Effects of antifouling compounds on the growth of macroalgae Undaria pinnatifida.

Seaweeds are some of the principal primary producers of marine environments, and they are important ecological elements of coastal ecosystems. The effects of harmful chemicals on seaweeds may adversely affect coastal ecosystems, hence we aimed to develop a new phytotoxicity test using the gametophytes of a common temperate kelp species, Undaria pinnatifida (KU-1630), for the widely used antifouling chemical substances Cybutryne, Diuron, Cu2+, and Zn2+. Toxicity to gametophytes of U. pinnatifida was assessed by comparing the relative growth rate (RGR) at the logarithmic growth phase. Fragmentation method, initial algal biomass, photon irradiance, and adhesive period were investigated for developing optimal test conditions. Cybutryne exposure tests were performed with seven replicates and control, the RGR ranging from 0.17 to 0.19, while mean 7-day EC50 and no observed effect concentration (NOEC) were 5.1 µg/L and 1.8 µg/L, respectively. The 7-day EC50 for other antifoulants was 14 µg/L for Diuron, 17 µg/L for Cu2+, and 1500 µg/L for Zn2+. This test method demonstrated high sensitivity and reproducibility, and it may be added to the routine methods used for toxicity evaluation of hazardous chemicals.

An investigation of arsenic contamination in Peninsular Malaysia based on Centella asiatica and soil samples.

The first objective of this study was to provide data of arsenic (As) levels in Peninsular Malaysia based on soil samples and accumulation of As in Centella asiatica collected from 12 sampling sites in Peninsular Malaysia. The second objective was to assess the accumulation of As in transplanted C. asiatica between control and semi-polluted or polluted sites. Four sites were selected which were UPM (clean site), Balakong (semi-polluted site), Seri Kembangan (semi-polluted site) and Juru (polluted site). The As concentrations of plant and soil samples were determined by Instrumental Neutron Activation Analysis. The As levels ranged from 9.38 to 57.05 µg/g dw in soils, 0.21 to 4.33 µg/g dw in leaves, 0.18 to 1.83 µg/g dw in stems and 1.32-20.76 µg/g dw in roots. All sampling sites had As levels exceeding the CCME guideline (12 µg/g dw) except for Kelantan, P. Pauh, and Senawang with P. Klang having the highest As in soil (57.05 µg/g dw). In C. asiatica, As accumulation was highest in roots followed by leaves and stems. When the As level in soils were higher, the uptake of As in plants would also be increased. After the transplantation of plants to semi-polluted and polluted sites for 3 weeks, all concentration factors were greater than 50 % of the initial As level. The elimination factor was around 39 % when the plants were transplanted back to the clean sites for 3 weeks. The findings of the present study indicated that the leaves, stems and roots of C. asiatica are ideal biomonitors of As contamination. The present data results the most comprehensive data obtained on As levels in Malaysia.

Concentrations of heavy metals (Cu, Cd, Zn and Ni) and PAHs in Perna viridis collected from seaport and non-seaport waters in the Straits of Johore.

In this study, the ranges of pollutants found in the soft tissues of Perna viridis collected from Kg. Masai and Kg. Sg. Melayu, both located in the Straits of Johore, were 0.85-1.58 µg/g dry weight (dw) for Cd, 5.52-12.2 µg/g dw for Cu, 5.66-8.93 µg/g dw for Ni and 63.4-72.3 µg/g dw for Zn, and 36.4-244 ng/g dry weight for ∑PAHs. Significantly (p < 0.05) higher concentrations of Cd, Cu, Ni, Zn and ∑PAHs in the mussels were found in the water of a seaport site at Kg. Masai than a non-seaport site at Kg. Sg. Melayu population. The ratios of low molecular weight/high molecular weight hydrocarbons (2.94-3.42) and fluoranthene/pyrene (0.43-0.45) in mussels from both sites indicated the origin of the PAHs to be mainly petrogenic. This study has demonstrated the utility of using the soft tissues of P. viridis as a biomonitor of PAH contamination and bioavailability in the coastal waters of Peninsular Malaysia.

