Digestive proteases from marine organisms have been poorly applied to biomedicine. Exceptions are trypsin and other digestive proteases from a few cold-adapted or temperate fish and crustacean species. These enzymes are more efficient than enzymes from microorganism and higher vertebrates that have been used traditionally. However, the biomedical potential of digestive proteases from warm environment species has received less research attention. This review aims to provide an overview of this unrealised biomedical potential, using the debridement application as a paradigm. Debridement is intended to remove nonviable, necrotic and contaminated tissue, as well as fibrin clots, and is a key step in wound treatment. We discuss the physiological role of enzymes in wound healing, the use of exogenous enzymes in debridement, and the limitations of cold-adapted enzymes such as their poor thermal stability. We show that digestive proteases from tropical crustaceans may have advantages over their cold-adapted counterparts for this and similar uses. Differences in thermal stability, auto-proteolytic stability, and susceptibility to proteinase inhibitors are discussed. Furthermore, it is proposed that the feeding behaviour of the source organism may direct the evaluation of enzymes for particular applications, as digestive proteases have evolved to fill a wide variety of feeding habitats, natural substrates, and environmental conditions. We encourage more research on the biomedical application of digestive enzymes from tropical marine crustaceans.
Vibrio cholerae still represents a significant threat to human health worldwide despite the advances in hygiene, consumer knowledge, food treatment and food processing. In Malaysia, statistics in year 2009 have shown that among the food and water borne diseases, food poisoning has the highest incidence rate of 36.17 per 100,000 populations and with a mortality rate of 0.01 per 100,000 populations. In this study, 22 seafood samples comprising of fish, squid, crustacean and mollusks purchased from wet market and supermarket were analyzed. The Most Probable Number (MPN) and real time PCR was used to enumerate the Vibrio cholerae in seafood sample. The results showed that MPN-real time PCR of the samples from wet market had a maximum of >1100 MPN/g compare to 93 MPN/g enumerated from the MPN plate. The MPN-real time PCR in the samples from supermarket indicated 290 MPN/g as compared to 240 MPN/g enumerated from the MPN plate. The standard curves showed that there was a good linear correlation between the Ct values. The minimum level of detection of Vibrio cholerae standard DNA at targeted gene was 3 x 10-5 ng/μl.
Aquaculture production of the Pacific white shrimp is the largest in the world for crustacean species. Crucial to the sustainable global production of this important seafood species is a fundamental understanding of the shrimp gut microbiota and its relationship to the microbial ecology of shrimp pond. This is especially true, given the recently recognized role of beneficial microbes in promoting shrimp nutrient intake and in conferring resistance against pathogens. Unfortunately, aquaculture-related microbiome studies are scarce in Southeast Asia countries despite the severe impact of early mortality syndrome outbreaks on shrimp production in the region. In this study, we employed the 16S rRNA amplicon (V3-V4 region) sequencing and amplicon sequence variants (ASV) method to investigate the microbial diversity of shrimp guts and pond water samples collected from aquaculture farms located in Malaysia and Vietnam. Substantial differences in the pond microbiota were observed between countries with the presence and absence of several taxa extending to the family level. Microbial diversity of the shrimp gut was found to be generally lower than that of the pond environments with a few ubiquitous genera representing a majority of the shrimp gut microbial diversity such as Vibrio and Photobacterium, indicating host-specific selection of microbial species. Given the high sequence conservation of the 16S rRNA gene, we assessed its veracity at distinguishing Vibrio species based on nucleotide alignment against type strain reference sequences and demonstrated the utility of ASV approach in uncovering a wider diversity of Vibrio species compared to the conventional OTU clustering approach.
Deterioration of water quality mainly due to high total ammonia nitrogen (TAN) and nitrite will affect the productivity of shrimp culture. In this study, three indigenous strains assigned as VCM5, VCM8 and VCM12 were evaluated for their ability to degrade TAN and nitrite. These strains were inoculated into shrimp aquaculture wastewater to enhance the degradation of TAN and nitrite. All the strains reduced TAN and nitrite level from the shrimp aquaculture wastewater significantly (p<0.05). Strain VCM5 (GenBank accession number KJ700465) and VCM8 (GenBank accession number KJ700464) showed 99.71% sequence similarity with the 16S rRNA gene type species Bacillus vietnamensis 15-1T (ABO99708) and strain VCM12 (GenBank accession number KJ700463) showed 99.05% sequence similarity with 16S rRNA gene sequence type species Gordonia bronchialis DSM43247T (CP001802).
