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Fulltext A review of colorectal cancer research in Malaysia

Lim KG

Med J Malaysia, 2014 Aug;69 Suppl A:23-32.
PMID: 25417948 MyJurnal

105 articles related to colorectal cancer(CRC) were found in a search through a database dedicated to indexing all original data relevant to medicine published in Malaysia between the years 2000-2013. 56 articles were selected and reviewed on the basis of clinical relevance and future research implications. Research into the genetic basis for colorectal cancer included studies in germline mutations of known syndromes as well as polymorphisms that conferred individuals a higher odds ratio for developing CRC. Several studies also documented the variety of somatic mutations seen in cases of sporadic CRC in Malaysia. Studies into the knowledge and attitudes of Malaysians regarding CRC revealed poor appreciation of the common symptoms, risk factors and available measures for its early detection. This may explain the observed facts that more Malaysians present with late stage CRC than seen in developed countries. The small amount of data recorded concerning the outcome of treatment also suggests overall survival of Malaysian CRC patients for comparable stage of CRC is lower than achieved in developed countries.

Matched MeSH terms: Polymorphism, Genetic
Mutations in mitochondrial NADH dehydrogenase subunit 1 (mtND1) gene in colorectal carcinoma

Yusnita Y, Norsiah MD, Rahman AJ

Malays J Pathol, 2010 Dec;32(2):103-10.
PMID: 21329181 MyJurnal

Mitochondrial Subunit ND1 (mtND1) gene is involved in the first step of the electron transport chain of oxidative phosphorylation (OXPHOS). Alteration of the electron transport components by mutations in mtDNA may compromise the normal electron flow. This could lead to an increase of bifurcation and generation of superoxidase radicals and increase oxidative stress in various types of cancer cells. Genomic DNA was extracted from thirty matched primary colorectal tumour tissues and matching non-tumour tissues. Blood samples were obtained from twenty-five normal people. The mtNDI coding region was amplified by step-down PCR. The purified products were then subjected to direct sequencing and subsequently, the DNA sequences obtained were compared with the revised Cambridge Reference Sequence (rCRS) and MITOMAP. From the analysis, the mtND1 gene showed 11 (45.8%) different mutations and also 13 (54.2%) polymorphisms. The heteroplasmic mutation A4123A/G (I273I/V) might have a pathogenic significance as it fulfills various pathogenic criteria. Three mutations, T3394C (Y30H), A3434G (Y43C) and C3497T (A64V) which occur in a highly conserved region were likely to alter the structure and function of the ND1 protein. We suggest that these mutations, and in combination with the polymorphic variance in mtDNA, may cause slight changes that generate subtly higher levels of toxic reactive oxygen species (ROS).

Matched MeSH terms: Polymorphism, Genetic
Fulltext Variants of organic anion transporter polypeptide 2 gene are not risk factors associated with severe neonatal hyperbilirubinemia

Wong FL, Boo NY, Ainoon O, Wang MK

Malays J Pathol, 2009 Dec;31(2):99-104.
PMID: 20514852 MyJurnal

This study aimed to determine the prevalence of four variants of organic anion transporter polypeptide 2 (OATP2) gene, and their association with severe hyperbilirubinemia.

Matched MeSH terms: Polymorphism, Genetic*
Fulltext Analysis of PCR-RAPD DNA and antibiotic susceptibility profiles of antrum and corpus isolates of Helicobacter pylori from Malaysian patients

