Sperm ultrastructure of the honey bee (Apis mellifera) (L) (Hymenoptera, Apidae) with emphasis on the nucleus-flagellum transition region

The flagellum of Apis mellifera (Hymenoptera, Apidae) consists of two mitochondrial derivatives, an axoneme and two accessory bodies. The mitochondrial derivatives are of unequal size and lie parallel to the axoneme. In the larger derivative four regions can be distinguished while in the smaller, only three. The region occurring only in the larger derivative consists of paracystalline material. The smaller mitochondrial derivative terminates anterior to the larger one. An extremely long centriolar adjunct is observed between the nucleus and the smaller mitochondrial derivative. This adjunct is compact, very electron dense and gradually tapers from base toward apex, finishing at the anterior extremity of the axonemal microtubules. In this flagellar region, there is only one accessory body present between the larger mitochondrial derivative and the axoneme. Anteriorly, the tips of the axonemal microtubules are inserted in a well developed mass of granular appearance. This material surrounds the nuclear base, separating it from the anterior end of the larger mitochondrial derivative. We believe that the structure identified here as a centriolar adjunct is homologous to that observed in Formicidae, Ichneumonoidea and Symphyta. Therefore, very probably, it is common to most Hymenoptera

Structural and ultrastructural characterization of male reproductive tracts and spermatozoa in fig wasps of the genus Pegoscapus (Hymenoptera, Chalcidoidea)

The three Pegoscapus species present the same internal reproductive tract features comprising testes with a single testicular tubule, seminal vesicles, vasa deferentia, accessory glands and an ejaculatory duct. The seminal vesicle shows two morphologically distinct portions although they do not resemble the separate chambers found in other Chalcidoidea. The anterior portion of the seminal vesicle shows a prominent epithelium and stores the mature spermatozoa, while the posterior region is formed by a thicker muscular sheath that participates on ejaculation. The sexual maturation in Pegoscapus is achieved at emergence, when the testicular degeneration occurs. The spermatozoa of Pegoscapus reveal a basic structure similar to that of other Chalcidoidea. In Pegoscapus sp1. and Pegoscapus sp2. they present the same features, whereas Pegoscapus tonduzi comprises some different characteristics. It measures approximately 160 μm in Pegoscapus sp1. and Pegoscapus sp2., while in P. tonduzi the spermatozoa measure about 360 μm. The extracellular sheath thickness is another difference among the species. While Pegoscapus sp1. and Pegoscapus sp2. show a thick extracellular sheath, in P. tonduzi this sheath is very thin resulting in a large space intervening between the extracellular sheath and the nucleus. Despite these differences, the three species analyzed share some characteristics that allow the establishment of an identity to the spermatozoon of the genus Pegoscapus: the seminal vesicle not divided in chambers; the absence of acrosomal structures in the spermatozoa; the length of the extracellular sheath; the central microtubules being the firsts to terminate in the sequence of microtubular cutoff at the final axonemal portion

Impact of plant genotype and plant habitat in shaping bacterial pathobiome: a comparative study in olive tree

Plant-inhabiting microorganisms interact directly with each other affecting disease progression. However, the role of host plant and plant habitat in shaping pathobiome composition and their implication for host susceptibility/resistance to a particular disease are currently unknown. For the elucidation of these questions, both epiphytic and endophytic bacterial communities, present in asymptomatic and symptomatic twigs from olive cultivars displaying different susceptibilities to olive knot (OK) disease, were investigated using culturing methods. OK disease was the main driver of the bacterial community, causing changes on their diversity, abundance and composition. OK disease effect was most notorious on OK-susceptible cultivar and when considering the endophytic communities. Plant habitat (epiphytes vs. endophytes) also contributed to the bacterial community assembling, in particular on symptomatic twigs (knots) of OK-susceptible cultivar. In contrast, host cultivar had little effect on the bacterial community composition, but OK-symptomatic twigs (knots) revealed to be more affected by this driver. Overall, the pathobiome seems to result from an intricate interaction between the pathogen, the resident bacteria, and the plant host. Specific bacterial genera were associated to the presence or absence of OK disease in each cultivar. Their ability to trigger and/or suppress disease should be studied in the future.The authors are grateful to the FEDER and Foundation for Science and Technology (FCT, Portugal) under Programme PT2020 for financial support to CIMO (UIDB/00690/2020) and BioISI (UID/MULTI/04046/2013), as well as the Horizon 2020, the European Union's Framework Programme for Research and Innovation, for financial support the project PRIMA/0002/2018 INTOMED -Innovative tools to combat crop pests in the Mediterranean. D. Mina thanks FCT, POPH-QREN and FSE for SFRH-BD-105341/2014 grant

