Applicability of ecological evaluation tools in estuarine ecosystems: the case of the lower Mondego estuary (Portugal)

In accordance with the Water Framework Directive guidelines (WFD, 2000, European Communities Official Journal L327 2000/60/EC), classification schemes and ecological evaluation tools (based on benthic invertebrate fauna data sets from 1990 to 2002) were applied in the lower Mondego estuary. Two distinct scenarios could be tested due to the implementation of mitigation practices in 1999, following a long eutrophication process, which started by the early 1980s. Some discrepancies in the results were found by the application of the different indices. The AMBI index (accounting for taxonomic composition) and the ABC method (accounting for abundance and biomass k-dominance patterns) classifications often disagreed with those based on species diversity (Margalef and Shannon-Wiener). The ambiguous results made the classification a complex task to achieve, contrary to the Directive’s objective of maintaining it simple and clear. Our results suggest the necessity of adjusting some of the indices and their ranges to estuarine characteristics, namely to account the typical dominance and abundance of some particular species. These aspects are not taken into consideration by some of the indices proposed, which are more adapted to typical marine conditions. Based on our results, these widely applied indices might still improve their efficiency in estuarine systems allowing their use in the resembling types already established within the new Directive agenda.     

Validation of the Macrophyte Quality Index (MaQI) set up to assess the ecological status of Italian marine transitional environments

The paper couples the results obtained by applying the expert and the rapid Macrophyte Quality Indices set up to assess the ecological status of the Italian transitional environments according to the requirements by the Water Framework Directive (2000/60/CE). The indices were validated by comparing the composition of the macrophyte assemblages and the values of some bio-physico-chemical parameters of the water column of 20 stations of the Venice lagoon sampled monthly for one year between 2003 and 2005. In 5 stations out of the 20, the ones which fall within the 5 classes of ecological status suggested by the Water Framework Directive, sedimentation rates, sediment grain-size, and nutrient and pollutant (metals, Polychloro-Dibenzo-Dioxins/Furans, Polycyclic Aromatic Hydrocarbons, Pesticides and Polychlorinated Biphenils) concentrations in surface sediments were also determined. Results showed strong relationships between the trends of these environmental parameters and the composition and structure of macrophyte associations, as well as with the Macrophyte Quality Index assessment. Chlorophyceae showed a trend opposite to Rhodophyceae whose presence was concentrated in oxygenated and transparent environments. Chlorophyceae and the species characterised by low scores prevailed in turbid areas where nutrient and pollutant concentrations were high. Results allowed the identification of the conditions of the “reference sites” (confinement areas and sites with high water renewal) and the integration of the dichotomic key used for the application of the R-MaQI. Handling editor: S. M. Thomaz     

Iles Eparses (SW Indian Ocean) as reference ecosystems for environmental research

TAAF ensures since 2007 the management of 5 small tropical islands lying in the southwestern Indian Ocean: the Iles Eparses. These islands share an exceptional natural heritage including many marine and terrestrial endemic species. At a regional scale the Iles Eparses are some of the most pristine ecosystems, largely preserved from anthropogenic impacts due to their geographical isolation and a historically very limited human occupation. In this context, TAAF wished that Iles Eparses become unique natural laboratories for earth scientists and environmental process observation – like climate change impacts - but also sustainable biodiversity sanctuaries for which the scientific community should provide baseline ecological data to inform on appropriate conservation tools. An inter-agency research consortium emerged in 2009 to meet this commitment for the Iles Eparses. This program was intended to set a science framework in accordance with France' objectives for Research and Conservation. It enabled between 2009 and 2014 the implementation of 18 cross-disciplinary research projects ranging from geology to ecology and represented by the variety of the proposed articles in this special issue. Altogether research projects have dramatically increased knowledge on the Iles Eparses' ecosystems and have provided the first overview of their diversity, their functions and their dynamics and its determinants. In particular applied research efforts have supplied a significant amount of ecological evidence that is now available to develop optimal conservation strategy to ensure the Iles Eparses' long-term biodiversity value. These findings point out that the continuation of research activity in the Iles Eparses should be considered a priority.     

