Significant multilocus linkage disequilibrium (mLD) for bothP. falciparum and P. vivax post-LLIN
For P. falciparum , matching haplotypes (allowing missing loci)
were seen in all post-LLIN datasets and the pre-LLIN ESP2 2005 dataset.
However, for P. vivax , matching haplotypes (allowing missing
loci) were rarely seen and only in post-LLIN data sets. Among the 332
complete P. falciparum multilocus haplotypes (9-loci successfully
genotyped) from all study sites, 16 repeated haplotypes were found, with
11 haplotypes represented two times, three represented three times, and
two represented four times. Clonal haplotypes were always found within
the same year and province, and in all cases except one in the same
catchment area, but not always in the same village (7/16 haplotypes
found in neighbouring villages). In ESP2 2005, one clonal haplotype was
found in two villages (Yenigo and Sengo, Figure 1) from different
catchment areas, roughly 40km apart. Among the 303 complete P.
vivax multilocus haplotypes (10-loci), two haplotypes were repeated,
with one haplotype represented two times (in two different villages in
ESP 2012), and one represented four times (three in one village, one in
a neighbouring village in MAD 2010).
To investigate whether inbreeding was present in these populations
(Smith, Smith, O’Rourke, & Spratt, 1993), non-random associations among
the microsatellite loci (mLD) were calculated for all complete and
unique haplotypes. Whilst mLD was absent from most pre-LLIN populations,
significant mLD was observed in P. falciparum and P. vivaxinfections post-LLIN (Table 1). In ESP 2012-13, mLD was high with unique
infections indicating the circulation of closely related haplotypes in
the population, suggesting near clonal transmission (low recombination
between diverse clones and high levels of inbreeding) or the presence of
high proportions of meiotic siblings among isolates (Bright et al.,
2014; Smith et al., 1993), as observed in the village of Sunuhu (Table
S3). Low, but significant mLD was found for P. falciparum in
Madang in 2006 (Table 1), however this population was structured
(Schultz et al., 2010), resulting in a phenomenon called the Wahlund
effect, confirmed by the fact that linkage equilibrium was restored when
mLD was analysed separately for subpopulations (Jennison et al., 2015;
Wahlund, 1928) (Table S4). In post-LLIN Madang, the observed mLD forP. falciparum is not the result of subpopulation structure, as
significant mLD remained in the subpopulations (Table S4). In the Mugil
area in 2014, significant mLD for P. falciparum remains due to
the circulation of a few very closely related haplotypes in the Megiar
village (Table S4, Dataset 1).