Prevendo mudanças climáticas: resposta da meiofauna a mudanças na distribuição espacial de espécies-chave

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Natural recovery of the benthic nematode assemblages associated to Zostera noltii seagrass beds after physical disturbance caused by digging harvest activity

Seagrass beds are productive ecosystems that maintain high levels of biodiversity, making them susceptible to anthropogenic pressures such as bivalve harvesting. Nematodes are considered great ecological indicators as changes in their density, diversity and structure may represent changes in the environment. This experimental fieldwork aimed to assess the impact of the bivalve harvesting on the nematodes assemblage of a seagrass bed in the Mira estuary by simulating the digging activity. Two plots were subjected to the digging (D1 and D19) and two plots were control (C11 and C18). The sampling took place in five occasions: T0 – before digging; T1 – 14 days; T2 – 45 days; T3 – 75 days; and T4 – 165 days after digging. The results showed no significant difference in the nematode assemblages’ density, diversity and trophic composition between treatments and sampling times, evidencing their high tolerance for naturally stressed environments and to the level of digging they were exposed; Recuperação natural das comunidades de nematodes bentónicos associados aos povoamentos de Zostera noltii após atividade de marisqueio Resumo: As pradarias marinhas são ecossistemas produtivos que suportam elevados níveis de biodiversidade, pelo que estão sujeitos a pressões antropogénicas. Os nematodes são bons indicadores ecológicos pois respondem rapidamente a qualquer perturbação por alterações na densidade, diversidade e estrutura. Este trabalho experimental teve como finalidade o estudo da recuperação natural das comunidades de nematodes associados aos povoamentos de Zostera noltii pela simulação da atividade de marisqueio. Dois plots foram sujeitos a revolvimento (D1 e D19) e dois plots serviram como controlo (C11 e C18) e foram efetuadas amostragens em cinco ocasiões: T0 – antes do revolvimento; T1 – 14 dias; T2 – 45 dias; T3 – 75 dias; e T4 – 165 dias após revolvimento. Os resultados obtidos não mostraram diferenças significativas na diversidade, densidade e composição trófica das comunidades de nematodes entre tratamentos e tempos de amostragem, evidenciando a sua elevada tolerância a ambientes naturalmente dinâmicos e ao nível de revolvimento a que foram expostas.

The Invertebrate Assemblage Associated with Macrocystis pyrifera and its Utilization as a Food Resource by Kelp Forest Fishes

The present study was designed to examine the following: (1) the taxonomic. spatial, and temporal patterns of availability of all invertebrate species associated with Macrocystis (excluding protozoans and nematodes); (2) the utilization of this invertebrate assemblage as food by kelp forest fishes within the Macrocystis "foliage- searching" feeding guild, as well as proximal mechanisms leading to observed patterns of resource partitioning; and (3) the dynamic relationship between availability and utilization of this food resource. The approach was largely descriptive. with observations collected during a 19-month period from June 1975 to December 1976. Chapter I is an investigation of the resource utilization patterns of four species of kelp forest fishes with respect to food-related resource dimensions. and tests aspects of current theory involving inter- and intraspecific competition. Chapter II is a detailed examination of the invertebrate assemblage associated with Macrocystis and presents life histories of the fishes examined during this study. (PDFs contains 387 pages, chapter 1 is 203 pages, chapter 2 is 184 pages)

The influence of tidal and rainfall cycles on intertidal nematodes: a case study in a tropical sandy beach

O presente estudo descreve a associação de nematódeos da Baía de Tamandaré (Brasil), praia arenosa tropical típica, durante as marés (baixa, enchente, alta e vazante) de dois ciclos de maré consecutivos, em quatro meses diferentes do ano (Maio, Julho, Setembro e Novembro). A associação de nematódeos foi dominada por Metachromadora e Perepsilonema e variou significativamente entre meses e marés. Densidades foram mais baixas em julho e as mudanças na associação ocorreram durante meses de transição entre chuvoso e seco mostrando a influência do ciclo de chuvas. No ciclo de maré a enchente e vazante pareceram exercer a maior influência, embora os padrões não fossem muito claros. Recomenda-se que os estudos devam ser feitos ao nível de gêneros/espécies para melhor compreensão dos padrões das associações de nematódeos durante ciclos de marés.