Towards a multimetric index for ecological assessment of Mediterranean flatland ponds: the use of macroinvertebrates as bioindicators

Fifty-five macroinvertebrate metrics were tested for their response to pond condition in 41 ponds of northwest Spain to develop a preliminary multimetric index for ecological assessment of Mediterranean flatland ponds. Stressor specific response of individual attributes to eutrophication and habitat alteration was also investigated to identify differences in the responses of metrics to single stressors and elucidate how this might affect the performance of the final index. Several combinations were tested using discrimination efficiency (25th percentile of slightly impaired sites for metrics decreasing with perturbation and 75th percentile of slightly impaired sites for metrics increasing with perturbation) and Mann–Whitney U-test with Bonferroni adjustment (P < 0.001). The final index comprised five measures (generic richness of Chironominae, generic richness of Dytiscidae + Odonata + Tanypodinae, relative richness of Chironomidae, % Macropelopini and Shannon index) and discriminated between acceptable (good) and unacceptable (moderate) conditions with more than 86% efficiency. Moreover, all the five measures included in the final index showed unidirectional responses to eutrophication, decreasing as eutrophication increased. In contrast, the effect of habitat alteration was less clear, especially in ponds in best available conditions where a vegetation belt of shrubs and trees prevented growing of macrophitic vegetation on shores and consequently associated fauna. Interestingly, none of the functional groups (e.g. % predators and % collector–gatherers) were sensitive to degradation. Handling editor: K. Martens     

A multimetric index based on benthic macroinvertebrates for evaluation of Atlantic Forest streams at Rio de Janeiro State, Brazil

This study describes the application of a protocol for biological assessment of water quality at first to third order streams at Serra dos órg?os, an area covered by Atlantic Forest in Rio de Janeiro State, Brazil. Major impacts in the region are domestic effluents and deforestation. Our main objective is to establish biocriteria for the establishment of the Serra dos órg?os Multimetric Index (SOMI) based on benthic macroinvertebrates. We used data from previous studies, sampled by experienced biologists, from 1999 through 2002. The benthic macroinvertebrate community was sampled in 12 reference sites and seven impaired sites in three river basins: Guapimirim, Macaé and Grande, all from the same bioregion. From the 22 tested metrics, 6 were included in the SOMI (% Diptera, % Coleoptera, Family Taxa, EPT Taxa, BMWP-CETEC and % Shredders). Scores (5, 3 or 1) were developed for these metrics to allow for aggregation into the index. Seven intermediately impaired sites were used for evaluating the applicability of the multimetric index. We concluded that the SOMI is a robust easy-to-apply tool for biomonitoring programs in the Serra dos órg?os region, south-east Brazil. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorised users. Handling editor: D. Dudgeon     

Validation of a fish-based index of biotic integrity for streams and rivers of central Mexico

An existing version of a fish assemblage-basedindex of biotic integrity (IBI) for the streamsand rivers of west central Mexico was testedwith independent data to validate itsusefulness as a measure of ecosystem qualityand to determine the geographic area where itis effective. Fish assemblages from 63 uplandsites in 10 basins in central Mexico(Armería, Ameca, Coahuayana, Marabasco,Purificación, Grande de Morelia, Grande deSantiago, Lerma, Balsas and Pánuco) wereassessed using the metrics and scoring criteriafrom the existing IBI and then compared withindependent evaluations of habitat and waterquality. IBI scores were congruent withhabitat and water quality values in theArmería, Purificación and Marabascobasins, where the IBI was first developed, aswell as in the adjacent Ameca and Coahuayanabasins. We conclude that the IBI can be usedwithout modification to assess environmentalquality in non-coastal streams and riverswithin these five basins. Further data areneeded from the Grande de Morelia, Grande deSantiago and middle Lerma basins, but ourresults suggest that the existing IBI may alsobe effective here. However, the existing IBIdoes not consistently reflect habitat and waterquality conditions in the Balsas and Pánucobasins and must be modified before it can beapplied there. Necessary modifications in theBalsas basin appear to be small and relatedprimarily to changes in the scoring criteriafor metrics. However, in the Pánuco basinmore substantive changes in the nature of themetrics are required. Changes in the IBI forthese basins are proposed. The IBI is nowvalidated for use in river monitoring,conservation and restoration efforts in 5basins in west central Mexico and suggestionsfor its application in other basins areavailable here.     

