Carabid patterns in olive orchards and woody semi-natural habitats: first implications for conservation biological control against <Emphasis Type="Italic">Bactrocera oleae</Emphasis>

The role of carabids (Coleoptera: Carabidae) for pest control service in perennial crop systems has been scarcely investigated. We addressed this knowledge gap exploring activity patterns and traits of adult carabids dwelling olive orchard agroecosystems as potential natural enemies of third instar larvae and pupae of Bactrocera oleae (Diptera: Tephritidae). Olive orchard supported a well-structured carabid assemblage, whose species phenologies revealed a temporal overlapping within the pest cycle. The assemblage of adjacent woody semi-natural habitats is more of conservation interest, but may play a weaker role in B. oleae control provisioning. We suggest the identification of carabids main traits for B. oleae conservation biological control as a cost-effective strategy for addressing future attention and resources only to those predators that satisfy basic requirements. This research may open new scenarios on management interventions to both conserve predators and encourage alternative approaches against olive orchards pest.     

<Emphasis Type="Italic">Bactrocera oleae</Emphasis> pupae predation by <Emphasis Type="Italic">Ocypus olens</Emphasis> detected by molecular gut content analysis

The pupae of Bactrocera oleae (Diptera: Tephritidae) complete their development during autumn and winter in the soil, rather than in the drupe, resulting susceptible to edaphic predators. Environmentally friendly methods to control this olive pest involve the identification of its natural enemies. This study evaluated the role of Ocypus olens (Coleoptera: Staphylinidae) in the predation of B. oleae pupae, by means of molecular gut content analysis. Modified dry pitfall traps were used to collect live specimens from low-input olive orchards in Tuscany (Italy). Sampling was fine-tuned with a degree-day model estimating the presence of pest pupae in the soil. PCR analyses carried out on field-collected specimens demonstrated that O. olens is a predator of B. oleae, at least during autumn. These results are consistent with predictions of the degree-day model. Knowledge on species composition, traits and complementarity of the natural enemies of B. oleae pupae needs further investigation to advance conservation biological control strategies.     

Genetic polymorphism at the cytochrome oxidase I gene in mediterranean populations of Batrocera Oleae (Diptera: Tephritidae)

The tephritid Bactrocera oleae (Gmelin) is a harmful pest of olive crops that cause important agricultural and economic losses in the Mediterranean area where 90% of the world olive trees are cultivated. The knowledge of the genetic diversity in insect pest species populations is critical for decisions concerning appropriate control management strategies. In the present work, the genetic variability within and among 7 populations ‐five from Spain, one from Italy and one from Tunisia‐, was assessed by sequencing 1151 bp of the COI gene. A total of 21 haplotypes were observed. The intraspecific diversity was high, particularly in the Spanish populations (haplotype and nucleotide diversity 0.84 and 0.00137, respectively). However, the genetic differentiation among the populations was low in the case of Spanish ones (Fst between 0 and 0.041), and higher –and statistically significant– when comparing with the Italian and Tunisian samples. The haplotypes distribution and the PCoA analysis show three clear groups of populations: Spanish, Italian and Tunisian. The results might indicate the length of time elapsed since B. oleae became established in the Mediterranean region, the large effective sizes expected of its populations and the high gene flow among Iberian populations. The information could be relevant for integrated control programmes coordination.     

Patterns of inner chorion structure in Anastrepha (Diptera: Tephritidae) eggs

The inner chorion structure of Anastrepha eggs from 16 species of various infrageneric taxonomic groups is described by scanning and transmission electron microscopy. The layers of the chorion, the outer egg membrane, are structurally similar. Furthermore, an additional trabecular layer (ATL) that exists in some species, together with other characteristics, facilitates the recognition of four patterns of chorion structuring: Pattern I, in which the ATL layer is absent, is found in Anastrepha amita, the Anastrepha fraterculus complex, Anastrepha obliqua, Anastrepha sororcula, Anastrepha suspensa and Anastrepha zenildae (fraterculus group), and Anastrepha bistrigata and Anastrepha striata (striata group); Pattern II in Anastrepha serpentina (serpentina group), Anastrepha grandis (grandis group) and Anastrepha pseudoparallela (pseudoparallela group), in which the ATL presents large open spaces with pillars; Pattern III, found in Anastrepha consobrina (pseudoparallela group), in which the ATL is composed of round cavities; and Pattern IV, found in Anastrepha alveata and Anastrepha pickeli (spatulata group), where the large ATL cavities are reticulated. Comparatively, the chorion structure in Anastrepha eggs is more complex than in eggs of other fruit flies, e.g., Bactrocera, Rhagoletis and Ceratitis.     

