Novel Haplotypes of genetic polymorphisms in alcohol metabolizing enzymes in Kampala, Uganda

Novel Haplotypes of genetic polymorphisms in alcohol metabolizing enzymes in Kampala, Uganda | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Short Report Novel Haplotypes of genetic polymorphisms in alcohol metabolizing enzymes in Kampala, Uganda Rodney Okwasiimire, Rhona K Baingana, Dennis M Kasozi This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5955120/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 01 Jul, 2025 Read the published version in BMC Research Notes → Version 1 posted 9 You are reading this latest preprint version Abstract Background Alcohol is metabolized to acetaldehyde by alcohol dehydrogenases (ADH) and subsequently to acetate by aldehyde dehydrogenases (ALDH). Single nucleotide polymorphisms (SNPs) in ADH1B, ADH1C and ALDH2 genes lead to haplotypes encoding isozymes which influence development of alcoholism. The distribution of these haplotypes in Uganda has not been documented. The aim of this study was to determine genotype, allele, and haplotype frequencies of SNPs in ADH1B, ADH1C , and ALDH2 genes in Uganda. Results Five SNPs: ADH1B (rs1229984 and rs2066702), ADH1C (rs1693482 and rs698) and ALDH2 (rs671) were analyzed by PCR-restriction fragment length polymorphism assays in 250 samples. The frequencies of the fast-metabolizing alleles ADH1C*1 , ADH1B*3 , and ADH1B*2 were 49.6%, 18.2% and 0.2% respectively. The nonprotective haplotype ADH1B *1 had a high frequency of 81.6% and ADH1C*2 was 10.6%. A novel allele ADH1C*new comprising G (Codon 349 Val) at ADH1C rs698 and G (Codon 271 Arg) at ADH1C rs1693482 was identified with a frequency of 39.8%. Of the seven ADH haplotype combinations identified, ADH1B *1- ADH1C *1 was the most prevalent (48.4%). Notably ADH1B*1–ADH1C* new , had the second highest frequency (25.2%). Conclusion Our study provides the first data on novel ADH1B-ADH1C haplotypes in alcohol metabolizing genes in the Ugandan population. alcohol ADH1B ADH1C ALDH2 haplotype PCR SNP Uganda Figures Figure 1 Figure 2 Introduction According to the World Health Statistics 2023 report, Uganda has a high per capita alcohol consumption of 12.21 liters of alcohol per year [ 1 ] and is ranked the first in the African region (average of 6.3 liters) and fifth globally (average of 6.18 liters) in alcohol consumption [ 1 ]. In the liver, ethanol is oxidized to acetaldehyde mainly by alcohol dehydrogenase (ADH) enzymes, with the microsomal ethanol oxidizing system (MEOS) in particular, cytochrome P450 II (CYP2E1) and peroxisomal catalase (CAT) enzymes accounting for minor oxidation. CYP2E1 and CAT encoded enzymes have higher Km values for ethanol compared to ADH encoded enzymes, and thus become significant at high ethanol concentrations in blood or when ADH enzymes become inactive or inhibited [ 2 , 3 ]. Acetaldehyde is then oxidized to acetate by aldehyde dehydrogenase (ALDH) enzymes. Acetaldehyde is highly toxic and is indicated in the pathogenesis of alcoholic liver disease, alcohol dependency and various types of cancer among other effects [ 4 ] and unpleasant effects such as Asian flushing, headache, tachycardia and hypotension in some individuals and not others. Since these outcomes are limiting factors to further alcohol consumption, they confer protective behaviors against harmful alcohol use patterns [ 5 – 7 ]. Alcohol dehydrogenases (ADH) are coded for by seven genes namely ADH1A, ADH1B, ADH1C, ADH4, ADH5, ADH6 , and ADH7 located on human chromosome 4 [ 8 ]. The seven ADH genes have been categorized into five classes (I - V) according to similarities in their amino acid sequences and catalytic properties. Class I ADH genes ( ADH1A, ADH1B , and ADH1C ) encode three subunits denoted α, β, and γ respectively. ADH isozymes are then formed by random dimeric association of any of the three subunits and belong to the low Km (< 4 mM) forms which are considered to play a major role in ethanol metabolism in the liver [ 2 , 8 , 9 ]. Functional single nucleotide polymorphisms (SNPs) have been identified in only ADH1B and ADH1C that encode the β and γ subunits respectively [ 9 ]. The extensively studied functional SNPs of these two variants include the ADH1B rs1229984 and ADH1B rs2066702 as well as ADH1C rs698 and ADH1C rs1693482. These polymorphisms result in the expression of enzyme variants that are 20 times and 2.5 times faster than their respective wild type enzymes [ 2 , 5 , 7 , 10 ]. Notably, the fast-metabolizing haplotypes ADH1B *2, ADH1B *3, and ADH1C *1 are considered protective against alcoholism [ 10 ] while haplotypes ADH1B *1 and ADH1C *2 which increase susceptibility to alcohol addiction [ 9 ]. The genes that encode ALDH enzymes are divided into nine families with the major ones being in family 1 ( ALDH1 ), that represents cytosolic ALDHs, family 2 ( ALDH2 ) that represents mitochondrial ALDHs, and family 3 ( ALDH3 ), which is found in the human stomach, saliva, and hepatocarcinoma, that represents major constitutive and inducible ALDH forms [ 2 , 7 , 11 , 12 ]. Although multiple molecular forms of ALDH enzymes are present in the human liver, only family I and II isozymes, encoded by ALDH1 and ALDH2 genes, are believed to be involved in acetaldehyde oxidation. The ALDH2 gene encodes a mitochondrial enzyme that is strongly expressed in various tissues with the highest levels found in liver. It plays a major role in acetaldehyde oxidation due to its low Km (< 5 mM) that facilitates rapid breakdown of acetaldehyde. The isozyme has two variants; the ALDH2 *1 which encodes a wild type allele and ALDH2 *2, that has a single base-pair mutation in exon 12 which results in catalytic inactivation of the enzyme [ 3 , 12 , 13 ]. Individuals who are homozygous for the mutated ALDH2 *2 variant totally lack ALDH2 activity, while individuals who are heterozygous for the ALDH2 *1/2 genotype usually express approximately 30–50% of the ALDH activity when compared to persons carrying the wild type gene [ 7 ]. Acetaldehyde dehydrogenase deficient individuals exhibit a loath response to ethanol consumption, which is likely caused by elevated levels of blood acetaldehyde. This naturally occurring response has been powerfully correlated with reduced alcohol consumption and incidence of alcoholism [ 2 , 3 , 7 , 11 , 13 ]. Despite the role of CYP2E1 and CAT, this study focused on ADH1B, ADH1C and ALDH2 due to their significant impact on alcohol metabolism. The frequencies of the genetic polymorphisms in ADH1B and ADH1C as well as ALDH2 in European, Asian and American populations are fairly well established [ 3 , 5 , 6 , 13 , 15 ]. In the African region, frequencies of genetic polymorphisms in ADH1B and ADH1C as well as ALDH2 are available for Esan in Nigeria, Gambian in Western Division, The Gambia, Luhya in Kenya, Mende in Sierra Leone and Yoruba in Ibadan, Nigeria [ 16 ]. However, this information is lacking in the Uganda population, which is ranked the first in the African region and fifth globally in terms of per capita alcohol consumption [ 1 ]. It was thus of interest to ascertain the frequencies of polymorphisms in genes for alcohol metabolizing enzymes in Uganda population. We therefore undertook this study to establish the allele, genotype and haplotype frequencies of polymorphisms in ADH1B (rs1229984 and rs2066702), ADH1C (rs1693482 and rs698) and ALDH2 (rs671) in members of a healthy Ugandan population. We also compared our genotype and allele frequency data with that from other populations around the world. Methods Study participants The study participants were 250 healthy blood donors from the Ugandan population. Located in the eastern part of Africa, Uganda has an ethnically diverse population. According to the Uganda National Population and Housing Census of 2014, nine major ethnic groups accounted for up to 68.6% of Uganda’s total population [ 14 ]. The study samples were obtained from Nakasero Blood Bank a branch of the Uganda Blood Transfusion Service. Sample preparation and DNA extraction Venous blood (≈ 3 mL) from each participant was received from the blood bank in a purple top blood collection tube containing Ethylenediaminetetraacetate (EDTA). The blood samples were centrifuged at 3000xg for 10 minutes, and the buffy coat layer was aliquoted off into a 1.5 mL micro-centrifuge tube for DNA extraction and subsequent molecular genotyping assays. Genomic DNA was manually isolated from leukocytes (buffy coat) following the protocol for Rapid Extraction of High Quality DNA from Whole Blood [ 15 ]. The extracted DNA was then stored at − 20°C until required for genotyping assays. Genotyping of ADH1B, ADH1C and ALDH2 polymorphisms All PCR assays were carried out in 20µL reaction volumes containing 1X Dream Taq PCR buffer (Thermo Scientific™, Waltham, Massachusetts, USA), 0.5U Dream Taq Polymerase (Thermo Scientific™, Waltham, Massachusetts, USA), 200µM dNTPs (Thermo Scientific™, Waltham, Massachusetts, USA), 0.2µM of each forward (F) and reverse (R) primers (Biolegio B.V, The Netherlands) as well as nuclease free water (Thermo Scientific™, Waltham, Massachusetts, USA), plus 1µL of the extracted genomic DNA. Restriction enzymes for restriction fragment length polymorphism (RFLP) assays were procured from Thermo Scientific™, Waltham, Massachusetts, USA and were used following the manufacturer’s instructions in 20µL reaction volumes. The accession numbers for ADH1B, ADH1C and ALDH2 were P00325, P00326 and P05091 respectively. The rs1229984 and rs2066702 polymorphisms in ADH1B and rs1693482 and rs698 SNPs in ADH1C were genotyped using the polymerase chain reaction-restriction fragment length polymorphism (PCR – RFLP) technique [ 10 , 16 , 17 ]. Table 1 shows the details of the primer sequences, annealing temperatures and restriction enzymes that were utilized to genotype each of the four polymorphisms in ADH1B and ADH1C . Genotype and haplotype combinations of SNPS in ADH1B (rs1229984 and rs2066702) and ADH1C (rs1693482 and rs698) were determined. The rs671 polymorphism in ALDH2 was genotyped using the allele specific polymerase chain reaction technique [ 18 , 19 ]. Details regarding the primer sequences and annealing temperature are given in Table 1. All PCR and RFLP digestion products were electrophoresed in 1X tris borate EDTA (TBE) buffer, at 200V for approximately 30 minutes and visualized on 1.5% agarose gels stained with ethidium bromide, and photographed on a UV trans-illuminator. Table 1: ADH1B, ADH1C and ALDH2 genotyping conditions Abbreviations. SNP: Single Nucleotide Polymorphism, NA: Not applicable Data analysis Data obtained from each sample regarding the observed genotype and allele was entered into Microsoft Excel 2013. After clean up, it was exported to GraphPad Prism 6 for statistical and graphical analysis. The SHEsis software [20,21] available online was used for haplotype analysis and linkage disequilibrium (LD) analysis. Hardy–Weinberg equilibrium for the allele and genotype frequencies of ADH1B and ADH1C polymorphisms was tested using the Chi Square test with Yates' continuity correction (p ≤ 0.05 was considered statistically significant). LD analysis to measure the extent of non-random association of alleles at the loci as well as haplotype analysis for polymorphisms in each of the four alcohol metabolizing enzymes was performed. Lewontin's coefficient D' and correlation coefficient r 2 were calculated as measures of LD. Results Allele and genotype frequencies of SNPs in ADH1B, ADH1C and ALDH2 Of the 250 participants, 73 (29.2%) were female. The distribution of age between female and male participants was not significantly different (p-value: 0.188). The genotype and allele frequencies of ADH1B (rs1229984 and rs2066702), ADH1C (rs1693482 and rs698) and ALDH2 (rs671) were analyzed by PCR-RFLP and AS PCR as shown in Fig.1. As shown in Table 2, the genetic loci at ADH1B and ADH1C were polymorphic while the ALDH2 locus was monomorphic. The genotype frequency distributions for ADH1B rs2066702 (X 2 = 1.94, p = 0.164) and ADH1C rs1693482 (X 2 = 2.14, p = 0.144), were in accordance