The concentration of meHg in freshwater fish and seafood was investigated, as well as the consumption patterns of fish and seafood by different demographic groups (age, ethnicity, gender). A potential alarm for human health hazards was also assessed, and the results were compared to the provisional tolerable weekly intakes (PTWIs) and the hazard quotient parameter (HQ). The results showed that meHg levels of 67 species ranged from 0.013 to 0.252 mg/kg of wet weight (WW) with significant differences between different fish and seafood groups (χ2KW = 49.09; p pelagic fish (0.0686 mg/kg WW) > freshwater fish 0.045 mg/kg WW) > cephalopods (0.0405 mg/kg WW) crustaceans (0.0356 mg/kg WW). The results revealed that older population (> 40 years old) consumed significantly (p = 0.000) more fish compared to younger generations and the elderly consumed the highest amounts of fish (104.0 ± 113.0 g/day). The adolescents (10-17 years old) consumed more than double of amount for both cephalopod and crustacean compared to the older populations (p
This study presents the first ever data of extracting chitin from the Chiton shell, which was then converted to the soluble chitosan by soaking in the 45% NaOH solution. The obtained chitin and chitosan were characterized by the seven different methods. Antioxidant activity of the extracted chitosan was also evaluated using the two methods. The shell content was divided into calcium carbonate (90.5 %), protein (5.2%), and chitin (4.3 %). Due to the results of element analysis and 1H NMR, the final degree of deacetylation of chitosan was 90%. Surprisingly, a significant amount of Fe was accidentally found in the shell after demineralization, and removed from the solution through the filtering. Nonetheless, remained Fe in the extracted chitin and chitosan was 20 times higher than those previously reported from the shell of shrimps and crabs. Presence of this amount of Fe could describe why the produced chitosan was darker compared to the commercial chitosan. Antioxidant activity tests showed that the IC50 of the extracted chitosan was higher than one estimated for the commercial chitosan. Antioxidant activity of the extracted chitosan is even better than the commercial version and may be used in pharmaceutical industry as a source of antioxidant.
Next-Gen sequencing was used to recover the complete mitochondrial genome of Cherax tenuimanus. The mitogenome consists of 15,797 base pairs (68.14% A + T content) containing 13 protein-coding genes, two ribosomal subunit genes, 22 transfer RNAs, and a 779 bp non-coding AT-rich region. Mitogenomes have now been recovered for all six species of Cherax native to Western Australia.
Zooplankton samples collected before (1985-86) and after (2013-14) the establishment of Kapar power station (KPS) were examined to test the hypothesis that increased sea surface temperature (SST) and other water quality changes have altered the zooplankton community structure. Elevated SST and reduced pH were detected between before and after impact pairs, with the greatest impact at the station closest to KPS. Present PAHs and heavy metal concentrations are unlikely causal factors. Water parameter changes did not affect diversity but community structure of the zooplankton. Tolerant small crustaceans, salps and larvaceans likely benefited from elevated temperature, reduced pH and shift to a more significant microbial loop exacerbated by eutrophication, while large crustaceans were more vulnerable to such changes. It is predicted that any further rise in SST will remove more large-bodied crustacean zooplankton, the preferred food for fish larvae and other meroplankton, with grave consequences to fishery production.
A study was carried out to investigate the fractionation of Cd, Cr, Cu, Fe, Mn, Pb, and Zn in shrimp aquaculture sludge from Selangor, Malaysia, using original (unmodified) and modified four-steps BCR (European Community Bureau of Reference, now known as the Standards Measurements and Testing Program) sequential extraction scheme. Step 2 of the unmodified BCR procedure (subsequently called Method A) involves treatment with 0.1 M hydroxylammonium chloride at pH 2, whereas 0.5 M hydroxylammonium chloride at pH 1.5 was used in the modified BCR procedure (subsequently called Method B). Metal analyses were carried out by flame atomic absorption spectrometry. A pseudo-total aqua-regia digest of BCR CRM 701 has also been undertaken for quality assurance purposes. The recovery of Method A for all metals studied ranges from 96.14% to 105.26%, while the recovery for Method B ranges from 95.94% to 122.40%. Our results reveal that Method A underestimated the proportion of metals bound to the easily reducible fraction except for copper. Therefore, the potential mobility of these elements is higher than others. Thus, to use this sludge as a fertilizer we have to first find a remediation for reduction of heavy metal contamination.
The significance of food specific serum IgG4 antibody in food allergy is unclear and this led us to investigate the relevance of specific IgG4, along with IgG and IgE antibodies to two common food allergens in Malaysia. Enzyme-linked immunosorbent assay (ELISA) was used to measure the serum antibodies in 143 allergic rhinitis patients' sera, of which 47 were from patients with clinical indication of shrimp allergy, 46 with clinical indication of crab allergy and 50 without indication to either allergy. Clinical indication of allergy was based on answers to a questionnaire or results of the skin prick test. We found that the elevation of specific IgE or IgG4 is associated with shrimp and crab allergies but elevation of specific IgG is not associated with either allergy. However, the clinical utility of elevated specific IgG and IgG4 levels is pending further investigation.
