Biofilmbildung bei Pflanzen-assoziierten Bakterien der Gattung Methylobacterium und molekulargenetisch-physiologische Charakterisierung eines neuen Marchantia-Isolats (Mtb. sp. JT1)

Bakterien existieren bevorzugt in Biofilmen. Das Zusammenleben in diesen Gemeinschaften bietet den einzelnen Mikroben einen wirksamen Schutz und ermöglicht die Ausbildung langfristiger, synergistischer Wechselwirkungen, die mit multizellulären Systemen verglichen werden können. Biofilme bestehen aus Mikrooganismen-Populationen, die sich an Grenzflächen ansammeln und typischerweise von einer Matrix aus extrazellulären polymeren Substanzen umgeben sind. Auch auf Pflanzen-Oberflächen bilden viele Bakterien Biofilme, um ihre Überlebenswahrscheinlichkeit zu erhöhen. In dieser Arbeit wurde die Biofilmbildung bei Pflanzen-assoziierten Bakterien der Gattung Methylobacterium (Mtb.) untersucht, wobei molekular- und mikrobiologische sowie mikroskopische Techniken eingesetzt wurden. Es zeigte sich, dass alle untersuchten Vertreter der Gattung Methylobacterium in unterschiedlichem Ausmaß Biofilme bilden. Die Ausprägung ist dabei Taxon (bzw. Isolat)-spezifisch und vor allem von der Stickstoff-Verfügbarkeit abhängig. Jedoch spielen auch andere Umweltfaktoren, wie die Versorgung der Zellen mit Phosphat und die Zelldichte, bei der Ausbildung der überzellulären Einheiten eine wichtige Rolle. Die Matrix der Biofilme wird meist durch ein fibrilläres Netzwerk gebildet. Dabei handelt es sich um Heteropolysaccharide, die von den Bakterien synthetisiert und sezerniert werden. Einige Isolate bilden zusätzlich zahlreiche Fimbrien (Auswüchse), durch die sie an andere Zellen oder Oberflächen binden können. Im zweiten Teil dieser Arbeit wurden mehrere neue Methylobacterium-Isolate physiologisch und molekulargenetisch charakterisiert (Nährstoffverwertung, DNA-Sequenzen verschiedener Gene, phylogenetische Analysen usw.). Im Vordergrund stand hierbei der von einer urtümlichen Landpflanze, dem Lebermoos (Marchantia polymorpha), isolierte Stamm Mtb. sp. JT1. Dabei zeigten sich deutliche Unterschiede in der Morphologie und Physiologie des Bakterienstamms JT1 und dem nahe verwandten Stamm 5b.2.20 zu den bereits beschriebenen Taxa der Gattung, so dass eine Spezies-Neubeschreibung erforderlich war. Als Artname wurde aufgrund der außergewöhnlichen Oberflächenstrukturen Mtb. fimbriae sp. nov. eingeführt. Auch andere Methylobakterien (unter anderem Isolat Mtb. sp. F3.2, isoliert vom Laubmoos Funaria hygrometrica) stellen wahrscheinlich Vertreter einer neue Spezies dar (Artname Mtb. funariae sp. nov.). Jedoch zeigen Mtb. fimbriae und Mtb. funariae nur geringe physiologische und morphologische Unterschiede und konnten auf Grundlage umfassender DNA-DNA-Hybridisierungs-Studien nicht eindeutig voneinander abgegrenzt werden.

Scavenging or infection? Possible host choosing by entomopathogenic nematodes

Entomopathogenic nematodes are able to survive by scavenging. We tested Steinernema feltiae, S. affine and Heterorhabditis megidis alone or in different combinations to evaluate the responses of these nematodes when dead or live Galleria mellonella larvae were offered. Steinernema feltiae and S. affine scavenged upon dead G. mellonella larvae and about 30% more dead larvae were penetrated than live ones. By contrast, H. megidis penetrated more live larvae than dead ones. When the nematode species were combined, the results varied among the combinations, but the dead larvae were always used as a host. The behaviour of natural field populations of S. feltiae and S. affine was also compared. Steinernema feltiae showed no difference between scavenging and performing 'normal infections', whereas S. affine scavenged to a reduced amount (around 60% less); this difference could be related to the particular foraging strategy of these nematodes.

Differences in infectivity of Pasteuria penetrans isolates produced on different populations of Meloidogyne

The level of Pasteuria penetrans spore attachment on juveniles of Meloidogyne javanica, M. incognita and M. arenaria was greater when the nematodes were exposed to spores of a population that had been multiplied on a mixture of these Meloidogyne species than where Pasteuria was multiplied on a single nematode population. When tomato plants were inoculated with M. javanica, M. incognita and M. arenaria juveniles encumbered with spores produced on different Meloidogyne species, tile incidence of root galling and productivity of egg-masses were less, and this was also reflected in increased infection of females of M. javanica, M. incognita and M. arenaria compared to the infection by Pasteuria populations produced on single nematode species and therefore assumed to have a narrower genetic base.