with the Hardy–Weinberg equilibrium among the participants. The observed genotype and allele frequencies between members of the Ugandan population were compared with those reported from elsewhere in the world by selected studies as presented in Supplementary Tables 1 to 5. Using the chi-square test, statistically significant associations with p-values ( p ≤ 0.05) were identified. Table 2: Genotype and allele frequency distribution for ADH1B, ADH1C and ALDH2 polymorphisms p-value was obtained by Chi-square test ( X 2 ) Haplotype frequencies and linkage disequilibrium analysis for SNPs between ADH1B and ADH1C Haplotype frequencies for ADH1B (rs1229984 and rs2066702) and ADH1C (rs1693482 and rs698) are presented in Table 3. A G-G combination with a frequency of 39.8% was identified in ADH1C and has been designated as “ADH1C*new” in this paper. Table 3: Distribution of ADH1B (rs1229984 and rs2066702) and ADH1C (rs1693482 and rs698) haplotypes Gene Haplotype Observed frequency, N (%) ADH1B rs1229984 ADH1B rs2066702 ADH1B *1 G (Arg) C (Arg) 408 (81.6) ADH1B *2 A (His) C (Arg) 1 (0.2) ADH1B *3 G (Arg) T (Cys) 91 (18.2) ADH1C rs1693482 ADH1C rs698 ADH1C *1 G (Arg) A (Ile) 248 (49.6) ADH1C *2 A (Gln) G (Val) 53 (10.6) ADH1C *new G (Arg) G (Val) 199 (39.8) As shown in Table 4, a total of seven (7) haplotype combinations in SNPs between ADH1B and ADH1C were identified with the ADH1B *1- ADH1C *1 (G-C-G-A) haplotype combination having the highest frequency (48.4%) while the ADH1B *2- ADH1C *new (A-C-G-G) haplotype combination had the lowest frequency (0.2%). Table 4: Distribution of ADH1B and ADH1C combined haplotypes Haplotype Loci Frequency, N (%) ADH1B rs1229984 ADH1B rs2066702 ADH1C rs1693482 ADH1C rs698 ADH1B *1- ADH1C *1 G (Arg) C (Arg) G (Arg) A (Ile) 242 (48.4) ADH1B *1- ADH1C *2 G (Arg) C (Arg) A (Gln) G (Val) 40 (8.0) ADH1B *1- ADH1C*new G (Arg) C (Arg) G (Arg) G (Val) 126 (25.2) ADH1B *2- ADH1C*new A (His) C (Arg) G (Arg) G (Val) 1 (0.2) ADH1B *3- ADH1C *1 G (Arg) T (Cys) G (Arg) A (Ile) 6 (1.2) ADH1B *3 -ADH1C*new G (Arg) T (Cys) G (Arg) G (Val) 72 (14.4) ADH1B *3- ADH1C *2 G (Arg) T (Cys) A (Gln) G (Val) 13 (2.6) Figure 2 shows a HapMap with LD analysis for SNPs present in genes ADH1B and ADH1C . Strong LDs between rs1229984 and rs2066702 (Dʹ= 1.000, r 2 =0.000); rs1229984 and rs1693482 (Dʹ= 1.000, r 2 =0.000), rs2066702 and rs1693482 (Dʹ= 0.957, r 2 =0.024) as well as rs1693482 and rs698 polymorphisms (Dʹ= 0.999, r 2 =0.116) were observed. Discussion This study is the first to document the prevalence of genotypes, alleles and haplotypes of SNPs in ADH1B, ADH1C and ALDH2 genes that encode ethanol metabolizing enzymes in a sample of healthy members of the Ugandan population. We found a high frequency of 81.6% of the nonprotective ADH1B allele ( ADH1B *1) and a low frequency of the protective ADH1B allele at 18.2 % ( ADH1B *3). Notably, the ADH1B *1- ADH1C *1 haplotype combination had the highest frequency (48.4%). The genotype and allele frequencies of the ADH1B rs1229984 polymorphism were not significantly different ( p >0.05) from those reported for the seven African sub-populations and for the Indian population [7, 27] but were different (p ˂0.05) from those of other populations including the Chinese[22], Polish[23], Spanish[24] and Hungarian[25] as detailed in Supplementary Table 1. Similarly, the genotype and allele frequencies of the ADH1B rs2066702 polymorphism were not significantly different from those reported for the seven African sub-populations but were different from those reported for other populations including the Indian[26] and Spanish[27] outside Africa (Supplementary Table 2). In contrast, the ADH1C rs1693482 polymorphism yielded variable observations for the general African population. The allele frequencies concurred with those from the current study ( p = 0.698) while the genotype frequencies were different ( p = 0.000). However, by comparing with the seven African sub-populations showed similarity ( p ˃0.05) with the current study’s findings (Supplementary Table 3). The ADH1C rs1693482 genetic locus presented findings that differed from those of other populations including the Polish[28] and American[29] outside Africa. Regarding the ADH1C rs698 genetic locus, findings from the sampled members of the Ugandan population were different ( p ˂ 0.05) from the generalized African population and the seven African sub-populations as well as other populations elsewhere in the world as shown in Supplementary Table 4. Results of the present study show that the ALDH2 rs671 genetic locus is monomorphic for the GG genotype in the sampled members of the Ugandan population. In line with our results, previous findings for the African, American, European and South Asian populations demonstrated 100% frequency of G-allele of ALDH2 rs671(Supplementary Table 5). By contrast, the ALDH2 rs671 polymorphism was significantly different from East Asian populations [30], Japanese [19,31], Indian [4], Taiwanese [32], Czech [33] and Chinese [34]. Our study did not find any genetic variations among the sampled members of the Ugandan population with respect to ALDH2 rs671 (100% GG genotype), which has been associated with full enzyme activity. Only individuals with one (GA) or two (AA) copies of the mutant nucleotide at this ALDH2 locus have been shown to present elevated amounts of acetaldehyde following alcohol intake [8]. This finding thus implies that the members of the Ugandan population who were genotyped are not protected against alcohol dependence and alcoholism because their genetic makeup suggests that any acetaldehyde generated from ethanol metabolism will be rapidly oxidized to acetate and they are therefore not likely to experience acetaldehyde related reactions such as severe flushing, nausea, headache, hypotension and tachycardia [2,8,9]. On a positive note, the ALDH2 *1 allele which is dominant among the Ugandans we sampled has been reported to have a protective effect against alcohol-induced tissue damage [35,36], esophageal, head, neck, lung, stomach and colon cancers [35,37,38]. These protective effects are due to rapid oxidation of acetaldehyde to acetate associated with the genotype. Interestingly, a total of seven different ADH1B and ADH1C haplotype combinations were detected (Table 4). The high activity ADH1B alleles were detected in four haplotypes; three of them involving the ADH1B *3 allele and one involving the ADH1B *2 allele. Only one haplotype involved high activity alleles of ADH1B and ADH1C ( ADH1B *3 - ADH1C *1) at a frequency of 1.2%. The remaining three combinations involved the low activity ADH1C alleles, with ADH1B *1 - ADH1C *new having the highest frequency of 25.2%. The low activity ADH1B allele ( ADH1B *1) was detected in three haplotypes; with the ADH1B *1 - ADH1C *1 haplotype being the most prevalent at a rate of 48.4%. Regarding kinetic properties of enzymes encoded by ADH1B and ADH1C alleles, the ADH1B *2 allele encodes the β2 subunit that has a 40-fold higher Vmax compared to the β1which is encoded by the ADH1B *1 allele. The ADH1B *3 allele encodes the β3 subunit that has a 30-fold higher Vmax compared to the one encoded by the ADH1B *1 allele. Likewise, the ADH1C *1 allele encodes the γ1 subunit while ADH1C *2 allele encodes the γ2 subunit. The Vmax of the γ1 sub unit is twice that of the γ2 subunit [2,6,8,39]. The ADH1C *2 allele has been associated with slower metabolism of alcohol [8] and an increased risk of alcohol dependence [40]. Results of the present study thus imply that the high activity and thus protective alleles of ADH1B are less prevalent ( ADH1B *2: 0.2%, ADH1B *3: 18.2%) while the high activity allele in ADH1C ( ADH1C *1: 49.6%) is moderately prevalent among members of the Ugandan population that we studied. The low prevalence of the high activity ADH1B *3 - ADH1C *1 haplotype coupled with the absence of the inactive ALDH2 *2 allele among the genotyped Ugandans suggests that following alcohol consumption, the studied members of the Ugandan population are not likely to accumulate the toxic acetaldehyde at a high rate. Any acetaldehyde generated is not likely to accumulate in their bodies since it is easily oxidized by the acetaldehyde dehydrogenase enzyme encoded by the predominant ALDH2 *1 allele. As a result, the studied members of the Ugandan population are not likely to experience severe effects of alcohol consumption, and this could predispose them to alcoholism, alcohol dependence and other complications or disorders related to chronic alcohol consumption. This observation is supported by data from the World Health Organization’s global status report on alcohol and health that places Uganda’s alcohol consumption per capita in persons aged 15 or older at 12.21 liters of pure alcohol. This average is higher than the African (6.3 liters) and the global (6.18 liters) rating by the same report [1]. The proximity of the rs1229984 (exon 3) and rs2066702 (exon 9) of ADH1B and ADH1C (rs1693482 and rs698) genes on the long arm of chromosome 4 (4q21 to 23) suggests a strong linkage disequilibrium (Figure 2). In this study, we observed that the LD was characterized by high Dʹ (Dʹ=1.000) but low r 2 =0.000 between rs1229984 with both rs2066702 and rs1693482. In addition, rs1229984 and rs698 showed LD (Dʹ= 0.016, r 2 =0.000). In line with our findings, results from the Ensembl genome browser [30] showed no LD between rs1229984 and rs2066702. Weak LD may be explained by fact that the minor allele frequency was zero hence not enough genotype data was available to calculate LD values and r 2 was below 0.05. Furthermore, the LD between rs2066702 and rs1693482 in this study was Dʹ= 0.957 (r2=0.024). Similarly, LD varies considerably between different ethnic populations, ranging from Han Chinese in Beijing, China (Dʹ= 0.80594, r 2 =0.08064), Japanese in Tokyo Japan (Dʹ= 0.772511, r2=0.12587) and Southern Han Chinese, China (Dʹ= 0.6834, r 2 =0.1120). Our study observed a strong LD (Dʹ= 0.999, r2=0.116) between rs1693482 and rs698. Our results on LD between rs1693482 and rs698 are in agreement with those of several populations in the Ensembl genome browser [30] with LD (Dʹ= 1, r 2 =1) explained by the fact that rs1693482 and rs698 of locus ADH1C are separated by 3 kb. Limitations of the study This study had some limitations. First, samples were obtained from a population of blood donors who were considered to be healthy. Samples from participants with alcohol use disorders would have given us an understanding of whether the SNPs have any associations in the Ugandan population. Secondly, given that Uganda is an ethnically diverse country [41], in obtaining samples from the central region only, the study may have failed to document SNPs present in the other ethnic groups of Uganda. Thirdly, besides ADH1B and ADH1C, at high concentrations ethanol is also metabolized by cytochrome P450 II, peroxisomal catalase and ALDH1A1 variants which were not investigated. Conclusion In conclusion, our study contributes new knowledge of the genotype, allele and haplotype frequencies of the alcohol metabolizing enzymes: ADH1B, ADH1C and ALDH2 in a sample obtained from the Ugandan population. This study, the first of its kind in Uganda has revealed variations in population frequencies of fast-metabolizing alleles; ADH1B *2 (0.2%), ADH1B *3 (18.2%), and ADH1C *1 (49.6%) which are considered protective against alcoholism. Notably, our study identified high population frequencies of ADH1B *1 (81.6%) and ALDH2 *1 (100 %) which are considered as non-protective because they increase susceptibility to alcohol addiction. Furthermore, our study identified three new ADH1B – ADH1C haplotypes including ADH1B *3- ADH1C *new, ADH1B *1- ADH1C *new and ADH1B *2- ADH1C *new. In future research, a multi-site study with alcohol use disorder participants may be necessary to investigate SNPs associated with alcoholism in diverse ethnic populations. Furthermore, application of deep learning techniques [42]would provide crucial valuable insights on the role of the novel allele ADH1C*new. Abbreviations ADH: Alcohol