Heavy metal concentrations in common freshwater snails of Azraq Oasis,Jordan

Heavy metal concentrations in three freshwater snails, an aquatic plant, sediment, and water from Azraq Oasis pools in the Jordanian desert were measured by atomic absorption spectrophotometry. There were significant differences in metal accumulation among snail species. These snails are ranked according to their metal accumulation capacity in the order: Melanoides tuberculata > Melanopsis praemorsa > Theodoxus jordani. Although there were no significant differences among the size classes of these snails in metal concentrations, larger snails contained generally higher levels of metals than the smaller ones. The concentration factors for the selected snails exceeded 10³, while it ranged from 0.1–10.2 × 10³ for Typha angustata plants. Among sampling sites, metal concentrations in M. praemorsa, T. angustata, sediment, and water were not significantly different, except for Fe in M. praemorsa which was highest at station 3, and Cd and Cr which were highest in water and sediment of station 4. The seasonal variation in metal content was found to be species and metal-dependent.     

First report of Melanoides tuberculata (Mollusca: Thiaridae) harboring a xiphidiocercaria in Brazil: A new parasite introduced in the Americas?

Xiphidiocercariae were found in the invasive snail Melanoides tuberculata collected during a malacological survey in Ceará-Mirim, State of Rio Grande do Norte, Northeastern Brazil in November 2018 and submitted to morphological and molecular analyses. The morphology revealed similarities between the larvae here reported for the first time in M. tuberculata from Brazil and other xiphidiocercariae described in thiarid snails from Asia and Africa. Phylogenetic analyses based on 28S and ITS-2 sequences revealed that the larvae correspond to an unidentified species of the family Lecithodendriidae. Aspects related to the morphology and taxonomy of xiphidiocercariae found in M. tuberculata are briefly discussed. It is possible that the parasite here reported is a newly introduced species transmitted by M. tuberculata in the American continent.     

Adsorptive Removal and Recovery of U(VI), Cu(II), Zn(II), and Co(II) from Water and Industry Effluents

Tamarind fruit shell (TFS) was converted to a cation exchanger (PGTFS-SP-COOH) having a carboxylate functional group at the chain end by grafting poly(hydroxyethylmethacrylate) onto TFS (a lignocellulosic residue) using potassium peroxydisulfate-sodium thiosulfate redox initiator, and in the presence of N, N ′-methylenebisacrylamide as a cross-linking agent, followed by functionalization. The chemical modification was investigated using Fourier transform infrared (FTIR), X-ray diffraction (XRD), and potentiometric titrations. The feasibility of PGTFS-SP-COOH for the removal of heavy metals such as U(VI), Cu(II), Zn(II), and Co(II) ions from aqueous solutions was investigated by batch process. The optimum pH range for the removal of meal ions was found to be 6.0. For all the metal ions, equilibrium was attained within 2 h. The kinetic and isotherm data, obtained at optimum pH value 6.0, could be fitted with pseudo-second-order equation and Sips isotherm model, respectively. The Sips maximum adsorption capacity for U(VI), Cu(II), Zn(II), and Co(II) ions at 30°C was found to be 100.79, 65.69, 65.97, and 58. 81 mg/g, respectively. Increase of ionic strength decreased the metal ion adsorption. Different wastewater samples were treated with PGTFS-SP-COOH to demonstrate its efficiency in removing metal ions from wastewater. The adsorbed metal ions on PGTFS-SP-COOH can be recovered by treating with 1.0 M NaCl + 0.5 M HCl for U(VI) ions and 0.2 M HCl for Cu(II), Co(II), and Zn(II) ions. Four adsorption/desorption cycles were performed without significant decrease in removal capacity. The results showed that PGTFS-SP-COOH developed in this study exhibited considerable adsorption potential for the removal of U(VI), Cu(II), Zn(II), and Co(II) ions from water and wastewaters.     