Assessment of Cu, Pb, and Zn contamination in sediment of north western Peninsular Malaysia by using sediment quality values and different geochemical indices.

Surface sediments were collected from the north western aquatic area (13 intertidal sites and 5 river drainages) of Peninsular Malaysia, which were suspected to have received different anthropogenic sources. These sites included town areas, ports, fishing village, industrial areas, highway sides, jetties and some relatively unpolluted sites. The present study revealed that 4.79-32.91 µg/g dry weight for Cu, 15.85-61.56 µg/g dry weight for Pb, and 33.6-317.4 µg/g dry weight for Zn based on 13 intertidal surface sediments while those based on 5 river drainage surface sediments were 10.24-119.6 µg/g dry weight for Cu, 26.7-125.7 µg/g dry weight for Pb and 88.7-484.1 µg/g dry weight for Zn. In general, the metal levels in the drainage sediments are higher than in the intertidal sediments, suggesting dilution factor in the intertidal sediment and direct effluent from point sources in the drainage sediment. In particular, the total concentrations of Cu, Pb, and Zn for the sampling site at Kuala Kurau Town exceeded the Effect Range Median values for Cu, Pb, and Zn for assessments of sediment quality values for freshwater sediment as proposed by MacDonald et al. (Arch Environ Contam Toxicol 39:20-31, 2000), thus adverse biological effects would be observed above this level. Assessment using enrichment factor (using Fe as a normalizer) and geoaccumulation index showed that the three metals at Kuala Kurau Town and Juru Industry drainage were evidenced as having more enrichment and mostly due to non-natural sources. However, caution should be exercised that the interpretation can only become valid when the ratios, indices, and sediment quality values are combined. This is due to the fact that not all the established indices are applicable and, to a certain extent, some of them should be further revised and improved to suit a different metal for Malaysian sediment. Undoubtedly, sites near drainages at Kuala Kurau Town and Juru River Basin need greater attention to mitigate the heavy metal pollution in the future.

Allozyme polymorphisms in horseshoe crabs, Carcinoscorpius rotundicauda, collected from polluted and unpolluted intertidal areas in Peninsular Malaysia.

It has been widely reported that allozyme frequency variation is a potential indicator of heavy metal-induced impacts in aquatic populations. In the present study, wild populations of horseshoe crab (Carcinoscorpius rotundicauda) were collected from contaminated and uncontaminated sites of Peninsular Malaysia. By adopting horizontal starch gel electrophoresis, seven enzyme systems were used to study allozyme polymorphisms. Nine polymorphic loci were observed in C. rotundicauda. The relationships of allozyme variations with the concentrations of Cd, Cu, Ni, and Zn in sediments and in muscle tissues of horseshoe crabs were determined. Based on genetic distance, the lower mean value of Nei's D (0.017) indicated that both of the contaminated populations of Kg. Pasir Puteh and Kuala Juru were very closely related when compared to the relatively uncontaminated Pantai Lido population. Higher heterozygosities were shown by the contaminated populations when compared to the uncontaminated population. Different allelic frequencies could be observed for the aldolase (ALD; E.C. 2.7.5.1) locus between the contaminated and uncontaminated populations of C. rotundicauda. The dendrogram of genetic relationships of the three populations of C. rotundicauda showed the same clustering pattern as the dendrograms are based on heavy metals in the sediments and in the horseshoe crabs' abdominal muscles. From the F statistics, the present study showed that the three populations of horseshoe crabs were considered to have undergone moderate genetic differentiation with a mean F (ST) value of 0.092 .The current results suggest that allozyme polymorphism in horseshoe crabs is a potential biomonitoring tool for metal contamination, although further validation is required.

Genetic characterization of Perna viridis L. in peninsular Malaysia using microsatellite markers.

