Rapid evolution and gene expression: a rapidly evolving Mendelian trait that silences field crickets has widespread effects on mRNA and protein expression.

A major advance in modern evolutionary biology is the ability to start linking phenotypic evolution in the wild with genomic changes that underlie that evolution. We capitalized on a rapidly evolving Hawaiian population of crickets (Teleogryllus oceanicus) to test hypotheses about the genomic consequences of a recent Mendelian mutation of large effect which disrupts the development of sound-producing structures on male forewings. The resulting silent phenotype, flatwing, persists because of natural selection imposed by an acoustically orienting parasitoid, but it interferes with mate attraction. We examined gene expression differences in developing wing buds of wild-type and flatwing male crickets using RNA-seq and quantitative proteomics. Most differentially expressed (DE) transcripts were down-regulated in flatwing males (625 up vs. 1716 down), whereas up- and down-regulated proteins were equally represented (30 up and 34 down). Differences between morphs were clearly not restricted to a single pathway, and we recovered annotations associated with a broad array of functions that would not be predicted a priori. Using a candidate gene detection test based on homology, we identified 30% of putative Drosophila wing development genes in the cricket transcriptome, but only 10% were DE. In addition to wing-related annotations, endocrine pathways and several biological processes such as reproduction, immunity and locomotion were DE in the mutant crickets at both biological levels. Our results illuminate the breadth of genetic pathways that are potentially affected in the early stages of adaptation.

Airflow accelerates bovine and human articular cartilage drying and chondrocyte death.

OBJECTIVE: Exposure of articular cartilage to static air results in changes to the extracellular matrix (ECM) and stimulates chondrocyte death, which may cause joint degeneration. However during open orthopaedic surgery, cartilage is often exposed to laminar airflow, which may exacerbate these damaging effects. We compared drying in static and moving air in terms of cartilage appearance, hydration and chondrocyte viability, and tested the ability of saline-saturated gauze to limit the detrimental effects of air exposure. DESIGN: Articular cartilage from bovine metatarsophalangeal joints (N = 50) and human femoral heads (N = 6) was exposed for 90 min to (1) static air (2) airflow (up to 0.34 m/s), or (3) airflow (0.18 m/s), covered with gauze. Following air exposure, cartilage was also rehydrated (0.9% saline; 120 min) to determine the reversibility of drying effects. The influence of airflow was assessed by studying macroscopic appearance, and quantifying superficial zone (SZ) chondrocyte viability and cartilage hydration. RESULTS: Airflow caused advanced changes to cartilage appearance, accelerated chondrocyte death, and increased dehydration compared to static air. These effects were prevented if cartilage was covered by saline-saturated gauze. Cartilage rehydration reversed macroscopic changes associated with drying but the chondrocyte death was not altered. Chondrocytes at the cut edge of cartilage were more sensitive to drying compared to cells distant from the edge. CONCLUSIONS: Airflow significantly increased articular cartilage dehydration and chondrocyte death compared to static air. As laminar airflow is routinely utilised in operating theatres, it is essential that articular cartilage is kept wet via irrigation or by covering with saline-saturated gauze to prevent chondrocyte death.

Transcriptional changes during Daphnia pulex development indicate that the maturation decision resembles a rate more than a threshold.

Maturation is a critical developmental process, and the age and size at which it occurs have important fitness consequences. Although maturation is remarkably variable, certain mechanisms, including a minimum size or state threshold, are proposed to underlie the process across a broad diversity of taxa. Recent evidence suggests that thresholds may themselves be developmentally plastic, and in the crustacean Daphnia pulex it is unclear whether maturation follows a threshold or is a gradual process more akin to a rate. Changes in gene expression across four instars before and during maturation were compared in a cDNA microarray experiment. Developmental stage was treated statistically both as a discontinuous and as a continuous variable, to determine whether genes showed gradual or discrete changes in expression. The continuous analysis identified a greater number of genes with significant differential expression (45) than the discontinuous analysis (11). The majority of genes, including those coding for histones, factors relating to transcription and cell cycle processes, and a putative developmental hormone showed continuous increases or decreases in expression from the first to the fourth instars that were studied, suggestive of a prolonged and gradual maturation process. Three genes coding for a fused vitellogenin/superoxide dismutase showed increases in expression following the second instar and coincided with the posited maturation threshold, but even their expression increased in a continuous fashion.