Trace Metal Concentration in a Temperate Freshwater Reservoir Seasonally Subjected to Blooms of Toxin-Producing Cyanobacteria

In situ interactions between cyanobacteria and metals were studied at Torrão reservoir (Tâmega River, North Portugal). The metal content of water and sediments from the reservoir was monitored monthly at Marco de Canaveses (seasonally subjected to toxic blooms of Microcystis aeruginosa) and upstream at Amarante (no blooms recorded), for 16 months. During the 16 months of the study period, M. aeruginosa bloomed twice at Marco de Canaveses, firstly forming a scum, and later with colonies scattered throughout the reservoir. Metals Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb and Zn were analysed in the sediment and in the water column. Cu-binding ligands in water were also determined. When no blooms were taking place, average metal levels for water and sediment were not statistically different at both locations. Therefore, it was considered that the absence of cyanobacteria blooms at Amarante was not due to differences in metal content. When blooms were taking place at Marco de Canaveses, a significant increase of metal levels in the sediment occurred simultaneously. Sediment quality guidelines showed that during this period, Cu and Pb concentrations (32.3 and 43.2 mg kg^?1, respectively) were potentially toxic. However, quantification of the exchangeable metal fraction indicated that these metals were probably not bioavailable. Concentration of Cu-binding ligands in water was higher during the blooms, indicating that cyanobacteria are capable of changing the metal speciation in situ in a reservoir.     

Biosorption of Cadmium from Aqueous Solution by Shell Dust of the Fresh Water Snail Melanoides tuberculata

ABSTRACT The metal bioadsorption potential of shell dust of the freshwater snail Melanoides tuberculata (MTSD) was evaluated under laboratory conditions using cadmium as a model metal. As bioadsorbent, MTSD exhibited a biosorption capacity of 27.03 mg g^?1 at pH 6, indicating potential to remove cadmium from aqueous solution. The adsorption data fit more to the Langmuir (R² = 0.998) equation than the Freundlich (R² = 0.761) equation at equilibrium condition. The kinetics of biosorption followed the pseudo-second-order model (R² = 0.999) better than the Lagergren model (R² = 0.676), as was evident from the regression analysis. The presence of calcium ions appears to have facilitated ion exchange with cadmium along with the binding of different functional groups, as revealed through Fourier transform infrared (FT-IR) analysis. It is apparent from these observations that MTSD can act as low-cost and efficient bioadsorbent for cadmium bioremediation from aquatic habitats. Use of the shells of M. tuberculata for metal biosorption will promote the utility of a waste material of biological origin for bioremediation of heavy metals such as cadmium.     

Clonal diversity driven by parasitism in a freshwater snail

One explanation for the widespread abundance of sexual reproduction is the advantage that genetically diverse sexual lineages have under strong pressure from virulent coevolving parasites. Such parasites are believed to track common asexual host genotypes, resulting in negative frequency‐dependent selection that counterbalances the population growth‐rate advantage of asexuals in comparison with sexuals. In the face of genetically diverse asexual lineages, this advantage of sexual reproduction might be eroded, and instead sexual populations would be replaced by diverse assemblages of clonal lineages. We investigated whether parasite‐mediated selection promotes clonal diversity in 22 natural populations of the freshwater snail Melanoides tuberculata. We found that infection prevalence explains the observed variation in the clonal diversity of M. tuberculata populations, whereas no such relationship was found between infection prevalence and male frequency. Clonal diversity and male frequency were independent of snail population density. Incorporating ecological factors such as presence/absence of fish, habitat geography and habitat type did not improve the predictive power of regression models. Approximately 11% of the clonal snail genotypes were shared among 2–4 populations, creating a web of 17 interconnected populations. Taken together, our study suggests that parasite‐mediated selection coupled with host dispersal ecology promotes clonal diversity. This, in return, may erode the advantage of sexual reproduction in M. tuberculata populations.     

Myriophyllum aquaticum as a biomonitor of water heavy metal input related to agricultural activities in the Xanaes River (Córdoba,Argentina)

The aim of the present study was to assess the temporal variation of the heavy metal content (Co, Cu, Fe, Mn, Ni, Pb, and Zn) in surface water and sediments in relation to agricultural practices in the Xanaes River (Córdoba, Argentina). A second objective was to analyze possible relationships between the input of heavy metals on surface water and sediment, heavy metal accumulation and physiological changes in the aquatic plant Myriophyllum aquaticum. Samples were taken from the river at two contrasting sites (between April 2010 and August 2010): (1) a pristine area (mountain site), and (2) an area with intensive agricultural activity located at 60 km down river (agricultural site). The total concentration of heavy metals in surface water was higher in samples collected at the agricultural site but in sediments only the Mn concentration was higher than at the mountain site. The Fe and Mn concentrations in surface water at the agricultural site exceeded the recommended values for Argentinean Legislation of 300 μg L⁻¹ for Fe and 100 μg L⁻¹ for Mn. The accumulations of Zn and Mn in M. aquaticum were higher at the agricultural site and more elevated than the Zn and Mn concentrations in sediments at the same sites and sampling times. At the agricultural site, temporal variations of Cu, Fe and Zn were relatively similar for plants and water column, but the levels of the metals in plants were displaced over time. These results suggest that the levels of pollutants in the river came in pulses from the riverbank. These results show the potential use of M. aquaticum as a suitable accumulation biomonitor at the early stages of heavy metal pollution in rivers.     