Australian water quality guidelines: a new approach for protecting ecosystem health

The Australian and New Zealand Environment and Conservation Council (ANZECC) and the Australian Water Resources Council (AWRC) have developed a National Water Quality Management Strategy which seeks to ensure that the nation's water resources are managed on a sustainable basis. An important element of this strategy are the Australian Water Quality Guidelines which focus on the protection of Australian freshwater and marine ecosystems. Here the aim is to protect biodiversity and maintain the ecological integrity of each marine and freshwater resource. Specific guidelines have been formulated in terms of key indicators of quality, with a single reference value or ranges of reference values provided for guidance. For those indicators where ranges are provided, it is the expectation that State environmental and resource management agencies will undertake local, site-specific investigations of their own systems to define the specific levels to be adopted. For the first time, specific and quantitative biological indicators have been introduced; these are species richness, species composition, primary production, and ecosystem function.As Australia progresses towards broader, more holistic, ecologically-based management of the nation's water resources, the present water quality guidelines must be extended to ecosystem or environmental guidelines, where the maintenance of adequate water quality is seen as only one (albeit important) component. Other considerations must include habitat protection, sediment quality, and stream flow maintenance. This increased emphasis on more ecologically-focused management of Australia's inland and coastal waters will present a number of challenges for the three major groups involved: the community, the managers, and the researchers. These challenges are discussed.Based on a paper presented at a Workshop on Biological Assessment of Aquatic Ecosystem Health, Sydney, 1–2 October 1992.     

Detecting the impact of oceano-climatic changes on marine ecosystems using a multivariate index: The case of the Bay of Biscay (North Atlantic-European Ocean)

Large‐scale univariate climate indices (such as NAO) are thought to outperform local weather variables in the explanation of trends in animal numbers but are not always suitable to describe regional scale patterns. We advocate the use of a Multivariate Oceanic and Climatic index (MOCI), derived from ‘synthetic’ and independent variables from a linear combination of the total initial variables objectively obtained from Principal Component Analysis. We test the efficacy of the index using long‐term data from marine animal populations. The study area is the southern half of the Bay of Biscay (43°–47°N; western Europe). Between 1974 and 2000 we monitored cetaceans and seabirds along 131000 standardized line transects from ships. Fish abundance was derived from commercial fishery landings. We used 44 initial variables describing the oceanic and atmospheric conditions and characterizing the four annual seasons in the Bay of Biscay. The first principal component of our MOCI is called the South Biscay Climate (SBC) index. The winter NAO index was correlated to this SBC index. Inter‐annual fluctuations for most seabird, cetacean and fish populations were significant. Boreal species (e.g. gadiformes fish species, European storm petrel and Razorbill …) with affinities to cold temperate waters declined significantly over time while two (Puffin and Killer Whale) totally disappeared from the area during the study period. Meridional species with affinities to hotter waters increased in population size. Those medium‐term demographic trends may reveal a regime shift for this part of the Atlantic Ocean. Most of the specific observed trends were highly correlated to the SBC index and not to the NAO. Between 40% and 60% of temporal variations in species abundance were explained by the multivariate SBC index suggesting that the whole marine ecosystem is strongly affected by a limited number of physical parameters revealed by the multivariate SBC index. Aside the statistical error of the field measurements, the remaining variation unexplained by the physical characteristics of the environment correspond to the impact of anthropogenic activities such overfishing and oil‐spills.     

Ultrastructural analysis of transitional endoplasmic reticulum and pre-Golgi intermediates: a highway for cars and trucks

Cargo selection and export from the endoplasmic reticulum occurs at specialized sites in cells. Export complexes consist of transitional elements of the endoplasmic reticulum and pre-Golgi intermediates. It is generally assumed that 60 to 80 nm initially COPII-coated vesicles derived from the transitional endoplasmic reticulum are the main carriers for transport of cargo to the Golgi apparatus. We have analyzed on serial ultrathin sections the transitional endoplasmic reticulum and pre-Golgi intermediates of beta cells of islets of Langerhans in mouse pancreas. In addition to Golgi-associated complexes, others were observed in the periphery of the cells or close to the nuclear envelope. Upon three-dimensional reconstruction, non-coated ribosome-free tubules with an average diameter of 115 nm (range 60–195 nm) and a length of up to 500 nm were detected in the pre-Golgi intermediates in addition to small vesiculo-tubular elements. Furthermore, evidence was found that the large tubular elements may directly arise from transitional elements of the endoplasmic reticulum. In a given cell, pre-Golgi intermediates were found to be composed solely of small vesiculo-tubular elements or additionally of tubules or solely of tubules. Immunogold labeling for proinsulin indicated that the large tubular elements contained cargo and thus appear to take part in ER-to-Golgi transport.Presented at the 43rd Symposium of the Society for Histochemistry, Vienna, Austria, 27–29 September 2001     

