caa a22 4500 445341998 CHVBK 20180317142816.0 cr unu---uuuuu 170323e20110601xx s 000 0 eng 10.1007/s00436-010-2208-0 doi (NATIONALLICENCE)springer-10.1007/s00436-010-2208-0 Leishmania sp. identification by PCR associated with sequencing of target SSU rDNA in paraffin-embedded skin samples stored for more than 30years [Elektronische Daten] [Ana de Lima, Ricardo Zampieri, Thaíse Tomokane, Márcia Laurenti, Fernando Silveira, Carlos Corbett, Lucile Floeter-Winter, Cláudia Gomes] Paraffin-embedded samples commonly stored at educational and research institutions constitute tissues banks for follow-up or epidemiological studies; however, the paraffin inclusion process involves the use of substances that can cause DNA degradation. In this study, a PCR protocol was applied to identify Leishmania strains in 33 paraffin-embedded skin samples of patients with American cutaneous leishmaniasis. DNA was obtained by the phenol-chloroform protocol following paraffin removal and then used in PCR or nested PCR based on the nucleotide sequence of the small subunit ribosomal RNA (SSU rDNA). The amplicons obtained were cloned and sequenced to determine the single nucleotide polymorphism that distinguishes between different Leishmania species or groups. This assay allowed to distinguish organisms belonging to the subgenus Viannia and identify L. (Leishmania) amazonensis and L. (L.) chagasi of the Leishmania subgenus. Of the 33 samples, PCR and nested PCR identified 91% of samples. After sequencing the PCR product of 26 samples, 16 were identified as L. (L.) amazonensis, the other 10 contain organisms belonging to the L. (Viannia) sub-genus. These results open a huge opportunity to study stored samples and promote relevant contributions to epidemiological studies. Springer-Verlag, 2010 de Lima Ana Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut Zampieri Ricardo Physiology Department, Biosciences Institute of São Paulo University, São Paulo, Brazil aut Tomokane Thaíse Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut Laurenti Márcia Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut Silveira Fernando Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut Corbett Carlos Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut Floeter-Winter Lucile Physiology Department, Biosciences Institute of São Paulo University, São Paulo, Brazil aut Gomes Cláudia Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut Parasitology Research Springer-Verlag 108/6(2011-06-01), 1525-1531 0932-0113 108:6<1525 2011 108 436 https://doi.org/10.1007/s00436-010-2208-0 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s00436-010-2208-0 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- de Lima Ana Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut NATIONALLICENCE 700 1- Zampieri Ricardo Physiology Department, Biosciences Institute of São Paulo University, São Paulo, Brazil aut NATIONALLICENCE 700 1- Tomokane Thaíse Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut NATIONALLICENCE 700 1- Laurenti Márcia Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut NATIONALLICENCE 700 1- Silveira Fernando Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut NATIONALLICENCE 700 1- Corbett Carlos Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut NATIONALLICENCE 700 1- Floeter-Winter Lucile Physiology Department, Biosciences Institute of São Paulo University, São Paulo, Brazil aut NATIONALLICENCE 700 1- Gomes Cláudia Pathology Department, Medical School of São Paulo University, São Paulo, Brazil aut NATIONALLICENCE 773 0- Parasitology Research Springer-Verlag 108/6(2011-06-01), 1525-1531 0932-0113 108:6<1525 2011 108 436 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer