Peter John Mazzoglio

Feminizing Wolbachia in Zyginidia pullula (Insecta, Hemiptera), a leafhopper with an XX/X0 sex-determination system

Zyginidia pullula is a grass-dwelling leafhopper characterized by a bisexual reproduction mode. In this species, some females collected in Northern Italy, when mated with males, gave origin to an exclusively female brood. Here, we demonstrated that in these lineages an infection by a new strain of Wolbachia pipientis—designated as wZygpul—was detected by amplifying and sequencing the wsp and 16S rRNA genes. About half of the female progeny were characterized by intersexual phenotypes, i.e. showing upper pygofer appendages, a typical male secondary sexual feature. The karyological analysis proved that while phenotypically normal females had a female genotype, those with upper pygofer appendages had a male genotype and were thus feminized males. The complete removal of W. pipientis after tetracycline treatment of morphologically normal females, and the consequent re-appearance of males in the brood, permitted us to connect the feminizing effect with the presence of the bacterium. This is the first case of feminization by W. pipientis in an XX/X0 sex-determination system, and is the second case reported in insects.

Unravelling the Wolbachia evolutionary role: the reprogramming of the host genomic imprinting.

Environmental factors can induce significant epigenetic changes that may also be inherited by future generations. The maternally inherited symbiont of arthropods Wolbachia pipientis is an excellent candidate as an ‘environmental’ factor promoting trans-generational epigenetic changes: by establishing intimate relationships with germ-line cells, epigenetic effects of Wolbachia symbiosis would be manifested as a ‘maternal effect’, in which infection of the mother modulates the offspring phenotype. In the leafhopper Zyginidia pullula, Wolbachia feminizes genetic males, leaving them as intersexes. With the exception of male chitinous structures that are present in the last abdominal segment, feminized males display phenotypic features that are typical of females. These include ovaries that range from a typical histological architecture to an altered structure. Methylation-sensitive random amplification of polymorphic DNA profiles show that they possess a female genomic imprint. On the other hand, some rare feminized males bear testes instead of ovaries. These specimens possess a Wolbachia density approximately four orders of magnitude lower than feminized males with ovaries and maintain a male genome—methylation pattern. Our results indicate that Wolbachia infection disrupts male imprinting, which dramatically influences the expression of genes involved in sex differentiation and development, and the alteration occurs only if Wolbachia exceeds a density threshold. Thus, a new Wolbachias role as an environmental evolutionary force, inducing epigenetic trans-generational changes, should now be considered.

Grey squirrel Sciurus carolinensis management in Italy - squirrel distribution in a highly fragmented landscape

American grey squirrels Sciurus carolinensis introduced to northern Italy in 1948 have caused damage to commercial poplar plantations and have replaced the native red squirrel Sciurus vulgaris from most of the 350 km2 of the Piedmont Po-plain they currently occupy. In order to plan a control programme aiming to stop grey squirrels from further spreading and to decrease their numbers, the current distribution and population size in the highly fragmented landscape of the Po-plain were studied. The probability of finding grey squirrels in woodland fragments increased with habitat quality (diversity of trees producing large, consumable seeds), woodlot size and the proportion of poplar. Adding isolation variables did not improve the fit of the logistic regression model that predicted squirrel presence. The density of squirrel dreys, an index of population density, in the large Stupinigi forest also increased with tree species diversity. An estimate of the minimum population size for all woodlots assessed for squirrel presence was 1,260 animals in the summer of 1996. This extrapolates to a total of ca 2,500 grey squirrels in Piedmont. Grey squirrels continue to increase their range and are getting close to the continuous mixed forests of the pre-Alps and to large hazel plantations. Control measures to stop the spread of grey squirrels, and eventually to eradicate them, should be implemented immediately.

ACTIVITY BUDGET AND FORAGING BEHAVIOR OF INTRODUCED CALLOSCIURUS FINLAYSONII (RODENTIA, SCIURIDAE) IN ITALY

Abstract We studied seasonal activity patterns and feeding behavior of a population of Finlaysons squirrel (Callosciurus finlaysonii) native to Indochina and introduced into Italy. Squirrels spent the majority of active time foraging, switching between resources according to availability. In winter, they fed mainly on buds or stripped bark, shifting to flowers when plants bloomed in spring, and to mature seeds and fruits from June to October. By November–December, bark-stripping became progressively more important. This activity accounted for 36.5% of the yearly feeding time. The flexibility to exploit a wide range of foods, the ability to change food habits throughout the year, and hoarding behavior are factors that could indicate a proclivity to use different habitats successfully.

Wolbachia is not all about sex: male-feminizing Wolbachia alters the leafhopper Zyginidia pullula transcriptome in a mainly sex-independent manner

Wolbachia causes the feminization of chromosomally male embryos in several species of crustaceans and insects, including the leafhopper Zyginidia pullula. In contrast to the relatively well-established ecological aspects of male feminization (e.g., sex ratio distortion and its consequences), the underlying molecular mechanisms remain understudied and unclear. We embarked on an exploratory study to investigate the extent and nature of Wolbachias effect on gene expression pattern in Z. pullula. We sequenced whole transcriptomes from Wolbachia-infected and uninfected adults. 18147 loci were assembled de novo, including homologs of several Drosophila sex determination genes. A number of transcripts were flagged as candidate Wolbachia sequences. Despite the resemblance of Wolbachia-infected chromosomal males to uninfected and infected chromosomal females in terms of sexual morphology and behavior, principal component analysis revealed that gene expression patterns did not follow these sexual phenotype categories. The principal components generated by differentially expressed genes specified a strong sex-independent Wolbachia effect, followed by a weaker Wolbachia-sexual karyotype interaction effect. Approaches to further examine the molecular mechanism of Wolbachia-host interactions have been suggested based on the presented findings.

Bark-stripping damage by Callosciurus finlaysonii introduced into Italy

Abstract The Finlayson’s squirrel Callosciurus finlaysonii was introduced into Italy during the 1980s and has established two viable populations. The diet of this species includes a high proportion of tree barks, suggesting an intensive debarking behavior. We reported a severe bark-stripping impact in both colonized areas, and we tested whether a preference for some tree species existed. Results of this work showed the presence of a wide spectrum of damaged species, without any strong preference, mainly with large wounds. Old deciduous plants and conifers, which presented a hard bark, were usually avoided.

Male or female? The epigenetic conflict between a feminizing bacterium and its insect host

In the leafhopper Zyginidia pullula, the maternally-inherited symbiont Wolbachia pipientis feminizes genetic males, leaving them as intersex females. The Wolbachia density is correlated with the feminization degree of males which either bear ovaries or testes. Methylation-sensitive RAPD profiles showed that while feminized males with ovaries possess a female imprinting pattern, those with testes maintain the same methylation pattern of males, indicating that the Wolbachia infection is able to modulate host genomic imprinting, but this occurs only if the bacterium exceeds a density threshold. Here we report methylation-sensitive RAPD analyses on gonads (testes and ovaries) from leafhopper uninfected male and female individuals, and males feminized by Wolbachia, discussing these additional insights and speculating on possible bacterium/host interactions.