Sample processing and data collection
Plant morphometrics: In those plants collected in the field we
recorded the number of branches, plant height, number of primary roots,
length of the longest root, and root volume. Sampled plants were dried
at 60ºC for 72 hrs., to record the dry biomass of roots and shoots.
AMF-colonization rates and spore identification: Thin secondary
roots were collected to determine the root colonization rates from AMF,
using a modified version of the Trypan Blue technique (see Ramos-Zapataet al. , 2011). In every site, we pooled the rhizospheric soil
from each of the five sampled plants per site, and then a sample of 100
g of this composite sample was used to extract spores using the wet
sieve method (see Mejía-Alva et al. , 2018). Afterward, the spores
were placed in microscope slides with PVLG-Melzer solution to estimate
spore density, richness, and diversity of AMF. Viable spores were
counted and identified as morphospecies based on the color, presence,
and type of ornamentation, and presence of bulbs using INVAM species
descriptions as a guide (INVAM, 2017).
Soil nutrients: From the pooled soil collected per site, a
sub-sample of 50 g was taken and sieved through a 1mm mesh to perform
soil nutrient and pH analyses. Nitrogen (N) and inorganic phosphorus (P)
were assessed by the Kjeldahl method and Olsen method (see
Estrada-Medina et al ., 2016), respectively. Meanwhile potassium
(K), sodium (Na), and calcium (Ca) were determined by flame photometry
(Helmke and Sparks, 1987). pH values were obtained with potentiometry
(see Estrada-Medina et al ., 2016).