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1412-033X (Printed Edition) ISSN: 2085-4722 (Electronic) ISSN: 1412-033X (printed edition) ISSN: 2085-4722 (electronic) Journal of Biological Diversity Volume 12 – Number 1 – January 2011 FIRST PUBLISHED: 2000 ISSN: 1412-033X (printed edition) 2085-4722 (electronic) EDITORIAL BOARD (COMMUNICATING EDITORS): Abdel Fattah N.A. Rabou (Palestine), Dato A. Latiff Mohamad (Malaysia), Alan J. Lymbery (Australia), Ali Saad Mohamed (Sudan), Bambang H. Saharjo (Indonesia), Charles H. Cannon Jr. (USA), Edi Rudi (Indonesia), Guofan Shao (USA), Hassan Poorbabaei (Iran), Hwan Su Yoon (USA), Jamaluddin (India), Joko R. Witono (Indonesia), Katsuhiko Kondo (Japan), Livia Wanntorp (Sweden), Mahendra K. Rai (India), María La Torre Cuadros (Peru), Mariela A. Marinoff (Argentine), Mochamad A. Soendjoto (Indonesia), Salvador Carranza (Spain), Shahabuddin (Indonesia), Sonia Malik (Brazil), Sugiyarto (Indonesia), Thaweesakdi Boonkerd (Thailand) EDITOR-IN-CHIEF: Sutarno EDITORIAL MEMBERS: English Literary Editor: I Made Sudiana ([email protected]) Technical Editor & Banking: Solichatun ([email protected]) Distribution & Marketing: Rita Rakhmawati ([email protected]) Webmaster: Ari Pitoyo ([email protected]) MANAGING EDITORS: Ahmad Dwi Setyawan ([email protected]) PUBLISHER: Department of Biology, Faculty of Mathematics and Natural Sciences, Sebelas Maret University, Surakarta and The Society for Indonesian Biodiversity ADDRESS: Jl. Ir. Sutami 36A Surakarta 57126. Tel. +62-271-7994097, Tel. & Fax.: +62-271-663375, Email: [email protected] BANKING: Solichatun, BNI KC Sebelas Maret, Acc. No. 0033691646 ONLINE: www.unsjournals.com ACCREDITED BY DECREE OF THE DIRECTORATE GENERAL OF HIGHER EDUCATION, THE MINISTRY OF NATIONAL EDUCATION, REPUBLIC OF INDONESIA No. 65a/DIKTI/Kep/2008 (valid until October 2011) EXPERTATION AND CORRESPONDING EMAIL OF THE COMMUNICATING EDITORS: GENETIC DIVERSITY: Alan J. Lymbery ([email protected]), Hwan Su Yoon ([email protected]), Mahendra K. Rai ([email protected]), Salvador Carranza ([email protected]), Sonia Malik ([email protected]). SPECIES DIVERSITY: Dato A. Latiff Mohamad ([email protected]), Joko R. Witono ([email protected]), Katsuhiko Kondo ([email protected]), Livia Wanntorp ([email protected]), Thaweesakdi Boonkerd ([email protected]). ECOSYSTEM DIVERSITY: Abdel Fattah N.A. Rabou ([email protected]), Ali Saad Mohamed ([email protected]), Bambang H. Saharjo ([email protected]), Charles H. Cannon Jr. ([email protected]), Edi Rudi ([email protected]), Guofan Shao ([email protected]), Hassan Poorbabaei ([email protected]), Jamaluddin ([email protected]), Mochamad A. Soendjoto ([email protected]), Shahabuddin ([email protected]), Sugiyarto ([email protected]). ETHNOBIOLOGY: María La Torre Cuadros ([email protected]), Mariela A. Marinoff ([email protected]). BIODIVERSITAS ISSN: 1412-033X (printed edition) Volume 12, Number 1, January 2011 ISSN: 2085-4722 (electronic) Pages: 1-6 DOI: 10.13057/biodiv/d120101 Identification and characterization of Salmonella typhi isolates from Southwest Sumba District, East Nusa Tenggara based on 16S rRNA gene sequences CHARIS AMARANTINI1,♥, LANGKAH SEMBIRING2, HARIPURNOMO KUSHADIWIJAYA3, WIDYA ASMARA4 1Faculty of Biology, Duta Wacana Christian University (UKDW). Jl. Dr. Wahidin Sudirohusodo No. 5-27, Yogyakarta 55224, Indonesia. Tel.: +62-274- 563929. Fax.: +62-0274-4513235. email: [email protected] 2Faculty of Biology, Gadjah Mada University (UGM), Yogyakarta 55284, Indonesia. 3Field Epidemiology Training Program, Department of Public Health, Faculty of Medicine, Gadjah Mada University (UGM), Yogyakarta 55284, Indonesia. 4Department of Microbiology, Faculty of Veterinary Medicine, Gadjah Mada University (UGM), Yogyakarta 55284, Indonesia. Manuscript received: 9 April 2010. Revision accepted: 23 August 2010. ABSTRACT Amarantini C, Sembiring L, Kushadiwijaya H, Asmara W (2011) Identification and characterization of Salmonella typhi isolates from Southwest Sumba District, East Nusa Tenggara based on 16S rRNA gene sequences. Biodiversitas 12: 1-6. The incidence rate of typhoid fever in the Southwest Sumba District, East Nusa Tenggara was approximately about 725/100,000. In spite of such rate, there was not much known-yet about the molecular epidemiology of the disease. Thus, having accurate data and a strong discriminatory ability was crucial to scrutinize the molecular epidemiology of S. typhi with a molecular phylogenetic approach based on 16S rRNA gene sequences. Sixteen isolates representative of S. typhi from different geographical regions in Southwest Sumba District along with the reference strain S. typhi NCTC 786 had been identified and characterized based on 16S rRNA gene sequences using PCR amplification and sequencing. The 16S rRNA sequences data