Secondary contact and asymmetrical gene flow in a cosmopolitan marine fish across the Benguela upwelling zone.

The combination of oceanographic barriers and habitat heterogeneity are known to reduce connectivity and leave specific genetic signatures in the demographic history of marine species. However, barriers to gene flow in the marine environment are almost never impermeable which inevitably allows secondary contact to occur. In this study, eight sampling sites (five along the South African coastline, one each in Angola, Senegal and Portugal) were chosen to examine the population genetic structure and phylogeographic history of the cosmopolitan bluefish (Pomatomus saltatrix), distributed across a large South-east Atlantic upwelling zone. Molecular analyses were applied to mtDNA cytochrome b, intron AM2B1 and 15 microsatellite loci. We detected uncharacteristically high genetic differentiation (FST 0.15-0.20; P<0.001) between the fish sampled from South Africa and the other sites, strongly influenced by five outlier microsatellite loci located in conserved intergenic regions. In addition, differentiation among the remaining East Atlantic sites was detected, although mtDNA indicated past isolation with subsequent secondary contact between these East Atlantic populations. We further identified secondary contact, with unidirectional gene flow from South Africa to Angola. The directional contact is likely explained by a combination of the northward flowing offshore current and endogenous incompatibilities restricting integration of certain regions of the genome and limiting gene flow to the south. The results confirm that the dynamic system associated with the Benguela current upwelling zone influences species distributions and population processes in the South-east Atlantic.

Evaluating the resolution power of new microsatellites for species identification and stock delimitation in the Cape hakes Merluccius paradoxus and Merluccius capensis (Teleostei: Merlucciidae).

The utility of 15 new and 17 previously published microsatellite markers was evaluated for species identification and stock delimitation in the deep-water hake Merluccius paradoxus and the shallow-water hake Merluccius capensis. A total of 14 microsatellites were polymorphic in M. paradoxus and 10 in M. capensis. Two markers could individually discriminate the species using Bayesian clustering methods and a statistical power analysis showed that the set of markers for each species is likely to detect subtle genetic differentiation (FST < 0·006) that will be valuable to delimit and characterize genetic stocks.

Identification of naturally occurring hybrids between two overexploited sciaenid species along the South African coast.

Hybridisation between fish species can play a significant role in evolutionary processes and can influence management and conservation planning, however, this phenomenon has been widely understudied, especially in marine organisms. The distribution limits of two sciaenid species (silver kob, Argyrosomus inodorus, and dusky kob, A. japonicus) partly overlap along the South African coast, where both species have undergone severe depletion due to overfishing. Following the identification of a number of possible cases of species misidentification or hybridisation (21 out of 422 individuals), nuclear and mitochondrial DNA data (12microsatellite loci and 562bp of the COI gene) were analysed to investigate the genetic composition of these individuals. Results indicated a field-based species misidentification rate of approximately 2.8% and a rate of natural hybridisation of 0.7%. Interestingly, all hybrid fish resulted from first-generation (F1) hybridisation events, which occurred exclusively between silver kob females and dusky kob males. Whether hybridisation is the result of natural events (such as secondary contact following a shift in distribution range), or anthropogenic activities (size-selective pressure due to overfishing), these findings have important implications for critical recovery and future management of these species in the wild.

Phylogeographic patterns and cryptic speciation across oceanographic barriers in South African intertidal fishes.

Biogeographic boundaries are the meeting zone of broadly distributed faunas, or the actual cause of a faunal break. In the latter case, closely related sister species should be found across such a boundary. To achieve such a situation, preliminary stages are expected, where phylogeographic breaks followed by genetic cryptic speciation would be observed. Biogeographic boundaries, in the Cape Point/Cape Agulhas region of southern Africa, offer an ideal system to test such predictions. Here, we studied two intertidal clinid fish species that are endemic to southern Africa, Clinus superciliosus (n = 127) and Muraenoclinus dorsalis (n = 114). Using mitochondrial control region, 16S rRNA, 12S rRNA and NADH2 genes and the nuclear rhodopsin and the first intron of the S7 ribosomal protein gene, we show both phylogeographic breaks and likely cryptic speciation in each species. Pairwise Φ(st) results suggest population genetic structuring for both species, with higher levels for M. dorsalis (Φ(st) = 0.34-0.93) than for C. superciliosus (Φ(st) = 0.1-0.74). Further, we recover two and three distinct lineages within M. dorsalis and C. superciliosus, respectively. Phylogenetic topologies, concordance between nuclear and mitochondrial markers and levels of sequence divergence, which are consistent with closely related sister species pairs, suggest the presence of cryptic species. Our results therefore meet the expectation for reduced gene flow at a biogeographic barrier, which translates into significant genetic breaks and cryptic sister species.

Development and characterization of polymorphic markers for the sap-stain fungus Ophiostoma quercus.

