Chironomus plumosus larvae increase fluxes of denitrification products and diversity of nitrate-reducing bacteria in freshwater sediment

Benthic invertebrates affect microbial processes and communities in freshwater sediment by enhancing sediment-water solute fluxes and by grazing on bacteria. Using microcosms, the effects of larvae of the widespread midge Chironomus plumosus on the efflux of denitrification products (N₂O and N₂ + N₂O) and the diversity and abundance of nitrate- and nitrous-oxide-reducing bacteria were investigated. Additionally, the diversity of actively nitrate- and nitrous-oxide-reducing bacteria was analyzed in the larval gut. The presence of larvae increased the total effluxes of N₂O and N₂ + N₂O up to 8.6- and 4.2-fold, respectively, which was mostly due to stimulation of sedimentary denitrification; incomplete denitrification in the guts accounted for up to 20% of the N₂O efflux. Phylotype richness of the nitrate reductase gene narG was significantly higher in sediment with than without larvae. In the gut, 47 narG phylotypes were found expressed, which may contribute to higher phylotype richness in colonized sediment. In contrast, phylotype richness of the nitrous oxide reductase gene nosZ was unaffected by the presence of larvae and very few nosZ phylotypes were expressed in the gut. Gene abundance of neither narG, nor nosZ was different in sediments with and without larvae. Hence, C. plumosus increases activity and diversity, but not overall abundance of nitrate-reducing bacteria, probably by providing additional ecological niches in its burrow and gut.     

Short-term influence of recolonisation by the polycheate worm [2pt] Nereis succinea on oxygen and nitrogen fluxes and [2pt] denitrification: a microcosm simulation

The short-term effects of sediment recolonisation by Nereis succinea on sediment-water column fluxes of oxygen and dissolved inorganic nitrogen, and rates of denitrification, were studied in microcosms of homogenised, sieved sediments. The added worms enhanced oxygen uptake by the sediments, due to the increased surface area provided by the burrow walls and the degree of stimulation was stable with time. Similarly, ammonium fluxes to the water column were stimulated by N. succinea, but declined over the 3 day incubation in all microcosms including the controls. Nitrate fluxes were generally greater in the faunated microcosms, but highly variable with time. Denitrification rates were positively stimulated by N. succinea populations, denitrification of water column nitrate was stimulated 10-fold in comparison to denitrification coupled to nitrification in the sediments. Rates of denitrification of water column nitrate were not significantly different from rates in undisturbed sediment cores with similar densities of N. succinea, whereas rates of coupled nitrification–denitrification were 3-fold lower in the experimental set-up. These results may reflect the relative growth rates of nitrifying and denitrifying bacteria, which allow more rapid colonisation of new burrow surfaces by denitrifier compared to nitrifier populations. The data indicate that recolonisation by burrowing macrofauna of the highly reduced sediments of the Sacca di Goro, Lagoon, Italy, following the annual dystrophic crisis, may play a significant role in the reoxidation and detoxification of the sediments. The increased rates of denitrification associated with the worm burrows, may promote nitrogen losses, but due to the low capacity of nitrifying bacteria to colonise the new burrow structures, these losses would be highly dependent upon water column nitrate concentrations.     

The influence of chironomus plumosus larvae on the exchange of dissolved substances between sediment and water

4th instar Chironomus plumosus larvae (about 1000·m^–2) were added to tubes containing sediment and overlying water. At a temperature of 20°C the larvae greatly increased the trasnport of silica, phosphorus and iron from the sediment to the water. Oxygen concentrations did not influence the exchange of silica. For two non-calcareous sediments the exchange of phosphorus and iron was much higher under anaerobic than under aerobic conditions while the difference was small for sediment from a hardwater lake. Exchange of inorganic nitrogen was little influenced by added chironomid larvae.     

