Gene drives may be capable of addressing ecological problems by altering entire populations of wild organisms, but their use has remained largely theoretical due to technical constraints. Here we consider the potential for RNA-guided gene drives based on the CRISPR nuclease Cas9 to serve as a general method for spreading altered traits through wild populations over many generations. We detail likely capabilities, discuss limitations, and provide novel precautionary strategies to control the spread of gene drives and reverse genomic changes. The ability to edit populations of sexual species would offer substantial benefits to humanity and the environment. For example, RNA-guided gene drives could potentially prevent the spread of disease, support agriculture by reversing pesticide and herbicide resistance in insects and weeds, and control damaging invasive species. However, the possibility of unwanted ecological effects and near-certainty of spread across political borders demand careful assessment of each potential application. We call for thoughtful, inclusive, and well-informed public discussions to explore the responsible use of this currently theoretical technology.

/pdf/concerning-rna-guided-gene-drives-for-the-alteration-of-wild-2la0voj6g0.pdf

Concerning RNA-guided gene drives for the alteration of wild populations

Site-specific selfish genes exploit host functions to copy themselves into a defined target DNA sequence, and include homing endonuclease genes, group II introns and some LINE-like transposable elements. If such genes can be engineered to target new host sequences, then they can be used to manipulate natural populations, even if the number of individuals released is a small fraction of the entire population. For example, a genetic load sufficient to eradicate a population can be imposed in fewer than 20 generations, if the target is an essential host gene, the knockout is recessive and the selfish gene has an appropriate promoter. There will be selection for resistance, but several strategies are available for reducing the likelihood of it evolving. These genes may also be used to genetically engineer natural populations, by means of population-wide gene knockouts, gene replacements and genetic transformations. By targeting sex-linked loci just prior to meiosis one may skew the population sex ratio, and by changing the promoter one may limit the spread of the gene to neighbouring populations. The proposed constructs are evolutionarily stable in the face of the mutations most likely to arise during their spread, and strategies are also available for reversing the manipulations.

/pdf/site-specific-selfish-genes-as-tools-for-the-control-and-56p3qs3uma.pdf

Site-specific selfish genes as tools for the control and genetic engineering of natural populations

Female Aedes aegypti mosquitoes infect more than 400 million people each year with dangerous viral pathogens including dengue, yellow fever, Zika and chikungunya. Progress in understanding the biology of mosquitoes and developing the tools to fight them has been slowed by the lack of a high-quality genome assembly. Here we combine diverse technologies to produce the markedly improved, fully re-annotated AaegL5 genome assembly, and demonstrate how it accelerates mosquito science. We anchored physical and cytogenetic maps, doubled the number of known chemosensory ionotropic receptors that guide mosquitoes to human hosts and egg-laying sites, provided further insight into the size and composition of the sex-determining M locus, and revealed copy-number variation among glutathione S-transferase genes that are important for insecticide resistance. Using high-resolution quantitative trait locus and population genomic analyses, we mapped new candidates for dengue vector competence and insecticide resistance. AaegL5 will catalyse new biological insights and intervention strategies to fight this deadly disease vector.

/pdf/improved-reference-genome-of-aedes-aegypti-informs-arbovirus-4twjg0mxs8.pdf

Improved reference genome of Aedes aegypti informs arbovirus vector control

Engineered gene drives - the process of stimulating the biased inheritance of specific genes - have the potential to enable the spread of desirable genes throughout wild populations or to suppress harmful species, and may be particularly useful for the control of vector-borne diseases such as malaria. Although several types of selfish genetic elements exist in nature, few have been successfully engineered in the laboratory thus far. With the discovery of RNA-guided CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats-CRISPR-associated 9) nucleases, which can be utilized to create, streamline and improve synthetic gene drives, this is rapidly changing. Here, we discuss the different types of engineered gene drives and their potential applications, as well as current policies regarding the safety and regulation of gene drives for the manipulation of wild populations.

/pdf/cheating-evolution-engineering-gene-drives-to-manipulate-the-429dkalaqo.pdf

Cheating evolution: engineering gene drives to manipulate the fate of wild populations

▪ Abstract Sex chromosome drive refers to the unequal transmission of X and Y chromosomes from individuals of the heterogametic sex, resulting in biased sex ratios among progeny and within populations. The presence of driving sex chromosomes can reduce mean fitness within a population, bring about intragenomic conflict between the X chromosome, the Y, and the autosomes, and alter the intensity or mode of sexual selection within species. Sex chromosome drive, or its genetic equivalent, is known in plants, mammals, and flies. Many species harboring driving X chromosomes have evolved Y-linked and autosomal suppressors of drive. If a drive polymorphism is not stable, then driving chromosomes may spread to fixation and cause the extinction of a species. Certain characteristics of species, such as population density and female mating rate, may affect the probability of fixation of driving chromosomes. Thus, sex chromosome drive could be an agent of species-level selection.

