Allelic bias when performing in-solution enrichment of ancient human DNA

In-solution hybridisation enrichment of genetic variation is a valuable methodology in human paleogenomics. It allows enrichment of endogenous DNA by targeting genetic markers that are comparable between sequencing libraries. Many studies have used the 1240k reagent—which enriches 1,237,207 genome-wide SNPs—since 2015, though access was restricted. In 2021, Twist Biosciences and Daicel Arbor Biosciences independently released commercial kits that enabled all researchers to perform enrichments for the same 1240 k SNPs. We used the Daicel Arbor Biosciences Prime Plus kit to enrich 132 ancient samples from three continents. We identified a systematic assay bias that increases genetic similarity between enriched samples and that cannot be explained by batch effects. We present the impact of the bias on population genetics inferences (e.g. Principal Components Analysis, ƒ-statistics) and genetic relatedness (READ). We compare the Prime Plus bias to that previously reported of the legacy 1240k enrichment assay. In ƒ-statistics, we find that all Prime-Plus-generated data exhibit artefactual excess shared drift, such that within-continent relationships cannot be correctly determined. The bias is more subtle in READ, though interpretation of the results can still be misleading in specific contexts. We expect the bias may affect analyses we have not yet tested. Our observations support previously reported concerns for the integration of different data types in paleogenomics. We also caution that technological solutions to generate 1240k data necessitate a thorough validation process before their adoption in the paleogenomic community.     

Local adaptation and pronounced genetic differentiation in an extremophile fish, Poecilia mexicana, inhabiting a Mexican cave with toxic hydrogen sulphide

We investigated genetic differentiation and migration patterns in a small livebearing fish, Poecilia mexicana, inhabiting a sulfidic Mexican limestone cave (Cueva del Azufre). We examined fish from three different cave chambers, the sulfidic surface creek draining the cave (El Azufre) and a nearby surface creek without the toxic hydrogen sulphide (Arroyo Cristal). Using microsatellite analysis of 10 unlinked loci, we found pronounced genetic differentiation among the three major habitats: Arroyo Cristal, El Azufre and the cave. Genetic differentiation was also found within the cave between different pools. An estimation of first-generation migrants suggests that (i) migration is unidirectional, out of the cave, and (ii) migration among different cave chambers occurs to some extent. We investigated if the pattern of genetic differentiation is also reflected in a morphological trait, eye size. Relatively large eyes were found in surface habitats, small eyes in the anterior cave chambers, and the smallest eyes were detected in the innermost cave chamber (XIII). This pattern shows some congruence with a previously proposed morphocline in eye size. However, our data do not support the proposed mechanism for this morphocline, namely that it would be maintained by migration from both directions into the middle cave chambers. This would have led to an increased variance in eye size in the middle cave chambers, which we did not find. Restricted gene flow between the cave and the surface can be explained by local adaptations to extreme environmental conditions, namely H2S and absence of light. Within the cave system, habitat properties are patchy, and genetic differentiation between cave chambers despite migration could indicate local adaptation at an even smaller scale.     

Adeno-Associated Virus Type 2 Rep68 Can Bind to Consensus Rep-Binding Sites on the Herpes Simplex Virus 1 Genome

Adeno-associated virus type 2 is known to inhibit replication of herpes simplex virus 1 (HSV-1). This activity has been linked to the helicase- and DNA-binding domains of the Rep68/Rep78 proteins. Here, we show that Rep68 can bind to consensus Rep-binding sites on the HSV-1 genome and that the Rep helicase activity can inhibit replication of any DNA if binding is facilitated. Therefore, we hypothesize that inhibition of HSV-1 replication involves direct binding of Rep68/Rep78 to the HSV-1 genome.     

An indigenous religious ritual selects for resistance to a toxicant in a livebearing fish

Human-induced environmental change can affect the evolutionary trajectory of populations. In Mexico, indigenous Zoque people annually introduce barbasco, a fish toxicant, into the Cueva del Azufre to harvest fish during a religious ceremony. Here, we investigated tolerance to barbasco in fish from sites exposed and unexposed to the ritual. We found that barbasco tolerance increases with body size and differs between the sexes. Furthermore, fish from sites exposed to the ceremony had a significantly higher tolerance. Consequently, the annual ceremony may not only affect population structure and gene flow among habitat types, but the increased tolerance in exposed fish may indicate adaptation to human cultural practices in a natural population on a very small spatial scale.     

