Parasite and virus dynamics in the honeybee Apis mellifera unicolor on a tropical island recently invaded by Varroa destructor.

In La Réunion, the established honeybee subspecies Apis mellifera unicolor, an endemic subspecies of African lineage, is facing considerable challenges. Since the introduction of the Varroa destructor mite in 2017 high colony losses have been recorded. We investigated the dynamics of V. destructor and two viruses, the Deformed Wing Virus (DWV), known to be transmitted by the mite, and the Chronic Bee Paralysis Virus (CBPV), in A. m. unicolor. Colonies from two apiaries located at 300 and 900 m a.s.l were monitored twice for one year without any acaricide treatment. The brood area, V. destructor infestation rates, DWV and CBPV prevalence and load were recorded monthly. A. m. unicolor maintained brood rearing throughout the year. Varroa destructor infestation resulted in high colony mortality (up to 85 %) and high phoretic mite rates (up to 52 mites per hundred bees). The establishment of DWV in colonies occurred after that of V. destructor and the mite infestation rate had a significant effect on the virus prevalence and load. CBPV appeared only transiently throughout the surveys. The data showed that, in tropical colonies with permanent brood rearing, V. destructor and DWV can reach high levels, but are still subject to seasonal variations that appear to be influenced by environmental conditions. This suggests that beekeeping practices could be adapted by favouring sites and periods for transhumance or acaricide treatment.

Development and characterization of twenty microsatellite markers for Phelsuma inexpectata (Squamata: Gekkonidae), a critically endangered gecko endemic to Reunion Island.

BACKGROUND: The Manapany day gecko (Phelsuma inexpectata) is endemic to the south of Reunion Island. Threatened by habitat fragmentation and loss, human activities and invasive species, P. inexpectata is considered as critically endangered. Conservation measures are required but data on the species are missing, notably on its genetic diversity and population structure for which no specific markers are available to date. Here, we aimed to develop molecular markers to allow genetic studies of P. inexpectata. METHODS AND RESULTS: We developed and characterized 20 polymorphic microsatellite markers based on 23 P. inexpectata individuals sampled from 10 sites. Then, the markers were tested on a total of 101 individuals, 30 from a natural site and 71 from an anthropized site. The mean values of Na, Ho and He were 2.3 (± 0.2), 0.353 (± 0.053) and 0.345 (± 0.046) in the natural site and 2.8 (± 0.3), 0.345 (± 0.051) and 0.338 (± 0.048) in the anthropized site, respectively. Based on the combined loci, the probability of identity (PID) for unrelated specimens were 2.7 × 10-7 and 2.6 × 10-7 in the natural and anthropized site, respectively. CONCLUSIONS: This work provides the first set of microsatellite markers for P. inexpectata, constituting a valuable tool to conduct classical genetic studies on the species, such as estimating genetic diversity, population structure and kinship relationships among individuals. Such studies will provide relevant information on P. inexpectata and will therefore be helpful in the implementation of conservation measures for this threatened species.

Large-scale mitochondrial DNA analysis of native honey bee Apis mellifera populations reveals a new African subgroup private to the South West Indian Ocean islands.

BACKGROUND: The South West Indian Ocean (SWIO) archipelagos and Madagascar constitute a hotspot of biodiversity with a high rate of endemism. In this area, the endemic subspecies A. m. unicolor has been described in Madagascar. It belongs to the African lineage, one of the four described evolutionary lineages in honey bees. Despite a long beekeeping tradition and several recorded European introductions, few studies have been carried out on the diversity and proportion of honey bee subspecies. In order to identify and define which evolutionary lineages and potential sub-lineages are present in the SWIO, the COI-COII intergenic region and the ND2 gene of the mtDNA were sequenced in honey bee colonies from three archipelagos. An extensive sampling (n = 1184 colonies) was done in the Mascarene (La Réunion, Mauritius, Rodrigues), Seychelles (Mahé, Praslin, La Digue) and Comoros (Grande Comore, Mohéli, Anjouan, Mayotte) archipelagos. Islands genetic diversity was compared to newly sampled populations from Madagascar, continental African and European populations. RESULTS: African lineage haplotypes were found in all islands (except for Rodrigues). Madagascar, Comoros and Seychelles had 100% of A lineage, 95.5% in La Réunion and 56.1% in Mauritius. Among all African colonies detected in the SWIO, 98.1% (n = 633) of COI-COII haplotypes described the presence of the subspecies A. M. unicolor. Both genetic markers revealed i) a new private AI mitochondrial group shared by the SWIO archipelagos and Madagascar distant from continental populations; ii) the private African haplotypes for each island suggested diversity radiation in the archipelagos; iii) the detection of the Comoros archipelago as a possible contact area between insular and continental African populations. The exotic European C and M lineages were only detected in the Mascarene archipelago, but striking differences of proportion were observed among islands. Merely 4.6% of European colonies were found in La Réunion whereas Mauritius cumulated 44%. Here, among the 84 observed COI-COII haplotypes, 50 were newly described including 13 which were private to the SWIO archipelagos and Madagascar. Similarly, 24 of the 34 found ND2 haplotypes were novel which included six haplotypes particular to the SWIO populations. CONCLUSION: A new African subgroup was described in the SWIO region with mitochondrial genetic evidence that A. m. unicolor is the indigenous subspecies of the archipelagos surrounding Madagascar.