A total of 19 polymorphic microsatellite loci were used to analyse levels of genetic variation for 10 populations of Perna viridis L. collected from all over peninsular Malaysia. The populations involved in this study included Pulau Aman in Penang, Tanjung Rhu in Kedah, Bagan Tiang in Perak, Pulau Ketam in Selangor, Muar, Parit Jawa, Pantai Lido and Kampung Pasir Puteh in Johore, and Kuala Pontian and Nenasi in Pahang state. The number of alleles per locus ranged from two to seven, with an average of 3.1. Heterozygote deficiencies were observed across all the 10 populations. Characterization of the populations revealed that local populations of P. viridis in peninsular Malaysia were genetically similar enough to be used as a biomonitoring agent for heavy metal contamination in the Straits of Malacca. Cluster analysis grouped the P. viridis populations according to their geographical distributions with the exception of Parit Jawa. The analysis also revealed that P. viridis from the northern parts of peninsular Malaysia were found to be the most distant populations among the populations of mussels investigated and P. viridis from the eastern part of peninsular Malaysia were closer to the central and southern populations than to the northern populations.

Telescopium telescopium as potential biomonitors of Cu, Zn, and Pb for the tropical intertidal area.

The distributions of Cu, Zn, and Pb concentrations in the selected soft tissues (foot, cephalic tentacle, mantle, muscle, gill, digestive caecum, and remaining soft tissues) and shells of the mud-flat snail Telescopium telescopium were determined in snails from eight geographical sites in the south-western intertidal area of Peninsular Malaysia. Generally, the digestive caecum compared with other selected soft tissues, accumulated higher concentration of Zn (214.35+/-14.56 microg/g dry weight), indicating that the digestive caecum has higher affinity for the essential Zn to bind to metallothionein. The shell demonstrated higher concentrations of Pb (41.23+/-1.20 microg/g dry weight) when compared to the selected soft tissues except gill from Kuala Sg. Ayam (95.76+/-5.32 microg/g dry weight). The use of different soft tissues also can solve the problem of defecation to reduce error in interpreting the bioavailability of heavy metals in the intertidal area.

Eleven novel polymorphic microsatellite DNA markers from the green-lipped mussel Perna viridis.

We report on the characterization of 11 polymorphic microsatellite loci in P. viridis, the first set of such markers developed and characterized for this species. The number of alleles per locus ranged from 2 to 7, whereas the observed heterozygosity ranged from 0.0447 to 0.4837. These markers should prove useful as powerful genetic markers for this species.

Patterns of rapd markers and heavy metal concentrations in Perna viridis (L.), collected from metal-contaminated and uncontaminated coastal waters: are they correlated with each other?

Genetic variation due to heavy metal contamination has always been an interesting topic of study. Because of the numerous contaminants being found in coastal and intertidal waters, there is always much discussion and argument as to which contaminant(s) caused the variations in the genetic structures of biomonitors. This study used a Single Primer Amplification Reaction (SPAR) technique namely Random Amplified Polymorphic DNA (RAPD) to determine the genetic diversity of the populations of the green-lipped mussel Perna viridis collected from a metal-contaminated site at Kg. Pasir Puteh and those from four relatively' uncontaminated sites (reference sites). Heavy metal levels (Cd, Cu, Pb and Zn) were also measured in the soft tissues and byssus of the mussels from all the sites. Cluster analyses employing UPGMA done based on the RAPD makers grouped the populations into two major clusters; the Bagan Tiang, Pantai Lido, Pontian and Kg. Pasir Puteh populations were in one cluster, while the Sg. Belungkor population clustered by itself. This indicated that the genetic diversity based on bands resulting from the use of all four RAPD primers on P. viridis did not indicate its potential use as a biomarker of heavy metal pollution in coastal waters. However, based on a correlation analysis between a particular metal and a band resulting from a specific RAPD primer revealed some significant (P < 0.01) correlations between the primers and the heavy metal concentrations in the byssus and soft tissues. Thus, the correlation between a particular metal and the bands resulting from the use of a specific RAPD primer on P. viridis could be used as biomonitoring tool of heavy metal pollution.