Norazah A, Rasinah WZ, Zaili Z, Aminuddin A, Ramelah M

Malays J Pathol, 2009 Jun;31(1):29-34.
PMID: 19694311 MyJurnal

This study was conducted to determine whether there was any genetic heterogeneity among Helicobacter pylori strains isolated from the antrum and corpus of the same individual in a Malaysian population and to determine the presence of heterogeneous susceptibility of the isolates by comparing PCR-RAPD and antibiotic profiles. Forty-four H. pylori isolates cultured from the antrum and corpus of 22 patients were analyzed. Antibiotic susceptibility testing was carried out by minimum inhibitory concentration determination, using E-Test method strips. PCR-RAPD was carried out on all the strains and the profiles generated were analysed for cluster analysis. Twenty-nine different PCR-RAPD profiles were observed in the 44 isolates. Fifteen pairs of the isolates from the same patients had the same PCR-RAPD patterns while in 7 pairs, the profiles were different. The strains were clustered into 2 separate clusters at a low coefficient of similarity, where most of the strains were in cluster 1. The degree of similarity was very low among most of the isolates. Most of the patients (16 of 22) were infected with strains that have the same antibiotic susceptibility profiles. Out of these, only 10 pairs shared the same PCR-RAPD and antibiotic profiles. Five pairs of isolates with similar PCR-RAPD profiles differed in their antibiotic profiles due to metronidazole resistance in one of the sites. A large degree of genetic heterogeneity was observed among H. pylori strains circulating among Malaysian patients. An individual patient can be infected with multiple strains and the strains can be antibiotic resistant.

Matched MeSH terms: Polymorphism, Genetic
Fulltext Molecular characterisation and frequency of Ggamma Xmn I polymorphism in Chinese and Malay beta-thalassaemia patients in Malaysia

Wong YC, George E, Tan KL, Yap SF, Chan LL, Tan JA

Malays J Pathol, 2006 Jun;28(1):17-21.
PMID: 17694955

The molecular basis of variable phenotypes in P-thalassaemia patients with identical genotypes has been associated with co-inheritance of alpha-thalassaemia and persistence of HbF production in adult life. The Xmn I restriction site at -158 position of the Ggamma-gene is associated with increased expression of the Ggamma-globin gene and higher production of HbF This study aims to determine the frequency of the digammaferent genotypes of the Ggamma Xmn I polymorphism in P-thalassaemia patients in two ethnic groups in Malaysia. Molecular characterisation and frequency of the Ggamma Xmn I polymorphism were studied in fifty-eight Chinese and forty-nine beta-thalassaemia Malay patients by Xmn I digestion after DNA amplification of a 650 bp sequence. The in-house developed technique did not require further purification or concentration of amplified DNA before restriction enzyme digestion. The cheaper Seakem LE agarose was used instead of Nusieve agarose and distinct well separated bands were observed. Genotyping showed that the most frequent genotype observed in the Malaysian Chinese was homozygosity for the absence of the Xmn I site (-/-) (89.7%). In the Malays, heterozygosity of the Xmn I site (+/-) was most common (63.3%). Homozygosity for the Xmn I site (+/+) was absent in the Chinese, but was confirmed in 8.2% of the Malays. The ratio of the (+) allele (presence of the Xmn I site) to the (-) allele (absence of the Xmn I site)) was higher in the Malays (0.66) compared to the Chinese (0.05). The (+/-) and (+/+) genotypes are more commonly observed in the Malays than the Chinese in Malaysia.

Matched MeSH terms: Polymorphism, Genetic*
Fulltext A Nested Allele-Specific Multiplex Polymerase Chain Reaction Method for the Detection of DRD2 Polymorphisms

Zahari Z, Salleh MR, Zahri Johari MK, Musa N, Ismail R

Malays J Med Sci, 2011 Oct;18(4):44-57.
PMID: 22589672 MyJurnal

The dopamine D2 receptor gene (DRD2) plays a role in many diseases such as schizophrenia, Parkinson's disease, and addictive behaviour. Methods currently available for the detection of DRD2 polymorphisms are costly and cannot detect all 8 polymorphisms of our research interest simultaneously (Val96Ala, Leu141Leu, Val154Ile, Pro310Ser, Ser311Cys, TaqI A, A-241G, and -141C Ins/Del). Therefore, we developed a nested multiplex polymerase chain reaction (PCR) for simultaneous detection of these polymorphisms.

Matched MeSH terms: Polymorphism, Genetic
Dietary Carbohydrate Promotes Cell Survival in Cancer Via the Up-Regulation of Fat Mass and Obesity-Associated Gene Expression Level

Doaei S, Gholamalizadeh M, Akbari ME, Akbari S, Feradova H, Rahimzadeh G, et al.