High-throughput sequencing of the mitochondrial genomes from archived fish scales: an example of the endangered putative species flock of Sevan trout <Emphasis Type="Italic">Salmo ischchan</Emphasis>

Burrowing benthic animals belonging to the same functional group may produce species-specific effects on microbially mediated nitrogen (N) processes depending upon different ecological traits. We investigated the effects of two tube-dwelling organisms, amphipods (Corophium insidiosum) and chironomid larvae (Chironomus plumosus), on benthic N cycling in bioturbated estuarine sediments. Aims of this work were to analyze the interactions among burrowers and N-related microbial processes in two distinct sedimentary environments colonized by benthic animals with different ecological traits. We hypothesized higher rates of nitrification and higher coupled nitrification–denitrification in sediments with C. insidiosum due to continuous ventilation rates. We expected higher denitrification of water column nitrate in sediments with C. plumosus due to lower and intermittent ventilation activity and lower oxygen levels in burrows. To this purpose, we combined process–specific (nitrification and denitrification) with net N flux measurements in intact and reconstructed sediments. Sediments with C. insidiosum had higher rates of oxygen demand and of potential nitrification and higher concentration of pore water NH₄⁺ as compared to sediments with C. plumosus. Sediments with both species displayed comparable net N₂ fluxes, mostly sustained by respiration of water column NO₃^? in sediments with chironomid larvae and by NO₃^? produced within sediments in sediments with corophiid amphipods. Corophium insidiosum stimulated nitrification nearly 15-fold more as compared to C. plumosus. Overall, our results demonstrate that sediments with burrowing fauna may display similar rates of denitrification, but underlying mechanisms may deeply vary and be species-specific.     

Denitrification in deep subsurface sediments

Dissimilatory nitrate reduction (denitrification) in subsurface sediments by indigenous microflora was investigated in samples obtained over a range of depths from 0 to 289 m. Denitrifying activity in sediment samples retrieved from similar stratigraphic horizons at four different sites was determined by measuring the accumulation of N₂O using the acetylene blockage technique. Denitrification was detected in unamended samples which received only prereduced deionized water at almost all depths in all sediments sampled. The surface sediments showed the highest denitrification activity. In the deeper sediments, denitrifying activity was much higher in saturated sandy samples and lower or absent in drier clay samples. Addition of nitrate enhanced denitrification activity in all samples from below the water table down to the maximum depth sampled (289 m), while addition of a carbon (succinate) source in general had no stimulatory effect. These results show that denitrifying microorganisms were present in all of the deep subsurface sediments tested in this study. Furthermore, these results suggest that adequate supplies of metabolizable organic carbon were available to support denitrifying activity. However, denitrification may be limited by inadequate supplies of nitrate in the sediments.     

Microelectrode Measurements of Nitrate Gradients in the Littoral and Profundal Sediments of a Meso-Eutrophic Lake (Lake Vechten, The Netherlands)

NO₃⁻ concentration profiles were measured in the sediments of a meso-eutrophic lake with a newly developed microelectrode. The depth of penetration of NO₃⁻ varied from only 1.3 mm in organic-rich profundal silty sediments to 5 mm in organic-poor littoral sandy sediments. The thickness of the zone of denitrification in the organic-rich sediments was <500 μm. Oxygen profiles measured simultaneously revealed that the zone of denitrification was directly adjacent to the aerobic zone. The results demonstrate high denitrification rates (0.26 to 1.31 mmol m⁻² day⁻¹) at in situ nitrate concentrations in the overlying water (0.030 mmol liter⁻¹) and limitation of denitrification by nitrate availability.     

Denitrification in Marl and Peat Sediments in the Florida Everglades

The potential for denitrification in marl and peat sediments in the Shark River Slough in the Everglades National Park was determined by the acetylene blockage assay. The influence of nitrate concentration on denitrification rate and N₂O yield from added nitrate was examined. The effects of added glucose and phosphate and of temperature on the denitrification potential were determined. The sediments readily denitrified added nitrate. N₂O was released from the sediments both with and without added acetylene. The marl sediments had higher rates than the peat on every date sampled. Denitrification was nitrate limited; however, the yields of N₂O amounted to only 10 to 34% of the added nitrate when 100 μM nitrate was added. On the basis of measured increases in ammonium concentration, it appears that the balance of added nitrate may be converted to ammonium in the marl sediment. The sediment temperature at the time of sampling greatly influenced the denitrification potential (15-fold rate change) at the marl site, indicating that either the number or the specific activity of the denitrifiers changed in response to temperature fluctuations (9 to 25°C) in the sediment. It is apparent from this study that denitrification in Everglades sediments is not an effective means of removing excess nitrogen which may be introduced as nitrate into the ecosystem with supply water from the South Florida watershed and that sporadic addition of nitrate-rich water may lead to nitrous oxide release from these wetlands.     

