Phylogeography of the southern African vlei rat, Otomys irroratus, inferred from chromosomal and DNA sequence data

Engelbrecht, Adriaan (2010-12)

Thesis (MSc)--University of Stellenbosch, 2010.

Thesis

ENGLISH ABSTRACT: This study examines the phylogeography of the southern African vlei rat, Otomys irroratus using the mtDNA cyt b gene and chromosomal data derived using G-, and C-banding, Ag-NOR staining and fluorescence in situ hybridization (FISH using flow sorts of Myotomys unisulcatus). A total of N = 102 specimens were used from the Western Cape, Eastern Cape, Northern Cape, Free State, Mpumalanga and KwaZulu-Natal provinces of South Africa. Of the N = 102, N = 55 comprised fresh material while N = 47 comprises museum material obtained from the Durban Natural Science Museum of South Africa. Cytogentic analysis of N = 55 specimens collected from seven localities in South Africa revealed intra-specific variation resulting from two rearrangements, namely pericentric inversions and heterochromatin variation. Of the 55 specimens that were analyzed 47% contained inversions or centromeric shifts on four autosomes (OIR1, OIR4, OIR6 and OIR10) which were present singly in specimens (i.e. none of the specimens contained all four inversions concurrently). These inversions were present in both homozygous and heterozygous state over a wide geographic range suggesting that they are floating polymorphisms. Given the potential role of inversions in post-mating isolation (through production of aneuploid gametes), the prevalence of inversions as floating polymorphisms in the vlei rats suggest that they are probably retained in the population through suppression of recombination in the inverted regions of the chromosomes. In addition, differences between populations is due to the presence or absence of heterochromatic arms (and not inversions), which cause variation in the NFa (40 – 49) and supernumerary B chromosomes, resulting in the variation in diploid number (2n = 28 – 32). Analysis of N = 55 specimens revealed Ag-NORs on 7 autosomal pairs 1, 2, 5, 7, 8 and 9 proximal to the centromere on the short arm of the chromosome. Pair 8 also displayed Ag-NOR at the distal end of the long arm of the chromosome in individuals from the Free State province. Pair 3 showed two Ag- NORs occurring proximal to the centromere on the short arm and on the terminal end of the long arm, respectively. I obtained 953bp of mtDNA cyt b from fresh material and 400bp from museum material. Using maximum parsimony and Bayesian inference two main clades were retrieved. Clade A specimens occur mainly in the Western and Eastern Cape provinces of South Africa. Clade B specimens occur in the Eastern Cape, Free State, KwaZulu-Natal, Northern Cape and Mpumalanga provinces of South Africa. The mean sequence divergence between the main clades (A and B) is 7.0% and between sub-clades comprising clade B is 4.8%, while within clade A the sequence divergence was 1.91%. Nested clade analysis revealed allopatric fragmentation within O. irroratus. Chromosomal characters also support the two evolutionary lineages as clade A has pericentric inversions which occur as floating polymorphisms which are absent in clade B. Clade B in turn is fixed for a complex tandem fusion rearrangement which is absent from clade A. Divergence date estimates indicate that the two clades separated around 1.1 MYA, which coincides with climate changes since the late Pliocene/Pleistocene epochs. Cladogenesis within this species complex could therefore have been influenced by habitat fragmentation. A full taxonomic review of O. irroratus is therefore warranted by this study.

AFRIKAANSE OPSOMMING: Die suider Afrikaanse vlei rot, Otomys irroratus word gekenmerk deur fenotipiese konservatisme regoor die spesie se verspreiding en het groot chromosomale variasie met diploied chromosoom getalle wat reeks vanaf 2n = 23 tot 2n = 32. Hierdie variasie binne O. irroratus het gelei tot die beskrywing van drie chromosomale groupe naamlik die A sitotipe wat gekenmerk word deur 'n akrosentriese komplement. Die tweede groep wat die B sitotipe genoem word besit ten minste agt chromosoom pare met heterokromatiese kort arms, onderwyl die derde group (die C sitotipe) vier chromosoom pare het met heterokromatiese kort arms. Hierdie studie bestudeer die bevolkings genetika struktuur van O. irroratus deur 102 monsters te analiseer wat gekollekteer was regoor die spesie se verspreiding binne Suid-Afrika en die mitochondriale merker sitokroom b sowel as chromosoom fluoressent hibridisasie te gebruik. Ek het 55 monsters van sewe lokaliteite binne Suid-Afrika sitogeneties geanaliseer deur gebruik te maak van G- en C-bandbepaling asook die hibridisasie patrone geproduseer deur die vloeisorteerde chromosoome van Myotomys unisulcatus. Die analise het gewys dat 47% van die monsters perisentromeriese inversies besit het, wat slegs aangetref was of die outosome OIR1, OIR4, OIR6 en OIR10. Hierdie inversies was nooit almal teenwoordig binne dieselfde monster nie en was gevind in beide heterosigotiese en homosigotiese vorm. Die inversies kom ook voor oor 'n wye verspreiding wat daarop aandui dat dit swerwende polymorfisme is. Omdat inversies lei tot die produksie van aneuploiede gamete speel hulle 'n belangrike rol in post-parings reproduktiewe isolasie, die verskyning van swerwende inversies binne vlei rotte dui dus daarop dat hulle onderhou word binne populasie verband deur die onderdrukking van rekombinasie in die gedeeltes van die chromosoom. Verdere verskille tussen populasies behels die voorkoms of afwesigheid van heterochromatiese kort arms wat (nie inversies) wat lei tot die variasies in die Nfa (40 – 49). Die variasie in diploied getal (2n = 28 – 32) is eksklusief as gevolg van B chromosoome. Ag-NOR banding het ook gewys dat daar twee evolusionêre lyne binne O. irroratus voorkom. Verder het filogenetiese analise van al die monsters verkryg deur volgorde-bepaling met behulp van maksimale parsimonie en Bayesian afleiding twee klades geidentifiseer. Klade A diere kom voor in die Wes en Oos-Kaap provinsies van Suid-Afrika terwyl klade B diere voorkom in die Oos-Kaap, Vrystaat, KwaZulu-Natal, Noord-Kaap en Mpumalanga provinsies onderskeidelik van Suid-Afrika. Die gemiddelde volgorde-bepalings verskille beloop 7% tussen die twee hoof klades (A en B) en tussen sub-klades 4.8%, terwyl binne klade A die verskille slegs 1.91% beloop het. Analise van die verwantskap tussen die klades het gewys dat allopatriese fragmentasie heel waarskynlik gelei het tot die populasie genetiese struktuur binne O. irroratus. Chromosoom karakters onderskraag die twee evolusionêre lyne waar klade A slegs perisentriese inversies besit wat swerwend wat ontbreek in klade B. Klade B op sy beurt besit 'n komplekse tandemme fusie wat glad nie voorkom in klade A nie. Molekulêre datering het verder gewys dat die twee klades omtrent 1.1 miljoen jaar gelede versprei het, wat ooreenstem met die klimaats veranderinge wat sedert die Peioceen en Pleistoceen plaasgevind het. Klade vorming binne die spesies komples kan daarom as gevolg van habitat fragmentasie plaasgevind het. Hierdie studie dus noodsaak 'n volle taksonomiese ondersoek van O. irroratus ten einde vas te stel hoeveel spesies binne die komplex voorkom.

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