Three in one: fixing marine nematodes for ecological, molecular, and morphological studies

Multidisciplinary benthic studies are still hindered by the lack of a unique fixative that satisfactorily preserves morphology and DNA, and that is simultaneously adequate for ecological surveys. The objective of this study is to test the performance of five fixatives: formalin, ethanol, dimethylsulfoxide with EDTA and NaCl salts (DESS), methanol with acetic acid (METHAC), and ethanol with acetic acid (ETHAC), for the preservation of estuarine and exclusively marine nematode assemblages for morphological, molecular, and ecological studies. The presence of the stain rose bengal in each fixative was also evaluated in the yield of PCR reactions. For molecular analyses, one species of each habitat was considered. Results revealed that fixative performance for morphological studies is habitat-and species-dependent. For studies of estuarine sediment nematodes, we recommend the use of pure ethanol, because it caused little morphological distortion (<10% of the assemblage), preserved all the species for ecological studies, and yielded high quality DNA sequences. For studies of exclusively marine environments, METHAC or DESS are the most adequate. The first performed better for morphological and ecological surveys, whereas the second was more appropriate for molecular research. For ecological studies, DESS should be used in comparison with formalin, in order to cross check the results. Finally, staining of samples with rose bengal is not recommended, because it hindered DNA amplification regardless of the fixative used.

Enzymatic activity on sediments and nematode assemblage responses during seagrass beds habitat recovery following the disturbance of the traditional digging activity of bivalves harvesting.

Sediment digging is an anthropogenic activity connected to the exploitation of living resources in estuarine and marine environments. The knowledge on the functional responses of the benthic assemblages to the physical disturbance is an important baseline to understand the ecological processes of the habitat recovery and restoration and to develop tools for the management of the harvesting activities. To investigate the effects of the digging activity of the bivalves on Zostera noltii seagrass beds a manipulative field experiment was conducted that included the enzymatic activity of sediments and the associated nematode assemblages. Four plots (two undisturbed serving as control and two dug to collect bivalves - treatment) with 18 subplots were randomly located at seagrass beds in the Mira estuary at the SW coast of Portugal. Samples were randomly and unrepeatably collected from three subplots of each plot in five different occasions, before sediment digging (T0) up to six months after disturbance (T5). Microbial activity in sediments was assess by determining the extracelular enzymatic activity of six hydrolytic enzymes (sulfatase, phosphatase, b -N-acetilglucosaminidase, b-glucosidase, urease, protease) and two oxidoreductases (phenol oxidase and peroxidase). The microbial community status was also assessed through the measurement of dehydrogenase, which reflects microbial respiration. The nematode assemblages composition, biodiversity and trophic composition at different sampling occasions were also analyzed. The fluorometric and biochemical parameters analysed of the Z. noltii plants during the experimental period showed a recovery of the seagrass beds, and it was detected an increase of the enzymatic activity of the sediments after disturbance. The nematodes assemblages were similar in all sampling occasions. The seagrass beds and the nematodes assemblages associated showed a high resilience to the stress caused by the traditional bivalves digging activity. The obtained results allow the development of a management programme for the commercial fishing activity to maintain the good environmental status and minimized the secondary environmental effects on marine and estuarine habitats through the establishment of a baseline for the regulation of the harvesting frequency.