were aligned with the corresponding available S. typhi sequences retrieved from the NCBI database by using CLUSTAL X software. Phylogenetic trees were generated with PHYLIP software package. Molecular phylogenetic analysis indicated that all the isolates belong to S. typhi species were suggested by their relativity with the type strain of S. typhi ATCC19430T. It was also found that the isolates which belong to S. typhi species formed several different centers of diversity within the 16S rRNA gene tree. Each clade consisted of the strains from different geographical places in the District. Thus, to conclude the inquiry, there was evident inter-geographical spread of the strains and it tended to spread further into more remote areas in the District. Key words: Salmonella typhi strains, typhoid fever, 16S rRNA gene sequences, molecular phylogenetic analysis. INTRODUCTION symptoms ranging from mild illness with slight fever, the body feels uncomfortable, coughing and the clinical There are 17 to 22 million cases of typhoid fever circumstances such as severe abdominal pain and worldwide per year and it causes 216,000 to 600,000 complications. This condition could often cause difficulties deaths annually (Steele 2008). Based on a preliminary to diagnose it when merely based only on clinical symptom survey, the fever was highly endemic in Southwest Sumba (Muliawan and Surjawidjaja 1999). In addition to such District, East Nusa Tenggara and it infected 725/ 100,000 high rate, information about the molecular epidemiology of people per year in the District according to the report made the disease was unknown. Thus, it was indispensable to by Karitas Hospital in 2006. Until recently however it has have accurate data and a strong discriminative ability to not been under controlled because the limitations of health distinguish the types of the strains of the pathogenic service units to diagnose the typhoid through laboratory bacteria needed in the epidemiological study. studies. The incidence rate in the area was higher than the The application of the molecular detection methods for incidence in rural areas (358/100,000) or semi rural areas the epidemiological study was advantageous because the (157/100,000 inhabitants). It was nearly the same as the genetic profile of the bacteria is a source of information to total number of people infected in urban areas map the spread of the bacteria in the community. (810/100,000) annually in Indonesia (WHO 2003). Henceforth it is beneficial to determine best-fitted control Typhoid fever is an acute systemic infection caused by strategies over the disease. With the given rationale, this Salmonella enterica subsp. enterica serotype typhi study was aimed to unravel the strain diversities belonging (Salmonella typhi). The acute systemic infection causes S. to the S. typhi species isolated from typhoid fever patients typhi get into the blood stream after passing through using a molecular phylogenetic approach based on 16S several organs in the host. This disease shows clinical rRNA gene sequences, and also to understand the spread of S. typhi based on its varieties and interrelations among the 2 BIODIVERSITAS 12 (1): 1-6, January 2011 strains isolated from the infected patients in Southwest edited and assembled with Finch TV 1.4.0 and DNA Baser Sumba District, East Nusa Tenggara. sequence analysis software. Complete assembled sequences were aligned with the corresponding S. typhi sequences retrieved from the NCBI database with CUSTAL X MATERIALS AND METHODS software (Thompson et al. 1997). Bacterial strains Construction of phylogenetic tree In this inquiry, there were 16 representative isolates of Based on 16S rRNA nucleotide sequences, a S. typhi from different geographical regions in Southwest phylogenetic tree was constructed with PHYLIP software Sumba District. They were from infected patients in package (Felsenstein 1993) with the neighbor-joining Karitas Hospital in Weetabula, a private clinic in Elopada algorithm (Saito and Nei 1987). The evolutionary distance Subdistrict in Southwest Sumba District, and Lende Moripa matrix for the neighbor-joining method was generated in Hospital in Waikabubak, West Sumba District. Specimen accordance with the description introduced by Jukes and collection and microbiological identification methods were Cantor (1969). The phylogenetic distances were obtained described in the journal article published previously by adding only the values of the horizontal components. (Amarantini et al. 2009). Eventually, the matrix of the nucleotide similarity and difference was generated with PHYDIT software (Chun Extraction of bacterial DNA, PCR amplification and 1999). DNA sequencing Bacterial DNA was extracted in accordance with the Mapping the
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