|dc.description.abstract||Bovine anaplasmosis caused by Anaplasma marginale is endemic in South Africa. This
endemicity is due to presence of tick vectors that transmit A. marginale the causal agent
of the disease and the high seroprevalence in Limpopo, Free State and North West
provinces. To date, the genetic diversity of A. marginale isolates infecting cattle in all
South African provinces, except Free State, are generally unknown. Recently, vaccines
based on the A. marginale major surface protein 1 a (MSP1 a) has been proposed as a
strategy for controlling bovine anaplasmosis. However, characterization of genetic
diversities of the A. marginale isolates in these regions is still needed before this protein
can be used for vaccine development. Therefore, the aim of this study was to determine
the prevalence, genetic diversity and phylogenetic relationship of A. marginale infecting
cattle in all South African provinces except the Free State.
A total of 280 whole blood samples were collected from cattle in all provinces with
exception of the Free State. Twenty six districts and municipalities were included in this
sampling. Anaplasma marginale genomic DNA was then extracted from the blood
sample using ZR Genomic DNA 1M Tissue Miniprep (Zymo Research, CA, USA). A
polymerase chain reaction (PCR) was done with primers targeting msp1a and msp4
genes and the PCR products were sequenced using genetic analyser (ABI, Life
technologies, CA, USA). The generated sequences were analysed by bioinformatics
and their phylogeny as well as genetic diversity index (GDI) was determined based on
the sequences of msp1a and msp4 genes.
Overall, the prevalence of A. marginale infection in cattle was 76% in all provinces
except for Northern Cape Province where the prevalence was zero. The prevalence per
province was as follows: Eastern Cape 19.1 %, Gauteng 9.6%, KwaZulu-Natal 23.0%,
Limpopo 15.3%, Mpumalanga 10.1%, North West 12.4% and Western Cape 10.5%.
The msp1 a revealed genetic variability with regions of different types of tandem repeats.
Some repeats were conserved amongst the A. marginale strains and revealed low
variable peptides in the MSP1 a tandem repeats. A polynomial correlation (R2=0. 76)
was observed between the GDI and anaplasmosis prevalence per province.
Interestingly, provinces with the highest prevalence were not the ones with highest or
The analysis of msp4 gene sequences, which provided evolutionary information about
geographically distinct A. marginale strains, was used in the present study for
phylogenetic analysis of samples from Limpopo (LP), Mpumalanga (MP), North West
(NW), Gauteng (GP), KwaZulu-Natal (KZN), Eastern Cape (EC) and Western Cape
(WC) provinces of South Africa. Two clades were observed which consisted of first
clade (LP, NW, GP, KZN and WC) and second clade (MP and EC) isolates.
In addition when DNA sequence variation of msp4 gene was analysed in combination
with isolates from other countries outside South Africa, important phylogeographic
information was observed. The South African strains had 100% identity with isola tes
from Kenya, Zimbabwe and Australia. Good representation of the Southern and
Northern Hemispheres was observed and demonstrated that the msp4 gene was a
good phylogeographic marker. These results indicated that A. marginale is widespread
in South Africa, and suggested that the analysis of msp1a and msp4 gene sequences
provided an understanding of the phylogeny and epidemiology of A. marginale in South