Why More Biologists Must Embrace Quantitative Modeling.

Biology as a field has transformed since the time of its foundation from an organized enterprise cataloging the diversity of the natural world to a quantitatively rigorous science seeking to answer complex questions about the functions of organisms and their interactions with each other and their environments. As the mathematical rigor of biological analyses has improved, quantitative models have been developed to describe multi-mechanistic systems and to test complex hypotheses. However, applications of quantitative models have been uneven across fields, and many biologists lack the foundational training necessary to apply them in their research or to interpret their results to inform biological problem-solving efforts. This gap in scientific training has created a false dichotomy of "biologists" and "modelers" that only exacerbates the barriers to working biologists seeking additional training in quantitative modeling. Here, we make the argument that all biologists are modelers and are capable of using sophisticated quantitative modeling in their work. We highlight four benefits of conducting biological research within the framework of quantitative models, identify the potential producers and consumers of information produced by such models, and make recommendations for strategies to overcome barriers to their widespread implementation. Improved understanding of quantitative modeling could guide the producers of biological information to better apply biological measurements through analyses that evaluate mechanisms, and allow consumers of biological information to better judge the quality and applications of the information they receive. As our explanations of biological phenomena increase in complexity, so too must we embrace modeling as a foundational skill.

2019 UK National Guideline for consultations requiring sexual history taking : Clinical Effectiveness Group British Association for Sexual Health and HIV.

This guideline is an update of a previous version published in 2013. In this new version, we have reflected changes in the way sexual health services are now provided by assuming an integrated Sexual Health/Sexual and Reproductive Healthcare service. There are new recommendations for online testing, female genital mutilation (FGM), chemsex and considerations for transgender (and non-binary) individuals. Previous versions rather assumed a cis-gender clientele and so we have taken a more mechanistic approach to sex and risk without assuming gender identification. We have updated our gender terminology in line with the British Association for Sexual Health and HIV 'sexual health standards for trans, including non-binary, people' although have retained the terminology of 'men' and 'women' in a few cases where it related to other guidelines, e.g. human papillomavirus vaccination and FGM.

Standardized human bone marrow-derived stem cells infusion improves survival and recovery in a rat model of spinal cord injury.

Spinal cord injury (SCI) is an incurable disorder with an unmet need of an effective treatment. Recently, autologous human bone marrow-derived stem cells have shown to promote functional improvement, due to their anti-inflammatory and regenerative/apocrine properties. In this study, the primary objective was to test whether a single intrathecal injection with a 100 µL suspension of 400,000 fresh human bone marrow-derived CD34+ and an equal number of CD105+ stem cells (Neuro-Cells (NC)), one day after balloon-compression of the spinal cord, improves motor function and reduces secondary damage in immunodeficient rats. During the first 5 weeks after this intervention, NC significantly improved locomotor recovery and induced less injury-associated adverse events compared to vehicle-treated rats. Histological analysis showed that NC reduced astrogliosis, and apoptosis early after administration (day 4), but not at a later stage (day 56) after SCI. Proteomic studies (at day 56) pointed to the release of paracrine factors and identified proteins involved in regenerative processes. As stem cells seem to reach their effects in acute lesions by mainly suppressing (secondary) inflammation, it is thus realistic to expect a lower magnitude of their eventual beneficial effect in T-cell deficient rats, a fact reinforcing the robustness of Neuro-Cells efficacy. Taken together, this study indicates that an intrathecal instillation of Neuro-Cells holds great promise as a neuro-regenerative intervention in a clinical setting with acute SCI patients.

Alignment-free genome comparison enables accurate geographic sourcing of white oak DNA.

BACKGROUND: The application of genomic data and bioinformatics for the identification of restricted or illegally-sourced natural products is urgently needed. The taxonomic identity and geographic provenance of raw and processed materials have implications in sustainable-use commercial practices, and relevance to the enforcement of laws that regulate or restrict illegally harvested materials, such as timber. Improvements in genomics make it possible to capture and sequence partial-to-complete genomes from challenging tissues, such as wood and wood products. RESULTS: In this paper, we report the success of an alignment-free genome comparison method, [Formula: see text] that differentiates different geographic sources of white oak (Quercus) species with a high level of accuracy with very small amount of genomic data. The method is robust to sequencing errors, different sequencing laboratories and sequencing platforms. CONCLUSIONS: This method offers an approach based on genome-scale data, rather than panels of pre-selected markers for specific taxa. The method provides a generalizable platform for the identification and sourcing of materials using a unified next generation sequencing and analysis framework.

