Molecular and morphological diagnoses of five species of Trichogramma: biological control agents of Chrysodeixis chalcites (Lepidoptera: Noctuidae) and Tuta absoluta (Lepidoptera: Gelechiidae) in the Canary Islands

Prospecting for potential natural enemies of the invasive lepidopteran tomato pest Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae) and the banana pest Chrysodeixis chalcites (Esper) (Lepidoptera: Noctuidae) on the Canary Islands archipelago, where no Trichogramma species were previously recorded, has led to the discovery of five distinct species. T. achaeae Nagaraja & Nagarkatti, T. bourarachae Pintureau & Babault, T. euproctidis (Girault) and T. evanescens Westwood are relatively widespread species. The fifth is close to T. brassicae Bezdenko, but differs sufficiently in the sequence of the ITS2 region of ribosomal RNA to warrant further investigation as a species probably new to science. Each species is treated in detail in order to facilitate identification in future using molecular and/or morphological characters, or a combination of both. All species are newly recorded for the Canary Islands, and the distribution of each within the islands and elsewhere is provided. Known host records are given within the Canary Islands and elsewhere. The most common species found, T. achaeae, is already being used in biological control programmes against T. absoluta in mainland Spain and field trials are ongoing to evaluate its effectiveness as a biological control agent of C. chalcites in banana crops.     

Salmon aquaculture and antimicrobial resistance in the marine environment

Antimicrobials used in salmon aquaculture pass into the marine environment. This could have negative impacts on marine environmental biodiversity, and on terrestrial animal and human health as a result of selection for bacteria containing antimicrobial resistance genes. We therefore measured the numbers of culturable bacteria and antimicrobial-resistant bacteria in marine sediments in the Calbuco Archipelago, Chile, over 12-month period at a salmon aquaculture site approximately 20 m from a salmon farm and at a control site 8 km distant without observable aquaculture activities. Three antimicrobials extensively used in Chilean salmon aquaculture (oxytetracycline, oxolinic acid, and florfenicol) were studied. Although none of these antimicrobials was detected in sediments from either site, traces of flumequine, a fluoroquinolone antimicrobial also widely used in Chile, were present in sediments from both sites during this period. There were significant increases in bacterial numbers and antimicrobial-resistant fractions to oxytetracycline, oxolinic acid, and florfenicol in sediments from the aquaculture site compared to those from the control site. Interestingly, there were similar numbers of presumably plasmid-mediated resistance genes for oxytetracycline, oxolinic acid and florfenicol in unselected marine bacteria isolated from both aquaculture and control sites. These preliminary findings in one location may suggest that the current use of large amounts of antimicrobials in Chilean aquaculture has the potential to select for antimicrobial-resistant bacteria in marine sediments.     

Future land use effects on the connectivity of protected area networks in southeastern Spain

Land-use change is a major driver of the global biodiversity crisis, mainly via the fragmentation and loss of natural habitat. Although land-use changes will accelerate to meet humankind's growing demand for agricultural products, conservation planning rarely considers future land uses and how they may affect the connectivity of ecological networks. Here, we integrate land-use models with landscape fragmentation and connectivity analyses, to assess the effects of past and future land-use changes on the connectivity of protected area networks for a highly dynamic region in southeast Spain. Our results show a continued geographical polarisation of land use, with agricultural intensification and urban development in the coastal areas, and the abandonment of traditional land use in the mountains (e.g., 1100 km² of natural vegetation are projected to be lost in coastal areas whereas 32 km² of natural vegetation would recover in interior areas from 1991 to 2015). As a result, coastal protected areas will experience increasing isolation. The connectivity analyses reveal that the two protected area networks in place in the study area, the European “Natura 2000” and the Andalusian “RENPA” networks, include many landscape connectors. However, we identify two areas that currently lack protection but contain several important patches for maintaining the region's habitat connectivity: the northwestern and the southwestern slopes of the Sierra Cabrera and Bédar protected area. Our results highlight the importance of considering future land-use trajectories in conservation planning to maintain connectivity at the regional scale, and to improve the resilience of conservation networks.     

Complementation of a Nonmotile flaB Mutant of Borrelia burgdorferi by Chromosomal Integration of a Plasmid Containing a Wild-Type flaB Allele

With the recent identification of antibiotic resistance phenotypes, the use of reporter genes, the isolation of null mutants by insertional inactivation, and the development of extrachromosomal cloning vectors, genetic analysis of Borrelia burgdorferi is becoming a reality. A previously described nonmotile, rod-shaped, kanamycin-resistant B. burgdorferi flaB::Km null mutant was complemented by electroporation with the erythromycin resistance plasmid pED3 (a pGK12 derivative) containing the wild-type flaB sequence and 366 bp upstream from its initiation codon. The resulting MS17 clone possessed erythromycin and kanamycin resistance, flat-wave morphology, and microscopic and macroscopic motility. Several other electroporations with plasmids containing wild-type flaB and various lengths (198, 366, or 762 bp) of sequence upstream from the flaB gene starting codon did not lead to functional restoration of the nonmotile flaB null mutant. DNA hybridization, PCR analysis, and sequencing indicated that the wild-type flaB gene in nonmotile clones was present in the introduced extrachromosomal plasmids, while the motile MS17 clone was a merodiploid containing single tandem chromosomal copies of mutated flaB::Km and wild-type flaB with a 366-bp sequence upstream from its starting codon. Complementation was thus achieved only when wild-type flaB was inserted into the borrelial chromosome. Several possible mechanisms for the failure of complementation for extrachromosomally located flaB are discussed.     