dehydrogenase ALDH: Aldehyde dehydrogenase CAT: Catalase CEDAT: College of Engineering, Design, Art and Technology CYP2E1: Cytochrome P450 Family 2 Subfamily E Member 1 DNA: Deoxyribonucleic acid dNTP: Deoxynucleotide triphosphate DRGT: Directorate of Research and Graduate Training EDTA: Ethylenediaminetetraacetic acid Km: Michaelis-Menten constant LD: Linkage Disequilibrium MAKSHSREC: Makerere University School of Health Science Research Ethics Committee MAPRONANO ACE: Africa Centre of Excellence in Materials, Product Development and Nanotechnology MEOS: Microsomal Ethanol Oxidizing System NCDs: Non-Communicable Diseases PCR: Polymerase chain reaction RFLP: Restriction Fragment Length Polymorphism SIDA: Swedish International Development Cooperation Agency SNP: Single Nucleotide Polymorphism TBE: Tris Borate EDTA UBTS: Uganda Blood Transfusion Services WHO: World Health Organization Declarations Ethics approval and consent to participate Ethical clearance for this study was obtained from the Makerere University College of Health Sciences, School of Health Sciences Research and Ethics Committee (Reference no. MAKSHSREC-2020-65). A waiver for informed consent to participate in this study was sought and granted by the MAKSHSREC. Consent for publication Not Applicable Availability of data and material All the relevant data to this study has been included in the article plus supplementary tables S1 – S5. The UniProtKB/Swiss-Prot accession numbers for ADH1B, ADH1C and ALDH2 were P00325, P00326 and P05091 respectively. Competing interests No conflict declared Funding This study was funded in part by the Swedish International Development Cooperation Agency (SIDA) and Makerere University under SIDA contribution No: 51180060. Specifically, the research received financial support from the DRGT 377 Project (Postdoctoral award to DMK) of SIDA Phase IV in the Makerere-Swedish Bilateral Programme (2015-2020). R.O received a partial Master’s scholarship from the Africa Center of Excellence in Materials, Product Development and Nanotechnology (MAPRONANO ACE), CEDAT-Makerere University. Author Contributions RO, RKB and DMK performed the following: Conceptualization, methodology, validation, formal analysis, investigation, data curation, writing – original draft, writing - review and editing. RO conducted the laboratory analysis as a research project for his Master of Science in Biochemistry degree of Makerere University. RKB and DMK supervised the study. Acknowledgements We thank the Ugandan blood donors who save endless lives through their donated blood and the Uganda Blood Transfusion Services (Nakasero Blood Bank) for granting us access to blood samples for use in this study. Author Information 1 Department of Biochemistry and Systems Biology, College of Natural Sciences, Makerere University. P.O. BOX 7062 Kampala, Uganda 2 Central Diagnostic Laboratory, College of Veterinary medicine, Animal resources and Biosecurity, Makerere University. P.O. BOX 7062 Kampala, Uganda 3 Department of Agricultural Sciences, University of Helsinki, P.O. Box 28, FI-00014, Helsinki, Finland References World Health Organization. Global status report on alcohol and health 2018. David Bramley, editor. 2018. Agarwal DP. Genetic polymorphisms of alcohol metabolizing enzymes. Pathol Biol. 2001;49:703–9. Norberg A, Jones AW, Hahn RG, Gabrielsson JL. Role of variability in explaining ethanol pharmacokinetics: Research and forensic applications. Clin Pharmacokinet. 2003;42:1–31. Vaswani M, Prasad P, Kapur S. Association of ADH1B and ALDH2 gene polymorphisms with alcohol dependence: a pilot study from India. Hum Genomics. 2009;3:213–20. Seitz HK, Becker P. Alcohol metabolism and cancer risk. Alcohol Res Heal. 2007;30:38–47. Zhakari S. Overview: How Is Alcohol Metabolised by the Body? Alcohol Res Heal. 2006;29:245–54. Gemma S, Vichi S, Testai E. Individual susceptibility and alcohol effects:biochemical and genetic aspects. Ann Ist Super Sanita. 2006;42:8–16. Edenberg HJ. The genetics of alcohol metabolism: Role of alcohol dehydrogenase and aldehyde dehydrogenase variants. Alcohol Res Heal. 2007;30:5–13. Hurley TD, Edenberg H. Gene encoding enzymes in alcohol metabolism. Alcohol Res. 2012;34:339–44. Osier M V., Pakstis AJ, Soodyall H, Comas D, Goldman D, Odunsi A, et al. A Global Perspective on Genetic Variation at the ADH Genes Reveals Unusual Patterns of Linkage Disequilibrium and Diversity. Am J Hum Genet. 2002;71:84–99. Harada S, Agarwal DP, Goedde HW. Aldehyde dehydrogenase polymorphism and alcohol metabolism in alcoholics. Alcohol. 1985;2:391–2. Vasiliou V, Pappa A, Petersen DR. Role of aldehyde dehydrogenases in endogenous and xenobiotic metabolism. Chem Biol Interact. 2000;129:1–19. Ramchandani VA, Bosron WF, Li TK. Research advances in ethanol metabolism. Pathol Biol. 2001;49:676–82. Uganda Bureau of Statistics. National Population and Housing Census 2014 - Main Report, kampala. Uganda. Uganda Bur Stat. 2016;1–105. Madhad VJ, Sentheil KP. The Rapid & Non-Enzymatic isolation of DNA from the Human peripheral whole blood suitable for Genotyping. Eur J Biotechnol Biosci. 2014;1:1–16. Wolf JM, Simon D, Béria JU, Tietzmann DC, Stein AT, Lunge VR. Analysis of the Association of Nonsynonymous Polymorphisms in ADH Genes with Hazardous Drinking in HIV-1-Positive Individuals. Alcohol Clin Exp Res. 2017;41:1866–74. Xu Y, Carr LG, Bosron WF, Li TK, Edenberg HJ. Genotyping of human alcohol dehydrogenases at the ADH2 and ADH3 loci following DNA sequence amplification. Genomics. 1988;2:209–14. Kayaalti Z, Söylemezoĝlu T. Distribution of ADH1B, ALDH2, CYP2E1*6, and CYP2E1*7B genotypes in Turkish population. Alcohol. 2010;44:415–23. Tamakoshi A, Hamajima N, Kawase H, Wakai K, Katsuda N, Saito T, et al. Duplex polymerase chain reaction with confronting two-pair primers (PCR-CTPP) for genotyping alcohol dehydrogenase β subunit (ADH2) and aldehyde dehydrogenase 2 (ALDH2). Alcohol Alcohol. 2003;38:407–10. Shi YY, He L. SHEsis, a powerful software platform for analyses of linkage disequilibrium, haplotype construction, and genetic association at polymorphism loci. Cell Res. 2005;15:97–8. Zhiqiang Li, Zhao Zhang, Zangdong He, Wei Tang, Tao Li, Zhen Zeng, Lin He YS. A partition-ligation-combination-subdivision EM algorithm for haplotype inference with multiallelic markers : update of the SHEsis ( http://analysis.bio-x.cn). Cell Res. 2009;519–23. Zhang J, Jin T, Yunus Z, Li X, Geng T, Wang H, et al. Genetic polymorphisms of VIP variants in the Tajik ethnic group of northwest China. BMC Genet. 2014;15:1–9. Fang FZ, Hou L, Terry MB, Lissowska J, Morabia A, Chen J, et al. Genetic polymorphisms in alcohol metabolism, alcohol intake and the risk of stomach cancer in Warsaw, Poland. Int J Cancer. 2007;121:2060–4. Jiménez-Jiménez FJ, Gómez-Tabales J, Alonso-Navarro H, Zurdo M, Turpín-Fenoll L, Millán-Pascual J, et al. Association between the rs1229984 polymorphism in the alcohol dehydrogenase 1B gene and risk for restless legs syndrome. Sleep. 2017;40. Tóth R, Fiatal S, Petrovski B, McKee M, Ádány R. Combined effect of ADH1B rs1229984, rs2066702 and ADH1C rs1693482/rs698 alleles on alcoholism and chronic liver diseases. Dis Markers. 2011;31:267–77. Kumar Dutta A. Genetic factors affecting susceptibility to alcoholic liver disease in an Indian population. Ann Hepatol [Internet]. 2013;12:901–7. Available from: http://dx.doi.org/10.1016/S1665-2681(19)31295-5 Martínez C, Galván S, Garcia-Martin E, Ramos MI, Gutiérrez-Martín Y, Agúndez JAG. Variability in ethanol biodisposition in whites is modulated by polymorphisms in the ADH1B and ADH1C genes. Hepatology. 2010;51:491–500. Cichoz-Lach H, Celinski K, Slomka M. Alcohol-metabolizing enzyme gene polymorphisms and alcohol chronic pancreatitis among Polish individuals. Hpb. 2008;10:138–43. Djoussé L, Levy D, Herbert AG, Wilson PWF, D’Agostino RB, Cupples LA, et al. Influence of alcohol dehydrogenase 1C polymorphism on the alcohol-cardiovascular disease association (from the Framingham Offspring Study). Am J Cardiol. 2005;96:227–32. Yates AD, Achuthan P, Akanni W, Allen J, Allen J, Alvarez-Jarreta J, et al. Ensembl 2020. Nucleic Acids Res. 2020;48:D682–8. Avinçsal MO, Shinomiya H, Teshima M, Kubo M, Otsuki N, Kyota N, et al. Impact of alcohol dehydrogenase-aldehyde dehydrogenase polymorphism on clinical outcome in patients with hypopharyngeal cancer. Head Neck. 2018;40:770–7. Chien HT, Young CK, Chen TP, Liao CT, Wang HM, Cheng S De, et al. Alcohol-metabolizing enzymes’ gene polymorphisms and susceptibility to multiple head and neck cancers. Cancer Prev Res. 2019;12:247–54. Hubáček JA, Šedová L, Olišarová V, Adámková V, Adámek V, Tóthová V. Distribution of adh1b genotypes predisposed to enhanced alcohol consumption in the Czech roma/gypsy population. Cent Eur J Public Health. 2018;26:284–8. Wang Y, Du F, Zhao H, Yu X, Liu J, Xiao Y, et al. Synergistic association between two alcohol metabolism relevant genes and coronary artery disease among Chinese hypertensive patients. PLoS One. 2014;9:1–7. Chao Y ‐C, Liou S ‐R, Chung Y ‐Y, Tang H ‐S, Hsu C ‐T, Li T ‐K, et al. Polymorphism of alcohol and aldehyde dehydrogenase genes and alcoholic cirrhosis in chinese patients. Hepatology. 1994;19:360–6. Nagata K, Yamazoe Y. Genetic Polymorphism of Human Cytochrome P450 Involved in Drug Metabolism. Drug Metab Pharmacokinet. 2002;17:167–89. Matsuo K, Hamajima N, Shinoda M, Hatooka S, Inoue M, Takezaki T, et al. Erratum: Gene-environment interaction between an aldehyde dehydrogenais-2 (ALDH2) polymorphism and alcohol consumption for the risk of esophageal cancer (Carcinogenesis (2001) vol. 22 (913-916)). Carcinogenesis. 2001. p. 1893. Yokoyama A, Muramatsu T, Omori T, Matsushita S, Yoshimizu H, Higuchi S, et al. Alcohol and aldehyde dehydrogenase gene polymorphisms influence susceptibility to esophageal cancer in Japanese alcoholics. Alcohol Clin Exp Res. 1999;23:1705–10. Edenberg HJ, Xuei X, Chen HJ, Tian H, Wetherill LF, Dick DM, et al. Association of alcohol dehydrogenase genes with alcohol dependence: A comprehensive analysis. Hum Mol Genet. 2006;15:1539–49. Whitfield JB. Acute reactions to alcohol. Addict Biol. 1997;2:377–86. Gurdasani D, Carstensen T, Fatumo S, Chen G, Franklin CS, Prado-Martinez J, et al. Uganda Genome Resource Enables Insights into Population History and Genomic Discovery in Africa. Cell. 2019;179:984-1002.e36. Mohammed F. Hassan, Amanda Elswick Gentry, Elizabeth C. Prom-Wormley, Roseann E. Peterson BTW. Machine Learning Approaches to Predict Alcohol Consumption from Biomarkers in the UK Biobank. medRxiv 2024122224319486; doi https//doi.org/101101/2024122224319486. 