Benthic nematode assemblage composition and diversity during a natural recovery process of Zostera noltii seagrass beds,

In 2008, the stable seagrass beds of the Mira estuary (SW Portugal) disappeared completely; however, during 2009, they have begun to present early symptoms of natural recovery, characterised by a strongly heterogeneous distribution. This study was designed to investigate the spatial and temporal variability patterns of species composition, densities and trophic composition of the benthic nematode assemblages in this early recovery process, at two sampling sites with three stations each and at five sampling occasions. Because of the erratic and highly patchy seagrass recovery and the high environmental similarity of the two sampling sites, we expected within-site variability in nematode assemblages to exceed between-site variability. However, contrary to that expectation, whilst nematode genus composition was broadly similar between sites, nematode densities differed significantly between sites, and this between-site variability exceeded within-site variability. This may be linked to differences in the Zostera recovery patterns between both sites. In addition, no clear temporal patterns of nematode density, trophic composition and diversity were evident.

Estimating local stream fish assemblage attributes: Sampling effort and efficiency at two spatial scales

As part of a wider study to develop an ecosystem-health monitoring program for wadeable streams of south-eastern Queensland, Australia, comparisons were made regarding the accuracy, precision and relative efficiency of single-pass backpack electrofishing and multiple-pass electrofishing plus supplementary seine netting to quantify fish assemblage attributes at two spatial scales (within discrete mesohabitat units and within stream reaches consisting of multiple mesohabitat units). The results demonstrate that multiple-pass electrofishing plus seine netting provide more accurate and precise estimates of fish species richness, assemblage composition and species relative abundances in comparison to single-pass electrofishing alone, and that intensive sampling of three mesohabitat units (equivalent to a riffle-run-pool sequence) is a more efficient sampling strategy to estimate reach-scale assemblage attributes than less intensive sampling over larger spatial scales. This intensive sampling protocol was sufficiently sensitive that relatively small differences in assemblage attributes (<20%) could be detected with a high statistical power (1-β > 0.95) and that relatively few stream reaches (<4) need be sampled to accurately estimate assemblage attributes close to the true population means. The merits and potential drawbacks of the intensive sampling strategy are discussed, and it is deemed to be suitable for a range of monitoring and bioassessment objectives.

Development and application of a predictive model of freshwater fish assemblage composition to evaluate river health in eastern Australia

Multivariate predictive models are widely used tools for assessment of aquatic ecosystem health and models have been successfully developed for the prediction and assessment of aquatic macroinvertebrates, diatoms, local stream habitat features and fish. We evaluated the ability of a modelling method based on the River InVertebrate Prediction and Classification System (RIVPACS) to accurately predict freshwater fish assemblage composition and assess aquatic ecosystem health in rivers and streams of south-eastern Queensland, Australia. The predictive model was developed, validated and tested in a region of comparatively high environmental variability due to the unpredictable nature of rainfall and river discharge. The model was concluded to provide sufficiently accurate and precise predictions of species composition and was sensitive enough to distinguish test sites impacted by several common types of human disturbance (particularly impacts associated with catchment land use and associated local riparian, in-stream habitat and water quality degradation). The total number of fish species available for prediction was low in comparison to similar applications of multivariate predictive models based on other indicator groups, yet the accuracy and precision of our model was comparable to outcomes from such studies. In addition, our model developed for sites sampled on one occasion and in one season only (winter), was able to accurately predict fish assemblage composition at sites sampled during other seasons and years, provided that they were not subject to unusually extreme environmental conditions (e.g. extended periods of low flow that restricted fish movement or resulted in habitat desiccation and local fish extinctions).