Solution-based nanosensors for in-field detection with the naked eye.

Nanomaterials are revolutionising analytical applications with low-cost tests that enable detecting a target molecule in a few steps and with the naked eye. With this approach, non-experts can perform analyses on-site and without utilising electronic readers. This is advantageous in point-of-care diagnostics, in-field measurements and analyses performed in resource-constrained settings. Here we review the main strategies adopted for detecting analytes with the naked eye and at the point of need using plasmonic nanosensors, catalytic nanoparticles and fluorescent nanomaterials. Examples of the detection of ions, glucose, small molecules, peptides and proteins with the nanosensors are explained in detail.

Understanding the dandruff scalp before and after treatment: an in vivo Raman spectroscopic study.

OBJECTIVES: To study and characterize the stratum corneum (SC) of dandruff scalp using in vivo Raman spectroscopy, to study how it compares with the non-dandruff scalp and to see the effect of treatment with a zinc pyrithione (ZnPTO)-based anti-dandruff shampoo. METHODS: The scalp skin was measured using a recently developed in vivo Raman probe. This method allows the inherent molecular components of the SC to be measured in vivo and confocally with depth, in particular the levels of natural moisturizing factors (NMF), lipids, lactic acid, urea and water. RESULTS: Depth-profile data for the skin components in dandruff SC in vivo are shown for the first time. The dandruff SC has lower NMF than the non-dandruff SC (0.16 compared with 0.39 a.u.), lower hydration, elevated levels of urea and lower levels of lactic acid. Treatment with an anti-dandruff shampoo containing 1% ZnPTO substantially restores the levels of each of these components close to the non-dandruff levels. Further to this, it is shown that sebum penetrates deeper into dandruff SC and at higher levels compared with non-dandruff SC. The levels of sebum localized within the SC are also brought closer to those of the non-dandruff condition after ZnPTO treatment. CONCLUSION: The in vivo Raman probe has allowed the direct measurement of dandruff-affected skin in situ for the first time. It has been shown that the dandruff SC is different from that of the non-dandruff scalp and that it is changed by treatment with shampoo containing ZnPTO and brought towards the characteristics of non-dandruff scalp. It offers novel insights into how the nature of a healthy scalp should be defined.

Wastewater-based epidemiology as a public health resource in low- and middle-income settings.

In the face of emerging and re-emerging diseases, novel and innovative approaches to population scale surveillance are necessary for the early detection and quantification of pathogens. The last decade has seen the rapid development of wastewater and environmental surveillance (WES) to address public health challenges, which has led to establishment of wastewater-based epidemiology (WBE) approaches being deployed to monitor a range of health hazards. WBE exploits the fact that excretions and secretions from urine, and from the gut are discharged in wastewater, particularly sewage, such that sampling sewage systems provides an early warning system for disease outbreaks by providing an early indication of pathogen circulation. While WBE has been mainly used in locations with networked wastewater systems, here we consider its value for less connected populations typical of lower-income settings, and in assess the opportunity afforded by pit latrines to sample communities and localities. We propose that where populations struggle to access health and diagnostic facilities, and despite several additional challenges, sampling unconnected wastewater systems remains an important means to monitor the health of large populations in a relatively cost-effective manner.

Detecting genes for variation in parasite burden and immunological traits in a wild population: testing the candidate gene approach.

Identifying the genes underlying phenotypic variation in natural populations can provide novel insight into the evolutionary process. The candidate gene approach has been applied to studies of a number of traits in various species, in an attempt to elucidate their genetic basis. Here, we test the application of the candidate gene approach to identify the loci involved in variation in gastrointestinal parasite burden, a complex trait likely to be controlled by many loci, in a wild population of Soay sheep. A comprehensive literature review, Gene Ontology databases, and comparative genomics resources between cattle and sheep were used to generate a list of candidate genes. In a pilot study, these candidates, along with 50 random genes, were then sequenced in two pools of Soay sheep; one with low gastrointestinal nematode burden and the other high, using a NimbleGen sequence capture experiment. Further candidates were identified from single nucleotide polymorphisms (SNPs) that were highly differentiated between high- and low-resistance sheep breeds. A panel of 192 candidate and control SNPs were then typed in 960 individual Soay sheep to examine whether they individually explained variation in parasite burden, as measured as faecal egg count, as well as two immune measures (Teladorsagia circumcincta-specific antibodies and antinuclear antibodies). The cumulative effect of the candidate and control SNPs were estimated by fitting genetic relationship matrices (GRMs) as random effects in animal models of the three traits. No more significant SNPs were identified in the pilot sequencing experiment and association study than expected by chance. Furthermore, no significant difference was found between the proportions of candidate or control SNPs that were found to be significantly associated with parasite burden/immune measures. No significant effect of the candidate or control gene GRMs was found. There is thus little support for the candidate gene approach to the identification of loci explaining variation in parasitological and immunological traits in this population. However, a number of SNPs explained significant variation in multiple traits and significant correlations were found between the proportions of variance explained by individual SNPs across multiple traits. The significant SNPs identified in this study may still, therefore, merit further investigation.