Linkages between organic matter mineralization and denitrification in eight riparian wetlands

Denitrification (N₂ production) and oxygen consumption rates were measured at ambient field nitrate concentrations during summer in sediments from eight wetlands (mixed hardwood swamps, cedar swamps, heath dominated shrub wetland, herbaceous peatland, and a wetland lacking live vegetation) and two streams. The study sites included wetlands in undisturbed watersheds and in watersheds with considerable agricultural and/or sewage treatment effluent input. Denitrification rates measured in intact cores of water-saturated sediment ranged from 20 to 260 mol N m^-2 h^-1 among the three undisturbed wetlands and were less variable (180 to 260 mol N M^-2 h^-1) among the four disturbed wetlands. Denitrification rates increased when nitrate concentrations in the overlying water were increased experimentally (1 up to 770 M), indicating that nitrate was an important factor controlling denitrification rates. However, rates of nitrate uptake from the overlying water were not a good predictor of denitrification rates because nitrification in the sediments also supplied nitrate for denitrification. Regardless of the dominant vegetation, pH, or degree of disturbance, denitrification rates were best correlated with sediment oxygen consumption rates (r ² = 0.912) indicating a relationship between denitrification and organic matter mineralization and/or sediment nitrification rates. Rates of denitrification in the wetland sediments were similar to those in adjacent stream sediments. Rates of denitrification in these wetlands were within the range of rates previously reported for water-saturated wetland sediments and flooded soils using whole core¹⁵N techniques that quantify coupled nitrification/denitrification, and were higher than rates reported from aerobic (non-saturated) wetland sediments using acetylene block methods.     

The effect of tubificids and chironomids on particle redistribution of lake sediment

The effects of chironomid larvae,Chironomus plumosus, and tubificid worms,Limnodrilus spp., on particle redistribution in lake sediment were investigated experimentally using pots containing sediments obtained from Lake Suwa, Japan. The chironomids and tubificids increased the water content of surface sediment. The chironomid larvae had no effect on particle size distribution, while tubificids continuously accumulated small particles on the surface sediment through their selective feeding activity. Particles larger than 0.125 mm were buried at a sediment depth of 6 cm. In Lake Suwa, long diatom frustules, large plant debris and blue-green algal flocs were found to accumulate in the deeper layer of the lake sediment inhabited by tubificids at high density.     

The Fate of 15N-Nitrate in Healthy and Declining Phragmites australis Stands

Abstract The dissimilatory nitrate-reducing processes, denitrification, and dissimilatory nitrate-reduction to ammonium were studied in freshwater lake sediments within healthy and degrading Phragmites australis (reed) stands. The samples from the healthy vegetation site contained roots and rhizomes. Cores were supplied with 1.9–5.2 μg ¹⁵N-NO₃ ⁻ g⁻¹ dry sediment in the laboratory and subsequently incubated for 8 h at 20°C, in the dark. The ¹⁵N compounds were determined before (natural percentage of ¹⁵N) and after 1 and 8 h of incubation. The uptake of ¹⁵N by the roots and rhizomes in the healthy vegetation was 61%. Nitrogen losses, interpreted as denitrification, accounted for 25 and 84% of the added ¹⁵N-NO₃ ⁻ in sediment from the healthy and degrading vegetation sites, respectively. The percentages of nitrate reduced to ammonium were 4 and 9% in sediment from the healthy vegetation and degrading vegetation sites, respectively. The percentage of ¹⁵N–total N in the sediment of the healthy vegetation site was 10%, whereas for the degrading vegetation site this percentage was 7%. The percentage of nitrate reduced to ammonium could be potentially underestimated by the percentage of ¹⁵N measured in the sediment. In this case, in healthy and degenerating P. australis stands, the percentage of produced ammonium accounted for 14–16%. The nitrate reduction rates were calculated based on an incubation period of one hour. The denitrification rate in sediment from the degrading vegetation site was higher than from the healthy vegetation site. The rate of dissimilatory nitrate reduction to ammonium was almost tenfold higher in sediment from the degrading vegetation site compared to sediment from the healthy vegetation site. The significantly lower percentages of dissimilatory nitrate reduction to ammonium and denitrification in the healthy stand compared to the degrading stand was probably due to the presence of roots and rhizomes. In the sediments of healthy and degrading P. australis stands, denitrification was the main nitrate-reducing process. Received: 24 July 1996; Accepted: 5 December 1996     