Host-delivered RNAi: An effective strategy to silence genes in plant parasitic nematodes

Root-knot nematodes (Meloidogyne spp.) are obligate, sedentary endoparasites that infect many plant species causing large economic losses worldwide. Available nematicides are being banned due to their toxicity or ozone-depleting properties and alternative control strategies are urgently required. We have produced transgenic tobacco (Nicotiana tabacum) plants expressing different dsRNA hairpin structures targeting a root-knot nematode (Meloidogyne javanica) putative transcription factor, MjTis11. We provide evidence that MjTis11 was consistently silenced in nematodes feeding on the roots of transgenic plants. The observed silencing was specific for MjTis11, with other sequence-unrelated genes being unaffected in the nematodes. Those transgenic plants able to induce silencing of MjTis11, also showed the presence of small interfering RNAs. Even though down-regulation of MjTis11 did not result in a lethal phenotype, this study demonstrates the feasibility of silencing root-knot nematode genes by expressing dsRNA in the host plant. Host-delivered RNA interference-triggered (HD-RNAi) silencing of parasite genes provides a novel disease resistance strategy with wide biotechnological applications. The potential of HD-RNAi is not restricted to parasitic nematodes but could be adapted to control other plant-feeding pests.

Beyond chemical dependency for managing plant-parasitic nematodes: Examples from the banana, pineapple and vegetable industries of tropical and subtropical Australia

Plant-parasitic nematodes are important pests of horticultural crops grown in tropical and subtropical regions of Australia. Burrowing nematode (Radopholus similis) is a major impediment to banana production and root-knot nematodes (predominantly Meloidogyne javanica and M. incognita) cause problems on pineapple and a range of annual vegetables, including tomato, capsicum, zucchini, watermelon, rockmelon, potato and sweet potato. In the early 1990s, nematode control in these industries was largely achieved with chemicals, with methyl bromide widely used on some subtropical vegetable crops, ethylene dibromide applied routinely to pineapples and non-volatile nematicides such as fenamiphos applied up to four times a year in banana plantations. This paper discusses the research and extension work done over the last 15 years to introduce an integrated pest management approach to nematode control in tropical and subtropical horticulture. It then discusses various components of current integrated pest management programs, including crop rotation, nematode monitoring, clean planting material, organic amendments, farming systems to enhance biological suppression of nematodes and judicious use of nematicides. Finally, options for improving current management practices are considered.

Root-lesion nematodes (Pratylenchus thornei and P. neglectus): A review of recent progress in managing a significant pest of grain crops in northern Australia

Two species of root-lesion nematode (predominantly Pratylenchus thornei but also P. neglectus) are widespread pathogens of wheat and other crops in Australia's northern grain belt, a subtropical region with deep, fertile clay soils and a summer-dominant rainfall pattern. Losses in grain yield from P. thornei can be as high as 70% for intolerant wheat cultivars. This review focuses on research which has led to the development of effective integrated management programs for these nematodes. It highlights the importance of correct identification in managing Pratylenchus species, reviews the plant breeding work done in developing tolerant and resistant cultivars, outlines the methods used to screen for tolerance and resistance, and discusses how planned crop sequencing with tolerant and partially resistant wheat cultivars, together with crops such as sorghum, sunflower, millets and canaryseed, can be used to reduce nematode populations and limit crop damage. The declining levels of soil organic matter in cropped soils are also discussed with reference to their effect on soil health and biological suppression of root-lesion nematodes.

Resistance to root-lesion nematodes (Pratylenchus thornei and P-neglectus) in synthetic hexaploid wheats and their durum and Aegilops tauschii parents.

