Building strong relationships between conservation genetics and primary industry leads to mutually beneficial genomic advances

Several reviews in the past decade have heralded the benefits of embracing high‐throughput sequencing technologies to inform conservation policy and the management of threatened species, but few have offered practical advice on how to expedite the transition from conservation genetics to conservation genomics. Here, we argue that an effective and efficient way to navigate this transition is to capitalize on emerging synergies between conservation genetics and primary industry (e.g., agriculture, fisheries, forestry and horticulture). Here, we demonstrate how building strong relationships between conservation geneticists and primary industry scientists is leading to mutually‐beneficial outcomes for both disciplines. Based on our collective experience as collaborative New Zealand‐based scientists, we also provide insight for forging these cross‐sector relationships.     

Characterization of Expressed Sequence Tag–Derived Simple Sequence Repeat Markers for Aspergillus flavus: Emphasis on Variability of Isolates from the Southern United States

Simple sequence repeat (SSR) markers were developed from Aspergillus flavus expressed sequence tag (EST) database to conduct an analysis of genetic relationships of Aspergillus isolates from numerous host species and geographical regions, but primarily from the United States. Twenty-nine primers were designed from 362 tri-nucleotide EST-SSR sequences. Eighteen polymorphic loci were used to genotype 96 Aspergillus species isolates. The number of alleles detected per locus ranged from 2 to 24 with a mean of 8.2 alleles. Haploid diversity ranged from 0.28 to 0.91. Genetic distance matrix was used to perform principal coordinates analysis (PCA) and to generate dendrograms using unweighted pair group method with arithmetic mean (UPGMA). Two principal coordinates explained more than 75?% of the total variation among the isolates. One clade was identified for A. flavus isolates (n?=?87) with the other Aspergillus species (n?=?7) using PCA, but five distinct clusters were present when the others taxa were excluded from the analysis. Six groups were noted when the EST-SSR data were compared using UPGMA. However, the latter PCA or UPGMA comparison resulted in no direct associations with host species, geographical region or aflatoxin production. Furthermore, there was no direct correlation to visible morphological features such as sclerotial types. The isolates from Mississippi Delta region, which contained the largest percentage of isolates, did not show any unusual clustering except for isolates K32, K55, and 199. Further studies of these three isolates are warranted to evaluate their pathogenicity, aflatoxin production potential, additional gene sequences (e.g., RPB2), and morphological comparisons.     

A decaepitope polypeptide primes for multiple CD8+ IFN-gamma and Th lymphocyte responses: evaluation of multiepitope polypeptides as a mode for vaccine delivery

Proteins are generally regarded as ineffective immunogens for CTL responses. We synthesized a 100-mer decaepitope polypeptide and tested its capacity to induce multiple CD8(+) IFN-gamma and Th lymphocyte (HTL) responses in HLA transgenic mice. Following a single immunization in the absence of adjuvant, significant IFN-gamma in vitro recall responses were detected for all epitopes included in the construct (six A2.1-, three A11-restricted CTL epitopes, and one universal HTL epitope). Immunization with truncated forms of the decaepitope polypeptide was used to demonstrate that optimal immunogenicity was associated with a size of at least 30-40 residues (3-4 epitopes). Solubility analyses of the truncated constructs were used to identify a correlation between immunogenicity for IFN-gamma responses and the propensity of these constructs to form particulate aggregates. Although the decaepitope polypeptide and a pool of epitopes emulsified in IFA elicited similar levels of CD8(+) responses using fresh splenocytes, we found that the decaepitope polypeptide more effectively primed for in vitro recall CD8(+) T cell responses. Finally, immunogenicity comparisons were also made between the decaepitope polypeptide and a corresponding gene encoding the same polypeptide delivered by naked DNA immunization. Although naked DNA immunization induced somewhat greater direct ex vivo and in vitro recall responses 2 wk after a single immunization, only the polypeptide induced significant in vitro recall responses 6 wk following the priming immunization. These studies support further evaluation of multiepitope polypeptide vaccines for induction of CD8(+) IFN-gamma and HTL responses.     

Predictor species: Improving assessments of rare species occurrence by modeling environmental co‐responses

Designing an effective conservation strategy requires understanding where rare species are located. Because rare species can be difficult to find, ecologists often identify other species called conservation surrogates that can help inform the distribution of rare species. Species distribution models typically rely on environmental data when predicting the occurrence of species, neglecting the effect of species' co‐occurrences and biotic interactions. Here, we present a new approach that uses Bayesian networks to improve predictions by modeling environmental co‐responses among species. For species from a European peat bog community, our approach consistently performs better than single‐species models and better than conventional multi‐species approaches that include the presence of nontarget species as additional independent variables in regression models. Our approach performs particularly well with rare species and when calibration data are limited. Furthermore, we identify a group of “predictor species” that are relatively common, insensitive to the presence of other species, and can be used to improve occurrence predictions of rare species. Predictor species are distinct from other categories of conservation surrogates such as umbrella or indicator species, which motivates focused data collection of predictor species to enhance conservation practices.     