2024;1–20. Additional Declarations No competing interests reported. Supplementary Files SupplementaryTable1.xlsx SupplementaryTable2.xlsx SupplementaryTable3.xlsx SupplementaryTable4.xlsx SupplementaryTable5.xlsx Cite Share Download PDF Status: Published Journal Publication published 01 Jul, 2025 Read the published version in BMC Research Notes → Version 1 posted Editorial decision: Revision requested 23 Apr, 2025 Reviews received at journal 23 Apr, 2025 Reviewers agreed at journal 18 Apr, 2025 Reviews received at journal 15 Apr, 2025 Reviewers agreed at journal 12 Apr, 2025 Reviewers agreed at journal 11 Apr, 2025 Reviewers invited by journal 11 Apr, 2025 Submission checks completed at journal 10 Apr, 2025 First submitted to journal 04 Apr, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5955120","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Short Report","associatedPublications":[],"authors":[{"id":442164679,"identity":"79015de7-1299-442c-8a5d-ea75590b5142","order_by":0,"name":"Rodney Okwasiimire","email":"","orcid":"","institution":"Makerere University","correspondingAuthor":false,"prefix":"","firstName":"Rodney","middleName":"","lastName":"Okwasiimire","suffix":""},{"id":442164680,"identity":"363787c4-a0b2-4e7d-a88e-d67bdebb997a","order_by":1,"name":"Rhona K Baingana","email":"","orcid":"","institution":"Makerere University","correspondingAuthor":false,"prefix":"","firstName":"Rhona","middleName":"K","lastName":"Baingana","suffix":""},{"id":442164681,"identity":"1a64f4f5-27a6-448a-98aa-867a71e9c004","order_by":2,"name":"Dennis M Kasozi","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABAUlEQVRIiWNgGAWjYDCCA0CcwMBgAOYkVDAwSJCo5QyxWhhgWhjbiNDCd7z32IMHfxiM+fsPH/7wcN42eckG5ocfGNtqcWqRPHMu3SCxjcFM4kZagkHittuGsxnYjCUY247j1GJwI8dMIrGBwYbhBo9BAlAL4zwGBjOgC4/h1nL/jZlEwh8GG/nzZwwOJM65bT+Pgf0bfi03eIBa2BjMDA7kGDYkNtxOnM3AA7KlBo9fQA5rkzA2vJGWzJBw7HbyzGaeYomEcwdwauE7fsZM8scfG8N55w8f/vij5rbtjOPtGz98KKvDqQUKkCODmQEUuYcJacEEBG0ZBaNgFIyCkQMA+txWhwGfbtQAAAAASUVORK5CYII=","orcid":"","institution":"Makerere University","correspondingAuthor":true,"prefix":"","firstName":"Dennis","middleName":"M","lastName":"Kasozi","suffix":""}],"badges":[],"createdAt":"2025-02-04 05:23:28","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5955120/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5955120/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s13104-025-07331-y","type":"published","date":"2025-07-01T15:57:24+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":80561206,"identity":"eb147568-6a8d-4aaf-b539-f4487fac033e","added_by":"auto","created_at":"2025-04-14 16:36:31","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":560165,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGenotyping of\u003c/strong\u003e\u003cem\u003e\u003cstrong\u003e ADH1B and ADH1C\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e by PCR-RFLP and \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eALDH2 rs671\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e by Allele specific PCR.\u003c/strong\u003e\u003cem\u003e\u003cstrong\u003e (\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003eA)\u003c/strong\u003e \u003cem\u003eADH1B\u003c/em\u003ers1229984 genotyping- Lane M: 100bp DNA ladder, Lanes 1\u0026amp;2: G/G genotype (undigested), Lanes 3\u0026amp;4: G/A genotype, Lane 5: Negative control \u003cstrong\u003e(B)\u003c/strong\u003e \u003cem\u003eADH1B \u003c/em\u003ers2066702 genotyping- Lane M: 50bp DNA ladder, Lanes 1\u0026amp;2: C/C genotype (undigested), Lanes 3\u0026amp;4: C/T genotype, Lanes 5\u0026amp;6: T/T genotype, Lane 7: Negative control \u003cstrong\u003e(C)\u003c/strong\u003e \u003cem\u003eADH1C\u003c/em\u003ers1693482 genotyping-Lane M: 50bp Molecular ladder, Lanes 1\u0026amp;2: G/G genotype, Lanes 3\u0026amp;4: G/A genotype, Lanes 5\u0026amp;6: A/A genotype, Lane 7: Negative control \u003cstrong\u003e(D)\u003c/strong\u003e \u003cem\u003eADH1C\u003c/em\u003e rs698 genotyping - Lane M: 50bp Molecular ladder, Lanes 1\u0026amp;2: A/A genotype, Lanes 3\u0026amp;4: A/G genotype, Lanes 5\u0026amp;6: G/G genotype (undigested), Lane 7: Negative control\u003cstrong\u003e (E) \u003c/strong\u003eALDH2 rs671 genotyping -\u003cstrong\u003e \u003c/strong\u003eLane N: Negative control,\u003cstrong\u003e \u003c/strong\u003eLane M: 100bp Molecular ladder, Lanes 1-4: G/G genotype\u003c/p\u003e","description":"","filename":"Fig1tiff300dpi.png","url":"https://assets-eu.researchsquare.com/files/rs-5955120/v1/fb55a94bacb0f9c59b7adc39.png"},{"id":80562496,"identity":"a96aa8d6-9464-4ea3-bfa7-ec640d1d3bb3","added_by":"auto","created_at":"2025-04-14 16:52:31","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":202276,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eLinkage disequilibrium plots (Hap map) for \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eADH1B\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e and \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eADH1C\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003epolymorphisms. \u003c/strong\u003eA strong LD was detected between SNPs rs1229984 and rs2066702 (Dʹ= 1.000), rs1229984 and rs1693482 (Dʹ= 1.000), rs2066702 and rs1693482 (Dʹ= 0.957) as well as rs1693482 and rs698 (Dʹ= 0.999) was detected. The SNPs rs2066702 and rs698 showed moderate LD (Dʹ= 0.669) while rs1229984 and rs698 showed a very weak LD (Dʹ= 0.016).\u003c/p\u003e","description":"","filename":"Fig2tiff300dpi.png","url":"https://assets-eu.researchsquare.com/files/rs-5955120/v1/0e66e962efa4c7f3d43e379a.png"},{"id":86180969,"identity":"9c7cef8c-11f9-4376-b7c6-53670d0d24a3","added_by":"auto","created_at":"2025-07-07 16:23:06","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1913293,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5955120/v1/8cc3396f-df33-4780-bc3f-827eb6cdf23f.pdf"},{"id":80562129,"identity":"5c97f458-a04d-44d8-bcbc-dd95a6e46e0f","added_by":"auto","created_at":"2025-04-14 16:44:31","extension":"xlsx","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":13207,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable1.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-5955120/v1/0a6da898dd0d01a1a8bbe9e6.xlsx"},{"id":80562131,"identity":"194cd293-6176-41b6-b01f-764b0f0482ee","added_by":"auto","created_at":"2025-04-14 16:44:31","extension":"xlsx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":12664,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable2.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-5955120/v1/db01cf19e0a7b6f0d8194c53.xlsx"},{"id":80563292,"identity":"ee16cda4-b9c0-44f7-84f1-4a89f9f39561","added_by":"auto","created_at":"2025-04-14 17:00:31","extension":"xlsx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":12572,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable3.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-5955120/v1/1b1b34d4aeb314185831924b.xlsx"},{"id":80561213,"identity":"5ea4c5fa-b3cf-4e03-8445-754a72dfedce","added_by":"auto","created_at":"2025-04-14 16:36:31","extension":"xlsx","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":12765,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable4.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-5955120/v1/be5127efd96842d3d71ac693.xlsx"},{"id":80562497,"identity":"be5356e7-e23b-424e-8e3d-c362e9b61c00","added_by":"auto","created_at":"2025-04-14 16:52:31","extension":"xlsx","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":12516,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable5.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-5955120/v1/98e0af615b8f7fb45764a005.xlsx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Novel Haplotypes of genetic polymorphisms in alcohol metabolizing enzymes in Kampala, Uganda","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAccording to the World Health Statistics 2023 report, Uganda has a high per capita alcohol consumption of 12.21 liters of alcohol per year [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e] and is ranked the first in the African region (average of 6.3 liters) and fifth globally (average of 6.18 liters) in alcohol consumption [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn the liver, ethanol is oxidized to acetaldehyde mainly by alcohol dehydrogenase (ADH) enzymes, with the microsomal ethanol oxidizing system (MEOS) in particular, cytochrome P450 II (CYP2E1) and peroxisomal catalase (CAT) enzymes accounting for minor oxidation. \u003cem\u003eCYP2E1\u003c/em\u003e and \u003cem\u003eCAT\u003c/em\u003e encoded enzymes have higher Km values for ethanol compared to \u003cem\u003eADH\u003c/em\u003e encoded enzymes, and thus become significant at high ethanol concentrations in blood or when ADH enzymes become inactive or inhibited [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Acetaldehyde is then oxidized to acetate by aldehyde dehydrogenase (ALDH) enzymes. Acetaldehyde is highly toxic and is indicated in the pathogenesis of alcoholic liver disease, alcohol dependency and various types of cancer among other effects [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e] and unpleasant effects such as Asian flushing, headache, tachycardia and hypotension in some individuals and not others. Since these outcomes are limiting factors to further alcohol consumption, they confer protective behaviors against harmful alcohol use patterns [\u003cspan additionalcitationids=\"CR6\" citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eAlcohol dehydrogenases (ADH) are coded for by seven genes namely \u003cem\u003eADH1A, ADH1B, ADH1C, ADH4, ADH5, ADH6\u003c/em\u003e, and \u003cem\u003eADH7\u003c/em\u003e located on human chromosome 4 [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. The seven \u003cem\u003eADH\u003c/em\u003e genes have been categorized into five classes (I - V) according to similarities in their amino acid sequences and catalytic properties. Class I \u003cem\u003eADH\u003c/em\u003e genes (\u003cem\u003eADH1A, ADH1B\u003c/em\u003e, and \u003cem\u003eADH1C\u003c/em\u003e) encode three subunits denoted α, β, and γ respectively. ADH isozymes are then formed by random dimeric association of any of the three subunits and belong to the low Km (\u0026lt;\u0026thinsp;4 mM) forms which are considered to play a major role in ethanol metabolism in the liver [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Functional single nucleotide polymorphisms (SNPs) have been identified in only \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e that encode the β and γ subunits respectively [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. The extensively studied functional SNPs of these two variants include the \u003cem\u003eADH1B\u003c/em\u003e rs1229984 and \u003cem\u003eADH1B\u003c/em\u003e rs2066702 as well as \u003cem\u003eADH1C\u003c/em\u003e rs698 and \u003cem\u003eADH1C\u003c/em\u003e rs1693482. These polymorphisms result in the expression of enzyme variants that are 20 times and 2.5 times faster than their respective wild type enzymes [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Notably, the fast-metabolizing haplotypes \u003cem\u003eADH1B\u003c/em\u003e*2, \u003cem\u003eADH1B\u003c/em\u003e*3, and \u003cem\u003eADH1C\u003c/em\u003e*1 are considered protective against alcoholism [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e] while haplotypes \u003cem\u003eADH1B\u003c/em\u003e*1 and \u003cem\u003eADH1C\u003c/em\u003e*2 which increase susceptibility to alcohol addiction [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe genes that encode ALDH enzymes are divided into nine families with the major ones being in family 1 (\u003cem\u003eALDH1\u003c/em\u003e), that represents cytosolic ALDHs, family 2 (\u003cem\u003eALDH2\u003c/em\u003e) that represents mitochondrial ALDHs, and family 3 (\u003cem\u003eALDH3\u003c/em\u003e), which is found in the human stomach, saliva, and hepatocarcinoma, that represents major constitutive and inducible ALDH forms [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Although multiple molecular forms of ALDH enzymes are present in the human liver, only family I and II isozymes, encoded by \u003cem\u003eALDH1\u003c/em\u003e and \u003cem\u003eALDH2\u003c/em\u003e genes, are believed to be involved in acetaldehyde oxidation. The \u003cem\u003eALDH2\u003c/em\u003e gene encodes a mitochondrial enzyme that is strongly expressed in various tissues with the highest levels found in liver. It plays a major role in acetaldehyde oxidation due to its low Km (\u0026lt;\u0026thinsp;5 mM) that facilitates rapid breakdown of acetaldehyde. The isozyme has two variants; the \u003cem\u003eALDH2\u003c/em\u003e*1 which encodes a wild type allele and \u003cem\u003eALDH2\u003c/em\u003e*2, that has a single base-pair mutation in exon 12 which results in catalytic inactivation of the enzyme [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. Individuals who are homozygous for the mutated \u003cem\u003eALDH2\u003c/em\u003e*2 variant totally lack ALDH2 activity, while individuals who are heterozygous for the \u003cem\u003eALDH2\u003c/em\u003e*1/2 genotype usually express approximately 30\u0026ndash;50% of the ALDH activity when compared to persons carrying the wild type gene [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Acetaldehyde dehydrogenase deficient individuals exhibit a loath response to ethanol consumption, which is likely caused by elevated levels of blood acetaldehyde. This naturally occurring response has been powerfully correlated with reduced alcohol consumption and incidence of alcoholism [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eDespite the role of CYP2E1 and CAT, this study focused on ADH1B, ADH1C and ALDH2 due to their significant impact on alcohol metabolism. The frequencies of the genetic polymorphisms in \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e as well as \u003cem\u003eALDH2\u003c/em\u003e in European, Asian and American populations are fairly well established [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e, \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. In the African region, frequencies of genetic polymorphisms in \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e as well as \u003cem\u003eALDH2\u003c/em\u003e are available for Esan in Nigeria, Gambian in Western Division, The Gambia, Luhya in Kenya, Mende in Sierra Leone and Yoruba in Ibadan, Nigeria [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. However, this information is lacking in the Uganda population, which is ranked the first in the African region and fifth globally in terms of per capita alcohol consumption [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. It was thus of interest to ascertain the frequencies of polymorphisms in genes for alcohol metabolizing enzymes in Uganda population. We therefore undertook this study to establish the allele, genotype and haplotype frequencies of polymorphisms in \u003cem\u003eADH1B\u003c/em\u003e (rs1229984 and rs2066702), \u003cem\u003eADH1C\u003c/em\u003e (rs1693482 and rs698) and \u003cem\u003eALDH2\u003c/em\u003e (rs671) in members of a healthy Ugandan population. We also compared our genotype and allele frequency data with that from other populations around the world.