Applications of biotope mapping for spatial environmental planning and policy: case studies in urban ecosystems in Korea

The purpose of this paper is to investigate the current status and method of biotope mapping for practical use for landscape planning and environmental policy in urban ecosystem in Korea. We examine current ecological mapping of Seoul, Seongnam, Daegu, and Yongin. Ecological mapping is examined closely in terms of investigation methodology, investigation subject, classification of urban landscape, and the present condition of application. Biotope mapping in Seoul and Seongnam were carried out by the city governments concerned with the pre-set budgets earmarked for mapping. In order to promote the utilization of biotope maps for city planning in Korea, the following actions should be considered. First, the survey method should be standardized by introducing a uniform standard with respect to the scope of survey, the quality of primary data used, the survey method, and the level of the survey. Second, it is necessary to identify a basic category of biotope for each area by consolidating the outcome of the previous surveys. Third, it is highly desirable to minimize the differences between the evaluation criteria and the assessment factors. Fourth, it is ideal to apply the results of the biotope evaluation to city planning in an indirect manner through reflecting the results first in the landscape plans. In order to facilitate this alternative utilization, it is necessary to strengthen the control provisions contained in the ordinances of the city concerned or to enact a set of new provisions in the ordinances so that biotope mapping could be used more widely as a criterion for the spatial environmental impact assessment.     

Use of macroinvertebrate-based multimetric indices for water quality evaluation in Spanish Mediterranean rivers: an intercalibration approach with the IBMWP index

For the European Parliament and Commission to implement the Water Framework Directive (WFD), the water-quality indices that are currently used in Europe need to be compared and calibrated. This will facilitate the comparative assessment of ecological status throughout the European Union. According to the WFD, biologic indices should respond consistently to human impacts, using multimetric approaches and water-quality classification boundaries adjusted to a common set of normative definitions. The European Commission has started an intercalibration exercise to review biologic indices and harmonize class boundaries. We used data from rivers in Spain to compare the IBMWP (Iberian Biological Monitoring Working Party) index, which is commonly used by water authorities in Spain and by several research centers, with the Intercalibration Common Multimetric Index (ICM-Star), which was used as a standard in the intercalibration exercise. We also used data from Spanish rivers to compare the multimetric indices ICM-7 (based on quantitative data) and ICM-9 (based on qualitative data) with the IBMWP. ICM-7 and ICM-9 were proposed by the Mediterranean Geographical Intercalibration Group (Med-GIG). Additionally, we evaluated two new multimetric indices, developed specifically for macroinvertebrate communities inhabiting Mediterranean river systems. One of these is based on quantitative data (ICM-10), while the other is based on qualitative data (ICM-11a). The results show that the IBMWP index responds well to the stressor gradient present in our data, and correlates well with ICM-Star. Moreover, the IBMWP quality class boundaries were consistent with the intercalibration requirements of the WFD. However, multimetric indices showed a more linear relation with the stressor gradient in our data, and less variation in reference values. In addition, they may provide more statistical power for detecting potential environmental impacts. Multimetric indices produced similar results for quantitative and qualitative data. Thus, ICM-10 (also named IMMi-T) and ICM-11a (also named IMMi-L) indices could be used to meet European Commission requirements for assessing the water quality in Spanish Mediterranean rivers. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Handling editor: Joel Trexler     