Wild rodents as a model to discover genes and pathways underlying natural variation in infectious disease susceptibility.

Individuals vary in their susceptibility to infectious disease, and it is now well established that host genetic factors form a major component of this variation. The discovery of genes underlying susceptibility has the potential to lead to improved disease control, through the identification and management of vulnerable individuals and the discovery of novel therapeutic targets. Laboratory rodents have proved invaluable for ascertaining the function of genes involved in immunity to infection. However, these captive animals experience conditions very different to the natural environment, lacking the genetic diversity and environmental pressures characteristic of natural populations, including those of humans. It has therefore often proved difficult to translate basic laboratory research to the real world. In order to further our understanding of the genetic basis of infectious disease resistance, and the evolutionary forces that drive variation in susceptibility, we propose that genetic research traditionally conducted on laboratory animals is expanded to the more ecologically valid arena of natural populations. In this article, we highlight the potential of using wild rodents as a new resource for biomedical research, to link the functional genetic knowledge gained from laboratory rodents with the variation in infectious disease susceptibility observed in humans and other natural populations.

Identification of putative markers of triclabendazole resistance by a genome-wide analysis of genetically recombinant Fasciola hepatica.

Despite years of investigation into triclabendazole (TCBZ) resistance in Fasciola hepatica, the genetic mechanisms responsible remain unknown. Extensive analysis of multiple triclabendazole-susceptible and -resistant isolates using a combination of experimental in vivo and in vitro approaches has been carried out, yet few, if any, genes have been demonstrated experimentally to be associated with resistance phenotypes in the field. In this review we summarize the current understanding of TCBZ resistance from the approaches employed to date. We report the current genomic and genetic resources for F. hepatica that are available to facilitate novel functional genomics and genetic experiments for this parasite in the future. Finally, we describe our own non-biased approach to mapping the major genetic loci involved in conferring TCBZ resistance in F. hepatica.

FORWARDS-1: an adaptive, single-blind, placebo-controlled ascending dose study of acute baclofen on safety parameters in opioid dependence during methadone-maintenance treatment-a pharmacokinetic-pharmacodynamic study.

BACKGROUND: Treatment of opiate addiction with opiate substitution treatment (e.g. methadone) is beneficial. However, some individuals desire or would benefit from abstinence but there are limited options to attenuate problems with opiate withdrawal. Preclinical and preliminary clinical evidence suggests that the GABA-B agonist, baclofen, has the desired properties to facilitate opiate detoxification and prevent relapse. This study aims to understand whether there are any safety issues in administering baclofen to opioid-dependent individuals receiving methadone. METHODS: Opiate-dependent individuals (DSM-5 severe opioid use disorder) maintained on methadone will be recruited from addiction services in northwest London (NHS and third sector providers). Participants will be medically healthy with no severe chronic obstructive pulmonary disease or type 2 respiratory failure, no current dependence on other substances (excluding nicotine), no current severe DSM-5 psychiatric disorders, and no contraindications for baclofen or 4800 IU vitamin D (placebo). Eligible participants will be randomised in a 3:1 ratio to receive baclofen or placebo in an adaptive, single-blind, ascending dose design. A Bayesian dose-escalation model will inform the baclofen dose (10, 30, 60, or 90 mg) based on the incidence of 'dose-limiting toxicity' (DLT) events and participant-specific methadone dose. A range of respiratory, cardiovascular, and sedative measures including the National Early Warning Score (NEWS2) and Glasgow Coma Scale will determine DLT. On the experimental day, participants will consume their usual daily dose of methadone followed by an acute dose of baclofen or placebo (vitamin D3) ~ 1 h later. Measures including oxygen saturation, transcutaneous CO2, respiratory rate, QTc interval, subjective effects (sedation, drug liking, craving), plasma levels (baclofen, methadone), and adverse events will be obtained using validated questionnaires and examinations periodically for 5 h after dosing. DISCUSSION: Study outcomes will determine what dose of baclofen is safe to prescribe to those receiving methadone, to inform a subsequent proof-of-concept trial of the efficacy baclofen to facilitate opiate detoxification. To proceed, the minimum acceptable dose is 30 mg of baclofen in patients receiving ≤ 60 mg/day methadone based on the clinical experience of baclofen's use in alcoholism and guidelines for the management of opiate dependence. TRIAL REGISTRATION: Clinicaltrials.gov NCT05161351. Registered on 16 December 2021.

