Engineering of complex polyketide biosynthesis — insights from sequencing of the monensin biosynthetic gene cluster

The biosynthesis of complex reduced polyketides is catalysed in actinomycetes by large multifunctional enzymes, the modular Type I polyketide synthases (PKSs). Most of our current knowledge of such systems stems from the study of a restricted number of macrolide-synthesising enzymes. The sequencing of the genes for the biosynthesis of monensin A, a typical polyether ionophore polyketide, provided the first genetic evidence for the mechanism of oxidative cyclisation through which polyethers such as monensin are formed from the uncyclised products of the PKS. Two intriguing genes associated with the monensin PKS cluster code for proteins, which show strong homology with enzymes that trigger double bond migrations in steroid biosynthesis by generation of an extended enolate of an unsaturated ketone residue. A similar mechanism operating at the stage of an enoyl ester intermediate during chain extension on a PKS could allow isomerisation of an E double bond to the Z isomer. This process, together with epoxidations and cyclisations, form the basis of a revised proposal for monensin formation. The monensin PKS has also provided fresh insight into general features of catalysis by modular PKSs, in particular into the mechanism of chain initiation. Journal of Industrial Microbiology & Biotechnology (2001) 27, 360–367. Received 18 March 2001/ Accepted in revised form 09 July 2001     

Genetic Diversity and Host Specificity Varies across Three Genera of Blood Parasites in Ducks of the Pacific Americas Flyway

Birds of the order Anseriformes, commonly referred to as waterfowl, are frequently infected by Haemosporidia of the genera Haemoproteus, Plasmodium, and Leucocytozoon via dipteran vectors. We analyzed nucleotide sequences of the Cytochrome b (Cytb) gene from parasites of these genera detected in six species of ducks from Alaska and California, USA to characterize the genetic diversity of Haemosporidia infecting waterfowl at two ends of the Pacific Americas Flyway. In addition, parasite Cytb sequences were compared to those available on a public database to investigate specificity of genetic lineages to hosts of the order Anseriformes. Haplotype and nucleotide diversity of Haemoproteus Cytb sequences was lower than was detected for Plasmodium and Leucocytozoon parasites. Although waterfowl are presumed to be infected by only a single species of Leucocytozoon, L. simondi, diversity indices were highest for haplotypes from this genus and sequences formed five distinct clades separated by genetic distances of 4.9%–7.6%, suggesting potential cryptic speciation. All Haemoproteus and Leucocytozoon haplotypes derived from waterfowl samples formed monophyletic clades in phylogenetic analyses and were unique to the order Anseriformes with few exceptions. In contrast, waterfowl-origin Plasmodium haplotypes were identical or closely related to lineages found in other avian orders. Our results suggest a more generalist strategy for Plasmodium parasites infecting North American waterfowl as compared to those of the genera Haemoproteus and Leucocytozoon.     

Study of response of Swiss Webster mice to light subunit of mushroom tyrosinase

The light subunit of mushroom, Agaricus bisporus, tyrosinase (LSMT), has been identified as an extrinsic component of the enzyme. Its function is unknown, but it can cross an epithelial cell layer, which suggests that it can be absorbed by the intestine. A similar capability has been demonstrated for the HA-33 component of the progenitor toxin from Clostridium botulinum, which is the closest structural homolog of LSMT. Unlike HA-33, LSMT appears to be non-immunogenic as shown by preliminary tests in Swiss Webster mice. We investigated the immunogenicity and histopathology of LSMT in mice to determine its safety in vivo. LSMT did not evoke generation of antibodies after prolonged periods of intraperitoneal administration. Histopathological observations confirmed the absence of responses in organs after twelve weekly administrations of LSMT. We found that LSMT is not toxic and is less immunogenic than the C. botulinum HA-33 protein, which supports further research and development for pharmaceutical application.     

Transmission of influenza reflects seasonality of wild birds across the annual cycle

Influenza A Viruses (IAV) in nature must overcome shifting transmission barriers caused by the mobility of their primary host, migratory wild birds, that change throughout the annual cycle. Using a phylogenetic network of viral sequences from North American wild birds (2008–2011) we demonstrate a shift from intraspecific to interspecific transmission that along with reassortment, allows IAV to achieve viral flow across successive seasons from summer to winter. Our study supports amplification of IAV during summer breeding seeded by overwintering virus persisting locally and virus introduced from a wide range of latitudes. As birds migrate from breeding sites to lower latitudes, they become involved in transmission networks with greater connectivity to other bird species, with interspecies transmission of reassortant viruses peaking during the winter. We propose that switching transmission dynamics may be a critical strategy for pathogens that infect mobile hosts inhabiting regions with strong seasonality.     