Population genetic simulation: Benchmarking frameworks for non-standard models of natural selection

Population genetic simulation has emerged as a common tool for investigating increasingly complex evolutionary and demographic models. Software capable of handling high-level model complexity has recently been developed, and the advancement of tree sequence recording now allows simulations to merge the efficiency and genealogical insight of coalescent simulations with the flexibility of forward simulations. However, frameworks utilizing these features have not yet been compared and benchmarked. Here, we evaluate various simulation workflows using the coalescent simulator msprime and the forward simulator SLiM, to assess resource efficiency and determine an optimal simulation framework. Three aspects were evaluated: (1) the burn-in, to establish an equilibrium level of neutral diversity in the population; (2) the forward simulation, in which temporally fluctuating selection is acting; and (3) the final computation of summary statistics. We provide typical memory and computation time requirements for each step. We find that the fastest framework, a combination of coalescent and forward simulation with tree sequence recording, increases simulation speed by over twenty times compared to classical forward simulations without tree sequence recording, although it does require six times more memory. Overall, using efficient simulation workflows can lead to a substantial improvement when modelling complex evolutionary scenarios—although the optimal framework ultimately depends on the available computational resources.     

Sex-specific SOD levels and DNA damage in painted dragon lizards (Ctenophorus pictus)

When groups of individuals differ in activities that may influence the production of reactive molecules, such as superoxide, we expect selection to result in congruent upregulation of antioxidant production in the group(s) most at risk of suffering concomitant erosion of essential tissue and biomolecules, such as DNA. We investigate this in a (near) annual lizard species, the Australian painted dragon (Ctenophorus pictus), in which males and females have fundamentally different lifestyles, with males being overtly conspicuous and aggressive, whereas females are placid and camouflaged. When kept in identical conditions to females in captivity, males had higher levels of superoxide dismutase (SOD) through the activity season, which is consistent with selection for a higher capacity of superoxide antioxidation and a lower level of DNA damage than females. Males, however, lacked the clear negative, linear relationship between SOD and DNA erosion observed in females, suggesting that female upregulation of SOD results in a more predictable antioxidation and a more immediate target for selection. Lastly, we analysed aspects of female reproduction from a DNA erosion perspective. Females closer to ovulation, hence with less remaining, circulating vitellogenin, had higher superoxide levels. Furthermore, a multiple regression analysis showed that females that produced more clutches over time suffered more DNA erosion, whereas females with higher SOD levels suffered less DNA erosion.     

Digit ratio, color polymorphism and egg testosterone in the Australian painted dragon

Variation in exposure to sex hormones during early development contributes to phenotypic plasticity in vertebrate offspring. As a proposed marker for prenatal sex hormone exposure and because of their association with various physiological and behavioral characteristics, digit ratio and/or digit length have received notable interest within the field of evolutionary ecology. However, the validity of digit measures as a proxy of prenatal sex hormone exposure is controversial and only few studies have provided direct evidence for the link between digit development and prenatal sex hormones. Here, we report morph- and sex-specific variation in digit ratio in wild painted dragon lizards (Ctenophorus pictus). Lizards expressing a yellow bib have significantly larger third-to-fourth toe ratios (3D:4D) than lizards without a bib. Males have significantly smaller 3D:4D than females. Furthermore, we show that experimental elevation of yolk testosterone significantly increases 3D:4D in hatchling painted dragon lizards, but has no influence on hatchling size. Our results provide direct and indirect evidence for the involvement of prenatal sex steroids in digit development and it is suggested that digit ratio may be used as a biomarker for prenatal steroid exposure in this reptilian species. As such, digit ratio may provide a useful tool to study temporal or spatial differences in the proximate hormonal mechanisms modulating physiological and behavioural phenotypes.     

Influence of black spot disease on shoaling behaviour in female western mosquitofish, <Emphasis Type="Italic">Gambusia affinis</Emphasis> (Poeciliidae,Teleostei)

Parasites can fundamentally alter the cost–benefit ratio of living in a group, e.g. if infected individuals increase the predation risk of shoal mates. Here, the effect of an infection with a trematode, Uvulifer sp. (Diplostomatidae) on the shoaling behaviour of female western mosquitofish, Gambusia affinis, was investigated. The parasite examined causes a direct phenotypical change of the host by forming black spots on its body surface. When given a choice between a stimulus shoal and no shoal, we found shoaling tendencies to be significantly reduced in infected focal fish. In another experiment, we tested for association preferences relative to the infection status of the stimulus fish. Given the choice between an infected and a healthy stimulus fish, both infected and healthy focal fish preferred to associate with non-infected stimulus fish. Our results suggest that (1) the cost–benefit ratio of shoaling might be different for infected and non-infected individuals. Infected fish may be more affected by competition for food within a shoal. (2) Associating with infected conspecifics appears to be costly for female mosquitofish, maybe due to increased predation risk.