City-wide wastewater genomic surveillance through the successive emergence of SARS-CoV-2 Alpha and Delta variants.

Genomic surveillance of SARS-CoV-2 has provided a critical evidence base for public health decisions throughout the pandemic. Sequencing data from clinical cases has helped to understand disease transmission and the spread of novel variants. Genomic wastewater surveillance can offer important, complementary information by providing frequency estimates of all variants circulating in a population without sampling biases. Here we show that genomic SARS-CoV-2 wastewater surveillance can detect fine-scale differences within urban centres, specifically within the city of Liverpool, UK, during the emergence of Alpha and Delta variants between November 2020 and June 2021. Furthermore, wastewater and clinical sequencing match well in the estimated timing of new variant rises and the first detection of a new variant in a given area may occur in either clinical or wastewater samples. The study's main limitation was sample quality when infection prevalence was low in spring 2021, resulting in a lower resolution of the rise of the Delta variant compared to the rise of the Alpha variant in the previous winter. The correspondence between wastewater and clinical variant frequencies demonstrates the reliability of wastewater surveillance. However, discrepancies in the first detection of the Alpha variant between the two approaches highlight that wastewater monitoring can also capture missing information, possibly resulting from asymptomatic cases or communities less engaged with testing programmes, as found by a simultaneous surge testing effort across the city.

Transcriptomic response of red grouse to gastro-intestinal nematode parasites and testosterone: implications for population dynamics.

A central issue in ecology is in understanding the relative influences of intrinsic and extrinsic effects on population regulation. Previous studies on the cyclic population dynamics of red grouse (Lagopus lagopus scoticus) have emphasized the destabilizing effects of either nematode parasites or territorial behaviour and aggression. The potential interacting effects of these processes, mediated through density-dependent, environmentally induced alterations of host immunocompetence influencing susceptibility to parasites have not been considered. Male red grouse at high density are more aggressive, associated with increased testosterone, which potentially could lead to reduced immunocompetence at a stage when parasites are most prevalent. This could depress individual condition, breeding performance and survival and thus drive or contribute to overall reductions in population size. Here, we characterize the transcriptomic response of grouse to nematode parasite infection and investigate how this is subsequently affected by testosterone, using a microarray approach contrasting red grouse with high and low parasite load at both high and low testosterone titre. A suite of 52 transcripts showed a significant level of up-regulation to either chronic parasite load or experimental parasite infection. Of these, 51 (98%) showed a reduced level of expression under conditions of high parasite load and high testosterone. The genes up-regulated by parasites and then down-regulated at high testosterone titre were not necessarily associated with immune response, as might be intuitively expected. The results are discussed in relation to the fitness and condition of individual red grouse and factors influencing the regulation of abundance in natural populations.

Rotation and translational motion prior to self-assembly: dynamics of ethanethiolate on Cu(111).

We investigate the dynamics of low-coverage ethanethiolate on Cu(111) using helium spin-echo spectroscopy. Above 210 K, the measurements are dominated by translational hopping with an activation energy of only 86 ± 5 meV. At lower temperatures (150-210 K) a further process becomes apparent which has the signature of confined motion. We demonstrate the experimental results are consistent with scattering from an anchored rotor, enabling identification of sixfold jump rotation of the ethyl tail group around a static sulfur adsorption site, with a rotational activation energy of 18 ± 8 meV. Our approach represents a new form of rotational spectroscopy which can be used to study rotational surface diffusion.