Evolutionary history of plant hosts and fungal symbionts predicts the strength of mycorrhizal mutualism.

Most plants engage in symbioses with mycorrhizal fungi in soils and net consequences for plants vary widely from mutualism to parasitism. However, we lack a synthetic understanding of the evolutionary and ecological forces driving such variation for this or any other nutritional symbiosis. We used meta-analysis across 646 combinations of plants and fungi to show that evolutionary history explains substantially more variation in plant responses to mycorrhizal fungi than the ecological factors included in this study, such as nutrient fertilization and additional microbes. Evolutionary history also has a different influence on outcomes of ectomycorrhizal versus arbuscular mycorrhizal symbioses; the former are best explained by the multiple evolutionary origins of ectomycorrhizal lifestyle in plants, while the latter are best explained by recent diversification in plants; both are also explained by evolution of specificity between plants and fungi. These results provide the foundation for a synthetic framework to predict the outcomes of nutritional mutualisms.

Erratum: Author Correction: Evolutionary history of plant hosts and fungal symbionts predicts the strength of mycorrhizal mutualism.

[This corrects the article DOI: 10.1038/s42003-018-0120-9.].

Disentangling genetic structure for genetic monitoring of complex populations.

Genetic monitoring estimates temporal changes in population parameters from molecular marker information. Most populations are complex in structure and change through time by expanding or contracting their geographic range, becoming fragmented or coalescing, or increasing or decreasing density. Traditional approaches to genetic monitoring rely on quantifying temporal shifts of specific population metrics-heterozygosity, numbers of alleles, effective population size-or measures of geographic differentiation such as FST. However, the accuracy and precision of the results can be heavily influenced by the type of genetic marker used and how closely they adhere to analytical assumptions. Care must be taken to ensure that inferences reflect actual population processes rather than changing molecular techniques or incorrect assumptions of an underlying model of population structure. In many species of conservation concern, true population structure is unknown, or structure might shift over time. In these cases, metrics based on inappropriate assumptions of population structure may not provide quality information regarding the monitored population. Thus, we need an inference model that decouples the complex elements that define population structure from estimation of population parameters of interest and reveals, rather than assumes, fine details of population structure. Encompassing a broad range of possible population structures would enable comparable inferences across biological systems, even in the face of range expansion or contraction, fragmentation, or changes in density. Currently, the best candidate is the spatial Λ-Fleming-Viot (SLFV) model, a spatially explicit individually based coalescent model that allows independent inference of two of the most important elements of population structure: local population density and local dispersal. We support increased use of the SLFV model for genetic monitoring by highlighting its benefits over traditional approaches. We also discuss necessary future directions for model development to support large genomic datasets informing real-world management and conservation issues.

Artificial urine and FBS supplemented media in cytocompatibility assays for PLGA-PEG-based intravesical devices using the urothelium cell line UROtsa.

European and German directives for approval of new medical devices require tests for cytotoxicity in relevant media, since urine can influence cytotoxicity of biodegradable devices. The aim of this study was to determine the long-term cytotoxicity of PLGA-b-mPEG (PLGA-PEG) polymer carriers and artificial urine (AU) to human UROtsa cells. Benign urothelial UROtsa cells were incubated in fetal bovine serum-containing RPMI 1640 medium supplemented with a range of concentrations of AU for 24 h and 7 days. Cell viability was determined by the XTT assay and by live/dead staining. The cytotoxicity of medium containing degradation products from PLGA-PEG carriers was also tested on the UROtsa cells in AU-containing and control medium. PLGA-PEG carriers exhibited no cytotoxicity to UROtsa cells after 24 h of incubation. However, after 7 days, cytotoxicity was observed, but this was largely attributable to the effects of 30% AU on the cells. Compared to phosphate buffer saline (PBS) and normalized to RPMI 1640 medium, significant cytotoxicity was observed by 24 h in medium containing 50% AU and by 7 days in medium containing 30% AU. Live/Dead staining confirmed proliferation results and no pH-changes could be observed. Here we demonstrate for the first time the impact of AU on standard cytotoxicity tests related to biomaterials for urinary-tract applications. Our study showed cytotoxic effects of high concentrations of 50% AU by 24 h and by physiological concentrations of AU (i.e., 30%) by 7 days. We have also demonstrated that PLGA-PEG has no cytotoxic effects in the appropriate AU-containing test environment. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2140-2147, 2018.