Sex Chromosome Meiotic Drive

A Giemsa C-banding technique applied to the mosquito, Aedes aegypti, has revealed a distinctive banding pattern which is described as a reliable means of distinguishing between the morphologically similar X and Y chromosomes during all stages of mitosis and meiosis. The essential difference is that the Y chromosome, unlike the X and the autosomes, is not C-banded in the centromere region. An intercalary band is also present in one arm of all X chromosomes and some Y chromosomes. The distribution of these cytological markers throughout meiosis indicates that the sex locus occurs somewhere within a pericentric region, the minimum extent of which includes both the intercalary band and the centromere.

X and Y chromosomes of Aedes aegypti (L.) distinguished by Giemsa C-banding.

Meiotic drive in Aedes aegypti (L.) is shown by a Giemsa C-banding technique to be associated with preferential isochromatid breakage of the X chromosome during male meiosis. These breaks remain open at least until anaphase-I and, since the range of cells affected is proportional to the sensitivity of the X chromosome to the Distorter gene, it is argued that they are directly related to the decreased number of spermatozoa found in distorting males. This reduction is considered to be attributable to the degeneration of more X- than Y-bearing spermatids but it is probable that some non-functional X-bearing spermatozoa are also produced. Chromosome breakage is almost completely confined to four sites, two adjacent to the centromere, one just proximal to the intercalary band and another about the centre of the unbanded arm. Although the first three of these lie within a region in which crossing-over does not take place, fragmentation occurs more frequently in a chiasmate arm than in one devoid of chromatid exchange.

A cytogenetic analysis of meiotic drive in the mosquito, Aedes aegypti (L.)

The separate identities of the male-determining factor, M, and the sex-linked Distorter gene, D, are established in an Accra strain of Aedes aegypti. Their to each other and to Giemsa C-bands. Thus, M is invariably inherited with the centromere, whereas D lies towards the intercalary band. Approximately 1.2% recombination occurs between M and D but, in a chromosome known for its distal localization of chiasmata, it is argued that he two are not necessarily as closely linked cytologically as this might imply. Evidence on the genetic effects of recombination in the region of M and D is also considered.

Cytological mapping of the M and D loci in the mosquito, Aedes aegypti (L.)

Giemsa C-banding of two liverwort species, Pellia epiphylla (L.) Corda (n=9) and P. neesiana (Gott.) Limpr. (n = X/Y + 8), identified each chromosome as unique. Almost the full complement of C-bands was defined by N-banding. Apart from minor variation in the prominence of certain bands, which were designated according to principles adopted in human cytology, the chief exceptions were the absence of bands 1p2, 1p4, 1q2, 1q4 and 7p6 from the N-banded karyotype of P. epiphylla and of band 7q25 from P. neesiana. Both quinacrine dihydrochloride and Hoechst 33258 showed reduced fluorescence in the constitutive heterochromatin of P. epiphylla. The effects of these two fluorochromes on P. neesiana, however, differed from each other and from their effect on P. epiphylla. Only one Q-band, the brightly fluorescing 1(X/Y)q24, occurred, whereas Hoechst 33258 resulted in considerable differentiation. The majority of C-bands were correlated with bright fluorescence with Hoechst 33258 but a few were dim. On the basis of these results, four major types of heterochromatin were identified in P. neesiana and two further types in P. epiphylla. They are discussed in the context of previously reported early replication of P. neesiana heterochromatin and point to considerable cytological evolution within these two species.

Heterochromatin diversity in two species of Pellia (Hepaticae) as revealed by C-, Q-, N- and Hoechst 33258-banding

In interphase cells of Aedes aegypti (L.) (2n=4+ XX/XY), only the nucleolus responded to selective silver staining. The secondary constriction on chromosome 3 remained unresponsive at all times but the six centromeres were identified throughout mitosis from early prophase as well as those stages of meiosis subsequent to diplotene. The centromeric blocks were not synonymous with the pericentric heterochromatin revealed by C-banding. X chromosomes without an intercalary C-band were newly discovered in Ae. aegypti in the Bangalore strain. Sequential Q-or Hoechst 33258/C-banding in this and the Trinidad-30 strain revealed intercalary heterochromatin diversity within and between strains and also differences between intercalary and pericentric heterochromatin.

M. E. Newton

Papers

X and Y chromosomes of Aedes aegypti (L.) distinguished by Giemsa C-banding.

A cytogenetic analysis of meiotic drive in the mosquito, Aedes aegypti (L.)

Cytological mapping of the M and D loci in the mosquito, Aedes aegypti (L.)

Heterochromatin diversity in two species of Pellia (Hepaticae) as revealed by C-, Q-, N- and Hoechst 33258-banding

Heterochromatin diversity and cyclic responses to selective silver staining in Aedes aegypti (L.)