How effective is recognition of siblings on the basis of genotype?

The ability to recognize kin based on genetic markers has been widely proposed as a mechanism to facilitate altruistic behaviour and inbreeding avoidance. Siblings are an important group of relatives to discriminate from unrelated individuals but present a problem, because siblings can share 0, 1 or 2 alleles at any single recognition locus. Here, we present a Bayesian model of kin recognition that defines the potential for genotypic information to convey kinship. Under the direct comparison model, where the signaller's genotype is compared with that of the receiver, the odds ratio that a pair of individuals were siblings was substantially increased if they shared both alleles at a single locus, but only a minority of siblings were recognized; increasing the number of recognition loci used could not increase both the odds ratio and the proportion of siblings recognized. A maternal comparison model, where the signaller's genotype is compared with that of the receiver's mother, performed poorly when only a single recognition locus was considered, but became increasingly effective with more recognition loci. Nevertheless, incorporating partial-matching information across multiple, independent loci are likely to be difficult. Further empirical work needs to establish the mechanistic basis of genetic kin recognition used by different taxa.

Cognitive modularity and genetic disorders.

This study challenges the use of adult neuropsychological models for explaining developmental disorders of genetic origin. When uneven cognitive profiles are found in childhood or adulthood, it is assumed that such phenotypic outcomes characterize infant starting states, and it has been claimed that modules subserving these abilities start out either intact or impaired. Findings from two experiments with infants with Williams syndrome (a phenotype selected to bolster innate modularity claims) indicate a within-syndrome double dissociation: For numerosity judgments, they do well in infancy but poorly in adulthood, whereas for language, they perform poorly in infancy but well in adulthood. The theoretical and clinical implications of these results could lead to a shift in focus for studies of genetic disorders.

The effect of infection history on the fitness of the gastrointestinal nematode Strongyloides ratti.

SUMMARY: Hosts in nature will often acquire infections by different helminth species over their lifetime. This presents the potential for new infections to be affected (particularly via the host immune response) by a host's history of previous con- or hetero-specific infection. Here we have used an experimental rat model to investigate the consequences of a history of primary infection with either Nippostrongylus brasiliensis, Strongyloides venezuelensis or S. ratti on the fitness of, and immunological response to, secondary infections of S. ratti. We found that a history of con-specific, but not hetero-specific, infection reduced the survivorship of S. ratti; the fecundity of S. ratti was not affected by a history of either con- or hetero-specific infections. We also found that a history of con-specific infection promoted Th2-type responses, as shown by increased concentrations of total IgE, S. ratti-specific IgG1, rat mast cell protease II (RMCPII), IL4 (but decreased concentrations of IFNgamma) produced by mesenteric lymph node cells in response to S. ratti antigen. Additionally, S. ratti-specific IgG1 was positively related to the intensity of both primary and secondary infections of S. ratti. Hetero-specific primary infections were only observed to affect the concentration of total IgE and RMCPII. The overall conclusion of these experiments is that the major immunological effect acting against an infection is induced by the infection itself and that there is little effect of prior infections of the host.

An open study to evaluate topical treatment of equine chorioptic mange with shampooing and lime sulphur solution.

Chorioptic mange caused by Chorioptes bovis is a common pruritic skin condition of the horse. This surface-browsing parasite usually affects the lower legs (leg mange) but can present as a generalized skin disease. Numerous anecdotal reports exist in the literature about the benefit of lime sulphur as a treatment for surface ectoparasites in the horse. This report studies the use of lime sulphur when applied as a 5% solution, some with and some without prior shampooing and clipping, to treat confirmed cases of chorioptic mange in 22 horses. Horses included in the trial had clinical signs indicative of chorioptic mange and positive identification of chorioptic mites on skin scrapings and tape preparations. Each horse was treated with sulphurated lime dip solution four times at 7-day intervals. Most horses were clipped and/or shampooed prior to treatment. Animals were assigned a score based on a scale of 1-10 to assess the severity of their lesions and degree of behavioural signs. The horses were again scored and examined for mites after four treatments. All animals showed a reduction in scores at the end of the trial and mites were not demonstrated from any animal.

Parasite interactions in natural populations: insights from longitudinal data.