Two-component collagen nerve guides support axonal regeneration in the rat peripheral nerve injury model.

Progress in material development has enabled the production of nerve guides that increasingly resemble the characteristics of an autologous nerve graft. In the present study, 20 mm adult rat sciatic nerve defects were bridged with the collagen-based, two-component nerve guide 'Neuromaix', the commercially available NeuraGen® nerve tube or an autologous nerve graft. Neuromaix was able to support structural as well as functional regeneration across this gap. The majority of the axons grew across the scaffold into the distal nerve segment and retrograde tracing confirmed that these axons were of somatosensory and motor origin. Histomorphology revealed that axons regenerating through Neuromaix exhibited reduced myelin sheath thickness, whereas axon diameter and axon density were comparable to those of the autograft. Neuromaix implantation resulted in reinnervation of the gastrocnemius muscle to a level that was not significantly different from that supported by the autograft, as demonstrated by electrophysiology. Our findings show that the use of the Neuromaix scaffold not only allowed axonal regeneration across large nerve gaps, but that the regenerating axons were also able to functionally reinnervate the muscles. These data provide a promising perspective for the first in human application of the materials. Copyright © 2016 John Wiley & Sons, Ltd.

Home-field advantage? evidence of local adaptation among plants, soil, and arbuscular mycorrhizal fungi through meta-analysis.

BACKGROUND: Local adaptation, the differential success of genotypes in their native versus foreign environment, arises from various evolutionary processes, but the importance of concurrent abiotic and biotic factors as drivers of local adaptation has only recently been investigated. Local adaptation to biotic interactions may be particularly important for plants, as they associate with microbial symbionts that can significantly affect their fitness and may enable rapid evolution. The arbuscular mycorrhizal (AM) symbiosis is ideal for investigations of local adaptation because it is globally widespread among most plant taxa and can significantly affect plant growth and fitness. Using meta-analysis on 1170 studies (from 139 papers), we investigated the potential for local adaptation to shape plant growth responses to arbuscular mycorrhizal inoculation. RESULTS: The magnitude and direction for mean effect size of mycorrhizal inoculation on host biomass depended on the geographic origin of the soil and symbiotic partners. Sympatric combinations of plants, AM fungi, and soil yielded large increases in host biomass compared to when all three components were allopatric. The origin of either the fungi or the plant relative to the soil was important for explaining the effect of AM inoculation on plant biomass. If plant and soil were sympatric but allopatric to the fungus, the positive effect of AM inoculation was much greater than when all three components were allopatric, suggesting potential local adaptation of the plant to the soil; however, if fungus and soil were sympatric (but allopatric to the plant) the effect of AM inoculation was indistinct from that of any allopatric combinations, indicating maladaptation of the fungus to the soil. CONCLUSIONS: This study underscores the potential to detect local adaptation for mycorrhizal relationships across a broad swath of the literature. Geographic origin of plants relative to the origin of AM fungal communities and soil is important for describing the effect of mycorrhizal inoculation on plant biomass, suggesting that local adaptation represents a powerful factor for the establishment of novel combinations of fungi, plants, and soils. These results highlight the need for subsequent investigations of local adaptation in the mycorrhizal symbiosis and emphasize the importance of routinely considering the origin of plant, soil, and fungal components.

MycoDB, a global database of plant response to mycorrhizal fungi.

Plants form belowground associations with mycorrhizal fungi in one of the most common symbioses on Earth. However, few large-scale generalizations exist for the structure and function of mycorrhizal symbioses, as the nature of this relationship varies from mutualistic to parasitic and is largely context-dependent. We announce the public release of MycoDB, a database of 4,010 studies (from 438 unique publications) to aid in multi-factor meta-analyses elucidating the ecological and evolutionary context in which mycorrhizal fungi alter plant productivity. Over 10 years with nearly 80 collaborators, we compiled data on the response of plant biomass to mycorrhizal fungal inoculation, including meta-analysis metrics and 24 additional explanatory variables that describe the biotic and abiotic context of each study. We also include phylogenetic trees for all plants and fungi in the database. To our knowledge, MycoDB is the largest ecological meta-analysis database. We aim to share these data to highlight significant gaps in mycorrhizal research and encourage synthesis to explore the ecological and evolutionary generalities that govern mycorrhizal functioning in ecosystems.

