Sampling and molecular work
Soil samples were collected in September, 2012, in Kinabalu and Crocker
Range Parks of Sabah, Borneo; in May, 2011 and 2013, in Jujuy, Salta and
Tucumán provinces of Argentina; and in June, 2016, in Bocas del Toro,
Chiriquí, Colón and Panamá provinces of Panama. The sampling sites
represent the entire elevation range of various types of forests in the
respective regions. Forest type, elevation, geographic coordinates, and
localities are shown in Table S1 and on maps corresponding to the three
regions (Figs. S1-3).
In Argentina and Borneo, 40 soil cores, 2 cm in diameter and ca. 20 cm
deep, were taken at each sampling site (ca. 10 × 25 m). Cores were
collected ca. 2 m from each other to minimize the probability of
sampling the same genet repeatedly. In Panama, 10 soil cores of the
above dimensions were collected in each site (ca. 4 × 5 m). Soil cores
taken at a given site were pooled, resulting in a composite soil sample
for each site. With respect to the samples collected in Argentina and
Panama, ca. 20 g of each sample was kept frozen until lyophilization ca.
2 weeks later. In Borneo, because of the more remote location, soil
samples were dried immediately at 30-35 °C. Because of these
differences, we handled the data separately for each region for the
analyses that are the focus of this paper. We did use the combined
dataset to provide the first characterization of the putative ‘core’
pantropical mycobiome of montane and lowland forests, i.e., fungi shared
among the Neotropical and Paleotropical regions and characteristic of
the different elevation zones.
Genomic DNA was extracted from 0.5 ml of dry soil from each sample with
the NucleoSpin® soil kit (Macherey-Nagel Gmbh & Co.,
Düren, Germany) according to manufacturer’s protocol. For each sample,
two independent DNA extractions were carried out and the extracts were
pooled. The ITS2 region (ca. 250 bp) of the nuclear ribosomal DNA repeat
was amplified via PCR with primers fITS7 (Ihrmark et al. 2012)
and ITS4 (White et al. 1990). The ITS4 primer was labelled with
sample-specific Multiplex Identification DNA-tags (MIDs). The amplicon
library was sequenced at Naturalis Biodiversity Center (Naturalis) with
an Ion 318TM Chip and an Ion Torrent Personal Genome
Machine (Life Technologies, Guilford, CT, U.S.A.). Chemical analyses of
soil samples from Argentina and Borneo were carried out as described in
Geml et al. (2014) and Geml et al. (2017), respectively.
The samples from Panama were analysed by the Instituto de Investigación
Agropecuaria de Panamá (IDIAP). Climate data were obtained from the
WorldClim database (www.worldclim.org) based on the geographic
coordinates of the sampling sites.