Abstract
Microbiological studies on the intercontinental transport of dust are confounded by the difficulty of obtaining sufficient material for analysis. Axenic samples of dust collected at high altitudes or historic specimens in museums are often so small and precious that the material can only be sacrificed when positive results are assured. With this in mind, we evaluated current methods and developed new ones in an attempt to catalogue all microbes present in small dust or sand samples. The methods used included classical microbiological approaches in which sand extracts were plated out on a variety of different media, polymerase chain reaction (PCR)-based amplification of 16S/18S rRNA sequences followed by construction of clone libraries, PCR amplification of 16S rRNA sequences followed by high-throughput sequencing (HtS) of the products and direct HtS of DNA extracted from the sand. A representative sand sample collected at Bahaï Wadi in the desert of the Republic of Chad was used. HtS with or without amplification showed the most promise and can be performed on ≤100 ng DNA. Since living microbes are often required, current best practices would involve geochemical and microscopic characterisation of the sample, followed by DNA isolation and direct HtS. Once the microbial content of the sample has been deciphered, growth conditions (including media) can be tailored to isolate the micro-organisms of interest.
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Acknowledgments
We would especially like to thank Dr. Agathe Stricker of the International Committee of the Red Cross in Geneva, Switzerland, for organising the collection of the “Sandman” sample C3 from the Republic of Chad. We thank Dora Gerber, Michal Parkan, Luiz Roesch and Wolfgang Streit for their unstinting help. In Switzerland, this work was supported by the Fonds National Suisse de la Recherche Scientifique (Projects 3100AO-104097 and 3100A0-116858), the Département de l’Instruction Publique du Canton de Genève and the Université de Genève. Work in the United States of America was made possible by grants from the National Science Foundation (Number MCB-0454030) and the United States Department of Agriculture (Numbers 2005-35319-16300, 00067345).
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10453_2012_9264_MOESM2_ESM.eps
Two litre “Leonard” jars planted with seedlings of Acacia albida, A. espinosa, A. saligna, A. schweinfurthii, A. tortilis and Vigna unguiculata (see Materials and methods). Panel A - jar inoculated with 1 g of sand from sample C3. Panel B - jar inoculated with 1 g sterile, quartz sand. Panel C – nodules on the roots of V. unguiculata. Panel D – nodules on the roots of A. espinosa. Panel E – nodules on the roots of A. albida. (EPS 18164 kb)
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Giongo, A., Favet, J., Lapanje, A. et al. Microbial hitchhikers on intercontinental dust: high-throughput sequencing to catalogue microbes in small sand samples. Aerobiologia 29, 71–84 (2013). https://doi.org/10.1007/s10453-012-9264-0
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DOI: https://doi.org/10.1007/s10453-012-9264-0