A new species of Viola L. (Violaceae) from South Anatolia

A new species of Viola L., Viola yildirimlii M. Dinç & Y. Bağcı sp. nov. from South Anatolia is described and illustrated. It is found on the rocky slopes of Aladağ National Park, in the county of Adana, south Turkey, at an elevation of 1800 m. It belongs to Viola , subsect. Viola , and is similar to the Turkish endemics Viola isaurica Contandr. & Quézel and V. kizildaghensis M. Dinç & Ş. Yıldırımlı. Diagnostic morphologic characters for a detailed discrimination from two similar taxa and other Turkish Eflagellatae species are discussed.  © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society , 2003, 141 , 477–482.     

Characterization of Spanish grapevine cultivar diversity using sequence-tagged microsatellite site markers.

A broad germplasm bank collection containing most of the autochthonous Spanish grapevine cultivars was analyzed using six sequence-tagged microsatellite site (STMS) loci: VVS2, VVMD5, VVMD7, ssrVrZAG47, ssrVrZAG62, and ssrVrZAG79. The number of alleles obtained ranged from 9 in ssrVrZAG47 to 13 in VVS2, and the observed genotypes per locus varied between 24 (ssrVrZAG47) and 41 (VVSS2). A total of 57 unique genotypes were obtained considering all 6 loci, and 40 varieties presented at least 1 of these specific genotypes. The genotypic combinations for the 6 loci have generated 163 different profiles in the 176 cultivars. Ten pairs of accessions and one group of four Garnacha's cultivars can not be differentiated. The observed heterozygosity varied between 75.6 (VVMD7) and 90.9% (VVMD5), without significant differences from the expected values for any loci. The VVMD5 locus was the most informative, and also showed the highest discrimination power. The cumulative discrimination power for all six loci was practically 1; however, in fact, these STMS loci have differentiated only about 93% of the accessions, probably owing to high relatedness of the plant material. Usefulness of this STMS set for characterization of a Spanish grapevine collection is emphasized, as well as the elaboration of databases with these molecular markers.     

Diversity in soil fungi from undisturbed and disturbed Celtis tala and Scutia buxifolia forests in the eastern Buenos Aires province (Argentina)

The rhizospheric soil microfungi from a native forest (undisturbed and disturbed) were studied using soil dilution plate and soil washing methods. Fungi were isolated using slightly acid and alkaline culture media. 54 taxa were isolated: 49 from undisturbed forest soil and 37 from disturbed forest soil. Acremonium sp., Aspergillus ustus, Coemansia pectinata, Doratomyces stemonitis, Fusarium solani, F. oxysporum, Gliocladium roseum, Humicola fusco-atra, Mortierella sp., Penicillium lilacinum, Trichoderma harzianum, and T. koningii, showed the highest frequency, in both, undisturbed and disturbed forests. In undisturbed soil forest the biodiversity index was 3.97 whereas in disturbed ones was 3.89.     

Digestive proteases during development of larvae of red palm weevil, Rhynchophorus ferrugineus (Olivier, 1790) (Coleoptera: Curculionidae).

The evolution of digestive proteases during larval development of Rhynchophorus ferrugineus (Olivier, 1790) has been studied. A progressive increase of protease activity has been found. The optimum pH for proteolytic activity against azocasein was determined. Caseinograms revealed an active complex of alkaline proteases from the early stages of the development. From the apparent molecular masses, three groups of proteases have been found - high molecular-mass proteases, medium molecular-mass proteases, and low molecular-mass proteases. Studies using specific protease inhibitors showed the major presence of serine proteases in gut extracts. The results obtained from larvae reared on different substrates have made possible a comparative assessment of the influence of diet on the development of the digestive enzymatic system. Larvae fed on an artificial diet showed a complete pattern of digestive proteases. Data suggest that this diet seems to be suitable for future research with this insect pest.     

Extracellular lipolytic activity in Phoma glomerata

Several properties of the lipolytic activity exhibited by the conidial fungus Phoma glomerata were studied. Lipolytic activity in an aqueous buffer medium was measured on triacylglycerol, phosphoglyceride and cholesterol ester under different experimental conditions. The effect of storage temperature on the stability of the hydrolytic activity, and optimal conditions of temperature and time of maximal activity were determined. The optimal conditions for maximal lipolytic activity were found to be 40–50 °C and 1 h. The activity released to the medium by 1 mg cells for 1 h at 40 °C was stated as the enzyme released unit (ERU). The protein fraction of MW > 50 kDa obtained by ultrafiltration of the medium, was active on the three substrates assayed, and it showed a non-specific hydrolytic activity on both the 1- and 2-acyl esters either in the neutral glyceride or in the phosphoglyceride. A protein of M _r approx. 75 kDa was the only one that showed esterase activity. The crude medium, stored at –15 °C, maintained its initial hydrolytic activity on triacylglycerol for at least 42 days, though when it was kept for 10 days at 4 °C, the activity fell to 50%. Kinetic parameters using substrates such as triolein (TO), dipalmitoyl phosphatidylcholine (DPPC) and cholesteryl oleate (ChoO), were comparatively evaluated. The activity of the enzyme in the hydrolysis of TO showed the highest values, whereas the maximal specific activities were less when the enzyme was assayed against DPPC and ChoO.