Accumulation of heavy metals by aquatic mosses. 2: Rhynchostegium riparioides

A study was made of general ecology and metal accumulation in the widespread aquatic moss Rhynchostegium riparioides, (Hedw.) C. Jens. with a view to developing the use of this species as a monitor of heavy metal pollution. In order to establish a data bank for statistical analysis, samples of water and moss were taken within a 6-week period from 105 sites (10-m reaches) in Northern England from streams and rivers of diverse physical and chemical types. Analyses were made of 14 metals (Na, Mg, K, Ca, Cr, Mn, Fe, Co, Ni, Cu, Zn, Cd, Ba, Pb) in both 2-cm tips and whole plants. The same 14 metals were also measured in both total and filtrable water, together with 12 other variables. Samples of tips were easier to prepare for analysis, but had significantly (p < 0.001) lower concentrations of all metals except Na and K. Significant correlations (p < 0.001) between metal in moss and aqueous metal were found for ten metals (Na, K, Mg, Ca, Mn, Cu, Zn, Cd, Ba, Pb). Correlations between metals in moss and in water were in general similar for tips and whole plants, but much higher for tips with Na, Zn and Cd; the relationship was quite similar whether total or filtrable water was considered, with the exception of Ba where the correlation was much higher with the latter. A multiple regression was used to suggest which variables in water and/ or moss may influence accumulation of Co, Ni, Cu, Zn, Cd, Ba and Pb in the moss. For instance, the variables which had a very highly significant effect on Pb in tips were Pb, filtrable reactive phosphate and Zn in the water. A discussion is included of how the data may be used for monitoring purposes.     

Determination of water quality,and trace metals in endemic Sarotherodon linellii,Pungu maclareni and Clarias maclareni,in crater Lake Barombi Mbo,Cameroon

Considering water pollution as a potential threat to some endemic cichlids of Lake Barombi Mbo, Cameroon, an investigation was done in 2011 to determine trace metals in its water, linking their uptake in gills and liver of fish to water chemistry. ICP-MS and ICP-OES analyses of trace metals based on total concentration of unfiltered lake water samples showed the presence of trace metals. All fish species accumulated Al, Mn and Sr in the highest concentrations in their gills, with Cu, Cd, Co, Cr, Pb and U highest in the liver. Pungu maclareni accumulated Al, Cr, Co, Sr and Pb in the highest concentrations. The highest mean gill Al concentration of 140 µg g^?1 dry weight was measured in P. maclareni gills, this being one of the critically endangered cichlids of the lake. Stable isotope analyses of carbon δ¹³C and nitrogen δ¹⁵N showed that P. maclareni had the highest mean δ¹³C (?30.2‰) and highest concentrations of Cr, Co, Pb and U in liver, probably linking the carbon source to the accumulation of metals. Though trace metal levels in the lake water were low, their presence in fish tissues suggest they are bioavailable, bioaccumulate and may pose a threat to the aquatic biota, and therefore should be monitored.     

Heavy Metal Distribution in Open Canals and Drains in the Upper Rio Grande Basin

The inclusion of reclaimed effluents for irrigated agriculture may allow communities along the U.S.-Mexico border to use water resources more efficiently and reduce urban costs of wastewater treatment. However, metal inflow from industry and urbanization may threaten food safety. This study examines metal concentrations in sediments from open canal systems charged with flow from the Rio Grande and from effluents discharged from border communities. At the surface of canal beds, sediments were collected from six canal segments that began at the fringe of the El Paso/Juarez metroplex to rural areas downstream, and ranged from 9 to 24?km in length. Sediments were analyzed for Cd, Co, Cr, Cu, Ni, Pb, and Zn. These metals rarely exceeded 20?mg kg^?1. Drainage and effluent conveyance increased the variability of metal concentrations in sediments. Geostatistical models did not significantly account for spatial variability of metals, except in Mexico. This may implicate multiple rather than single inflow sources of metals. Peaks in metal concentrations often coincided with growing rural communities. However, most metal concentrations were within conventional global ranges and were not at levels high enough to threaten food safety. Future sampling strategies will require reduced sampling intervals from 1 to <0.13?km.     