Structural Changes in the Male Reproductive Tract of the Stingless Bee Scaptotrigona xanthotricha Moure 1950 (Meliponini, Apidae) During Sexual Maturation

Stingless bees are social insects widely distributed in the Neotropical region but the loss of habitats and the increased use of pesticides in agriculture have threatened the bee populations. Thus, the knowledge of the biology of these insects, including reproductive biology is important for their conservation and management. This study describes the morphological changes in the male reproductive tract during the sexual maturation in the stingless bee Scaptotrigona xanthotricha (Moure 1950). The reproductive tract was investigated under light microscopy from young pupae to 22 days old adults. The male reproductive organs in S. xanthotricha are a pair of testes, each with four follicles, pair of vasa deferentia with enlarged region forming the seminal vesicle that open in an ejaculatory duct. In the male reproductive system of these bees there is no structure that can be recognized as accessory glands as seen in insects in general. From brown-eyed pupae to newly-emerged adults, the epithelia of the seminal vesicles and post-vesicular vasa deferentia have prismatic cells that release secretion to the organs lumen. In 5-days old adults the testes undergo degeneration, the seminal vesicles filled with secretion and spermatozoa, and the epithelium has cubic cells rich in inclusions in the basal region suggesting that this is the age in which males reach the sexual maturation. Structural modifications in the reproductive tract during sexual maturation are discussed concerning the reproductive biology of Meliponini

Biological control of Pseudomonas savastanoi pv. savastanoi by two bacterial isolated from olive tree phyllosphere

Olive knot disease, caused by the bacterium Pseudomonas savastanoi pv. savastanoi (Psv), has been responsible for high economical crop losses in olive orchards, especially in Mediterranean countries. Olive knot cannot be eradicated once it is established in an orchard, and therefore its control is based on preventive measures. Laboratory experiments previously performed by us have showed the capacity of some bacterial species, isolated from olive tree phyllosphere, to inhibited Psv growth. In the present work, the two most promising bacterial isolates (i.e. Frondihabitans sp. and Paenibacillus sp.) were evaluated for the control of Psv in olive plantlets (Olea europaea L.) under greenhouse conditions, to predict their effect in a more natural condition. In pot experiments, two-year-old olive plants cv. Cobrançosa were inoculated with the antagonistic bacteria and Psv individually or in combination. Inoculations were performed in wounds previously made in three different sites of the main stem. Thirty replicate plants were used per strain. The plants were observed for symptom development and the number of bacteria on the inoculation sites was periodically evaluated, for up to 120 days after inoculation. To quantify the reduction of symptom expression, knots were excised from stems and their weights were compared between treatments. Inoculation with Psv resulted in the formation of knots with significantly higher weight values compared to plants inoculated simultaneously with Psv and antagonistic bacteria. Both tested bacterial also showed the capacity to reduce the amount of Psv in the inoculation sites, suggesting their effectiveness in controlling the multiplication of the pathogen. Data presented here demonstrate for the first time the potential of these bacteria for suppression of olive knot, and these two species should be considered in the future as potential biocontrol agents against Psv.info:eu-repo/semantics/publishedVersio

Ability of fungi isolated from olive tree cultivars with different susceptibilities to olive knot to inhibit Pseudomonas savastanoi pv. savastanoi