Malays J Med Sci, 2019 Mar;26(2):8-17.
PMID: 31447604 DOI: 10.21315/mjms2019.26.2.2

Cancer cells are mainly dependent on glycolysis for their growth and survival. Dietary carbohydrates play a critical role in the growth and proliferation of cancer and a low-carbohydrate diet may help slow down the growth of tumours. However, the exact mechanisms behind this effect are unclear. This review study aimed to investigate the effect of fat mass and obesity-associated (FTO) gene in the association between dietary carbohydrates and cancer. This study was carried out using keywords such as polymorphism and/or cancer and/or dietary carbohydrate and/or FTO gene. PubMed and Science Direct databases were used to collect all related articles published from 1990 to 2018. Recent studies showed that the level of FTO gene expression in cancer cells is dramatically increased and may play a role in the growth of these cells through the regulation of the cellular metabolic pathways, including the phosphoinositide 3-kinases/protein kinaseB (PI3K/AKT) signaling pathway. Dietary carbohydrate may influence the FTO gene expression by eliminating the inhibitory effect of adenosine monophosphate-activated protein kinase (AMPK) on the FTO gene expression. This review summarised what has been recently discovered about the effects of dietary carbohydrate on cancer cells and tried to determine the mediating role of the FTO gene in these effects.

Matched MeSH terms: Polymorphism, Genetic
Fulltext Influence of CYP2D6 polymorphisms on symptomatology and side-effects of patients with schizophrenia in Malaysia

Zahari Z, Salleh MR, Teh LK, Ismail R

Malays J Med Sci, 2009 Jul;16(3):12-20.
PMID: 22589660 MyJurnal

Our objective was to investigate the association of CYP2D6 polymorphisms with symptoms and side-effects of patients with schizophrenia.

Matched MeSH terms: Polymorphism, Genetic
Fulltext Low Levels of Polymorphisms and Negative Selection in Plasmodum knowlesi Merozoite Surface Protein 8 in Malaysian Isolates

Atique Ahmed M, Kang HJ, Quan FS

Korean J Parasitol, 2019 Aug;57(4):445-450.
PMID: 31533414 DOI: 10.3347/kjp.2019.57.4.445

Human infections due to the monkey malaria parasite Plasmodium knowlesi is increasingly being reported from most Southeast Asian countries specifically Malaysia. The parasite causes severe and fatal malaria thus there is a need for urgent measures for its control. In this study, the level of polymorphisms, haplotypes and natural selection of full-length pkmsp8 in 37 clinical samples from Malaysian Borneo along with 6 lab-adapted strains were investigated. Low levels of polymorphism were observed across the full-length gene, the double epidermal growth factor (EGF) domains were mostly conserved, and non-synonymous substitutions were absent. Evidence of strong negative selection pressure in the non-EGF regions were found indicating functional constrains acting at different domains. Phylogenetic haplotype network analysis identified shared haplotypes and indicated geographical clustering of samples originating from Peninsular Malaysia and Malaysian Borneo. This is the first study to genetically characterize the full-length msp8 gene from clinical isolates of P. knowlesi from Malaysia; however, further functional characterization would be useful for future rational vaccine design.

Matched MeSH terms: Polymorphism, Genetic
The chromosomes of four species of the nasuta complex of Drosophila. I. Chromosome maps and inversion polymorphism

Lambert DM

J Hered, 1976 3 1;67(2):92-8.
PMID: 5483

The salivary chromosomes of four species of the nasuta complex of Drosophila, D. sulfurigaster albostrigata, D, kohkoa, D. albomicans, and D. kepulauana were studied and chromosome maps of each species are presented; the maps of the latter three species are based on the map of D. sulfurigaster albostrigata. Three of the species D. sulfurigaster albostrigata, D. albomicans, and D. kohkoa were shown to be highly polymorphic for chromosomal inversions while the available evidence indicated that D. kepulauana is much less polymorphic. These facts are correlated with the geographic distribution of the species. Transitional homoselection has not been complete in the evolution of three of the species since D. sulfurigaster albostrigata, D. kohkoa, and D. albomicans have a number of naturally occurring polymorphisms in common.

Matched MeSH terms: Polymorphism, Genetic*
Genome-Wide Association for HbA1c in Malay Identified Deletion on SLC4A1 that Influences HbA1c Independent of Glycemia

Chai JF, Kao SL, Wang C, Lim VJ, Khor IW, Dou J, et al.