Bioaccumulation of alkaline soil metals (Ca,Mg) and heavy metals (Cd,Ni, Pb) patterns expressed by freshwater species of Ulva (Wielkopolska,Poland)

The paper includes results of the first study on the accumulation of selected metals in freshwater populations of Ulva taxa at 16 sites in Poland. The thalli examined contained very high concentrations of Ca and Mg, owing to well‐developed surface incrustations of carbonate. Among the heavy metals investigated, the most significant concentration in the thalli was Ni, whereas the lowest was Pb. The median concentrations of Ni and Cd in the freshwater Ulva thalli were significantly higher than those reported in the available literature for marine taxa of the same genus. Several statistically significant correlations were found when the concentrations of different metals in the thalli and the water were analysed (© 2012 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Isolation and identification of a plant growth inhibitor from Tinospora tuberculata Beumee

Tinospora tuberculata Beumee has been used widely as a folk medicine and several bioactive compounds have been isolated. However, no herbicidal compound has been reported in this species. Therefore, we investigated the presence of phytotoxins in T. tuberculata. The aqueous methanol extracts of T. tuberculata inhibited the growth of roots and shoots of cress (Lepidum sativum L.), lettuce (Lactuca sativa L.), timothy (Phleum pratense L.) and barnyard grass (Echinochloa crus-galli (L.) Beauv.). The extract was then purified by several chromatographic runs with monitoring the inhibitory activity and the main phytotoxic substance was isolated. The chemical structure of the compound was determined by spectral data as syringin (4-[(1E)-3-Hydroxyprop-1-en-1-yl]-2,6-dimethoxyphenyl β-d-glucopyranoside). It inhibited the root and shoot growth of all test plant species at concentrations >10 µM. The concentrations required for 50 % inhibition of root and shoot growth of cress and lettuce ranged from 78.2 to 412 μM, and that of timothy and barnyard grass renged from 9.8 to 73.2 µM. Effectiveness of syringin on monocotyledonous (timothy and barnyard grass) plants was greater than that on dicotyledonous (cress and lettuce) plants. These results suggest that syringin may contribute to the allelopathic effect caused by the T. tuberculata extract.     

Eye trematode infection in small passerines in Peru caused by Philophthalmus lucipetus,an agent with a zoonotic potential spread by an invasive freshwater snail

Until now, four species of eye trematodes have been found in South America. Of them, Philophthalmus lucipetus (synonymized with Philophthalmus gralli) displays a broad host spectrum, with at least 30 bird species (prevalently large water birds), five mammal species and humans serving as definitive hosts, and with snails Fagotia (Microcolpia) acicularis, Amphimelania holandri, Melanopsis praemorsa and Melanoides tuberculata serving as intermediate hosts. When examining a total of 50 birds of ten species in the wetland of Pantanos de Villa, Lima, Peru in July 2011, eye trematodes were identified visually in the edematous conjunctival sac of 11 (48%) out of 23 resident many-colored rush tyrants Tachuris rubrigastra. Based on morphometric characteristics, the trematodes were identified as P. lucipetus. ITS2 and CO1 gene of the examined specimens combined showed a 99% similarity to an Iranian isolate of Philophthalmus sp. from the intermediate host Melanoides tuberculata, an invasive freshwater snail, suggesting that these two isolates represent the same species with a wide geographical range. Moreover, the prevalence of infection with the philophthalmid cercariae was 31% in 744 Melanoides tuberculata examined in Pantanos de Villa in 2010. It is evident that P. lucipetus occurs throughout the world as well as locally, including Eurasia and South America. Here we report this trematode for the first time in Peru, and we were the first to sequence any of the South American eye trematodes. Low host specificity of P. lucipetus and the invasive character of Melanoides tuberculata as a competent intermediate host suggest that eye trematodosis caused by P. lucipetus may emerge frequently in various parts of the world, especially in the tropics. Increase of the zoonotic potential of the P. lucipetus associated with this invasive snail spreading across the world is predictable and should be of interest for further research.     