Efficient bridging of 20 mm rat sciatic nerve lesions with a longitudinally micro-structured collagen scaffold.

An increasing number of biomaterial nerve guides has been developed that await direct comparative testing with the 'gold-standard' autologous nerve graft in functional repair of peripheral nerve defects. In the present study, 20 mm rat sciatic nerve defects were bridged with either a collagen-based micro-structured nerve guide (Perimaix) or an autologous nerve graft. Axons regenerated well into the Perimaix scaffold and, the majority of these axons grew across the 20 mm defect into the distal nerve segment. In fact, both the total axon number and the number of retrogradely traced somatosensory and motor neurons extending their axons across the implant was similar between Perimaix and autologous nerve graft groups. Implantation of Schwann cell-seeded Perimaix scaffolds provided only a beneficial effect on myelination within the scaffold. Functional recovery supported by the implanted, non-seeded Perimaix scaffold was as good as that observed after the autologous nerve graft, despite the presence of thinner myelin sheaths in the Perimaix implanted nerves. These findings support the potential of the Perimaix collagen scaffold as a future off-the-shelf device for clinical applications in selected cases of traumatic peripheral nerve injury.

Frequency-dependent selection drives HBeAg seroconversion in chronic hepatitis B virus infection.

HBeAg seroconversion is an important stage in the evolution of a chronic hepatitis B virus (HBV) infection that usually leads to control of viral replication and a reduced risk for liver cirrhosis and cancer. Since current therapies for the HBV-associated liver inflammation that is known as chronic hepatitis B (CHB). Rarely induce permanent HBeAg seroconversion, there is a need to understand the mechanisms responsible for the purpose of identifying new therapeutic targets. Currently, the most widely accepted hypothesis is that the patient's humoral and cellular immune responses to the HBV initiate HBeAg seroconversion. Although we accept that this hypothesis cannot be excluded, we propose an alternative that is consistent with published data on HBeAg seroconversion. We postulate, as others have, that the HBeAg suppresses the immune response to the HBV. However, production of the HBeAg incurs a metabolic cost to the hepatocyte which reduces the replicative capacity of the virus. Consequently, HBeAg-negative viruses replicate faster than HBeAg-positive viruses. HBeAg-negative variants arise de novo; and when their frequency in the population is low they have a replicative advantage. However, they also benefit from the immunosuppressive effects of the HBeAg-positive viruses in the population. As HBeAg-negative variants increase in frequency and HBeAg levels fall, the immune system recognizes the HBV, and HBeAg seroconversion occurs as a consequence of frequency-dependent selection acting on HBeAg-negative variants. This hypothesis explains the wide inter-individual variation in age of seroconversion, the increased rate of seroconversion during anti-viral treatment and the phenomena of both spontaneous and post-treatment HBeAg reversions (in which patients cycle between the HBeAg-positive and negative phases of their infection).

Repeat antenatal HIV testing in the third trimester: a study of feasibility and maternal uptake rates.

OBJECTIVES: The study aimed to assess the feasibility and acceptability of third-trimester antenatal HIV testing within our service after two cases of HIV seroconversion in pregnancy were noted in 2008. North American Guidelines recommend universal third-trimester HIV testing in areas with an HIV prevalence of more than 1 per 1000. The HIV prevalence rate in our area is 3.01 per 1000. METHODS: Pregnant women prior to 28 weeks of gestation were recruited at booking between 1 September 2008 and 31 August 2009 and offered an additional third-trimester HIV test. Consent was obtained and testing was performed by hospital and community midwives. Information was entered into a modified existing electronic maternity database. A qualitative e-mail survey of midwives investigated barriers to participation in the study. RESULTS: A total of 4134 women delivered; three (< 0.1%) declined first-trimester testing. Twenty-two women (0.5%) tested HIV positive, of whom six were newly diagnosed. Overall, 2934 of 4134 women (71%) were offered and accepted a third-trimester HIV test and had results available. Data were unavailable for 195 women (4.7%). A total of 663 of 4131 women (16%) were not offered a third-trimester test. Of 3273 women documented as having been offered a test, 3177 (97.1%) accepted. There were no positive third-trimester tests. Forty of 50 (80%) midwives surveyed responded with questionnaire feedback and cited lack of national policy and extra workload as barriers to performing third-trimester testing. CONCLUSIONS: Third-trimester testing was feasible and consent rates were high in those offered repeat testing. Third-trimester testing has the potential to prevent paediatric HIV infection and universal testing should be considered in high-prevalence areas.