Spread and strain determination of Varroa destructor (Acari: Varroidae) in Madagascar since its first report in 2010.

Varroa destructor is a major pest in world beekeeping. It was first detected in Madagascar in 2010 on the endemic honeybee Apis mellifera unicolor. To evaluate V. destructor spread dynamics in Madagascar a global survey was conducted in 2011-2012. A total of 695 colonies from 30 districts were inspected for the presence of mites. 2 years after its introduction, nine districts were found infested. Varroa destructor spread was relatively slow compared to other countries with a maximum progression of 40 km per year, the five newly infested districts being located next to the first infested ones. The incidence of mite infestation was also investigated by monitoring 73 colonies from five apiaries during 1 year (2011-2012). Sixty percent of local colony mortality was recorded after 1 year of survey. Varroa destructor strain determination was done by partial sequencing of the cytochrome oxidase I gene of 13 phoretic mites sampled in five districts. A single V. destructor mitochondrial haplotype was detected, the Korean type, also present in the closest African countries. A global pathogen survey was also conducted on the colonies inspected for mite presence. The greater wax moth, Galleria mellonella has been found in all colonies all over the country. Two other pathogens and morphological abnormalities in workers, such as deformed wings, were found associated with only V. destructor presence. A prevention management plan must be implemented to delimit mite spread across the island.

Geographic population structure of the honeybee microsporidian parasite Vairimorpha (Nosema) ceranae in the South West Indian Ocean.

The microsporidian Vairimorpha (Nosema) ceranae is one of the most common parasites of the honeybee. A single honeybee carries many parasites and therefore multiple alleles of V. ceranae genes that seem to be ubiquitous. As a consequence, nucleotide diversity analyses have not allowed discriminating genetic structure of parasite populations. We performed deep loci-targeted sequencing to monitor the haplotype frequencies of genome markers in isolates from discontinuous territories, namely the tropical islands of the South West Indian Ocean. The haplotype frequency distribution corroborated the suspected tetraploidy of the parasite. Most major haplotypes were ubiquitous in the area but with variable frequency. While oceanic isolates differed from European and Asian outgroups, parasite populations from distinct archipelagoes also differed in their haplotype distribution. Interestingly an original and very divergent Malagasy isolate was detected. The observed population structure allowed formulating hypotheses upon the natural history of V. ceranae in this oceanic area. We also discussed the usefulness of allelic distribution assessment, using multiple informative loci or genome-wide analyses, when parasite population is not clonal within a single host.

Experience influences aggressive behaviour in the Argentine ant.

All animals interact with conspecifics during their life, and nearly all also display some form of aggression. An enduring challenge, however, is to understand how the experiences of an individual animal influence its later behaviours. Several studies have shown that prior winning experience increases the probability of initiating fights in later encounters. Using behavioural assays in the laboratory, we provide evidence that, in Argentine ants (Linepithema humile), the mere exposure to an opponent, without the encounter escalating to a fight, also increases the probability that it will display aggression in later encounters. Argentine ant workers differ in their propensity to attack non-colonymates, with some ants repeatedly aggressive and others consistently more docile. Although 78 per cent of the workers were consistent in their behaviour from one encounter to the next, workers that did change their behaviour after an encounter with a non-colonymate more often changed from non-aggressive to aggressive, rather than the reverse. Surprisingly, a single encounter with a non-colonymate increased a worker's propensity to fight in encounters up to a week later. An encounter with a non-colonymate also increased the probability that a worker would attack ants from a colony that it had not previously encountered. Thus, these interactions lowered the overall aggression threshold, rather than stimulating a specific aggressive response to a particular foreign colony. Finally, our data suggest that aggression towards non-colonymates increases with age.