Odonata (Insecta) as a tool for the biomonitoring of environmental quality

Despite the fundamental dependence of human populations on water resources, a range of anthropogenic impacts, in particular the removal of riparian vegetation, threaten freshwater environments. One of the most effective means of evaluating the effects of anthropogenic disturbance in aquatic ecosystems is the use of bioindicators, and the insects of the order Odonata are among the most efficient indicators, due to their enormous sensitivity to environmental changes. In this context, the present study aimed to verify which parameters of the odonate community (species richness, abundance/biomass, composition, taxonomic diversity and taxonomic/phylogenetic distinctness) are most effective for the evaluation of the loss of environmental integrity. The study focused on 50 streams in the northeast of the Brazilian state of Pará. The streams were sampled during the dry season, between June and August 2011. The physical characteristics of each stream were evaluated using a Habitat Integrity Index (HII). The species composition provided the best parameter for the evaluation of ecological integrity, providing a relatively accurate assessment at a lower mean research cost than other parameters. Taxonomic diversity and distinctness also provided relatively reliable results, contributed additional information on the evolutionary relationships among the odonate taxa, and also provided a low-cost approach. Deconstructing communities is necessary to detect impacts, considering the considerable variation in the environmental requirements of the different species. Overall, the parameter that best responded to gradients of disturbance was species composition, followed by diversity and taxonomic distinctness. Given these findings, odonate-based biomonitoring should focus on these parameters to guarantee the optimal detection and evaluation of habitat alterations.     

Developing a scuba trail vulnerability index (STVI): a case study from a Mediterranean MPA

Scuba diving is now one of the major form of commercial use of marine protected areas (MPAs) around the world and the control of its potential impacts on the marine environment represents a fundamental key to manage this recreational activity in highly dived areas. A potential tool to tackle such issues has been thought to be the definition of a value of recreational carrying capacity of an area, but this approach has been rarely considered management-effective. Therefore, the first step for effectively managing scuba-diving should be ‘bottom-up’: characterizing the benthic communities potentially affected by diving and evaluating their vulnerability. Aim of this paper is to propose a tool to define an index of vulnerability for dive trails (STVI: scuba trail vulnerability index). This has taken into consideration both physical and biological features of each trail. All the considered features are represented by non-quantitative variables, because either they are purely qualitative or their quantitative measurement is impractical. The management of such qualitative information and its translation into a formal methodology was performed by means of fuzzy logic, which has been repeatedly proposed as a powerful technique to develop indices of environmental quality. The approach adopted in this study provided a useful tool for the preliminary assessment of the potential vulnerability of benthic assemblages to scuba-diving and may represent an alternative method to the assessment of carrying capacity. The application of this index will enable management strategies for potentially reducing the degradation of benthic organisms/assemblages, and allowing a sustainable use of MPAs.     

The regulation index: a new method for assessing the relationship between oxygen consumption and environmental oxygen

Critical oxygen pressure (P(C)) is used in respiratory physiology to measure the response to hypoxia. P(C) defines the partial pressure of oxygen (Po(2)) at which an oxygen regulator switches to a conformer. However, not all animals show such clear patterns in oxygen consumption rate (Mo2), and there are many methods for determining P(C). This study assesses two methods that determine regulatory ability and four that calculate P(C). A new method, the regulation index (RI), assigns to an animal a relative measure of regulatory ability by calculating the area under the Mo2 versus Po(2) curve that is greater than a linear trend. The six methods are applied to developmental Mo2 data of two amphibians, Pseudophryne bibronii and Crinia georgiana. The four methods used to determine P(C) produced similar results but failed to identify the increase in regulation on hatching in C. georgiana or the greater regulation in larval C. georgiana compared with P. bibronii. Of the two methods that evaluated regulation, only the RI satisfactorily represented the entire range of Po(2). The RI is advantageous because it has clearly defined limits and does not constrain data to fit any single pattern. The RI can be used in concert with P(C), which can be easily calculated during the RI analysis, to provide a clearer definition of the Mo2 response to environmental Po(2).     

The response of Chironomidae (Diptera) to a long-term exclusion of terrestrial organic matter