Study site: Allergic rhinitis clinic, University Malaya Medical Centre (UMMC), Kuala Lumpur, Malaysia
Crude extracts (methanol) of various parts, viz. the leaves, fruits, roots, stem and trunk bark, of Garcinia atroviridis were screened for antimicrobial, cytotoxic, brine shrimp toxic, antitumour-promoting and antioxidant activities. The crude extracts exhibited predominantly antibacterial activity with the root extract showing the strongest inhibition against the test bacteria at a minimum inhibitory dose (MID) of 15.6 microg/disc. Although all the extracts failed to inhibit the growth of most of the test fungi, significant antifungal activity against Cladosporium herbarum was exhibited by most notably the fruit (MID: 100 microg), and the leaf (MID: 400 microg) extracts. None of the extracts were significantly cytotoxic, and lethal towards brine shrimps. The root, leaf, trunk and stem bark extracts (except for the fruits) showed strong antioxidant activity exceeding that of the standard antioxidant, alpha-tocopherol. Antitumour-promoting activity (>95% inhibition) was shown by the fruit, leaf, stem and trunk bark extracts.
Majority of the studies on the effect of chitin and chitosan on growth and carcass characteristics of broiler chickens has concentrated more on shrimp chitin and shrimp chitosan, and often with contradictory results. Therefore, the objective of this present study is to evaluate and compare the effect of dietary chitin and chitosan from cricket and shrimp on growth performance, carcass, and organ characteristics of broiler chickens. One hundred fifty-day-old male Cobb500 broiler chicks of similar average weight were randomly allotted into one of the five dietary treatments with three replicates. Treatment 1 (T1) chicks were fed basal diet only (control), treatment 2 and 3 (T2 and T3) chicks were given basal diet with 0.5 g/kg diet of cricket chitin and cricket chitosan, respectively, while treatment 4 and 5 (T4 and T5) chicks were served basal diet with 0.5 g/kg diet of shrimp chitin and shrimp chitosan respectively. No significant variation occurred between cricket chitin and shrimp chitin, although data on growth performance were higher in cricket chitin, but growth performance varied significantly between cricket chitosan and shrimp chitosan. This study revealed that cricket chitin at 0.5 g/kg significantly improved growth performance, carcass quality, and organ characteristics of broilers more than chitosan. Birds fed basal diet alone, although gained more weight, also accumulated more fat having the poorest feed conversion ratio (FCR) and the highest mortality. However, carcass of birds fed cricket chitin was the leanest and thus economically beneficial as they consumed the least amount of feed with the best FCR.
Astaxanthin is one of the main carotenoid pigments. It has beneficial effects on the immune system of the human body due to its powerful antioxidant properties. The application of this bioactive compound can be found to be significant in the food, pharmaceutical, and cosmetics industries. The aim of this research was to investigate astaxanthin yield from six species of Malaysian shrimp carapace. Six types of shrimp species-Parapenaeopsis sculptili, Metapenaeus lysianassa, Macrobrachium rosenbergii, Metapenaeopsis hardwickii, Penaeus merguiensis, and Penaeus monodon-were used to investigate total carotenoid content and astaxanthin yield. The investigation was carried out using chemical extraction and high-pressure processing (HPP) methods at 210 MPa, for a period of 10 min with a solvent mixture of acetone and methanol (7:3, v/v). HPP was proven to have a significant impact in increasing the total carotenoid content and astaxanthin yield. The highest total carotenoid content and astaxanthin yield is shown to be contained in the Penaeus monodon species. Total carotenoid was increased from 46.95 µg/ml using chemical extraction to 68.26 µg/ml using HPP; yield of astaxanthin was increased from 29.44 µg/gdw using chemical extraction to 59.9744 µg/gdw using HPP. Therefore, comparison between the HPP and chemical extraction methods showed that HPP is more advantageous with higher astaxanthin yield, higher quality, and shorter extraction time.
The vp28 gene encoding an envelope protein (28 kDa) of white spot syndrome virus (WSSV) was amplified from WSSV-infected tiger shrimp that originated from Malaysia. Recombinant VP28 protein (r-28) was expressed in Escherichia coli and used as an antigen for preparation of monoclonal antibodies (MAbs). Three murine MAbs (6F6, 6H4 and 9C10) that were screened by r-28 antigen-based enzyme-linked immunosorbent assay (ELISA) were also able to recognize viral VP28 protein as well as r-28 on Western blot. Three non-overlapping epitopes of VP28 protein were determined using the MAbs in competitive ELISA; thus, an antigen-capture ELISA (Ac-ELISA) was developed by virtue of these MAbs. Ac-ELISA can differentiate WSSV-infected shrimp from uninfected shrimp and was further confirmed by a polymerase chain reaction (PCR) and Western blot. Approximately 400 pg of purified WSSV sample and 20 pg of r-28 could be detected by Ac-ELISA, which is comparable in sensitivity to PCR assay but more sensitive than Western blot in the detection of purified virus. Hemolymph and tissue homogenate samples collected from a shrimp farm in Malaysia during December 2000 and July 2001 were also detected by Ac-ELISA and PCR with corroborating results.
The lipid fraction of rubber (Hevea brasiliensis (kunth. Muell)) seed was extracted and analyzed for toxicological effect. The toxicological compound such as linamarin in rubber seed oil (RSO) extracted using different solvents, such as hexane (RSOh), mixture of chloroform + methanol (RSOchl+mth) and ethanol (RSOeth) were also studied. Various methods analysis such as Fourier transforms infrared spectroscopy (FTIR) and colorimetric methods were carried out to determine the present of such compounds.