Cryptic genetic diversity in Rattus of the San Francisco Bay region, California

Invasive species can have complex invasion histories, harbor cryptic levels of diversity, and pose taxonomic problems for pest management authorities. Roof rats, Rattus rattus sensu lato, are common invasive pests of the San Francisco Bay Area in California, USA. They are a significant health risk and pest management efforts impose a large financial investment from public institutions and private individuals. Recent molecular genetic and taxonomic studies of black rats in their native range in Asia have shown that the species is a complex of two karyotypic forms and four mitochondrial genetic lineages that may represent four distinct species. We used mtDNA sequences and nuclear microsatellite variation to identify which mitochondrial lineages of the R. rattus group are present in the San Francisco Bay Area and to test for gene flow among them. We recovered specimens with mtDNA sequences representing two of the major mtDNA lineages of the R. rattus group. Microsatellite variation, however, was not structured in concordance with mtDNA lineages, suggesting a more complex history involving hybridization and introgression between these lineages. Although Aplin et al. (2011) and Lack et al. (2012) reported R. rattus Lineage II in North America, this is the first detailed examination of possible gene flow amongst lineages in this region.     

DNA barcoding of sunn pest adult parasitoids using cytochrome c oxidase subunit I (COI)

In this study, DNA barcoding was used in the identification of potential biological control agents of sunn pest adult parasitoid species, including Eliozeta helluo (F.), Phasia subcoleoptrata (L.), Ectophasia crassipennis (F.) and Elomyia lateralis (Meig). DNA analyses were assessed by sequencing cytochrome c oxidase subunit I (COI) gene. The obtained sequences were analyzed in terms of nucleotide composition, nucleotide pair frequency and haplotype diversity. Genetic divergence among haplotypes was estimated by constructing genetic distance matrix using DNA sequence variations, by Kimura 2-parameter model. Variable sites and average variations of the sequenced 603 base pair long DNA fragment were calculated. All COI barcodes were matched with reference sequences of expected species according to morphological identification. Neighbor-joining tree was drawn based on DNA barcodes and all the specimens clustered in agreement with their taxonomic classification at species level. The evolutionary history inferred using the UPGMA method indicated two distinct mitochondrial haplotype lineages. The genetic variation between sunn pest adult parasitoids will be useful in sunn pest management, regulatory and environmental applications.     

Marked Genetic Differentiation between Western Iberian and Italic Populations of the Olive Fly: Southern France as an Intermediate Area

The olive fly, Bactrocera oleae, is the most important pest affecting the olive industry, to which it is estimated to cause average annual losses in excess of one billion dollars. As with other insects with a wide distribution, it is generally accepted that the understanding of B. oleae population structure and dynamics is fundamental for the design and implementation of effective monitoring and control strategies. However, and despite important advances in the past decade, a clear picture of B. oleae''s population structure is still lacking. In the Mediterranean basin, where more than 95% of olive production is concentrated, evidence from several studies suggests the existence of three distinct sub-populations, but the geographical limits of their distributions, and the level of interpenetration and gene flow among them remain ill-characterized. Here we use mitochondrial haplotype analysis to show that one of the Mediterranean mitochondrial lineages displays geographically correlated substructure and demonstrate that Italic populations, though markedly distinct from their Iberian and Levantine counterparts are more diverse than previously described. Finally, we show that this distinction does not result from extant hypothetical geographic limits imposed by the Alps or the Pyrenees nor, more generally, does it result from any sharp boundary, as intermixing is observed in a broad area, albeit at variable levels. Instead, Bayesian phylogeographic analysis suggests the interplay between isolation-mediated differentiation during glacial periods and bi-directional dispersal and population intermixing in the interglacials has played a major role in shaping current olive fly population structure.     

Prospects for improving biological control of olive fruit fly,Bactrocera oleae (Diptera: Tephritidae), with introduced parasitoids (Hymenoptera)

Olive fruit fly is a key pest of olive and consequently a serious threat to olive fruit and oil production throughout the Mediterranean region. With the establishment of Bactrocera oleae in California a decade ago, interest was renewed in classical (introduction) biological control of the pest. Here we discuss the prospects of identifying natural enemies of B. oleae in Africa and Asia that may help reduce B. oleae populations in California and elsewhere. Based on the current understanding of Bactrocera phylogenetics, early opinions that B. oleae originated in Africa or western Asia rather than the Mediterranean region or the Near East are taxonomically and ecologically supportable. Closely related to cultivated olive, the wild olive Olea europaea cuspidata is widely distributed in southern and eastern Africa, the Arabian Peninsula, and eastwards into Asia as far as southwestern China. Little is known regarding the biology and ecology of B. oleae in Africa and eastern Asia, especially in wild olives. While the diversity of parasitoids of B. oleae in the Mediterranean region is low and unspecialized, a diverse assemblage of parasitoids is known from B. oleae in Africa. Conversely, regions in Asia have remained largely unexplored for B. oleae and its natural enemies.     