The physiological and immunological state of an animal can be influenced by current infections and infection history. Consequently, both ongoing and previous infections can affect host susceptibility to another parasite, the biology of the subsequent infection (e.g. infection length) and the impact of infection on host morbidity (pathology). In natural populations, most animals will be infected by a succession of different parasites throughout the course of their lives, with probably frequent concomitant infections. The relative timing of different infections experienced by a host (i.e. the sequence of infection events), and the effects on factors such as host susceptibility and host survival, can only be derived from longitudinal data on individual hosts. Here we review some of the evidence for the impact of co-infection on host susceptibility, infection biology and pathology focusing on insights obtained from both longitudinal studies in humans and experiments that explicitly consider the sequence of infection. We then consider the challenges posed by longitudinal infection data collected from natural populations of animals. We illustrate their usefulness using our data of microparasite infections associated with field vole (Microtus agrestis) populations to examine impacts on susceptibility and infection length. Our primary aim is to describe an analytical approach that can be used on such data to identify interactions among the parasites. The preliminary analyses presented here indicate both synergistic and antagonistic interactions between microparasites within this community and emphasise that such interactions could have significant impacts on host-parasite fitness and dynamics.

Fat transforms ascorbic acid from inhibiting to promoting acid-catalysed N-nitrosation.

BACKGROUND: The major potential site of acid nitrosation is the proximal stomach, an anatomical site prone to a rising incidence of metaplasia and adenocarcinoma. Nitrite, a pre-carcinogen present in saliva, can be converted to nitrosating species and N-nitroso compounds by acidification at low gastric pH in the presence of thiocyanate. AIMS: To assess the effect of lipid and ascorbic acid on the nitrosative chemistry under conditions simulating the human proximal stomach. METHODS: The nitrosative chemistry was modelled in vitro by measuring the nitrosation of four secondary amines under conditions simulating the proximal stomach. The N-nitrosamines formed were measured by gas chromatography-ion-trap tandem mass spectrometry, while nitric oxide and oxygen levels were measured amperometrically. RESULTS: In absence of lipid, nitrosative stress was inhibited by ascorbic acid through conversion of nitrosating species to nitric oxide. Addition of ascorbic acid reduced the amount of N-nitrosodimethylamine formed by fivefold, N-nitrosomorpholine by >1000-fold, and totally prevented the formation of N-nitrosodiethylamine and N-nitrosopiperidine. In contrast, when 10% lipid was present, ascorbic acid increased the amount of N-nitrosodimethylamine, N-nitrosodiethylamine and N-nitrosopiperidine formed by approximately 8-, 60- and 140-fold, respectively, compared with absence of ascorbic acid. CONCLUSION: The presence of lipid converts ascorbic acid from inhibiting to promoting acid nitrosation. This may be explained by nitric oxide, formed by ascorbic acid in the aqueous phase, being able to regenerate nitrosating species by reacting with oxygen in the lipid phase.

Brainstem auditory evoked responses in 37 dogs with otitis media before and after topical therapy.

OBJECTIVES: The objective of this study was to determine whether intra-aural administration of aqueous solutions of marbofloxacin, gentamicin, tobramycin and ticarcillin (used off-licence) was associated with changes in hearing as measured by brainstem auditory evoked responses. MATERIALS AND METHODS: Dogs diagnosed with otitis media (n=37) underwent brainstem auditory evoked response testing and then were treated for their ear disease. First, the external ear canal and middle ear were flushed with sterile saline followed by EDTA tris with 0·15% chlorhexidine. Then, a combination of aqueous antibiotic mixed with an aqueous solution of EDTA tris was instilled into the middle ear. Follow-up examinations were undertaken for each dog, and treatment was continued until there were no detected infectious organisms or inflammatory infiltrate. Brainstem auditory evoked response testing was repeated after resolution of the infection and discontinuation of therapy. RESULTS: Brainstem auditory evoked responses in dogs treated with aqueous solutions of marbofloxacin or gentamicin remained unchanged or improved after therapy of otitis media but were impaired in dogs treated with ticarcillin or tobramycin. CLINICAL SIGNIFICANCE: If off-licence use of topical antibiotics is deemed necessary in cases of otitis media, aqueous solutions of marbofloxacin and gentamicin appear to be less ototoxic than aqueous solutions of ticarcillin or tobramycin.