When European meets African honeybees (Apis mellifera L.) in the tropics: Morphological changes related to genetics in Mauritius Island (South-West Indian Ocean).

The previous genetic characterization of the honeybee population of Mauritius Island (Indian Ocean) revealed an ongoing process of hybridization between the first established African subspecies Apis mellifera unicolor and recently imported European subspecies (A. m. ligustica, A. m. carnica and A. m. mellifera). This context offers the rare opportunity to explore the influence of hybridization between African and European honeybees on phenotypic traits out of the case largely studied of the Africanized honeybee (hybrid between A. m. scutellata from South Africa and European subspecies). We thus conducted geometric morphometric analyses on forewings of 283 workers genetically characterized at 14 microsatellite loci to evaluate (1) if the morphological variability coincides well with the neutral genetic variability, (2) if hybrids exhibited rather parental, intermediate or transgressive traits, and (3) to test if fluctuating asymmetry (FA) of size and shape, as a measure of developmental stability, was elevated in hybrids (due to genetic stress) and/or European bees (due to unsuitable environment) compared to African bees. A strong concordance was found between morphological variability and neutral genetic variability, especially for wing shape, based on partial least-square analyses (PLS). However, on average, the morphology of hybrids was more similar to the African bees, potentially reflecting the dynamics and direction of introgression. Significant FA for wing size as well as wing shape was detected, suggesting the overall presence of stress during the development of the studied individuals. In contrast, the asymmetry levels do not differ according to the ancestry (African, European or hybrid) of the individuals. Therefore, if ongoing hybridization contributed to increasing the genetic and phenotypic diversity of the populations and influences its adaptive potential, developmental stressors could not be identified and their evolutionary consequences remain uncertain.

Autosomal and Mitochondrial Adaptation Following Admixture: A Case Study on the Honeybees of Reunion Island.

The honeybee population of the tropical Reunion Island is a genetic admixture of the Apis mellifera unicolor subspecies, originally described in Madagascar, and of European subspecies, mainly A. m. carnica and A. m. ligustica, regularly imported to the island since the late 19th century. We took advantage of this population to study genetic admixing of the tropical-adapted indigenous and temperate-adapted European genetic backgrounds. Whole genome sequencing of 30 workers and 6 males from Reunion, compared with samples from Europe, Madagascar, Mauritius, Rodrigues, and the Seychelles, revealed the Reunion honeybee population to be composed on an average of 53.2 ± 5.9% A. m. unicolor nuclear genomic background, the rest being mainly composed of A. m. carnica and to a lesser extent A. m. ligustica. In striking contrast to this, only 1 out of the 36 honeybees from Reunion had a mitochondrial genome of European origin, suggesting selection has favored the A. m. unicolor mitotype, which is possibly better adapted to the island's bioclimate. Local ancestry was determined along the chromosomes for all Reunion samples, and a test for preferential selection for the A. m. unicolor or European background revealed 15 regions significantly associated with the A. m. unicolor lineage and 9 regions with the European lineage. Our results provide insights into the long-term consequences of introducing exotic specimen on the nuclear and mitochondrial genomes of locally adapted populations.

Genetic diversity and differentiation among insular honey bee populations in the southwest Indian Ocean likely reflect old geographical isolation and modern introductions.

