Electronic Supplementary Material
Methods
PCR reactions
For malaria parasite detection, two µL of DNA extract were used to seed a first PCR reaction
targeting a 170bp-long fragment of malaria parasite mtDNA (qPlasm1f 5’CTGACTTCCTGGCTAAACTTCC-3’ and qPlasm1r 5’CATGTGATCTAATTACAGAAYAGGA-3’). This reaction was run using Platinum Taq
(Invitrogen) according to manufacturer’s instructions and under the following conditions: 3 min
at 95°C, 15 cycles [1 min at 95°C, 1 min at 54°C, 1 min at 72°C], 7 min at 72°C. Using 1 µL of
this first reaction, a nested PCR targeting a 90bp-long fragment (qPlasm2f 5’AGAAAACCGTCTATATTCATGTTTG-3’ and qPlasm2r 5’ATAGACCGAACCTTGGACTC-3’) was then performed using GoTaq qPCR Master Mix
(Promega, Fitchburg, WI) under the following cycling conditions: 5 min at 95°C, 40 cycles [15 s
at 95°C, 30 s at 57°C, 30 s at 60°C], ramp increase from 55 to 95°C (for dissociation analysis).
Positive samples were then subjected to a semi-nested PCR targeting a longer malaria parasite
mtDNA fragment (350bp). In the first round primers Plasmseq1f 5’GGATTTAATGTAATGCCTAGACGTA-3’ and Plasmseq1r 5’ATCTAAAACACCATCCACTCCAT-3’ were used; in the second round Pspcytbf1 5’TGCCTAGACGTATTCCTGATTATCCAG-3’ [1] and Plasmseq1r. The first reaction was run
from two µL of DNA extract while the second was seeded with two µL of the first round
reaction. Both PCRs were run using Platinum Taq under the following conditions: 3 min at
95°C, 50 cycles [40 s at 95°C, 45 s at 57°C, 45 s at 72°C], 7 min at 72°C.
1.
Kaiser M., Lowa A., Ulrich M., Ellerbrok H., Goffe A.S., Blasse A., Zommers Z., Couacy-
Hymann E., Babweteera F., Zuberbuhler K., et al. 2010 Wild chimpanzees infected with 5 Plasmodium
species. Emerg Infect Dis 16(12), 1956-1959. (doi:10.3201/eid1612.100424).
Tables
Table S1. Estimated coefficients from a model with total DNA as an offset term and
sampling period included as a random effect. Age was not significant (full reduced model
comparison: χ2=11.44, df=2, P=0.003). Age and daytime were z-transformed.
term
intercept
age
sex (F=0; M=1)
daytime
daytime2
autocorrelation
estimate
-0.923
-1.317
0.463
0.238
-0.120
0.523
SE
0.575
0.515
0.572
0.217
0.218
0.192
z
P
-2.557
0.808
1.098
-0.550
2.726
0.011
0.419
0.272
0.582
0.006
Table S2. Estimated coefficients from a model with total mitochondrial mammal DNA as
an offset term and sampling period included as a random effect. Age was not significant (full
reduced model comparison: χ2=12.2, df=2, P=0.002). Age and daytime were z-transformed.
term
intercept
age
sex (F=0, M=1)
daytime
daytime2
autocorrelation
estimate
-1.193
-1.103
0.760
0.212
0.030
0.703
SE
0.543
0.453
0.545
0.222
0.223
0.202
z
P
-2.433
1.395
0.956
0.135
3.482
0.015
0.163
0.339
0.893
<0.001