Eight polymorphic markers were developed from South African isolates of Ophiostoma quercus. The genome was screened for repeat regions using the fast isolation by amplified fragment length polymorphism of sequences containing repeats protocol and 20 de novo primer pairs flanking putative microsatellite regions were designed. Eight loci were optimized and their polymorphisms evaluated by sequencing. The repeat and flanking regions were highly polymorphic containing both indels and base-pair substitutions revealing a total of 46 alleles in 14 isolates and an average heterozygosity of 0.68. Substantial sequence variability makes these markers useful for genotyping populations in order to calculate diversity and monitor global movement of O. quercus.

Isolation and characterization of polymorphic tetranucleotide microsatellite loci in the pelagic perciform fish Pomatomus saltatrix (Linnaeus, 1766) from South Africa.

Eight polymorphic microsatellite loci, containing simple tetranucleotide repeats, were isolated de novo from a Pomatomus saltatrix partial genomic library using the fast isolation by amplified fragment length polymorphism of sequences containing repeats protocol. These loci were further characterized in 100 individuals from two putative populations off the South African east coast. The loci are highly polymorphic with 18-37 alleles (on average 24 alleles/locus) and the observed heterozygosity in both populations was high (0.79). These loci will be used to assess population structuring in P. saltatrix along the southern African coast with consideration of implications for future management of this important linefish species.

Mitochondrial DNA differentiation in the critically endangered Berg River redfin (Pseudobarbus burgi).

The Berg River redfin (Pseudobarbus burgi) is a critically endangered endemic cyprinid from South Africa. We investigated mitochondrial DNA control region variation among specimens representative of five populations drawn from two adjacent river systems. Phylogenetic analyses, a minimum spanning network, and an analysis of molecular variance underscore the pronounced genetic separation of redfins originating from the geographically closely allied Verlorevlei and Berg Rivers, two populations that may have remained isolated since the Pleistocene. Despite a lack of geographic structuring within the Berg River, historic female gene flow among the upper and middle/lower parts of the river appears to be limited and the contemporary populations are probably isolated due to deterioration of the mainstream of the river. Our results suggest that the Berg and Verlorevlei populations should be managed as distinct conservation units. We encourage the use of sanctuaries, particularly by private landowners within both river systems, as this approach may contribute effectively to preserving genetic diversity within the species.

Francolin phylogenetics: molecular, morphobehavioral, and combined evidence.

The phylogenetics of francolins (Francolinus species) were reassessed by obtaining 660 bp of sequence of the mitochondrial DNA (mtDNA) cytochrome b gene from 20 species, the Common Quail Coturnix coturnix africana, and the Madagascar Partridge Margaroperdix madagarensis. Published sequences of the Japanese Quail C. c. japonica, Alectoris partridges, and the Junglefowl Gallus gallus were also included. Separate analysis of the 200 phylogenetically informative cytochrome b characters and the 25 informative morphobehavioral characters, as well as a combined analysis of molecular and morphobehavioral data, do not support francolin monophyly but provide strong evidence for two previously suggested clades--the quail-francolins (or partridges) and the partridge-francolins (pheasants/francolins). The quail-francolin clade comprises three groups of African francolins and three Asian species that were previously considered more closely related to the partridge-francolins. The partridge-francolin clade, which includes four groups of African francolins, forms a sister group to the Coturnix quails, the Madagascar Partridge, and the Alectoris partridges. The molecular data suggest that the two francolin clades diverged approximately 3-6 MYA. Climatic fluctuations of the past 2.5 MYA may have led to the diversification of the ecologically different francolin species groups and speciation within them.

An evaluation of the Gilford 3400 automatic enzyme analyser.

An assessment of the Gilford Automatic Enzyme Analyser was conducted over a period of one year. The optics of the instrument were satisfactory with regard to accuracy of wavelength selection and linearity of absorbance response. Excellent precision was obtained for both absorbance readings and operation of the dispenser pump. Carry-over within the microflow-cell was low. The method of operation recommended by the manufacturers for enzyme determinations failed to take account of endogenous blank reactions which could lead to significant error. When revised methods utilising a pre-incubation stage and initiation with a single substrate were employed, the results correlated well with those obtained with standard automatic (LKB 8600) and manual (Pye Unicam SP 800) kinetic systems for aspartate and alanine aminotransferase, creatine phosphokinase and alpha-hydroxybutyrate dehydrogenase, and the precision at all activity levels was satisfactory. Acceptable precision could not be obtained over the clinical range for enzyme assays requiring a blank determination on each sample (5'-nucleotidase and adenosine deaminase) and those with very low normal serum activities (isocitrate dehydrogenase and glutamate dehydrogenase). These limitations appeared to be due to relative insensitivity of the transducer response and liability to optical disturbance. This apart, the instrument has many advantages over alternative equipment.