In vitro effects of extracts and purified tannins of sainfoin (Onobrychis viciifolia) against two cattle nematodes

Sainfoin (Onobrychis viciifolia) is a condensed tannin (CT)-containing legume and has anthelmintic potential against gastrointestinal nematodes of ruminants. This study investigated in vitro effects of acetone/water extracts and derived CT fractions from different types of sainfoin (i.e. accessions) against larvae of Cooperia oncophora and Ostertagia ostertagi by applying the larval feeding inhibition assay (LFIA). Seven sainfoin accessions were extracted and tested with L1 larvae at 10 and 40 μg extract/ml. In addition, CT in extracts from 4 accessions were fractionated according to polymer size and tested by LFIA at two concentrations (2 and 10 μg CT fraction/ml). All sainfoin extracts caused significant inhibition of L1-feeding of both C. oncophora and O. ostertagi with varying intensity compared to the control (phosphate buffered saline). For both nematode species the in vitro effect was positively correlated with CT content in the extracts, but not with any of the structural CT parameters. In contrast, the 16 CT fractions revealed significant correlations between in vitro effect and CT content, polymer size (mean degree of polymerisation, mDP) and monomeric composition (prodelphinidin percentage, % PD). These differences between crude extracts and purified fractions may stem from the fact that extracts contain complex CT mixtures, which may mask and thus suppress CT structural effects. This study provides the first indication that, apart from CT and % PD content, polymer size also contributes to anthelmintic activity of CTs. The results, therefore, suggest that the inter-accession variability in CT content and composition needs to be taken into account in future plant breeding programmes which seek to enhance the anthelmintic properties of sainfoin

Evidence that nematodes may vector the soft rot-causing enterobacterial phytopathogens

Bacterial soft rot is a globally significant plant disease that causes major losses in the production of many popular crops, such as potato. Little is known about the dispersal and ecology of soft-rot enterobacteria, and few animals have been identified as vectors for these pathogens. This study investigates whether soil-living and bacterial-feeding nematodes could act as vectors for the dispersal of soft-rot enterobacteria to plants. Soft-rot enterobacteria associated with nematodes were quantified and visualized through bacterial enumeration, GFP-tagging, and confocal and electron scanning microscopy. Soft-rot enterobacteria were able to withstand nematode grazing, colonize the gut of Caenorhabditis elegans and subsequently disperse to plant material while remaining virulent. Two nematode species were also isolated from a rotten potato sample obtained from a potato storage facility in Finland. Furthermore, one of these isolates (Pristionchus sp. FIN-1) was shown to be able to disperse soft-rot enterobacteria to plant material. The interaction of nematodes and soft-rot enterobacteria seems to be more mutualistic rather than pathogenic, but more research is needed to explain how soft-rot enterobacteria remain viable inside nematodes.

Organic soils promote the efficacy of entomopathogenic nematodes, with different foraging strategies, in the control of a major forest pest: a meta-analysis of field trial data

The large pine weevil, Hylobius abietis, is a serious pest of reforestation in northern Europe. However, weevils developing in stumps of felled trees can be killed by entomopathogenic nematodes applied to soil around the stumps and this method of control has been used at an operational level in the UK and Ireland. We investigated the factors affecting the efficacy of entomopathogenic nematodes in the control of the large pine weevil spanning 10 years of field experiments, by means of a meta-analysis of published studies and previously unpublished data. We investigated two species with different foraging strategies, the ‘ambusher’ Steinernema carpocapsae, the species most often used at an operational level, and the ‘cruiser’ Heterorhabditis downesi. Efficacy was measured both by percentage reduction in numbers of adults emerging relative to untreated controls and by percentage parasitism of developing weevils in the stump. Both measures were significantly higher with H. downesi compared to S. carpocapsae. General linear models were constructed for each nematode species separately, using substrate type (peat versus mineral soil) and tree species (pine versus spruce) as fixed factors, weevil abundance (from the mean of untreated stumps) as a covariate and percentage reduction or percentage parasitism as the response variable. For both nematode species, the most significant and parsimonious models showed that substrate type was consistently, but not always, the most significant variable, whether replicates were at a site or stump level, and that peaty soils significantly promote the efficacy of both species. Efficacy, in terms of percentage parasitism, was not density dependent.

First record of molluscs naturally infected with Angiostrongylus cantonensis (Chen, 1935) (Nematoda: Metastrongylidae) in Brazil

Seeking the identification of Angiostrongylus cantonensis as a potential etiological agent of three clinical cases of eosinophilic meningitis, mollusc specimens were collected in the state of Espírito Santo, Brazil. The snails were identified as Sarasinula marginata (45 specimens), Subulina octona (157), Achatina fulica (45) and Bradybaena similaris (23). Larvae obtained were submitted to polymerase chain reaction and restriction fragment length polymorphism diagnosis. Their genetic profile were corresponded to A. cantonensis. Rattus norvegicus experimentally infected with third-stage larvae, developed menigoencephalitis, and parasites became sexually mature in the lungs. Additionally, larvae obtained from A. fulica snails, from São Vicente, state of São Paulo, also showed genetic profiles of this nematode. This is the first record of Brazilian molluscs infected with this nematode species.

