Antimicrobial resistance in Antarctica: is it still a pristine environment?

Although the rapid spread of antimicrobial resistance (AMR), particularly in relation to clinical settings, is causing concern in many regions of the globe, remote, extreme environments, such as Antarctica, are thought to be relatively free from the negative impact of human activities. In fact, Antarctica is often perceived as the last pristine continent on Earth. Such remote regions, which are assumed to have very low levels of AMR due to limited human activity, represent potential model environments to understand the mechanisms and interactions underpinning the early stages of evolution, de novo development, acquisition and transmission of AMR. Antarctica, with its defined zones of human colonisation (centred around scientific research stations) and large populations of migratory birds and animals, also has great potential with regard to mapping and understanding the spread of early-stage zoonotic interactions. However, to date, studies of AMR in Antarctica are limited. Here, we survey the current literature focussing on the following: i) Dissection of human-introduced AMR versus naturally occurring AMR, based on the premise that multiple drug resistance and resistance to synthetic antibiotics not yet found in nature are the results of human contamination ii) The potential role of endemic wildlife in AMR spread There is clear evidence for greater concentrations of AMR around research stations, and although data show reverse zoonosis of the characteristic human gut bacteria to endemic wildlife, AMR within birds and seals appears to be very low, albeit on limited samplings. Furthermore, areas where there is little, to no, human activity still appear to be free from anthropogenically introduced AMR. However, a comprehensive assessment of AMR levels in Antarctica is virtually impossible on current data due to the wide variation in reporting standards and methodologies used and poor geographical coverage. Thus, future studies should engage directly with policymakers to promote the implementation of continent-wide AMR reporting standards. The development of such standards alongside a centralised reporting system would provide baseline data to feedback directly into wastewater treatment policies for the Antarctic Treaty Area to help preserve this relatively pristine environment. Video Abstract.

Sources of elevated heavy metal concentrations in sediments and benthic marine invertebrates of the western Antarctic Peninsula.

Antarctica is one of the least anthropogenically-impacted areas of the world. Metal sources to the marine environment include localised activities of research stations and glacial meltwater containing metals of lithogenic origin. In this study, concentrations of nine metals (Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Zn) were examined in three species of benthic invertebrates collected from four locations near Rothera Research Station on the western Antarctic Peninsula: Laternula elliptica (mudclam, filter feeder), Nacella concinna (limpet, grazer) and Odontaster validus (seastar, predator and scavenger). In addition, metals were evaluated in sediments at the same locations. Metal concentrations in different body tissues of invertebrates were equivalent to values recorded in industrialized non-polar sites and were attributed to natural sources including sediment input resulting from glacial erosion of local granodioritic rocks. Anthropogenic activities at Rothera Research Station appeared to have some impact on metal concentrations in the sampled invertebrates, with concentrations of several metals higher in L. elliptica near the runway and aircraft activities, but this was not a trend that was detected in the other species. Sediment analysis from two sites near the station showed lower metal concentrations than the control site 5 km distant and was attributed to differences in bedrock metal content. Differences in metal concentrations between organisms were attributed to feeding mechanisms and habitat, as well as depuration routes. L. elliptica kidneys showed significantly higher concentrations of eight metals, with some an order of magnitude greater than other organs, and the internal structure of O. validus had significantly higher Ni. This study supports previous assessments of N. concinna and L. elliptica as good biomonitors of metal concentrations and suggests O. validus as an additional biomonitor for use in future Antarctic metal monitoring programs.

Habituation underpins preference for mates with novel phenotypes in the guppy.