Morphological and allozyme studies of small terrestrial snails (Opeas sp., Subulina sp. and Huttonella bicolor) collected from Peninsular Malaysia.

Shell morphological characters and allozyme electrophoresis were used to study the relationships among six geographical populations of land snails collected from Peninsular Malaysia. Allozyme electrophoresis was used to study the genetic variations to complement the morphological features studied that included shell lengths, numbers of whorls and shell colour. Ten loci coding for six enzymes (MDH, LAP, ALP, PGM, G6PDH and EST) could be reliably scored in samples from the six populations studied. The dendrogram showed two major clusters with one cluster comprising Subulinidae populations from Perak, Selangor, Johor, Terengganu and Pahang while the other cluster included only the Streptaxidae Huttonella bicolor (red) population. The Subulinidae populations were grouped into two subclusters: one subcluster included the Subulina sp. populations from Perak, Selangor an Johor while the other subcluster included the Opeas sp. populations from Terengganu and Pahang. Morphological features can identify the different families and therefore they can complement the allozyme genetic studies on the land snail populations. Like other reports in the literature, our results also underline the importance of a genetic approach in conjunction with a morphological approach, for discriminating land snail species. The present results suggest that small land snails, which were similar in colour but different in sizes, were not of the same family/genus.

Crystalline style and tissue redistribution in Perna viridis as indicators of Cu and Pb bioavailabilities and contamination in coastal waters.

The concentrations of Cu, Pb, and Zn in the crystalline style (CS) and in the remaining soft tissues (ST) of the green-lipped mussel Perna viridis from 10 geographical sites along the coastal waters off peninsular Malaysia were determined. The CS, compared with the remaining ST, accumulated higher levels of Cu in both contaminated and uncontaminated samples, indicating that the style has a higher affinity for the essential Cu to bind with metallothioneins. The similar pattern of Cu accumulation in the different ST of mussels collected from clean and Cu-contaminated sites indicated that the detoxification capacity of the metallothioneins had not been overloaded. For Pb, higher levels of the metal in the CS than in the remaining ST were found only in mussels collected from a contaminated site at Kg. Pasir Puteh. This indicated a tissue redistribution of Pb due to its binding to metallothioneins for Pb detoxification and the potential of the CS as an indicator organ of Pb bioavailability and contamination. For Zn, the above two phenomena were not found since no obvious patterns were observed (lower levels of Zn in the CS than in the remaining ST) in contaminated and uncontaminated samples due to the mechanism of partial regulation. Generally, all the different STs studied (foot, mantle, gonad, CS, gill, muscle, and byssus) are good biomonitoring tissues for Cu and Pb bioavailabilities and contamination. Among these organs, the CS was found to be the best organ for biomonitoring Cu. The present data also suggest the use of the tissue redistribution of Pb in P. viridis as an indicator of Pb bioavailability and contamination in coastal waters.

Anthropogenic impacts on the distribution and biodiversity of benthic macroinvertebrates and water quality of the Langat River, Peninsular Malaysia.