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStudy participants\u003c/h2\u003e \u003cp\u003eThe study participants were 250 healthy blood donors from the Ugandan population. Located in the eastern part of Africa, Uganda has an ethnically diverse population. According to the Uganda National Population and Housing Census of 2014, nine major ethnic groups accounted for up to 68.6% of Uganda\u0026rsquo;s total population [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. The study samples were obtained from Nakasero Blood Bank a branch of the Uganda Blood Transfusion Service.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eSample preparation and DNA extraction\u003c/h3\u003e\n\u003cp\u003eVenous blood (\u0026asymp;\u0026thinsp;3 mL) from each participant was received from the blood bank in a purple top blood collection tube containing Ethylenediaminetetraacetate (EDTA). The blood samples were centrifuged at 3000xg for 10 minutes, and the buffy coat layer was aliquoted off into a 1.5 mL micro-centrifuge tube for DNA extraction and subsequent molecular genotyping assays. Genomic DNA was manually isolated from leukocytes (buffy coat) following the protocol for Rapid Extraction of High Quality DNA from Whole Blood [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. The extracted DNA was then stored at \u0026minus;\u0026thinsp;20\u0026deg;C until required for genotyping assays.\u003c/p\u003e\n\u003ch3\u003eGenotyping of ADH1B, ADH1C and ALDH2 polymorphisms\u003c/h3\u003e\n\u003cp\u003eAll PCR assays were carried out in 20\u0026micro;L reaction volumes containing 1X Dream Taq PCR buffer (Thermo Scientific\u0026trade;, Waltham, Massachusetts, USA), 0.5U Dream Taq Polymerase (Thermo Scientific\u0026trade;, Waltham, Massachusetts, USA), 200\u0026micro;M dNTPs (Thermo Scientific\u0026trade;, Waltham, Massachusetts, USA), 0.2\u0026micro;M of each forward (F) and reverse (R) primers (Biolegio B.V, The Netherlands) as well as nuclease free water (Thermo Scientific\u0026trade;, Waltham, Massachusetts, USA), plus 1\u0026micro;L of the extracted genomic DNA. Restriction enzymes for restriction fragment length polymorphism (RFLP) assays were procured from Thermo Scientific\u0026trade;, Waltham, Massachusetts, USA and were used following the manufacturer\u0026rsquo;s instructions in 20\u0026micro;L reaction volumes.\u003c/p\u003e \u003cp\u003eThe accession numbers for ADH1B, ADH1C and ALDH2 were P00325, P00326 and P05091 respectively. The rs1229984 and rs2066702 polymorphisms in \u003cem\u003eADH1B\u003c/em\u003e and rs1693482 and rs698 SNPs in \u003cem\u003eADH1C\u003c/em\u003e were genotyped using the polymerase chain reaction-restriction fragment length polymorphism (PCR \u0026ndash; RFLP) technique [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. Table\u0026nbsp;1 shows the details of the primer sequences, annealing temperatures and restriction enzymes that were utilized to genotype each of the four polymorphisms in \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e. Genotype and haplotype combinations of SNPS in \u003cem\u003eADH1B\u003c/em\u003e (rs1229984 and rs2066702) and \u003cem\u003eADH1C\u003c/em\u003e (rs1693482 and rs698) were determined.\u003c/p\u003e \u003cp\u003eThe rs671 polymorphism in \u003cem\u003eALDH2\u003c/em\u003e was genotyped using the allele specific polymerase chain reaction technique [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Details regarding the primer sequences and annealing temperature are given in Table\u0026nbsp;1. All PCR and RFLP digestion products were electrophoresed in 1X tris borate EDTA (TBE) buffer, at 200V for approximately 30 minutes and visualized on 1.5% agarose gels stained with ethidium bromide, and photographed on a UV trans-illuminator.\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eTable 1: \u003cem\u003eADH1B,\u003c/em\u003e \u003cem\u003eADH1C\u003c/em\u003e and\u003c/strong\u003e \u003cstrong\u003e\u003cem\u003eALDH2\u003c/em\u003e genotyping conditions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cimg src=\"data:image/png;base64,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\"\u003e\u003c/strong\u003e\u003cbr\u003e\u003c/p\u003e\n\u003cp\u003eAbbreviations.\u0026nbsp;SNP: Single Nucleotide Polymorphism, NA: Not applicable\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eData analysis\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eData obtained from each sample regarding the observed genotype and allele was entered into Microsoft Excel 2013. After clean up, it was exported to GraphPad Prism 6 for statistical and graphical analysis. The SHEsis software [20,21] available online \u0026nbsp;was used for haplotype analysis and linkage disequilibrium (LD) analysis.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eHardy\u0026ndash;Weinberg equilibrium for the allele and genotype frequencies of \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e polymorphisms was tested using the Chi Square test with Yates\u0026apos; continuity correction (p \u0026le; 0.05 was considered statistically significant). LD analysis to measure the extent of non-random association of alleles at the loci as well as haplotype analysis for polymorphisms in each of the four alcohol metabolizing enzymes was performed. Lewontin\u0026apos;s coefficient D\u0026apos; and correlation coefficient r\u003csup\u003e2\u003c/sup\u003e were calculated as measures of LD.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cem\u003eAllele and genotype frequencies of SNPs in\u0026nbsp;\u003c/em\u003e\u003cem\u003eADH1B,\u0026nbsp;\u003c/em\u003e\u003cem\u003eADH1C\u0026nbsp;\u003c/em\u003e\u003cem\u003eand ALDH2\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eOf the 250 participants, 73 (29.2%) were female. The distribution of age between female and male participants was not significantly different (p-value: 0.188). The genotype and allele frequencies of \u003cem\u003eADH1B\u003c/em\u003e (rs1229984 and rs2066702), \u003cem\u003eADH1C\u003c/em\u003e (rs1693482 and rs698) and \u003cem\u003eALDH2\u003c/em\u003e (rs671) were analyzed by PCR-RFLP and AS PCR as shown in Fig.1. \u0026nbsp;As shown in Table 2, the genetic loci at \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e were polymorphic while the \u003cem\u003eALDH2\u003c/em\u003e locus was monomorphic. The genotype frequency distributions for \u003cem\u003eADH1B\u0026nbsp;\u003c/em\u003ers2066702 (X\u003csup\u003e2\u003c/sup\u003e = 1.94, \u003cem\u003ep\u003c/em\u003e = 0.164) and \u003cem\u003eADH1C\u003c/em\u003ers1693482 (X\u003csup\u003e2\u003c/sup\u003e = 2.14, \u003cem\u003ep\u003c/em\u003e = 0.144), were in accordance with the Hardy\u0026ndash;Weinberg equilibrium among the participants. The observed genotype and allele frequencies between members of the Ugandan population were compared with those reported from elsewhere in the world by selected studies as presented in Supplementary Tables 1 to 5. Using the chi-square test, statistically significant associations with \u003cem\u003ep-values\u003c/em\u003e (\u003cem\u003ep\u003c/em\u003e \u0026le; 0.05) were identified.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2: Genotype and allele frequency distribution for \u003cem\u003eADH1B, ADH1C\u003c/em\u003e and \u003cem\u003eALDH2\u003c/em\u003e polymorphisms\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cimg src=\"data:image/png;base64,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\"\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003ep-value\u003c/em\u003e was obtained by Chi-square test (\u003cem\u003eX\u003csup\u003e2\u003c/sup\u003e\u003c/em\u003e)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eHaplotype frequencies and linkage disequilibrium analysis for SNPs between ADH1B and ADH1C\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eHaplotype frequencies for ADH1B (rs1229984 and rs2066702) and ADH1C (rs1693482 and rs698) are presented in Table 3. A G-G combination with a frequency of 39.8% was identified in ADH1C and has been designated as \u0026ldquo;ADH1C*new\u0026rdquo; in this paper.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"631\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"4\" style=\"width: 631px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eTable 3: Distribution of \u003cem\u003eADH1B\u003c/em\u003e (rs1229984 and rs2066702) and \u003cem\u003eADH1C\u003c/em\u003e (rs1693482 and rs698) haplotypes\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 104px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGene\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 321px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eHaplotype\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 206px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eObserved frequency, N (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eADH1B\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;rs1229984\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eADH1B\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003ers2066702\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eC (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 206px;\"\u003e\n \u003cp\u003e408 (81.6)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eA (His)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eC (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 206px;\"\u003e\n \u003cp\u003e1 (0.2)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eT (Cys)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 206px;\"\u003e\n \u003cp\u003e91 (18.2)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eADH1C\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;rs1693482\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eADH1C\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;rs698\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 206px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1C\u003c/em\u003e*1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eA (Ile)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 206px;\"\u003e\n \u003cp\u003e248 (49.6)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1C\u003c/em\u003e*2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eA (Gln)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eG (Val)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 206px;\"\u003e\n \u003cp\u003e53 (10.6)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1C\u003c/em\u003e*new\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 161px;\"\u003e\n \u003cp\u003eG (Val)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 206px;\"\u003e\n \u003cp\u003e199 (39.8)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eAs shown in Table 4, a total of seven (7) haplotype combinations in SNPs between \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e were identified with the \u003cem\u003eADH1B\u003c/em\u003e*1-\u003cem\u003eADH1C\u003c/em\u003e*1 (G-C-G-A) haplotype combination having the highest frequency (48.4%) while the \u003cem\u003eADH1B\u003c/em\u003e*2-\u003cem\u003eADH1C\u003c/em\u003e*new (A-C-G-G) haplotype combination had the lowest frequency (0.2%).