We examined the effects of a seven-year detrital exclusion on chironomid assemblages in an Appalachian headwater stream. We hypothesized that litter exclusion would lead to a reduction in all chironomids at both the subfamily and generic levels because organic matter serves as both food and habitat in these headwater streams. Tanytarsini total abundance and biomass significantly declined after litter exclusion. Before litter exclusion, Tanytarsini average abundance was 4271 ± 1135 S.E. m⁻² and 625 ± 98 after litter exclusion. Biomass was 3.57 ± 0.96 mg AFDM m⁻² before litter exclusion and 1.03 ± 0.9 after exclusion. In contrast, Orthocladiinae abundance and biomass did not change because a psammanophilic chironomid, Lopescladius sp., and other Orthocladiinae genera did not decline significantly. Overall chironomid taxa richness and diversity did not change as a result of litter exclusion. However, Canonical Correspondence Analysis (CCA) of genus-level biomass did show a clear separation between the litter exclusion stream and a reference stream. Separation of taxa between the two streams was due to differences in fine (r ² = 0.39) and coarse (r ² = 0.36) organic matter standing stocks and the proportion of small inorganic substrates (r ² = 0.39) present within a sample. As organic matter declined in the litter exclusion stream, overall chironomid biomass declined and the chironomid community assemblage changed. Tanytarsini were replaced by Orthocladiinae in the litter exclusion stream because they were better able to live and feed on biofilm associated with inorganic substrates. Handling editor: K. Martens     

Development of a preliminary index of biotic integrity (IBI) based on fish assemblages to assess ecosystem condition in the lakes of central Mexico

The lakes of central Mexico have great cultural, economic, and biological value, but they are being degraded at an accelerating rate. We employed historical data on fish communities from 19 of these lakes and case studies of community responses to environmental degradation from four of the best-studied, Xochimilco, Cuitzeo, Chapala, and Pátzcuaro, to construct a preliminary index of biotic integrity (IBI). This IBI was designed to be an easily applied method for assessing lake ecosystem health and evaluating restoration efforts. The IBI had 10 metrics: number of total native species, number of common native species, number of native Goodeidae species, number of native Chirostoma species, number of native sensitive species, percent of biomass as tolerant species, percent of biomass as exotic species, percent of biomass as native carnivorous species, maximum standard length of native species, and percent of exotic invertebrate parasite species on or in native fishes. Initial applications of the index showed promise, accurately ranking the relative degradation of the four case-study lakes. Further tests of the index are warranted, and more data are needed to standardize sampling procedures, improve species classifications, and refine metric scoring criteria.     

The genus Antillophos Woodring, 1928 (Gastropoda: Buccinidae) from the China seas,with description of a new species

The species of genus Antillophos Woodring, 1928 Woodring, W.P. (1928) Miocene Mollusks from Bowden, Jamaica. 2. Gastropods and Discussion of Results. Contributions to the Geology and Paleontology of the West Indies. Carnegie Institute of Washington, Washington D.C. [Google Scholar] from the China seas are studied. Six species, Antillophos liui n. sp., Antillophos lucubratonis Fraussen & Poppe, 2005 Fraussen, K. & Poppe, G.T. (2005) Revision of Phos and Antillophos (Buccinidae) from the Central Philippines. Visaya 1, 76–115. [Google Scholar], Antillophos monsecourorum Fraussen & Poppe, 2005 Fraussen, K. & Poppe, G.T. (2005) Revision of Phos and Antillophos (Buccinidae) from the Central Philippines. Visaya 1, 76–115. [Google Scholar], Antillophos pyladeum (Kato, 1995 Kato, S. (1995) Discussion of the genus Phos. Hitaciobi 70, 15–20. [Google Scholar]), Antillophos roseatus (Hinds, 1844 Hinds, R.B. (1844) Mollusca. In: Hinds, R.B. (Ed.) The Zoology of the Voyage of H.M.S. Sulphur During the years 1836–1842. 2. Smith, Elder and Co., London, pp. 1–72. [Google Scholar]) and Antillophos sp., are described and illustrated.

http://zoobank.org/urn:lsid:zoobank.org:pub:51481997-A841-4F37-8E15-B753DC99CB4D     

Migratory flows in apulia: Application of a new index for the evaluation of socioanthropological stress

The evaluation of the socio-anthropological stress of every ethnic group which has recently immigrated to Apulia, (South Italy), is part of the attempt to quantify some aspects of the migratory phenomenon and to offer an overall picture in a large territory such as that of Apulia. This analysis was carried out thanks to the proposal of multifactor index elaborated to start from a series of brief and easily understood key questions administered to individual immigrants in Apulia; the index of synthetic stress (IDN), the theoretical parameter that measures the overage stress in the whole population, is, therefore, given by the average of the indexes of the units forming the sample. in Atti del XIII Congresso degli Antropologi Italiani, Roma, 4–7 ottobre 1999.