Populations harbour enormous genetic diversity in ecologically important traits. Understanding the processes that maintain this variation is a long-standing challenge in evolutionary biology. Recent evidence indicates that a mating preference for novel sexual signals can be a powerful force maintaining genetic diversity. However, the proximate underpinnings of this preference, and its generality, remain unclear. Here, we test the hypothesis that preference for novel sexual signals is underpinned by habituation, a nearly ubiquitous form of learning whereby individuals become less responsive to repetitive stimuli. We use the Trinidadian guppy ( Poecilia reticulata), in which male colour patterns are diverse yet heritable. We show that repeated exposure to males with a given colour pattern reduces female interest in males with that pattern, and that interest recovers following brief isolation. These results fulfil two core criteria of habituation: responsiveness decline and spontaneous recovery. To distinguish habituation from sensory adaptation and fatigue, we also demonstrate stimulus specificity and dishabituation. These results provide the first evidence that habituation causes a preference for novel sexual signals, addressing the mechanistic underpinnings of this mating preference. Given the pervasiveness of habituation among taxa and sensory contexts, our findings suggest that preference for novelty may play an underappreciated role in mate choice and the maintenance of genetic variation.

The role of learning by a predator, Rivulus hartii, in the rare-morph survival advantage in guppies.

Negative frequency-dependent selection (NFDS), where rare types are favoured by selection, can maintain diversity. However, the ecological processes that mediate NFDS are often not known. Male guppies (Poecilia reticulata) exhibit extreme diversity of colour patterning and, in a previous field experiment, rare morphs had a survival advantage. Here, we test the hypothesis that predators impose NFDS because they are efficient at capturing familiar prey morphs, but are less efficient at capturing unfamiliar morphs. Over a series of trials, we presented Rivulus hartii, a natural predator of guppies, with male guppies with the same colour patterning (A trials); then, for a second series of trials, we presented the rivulus with guppies with a new colour pattern (B trials). The success of rivulus at capturing guppies on the first attack increased over successive A trials. First attack success decreased significantly for the early B trials, and then increased during successive B trials, eventually reaching the same level as in the best A trials. This experiment demonstrates that learning, perhaps through long-term search image formation, plays a role in predation success on familiar vs. unfamiliar prey morphs. These results support the hypothesis that predator learning contributes to the maintenance of the extreme male guppy polymorphism seen in nature.

Biodiversity of air-borne microorganisms at Halley Station, Antarctica.

A study of air-borne microbial biodiversity over an isolated scientific research station on an ice-shelf in continental Antarctica was undertaken to establish the potential source of microbial colonists. The study aimed to assess: (1) whether microorganisms were likely to have a local (research station) or distant (marine or terrestrial) origin, (2) the effect of changes in sea ice extent on microbial biodiversity and (3) the potential human impact on the environment. Air samples were taken above Halley Research Station during the austral summer and austral winter over a 2-week period. Overall, a low microbial biodiversity was detected, which included many sequence replicates. No significant patterns were detected in the aerial biodiversity between the austral summer and the austral winter. In common with other environmental studies, particularly in the polar regions, many of the sequences obtained were from as yet uncultivated organisms. Very few marine sequences were detected irrespective of the distance to open water, and around one-third of sequences detected were similar to those identified in human studies, though both of these might reflect prevailing wind conditions. The detected aerial microorganisms were markedly different from those obtained in earlier studies over the Antarctic Peninsula in the maritime Antarctic.

Low temperature bioremediation of oil-contaminated soil using biostimulation and bioaugmentation with a Pseudomonas sp. from maritime Antarctica.

AIMS: To identify native Antarctic bacteria capable of oil degradation at low temperatures. METHODS AND RESULTS: Oil contaminated and pristine soils from Signy Island (South Orkney Islands, Antarctica) were examined for bacteria capable of oil degradation at low temperatures. Of the 300 isolates cultured, Pseudomonas strain ST41 grew on the widest range of hydrocarbons at 4 degrees C. ST41 was used in microcosm studies of low temperature bioremediation of oil-contaminated soils. Microcosm experiments showed that at 4 degrees C the levels of oil degradation increased, relative to the controls, with (i) the addition of ST41 to the existing soil microbial population (bioaugmentation), (ii) the addition of nutrients (biostimulation) and to the greatest extent with (iii) a combination of both treatments (bioaugmentation and biostimulation). Addition of water to oil contaminated soil (hydration) also enhanced oil degradation, although less than the other treatments. Analysis of the dominant species in the microcosms after 12 weeks, using temporal temperature gradient gel electrophoresis, showed Pseudomonas species to be the dominant soil bacteria in both bioaugmented and biostimulated microcosms. CONCLUSIONS: Addition of water and nutrients may enhance oil degradation through the biostimulation of indigenous oil-degrading microbial populations within the soil. However, bioaugmentation with Antarctic bacteria capable of efficient low temperature hydrocarbon degradation may enhance the rate of bioremediation if applied soon after the spill. SIGNIFICANCE AND IMPACT OF THE STUDY: In the future, native soil bacteria could be of use in bioremediation technologies in Antarctica.

