Peter M. Grewe

Appraisal of molecular genetic techniques in fisheries

Molecular genetic approaches began to be used in fisheries in the 1950s. These initial studies were of blood group variants, primarily in tunas, salmonids and cod (reviewed by de Ligny, 1969), and successfully demonstrated the existence of genetically controlled variation which could be used in analyses of population structure. However these serological procedures were not enthusiastically adopted by fisheries biologists and most were subsequently abandoned in favour of electrophoretic procedures revealing genetically determined protein polymorphism. One of the first of these studies was that of Sick (1961), who described haemoglobin variants in whiting (Gadus melangus, Gadidae) and cod (Gadus morhua, Gadidae), but the impetus for more extensive work was provided by the seminal papers of Harris (1966) and Lewontin and Hubby (1966). They showed humans and the fruitfly Drosophila pseudoobscura, respectively, to be highly polymorphic for electrophoretically detectable allozyme variation. Protein electrophoresis was found to be quick, reasonably inexpensive, and reproducible. Most plant and animal species, including commercially valuable finfish and shellfish, showed sufficient variation that population structure could be examined quickly.

GENETIC CHARACTERIZATION OF STRAINS OF CYANOBACTERIA USING PCR-RFLP OF THE cpcBA INTERGENIC SPACER AND FLANKING REGIONS1

Oligonucleotide primers, specific for conserved regions of the genes encoding the β‐ and α‐phycocyanin subunits of phycobilisomes (cpcB and cpcA) of cyanobacteria, were used to amplify a DNA fragment containing the intervening intergenic spacer region (cpcBA‐IGS) of 19 strains of three morphospecies of cyanobacteria. Six Australian strains were identified as Anabaena circinalis Rabenhorst, six strains were identified as Microcystis aeruginosa Kützing, and seven strains were identified as Nodularia spumigena Mertens. Restriction enzyme digestion of the amplification products from the strains revealed restriction fragment length polymorphism (RFLP) within all three morphospecies. Strains corresponding to M. aeruginosa were highly polymorphic: 11 of the 14 restriction enzymes used displayed RFLPs. The A. circinalis and N. spumigena strains were less variable: three of 14 enzymes and seven of 14 enzymes, respectively, showed RFLPs. The presence of genetic variation between strains within these three divergent morphospecies, which span two orders of cyanobacteria (Chroococcales Wettstein and Nostocales (Borzi) Geitler), show that the cpcBA‐ IGS fragment has broad application as a molecular marker for intrageneric studies of cyanobacteria systematics and genetics.

Allozyme and mitochondrial DNA variation in yellowfin tuna (Thunnus albacares) from the Pacific Ocean

Samples of yellowfin tuna (Thunnus albacares) collected in 1991 and 1992 from the western, central and eastern regions of the Pacific Ocean were examined for genetic variability. Four polymorphic allozyme loci (ADA*, FH*, GPI-S* and GPI-F*) were examined in all samples and a fifth polymorphism (GDA*) was examined in western and central samples only. Samples were also screened for mitochondrial DNA variation following restriction analysis by two enzymes (BcII and EcoRI) detecting polymorphic cut sites. Eighteen mtDNA haplotypes were revealed, with an overall nucleon diversity of 0.678. A subset of individuals screened for eight restriction enzymes had an overall nucleon diversity of 0.724 and a mean nucleotide diversity per sample of 0.359%. No significant spatial heterogeneity was detected for alleles at the ADA*, FH*, GPI-S* and GDA* loci nor for the mtDNA haplotypes. Significant heterogeneity was detected for GPI-F*. At this locus, the two eastern samples (southern California and northern Mexico) were not significantly different from each other but were significantly different (P<0.001) from the five western/central samples (Philippines, Coral Sea, Kiribati, Hawaii-91 and Hawaii-92). GPI-F*100 was the most common allele in western and central regions, GPI-F*75 the most common in eastern samples.

