Genetics & Applications

Few case studies as ideas for zero-waste from food production and processing

Zoran Popovski — 2023-12-20

Globally, the amount of agricultural waste is huge but not properly utilized yet. Precisely, about one billion tons of food produced for human consumption is wasted each year. This wastage global economy costs are estimated at US$ 1 trillion on annual level. The definition of food waste varies globally depending on where food waste occurs in the food supply and consumption chain, how it is generated, and what it covers. This review is a compilation of few different approaches in the context of zero waste from food production and processing. Soybean hull exploitation due to mass production of tofu, soymilk, edamame, tofu sausages and burgers, soy breads, soy pasta and soymilk yoghurts and cheeses, animal feed leads to a formation of high waste load. There are several available ways for soybean hull valorization: as a biofertilizer, as a substrate for microbial growth, as an adsorbent, for extraction of antioxidants and in our case for extraction of enzymes. The global consumption of coffee is approximately 10 million tons, resulting in a substantial production of spent coffee grounds (SCG) worldwide. However, this by-product has recently gained attention as a valuable source of usable compounds, particularly in our case for essential oils. During the cheese production process, the whey obtained as a by-product can be used as a raw material for development of new dairy products and in the same time to prevent the potential pollution of natural watercourses. The valorisation of the waste whey obtained in the cheese production by using it in the development of new products also contributes in the environment protection. In N. Macedonia, about 1000 tons of blood waste from industrial slaughterhouses flow into natural watercourses every year and it can find application in various industries. These are just a few examples that show that the possibilities for the implementation of the zero-waste concept are unlimited.

Association between insertion/deletion polymorphism of the ace gene with risk of hypertension

Jovan Kulić — 2023-12-07

This study conducted an initial investigation into the association between ACE gene insertion/deletion (I/D) polymorphisms (rs1799752) and hypertension in the Republic of Srpska, Bosnia and Herzegovina. The study featured two distinct groups, each with 100 subjects, systematically categorized based on hypertension status and gender. DNA was extracted, PCR-amplified, and analyzed by gel electrophoresis. Results revealed a higher prevalence of the DD genotype and the D allele in the hypertensive group, although statistical significance was not observed. The II genotype occurred in 18% of the hypertension group and 21% in the control group. A significant difference was found in allele I frequencies between the two groups (p=0.004), with no gender-related variations in ACE alleles. The limited sample size may have constrained the ability to detect statistically significant differences. The odds ratio for the (DD + ID) genotype compared to II was 1.2110 (95% CI: 0.6006 to 2.4418; p=0.5927), indicating no statistical significance. Furthermore, no significant associations were identified between ACE genotypes and alleles and gender. In summary, this preliminary study suggests a potential trend towards a higher prevalence of the ACE gene D allele and DD genotype in hypertensive individuals. However, due to the small sample size, these associations did not achieve statistical significance in this population. Larger-scale investigations are needed for more definitive insights into the relationship between ACE gene polymorphisms and hypertension.

Does the geographic distance affect the genetic differentiation among bilberry populations sampled in Bosnia and Herzegovina?

Adnan Hodzic — 2023-12-08

Isolation-by-distance (IBD) pattern among bilberry (Vaccinium myrtillus L.) populations has previously been reported for this species in northern Europe. However, the number of molecular studies conducted on bilberry, using everything from isoenzymes, RAPDs to microsatellite markers, are very few and far between. Considering that Bosnia and Herzegovina (B&H) is a country rich with diverse fruit genetic resources, conducting a genetic characterization of the naturally occurring V. myrtillus populations could yield valuable data for the conservation and utilization of this resource. This study entailed genotyping samples collected from three bilberry populations located in Fojnica, Kladanj, and Srebrenica municipalities using seven polymorphic microsatellite or SSR (simple sequence repeats) markers. The obtained molecular data was used to calculate the correlation between the physical distance of the individual B&H populations and a parameter of the genetic differentiation (pairwise Fst). The results of the correlation analyses revealed an absence of a significant isolation-by-distance pattern among the three examined B&H bilberry populations. In addition, the most pronounced genetic differentiation was detected between the Srebrenica and each of the two remaining B&H populations. At the same time, the values for pFst were significant, albeit much lower, between the Fojnica and Kladanj populations. Bilberries from the sampled Srebrenica population appear to be distinct from the other B&H populations, possibly due to the different genetic origin of this population.

Automated DNA extraction from recent and archaeological human skeletal remains using QIAGEN's EZ2 Connect Fx Instrument – an experience report

Esma Fočak — 2023-12-13

The aim of this study was to investigate the efficiency of the automated DNA extraction method using the EZ2 Connect Fx instrument (QIAGEN), showcasing its ability to achieve swift and reliable results from bone samples. Genomic DNA extraction from 16 archaelogical and recent bone samples was performed with a commercial EZ1&2^® DNA Investigator^® Kit (QIAGEN). Prior to PCR amplification, DNA concentration was quantified using Qubit™ Fluorometer (Thermo Fisher Scientific). Amplification was carried out with the Investigator 24plex Kit (QIAGEN). DNA profiles were generated using 3500 Genetic Analyzer (Applied Biosystems) and analysed with GeneMapper™ ID-X 1.6 Software (Applied Biosystems). For nine archaeological dental samples, eight exhibited partial profiles, and one showed no amplified loci. Among the seven recent femoral bone samples, one exhibited a partial profile, while six presented complete DNA profiles. In this study, we investigated the benefits of automated DNA extraction, particularly its ability to generate timely and reliable results. Additionally, this method reduces pipette and tip usage, almost completely minimising the risk of human error, exogenous DNA contamination and cross-contamination. Such efficiency is of paramount importance, especially when working with skeletal remains, namely archaeological skeletal remains that pose a challenge for molecular genetic analyses.