Genetic and environmental effects on secondary sex traits in guppies (Poecilia reticulata).

Male guppies (Poecilia reticulata) exhibit extreme phenotypic and genetic variability for several traits that are important to male fitness, and several lines of evidence suggest that resource level affects phenotypic expression of these traits in nature. We tested the hypothesis that genetic variation for male secondary sex traits could be maintained by genotype-specific effects of variable resource levels (genotype-environment interaction). To do this, we measured genetic variation and covariation under two environmental conditions--relatively low and relatively high food availability. We found high levels of genetic variation for most traits, but we only found a significant G x E interaction across food levels for one trait (body size) for one population. The across-environment correlations for size were large and positive, indicating that the reaction norms for size did not cross. We also found that male colour pattern elements had nearly an order of magnitude more genetic variation than did male size. Heritability estimates indicated that Y-linked genes are responsible for some of the genetic variation in male size and colour traits. We discuss implications of these results for theories of the maintenance of genetic variation in male secondary sexual traits in guppies.

A preliminary study of airborne microbial biodiversity over Peninsular Antarctica.

This study used PCR-based molecular biological identification techniques to examine the biodiversity of air sampled over Rothera Point (Antarctic Peninsula). 16S rDNA fragments of 132 clones were sequenced and identified to reveal a range of microorganisms, including cyanobacteria, actinomycetes, diatom plastids and other uncultivated bacterial groups. Matches for microorganisms that would be considered evidence of human contamination were not found. The closest matches for many of the sequences were from Antarctic clones already in the databases or from other cold environments. Whilst the majority of the sequences are likely to be of local origin, back trajectory calculations showed that the sampled air may have travelled over the Antarctic Peninsula immediately prior to reaching the sample site. As a result, a proportion of the detected biota may be of non-local origin. Conventional identification methods based on propagule morphology or culture are often inadequate due to poor preservation of characteristic features or loss of viability during airbome transfer. The application of molecular biological techniques in describing airbome microbial biodiversity represents a major step forward in the study of airborne biota over Antarctica and in the distribution of microorganisms and propagules in the natural environment.

Absence of ST7 mutations in tumor-derived cell lines and tumors.

The gene ST7 has been proposed as the multi-tissue tumor-suppressor gene (TSG) at chromosome 7q31.1. However, we have sought and failed to detect the truncating mutations reported to exist in this gene.

Male genotype affects female longevity in Drosophila melanogaster.

Several recent studies suggest that interactions with conspecific males can reduce the longevity of female Drosophila melanogaster or support the idea that male and female fitness components are involved in antagonistic interactions. Here we report that males from third-chromosome isogenic lines demonstrated significant genetic variation in male reproductive performance and in the longevity of their mates. Increased male performance was marginally significantly associated with one measure of increased female survival rate. However, there was no indication of tradeoffs or negative correlations between male reproductive success and female survival. We discuss alternative hypotheses for the cause of the induced variation in female longevity.

Phenylboronic acid-salicylhydroxamic acid bioconjugates. 1. A novel boronic acid complex for protein immobilization.