J Clin Endocrinol Metab, 2020 Dec 01;105(12).
PMID: 32936915 DOI: 10.1210/clinem/dgaa658

CONTEXT: Glycated hemoglobin A1c (HbA1c) level is used to screen and diagnose diabetes. Genetic determinants of HbA1c can vary across populations and many of the genetic variants influencing HbA1c level were specific to populations.
OBJECTIVE: To discover genetic variants associated with HbA1c level in nondiabetic Malay individuals.
DESIGN AND PARTICIPANTS: We conducted a genome-wide association study (GWAS) analysis for HbA1c using 2 Malay studies, the Singapore Malay Eye Study (SiMES, N = 1721 on GWAS array) and the Living Biobank study (N = 983 on GWAS array and whole-exome sequenced). We built a Malay-specific reference panel to impute ethnic-specific variants and validate the associations with HbA1c at ethnic-specific variants.
RESULTS: Meta-analysis of the 1000 Genomes imputed array data identified 4 loci at genome-wide significance (P

Matched MeSH terms: Polymorphism, Genetic
Polymorphisms in the host CYP2C19 gene and antibiotic-resistance attributes of Helicobacter pylori isolates influence the outcome of triple therapy

Ram M R, Teh X, Rajakumar T, Goh KL, Leow AHR, Poh BH, et al.

J Antimicrob Chemother, 2019 01 01;74(1):11-16.
PMID: 30403784 DOI: 10.1093/jac/dky401

Objectives: Eradication of Helicobacter pylori is influenced by susceptibility to antimicrobial agents, elevated bacterial load and degree of acid inhibition, which can be affected by genotypes of drug-metabolizing enzymes [cytochrome P450 (CYP) 2C19 polymorphism]. Theoretically, the choice and dose of proton pump inhibitor may also influence the suppression of H. pylori infection. The CYP2C19 genotype has recently been found to have an impact on peptic ulcer healing, H. pylori eradication and therapeutic efficacy of proton pump inhibitors.
Methods: Here, we investigated the impact of the CYP2C19 genotype polymorphism and the success of triple therapy (fluoroquinolones/metronidazole/clarithromycin) on antibiotic-resistant strains in eradicating H. pylori in human subjects with non-ulcer dyspepsia (NUD), in human subjects with peptic ulcer disease (PUD) and in asymptomatic human subjects (positive and negative for H. pylori infection).
Results: Based on the CYP2C19 genotypes, determined by Droplet Digital PCR (ddPCR) analysis, we found 11.2%, 62.5% and 26.3% corresponding to rapid metabolizers, intermediate metabolizers and poor metabolizers, respectively. However, we did not find any significant effect for homozygous ABCB1 or CYP2C19*2 and CYP2C19*3 alleles. We detected several participants heterozygous for both ABCB1 and CYP2C19*2, CYP2C19*3 and CYP2C19*17 loci. The participants heterozygous for both ABCB1 and CYP2C19*2 and *3 loci should be defined as intermediate and poor metabolizers according to the haplotype analysis in the NUD, PUD and asymptomatic subjects.
Conclusions: Consequently, fluoroquinolones/metronidazole/clarithromycin-based triple therapies can be used to eradicate H. pylori infection, if one does not know the CYP2C19 genotype of the patient.

Matched MeSH terms: Polymorphism, Genetic*
Genetic polymorphisms and drug interactions leading to clopidogrel resistance: why the Asian population requires special attention

Hasan MS, Basri HB, Hin LP, Stanslas J

Int J Neurosci, 2013 Mar;123(3):143-54.
PMID: 23110469 DOI: 10.3109/00207454.2012.744308

Ischemic heart disease and stroke are the two leading causes of death worldwide. Antiplatelet therapy plays the most significant role in the management of these cardiovascular and cerebrovascular occlusive events to prevent recurrent ischemic attack. Clopidogrel, an antiplatelet drug, is widely prescribed either alone or in combination with aspirin as dual antiplatelet therapy for the prevention of vascular occlusive events. The antiplatelet response to clopidogrel varies widely. Hyporesponders and nonresponders are likely to have adverse cardiovascular events during follow-up. Some drugs, such as proton pump inhibitors (omeprazole), calcium channel blockers, selective serotonin reuptake inhibitors (nefazadone), coumarin derivatives (phenprocoumon), benzodiazepines, sulfonylurea, erythromycin, and itraconazole, decrease the antiplatelet effect of clopidogrel when administered concomitantly. Decreased response to clopidogrel is common among Asians due to genetic polymorphisms associated with clopidogrel resistance, and it is nearly 70% in some of the Asian communities. It is necessary to study Asian populations, because there are a large number of Asians throughout the world due to increased migration. Current guidelines do not make genetic testing or platelet response testing mandatory prior to clopidogrel prescription. Therefore, it is important for clinicians treating Asian patients to keep in mind the interindividual variability in response to clopidogrel when prescribing the drug.