Genetic differentiation between Tasmanian cultured Atlantic salmon (Salmo salar L.) and their ancestral Canadian population: comparison of microsatellite DNA and allozyme and mitochondrial DNA variation

Abstract Atlantic salmon ( Salmo salar ) were imported to Australia from the River Philip, Nova Scotia, in the mid-1960s. A population was established in New South Wales, and in the mid-1980s ova from this population were used to found the Tasmanian salmon aquaculture industry. An allozyme and mitochondrial DNA examination of the Tasmanian and parent Canadian populations in 1993 showed some small but significant allele frequency differences between the two samples for one of seven polymorphic allozymes and for mitochondrial DNA haplotypes. However, there was no evidence of reduced genetic variability in the Tasmanian sample. The same individual fish from both populations have now been examined for eight polymorphic microsatellite loci. Small but significant differences in allele frequencies between the two samples were found for four of the eight loci, and there was evidence of a small overall loss of genetic variation (both heterozygosity and alleles) in the Tasmanian sample. Mean heterozygosity per microsatellite locus was more than twice that per allozyme locus: for the Tasmanian fish 0.434 ( n =63) (0.207 for allozymes) and for the Canadian fish 0.509 ( n =63) (0.182 for allozymes). Estimates of per-generation effective population sizes were calculated to be 65.2±19.7 (s.d.) from the microsatellite data, 106.1±74.4 for the allozymes data and 70.1±19.4 for the combined data set, assuming allele frequencies in the River Philip sample represent those in the progenitor population and ten generations of isolation. Microsatellite loci, with higher numbers of alleles and higher heterozygosities, are more sensitive than allozyme loci to changes in effective population size.

Evidence of discrete yellowfin tuna (Thunnus albacares) populations demands rethink of management for this globally important resource.

Tropical tuna fisheries are central to food security and economic development of many regions of the world. Contemporary population assessment and management generally assume these fisheries exploit a single mixed spawning population, within ocean basins. To date population genetics has lacked the required power to conclusively test this assumption. Here we demonstrate heterogeneous population structure among yellowfin tuna sampled at three locations across the Pacific Ocean (western, central, and eastern) via analysis of double digest restriction-site associated DNA using Next Generation Sequencing technology. The differences among locations are such that individuals sampled from one of the three regions examined can be assigned with close to 100% accuracy demonstrating the power of this approach for providing practical markers for fishery independent verification of catch provenance in a way not achieved by previous techniques. Given these results, an extended pan-tropical survey of yellowfin tuna using this approach will not only help combat the largest threat to sustainable fisheries (i.e. illegal, unreported, and unregulated fishing) but will also provide a basis to transform current monitoring, assessment, and management approaches for this globally significant species.

Sex-ratio-biasing constructs for the control of invasive lower vertebrates.

volume 32 NumBeR 5 mAy 2014 nature biotechnology editors of the top-ranked biotechnology and applied microbiology journals who kindly gave us their opinion on synthetic biology and iGEM. We are indebted to Càtedra de Divulgació de la Ciència (UCC+i) and VLC/Campus—Valencia International Campus of Excellence (Universitat de València) for their enthusiastic support. C.V. was funded with a FPU (Formación de Personal Universitario) grant from the Spanish MECD (Ministerio de Educación, Cultura y Deporte). This work was partially funded by the European Project ST-FLOW (FP7-KBBE-2011-5).