The Olive Knot is distributed in olive-growing regions worldwide. This disease is caused by the Pseudomonas savastanoi pv. sovastonoí (Psv), a bacterium that produces galls especially on stems and twigs of olive tree. The use of biological control agents (BCAs)to control plant diseases have gained increasing attention during the last decades, due the negative impact of pesticides on environment and human health. This strategy can offer an alternative in order to achieve a sustainable and environ mentally friendly agriculture. The knowledge of fungal communities inhabiting the phyllosphere of olive tree and the type of interaction that they established with the host plant is essential to explore the use of these microorganisms as BCAs against Olive Knot. Previously, we have characterized the fungal communities inhabiting either the surface (epiphytes) or internal tissues of twigs (endophytes)from olive tree cultivars with different susceptibilities to Olive Knot (Cobrançosa moderately tolerant; Verdeal Transmontana susceptible). In this work, 43 endo- and epiphytes of this fungal community were screened for the suppression of Psv, and several mechanisms behind this activity were also studied by evaluating lytic enzymes production. The interspecific interaction was assessed on two different culture media, potato dextrose agar (PDA)and olive leaf twigs extract (OlTE), with agar overlays and by measuring the zone of growth inhibition. Lytic enzymes were evaluated qualitatively. In general, 95% of the fungal species tested inhibited significantly Psv and above an inhibition index (11) threshold of 1.5, being Dothiorella iberica, Rhinocladiella similis, Phoma aloes and Quambalaria cyanescens the species thatexhibited the maximum values (11 ranging from 31 to 19).The inhibition was observed to be affected by growth medium and correlated with the cultivar from which fungi were isolated. In OlTE was observed higher growth inhibition (up to 1.7 fold) and number of species with 11> 5 (up to 1.9 fold), compared to medium without host plant extracts. Similarly, fungal species isolated from the tolerant cultivar Cobrançosa showed higher capacity to inhibit Psv (up to 1.3 fold) when compared to isolates from cv.Verdeal Transmontana. This effect was noticed either for endophytes or epiphytes. The production of lipase, protease, amylase, and cellulase by fungi contributed to the antagonistic activity against Psv, whose production was influenced by olive tree extracts. Overall, results revealed that intimate physical interaction between fungal endophyte/epiphyte with Psvin the presence of olive tree extracts leads to the production of fungal secondary metabolite compounds that inhibit the pathogen growth, which were otherwise not produced. The potential of these fungal species as biological control agents should be considered and further evaluated under "in vivo" conditions.This work is funded by FEDER funds through COMPETE (Programa Operacional Factores de Competitividade) and by national funds by FCT (Fundação para a Ciência e a Tecnologia) in the framework of the project EXCL/AGR-PRO/0591/2012.This work is funded by FEDERfunds through COMPETE(Programa Operacional Factores de Competitividade) and by national funds by FCT(Fundação para a Ciência ea Tecnologia) inthe framework of the project EXCL/AGR-PRO!0591/2012.T. Gomes thanks FCT, POPH-QRENand FSE for PhD SFRH/BD/98127/2013 Grant.info:eu-repo/semantics/publishedVersio

Morphology of the male reproductive system and spermiogenesis in Hypanthidium foveolatum (Alfken, 1930) (Hymenoptera : Apidae : Megachilinae)

The morphological aspects of male reproductive tract, spermiogenesis and spermatozoa are typical for each species and reflect its evolution, establishing a unique source of characters, which has been used to help solve phylogenetic problems. In Hypanthidium foveolatum the reproductive tract is composed of the testes comprising 28 testicular tubules, deferent ducts, seminal vesicles, accessory glands and an ejaculatory duct. The differentiation of spermatids occurs within cysts of up to 128 germ line cells each one. During the early spermatid phase, the nucleus resembles that of somatic cells. There follows a gradual chromatin condensation with an increase in nuclear electron density. In the spermatozoon, the nucleus contains heterogeneous chromatin with a loose appearance. The acrosome, shaped with the active participation of the Golgi complex, shows an electron-dense perforatorium involved by four electron-lucent acrosomal vesicle projections. The sperm tail presents an axoneme with a 9 + 9 + 2 microtubule pattern and two mitochondrial derivatives, which appear with different sizes. A dense crystalloid is formed initially in the mitochondrial matrix of the large derivative. The mitochondrial derivatives’ differentiation occurs concomitantly with an axoneme outgrowth. The centriolar adjunct is observed near the axoneme, anterior to the smaller mithocondrial derivative and exhibits an approximately triangular shape in cross-sections. Microtubules were observed around the head region and flagellar components during spermiogenesis

Potential of the endophyte Penicillium commune in the control of olive anthracnose via induction of antifungal volatiles in host plant