Evolutionary dynamics of rRNA gene clusters in cichlid fish

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Nematodes of Lizards (Reptilia: Squamata) from Caatinga Biome, Northeastern Brazil

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Helminths of Lizards (Reptilia: Squamata) from Mato Grosso State, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Helminths of toad Rhinella icterica (Bufonidae), from the municipality of Botucatu, São Paulo State, Brazil.

Fifteen specimens of bufonid toad Rhinella icterica (Spix, 1824), 7 males and 8 females, collected in the municipality of Botucatu, São Paulo State, Brazil, from March 2002 to January 2003, were examined for helminth parasites. All toads were infected with at least one parasite species (overall prevalence = 100%). Five helminth species were found, including four nematode species, Rhabdias fuelleborni Travassos, 1926, Oswaldocruzia subauricularis (Rudolphi, 1819), gen. sp. no identified Cosmocercidae Travassos, 1925 and larvae of Physaloptera Rudolphi, 1819, and one acanthocephalan species, Acanthocephalus saopaulensis Smales, 2007. Rhabdias fuelleborni and A. saopaulensis were the most abundant species. Except for Physalopteridae larvae, all helminths occur in high prevalence (> 70%). All helminth species presented a distribution toward to more aggregated pattern in these host samples.

Endoparasites new to the Neotropical freshwater turtle, Mesoclemmys vanderhaegei (Bour 1973) (Testudines, Chelidae), from central Brazil.

Information on endoparasites infecting Neotropical turtles is scarce. The present paper reports the occurrence of endoparasites in three adult individuals of Vanderhaege's Toad-Headed Turtle (Mesoclemmys vanderhaegei). The records include an undescribed nematode species in the genus Spiroxys, not previously reported for the family Chelidae, and the first South American record of a monogenetic trematode of the genus Neopolystoma.

Nematóides associados a plantas daninhas na região de Jaboticabal - SP.: primeiro relato

Realizou-se, na região de Jaboticabal (S.P.), um levantamento dos principais gêneros e espécies de nematóides fitoparasitos encontrados em associação com plantas daninhas de ocorrência genera lizada nas principais culturas locais. Para tal, utilizou -se, como materiais, amostras do solo da rizosfera e raízes de plantas daninhas. Foram identificadas 13 espécies de nematóides de importância agrícola, pertencentes a nove gêneros, em 27 espécies daninhas.

Extensive polymorphism and chromosomal characteristics of ribosomal DNA in the characid fish Triportheus venezuelensis (Characiformes, Characidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Comparative efficacy evaluation of moxidectin gel and ivermectin paste against internal parasites of equines in Brazil

A total of 24 male and female equines of mixed breed, 10-20 months of age and naturally infected with internal parasites was utilized in a controlled test to evaluate the efficacy of a moxidectin 2% gel formulation at the dosage of 0.4 mg moxidectin per kg of live weight and an ivermectin 1.87% commercial paste formulation at the dosage 0.2 mg ivermectin per kg applied orally Animals were allocated into three groups of eight horses each based on pre-treatment eggs per gram (EPG) counts and treatments were randomized among the groups. One group was kept as untreated controls. One animal in the moxidectin-treated group died before the end of the trial from a cause unrelated to treatment leaving a total of seven animals in this group. Fecal egg counts were performed three times post-treatment and the number of parasites remaining in each animal was determined. Statistical analyses using geometric means were performed at the 1% level of significance. Both moxidectin and ivermectin preparations reduced initial EPG from a mean of 1600 to 0 on Days 5, 7 and at the end of the trial on Day 14. Efficacy percentages of moxidectin and ivermectin against immature and adult nematodes were as follows: Trichostrongylus axel, Parascaris equorum, Strongylus edentatus, S. vulgaris, Triodontophorus spp. and Gyalocephalus capitatus, 100% for both products; Habronema muscae 99.5 and 99.6%, respectively, Strongyloides westeri, 100 and 99.2%, respectively; Oxyuris equi, 99.6 and 100%, respectively; small strongyles, 99.7% for both products. of the latter, the most numerous were: Cylicocyclus insigne, Cylicostephanus longibursatus and Cyathostomum catinatum. No Gasterophilus nasalis were found in horses from either treated group, while two of eight control horses had infections with this parasite. Moxidectin showed greater efficacy (84.9%) than ivermectin (67.8%) against Strongylus vulgaris larvae found in the mesenteric artery aneurisms, but the difference was not statistically significant. Total parasite counts for both treated groups were significantly lower (p<0.01) than in the non-treated group. No significant differences were noted between moxidectin and ivermectin. Efficacy against the 30 nematode species found in this study was very evident for both products. As expected, neither moxidectin nor ivermectin was effective in controlling the tapeworm Anoplocephala perfoliata. No adverse reactions were observed during the experimental period. (C) 1998 Elsevier B.V. B.V. All rights reserved.