Environmental gradients determining the distribution of benthic macroinvertebrates in Lake Takkobu, Kushiro wetland, northern Japan

Effects of environmental variables on the distribution of benthic macroinvertebrates inhabiting sediments were studied at 25 sites along the shoreline of Lake Takkobu in the Kushiro wetland of northern Japan in summer 2003. During the last decade, the lake’s status has undergone a drastic shift from clear water dominated by submerged macrophytes to turbid water dominated by phytoplankton. The canonical correspondence analysis showed that four environmental variables explained the significant variation in the macroinvertebrate species composition: submerged plant biomass, bottom sediment organic matter content (OMC), distance from the mouth of the Takkobu River, and bottom-layer pH. Five species of Chironomidae [Chironomus sp. (except plumosus group), Psectrocladius sp., Corynoneura sp., Parachironomus sp. arcuatus group, and Zavreliella sp.] occurred in sites with relatively lower pH and a high submerged plant biomass, whereas three species of Tubificidae (Tubifex tubifex, Aulodrilus limnobius and Aulodrilus sp.) and two of Chironomidae (Nanocladius sp. and Monodiamesa sp.) occurred in sites with high pH and little vegetation. The three Tubificidae species also preferred organic-rich sediments. Irrespective of aquatic vegetation, Sphaerium sp. (Bivalvia) and Monodiamesa sp. (Chironomidae) occurred in low-OMC sites, whereas Tanypus sp. (Chironomidae) preferred high-OMC sites. The number of macroinvertebrate taxa showed the highest correlation with the number of submerged plants, suggesting that macroinvertebrate species richness was related mostly to submerged plant species diversity in this lake. The quantity and species richness of submerged plants and OMC are thus important determinants of the community structure of macroinvertebrates inhabiting sediments in Lake Takkobu.     

The denitrification capacity of sediment from a hypereutrophic lake

SUMMARY.

• 1 In sediment from Wintergreen Lake, Michigan, denitrification was not detectable by the acetylene inhibition method at in situ nitrate concentrations. When nitrate was added to sediment slurries, denitrification capacities up to 18.8μg N g^-1 h^-1 were measured. The denitrification capacities decreased with increasing sediment depth and distance from shore.
• 2 The high denitrification capacities in these sediments which under natural conditions had no supply of nitrate and oxygen suggested that denitrifies with alternative mechanisms for anaerobic energy conversion were present. Nitrous oxide was a significant portion of the N-gas produced immediately after the nitrate addition. Small amounts (4–5% of the total N-gas production) of nitric oxide accumulated in the early phase of nitrate reduction. Presumably after depletion of nitrate and nitrite both N₂O and NO were further reduced to N₂.
• 3 About 70%r of the added nitrate was denitrified, and the remainder was assumed to have been reduced to ammonium.

     

Spatio-temporal variability in benthic mineralization processes in the eastern English Channel

The effect of phytodetritus derived from Phaeocystis sp. bloom on benthic mineralization processes has been determined at four intertidal stations along the French coast of the eastern English Channel. Sites were chosen to offer a diversity of sediment types, from permeable sandy beach to estuarine mudflats. Sediment Oxygen Demand (SOD) as well as total fluxes of Dissolved Inorganic Nitrogen (DIN) at the sediment–water interface were determined by using whole core incubation technique and diffusive fluxes were predicted from interstitial water concentrations. In the absence of phytodetritus deposits, a marked gradient of granulometric characteristics and organic matter contents were observed, and resulted in more intensive mineralization processes in muddy sediments. Highly significant correlations (P < 0.05) were evidenced between SOD and porosity, bacterial biomass, Organic Carbon and Organic Nitrogen, evidencing the direct link between sediment texture, organic matter accumulation and microbial activity. The spring bloom led to a massive input of organic matter in surficial sediments and mineralization rates significantly increased while higher DIN release towards the water column was observed. A modification of the mineralization pathways was evidenced but clearly depended on the sediment type. With a global view, benthic mineralization processes in the intertidal zone provided significant a part of DIN inputs in the coastal zone while water column was depleted in nutrients.     