A chemical affinity system exhibiting antibody-like properties is described. The system exploits bioconjugates with appended phenylboronic acid (PBA) moieties and a support-bound phenylboronic acid complexing reagent derived from salicylhydroxamic acid (SHA) for protein immobilization on a chromatographic support. The structure of the PBA.SHA complex was characterized by 11B NMR and mass spectrometry and compared with complexes derived from model compounds. Protein modification reagents were synthesized from 3-aminophenylboronic acid and utilized to prepare bioconjugates from alkaline phosphatase (AP) and horseradish peroxidase (HRP). AP obtained from one source afforded PBA bioconjugates exhibiting significant loss of enzymatic activity, whereas AP obtained from a second source afforded PBA bioconjugates exhibiting only a modest loss of enzymatic activity. Conversely, HRP afforded PBA bioconjugates exhibiting no loss of enzymatic activity. SHA-modified Sepharose was prepared by reaction of methyl 4-[(6-aminohexanoylamino)methyl]salicylate with CNBr-activated Sepharose 4B, followed by treatment with aqueous alkaline hydroxylamine. PBA-AP and PBA-HRP conjugates were efficiently immobilized on SHA-Sepharose at pH 8.3. PBA-AP conjugates were retained after washing with acidic buffers at pH 6.7, 4.2, and 2.5, whereas PBA-HRP conjugates were retained after washing with buffer at pH 6.7, but were eluted to some extent at and below pH 4.2. The results are interpreted in terms of multivalent interactions involving boronic acid complex formation between the enzyme bioconjugates and immobilized complexing reagent.

Phenylboronic acid-salicylhydroxamic acid bioconjugates. 2. Polyvalent immobilization of protein ligands for affinity chromatography.

Phenylboronic acid bioconjugates prepared from alkaline phosphatase by reaction with either 2,5-dioxopyrrolidinyl 3-[N-[3-(1,3,2-dioxaboran-2-yl)phenyl]carbamoyl]propanoate (PBA-XX-NHS) or 2,5-dioxopyrrolidinyl 6-[[3,5-di-(1,3,2-dioxaboran-2-yl)phenyl]carbonylamino]hexanoate (PDBA-X-NHS) were compared with respect to the efficiency with which they were immobilized on salicylhydroxamic acid-modified Sepharose (SHA-X-Sepharose) by boronic acid complex formation. When immobilized on moderate capacity SHA-X-Sepharose (5.4 micromol of SHA/mL of gel), PDBA-alkaline phosphatase conjugates were shown to be stable with respect to both the alkaline (pH 11.0) and acidic (pH 2.5) buffers utilized to recover anti-alkaline phosphatase during affinity chromatography. Boronic acid complex formation was compared to covalent immobilization of alkaline phosphatase on Affi-Gel 10 and Affi-Gel 15. PDBA-AP.SHA-X-Sepharose was shown to afford superior performance to both Affi-Gel 10 and Affi-Gel 15 with respect to immobilization of alkaline phosphatase, retention of anti-alkaline phosphatase and recovery of anti-alkaline phosphatase under alkaline conditions. High capacity SHA-X-Sepharose (> or = 7 micromol of SHA/mL of gel) was shown to afford superior performance to moderate capacity SHA-X-Sepharose (4.5 micromol of SHA/mL of gel) with respect to stability at pH 11.0 and pH 2.5 when a PDBA-alphaHuman IgG conjugate with a low incorporation ratio of only 1.5:1 was immobilized on SHA-X-Sepharose and subsequently utilized for affinity chromatography of Human IgG. The results are interpreted in terms of either a bivalent or trivalent interaction involving boronic acid complex formation.

Small molecule-based chemical affinity system for the purification of proteins.

A new chemical affinity system is described for the purification of proteins. The Linx Affinity Purification System enables researchers to quickly and easily bind a protein ligand to a chromatographic matrix and use the resulting affinity resin to purify a second protein from crude mixtures. The entire process takes approximately 2 h.

Salmon: a new autosomal mutation demonstrating incomplete dominance in the boine snake Boa constrictor.