Neritid and thiarid gastropods from French Polynesian streams: how reproduction (sexual, parthenogenetic) and dispersal (active, passive) affect population structure

• 1 The streams of French Polynesia contain several species of Neritidae and Thiaridae (Mollusca: Gastropoda). The neritids are dioecious and amphidromous with a freshwater adult stage and a poorly known, marine larval stage. The thiarids are parthenogenetic and viviparous, and rely on passive dispersal for colonisation of new habitats.
• 2 Populations of the neritid Clithon spinosus and the thiarids Melanoides tuberculata and Thiara granifera were analysed using mitochondrial DNA sequences from COI to compare the population structure of the snails at three different scales: between streams (N = 9), between islands (N = 4), and between age and distance of paired islands.
• 3 The amphidromous C. spinosus showed no evidence of genetic isolation at any of the scales tested (F_st values 0.02). Parsimony analyses resulted in two haplotype clusters separated by a three‐step segment, which were not linked to geographic isolation. The larval phase of C. spinosus is most likely a long‐lived planktotroph and a very effective disperser.
• 4 Two haplotypes of M. tuberculata, separated by 16 base pairs, were found. Both haplotypes were found in snails on all islands, and individuals representing both were often collected in the same habitat. One haplotype of T. granifera was found. M. tuberculata has the characteristics of the ‘general‐purpose genotype’ of clonal population structure and although it relies on passive dispersal, it has colonised nearly all freshwater habitats on the islands.

     

Effects of fine-scale environmental heterogeneity on local genetic structure in Macoma balthica from the Gulf of Gdañsk (southern Baltic Sea)

Surveys of unionid populations have been incorporated into the Swedish national environmental monitoring program. Nevertheless there is still lack of knowledge of important aspects of the biology of many unionid species. There may also be differences between species. This work compares the spatial distribution of two unionid species, Anodonta anatina and Unio tumidus, at four sites in Sweden. Samples were taken at transects along the shoreline at different water depths on the surface bed and in the sediments. Individual mussel ages were determined. Our results indicated that younger mussel individuals do not show differences in horizontal spatial distribution compared to older mussel individuals. However, they showed a preference to be burrowed in the sediment compared to older individuals that were predominantly found at the sediment surface. We also found a large difference in burrowing frequency between species with a higher frequency of burrowed adult individuals of A. anatina than U. tumidus. This result may be due to the timing of our field survey that coincided with the period of glochidia release for U. tumidus. We can conclude that a monitoring program for freshwater mussel populations needs to be carefully planned in time.     

Bioaccumulation of heavy metals and their toxicity assessment in Mystus species

This study was conducted on two native fish species namely Mystus vittatus and Mystus tengara inhabiting challenging environment of Yamuna River. The heavy metals concentrations in the river water were found to be as follows: Fe > Mn > Zn > Cu > Ni > Cr > Cd, all above the Bureau of Indian Standards (BIS) and World Health Organization (WHO) guidelines. The high metal pollution index in gill, liver, and kidney of M. vittatus was recorded compared to M. tengara. The pathology caused by the accumulation of heavy metals resulted significantly (p < 0.05) higher enzyme activities of alkaline phosphatase (ALP), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and creatinine kinase (CK) in M. tengara as compared to M. vittatus. However, albumin: globulin ratio was found to be below 0.8 in both fishes. Higher total leukocyte (TLC) (48.5 × 10³/mm³), lymphocytes (40%), respiratory burst activity (1.9), and nitric oxide synthase (NOS) activity (13.11 U/L) in M. vittatus reflect high immune response. In addition, chromosomal breakage study showed significantly (p < 0.05) low micronuclei frequency, lobed nuclei, and kidney-shaped nuclei (KSN) in M. vittatus. These results indicate that under the same challenging conditions M. vittatus have more capability of resistance and its continuous survival points towards its suitability to serve as a bioindicator than M. tengara.     

Trace Metal Concentrations in Marsh Profiles Under the Influence of an Emerging Delta (Atchafalaya River and Wax Lake Delta) Overlying a Several Thousand Year Old (Former)Mississippi River Delta Lobe