Matched MeSH terms: Polymorphism, Genetic/genetics*
Polymorphisms of UGT1A1*6, UGT1A1*27 & UGT1A1*28 in three major ethnic groups from Malaysia

Teh LK, Hashim H, Zakaria ZA, Salleh MZ

Indian J Med Res, 2012 Aug;136(2):249-59.
PMID: 22960892

Genetic polymorphisms of uridine diphosphate glucuronyltransferase 1A1 (UGT1A1) have been associated with a wide variation of responses among patients prescribed with irinotecan. Lack of this enzyme is known to be associated with a high incidence of severe toxicity. The objective of this study was to investigate the prevalence of three different variants of UGT1A1 (UGT1A1*6, UGT1A1*27 and UGT1A1*28), which are associated with reduced enzyme activity and increased irinotecan toxicity, in the three main ethnic groups in Malaysia (Malays, Chinese and Indians).

Matched MeSH terms: Polymorphism, Genetic
Fulltext Population Genetics of Identifiler System in Malaysia

Nakamura Y, Samejima M, Minaguchi K, Nambiar P

Bull. Tokyo Dent. Coll., 2016;57(4):233-239.
PMID: 28049971 DOI: 10.2209/tdcpublication.2016-1400

Short tandem repeat (STR) polymorphisms were investigated in 341 unrelated Malay individuals (218 males and 123 females) living in or around Kuala Lumpur by using a forensic analysts kit. The following STRs were targeted: D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, vWA, TPOX, D18S51, D5S818, and FGA. The purpose of this study was to elucidate population genetics in Malaysia and calculate statistical parameters for forensic and anthropological research. Data on these STRs in the target population were obtained and subjected to statistical analysis. Accordance with the Hardy-Weinberg equilibrium was proven for all the loci targeted. The combined power of discrimination was greater than 0.9999999999, indicating that this multiplex system is an excellent tool for forensic casework. The allele frequency in the data were weighed against that in four other local populations (Chinese, Iranian, Belgian, and African). The average coefficient of correlation was strongest in the order of Africa (0.092522), Belgium (0.264822), Iran (0.404363), and China (0.706661). These results are consistent with what is known about the anthropological history of and prehistoric human migration in the Malay region. We believe that these data offer a valuable anthropological resource, being applicable to the statistical evaluation of DNA evidence in human identification, as well as the determination of ethnicity in healthy populations.

Matched MeSH terms: Polymorphism, Genetic
Allelic dimorphism in the merozoite surface protein-3alpha in Korean isolates of Plasmodium vivax

Han ET, Song TE, Park JH, Shin EH, Guk SM, Kim TY, et al.

Am J Trop Med Hyg, 2004 Dec;71(6):745-9.
PMID: 15642964

To study the genetic diversity of re-emerging Plasmodium vivax in the Republic of Korea, nucleotide sequence variations at the merozoite surface protein-3alpha (PvMSP-3alpha) locus were analyzed using 24 re-emerging isolates and 4 isolates from imported cases. Compared with the well known Belem strain (Brazil), a large number of amino acid substitutions, deletions, and insertions were found at the locus of the isolates examined. The Korean isolates were divided into two allelic types; type I (15 isolates), similar to the Belem strain, and type II (9), similar to the Chess strain (New Guinea). Isolates from imported cases were classified into three types; type III (1 from Malaysia), similar to type B from western Thailand, type IV (1 each from Indonesia and India), and type V (1 from Pakistan), both being new types. Our results have shown that the MSP-3alpha locus of re-emerging Korean P. vivax is dimorphic with two allelic types coexisting in the endemic area.

Matched MeSH terms: Polymorphism, Genetic
Fulltext The risk of recurrence in breast cancer patients treated with tamoxifen: polymorphisms of CYP2D6 and ABCB1

Teh LK, Mohamed NI, Salleh MZ, Rohaizak M, Shahrun NS, Saladina JJ, et al.