An unusual and attractive pigmentation pattern mutation termed "salmon" has been identified in the United States in several captive colonies of the common neotropical boine snake boa constrictor [Boa constrictor (Boidae)]. Boa constrictors expressing the Sa pigmentation pattern appear to be restricted to regions of Panama. Animals with the Sa phenotype exhibit a sharp decrease in melanophore pigments (e.g., melanin) and an increase of xanthophore pigments (e.g., pteridines and carotenoids) throughout the body, including ventral and caudal regions. Moreover, the dorsal saddles (blotches) and lateral diamond patterns are greatly reduced and/or absent. Our study was initiated using a female B. constrictor born in captivity and expressed the Sa pigmentation pattern. Results from breeding experiments indicated an inheritance mode of autosomal incomplete dominance for the Sa and Wt alleles.

Toward reconciling inferences concerning genetic variation in senescence in Drosophila melanogaster.

Standard models for senescence predict an increase in the additive genetic variance for log mortality rate late in the life cycle. Variance component analysis of age-specific mortality rates of related cohorts is problematic. The actual mortality rates are not observable and can be estimated only crudely at early ages when few individuals are dying and at late ages when most are dead. Therefore, standard quantitative genetic analysis techniques cannot be applied with confidence. We present a novel and rigorous analysis that treats the mortality rates as missing data following two different parametric senescence models. Two recent studies of Drosophila melanogaster, the original analyses of which reached different conclusions, are reanalyzed here. The two-parameter Gompertz model assumes that mortality rates increase exponentially with age. A related but more complex three-parameter logistic model allows for subsequent leveling off in mortality rates at late ages. We find that while additive variance for mortality rates increases for late ages under the Gompertz model, it declines under the logistic model. The results from the two studies are similar, with differences attributable to differences between the experiments.

Bacteriophage and associated polysaccharide depolymerases--novel tools for study of bacterial biofilms.

Bacteriophage for three representative strains of Gram-negative biofilm bacteria have proved to be of widespread occurrence. Lytic bacteriophage have been isolated from local sewage for the bacterium 1.15, an exopolysaccharide (EPS)-producing pseudomonad found originally as a component of biofilms in a local river, and for two Enterobacter agglomerans strains from industrial biofilms. Representative examples of all three bacteriophage possess a relatively low burst size and on solid media, exhibit very large plaques surrounded by a wide halo (5-20 mm) indicative of polysaccharide depolymerase action. The bacteriophage are thus similar to other viruses for EPS-producing bacteria in inducing the synthesis of enzymes degrading the polymers which occlude the bacterial cell surface. In each preparation, the polysaccharase activity was associated both with sedimented phage particles and with the supernate of bacterial lysates. The enzymes have been partially purified and used to prepare polysaccharide digests in which the major products from each polysaccharide are the presumed repeat units of the polymers or oligomers of these. The soluble phage enzymes each degrade their substrate by acting as endo-glycanohydrolases. The phage and their associated enzymes thus provide very useful highly specific tools for studies of biofilms incorporating the bacterial host strains. Their potential applications in studies on bacterial biofilms are discussed.

The effects of spontaneous mutation on quantitative traits. II. Dominance of mutations with effects on life-history traits.

We studied the dominance of the effects of chromosomes carrying unselected mutations on five life-history traits in Drosophila melanogaster. Mutations were accumulated on the second chromosome for 44 generations in the absence of natural selection. Traits studied were female fecundity early and late in adult life, male mating ability, and male and female longevity. Homozygous effects were estimated for 50 mutant lines, and heterozygous effects were estimated by crossing these lines in a partial diallel scheme. Direct estimates of dominance showed that the effects of mutants are at least partially recessive. Heterozygotes had higher trait means than homozygotes in all five cases, and these differences were significant for late fecundity and female longevity. For all traits, genetic variance was larger among homozygous crosses than among heterozygous crosses. These results are consistent with those of many other studies that suggest that both unselected mutations and those found segregating in natural populations are partially recessive.