Mitogenomics of the Speartooth Shark challenges ten years of control region sequencing

BackgroundMitochondrial DNA markers have long been used to identify population boundaries and are now a standard tool in conservation biology. In elasmobranchs, evolutionary rates of mitochondrial genes are low and variation between distinct populations can be hard to detect with commonly used control region sequencing or other single gene approaches. In this study we sequenced the whole mitogenome of 93 Critically Endangered Speartooth Shark Glyphis glyphis from the last three river drainages they inhabit in northern Australia.ResultsGenetic diversity was extremely low (π =0.00019) but sufficient to demonstrate the existence of barriers to gene flow among river drainages (AMOVA ΦST =0.28283, P <0.00001). Surprisingly, the comparison with single gene sub-datasets revealed that ND5 and 12S were the only ones carrying enough information to detect similar levels of genetic structure. The control region exhibited only one mutation, which was not sufficient to detect any structure among river drainages.ConclusionsThis study strongly supports the use of single river drainages as discrete management units for the conservation of G. glyphis. Furthermore when genetic diversity is low, as is often the case in elasmobranchs, our results demonstrate a clear advantage of using the whole mitogenome to inform population structure compared to single gene approaches. More specifically, this study questions the extensive use of the control region as the preferential marker for elasmobranch population genetic studies and whole mitogenome sequencing will probably uncover a large amount of cryptic population structure in future studies.

A comparison of allozymes and mitochondrial DNA in Atlantic salmon from Tasmania and from the ancestral population in Canada

Abstract About 30 years ago, Atlantic salmon ova were imported from River Philip, Nova Scotia, to found a population in New South Wales, Australia. Ova from the New South Wales stock were subsequently used to found the Tasmanian stock. Allozyme analysis of 7 variable nuclear loci of parr from Tasmania and from their ancestral Canadian population showed that the two samples were very similar to each other, although some minor differences were apparent. One of 18 restriction enzymes ( Hin PI) used in mitochondrial DNA analysis revealed polymorphism, with the Tasmanian sample having a small but significant increase in the frequency of the rarer haplotype (0.194±0.046 versus 0.056±0.028). There was no evidence of reduced nuclear or mitochondrial DNA variability in the Tasmanian stock compared to its ancestral stock.

Multi-genetic marker approach and spatio-temporal analysis suggest there is a single panmictic population of swordfish Xiphias gladius in the Indian Ocean.

Genetic population structure of swordfish Xiphias gladius was examined based on 2231 individual samples, collected mainly between 2009 and 2010, among three major sampling areas within the Indian Ocean (IO; twelve distinct sites), Atlantic (two sites) and Pacific (one site) Oceans using analysis of nineteen microsatellite loci (n = 2146) and mitochondrial ND2 sequences (n = 2001) data. Sample collection was stratified in time and space in order to investigate the stability of the genetic structure observed with a special focus on the South West Indian Ocean. Significant AMOVA variance was observed for both markers indicating genetic population subdivision was present between oceans. Overall value of F-statistics for ND2 sequences confirmed that Atlantic and Indian Oceans swordfish represent two distinct genetic stocks. Indo-Pacific differentiation was also significant but lower than that observed between Atlantic and Indian Oceans. However, microsatellite F-statistics failed to reveal structure even at the inter-oceanic scale, indicating that resolving power of our microsatellite loci was insufficient for detecting population subdivision. At the scale of the Indian Ocean, results obtained from both markers are consistent with swordfish belonging to a single unique panmictic population. Analyses partitioned by sampling area, season, or sex also failed to identify any clear structure within this ocean. Such large spatial and temporal homogeneity of genetic structure, observed for such a large highly mobile pelagic species, suggests as satisfactory to consider swordfish as a single panmictic population in the Indian Ocean.

Complete mitochondrial genome of the Critically Endangered speartooth shark Glyphis glyphis (Carcharhiniformes: Carcharhinidae)

Abstract In this study we present the first complete mitogenome for the speartooth shark Glyphis glyphis, a rare euryhaline elasmobranch from northern Australia and Papua New Guinea. The mitogenome is 16,702 bp in length and the overall base composition is 31.5% A; 26.0% C; 13.0% G and 29.5% T. It includes 2 ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes, 13 protein-coding genes and a putative 1066 bp long control region. The COI gene is initiated by GTG codon whereas the remaining protein-coding genes started with the ATG codon. This study will help elucidate the taxonomy of this poorly known group of sharks.