With globalization the Western honey bee has become a nearly cosmopolitan species, but it was originally restricted to the Old World. This renowned model of biodiversity has diverged into five evolutionary lineages and several geographic "subspecies." If Apis mellifera unicolor is indubitably an African subspecies endemic to Madagascar, its relationship with honey bees from three archipelagos in the southwest Indian Ocean (SWIO) hotspot of biodiversity is misunderstood. We compared recent mtDNA diversity data to an original characterization of the nuclear diversity from honey bees in the Mascarenes and Comoros archipelagos, using 14 microsatellites, but also additional mtDNA tRNALeu-cox2 analysis. Our sampling offers the most comprehensive dataset for the SWIO populations with a total of 3,270 colonies from 10 islands compared with 855 samples from Madagascar, 113 from Africa, and 138 from Europe. Comprehensive mitochondrial screening confirmed that honey bees from La Réunion, Mauritius, and Comoros archipelagos are mainly of African origin (88.1% out of 2,746 colonies) and that coexistence with European lineages occurs only in the Mascarenes. PCA, Bayesian, and genetic differentiation analysis showed that African colonies are not significantly distinct on each island, but have diversified among islands and archipelagos. FST levels progressively decreased in significance from European and African continental populations, to SWIO insular and continental populations, and finally among islands from the same archipelago. Among African populations, Madagascar shared a nuclear background with and was most closely related to SWIO island populations (except Rodrigues). Only Mauritius Island presented clear cytoplasmic disequilibrium and genetic structure characteristic of an admixed population undergoing hybridization, in this case, between A. m. unicolor and A. m. ligustica, A. m. carnica and A. m. mellifera-like individuals. Finally, global genetic clustering analysis helped to better depict the colonization and introduction pattern of honey bee populations in these archipelagos.

Aggressions and size-related fecundity of queenless workers in the ant Cataglyphis cursor.

In social hymenoptera, the reproductive division of labor is often linked to differences in individual body size with the reproductive caste (the queen) being larger than the workers. Likewise, the reproductive potential may vary with size within the worker caste and could affect the evolution of worker size in social insects. Here, we tested the relationship between worker size and reproductive potential in the facultative parthenogenetic ant Cataglyphis cursor. Colonies are headed by a multiply mated queen, but workers can produce gynes (virgin queens) and workers by thelytokous parthenogenesis after the queen's death. We observed the behaviour of workers (n = 357) until the production of gynes (212 h over 3 months) in an orphaned colony (mated queen not present). The size of workers was measured, and their paternal lineage determined using six microsatellite markers, to control for an effect of patriline. Larger workers were more likely to reproduce and lay more eggs indicating that individual level selection could take place. However, paternal lineage had no effect on the reproductive potential and worker size. From the behavioural and genetic data, we also show for the first time in this species, evidence of aggressive interactions among workers and a potential for nepotism to occur in orphaned colonies, as the five gynes produced belonged to a single paternal lineage.

Habitat-related microgeographic variation of worker size and colony size in the ant Cataglyphis cursor.

In social insects, colony size is a crucial life-history trait thought to have major implications for the evolution of social complexity, especially in relation to worker size polymorphism. Yet, little is known about how ecological factors can affect and constrain colony. Here, we explored the pattern of colony-size and worker-size variation in the Mediterranean ant Cataglyphis cursor, in relation to the type of habitats colonized (seaside vs. vineyard). The high level of the water table in the seaside habitat could constrain the depth of C. cursor underground nests and directly constrain its colony size. If worker size increases with colony size, as observed in other ant species, larger colony size and larger workers should be found in the vineyard populations. By comparing worker size among 16 populations, we verified that workers were significantly larger in the vineyard populations. We further determined that the morphological similarities detected among populations from the same habitat type were not due to geographic or genetic proximity. In two populations from each habitat type, the depth of nests was positively correlated with colony size and colony size with worker size. Using a type II regression approach, we further showed that the difference between the two populations in the depth of nest was sufficient to explain the difference in colony size, and similarly, variation in colony size was sufficient to explain variation in worker size. Our results suggest that a single proximate ecological factor could lead to significant variation in major life-history parameters.

Taste, movement, and death: varying effects of new prospero mutants during Drosophila development.

The PGal4 transposon inserted upstream of the pan-neural gene prospero (pros) causes several neural and behavioral defects in the Voila(1) strain. The precise excision of the transposon simultaneously rescued all these defects whereas its unprecise excision created new pros(V) alleles, including the null allele pros(V17). Here, we describe the relationship between the genetic structure of pros locus, larval locomotion, and larval gustatory response. These two behaviors showed varying degrees of variation depending upon the pros allele. We also found a good relation between behavioral alteration, the level of Pros protein in the embryo, and the degree of disorganization in the larval neuromuscular junction. These data suggest that the complete development of the nervous system requires a full complement of Pros, and that a gradual decrease in the levels of this protein can proportionally alter the development and the function of the nervous system.