Is phylogeography helpful for invasive species risk assessment? The case study of the bark beetle genus Dendroctonus

Despite evidence that conspecific lineages may display different climatic tolerances, most invasion risk assessment tools are calibrated without considering phylogeographic information. This study aims to investigate the existence of intraspecific niche divergence within a group of insect pests and to explore how the inclusion of phylogeographic information into species distribution models may alter the estimation of the potential distribution of a species. We studied North American bark beetles belonging to the genus Dendroctonus, a group of pests of conifers that are listed as quarantine species in numerous countries. Most Dendroctonus species exhibit strong genetic divergence that appears to be geographically structured and shaped by historical events and biotic factors. We modeled all lineage distributions within five species, using MaxEnt and Boosted Regression Trees, and compared the results with the models fitted at the species scale. Multivariate analysis and niche similarity and equivalency tests were additionally performed to investigate the existence and magnitude of climatic niche divergence between conspecific lineages. We also tested the ability of lineage‐based models to predict the region invaded by D. valens in China. Conspecific lineages showed a climatic niche more similar than expected by chance, but displayed different climatic envelopes in their native range and, consequently, different estimates of potential distributions. We also observed that classical models calibrated using the entire range of the species could potentially under‐ or overestimate the potential range of the species when compared to a global prediction built by aggregating lineage‐based projections. This study showed that the invasive phylogeographic lineage of D. valens has invaded regions characterized by climatic conditions highly similar to those encountered in its native range suggesting that preadaptations to environment might have played a role in this invasion. This study highlights how our perception of the invasion risk of pests may be altered when integrating phylogeographic information.     

Cryptic diversity,intraspecific phenetic plasticity and recent geographical translocations in Branchiomma (Sabellidae,Annelida)

The importance of identifying biological diversity accurately and efficiently is becoming more evident. It is therefore critical to determine the species boundaries between closely related taxa and to establish diagnostic characters that allow us to define species. This is not an easy task when species exhibit high intraspecific phenotypic plasticity or when distinct evolutionary lineages with an unusually large amount of genetic distinctiveness show no apparent morphological diversity (cryptic species). These phenomena appear to be common in the genus of fan worms Branchiomma (Sabellidae, Annelida), and consequently, taxonomic errors are widespread in the group. Moreover, some Branchiomma species have been unintentionally translocated outside the area where natural range extension is expected, increasing the taxonomic problems. We have performed a range of analytical methods including genetic distances, Bayesian inference, maximum likelihood, maximum parsimony, statistical parsimony analyses and general mixed Yule coalescent model to clarify the taxonomic status and assess the species boundaries of Branchiomma in Australia. This study shows that the traditional diagnostic morphological features are greatly homoplastic. Results also indicate that the diversity of Branchiomma in Australia is higher than previously reported and evidence some cases of high phenetic plasticity (in features previously considered as stable within species), high intraspecific genetic variability, cryptic species and several unexpected cases of translocations.     

Lineage diversity within a widespread endemic Australian skink to better inform conservation in response to regional‐scale disturbance

Much attention is paid in conservation planning to the concept of a species, to ensure comparability across studies and regions when classifying taxa against criteria of endangerment and setting priorities for action. However, various jurisdictions now allow taxonomic ranks below the level of species and nontaxonomic intraspecific divisions to be factored into conservation planning—subspecies, key populations, evolutionarily significant units, or designatable units. Understanding patterns of genetic diversity and its distribution across the landscape is a key component in the identification of species boundaries and determination of substantial geographic structure within species. A total of 12,532 reliable polymorphic SNP loci were generated from 63 populations (286 individuals) covering the distribution of the Australian eastern three‐lined skink, Bassiana duperreyi, to assess genetic population structure in the form of diagnosable lineages and their distribution across the landscape, with particular reference to the recent catastrophic bushfires of eastern Australia. Five well‐supported diagnosable operational taxonomic units (OTUs) existed within B. duperreyi. Low levels of divergence of B. duperreyi between mainland Australia and Tasmania (no fixed allelic differences) support the notion of episodic exchange of alleles across Bass Strait (ca 60 m, 25 Kya) during periods of low sea level during the Upper Pleistocene rather than the much longer period of isolation (1.7 My) indicated by earlier studies using mitochondrial sequence variation. Our study provides foundational work for the detailed taxonomic re‐evaluation of this species complex and the need for biodiversity assessment to include an examination of cryptic species and/or cryptic diversity below the level of species. Such information on lineage diversity within species and its distribution in the context of disturbance at a regional scale can be factored into conservation planning regardless of whether a decision is made to formally diagnose new species taxonomically and nomenclaturally.     