AAPS J, 2012 Mar;14(1):52-9.
PMID: 22183189 DOI: 10.1208/s12248-011-9313-6

CYP2D6 plays a major role in the metabolism of tamoxifen, and polymorphism of P-glycoprotein has been associated with resistance of many drug therapies. This study investigates the clinical impact of genetic variants of CYP2D6 and ABCB1 in breast cancer patients treated with tamoxifen. Blood samples from 95 breast cancer patients treated with tamoxifen were collected and genotyped for CYP2D6 and ABCB1 variants using allele-specific PCR method. Recurrence risks were calculated using Kaplan-Meier analysis and compared using the log-rank test. Patients carrying CYP2D6*10/*10 and heterozygous null allele (IM) showed higher risks of developing recurrence and metastasis (OR 13.14; 95% CI 1.57-109.94; P = 0.004) than patients with CYP2D6*1/*1 and *1/*10 genotypes. Patients with homozygous CC genotypes of ABCB1 C3435T showed a shorter time to recurrence. Patients who were CYP2D6 IM and homozygous CC genotype of C3435T have statistically significant higher risks of recurrence (P = 0.002). Similarly, median time to recurrence in these patients was only 12 months (95% CI = 0.79-23.2) compared to those without this combination which was 48 months (95% CI = 14.7-81.2). Patients with CYP2D6 IM and homozygous CC genotype of ABCB1 C3435T have shorter times to recurrence. The results confirmed the findings of previous studies and support FDA recommendation to perform pre-genotyping in patients before the choice of therapy is determined in breast cancer patients.

Matched MeSH terms: Polymorphism, Genetic
Lack of association between synapsin II (SYN2) gene polymorphism and susceptibility epilepsy: a case-control study and meta-analysis

Haerian BS, Lim KS, Tan HJ, Wong CP, Wong SW, Tan CT, et al.

Synapse, 2011 Oct;65(10):1073-9.
PMID: 21465568 DOI: 10.1002/syn.20939

The SYN2 rs3773364 A>G polymorphism has been proposed to be involved in susceptibility to epilepsy, but research results have been inconclusive. The aim of this study was to investigate the association between the SYN2 rs3773364 A>G polymorphism and susceptibility against epilepsy in a case-control study and a meta-analysis.

Matched MeSH terms: Polymorphism, Genetic/genetics*
Fulltext CYP2C9 polymorphism: prevalence in healthy and warfarin-treated Malay and Chinese in Malaysia

Ngow HA, Wan Khairina WM, Teh LK, Lee WL, Harun R, Ismail R, et al.

Singapore Med J, 2009 May;50(5):490-3.
PMID: 19495518

Genetic polymorphisms of CYP2C9 among different populations in different geographical regions could be different. CYP2C9 has been reported to be the enzyme responsible for the metabolism of many drugs, including warfarin and other drugs with a narrow therapeutic index. Realising the importance of inter-individual differences in the genetic profile in determining the outcome of a drug therapy, this study was conducted to explore the types and frequencies of CYP2C9 alleles in healthy and warfarin-treated Malays and Chinese, the two major ethnic groups in Malaysia. We aimed to evaluate the prevalence of the types and frequencies of common CYP2C9 alleles (*1, *2, *3 and *4) among the healthy unrelated individuals and diseased patients prescribed with warfarin.

Matched MeSH terms: Polymorphism, Genetic*
Fulltext Mutation analysis of the BRCA1 gene in Malaysian breast cancer patients

Balraj P, Khoo AS, Volpi L, Tan JA, Nair S, Abdullah H

Singapore Med J, 2002 Apr;43(4):194-7.
PMID: 12188064

Thirty patients with early onset breast cancer or familial breast cancer from Malaysia were analysed for germline mutation in the early onset breast cancer I gene (BRCA1). Direct sequencing of the entire coding region of BRCA1 identified a frameshift mutation, c.5447-5448insC (insC5447) (codon 1776 of exon 21) in a patient aged 32 of the Malay ethnic origin, who had no family history of breast and/or ovarian cancer. Eight polymorphisms (2201C > T, 2430T > C, P871L, E1038G, K1183R, 4427T > C, S1613G and IVS8-57delT) were identified in the samples tested.

Matched MeSH terms: Polymorphism, Genetic

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