Chironomidae communities of the near-shore zone in Zegrzyńsky Reservoir,Poland

The structural differences in chironomid communities of the near-shore zone in Zegrzyski Reservoir corresponded to variations in sediments composition. Sandy bottom areas covered by a thin layer of mud were inhabited mostly byPolypedilum, Cryptochironomus andGlyptotendipes larvae.Cladotanytarsus andStictochironomus were often the most numerous in the shallower zone (0.2–0.5 m). Predatory Tanypodinae were found most commonly on the sandy bottom covered by well mineralized plant debris and loose silt. Extremely high numbers and biomass ofChironomus andGlyptotendipes larvae occupied the clay bottom covered by a thick layer of mud. On the basis of developmental stage distribution it was found that at least three generations ofChironomus andGlyptotendipes occurred during the study period (April–October 1988). The combined production estimate for these two taxa exceeds any reported for Chironomidae in lentic or lotic habitats.     

Instar distribution and biomass of Chironomidae larvae in Lago El Junco, Isla San Cristobal, the Galápagos

The distribution, abundance and standing crop biomass of chironomid larvae were determined at one-meter depth intervals along three radial transects. Samples were collected by coring soft sediments while diving. Three genera were found in the lake: Chironomus sp. (collector-filtering larvae), Ablabesmyia sp. (predatory larvae) and Goeldichironomus sp. (collector-filtering larvae). Standing crop densities of chironomids, averaged over the entire lake, varied from 30,594 larvae/m² to 11,428 larvae/m² at different depths. No statistically significant zonation in density was found for the two most common taxa, Chironomus sp. (87.8% of specimens) and Ablabesmyia sp. (9.0%), however the deepest zones (>4 m) had the lowest estimated densities. Significant differences in standing crop biomass were detected, with the 6 m depth having greatest biomass. The increase in standing crop biomass was a function of (1) lower frequency of first instars of Chironomus sp. and Ablabesmyia sp. at 6 m (2) higher average larval biomass of both species at 6 m and (3) very significant increase in average biomass of fourth instars of Chironomus sp. at 6 m compared to fourth instars at shallower depths. These results indicate that the lentic chironomids of this isolated oceanic habitat consist of a small number of species that are ecological generalists and tolerant of low oxygen concentrations.     

Vertical Distribution of Denitrification in an Estuarine Sediment: Integrating Sediment Flowthrough Reactor Experiments and Microprofiling via Reactive Transport Modeling

Denitrifying activity in a sediment from the freshwater part of a polluted estuary in northwest Europe was quantified using two independent approaches. High-resolution N₂O microprofiles were recorded in sediment cores to which acetylene was added to the overlying water and injected laterally into the sediment. The vertical distribution of the rate of denitrification supported by nitrate uptake from the overlying water was then derived from the time series N₂O concentration profiles. The rates obtained for the core incubations were compared to the rates predicted by a forward reactive transport model, which included rate expression for denitrification calibrated with potential rate measurements obtained in flowthrough reactors containing undisturbed, 1-cm-thick sediment slices. The two approaches yielded comparable rate profiles, with a near-surface, 2- to 3-mm narrow zone of denitrification and maximum in situ rates on the order of 200 to 300 nmol cm⁻³ h⁻¹. The maximum in situ rates were about twofold lower than the maximum potential rate for the 0- to 1-cm depth interval of the sediment, indicating that in situ denitrification was nitrate limited. The experimentally and model-derived rates of denitrification implied that there was nitrate uptake by the sediment at a rate that was on the order of 50 (± 10) nmol cm⁻² h⁻¹, which agreed well with direct nitrate flux measurements for core incubations. Reactive transport model calculations showed that benthic uptake of nitrate at the site is particularly sensitive to the nitrate concentration in the overlying water and the maximum potential rate of denitrification in the sediment.     