\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"623\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"6\" style=\"width: 623px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eTable\u003cem\u003e\u0026nbsp;\u003c/em\u003e4: Distribution of \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e combined haplotypes\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eHaplotype\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"4\" style=\"width: 342px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eLoci\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 102px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eFrequency, N (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eADH1B\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;rs1229984\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eADH1B\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;rs2066702\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eADH1C\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;rs1693482\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 78px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eADH1C\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;rs698\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*1-\u003cem\u003e\u0026nbsp;ADH1C\u003c/em\u003e*1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eC (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 78px;\"\u003e\n \u003cp\u003eA (Ile)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e242 (48.4)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*1-\u003cem\u003e\u0026nbsp;ADH1C\u003c/em\u003e*2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eC (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003eA (Gln)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 78px;\"\u003e\n \u003cp\u003eG (Val)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e40 (8.0)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*1-\u003cem\u003e\u0026nbsp;ADH1C*new\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eC (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 78px;\"\u003e\n \u003cp\u003eG (Val)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e126 (25.2)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*2-\u003cem\u003e\u0026nbsp;ADH1C*new\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eA (His)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eC (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 78px;\"\u003e\n \u003cp\u003eG (Val)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e1 (0.2)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*3-\u003cem\u003e\u0026nbsp;ADH1C\u003c/em\u003e*1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eT (Cys)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 78px;\"\u003e\n \u003cp\u003eA (Ile)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e6 (1.2)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*3\u003cem\u003e\u0026nbsp;-ADH1C*new\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eT (Cys)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 78px;\"\u003e\n \u003cp\u003eG (Val)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e72 (14.4)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cem\u003eADH1B\u003c/em\u003e*3-\u003cem\u003e\u0026nbsp;ADH1C\u003c/em\u003e*2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eG (Arg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003eT (Cys)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003eA (Gln)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 78px;\"\u003e\n \u003cp\u003eG (Val)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e13 (2.6)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eFigure 2 shows a HapMap with LD analysis for SNPs present in genes \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e. Strong LDs between rs1229984 and rs2066702 (Dʹ= 1.000, r\u003csup\u003e2\u003c/sup\u003e=0.000);\u0026nbsp;rs1229984 and rs1693482 (Dʹ= 1.000, r\u003csup\u003e2\u003c/sup\u003e=0.000), rs2066702 and rs1693482 (Dʹ= 0.957, r\u003csup\u003e2\u003c/sup\u003e=0.024) as well as rs1693482 and rs698 polymorphisms (Dʹ= 0.999, r\u003csup\u003e2\u003c/sup\u003e=0.116) were observed.\u0026nbsp;\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThis study is the first to document the prevalence of genotypes, alleles and haplotypes of SNPs in \u003cem\u003eADH1B, ADH1C\u003c/em\u003e and \u003cem\u003eALDH2\u003c/em\u003e genes that encode ethanol metabolizing enzymes in a sample of healthy members of the Ugandan population. We found a high frequency of 81.6% of the nonprotective \u003cem\u003eADH1B\u0026nbsp;\u003c/em\u003eallele (\u003cem\u003eADH1B\u003c/em\u003e*1) and a low frequency of the protective \u003cem\u003eADH1B\u003c/em\u003e allele at 18.2 % (\u003cem\u003eADH1B\u003c/em\u003e*3). Notably, the \u003cem\u003eADH1B\u003c/em\u003e*1-\u003cem\u003eADH1C\u003c/em\u003e*1 haplotype combination had the highest frequency (48.4%).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe genotype and allele frequencies of the \u003cem\u003eADH1B\u003c/em\u003e rs1229984 polymorphism were not significantly different (\u003cem\u003ep\u003c/em\u003e \u0026gt;0.05) from those reported for the seven African sub-populations and for the Indian population [7, 27] but were different (p ˂0.05) from those of other populations including the Chinese[22], Polish[23], Spanish[24] and Hungarian[25] as detailed in Supplementary Table 1. Similarly, the genotype and allele frequencies of the \u003cem\u003eADH1B\u003c/em\u003e rs2066702 polymorphism were not significantly different from those reported for the seven African sub-populations but were different from those reported for other populations including the Indian[26]\u0026nbsp;and Spanish[27]\u0026nbsp; outside Africa (Supplementary Table 2). In contrast, the \u003cem\u003eADH1C\u003c/em\u003e rs1693482 polymorphism yielded variable observations for the general African population. The allele frequencies concurred with those from the current study (\u003cem\u003ep\u003c/em\u003e = 0.698) while the genotype frequencies were different (\u003cem\u003ep\u003c/em\u003e = 0.000). However, by comparing with the seven African sub-populations showed similarity (\u003cem\u003ep\u003c/em\u003e ˃0.05) with the current study\u0026rsquo;s findings (Supplementary Table 3). The ADH1C rs1693482 genetic locus presented findings that differed from those of other populations including the Polish[28]\u0026nbsp;and American[29]\u0026nbsp; outside Africa. Regarding the \u003cem\u003eADH1C\u003c/em\u003e rs698 genetic locus, findings from the sampled members of the Ugandan population were different (\u003cem\u003ep\u003c/em\u003e ˂ 0.05) from the generalized African population and the seven African sub-populations as well as other populations elsewhere in the world as shown in Supplementary Table 4.\u003c/p\u003e\n\u003cp\u003eResults of the present study show that the \u003cem\u003eALDH2\u003c/em\u003e rs671 genetic locus is monomorphic for the GG genotype in the sampled members of the Ugandan population. In line with our results, previous findings for the African, American, European and South Asian populations demonstrated 100% frequency of G-allele of \u003cem\u003eALDH2\u003c/em\u003e rs671(Supplementary Table 5). By contrast, the \u003cem\u003eALDH2\u003c/em\u003e rs671 polymorphism was significantly different from East Asian populations [30], Japanese [19,31], Indian [4], Taiwanese [32], Czech [33] and Chinese [34].\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eOur study did not find any genetic variations among the sampled members of the Ugandan population with respect to \u003cem\u003eALDH2\u003c/em\u003e rs671 (100% GG genotype), which has been associated with full enzyme activity. Only individuals with one (GA) or two (AA) copies of the mutant nucleotide at this \u003cem\u003eALDH2\u0026nbsp;\u003c/em\u003elocus have been shown to present elevated amounts of acetaldehyde following alcohol intake [8]. This finding thus implies that the members of the Ugandan population who were genotyped are not protected against alcohol dependence and alcoholism because their genetic makeup suggests that any acetaldehyde generated from ethanol metabolism will be rapidly oxidized to acetate and they are therefore not likely to experience acetaldehyde related reactions such as severe flushing, nausea, headache, hypotension and tachycardia [2,8,9]. On a positive note, the \u003cem\u003eALDH2\u003c/em\u003e*1 allele which is dominant among the Ugandans we sampled has been reported to have a protective effect against alcohol-induced tissue damage [35,36], esophageal, head, neck, lung, stomach and colon cancers [35,37,38]. These protective effects are due to rapid oxidation of acetaldehyde to acetate associated with the genotype.\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;Interestingly, a total of seven different \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e haplotype combinations were detected (Table 4). The high activity \u003cem\u003eADH1B\u003c/em\u003e alleles were detected in four haplotypes; three of them involving the \u003cem\u003eADH1B\u003c/em\u003e*3 allele and one involving the \u003cem\u003eADH1B\u003c/em\u003e*2 allele. Only one haplotype involved high activity alleles of \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e (\u003cem\u003eADH1B\u003c/em\u003e*3 - \u003cem\u003eADH1C\u003c/em\u003e*1) at a frequency of 1.2%. The remaining three combinations involved the low activity \u003cem\u003eADH1C\u003c/em\u003e alleles, with \u003cem\u003eADH1B\u003c/em\u003e*1 - \u003cem\u003eADH1C\u003c/em\u003e*new having the highest frequency of 25.2%. The low activity \u003cem\u003eADH1B\u0026nbsp;\u003c/em\u003eallele (\u003cem\u003eADH1B\u003c/em\u003e*1) was detected in three haplotypes; with the \u003cem\u003eADH1B\u003c/em\u003e*1 - \u003cem\u003eADH1C\u003c/em\u003e*1 haplotype being the most prevalent at a rate of 48.4%.\u003c/p\u003e\n\u003cp\u003eRegarding kinetic properties of enzymes encoded by \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e alleles, the \u003cem\u003eADH1B\u003c/em\u003e*2 allele encodes the \u0026beta;2 subunit that has a 40-fold higher Vmax compared to the \u0026beta;1which is encoded by the \u003cem\u003eADH1B\u003c/em\u003e*1 allele. The \u003cem\u003eADH1B\u003c/em\u003e*3 allele encodes the \u0026beta;3 subunit that has a 30-fold higher Vmax compared to the one encoded by the \u003cem\u003eADH1B\u003c/em\u003e*1 allele. Likewise, the \u003cem\u003eADH1C\u003c/em\u003e*1 allele encodes the \u0026gamma;1 subunit while \u003cem\u003eADH1C\u003c/em\u003e*2 allele encodes the \u0026gamma;2 subunit. The Vmax of the \u0026gamma;1 sub unit is twice that of the \u0026gamma;2 subunit [2,6,8,39]. The \u003cem\u003eADH1C\u003c/em\u003e*2 allele has been associated with slower metabolism of alcohol [8] and an increased risk of alcohol dependence [40]. Results of the present study thus imply that the high activity and thus protective alleles of \u003cem\u003eADH1B\u003c/em\u003e are less prevalent (\u003cem\u003eADH1B\u003c/em\u003e*2: 0.2%, \u003cem\u003eADH1B\u003c/em\u003e*3: 18.2%) while the high activity allele in \u003cem\u003eADH1C\u0026nbsp;\u003c/em\u003e(\u003cem\u003eADH1C\u003c/em\u003e*1: 49.6%) is moderately prevalent among members of the Ugandan population that we studied. The low prevalence of the high activity \u003cem\u003eADH1B\u003c/em\u003e*3 - \u003cem\u003eADH1C\u003c/em\u003e*1 haplotype coupled with the absence of the inactive \u003cem\u003eALDH2\u003c/em\u003e*2 allele among the genotyped Ugandans suggests that following alcohol consumption, the studied members of the Ugandan population are not likely to accumulate the toxic acetaldehyde at a high rate. Any acetaldehyde generated is not likely to accumulate in their bodies since it is easily oxidized by the acetaldehyde dehydrogenase enzyme encoded by the predominant \u003cem\u003eALDH2\u003c/em\u003e*1 allele. As a result, the studied members of the Ugandan population are not likely to experience severe effects of alcohol consumption, and this could predispose them to alcoholism, alcohol dependence and other complications or disorders related to chronic alcohol consumption. This observation is supported by data from the World Health Organization\u0026rsquo;s global status report on alcohol and health that places Uganda\u0026rsquo;s alcohol consumption per capita in persons aged 15 or older at 12.21 liters of pure alcohol. This average is higher than the African (6.3 liters) and the global (6.18 liters) rating by the same report [1].