There are concerns over the increasing concentrations of trace metals being found in the environment. Deltas are essentially integrators of watershed contamination as they are the repositories of sediment transported from and through the watershed. In order to assess changes in trace metal concentrations transported by the Mississippi River–Atchafalaya River systems, vibracores were collected from three coastal freshwater marsh sites under the influence of the Atchafalaya River and emerging Wax Lake Delta (WLD). The cores extended to a depth of 4 m which included deposits of an earlier Mississippi River Delta lobe. C-14 dating showed an age at the lower depth corresponding to approximately 3500 years ago. Vertical profile distribution of metals and metalloids were measured and comparisons between older deposits and concentrations in recent deposits were made. Concentrations of As, Cr, Cu, Ni, Zn, Mo, V, Co, and Hg metals were measured in the profiles along with Fe, Mn, and Al. There was no significant increase in heavy metals or metalloids in recent years as compared to more than 3000-year-old sediment associated with an earlier Mississippi River Delta lobe. Results show that sediment diversion through the Wax Lake Outlet did not increase concentrations of these metals in surface marsh soils. The metal concentrations in the marsh profile were compared to ERL (effects range low) and ERM (effects range medium) values to allow an ecotoxicological assessment. Arsenic was below the ERM values but was greater than the ERL which indicate a potential toxicity under certain conditions. All other metal/metalloids measured were below ERL limits. This study suggests that planned Mississippi River sediment diversions designed to slow the rate of coastal land loss are not likely to lead to trace metals contamination.     

Drought-induced release of metals from peatlands in watersheds recovering from historical metal and sulphur deposition

Climate change is predicted to cause an increase in frequency and severity of droughts in the boreal ecozone, which can result in the lowering of water tables and subsequent release of acidic, metal-contaminated waters from wetlands. We believe that in areas where historical deposition of metals and sulphur was severe, these episodic pulses of metals could reach concentrations sufficiently high to severely affect aquatic communities in receiving waters and cause a delay in biological recovery. The objective of this study is to evaluate the impact of drought on the chemistry of water draining from two Sudbury peatlands with widely contrasting peat organic matter content to determine the response of stream water chemistry to drought from peatland types in the region. Stream samples were collected using ISCO? automated water collectors from June to November 2011. Following a period of drought, there was a decline in pH and a large increase in concentrations of sulphate and metal ions (Al, Co, Cu, Fe, Mn, Ni, and Zn) in water draining both peatlands, with extreme concentrations occurring over a period of about two weeks. At the site with the higher peat organic matter content there was an increase in metals that have a high affinity to bind to DOC (Al, Cu, and Fe) during the onset of drought. This study demonstrates a dramatic response to drought at two sites that differ in metal and nutrient pool sizes, hydrology, and topography, suggesting the potential for a majority of peatlands in the region to experience this response. Efforts to restore aquatic ecosystems and protect freshwater resources must take into account these processes, as disruptions to biogeochemical cycles are likely to become more prevalent in a changing climate.     

Mechanistic role of each metal ion in Streptomyces dinuclear aminopeptidase: Peptide hydrolysis and 7 × 10-fold rate enhancement of phosphodiester hydrolysis

The dinuclear aminopeptidase from Streptomyces griseus (SgAP) and its metal derivatives catalyze the hydrolysis of the phosphoester bis(p-nitrophenyl) phosphate (BNPP) and the phosphonate ester p-nitrophenyl phenylphosphonate with extraordinary rate enhancements at pH 7.0 and 25 °C [A. Ercan, H. I. Park, L.-J. Ming, Biochemistry 45, (2006) 13779-13793.], reaching 6.7 billion-fold in terms of the first-order rate constant of the di-Co(II) derivative with respect to the autohydrolytic rates. Since phosphoesters are transition state-like inhibitors in peptide hydrolysis, their hydrolysis by SgAP is quite novel. Herein, we report the investigation of this proficient alternative catalysis of SgAP and the role of each metal ion in the dinuclear site toward peptide and BNPP hydrolysis. Mn(II) selectively binds to one of the dinuclear metal sites (M1), affording MnE-SgAP with an empty (E) second site for the binding of another metal (M2), including Mn(II), Co(II), Ni(II), Zn(II), and Cd(II). Peptide hydrolysis is controlled by M2, wherein the k_cat values for the derivatives MnM2-SgAP are different yet similar between MnCo- and CoCo-SgAP and pairs of other metal derivatives. On the other hand, BNPP hydrolysis is affected by metals in both sites. Thus, the two hydrolytic catalyses must follow different mechanisms. Based on crystal structures, docking, and the results presented herein, the M1 site is close to the hydrophobic specific site and the M2 site is next to Tyr246 that is H-bonded to a coordinated nucleophilic water molecule in peptide hydrolysis; whereas a coordinated water molecule on M1 becomes available as the nucleophile in phosphodiester hydrolysis.