Body temperature null distributions in reptiles with nonzero heat capacity: seasonal thermoregulation in the American alligator (Alligator mississippiensis)

Regulation of body temperature may increase fitness of animals by ensuring that biochemical and physiological processes proceed at an optimal rate. The validity of current methods of testing whether or not thermoregulation in reptiles occurs is often limited to very small species that have near zero heat capacity. The aim of this study was to develop a method that allows estimation of body temperature null distributions of large reptiles and to investigate seasonal thermoregulation in the American alligator (Alligator mississippiensis). Continuous body temperature records of wild alligators were obtained from implanted dataloggers in winter (n=7, mass range: 1.6-53.6 kg) and summer (n=7, mass range: 1.9-54.5 kg). Body temperature null distributions were calculated by randomising behavioural postures, thereby randomly altering relative animal surface areas exposed to different avenues of heat transfer. Core body temperatures were predicted by calculations of transient heat transfer by conduction and blood flow. Alligator body temperatures follow regular oscillations during the day. Occasionally, body temperature steadied during the day to fall within a relatively narrow range. Rather than indicating shuttling thermoregulation, however, this pattern could be predicted from random movements. Average daily body temperature increases with body mass in winter but not in summer. Daily amplitudes of body temperature decrease with increasing body mass in summer but not in winter. These patterns result from differential exposure to heat transfer mechanisms at different seasons. In summer, alligators are significantly cooler than predictions for a randomly moving animal, and the reverse is the case in winter. Theoretical predictions show, however, that alligators can be warmer in winter if they maximised their sun exposure. We concluded that alligators may not rely exclusively on regulation of body temperature but that they may also acclimatise biochemically to seasonally changing environmental conditions.     

Long-range intra-protein communication can be transmitted by correlated side-chain fluctuations alone

Allosteric regulation is a key component of cellular communication, but the way in which information is passed from one site to another within a folded protein is not often clear. While backbone motions have long been considered essential for long-range information conveyance, side-chain motions have rarely been considered. In this work, we demonstrate their potential utility using Monte Carlo sampling of side-chain torsional angles on a fixed backbone to quantify correlations amongst side-chain inter-rotameric motions. Results indicate that long-range correlations of side-chain fluctuations can arise independently from several different types of interactions: steric repulsions, implicit solvent interactions, or hydrogen bonding and salt-bridge interactions. These robust correlations persist across the entire protein (up to 60 Å in the case of calmodulin) and can propagate long-range changes in side-chain variability in response to single residue perturbations.     

Molecular identification and virulence of three Aeromonas hydrophila isolates cultured from infected channel catfish during a disease outbreak in west Alabama (USA) in 2009

Three isolates (AL09-71, AL09-72, and AL09-73) of Aeromonas hydrophila were cultured from infected channel catfish Ictalurus punctatus during a disease outbreak in west Alabama, USA, in August 2009. Sequence analysis of the 16S-23S rDNA intergenic spacer region (ISR), cpn60, gyrB, and rpoD genes of the 3 strains revealed that the 3 strains were closely related to each other, sharing 97 to 99% nucleotide sequence similarities. However, ISR sequences of the 3 isolates from 2009 shared only 64% nucleotide sequences with AL98-C1B, a 1998 isolate of A. hydrophila cultured from diseased fish in Alabama. Sequences of cpn60, gyrB, and rpoD from the 3 isolates from 2009 shared 91 to 95% homologies with AL98-C1B. Based on both LD50 and LD95 values of intraperitoneal injection assays, the virulences of the 3 isolates from 2009 were not significantly different from each other, but were at least 200-fold more virulent than AL98-C1B, indicating that the 3 west Alabama isolates of A. hydrophila from 2009 were highly virulent to channel catfish.     

Mechanism of Action of Secreted Newt Anterior Gradient Protein

Anterior gradient (AG) proteins have a thioredoxin fold and are targeted to the secretory pathway where they may act in the ER, as well as after secretion into the extracellular space. A newt member of the family (nAG) was previously identified as interacting with the GPI-anchored salamander-specific three-finger protein called Prod1. Expression of nAG has been implicated in the nerve dependence of limb regeneration in salamanders, and nAG acted as a growth factor for cultured newt limb blastemal (progenitor) cells, but the mechanism of action was not understood. Here we show that addition of a peptide antibody to Prod1 specifically inhibit the proliferation of blastema cells, suggesting that Prod1 acts as a cell surface receptor for secreted nAG, leading to S phase entry. Mutation of the single cysteine residue in the canonical active site of nAG to alanine or serine leads to protein degradation, but addition of residues at the C terminus stabilises the secreted protein. The mutation of the cysteine residue led to no detectable activity on S phase entry in cultured newt limb blastemal cells. In addition, our phylogenetic analyses have identified a new Caudata AG protein called AG4. A comparison of the AG proteins in a cell culture assay indicates that nAG secretion is significantly higher than AGR2 or AG4, suggesting that this property may vary in different members of the family.     

Temporal patterns of diversification and microendemism in Eastern Highland endemic barcheek darters (Percidae: Etheostomatinae)

Eastern North America is the location of the world's most species-rich temperate freshwater fish fauna. Hypotheses regarding the geographic and temporal scale of teleost diversification in this region have not been broadly investigated using absolute divergence time estimates among the constituent lineages. This study used time-calibrated molecular phylogenies estimated from mitochondrial and nuclear genes to investigate the temporal and geographic signatures of diversification within barcheek darters, a clade of allopatrically distributed species endemic to the Eastern Highlands. Results from divergence time estimates using an uncorrelated lognormal model suggest that the barcheek darters are an ancient group with a crown node estimate of 16.3 mya, 95% highest posterior density (HPD): [12.4, 20.5], and the clade is characterized by substantial intraspecific divergence times within several species. In particular, the Caney Fork endemic Etheostoma basilare comprises five strongly supported and deeply divergent clades with a most recent common ancestor estimated at 8.0 mya, 95% HPD: [5.6, 10.7]. These results are concordant with the hypothesis that geologically stable areas of eastern North America have facilitated both the generation and preservation of lineages across a substantial breadth of evolutionary time, and that allopatric speciation in darters has occurred at much smaller spatial scales than previously realized.