Root-lesion nematodes (Pratylenchus thornei Sher and Allen and P. neglectus (Rensch) Filipijev and Schuurmans Stekhoven) cause substantial yield loss to wheat crops in the northern grain region of Australia. Resistance to P. thornei for use in wheat breeding programs was sought among synthetic hexaploid wheats (2n= 6x = 42, AABBDD) produced through hybridisations of Triticum turgidum L. subsp. durum (Desf.) Husn (2n= 4x = 28, AABB) with Aegilops tauschii Coss. (2n= 2x = 14, DD). Resistance was determined for the synthetic hexaploid wheats and their durum and Ae. tauschii parents from the numbers of nematodes in the roots of plants grown for 16 weeks in pots of pasteurised soil inoculated with P. thornei. Fifty-nine (32%) of 186 accessions of synthetic hexaploid wheats had lower numbers of nematodes than Gatcher Selection 50a (GS50a), a partially resistant bread wheat. Greater frequencies of partial resistance were present in the durum parents (72% of 39 lines having lower nematode numbers than GS50a) and in the Ae. tauschii parents (55% of 53 lines). The 59 synthetic hexaploids were re-tested in a second experiment along with their parents. In a third experiment, 11 resistant synthetic hexaploid wheats and their F-1 hybrids with Janz, a susceptible bread wheat, were tested and the F(1)s were found to give nematode counts intermediate between the respective two parents. Synthetic hexaploid wheats with higher levels of resistance resulted from hybridisations where both the durum and Ae. tauschii parents were partially resistant, rather than where only one parent was partially resistant. These results suggest that resistance to P. thornei in synthetic hexaploid wheats is polygenic, with resistances located both in the D genome from Ae. tauschii and in the A and/or B genomes from durum. Five synthetic hexaploid wheats were selected for further study on the basis of (1) a high level of resistance to P. thornei of the synthetic hexaploid wheats and of both their durum and Ae. tauschii parents, (2) being representative of both Australian and CIMMYT (International Maize and Wheat Improvement Centre) durums, and (3) being representative of the morphological subspecies and varieties of Ae. tauschii. These 5 synthetic hexaploid wheats were also shown to be resistant to P. neglectus, whereas GS50a and 2 P. thornei-resistant derivatives were quite susceptible. Results of P. thornei resistance of F(1)s and F(2)s from a half diallel of these 5 synthetic hexaploid wheats, GS50a, and Janz from another study indicate polygenic additive resistance and better general combining ability for the synthetic hexaploid wheats than for GS50a. Published molecular marker studies on a doubled haploid population between the synthetic hexaploid wheat with best general combining ability (CPI133872) and Janz have shown quantitative trait loci for resistance located in all 3 genomes. Synthetic hexaploid wheats offer a convenient way of introgressing new resistances to P. thornei and P. neglectus from both durum and Ae. tauschii into commercial bread wheats.

In vitro infection of sheep lice (Bovicola ovis Schrank) by Steinernematid and Heterorhabditid nematodes.

Control of sheep lice with conventional pesticides can be compromised by difficulty in contacting lice in the dense water repellent fleeces of sheep, particularly when sheep have not been recently shorn. Entomopathogenic nematodes (ENs) are motile and are able to actively seek out insect hosts. They have particular advantages for the control of pests in cryptic habitats, such as the fleeces of sheep and avoid many of the problems frequently associated with chemical controls. This study investigated whether ENs were able infect and kill Bovicola ovis and compared the effectiveness of different species at different temperatures and when applied to wool. Four species of nematodes, Steinernema carpocapsae, Steinernema riobrave, Steinernema feltiae and Heterorhabditis bacteriophora were tested. All were shown to infect and kill lice in Petri dish assays at 30C. At 35C, the percent infection for S. carpocapsae and S. riobrave was significantly higher than for the other two species and percent infection by S. feltiae was significantly greater than for H. bacteriophora (P<0.05). At 37C the percent mortality induced by S. riobrave was significantly greater than for S. carpocapsae (P<0.05). All species were able to locate and infect lice in wool when formulated in water with 8% Tween 80. In wool assays the percent lice infected with nematodes was significantly greater for S. riobrave than H.bacteriophora at 25C, but there were no other differences between species (P=0.05). S. carpocapsae, S. riobrave and S. feltiae caused significantly higher lice mortality than H. bacteriophora at both 25 and 35C in wool assays, but mortality induced by the three steinernematid species did not differ significantly (P>0.05). It is concluded that of the ENs studied S. riobrave is likely to be most effective against B. ovis when applied to live sheep because of its greater tolerance to high temperatures and 'cruiser' foraging strategy .