The genetic diversity and complexity of P. falciparum infections is to a very large extent an important indicator of malaria transmission intensity in a region and is a very useful marker for assessing naturally-acquired anti-malarial immunity as well as the impacts of intervention programmes [11, 33–37]. Numerous studies from different regions have devoted efforts at characterizing the genetic complexity of P. falciparum infections at the community level, but very little attention has been given to studying genetic diversity at the level of the microenvironment. In this study, we investigated the genetic diversity and complexities of P. falciparum infections in the microenvironment among siblings of the same household. To our knowledge, this is the first study to provide information on the genetic diversity and complexity of P. falciparum infection at the level of the microenvironment in Nigeria, and it will certainly be a great addition to the limited data available on the subject globally.
Our findings showed that P. falciparum isolates exhibit a remarkable degree of genetic diversity in the microenvironment. Interestingly, we found that the pattern of distribution of parasite populations within households may be categorized into two based on the prevalence of MSP-2 allelic families. The first category were households where both MSP-2 allele types (FC27 and 3D7) were present, while the second were households where only one MSP-2 allele type (FC27 or 3D7) was present. The majority of the households (88.4%) investigated belonged to the first category where both MSP-2 alleles were present, showing that parasite clones carrying FC27 and 3D7 alleles are widely distributed in the study region. This observation was in agreement with a previous study in Tanzania where most of the households investigated had parasites of mixed genotypes . An important observation in the households where both MSP-2 allelic families were prevalent was that the FC27 and 3D7 alleles were disproportionately distributed among the infected children. Thus, in some households, all the infected children had mixed allelic infections with parasites carrying both FC27 and 3D7 alleles. In contrast, in other households, one of the children had parasite isolates carrying a particular type of MSP-2 allele and the other child had parasite isolates carrying the other type of MSP-2 allele. Nevertheless, in some other households, one or two of the children may be infected with multiple parasite clones or genotypes which may belong to either of the MSP-2 alleles or both. Although about 65% of the households have at least one child with isolates carrying both the FC27 and the 3D7 allele types, only a few households (30.2%) were observed to have all the children carrying isolates belonging to both the FC27 and the 3D7 allelic families. In a previous study in Gabon, it was observed that about 80% of the members of the household investigated had parasite isolates carrying both FC27 and 3D7 alleles  although the study examined only one household. The observed high prevalence of MSP-2 multiclonal infection in this study is an indication of a high ongoing parasite transmission, suggestive of effective genetic recombination of the parasite population within the female Anopheles mosquito vector .
It was also interesting to note that a few households (11.6%) belonged to the second category, where all the children had parasites carrying only one MSP-2 allele type (FC27 or 3D7). This was also consistent with the findings from Tanzania, where they observed a few instances in which different people in the same household had parasites of similar genotypes . Apart from carrying isolates of the same allele, we also found instances where identical genotypes or clones (identical fragment sizes) of the same allele were found in all the children in a household. Such infections with parasites of similar genotypes within households might possibly suggest inoculation by a single or related mosquito.
Majority of the participants in this study were infected with a mixture of more than one parasite clone. On the whole, we found that about 65% of the study participants had polyclonal infections consisting of 2–6 clones with an overall MOI of 2.31 clones per infected child. This observation was not different from our previous report  and is also consistent with other data from Nigeria [40, 41] and from other parts of Africa [14, 24, 42–50]. The simultaneous infection of large number of individuals with multiple parasite genotypes in areas of high transmission intensity has been suggested to be attributable to either multiple inoculations of single clones, or by a single inoculation of multiple clones that may have undergone crossing and recombination in the female Anopheles mosquito vector [51, 52]. Recombination events during the sexual stage of the malaria parasites in the Anopheles mosquito can lead to independent chromosomal re-assortment of genes and is the principal mechanism for generating a novel combination of genes, and consequently, new parasite strains with novel genotypes [51–54]. However, there are suggestions also that high genetic diversity in the parasite population might lead to a gradual selection of more virulent strains which in turn can lead to the emergence and proliferation of drug-resistant parasites [41, 55, 56].
The present study has some limitations, including (i) the use of a single genetic marker and (ii) the fact that PCR may not be able to resolve between alleles of similar size but different sequences or those with a size difference of about 10 bp. All of these may potentially underestimate the complexity of infections in this study. However, our data has provided more insight into the genetic complexity of P. falciparum in the microenvironment. Future studies will need to take cognisance of the above limitations, use more robust techniques, and consider other regions with different malaria transmission intensity.