Species distributions represent intraspecific genetic diversity of freshwater fish in conservation assessments

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Morphogenetic diversity of the ectomycorrhizal genus <Emphasis Type="Italic">Cortinarius</Emphasis> section <Emphasis Type="Italic">Calochroi</Emphasis> in the Iberian Peninsula

The taxonomic and genetic diversity of Cortinarius section Calochroi in one of the most biodiversity-rich regions in Europe, the Iberian Peninsula, was investigated through morphological and phylogenetic methods. This combined methodological approach allowed the identification of 15 known species and one new species, Cortinarius ortegae, which is described here. A dichotomous key is provided for field recognition of Calochroi species inhabiting different Iberian and Balearic Island woodlands. Polymorphism analyses within some studied species showed that the distribution of intraspecific lineages is dependent on geography and, in some cases, tree host. Furthermore, the dating analysis suggested that diversification within Calochroi started in the Pliocene and most of the current genetic diversity originated in the Pleistocene. This temporal scenario supports hypotheses in which climatic oscillations in the Quaternary may have been driving the evolution of this group of ectomycorrhizal fungi.     

DNA Barcode Analysis of Thrips (Thysanoptera) Diversity in Pakistan Reveals Cryptic Species Complexes

Although thrips are globally important crop pests and vectors of viral disease, species identifications are difficult because of their small size and inconspicuous morphological differences. Sequence variation in the mitochondrial COI-5ʹ (DNA barcode) region has proven effective for the identification of species in many groups of insect pests. We analyzed barcode sequence variation among 471 thrips from various plant hosts in north-central Pakistan. The Barcode Index Number (BIN) system assigned these sequences to 55 BINs, while the Automatic Barcode Gap Discovery detected 56 partitions, a count that coincided with the number of monophyletic lineages recognized by Neighbor-Joining analysis and Bayesian inference. Congeneric species showed an average of 19% sequence divergence (range = 5.6% - 27%) at COI, while intraspecific distances averaged 0.6% (range = 0.0% - 7.6%). BIN analysis suggested that all intraspecific divergence >3.0% actually involved a species complex. In fact, sequences for three major pest species (Haplothrips reuteri, Thrips palmi, Thrips tabaci), and one predatory thrips (Aeolothrips intermedius) showed deep intraspecific divergences, providing evidence that each is a cryptic species complex. The study compiles the first barcode reference library for the thrips of Pakistan, and examines global haplotype diversity in four important pest thrips.     

Geographic distribution of phylogenetically-distinct legume pod borer, Maruca vitrata (Lepidoptera: Pyraloidea: Crambidae)

Maruca vitrata Fabricius is a pantropical lepidopteran pest of legumes. Phylogenetic analysis of a mitochondrial cytochrome c oxidase-I gene (cox1) fragment indicates that three Maruca sp. mitochondrial lineages have unique geographic distributions [lineages 1 and 2: Australia, Taiwan, and West Africa (Niger, Nigeria, and Burkina Faso), and lineage 3: Puerto Rico]. The haplotype (T30, T114) is specific to lineages 1&2 and was assayed by NsiI and SacI polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) within population samples; it was not observed in the Puerto Rican samples, but was nearly fixed among samples from West Africa, Australia and Taiwan (85.5?C100%). Re-sequencing and phylogenetic analyses of PCR-RFLP defined cox1 haplotypes indicate that nucleotide diversity is highest among samples from West Africa. Phylogenetic reconstruction based upon ribosomal DNA (rDNA) internal transcribed spacer-2 (ITS-2) sequences provided additional evidence for three Maruca sp. clades. These data suggest that multiple unique Maruca species or subspecies are present worldwide, which has implications for the management of this pest species-complex.     

Potential of pmoA Amplicon Pyrosequencing for Methanotroph Diversity Studies

We analyzed the potential of pmoA amplicon pyrosequencing compared to that of Sanger sequencing with paddy soils as a model environment. We defined operational taxonomic unit (OTU) cutoff values of 7% and 18%, reflecting methanotrophic species and major phylogenetic pmoA lineages, respectively. Major lineages were already well covered by clone libraries; nevertheless, pyrosequencing provided a higher level of diversity at the species level.     

Spatio‐temporal analysis of the relationship between landscape structure and the olive fruit fly Bactrocera oleae (Diptera: Tephritidae)

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