Retrospective study of the effect of enhanced systematic sexually transmitted infection screening, facilitated by the use of electronic patient records, in an HIV-infected cohort.

OBJECTIVES: The aim of the study was to assess the impact of electronic checklists in enhancing sexually transmitted infection (STI) screening in routine HIV care. METHODS: This was a retrospective cohort study. In two HIV clinics, new STIs were recorded for three consecutive 12-month periods between 2009 and 2012 in a cohort of 882 HIV-infected patients. These three years coincided with the introduction of enhanced STI screening based on prompts within the electronic patient record (EPR) system. RESULTS: The number of diagnoses and the incidence of STIs more than doubled between 2010-2011 and 2011-2012 in both men who have sex with men (MSM) [from 18 of 115 (15%) to 42 of 132 (32%), a rise in STI incidence from 15.6 to 31.8/100 person-years; P < 0.001] and heterosexual patients [from six of 716 (0.8%) to 19 of 749 (2.5%), a rise in STI incidence from 0.8 to 2.5/100 person-years; P < 0.005]. The rise was significant in MSM for infections with chlamydia [from seven of 115 (6%) to 14 of 132 (11%), a rise in incidence from 6.0 to 10.6/100 person-years; P < 0.05], gonorrhoea [from five of 115 (4%) to 12 of 132 (9%), a rise in STI incidence from 4.3 to 9.1/100 person-years; P < 0.05] and early syphilis [from four of 115 (3%) to 13 of 132 (10%), a rise in incidence from 3.5 to 9.8/100 person-years; P < 0.001], but not for hepatitis C virus (HCV) and Lymphogranuloma venereum (LGV) infections. The rise was significant in heterosexual patients for infection with chlamydia [from four of 716 (0.6%) to 13 of 749 (1.7%), a rise in incidence from 0.6 to 1.7/100 person-years; P < 0.0001] but not for gonorrhoea, syphilis or Trichomonas vaginalis (TV). CONCLUSIONS: These data show that implementing systematic, frequent and routine STI screening led to a large increase in detected STIs in this HIV-infected cohort. This process is greatly enhanced by the use EPRs.

Motor outcome and allodynia are largely unaffected by novel olfactory ensheathing cell grafts to repair low-thoracic lesion gaps in the adult rat spinal cord.

Olfactory ensheathing cells (OEC) are a promising graftable cell population for improving functional outcomes after experimental spinal cord injury. However only few studies have focused on experimental models with large cavitations, which require bridging substrates to transfer and maintain the donor cells within the lesion site. Here, a state-of-the-art collagen-based multi-channeled three dimensional scaffold was used to deliver olfactory ensheathing cells to 2 mm long unilateral low-thoracic hemisection cavities. For a period of 10 weeks, allodynia of the hindpaws was monitored using the von Frey hair filament test, while an extensive analysis of motor ability was performed with use of the CatWalk gait analysis system and the BBB locomotor scale. No substantial improvement or deterioration of motor functions was induced and there was no effect on lesion-induced allodynia. On the basis of these data, we conclude that relatively large spinal cord lesions with cavitation may present additional hurdles to the therapeutic effect of OEC. Future studies are needed to address the nature that such lesion cavities place on cell grafts.

Spinal cord injury and its treatment: current management and experimental perspectives.

Clinical management of spinal cord injury (SCI) has significantly improved its general prognosis. However, to date, traumatic paraplegia and tetraplegia remain incurable, despite massive research efforts. Current management focuses on surgical stabilisation of the spine, intensive neurological rehabilitation, and the prevention and treatment of acute and chronic complications. Prevention remains the most efficient strategy and should be the main focus of public health efforts. Nevertheless, major advances in the understanding of the pathophysiological mechanisms of SCI open promising new therapeutic perspectives. Even if complete recovery remains elusive due to the complexity of spinal cord repair, a strategy combining different approaches may result in some degree of neurological improvement after SCI. Even slight neurological recovery can have high impact on the daily functioning of severely handicapped patients and, thus, result in significant improvements in quality of life.The main investigated strategies are: [1] initial neuroprotection, in order to decrease secondary injury to the spinal cord parenchyma after the initial insult; [2] spinal cord repair, in order to bridge the lesion site and reestablish the connection between the supraspinal centres and the deafferented cord segment below the lesion; and [3] re-training and enhancing plasticity of the central nervous system circuitry that was preserved or rebuilt after the injury.Now and in the future, treatment strategies that have both a convincing rationale and seen their efficacy confirmed reproducibly in the experimental setting must carefully be brought from bench to bedside. In order to obtain clinically significant results, their introduction into clinical research must be guided by scientific rigour, and their coordination must be rationally structured in a long-term perspective.