High fidelity does not preclude colonization: range expansion of molting Black Brant on the Arctic coast of Alaska

High rates of site fidelity have been assumed to infer static distributions of molting geese in some cases. To test this assumption, we examined movements of individually marked birds to understand the underlying mechanisms of range expansion of molting Black Brant (Branta bernicla nigricans) on the Arctic Coastal Plain (ACP) of Alaska. The Teshekpuk Lake Special Area (TLSA) on the ACP was created to protect the primary molting area of Brant. When established in 1977, the TLSA was thought to include most, if not all, wetlands used by molting Brant on the ACP. From 2010 to 2013, we surveyed areas outside the TLSA and counted an average of 9800 Brant per year, representing 29–37% of all molting Brant counted on the ACP. We captured and banded molting Brant in 2011 and 2012 both within the TLSA and outside the TLSA at the Piasuk River Delta and Cape Simpson to assess movements of birds among areas across years. Estimates of movement rates out of the TLSA exceeded those into the TLSA, demonstrating overall directional dispersal. We found differences in sex and age ratios and proportions of adult females with brood patches, but no differences in mass dynamics for birds captured within and outside the TLSA. Overall fidelity rates to specific lakes (0.81, range = 0.49–0.92) were unchanged from comparable estimates obtained in the early 1990s. We conclude that Brant are dispersing from the TLSA into new molting areas while simultaneously redistributing within the TLSA, likely as a consequence of changes in relative habitat quality. Shifts in distribution resulted from colonization of new areas by young birds as well as low levels of directional dispersal of birds that previously molted in the TLSA. Based on combined counts, the overall number of molting Brant across the ACP has increased substantially.     

Molecular evolution of genes in avian genomes

Background

Obtaining a draft genome sequence of the zebra finch (Taeniopygia guttata), the second bird genome to be sequenced, provides the necessary resource for whole-genome comparative analysis of gene sequence evolution in a non-mammalian vertebrate lineage. To analyze basic molecular evolutionary processes during avian evolution, and to contrast these with the situation in mammals, we aligned the protein-coding sequences of 8,384 1:1 orthologs of chicken, zebra finch, a lizard and three mammalian species.

Results

We found clear differences in the substitution rate at fourfold degenerate sites, being lowest in the ancestral bird lineage, intermediate in the chicken lineage and highest in the zebra finch lineage, possibly reflecting differences in generation time. We identified positively selected and/or rapidly evolving genes in avian lineages and found an over-representation of several functional classes, including anion transporter activity, calcium ion binding, cell adhesion and microtubule cytoskeleton.

Conclusions

Focusing specifically on genes of neurological interest and genes differentially expressed in the unique vocal control nuclei of the songbird brain, we find a number of positively selected genes, including synaptic receptors. We found no evidence that selection for beneficial alleles is more efficient in regions of high recombination; in fact, there was a weak yet significant negative correlation between ω and recombination rate, which is in the direction predicted by the Hill-Robertson effect if slightly deleterious mutations contribute to protein evolution. These findings set the stage for studies of functional genetics of avian genes.     

Phylogenetic evidence for role-reversals of gender-associated mitochondrial DNA in Mytilus (Bivalvia: Mytilidae)

Distinct gender-associated mitochondrial DNA (mtDNA) lineages (i.e., lineages which are transmitted either through males or through females) have been demonstrated in two families of bivalves, the Mytilidae (marine mussels) and the Unionidae (freshwater mussels), which have been separated for more than 400 Myr. The mode of transmission of these M (for male-transmitted) and F (for female-transmitted) molecules has been referred to as doubly uniparental inheritance (DUI), in contrast to standard maternal inheritance (SMI), which is the norm in animals. A previous study suggested that at least three origins of DUI are required to explain the phylogenetic pattern of M and F lineages in freshwater and marine mussels. Here we present phylogenetic evidence based on partial sequences of the cytochrome c oxidase subunit I gene and the 16S RNA gene that indicates the DUI is a dynamic phenomenon. Specifically, we demonstrate that F lineages in three species of Mytilus mussels, M. edulis, M. trossulus, and M. californianus, have spawned separate lineages which are now associated only with males. This process is referred to as "masculinization" of F mtDNA. By extension, we propose that DUI may be a primitive bivalve character and that periodic masculinization events combined with extinction of previously existing M types effectively reset the time of divergence between conspecific gender-associated mtDNA lineages.      

Do hunters target auxiliary markers? An example using black brant

Auxiliary markers play an essential role in understanding migration, movement, demography, and behavior of migratory birds. Use of such markers relies on the assumption that the markers do not affect the traits of interest. Neck collars, among the most conspicuous of markers, substantially affect risk of harvest, and survival even in the absence of harvest. Effects of less-conspicuous markers, such as colored plastic tarsal bands, are not well understood. We used 30 years (1986–2015) of banding, recovery, and recapture data from the Yukon-Kuskokwim Delta in Alaska, USA, to assess differences in direct band recovery rates (DRRs) between black plastic and brightly colored plastic bands applied to black brant (Branta bernicla nigricans). We also assessed the effect of the color of plastic tarsal bands on annual survival, risks of natural mortality harvest, and fidelity to the breeding colony of adult female black brant. When assessing only DRRs we found that brightly colored bands were recovered at higher rates than black plastic bands in the early 2000s, but DRRs for black bands increased more rapidly through time, resulting in similar DRRs for the 2 band colors at the end of the study. Using a Burnham model structure, our results demonstrated that individuals fitted with colored bands had slightly lower hazards of dying from natural causes or hunting than individuals carrying less-conspicuous black tarsal bands. Differences on annual probability scales were small and credible intervals broadly overlapped between band types, indicating minimal differences between individuals with different band types; however, we could not resolve all confounding in our study design and we suggest that specific studies directed at assessing marker effects are warranted. We encourage education of hunters about their roles as citizen scientists and the potentially detrimental effect of targeting birds with auxiliary markers.