A study of the impacts of anthropogenic activities on the distribution and biodiversity of benthic macroinvertebrates and water quality of the Langat River (Peninsular Malaysia) was conducted. Four pristine stations from the upstream and 4 stations at the downstream receiving anthropogenic impacts were selected along the river. For 4 consecutive months (March-June 1999), based on the Malaysian DOE (Malaysia Environmental Quality Report 2000, Department of Environment, Ministry of Science, Technology and Environment Malaysia. Maskha Sdn. Bhd. Kuala Lumpur, 86pp; Malaysia Environmental Quality Report 2001, Department of Environment, Ministry of Science, Technology and the Environment Malaysia) water quality index classes, the upstream stations recorded significantly (P<0.05) higher Biological Monitoring Working Party scores and better water quality indices than those of the downstream. The total number of macrobenthic taxa and their overall richness indices and diversity indices were significantly (P<0.05) higher at the upstream stations (54 taxa) than at the downstream stations (5 taxa). The upstream of the Langat River was dominated by Ephemeroptera and chironomid dipterans while other orders found in small quantities included Trichoptera, Diptera, Plecoptera, Odonata, Ephemeraptera, Coleoptera, and Gastropoda. On the other hand, the downstream of the river was mainly inhabited by the resistant Oligochaeta worms Limnodrilus spp. and Branchiodrilus sp. and Hirudinea in small numbers. The relationships between the physicochemical and the macrobenthic data were investigated by Pearson correlation analysis and multiple stepwise regression analysis. These statistical analyses showed that the richness and diversity indices were generally influenced by the total suspended solids and the conductivity of the river water. This study also highlighted the impacts of anthropogenic land-based activities such as urban runoff on the distribution and species diversity of macrobenthic invertebrates in the downstream of the Langat River. The data obtained in this study supported the use of the bioindicator concept for Malaysian rivers. Some sensitive (Trichopteran caddisflies and Ephemeraptera) and resistant species (Oligochaeta such as Limnodrilus spp.) are identified as potential bioindicators of clean and polluted river ecosystems, respectively, for Malaysian rivers.

Allozyme polymorphisms and heavy metal levels in the green-lipped mussel Perna viridis (Linnaeus) collected from contaminated and uncontaminated sites in Malaysia.

It has been widely reported that heavy metal contamination in coastal waters can modify the allozyme profiles of marine organisms. Previous studies have recorded elevated metal concentrations in sediments and mussel tissues off Peninsular Malaysia. In the present study, horizontal starch gel electrophoresis was carried out to estimate the levels of allelic variation of the green-lipped mussel, Perna viridis, collected from one contaminated and three relatively uncontaminated sites off Peninsular Malaysia. Fourteen polymorphic loci were observed. In addition, the concentrations of cadmium, copper, lead, mercury and zinc were determined in the sediments and in the soft tissues of the mussels. Mussels from contaminated site, evidenced by high metal pollution indices (MPI) of the sediment and the mussel tissues, showed the highest percentage of polymorphic loci (78.6%), while those collected from the uncontaminated sites had lower MPI of the sediment and mussel tissue, and exhibited lower percentages of polymorphic loci (35.7-57.1%). The population from the contaminated site showed the highest excess of heterozygosity (0.289) when compared to that of the populations from the three uncontaminated sites (0.108-0.149). Allozyme frequencies at the phosphoglucomutase (PGM; E.C. 2.7.5.1) locus also differed between the contaminated and uncontaminated populations. Previous studies have shown that exposure to heavy metals can select or counter-select for particular alleles at this locus. The present results suggest that allozyme polymorphism in P. viridis is a potential biomonitoring tool for heavy metal contamination but further validation is required.

The correlation between cytopenia and esophageal varices in patients with liver cirrhosis.

This retrospective study analysed the case records of 200 patients in the Department of Gastroenterology, Singapore General Hospital from February 2000 to January 2001 who had liver cirrhosis and underwent gastroscopy for the detection of varices. The aim of this study was to determine any relationship between leucopenia, thrombocytopenia and the occurrence of esophageal varices in a cirrhotic population. Our results showed that the diagnostic yield of varices grade 2 and 3 was 6.3% if platelet count was > 150,000/mm3, 25% if platelet count was 100,000 to 150,000/mm3, 38.9% if platelet count was 50,000-99,000/mm3 and 100% if platelet count was <50,000/mm3. Similarly, the diagnostic yield of varices grade 2 and 3 was 19.4% if total white count was > 4,000/mm3, 66.7% if total white count was 3,000- 4,000/mm3 and 94.8% if total white count was < 3,000/mm5. We conclude that thrombocytopenia and leucopenia can be used to stratify risk for occurrence of esophageal varices in cirrhotic patients and gastroscopy will have a high yield for varices when platelet count is < 150,000/mm3 or total white is < 4,000/mm.