\u003c/p\u003e\n\u003cp\u003eThe proximity of the rs1229984 (exon 3) and rs2066702 (exon 9) of \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C\u003c/em\u003e (rs1693482 and rs698) genes on the long arm of chromosome 4 (4q21 to 23) suggests a strong linkage disequilibrium (Figure 2). In this study, we observed that the LD was characterized by high Dʹ (Dʹ=1.000) but low r\u003csup\u003e2\u003c/sup\u003e=0.000 between rs1229984 with both rs2066702 and rs1693482. In addition, rs1229984 and rs698 showed LD (Dʹ= 0.016, r\u003csup\u003e2\u003c/sup\u003e=0.000). In line with our findings, results from the Ensembl genome browser\u0026nbsp;[30]\u0026nbsp;showed no LD between rs1229984 and rs2066702. Weak LD may be explained by fact that the minor allele frequency was zero hence not enough genotype data was available to calculate LD values and r\u003csup\u003e2\u003c/sup\u003e was below 0.05. \u0026nbsp;Furthermore, the LD between rs2066702 and rs1693482 in this study was Dʹ= 0.957 (r2=0.024). Similarly, LD varies considerably between different ethnic populations, ranging from Han Chinese in Beijing, China (Dʹ= 0.80594, r\u003csup\u003e2\u003c/sup\u003e=0.08064), Japanese in Tokyo Japan (Dʹ= 0.772511, r2=0.12587) and Southern Han Chinese, China (Dʹ= 0.6834, r\u003csup\u003e2\u003c/sup\u003e=0.1120). Our study observed a strong LD (Dʹ= 0.999, r2=0.116) between rs1693482 and rs698. Our results on LD between rs1693482 and rs698 are in agreement with those of several populations in the Ensembl genome browser [30]\u0026nbsp;with LD (Dʹ= 1, r\u003csup\u003e2\u003c/sup\u003e=1) explained by the fact that rs1693482 and rs698 of locus \u003cem\u003eADH1C\u003c/em\u003e are separated by 3 kb.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e\u0026nbsp;Limitations of the study \u0026nbsp; \u0026nbsp;\u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThis study had some limitations. First, samples were obtained from a population of blood donors who were considered to be healthy. Samples from participants with alcohol use disorders would have given us an understanding of whether the SNPs have any associations in the Ugandan population. Secondly, given that Uganda is an ethnically diverse country [41], in obtaining samples from the central region only, the study may have failed to document SNPs present in the other ethnic groups of Uganda. Thirdly, besides \u003cem\u003eADH1B\u003c/em\u003e and \u003cem\u003eADH1C,\u003c/em\u003e at high concentrations ethanol is also metabolized by cytochrome P450 II, peroxisomal catalase and \u003cem\u003eALDH1A1\u003c/em\u003e variants which were not investigated. \u0026nbsp;\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eIn conclusion, our study contributes new knowledge of the genotype, allele and haplotype frequencies of the alcohol metabolizing enzymes: \u003cem\u003eADH1B, ADH1C\u003c/em\u003e and \u003cem\u003eALDH2\u003c/em\u003e in a sample obtained from the Ugandan population. This study, the first of its kind in Uganda has revealed variations in population frequencies of fast-metabolizing alleles; \u003cem\u003eADH1B\u003c/em\u003e*2 (0.2%), \u003cem\u003eADH1B\u003c/em\u003e*3 (18.2%), and \u003cem\u003eADH1C\u003c/em\u003e*1 (49.6%) which are considered protective against alcoholism. Notably, our study identified high population frequencies of \u003cem\u003eADH1B\u003c/em\u003e*1 (81.6%) and \u003cem\u003eALDH2\u003c/em\u003e*1 (100 %) which are considered as non-protective because they increase susceptibility to alcohol addiction. Furthermore, our study identified three new \u003cem\u003eADH1B\u003c/em\u003e \u0026ndash;\u003cem\u003eADH1C\u003c/em\u003e haplotypes including \u003cem\u003eADH1B\u003c/em\u003e*3-\u003cem\u003eADH1C\u003c/em\u003e*new, \u003cem\u003eADH1B\u003c/em\u003e*1-\u003cem\u003eADH1C\u003c/em\u003e*new and \u003cem\u003eADH1B\u003c/em\u003e*2-\u003cem\u003eADH1C\u003c/em\u003e*new. In future research, a multi-site study with alcohol use disorder participants may be necessary to investigate SNPs associated with alcoholism in diverse ethnic populations. Furthermore, application of deep learning techniques [42]would provide crucial valuable insights on the role of the novel allele \u003cem\u003eADH1C*new.\u003c/em\u003e\u003c/p\u003e\n"},{"header":"Abbreviations","content":"\u003cp\u003eADH: Alcohol dehydrogenase\u003c/p\u003e\n\u003cp\u003eALDH: Aldehyde dehydrogenase\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eCAT: Catalase\u003c/p\u003e\n\u003cp\u003eCEDAT: College of Engineering, Design, Art and Technology\u003c/p\u003e\n\u003cp\u003eCYP2E1: Cytochrome P450 Family 2 Subfamily E Member 1\u003c/p\u003e\n\u003cp\u003eDNA: Deoxyribonucleic acid\u003c/p\u003e\n\u003cp\u003edNTP: Deoxynucleotide triphosphate\u003c/p\u003e\n\u003cp\u003eDRGT: Directorate of Research and Graduate Training\u003c/p\u003e\n\u003cp\u003eEDTA: Ethylenediaminetetraacetic acid\u003c/p\u003e\n\u003cp\u003eKm: Michaelis-Menten constant\u003c/p\u003e\n\u003cp\u003eLD: Linkage Disequilibrium\u003c/p\u003e\n\u003cp\u003eMAKSHSREC: Makerere University School of Health Science Research Ethics Committee\u003c/p\u003e\n\u003cp\u003eMAPRONANO ACE: Africa Centre of Excellence in Materials, Product Development and Nanotechnology\u003c/p\u003e\n\u003cp\u003eMEOS: Microsomal Ethanol Oxidizing System\u003c/p\u003e\n\u003cp\u003eNCDs: Non-Communicable Diseases\u003c/p\u003e\n\u003cp\u003ePCR: Polymerase chain reaction\u003c/p\u003e\n\u003cp\u003eRFLP: Restriction Fragment Length Polymorphism\u003c/p\u003e\n\u003cp\u003eSIDA: Swedish International Development Cooperation Agency\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eSNP: Single Nucleotide Polymorphism\u003c/p\u003e\n\u003cp\u003eTBE: Tris Borate EDTA\u003c/p\u003e\n\u003cp\u003eUBTS: Uganda Blood Transfusion Services\u003c/p\u003e\n\u003cp\u003eWHO: World Health Organization\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEthical clearance for this study was obtained from the Makerere University College of Health Sciences, School of Health Sciences Research and Ethics Committee (Reference no. MAKSHSREC-2020-65). A waiver for informed consent to participate in this study was sought and granted by the MAKSHSREC.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot Applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and material\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll the relevant data to this study has been included in the article plus supplementary tables S1 \u0026ndash; S5. The UniProtKB/Swiss-Prot accession numbers for ADH1B, ADH1C and ALDH2 were P00325, P00326 and P05091 respectively.\u003cem\u003e\u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNo conflict declared\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was funded in part by the Swedish International Development Cooperation Agency (SIDA) and Makerere University under SIDA contribution No: 51180060. Specifically, the research received financial support from the DRGT 377 Project (Postdoctoral award to DMK) of SIDA Phase IV in the Makerere-Swedish Bilateral Programme (2015-2020). R.O received a partial Master\u0026rsquo;s scholarship from the Africa Center of Excellence in Materials, Product Development and Nanotechnology (MAPRONANO ACE), CEDAT-Makerere University.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eRO, RKB and DMK performed the following: Conceptualization, methodology, validation, formal analysis, investigation, data curation, writing \u0026ndash; original draft, writing - review and editing. RO conducted the laboratory analysis as a research project for his Master of Science in Biochemistry degree of Makerere University. RKB and DMK supervised the study.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank the Ugandan blood donors who save endless lives through their donated blood and the Uganda Blood Transfusion Services (Nakasero Blood Bank) for granting us access to blood samples for use in this study.\u003cstrong\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Information\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003csup\u003e1\u003c/sup\u003eDepartment of Biochemistry and Systems Biology, College of Natural Sciences, Makerere University. P.O. BOX 7062 Kampala, Uganda\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003csup\u003e2\u003c/sup\u003e\u003c/strong\u003eCentral Diagnostic Laboratory, College of Veterinary medicine, Animal resources and Biosecurity, Makerere University. P.O. BOX 7062 Kampala, Uganda\u003c/p\u003e\n\u003cp\u003e\u003csup\u003e3\u0026nbsp;\u003c/sup\u003eDepartment of Agricultural Sciences, University of Helsinki, P.O. Box 28, FI-00014, Helsinki, Finland\u003c/p\u003e\n"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eWorld Health Organization. Global status report on alcohol and health 2018. David Bramley, editor. 2018. \u003c/li\u003e\n\u003cli\u003eAgarwal DP. Genetic polymorphisms of alcohol metabolizing enzymes. Pathol Biol. 2001;49:703\u0026ndash;9. \u003c/li\u003e\n\u003cli\u003eNorberg A, Jones AW, Hahn RG, Gabrielsson JL. Role of variability in explaining ethanol pharmacokinetics: Research and forensic applications. Clin Pharmacokinet. 2003;42:1\u0026ndash;31. \u003c/li\u003e\n\u003cli\u003eVaswani M, Prasad P, Kapur S. Association of ADH1B and ALDH2 gene polymorphisms with alcohol dependence: a pilot study from India. Hum Genomics. 2009;3:213\u0026ndash;20. \u003c/li\u003e\n\u003cli\u003eSeitz HK, Becker P. Alcohol metabolism and cancer risk. Alcohol Res Heal. 2007;30:38\u0026ndash;47. \u003c/li\u003e\n\u003cli\u003eZhakari S. Overview: How Is Alcohol Metabolised by the Body? Alcohol Res Heal. 2006;29:245\u0026ndash;54. \u003c/li\u003e\n\u003cli\u003eGemma S, Vichi S, Testai E. Individual susceptibility and alcohol effects:biochemical and genetic aspects. Ann Ist Super Sanita. 2006;42:8\u0026ndash;16. \u003c/li\u003e\n\u003cli\u003eEdenberg HJ. The genetics of alcohol metabolism: Role of alcohol dehydrogenase and aldehyde dehydrogenase variants. Alcohol Res Heal. 2007;30:5\u0026ndash;13. \u003c/li\u003e\n\u003cli\u003eHurley TD, Edenberg H. Gene encoding enzymes in alcohol metabolism. Alcohol Res. 2012;34:339\u0026ndash;44. \u003c/li\u003e\n\u003cli\u003eOsier M V., Pakstis AJ, Soodyall H, Comas D, Goldman D, Odunsi A, et al. A Global Perspective on Genetic Variation at the ADH Genes Reveals Unusual Patterns of Linkage Disequilibrium and Diversity. Am J Hum Genet. 2002;71:84\u0026ndash;99. \u003c/li\u003e\n\u003cli\u003eHarada S, Agarwal DP, Goedde HW. Aldehyde dehydrogenase polymorphism and alcohol metabolism in alcoholics. Alcohol. 1985;2:391\u0026ndash;2. \u003c/li\u003e\n\u003cli\u003eVasiliou V, Pappa A, Petersen DR. Role of aldehyde dehydrogenases in endogenous and xenobiotic metabolism. Chem Biol Interact. 