods to identify the causative parasites were as follows : 1)extraction of genomic DNA from the biopsy specimens and/or separately cultured parasites ; 2)PCR with consensus primers of Leishmania cytochrome b gene ; 3)direct sequencing of the amplified PCR products to determine the base sequences ; 4)identification of the causative parasites by NCBI/BLAST analysis.
Results and conclusion
The results were as follows : There were 16 cases of dry type, of which 6 cases (37.5%) were due to L.(L.)tropica. The remaining 10 cases (62.5%) were due to L.(L.) major, which was contrary to the descriptions of the traditional textbooks. There were 34 cases of wet type, of which 27 cases (79.4%) were due to L.(L.) major, while the remaining 6 cases (17.6%) were due to L.(L.)tropica. There were 10 cases of wet and dry type, of which one case (10%) was due to L.(L.)tropica, while the remaining 9 cases (90%) were due to L.(L.)major. As for a relation between the kinds of the parasites and the geographical factors of the patients, 45 (96%) out of 47 cases were due to L.(L.)major, which were found in Larkana city located in Indus Valley and in the desert area surrounding Sukkur city. Twelve (92%) out of 13 cases were due to L.(L.)tropica, which were found in Quetta city, a mountainous region near Afghanistan. As a conclusion, it seemed incorrect to infer the causative parasites from the clinical findings such as dry or wet types as described by the textbooks, but the geographical factors of the patients might be more influential for the diagnoses. Less