Anti-lipopolysaccharide factors (ALFs) are antimicrobial peptides with a central β-hairpin structure able to bind to microbial components. Mining sequence databases for ALFs allowed us to show the remarkable diversity of ALF sequences in shrimp. We found at least seven members of the ALF family (Groups A to G), including two novel Groups (F and G), all of which are encoded by different loci with conserved gene organization. Phylogenetic analyses revealed that gene expansion and subsequent diversification of the ALF family occurred in crustaceans before shrimp speciation occurred. The transcriptional profile of ALFs was compared in terms of tissue distribution, response to two pathogens and during shrimp development in Litopenaeus vannamei, the most cultivated species. ALFs were found to be constitutively expressed in hemocytes and to respond differently to tissue damage. While synthetic β-hairpins of Groups E and G displayed both antibacterial and antifungal activities, no activity was recorded for Group F β-hairpins. Altogether, our results showed that ALFs form a family of shrimp AMPs that has been the subject of intense diversification. The different genes differ in terms of tissue expression, regulation and function. These data strongly suggest that multiple selection pressures have led to functional diversification of ALFs in shrimp.

Protozoa and fungi are known to have extraordinarily diverse mechanisms of genetic exchange. However, the presence and epidemiological relevance of genetic exchange in Trypanosoma cruzi, the agent of Chagas disease, has been controversial and debated for many years. Field studies have identified both predominantly clonal and sexually recombining natural populations. Two of six natural T. cruzi lineages (TcV and TcVI) show hybrid mosaicism, using analysis of single-gene locus markers. The formation of hybrid strains in vitro has been achieved and this provides a framework to study the mechanisms and adaptive significance of genetic exchange. Using whole genome sequencing of a set of experimental hybrids strains, we have confirmed that hybrid formation initially results in tetraploid parasites. The hybrid progeny showed novel mutations that were not attributable to either (diploid) parent showing an increase in amino acid changes. In long-term culture, up to 800 generations, there was a variable but gradual erosion of progeny genomes towards triploidy, yet retention of elevated copy number was observed at several core housekeeping loci. Our findings indicate hybrid formation by fusion of diploid T. cruzi, followed by sporadic genome erosion, but with substantial potential for adaptive evolution, as has been described as a genetic feature of other organisms, such as some fungi.