2000;129:1\u0026ndash;19. \u003c/li\u003e\n\u003cli\u003eRamchandani VA, Bosron WF, Li TK. Research advances in ethanol metabolism. Pathol Biol. 2001;49:676\u0026ndash;82. \u003c/li\u003e\n\u003cli\u003eUganda Bureau of Statistics. National Population and Housing Census 2014 - Main Report, kampala. Uganda. Uganda Bur Stat. 2016;1\u0026ndash;105. \u003c/li\u003e\n\u003cli\u003eMadhad VJ, Sentheil KP. The Rapid \u0026amp; Non-Enzymatic isolation of DNA from the Human peripheral whole blood suitable for Genotyping. Eur J Biotechnol Biosci. 2014;1:1\u0026ndash;16. \u003c/li\u003e\n\u003cli\u003eWolf JM, Simon D, B\u0026eacute;ria JU, Tietzmann DC, Stein AT, Lunge VR. Analysis of the Association of Nonsynonymous Polymorphisms in ADH Genes with Hazardous Drinking in HIV-1-Positive Individuals. Alcohol Clin Exp Res. 2017;41:1866\u0026ndash;74. \u003c/li\u003e\n\u003cli\u003eXu Y, Carr LG, Bosron WF, Li TK, Edenberg HJ. Genotyping of human alcohol dehydrogenases at the ADH2 and ADH3 loci following DNA sequence amplification. Genomics. 1988;2:209\u0026ndash;14. \u003c/li\u003e\n\u003cli\u003eKayaalti Z, S\u0026ouml;ylemezoĝlu T. Distribution of ADH1B, ALDH2, CYP2E1*6, and CYP2E1*7B genotypes in Turkish population. Alcohol. 2010;44:415\u0026ndash;23. \u003c/li\u003e\n\u003cli\u003eTamakoshi A, Hamajima N, Kawase H, Wakai K, Katsuda N, Saito T, et al. Duplex polymerase chain reaction with confronting two-pair primers (PCR-CTPP) for genotyping alcohol dehydrogenase \u0026beta; subunit (ADH2) and aldehyde dehydrogenase 2 (ALDH2). Alcohol Alcohol. 2003;38:407\u0026ndash;10. \u003c/li\u003e\n\u003cli\u003eShi YY, He L. SHEsis, a powerful software platform for analyses of linkage disequilibrium, haplotype construction, and genetic association at polymorphism loci. Cell Res. 2005;15:97\u0026ndash;8. \u003c/li\u003e\n\u003cli\u003eZhiqiang Li, Zhao Zhang, Zangdong He, Wei Tang, Tao Li, Zhen Zeng, Lin He YS. A partition-ligation-combination-subdivision EM algorithm for haplotype inference with multiallelic markers : update of the SHEsis ( http://analysis.bio-x.cn). Cell Res. 2009;519\u0026ndash;23. \u003c/li\u003e\n\u003cli\u003eZhang J, Jin T, Yunus Z, Li X, Geng T, Wang H, et al. Genetic polymorphisms of VIP variants in the Tajik ethnic group of northwest China. BMC Genet. 2014;15:1\u0026ndash;9. \u003c/li\u003e\n\u003cli\u003eFang FZ, Hou L, Terry MB, Lissowska J, Morabia A, Chen J, et al. Genetic polymorphisms in alcohol metabolism, alcohol intake and the risk of stomach cancer in Warsaw, Poland. Int J Cancer. 2007;121:2060\u0026ndash;4. \u003c/li\u003e\n\u003cli\u003eJim\u0026eacute;nez-Jim\u0026eacute;nez FJ, G\u0026oacute;mez-Tabales J, Alonso-Navarro H, Zurdo M, Turp\u0026iacute;n-Fenoll L, Mill\u0026aacute;n-Pascual J, et al. Association between the rs1229984 polymorphism in the alcohol dehydrogenase 1B gene and risk for restless legs syndrome. Sleep. 2017;40. \u003c/li\u003e\n\u003cli\u003eT\u0026oacute;th R, Fiatal S, Petrovski B, McKee M, \u0026Aacute;d\u0026aacute;ny R. Combined effect of ADH1B rs1229984, rs2066702 and ADH1C rs1693482/rs698 alleles on alcoholism and chronic liver diseases. Dis Markers. 2011;31:267\u0026ndash;77. \u003c/li\u003e\n\u003cli\u003eKumar Dutta A. Genetic factors affecting susceptibility to alcoholic liver disease in an Indian population. Ann Hepatol [Internet]. 2013;12:901\u0026ndash;7. Available from: http://dx.doi.org/10.1016/S1665-2681(19)31295-5\u003c/li\u003e\n\u003cli\u003eMart\u0026iacute;nez C, Galv\u0026aacute;n S, Garcia-Martin E, Ramos MI, Guti\u0026eacute;rrez-Mart\u0026iacute;n Y, Ag\u0026uacute;ndez JAG. Variability in ethanol biodisposition in whites is modulated by polymorphisms in the ADH1B and ADH1C genes. Hepatology. 2010;51:491\u0026ndash;500. \u003c/li\u003e\n\u003cli\u003eCichoz-Lach H, Celinski K, Slomka M. Alcohol-metabolizing enzyme gene polymorphisms and alcohol chronic pancreatitis among Polish individuals. Hpb. 2008;10:138\u0026ndash;43. \u003c/li\u003e\n\u003cli\u003eDjouss\u0026eacute; L, Levy D, Herbert AG, Wilson PWF, D\u0026rsquo;Agostino RB, Cupples LA, et al. Influence of alcohol dehydrogenase 1C polymorphism on the alcohol-cardiovascular disease association (from the Framingham Offspring Study). Am J Cardiol. 2005;96:227\u0026ndash;32. \u003c/li\u003e\n\u003cli\u003eYates AD, Achuthan P, Akanni W, Allen J, Allen J, Alvarez-Jarreta J, et al. Ensembl 2020. Nucleic Acids Res. 2020;48:D682\u0026ndash;8. \u003c/li\u003e\n\u003cli\u003eAvin\u0026ccedil;sal MO, Shinomiya H, Teshima M, Kubo M, Otsuki N, Kyota N, et al. Impact of alcohol dehydrogenase-aldehyde dehydrogenase polymorphism on clinical outcome in patients with hypopharyngeal cancer. Head Neck. 2018;40:770\u0026ndash;7. \u003c/li\u003e\n\u003cli\u003eChien HT, Young CK, Chen TP, Liao CT, Wang HM, Cheng S De, et al. Alcohol-metabolizing enzymes\u0026rsquo; gene polymorphisms and susceptibility to multiple head and neck cancers. Cancer Prev Res. 2019;12:247\u0026ndash;54. \u003c/li\u003e\n\u003cli\u003eHub\u0026aacute;ček JA, \u0026Scaron;edov\u0026aacute; L, Oli\u0026scaron;arov\u0026aacute; V, Ad\u0026aacute;mkov\u0026aacute; V, Ad\u0026aacute;mek V, T\u0026oacute;thov\u0026aacute; V. Distribution of adh1b genotypes predisposed to enhanced alcohol consumption in the Czech roma/gypsy population. Cent Eur J Public Health. 2018;26:284\u0026ndash;8. \u003c/li\u003e\n\u003cli\u003eWang Y, Du F, Zhao H, Yu X, Liu J, Xiao Y, et al. Synergistic association between two alcohol metabolism relevant genes and coronary artery disease among Chinese hypertensive patients. PLoS One. 2014;9:1\u0026ndash;7. \u003c/li\u003e\n\u003cli\u003eChao Y ‐C, Liou S ‐R, Chung Y ‐Y, Tang H ‐S, Hsu C ‐T, Li T ‐K, et al. Polymorphism of alcohol and aldehyde dehydrogenase genes and alcoholic cirrhosis in chinese patients. Hepatology. 1994;19:360\u0026ndash;6. \u003c/li\u003e\n\u003cli\u003eNagata K, Yamazoe Y. Genetic Polymorphism of Human Cytochrome P450 Involved in Drug Metabolism. Drug Metab Pharmacokinet. 2002;17:167\u0026ndash;89. \u003c/li\u003e\n\u003cli\u003eMatsuo K, Hamajima N, Shinoda M, Hatooka S, Inoue M, Takezaki T, et al. Erratum: Gene-environment interaction between an aldehyde dehydrogenais-2 (ALDH2) polymorphism and alcohol consumption for the risk of esophageal cancer (Carcinogenesis (2001) vol. 22 (913-916)). Carcinogenesis. 2001. p. 1893. \u003c/li\u003e\n\u003cli\u003eYokoyama A, Muramatsu T, Omori T, Matsushita S, Yoshimizu H, Higuchi S, et al. Alcohol and aldehyde dehydrogenase gene polymorphisms influence susceptibility to esophageal cancer in Japanese alcoholics. Alcohol Clin Exp Res. 1999;23:1705\u0026ndash;10. \u003c/li\u003e\n\u003cli\u003eEdenberg HJ, Xuei X, Chen HJ, Tian H, Wetherill LF, Dick DM, et al. Association of alcohol dehydrogenase genes with alcohol dependence: A comprehensive analysis. Hum Mol Genet. 2006;15:1539\u0026ndash;49. \u003c/li\u003e\n\u003cli\u003eWhitfield JB. Acute reactions to alcohol. Addict Biol. 1997;2:377\u0026ndash;86. \u003c/li\u003e\n\u003cli\u003eGurdasani D, Carstensen T, Fatumo S, Chen G, Franklin CS, Prado-Martinez J, et al. Uganda Genome Resource Enables Insights into Population History and Genomic Discovery in Africa. Cell. 2019;179:984-1002.e36. \u003c/li\u003e\n\u003cli\u003eMohammed F. Hassan, Amanda Elswick Gentry, Elizabeth C. Prom-Wormley, Roseann E. Peterson BTW. Machine Learning Approaches to Predict Alcohol Consumption from Biomarkers in the UK Biobank. medRxiv 2024122224319486; doi https//doi.org/101101/2024122224319486. 2024;1\u0026ndash;20. \u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-research-notes","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"resn","sideBox":"Learn more about [BMC Research Notes](http://bmcresnotes.biomedcentral.com)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/resn/default.aspx","title":"BMC Research Notes","twitterHandle":"@BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"alcohol, ADH1B, ADH1C, ALDH2, haplotype, PCR, SNP, Uganda","lastPublishedDoi":"10.21203/rs.3.rs-5955120/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5955120/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003eAlcohol is metabolized to acetaldehyde by alcohol dehydrogenases (ADH) and subsequently to acetate by aldehyde dehydrogenases (ALDH). Single nucleotide polymorphisms (SNPs) in \u003cem\u003eADH1B, ADH1C\u003c/em\u003e and \u003cem\u003eALDH2\u003c/em\u003e genes lead to haplotypes encoding isozymes which influence development of alcoholism. The distribution of these haplotypes in Uganda has not been documented. The aim of this study was to determine genotype, allele, and haplotype frequencies of SNPs in \u003cem\u003eADH1B, ADH1C\u003c/em\u003e, and \u003cem\u003eALDH2\u003c/em\u003e genes in Uganda.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eFive SNPs: \u003cem\u003eADH1B\u003c/em\u003e (rs1229984 and rs2066702), \u003cem\u003eADH1C\u003c/em\u003e (rs1693482 and rs698) and \u003cem\u003eALDH2\u003c/em\u003e (rs671) were analyzed by PCR-restriction fragment length polymorphism assays in 250 samples. The frequencies of the fast-metabolizing alleles \u003cem\u003eADH1C*1\u003c/em\u003e, \u003cem\u003eADH1B*3\u003c/em\u003e, and \u003cem\u003eADH1B*2\u003c/em\u003e were 49.6%, 18.2% and 0.2% respectively. The nonprotective haplotype \u003cem\u003eADH1B\u003c/em\u003e*1 had a high frequency of 81.6% and \u003cem\u003eADH1C*2\u003c/em\u003e was 10.6%. A novel allele \u003cem\u003eADH1C*new\u003c/em\u003e comprising G (Codon 349 Val) at \u003cem\u003eADH1C rs698\u003c/em\u003e and G (Codon 271 Arg) at \u003cem\u003eADH1C rs1693482\u003c/em\u003e was identified with a frequency of 39.8%. Of the seven ADH haplotype combinations identified, \u003cem\u003eADH1B\u003c/em\u003e*1-\u003cem\u003eADH1C\u003c/em\u003e*1 was the most prevalent (48.4%). Notably \u003cem\u003eADH1B*1\u0026ndash;ADH1C* new\u003c/em\u003e, had the second highest frequency (25.2%).\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eOur study provides the first data on novel \u003cem\u003eADH1B-ADH1C\u003c/em\u003e haplotypes in alcohol metabolizing genes in the Ugandan population.\u003c/p\u003e","manuscriptTitle":"Novel Haplotypes of genetic polymorphisms in alcohol metabolizing enzymes in Kampala, Uganda","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-14 16:36:26","doi":"10.21203/rs.3.rs-5955120/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-04-23T09:02:21+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-04-23T06:33:53+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"103356128911830870541133003830258671467","date":"2025-04-18T05:30:02+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-04-15T05:04:01+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"53696378682896298056054980825280230973","date":"2025-04-13T02:55:36+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"180587170332839588641944006782516036976","date":"2025-04-11T08:44:53+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-04-11T08:43:26+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-04-11T01:57:45+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Research Notes","date":"2025-04-04T05:22:10+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-research-notes","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"resn","sideBox":"Learn more about [BMC Research Notes](http://bmcresnotes.biomedcentral.com)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/resn/default.aspx","title":"BMC Research Notes","twitterHandle":"@BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"07058eaa-a6b3-42a5-9d47-08ba9a8c75a5","owner":[],"postedDate":"April 14th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-07-07T16:20:56+00:00","versionOfRecord":{"articleIdentity":"rs-5955120","link":"https://doi.org/10.1186/s13104-025-07331-y","journal":{"identity":"bmc-research-notes","isVorOnly":false,"title":"BMC Research Notes"},"publishedOn":"2025-07-01 15:57:24","publishedOnDateReadable":"July 1st, 2025"},"versionCreatedAt":"2025-04-14 16:36:26","video":"","vorDoi":"10.1186/s13104-025-07331-y","vorDoiUrl":"https://doi.org/10.1186/s13104-025-07331-y","workflowStages":[]},"version":"v1","identity":"rs-5955120","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5955120","identity":"rs-5955120","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}