The relationships between relative growth rate,meristematic potential and compensatory growth of semiarid-land shrubs

The submersed macrophyte Vallisneria americana was grown for seven weeks in a greenhouse to test for differences in the ability of three different sediments to support growth stimulation in response to CO₂ enrichment at low pH. Plants accumulated 21- to 24-fold greater biomass at 10 × ambient CO₂ concentrations than at ambient CO₂ on all sediments. At both CO₂ levels, plants grown on sediment from an acidified lake accumulated ca. 81%, and those grown on oligotrophic lake sediment ca. 47% as much biomass as plants grown on alkaline lake sediment. Despite striking CO₂ and sediment effects on biomass accumulation, there was no significant interaction (using log-transformed data) between CO₂ and sediment effects, indicating that all sediments allowed similar proportionate growth responses to CO₂ enrichment. Plants grown on the less fertile sediments showed greater relative allocation to horizontal versus vertical growth by producing more rosette-bearing stolons in relation to plant height (leaf length) than plants grown on relatively fertile, alkaline lake sediment. Tissue analysis suggested that sediment effects on Vallisneria growth could be attributed neither to mineral putrient (nitrogen and phosphorus) limitation nor to aluminum toxicity in these low pH treatments. In any case, CO₂ availability can be an important regulator of submersed macrophyte growth at low pH on a variety of sediment types, including those from oligotrophic and acidic lakes.     

High rates of denitrification and nitrate removal in cold seep sediments

We measured denitrification and nitrate removal rates in cold seep sediments from the Gulf of Mexico. Heterotrophic potential denitrification rates were assayed in time-series incubations. Surficial sediments inhabited by Beggiatoa exhibited higher heterotrophic potential denitrification rates (32 μ N reduced day⁻¹) than did deeper sediments (11 μ N reduced day⁻¹). Nitrate removal rates were high in both sediment horizons. These nitrate removal rates translate into rapid turnover times (<1 day) for the nitrate pool, resulting in a faster turnover for the nitrate pool than for the sulfate pool. Together, these data underscore the rigorous nature of internal nitrogen cycling at cold seeps and the requirement for novel mechanisms to provide nitrate to the sediment microbial community.     

Resource allocation in the submerged plant Vallisneria natans related to sediment type, rather than water-column nutrients

1. Phenotypic plasticity in resource allocation by Vallisneria natans was investigated in a greenhouse experiment, using three types of sediment [sandy loam, clay, and a 50 : 50 (by volume) mixture of the two sediments] and two levels of water‐column nutrient. The clay was collected from a highly eutrophic lake in Jiangsu Province, China, and the N and P concentrations applied in nutrient media were at the upper limits observed in most lakes of China. 2. Growth and biomass allocation were significantly affected by sediment type, rather than water‐column nutrients. Plant growth in clay and the mixture were similar, and 2.4–3.4 times higher than that in sandy loam. Compared with the plants grown in clay or the mixed sediments, the plants grown in sandy loam allocated relatively more biomass to root (11–17% versus 7–8% of total biomass), and relatively less to leaf (76–82% versus 86–87% of total biomass). Plastic variations in root area were induced by sediment type alone (P < 0.05), whereas the impacts of sediment type and water‐column nutrients on leaf area were insignificant (P > 0.05). 3. Plant N and P concentrations were significantly affected by both sediment type and water‐column nutrients. Increased nutrient availability in the water column enhanced plant N concentration by 3.5–20.2%, and plant P concentration by 19.1–25.8%. 4. Biomass accumulation and plant nutrient concentration in plants grown in different sediment types and water‐column nutrients indicate that sediment type had more significant impacts on growth and N and P concentrations of V. natans than did water‐column nutrients. Changes in phenotype are a functional response to nutrient availability in sediment, rather than to water‐column nutrients.     

Function of Ventral and Lateral Processes in Larvae of Chironomus (Diptera, Chironomidae)

Oxygen consumption by larvae of five Chironomus species from the Plumosus group belonging to different larval forms was studied. Larvae of C. agilis ( f. l. reductus) were unable to utilize oxygen at its concentrations lower than 1 mg/l. Larvae of C. plumosus ( f. l. plumosus) used oxygen from water at concentrations higher than 0.7 mg/l; pupas of this species and larvae of C. borokensis ( f. l. plumosus) utilized oxygen at concentrations lower than 0.4 mg/l. C. entis and C. muratensis (both-- f. l. semireductus) utilized oxygen from water at its concentrations up to 0.05 mg/l. The lack of correlation between the rate and ability of larvae to use oxygen at low concentrations and the degree of development of processes allows suggesting that these structures are not additional respiratory organs, as is commonly accepted. At the same time, ability of species to inhabit under conditions of oxygen deficiency correlates directly with the size of the processes. Therefore, ventral and lateral processes are suggested to perform a function of excretion of anaerobic metabolism metabolites.