Toxicities and tolerances of Cd, Cu, Pb and Zn in a primary producer (Isochrysis galbana) and in a primary consumer (Perna viridis).

Studies on toxicities and tolerances of cadmium (Cd), copper (Cu), lead (Pb) and zinc (Zn) in the brown alga Isochrysis galbana and in the green-lipped mussel Perna viridis were conducted by short-term bioassays using endpoints growth production and mortality, respectively. The 5-day EC(50) and 24-h LC(50) of these heavy metals were determined in the brown alga and mussel, respectively. The EC(50) values calculated for the alga were 0.74 mg/l for Cd, 0.91 mg/l for Cu, 1.40 mg/l for Pb and 0.60 mg/l for Zn. The LC(50) values for the mussels were 1.53 mg/l for Cd, 0.25 mg/l for Cu, 4.12 mg/l for Pb and 3.20 mg/l for Zn. These LC(50) values were within the concentration ranges as reported by other authors who used P. viridis as the test organism. Based on these EC(50) and LC(50) values, the alga was most sensitive to Zn, followed by Cd, Cu and Pb while the mussel was most sensitive to Cu, followed by Cd, Zn and Pb. Differences in the trophic levels, metal handling strategies, biology and ecology of the primary producer (brown alga) and the primary consumer (mussel) are believed to be the plausible causes for the different toxicities and tolerances of the metals studied.

A case of penicillamine-induced dermopathy.

INTRODUCTION: We describe an interesting patient with penicillamine-induced dermopathy. CLINICAL PICTURE: A 49-year-old woman presented with a 1-year history of recurrent haemorrhagic blisters, milia and purpura over both her elbows, while on long-term penicillamine therapy (1.5 g daily) for Wilson's disease. Histologically, dermal elastin fibres were markedly reduced in the affected areas, consistent with penicillamine-induced elastolysis. TREATMENT AND OUTCOME: The patient's lesions improved significantly after reduction of her penicillamine dose to 500 mg daily. CONCLUSIONS: The cutaneous side effects of long-term penicillamine therapy are important to recognise as they may be associated with significant morbidity and may be markers of more ominous underlying systemic elastic fibre damage.

Can the byssus of green-lipped mussel Perna viridis (Linnaeus) from the west coast of Peninsular Malaysia be a biomonitoring organ for Cd, Pb and Zn? Field and laboratory studies.

Concentrations of cadmium (Cd), lead (Pb) and zinc (Zn) in total soft tissues (ST) and byssus (BYS) of the green-lipped mussel Perna viridis from 11 different geographical locations off the west coast of Peninsular Malaysia were determined. The metal concentrations distributed between the BYS and ST were compared. The results of this study indicated that higher levels of Cd (1.31 microg/g), Pb (38.49 microg/g) and Zn (206.52 microg/g) were accumulated in the BYS than in the total ST (Cd: 0.29 microg/g; Pb: 8.27 microg/g; Zn: 102.6 microg/g). Semi-static and short period controlled laboratory experiments were also conducted for the accumulation and depuration of Cd, Pb and Zn in the total ST and BYS of P. viridis. The ratios (BYS/ST) for Pb and Cd from the laboratory experiments showed that the total ST accumulated more metals than the BYS. Therefore, these laboratory results disagreed with those found for the field samples. However, the laboratory results for the Zn ratio (BYS/ST) agreed with those of the field samples. It was evident that when compared to the ST, the BYS was a more sensitive biomonitoring organ for Zn while it could be a complementary organ for Cd and Pb in the total ST. Since total ST of P. viridis had been reported to have regulative mechanism for Zn, its BYS can be used as a biomonitoring organ for the identification of coastal areas exposed to Zn pollution.