The importance of distinguishing between black Caribbeans and Africans in understanding sexual risk and care-seeking behaviours for sexually transmitted infections: evidence from a large survey of people attending genitourinary medicine clinics in England.

BACKGROUND: In the UK, black Caribbean and African populations experience disproportionately high rates of sexually transmitted infections (STIs) and HIV. Often studies do not differentiate between these populations notwithstanding differences in STI epidemiology and sociodemographics. METHODS: Patterns of care-seeking behaviour for STIs were explored separately for black Caribbean (n = 345), black African (n = 193) and white people through a cross-sectional survey of 2824 people attending five genitourinary medicine (GUM) clinics in England. RESULTS: Black Caribbean men were least likely to use, or try to use, their general practice surgery prior to GUM clinic attendance (16.6%). Symptomatic black Caribbean and African men were least likely to delay seeking care (30.8 and 26.3%, respectively). Symptomatic black Caribbean men faced the least provider delay in accessing care (27.3%). Black Caribbean men and women were most likely, and black African men and women least likely, to be diagnosed with an STI (49.7 and 32.0% versus 26.8 and 16.3%, respectively). Among symptomatic women, black Caribbeans and, among symptomatic men, black Africans were most likely to report abstaining from sex (46.3 and 73.1%, respectively). CONCLUSIONS: Our analyses highlight the importance of distinguishing between black ethnic groups and the need for future studies to ensure sufficiently large samples to permit such analyses.

The need for innovative sexually transmitted infection screening initiatives for young men: evidence from genitourinary medicine clinics across England.

The study objectives were to ascertain behavioural, access-related, health-seeking factors and sexually transmitted infection (STI) prevalence in young men (<25 years) attending genitourinary (GU) medicine clinics and compare them with older men (≥ 25 years) and young women (<25 years). Between October 2004 and March 2005, 4600 new attendees at seven sociodemographically and geographically contrasting GU medicine clinics across England completed questionnaires, which were linked to routine clinical data. Young men waited significantly less time to be seen in clinic compared with older men and young women. They were less likely to report symptoms than older men (P = 0.021) yet more likely to be diagnosed with chlamydia (P = 0.001) and gonorrhoea (P = 0.007). They were also more likely to be diagnosed with an acute STI relative to young women (P = 0.007). Our data confirm the need to make comprehensive STI screening readily available for young men and to develop effective and innovative screening strategies in different settings.

Evidence for reduced selection pressure on the hepatitis B virus core gene in hepatitis B e antigen-negative chronic hepatitis B.

The mechanisms underlying the high levels of hepatitis B virus (HBV) replication that cause hepatitis B e antigen (HBeAg)-negative chronic hepatitis B (e-CHB) are unknown. Impaired anti-HBV immunity, which may be measurable as a relaxation of selection pressure on the virus, is possible. A group of Tongans (n = 345) with a chronic HBV infection, including seven with e-CHB, were genotyped at HLA class I. The repertoire of HBV core-gene codons under positive selection pressure was defined by phylogenetic analysis (by using the paml program) of 708 cloned sequences extracted from the 67 of these 345 subjects with the same repertoire of HLA class I alleles as the seven e-CHB individuals and matched controls (see below). The frequency of non-synonymous mutations at these codons was measured in longitudinal data from 15 subjects. Finally, the number of non-synonymous mutations at these codons was compared in seven groups comprised of one subject with e-CHB and 1-3 HLA class I-matched controls with an inactive, HBeAg-negative chronic HBV infection (e-InD). Nineteen codons in the core gene were under positive selection pressure. There was a high frequency of new non-synonymous mutations at these codons (P<0.0001) in longitudinal data. The mean number of these 19 codons with non-synonymous mutations was lower (P = 0.02) in HBV from subjects with e-CHB (4.4±0.5 codons per subject) versus those with e-InD (6.4±0.4 codons per subject). There is a subtle relaxation in selection pressure on the HBV core gene in e-CHB. This may be due to impaired antiviral immunity, and could contribute to the high levels of viral replication that cause liver inflammation in this disease.