Olive anthracnose, caused by several Colletotrichum species, is the most economically harmful fruit disease of the olive crop. This work aimed to evaluate the ability of the endophyte Penicillium commune CIMO 14FM009 to protect the olive tree against Colletotrichum nymphaeae via induction of plant volatile organic compounds (VOCs). Accordingly, olive tree branches were inoculated with the endophyte and one month later with the pathogen. After 0, 3, and 24 h of pathogen inoculation, the volatile composition of leaves and fruits was analyzed by HSSPME- GC/MS, and compared with controls (branches inoculated with buffer, endophyte, or pathogen). The effect of plant-derived volatiles on C. nymphaeae was also evaluated. Penicillium commune induced the release of VOCs on the olive trees, with the capacity to reduce significantly the growth (up to 1.4-fold) and sporulation (up to 1.2-fold) of C. nymphaeae. This effect was most notorious on olives than on leaves, and occurred 3 h after pathogen-challenge, suggesting the need for a stressful stimulus for the production of antifungal VOCs. The observed inhibition was associated to a specific set of VOCs released from olives (mostly belonging to the alcohols and esters chemical classes) and leaves (mostly belonging to the alkenes). Curiously, a set of VOCs belonging to alkene, alkane and ester classes, were emitted exclusively in olive branches inoculated with C. nymphaeae. These findings provide new possibilities for controlling olive anthracnose using P. commune and/or volatiles, which efficacy should be tested in future works.This work is supported by FEDER funds through the COMPETE (Operational Program for Competitiveness Factors) and by National funds through the FCT (Foundation for Science and Technology) in the scope of the project POCI-01–0145-FEDER-031133 “MicOlives - Exploiting plant induced resistance by beneficial fungi as a new sustainable approach to olive crop protection”, Horizon 2020, the European Union’s Framework Programme for Research and Innovation, in the scope of the project PRIMA/0002/2018 “INTOMED- Innovative tools to combat crop pests in the Mediterranean”, and IFAP via the project “Bio4Med - Implementation of innovative strategies to increase sustainability in perennial Mediterranean crops”, as well as the Mountain Research Center - CIMO (UIDB/00690/2020; UIDP/00690/2020) and SusTEC (LA/P/0007/2020).info:eu-repo/semantics/publishedVersio

Spermatozoa morphology of the giant water bug Belostoma anurum (Herrich-Schäffer, 1848) (Heteroptera: Belostomatidae)

Giant water bugs (Belostomatidae) have important functions in aquatic ecosystems with wide tropical and subtropical distribution. This study describes the spermatozoa morphology of the predator B. anurum. The spermatozoa have not polymorphism and they are scattered without bundles into the seminal vesicle. The mean length of the spermatozoa is 510 μm, including the 35 μm length of the nucleus, which is the longest germ cell, reported for Belostomatidae. The head of the spermatozoa is formed by the acrosome and the nucleus with strongly condensed chromatin. In the nucleus–flagellar transition occurs a C-shaped centriole adjunct, partially surrounding the posterior end of the nucleus and totally surrounding the anterior ends of the two mitochondrial derivatives. In the flagellum, the axoneme has a 9 + 9 + 2 microtubular pattern, and it is linked to the two mitochondrial derivatives by protein bridges. The mitochondrial derivatives are symmetrical surrounding almost entirely the axoneme. The morphology of B. anurum spermatozoa differs from those found in other Pentatomomorpha by the acrosome that does not protrude along with the nucleus and by the single centriole adjunct. © 2020 Elsevier Gmb

Screening of potential biocontrol bacterial against Pseudomonas savastanoi pv. savastanoi and elucidation of their mode of action

Over the last decades, the olive knot disease, caused by the bacterium Pseudomonas savastanoi pv. savastanoi (Psv), has been responsible for irreversible damages on olive orchards. Reduced vigor and stem dryness caused by this phytopathogen lead to a decrease in olive fruit production, conducting to countless losses for farmers. In this work, bacterial endophytes and epiphytes of olive tree phyllosphere were screened for the suppression of Psv, and several mechanisms behind this activity was also studied by evaluating indoleacetic acid (IAA), siderophore and lytic enzymes production. Interspecific interaction was assessed on solid media with agar overlays. IAA was estimated spectrophotometrically, whereas siderophores and lytic enzymes were evaluated qualitatively. Several bacterial species tested showed to reduce Psv growth up to 70%, as well as its viability. The highest inhibition was observed for Frondihabitans sp. and Paenibacillus sp. A reduction on production of both IAA and siderophore, which are associated with knot development, by Psv was noticed in the presence of the most efficient bacterial. Production of lytic enzymes by antagonists such as lipase, chitinase, protease and amylase was also identified. Altogether the results indicate that some of the bacterial tested have great potential as biocontrol agents due to their capacity to produce metabolites/lytic enzymes that can interfere with Psv growth and/or development of knots. These potential biological agents should be further evaluated under natural conditions.This work is funded by FEDER funds through COMPETE (Programa Operacional Factores de Competitividade) and by national funds by FCT (Fundação para a Ciência e a Tecnologia) in the framework of the project EXCL/AGR-PRO/0591/2012. D. MINA thanks the Fundação para a Ciência e Tecnologia (FCT), Portugal for the Ph.D. grant SFRH/BD/105341/2014info:eu-repo/semantics/publishedVersio