Odontological analysis of medieval archaeological samples from Krešića greblje near Čitluk, Bosnia and Herzegovina - An auxiliary method to genetic analysis

Selma Zukić — 2023-12-18

Previous research on archaeological skeletal remains in Bosnia and Herzegovina rarely included odontological analysis. The objective of this research was biological profiling through odontological analysis (dental morphological characteristics, estimation of sex and dental age at death) in medieval archaeological skeletal remains samples from Krešića greblje in Paoča (Čitluk municipality, Bosnia and Herzegovina). After the excavations of the archaeological site, odontological analysis was performed before genetic analysis. The sample comprised 22 permanent teeth from 6 individuals. Odontological analysis included odontometric estimation of sex, estimation of age at death by two morphological methods, and an analysis of dental traits and dental pathology. Genetic analysis was used for sex determination. The results of odontological analysis classified all individuals as adults, with estimated age at death 20-25 in one individual, 30-35 in three individuals, 35-45 in one individual, and one individual older than 45 years of age. Odontometric sex estimation was in high concordance with sex determination by genetic analysis, and correctly classified five individuals’ sex (four females and one male). In one individual odontometric sex was assessed as female while genetic analysis revealed male. Morphological dental traits and dental pathology were investigated as well, adding to a better understanding of the odontological characteristics of the medieval population in the area of Bosnia and Herzegovina. Sex and dental age estimation given by odontological analysis provide fast and cost-effective information, and should be performed as an auxiliary before genetic analysis.

Presence of HLA-DQ2 and HLA-DQ8 /DR4 celiac disease predisposing alleles in tested group of patients in Bosnia and Herzegovina

Tea Bećirević — 2023-12-18

Celiac disease (CD) is an autoimmune disease characterized by gluten intolerance. The main cause of this immune - mediated enteropathy is gluten, a protein mainly present in wheat, rye, barley and spelt. The predisposition to celiac disease is determined by HLA class II genes encoding MHC II heterodimer molecules, more specifically HLA-DQ2 and HLA-DQ8. Up to date the exact number of people suffering from celiac disease in Bosnia and Herzegovina is still not known because there is no public registry for this disease. The aim of this study was to evaluate the HLA-DQ2 and HLA-DQ8/DR4 presence in tested group of patients (n = 23) referred to Polyclinic Atrijum in Sarajevo in period from August 2022. to August 2023. HLA-DQ2 and DQ8/DR4 allele identification was performed using Real-time PCR technique. According to the obtained results a total of 26% (n=6) of tested patients were positive for HLA-DQ2 and HLA-DQ8/DR4 alleles. Two of the patients were positive for HLA-DQ2 alleles and four patients were positive for HLA-DQ8/DR4 alleles. To our knowledge this is the first study evaluating the presence of HLA-DQ2 and HLA-DQ8 genotypes in tested population in Bosnia and Herzegovina.

A retrospective analysis of spontaneous chromosomal aberrations in human lymphocyte cultures of individuals from Bosnia and Herzegovina

Sanin Haverić — 2023-12-19

Spontaneous chromosomal aberrations are structural or numerical changes of chromosomes that occur naturally, without exposure to external genotoxic factors. They are not inherited, occur randomly in the karyotype, and do not have direct clinical significance. However, they can affect genomic instability and disease predisposition. They can result from DNA replication or repair processes errors, and typically are observed in cells that are actively dividing. Spontaneous chromosomal aberrations may arise due to the natural chromosomal instability and can be elevated in individuals exposed to mutagens. We analyzed frequencies of spontaneous chromosomal aberrations in 137 individuals subjected to karyotype analysis at the Laboratory for Cytogenetics and Genotoxicology, University of Sarajevo – Institute for Genetic Engineering and Biotechnology, during 2008-2023. Whole blood samples were cultivated for 72 hours with the thymidine added in the 48th hour. Metaphases were arrested by colcemid 60 minutes before harvesting. GTG banding was performed and slides were analyzed under 1000x magnification in accordance with An International System for Human Cytogenetic Nomenclature and E.C.A. Cytogenetic Guidelines and Quality Assurance. Constitutionally aberrant karyotypes were found in 2.92% of analysed individuals as well as altered karyotypes considered as normal chromosomal variants. In the total of 3092 analyzed metaphases, 20 spontaneous chromosomal aberrations were found in 13 individuals. This study contributes to the limited knowledge of the cytogenetic status of the Bosnian and Herzegovinian population. Further monitoring of spontaneous chromosomal aberrations incidences is recommended.

Highly Sensitive Detection of DNA Fragments by an Inexpensive and Simple Method Fabricated In-House Blue LED Transilluminator

Ryosuke Ishii — 2023-11-10

Agarose gel electrophoresis is a routinely employed experimental technique for visualizing nucleic acids, typically combined with ethidium bromide or some other molecular dye staining and a UV transilluminator. However, the utilization of a UV transilluminator is restricted due to high cost and safety considerations. In this study, an inexpensive and simple in-house blue Light Emitting Diode (LED) transilluminator was developed using a high-brightness blue LED, carbon resistor, and smartphone charging cable. Its sensitivity for DNA detection was compared with conventional methods. A detection limit of 4 ng/band was confirmed by observing a 100 bp DNA fragment, which is comparable to ethidium bromide staining and UV transilluminator. Additionally, the in-house blue LED transilluminator can be attached to existing electrophoresis chambers, allowing real-time observation of electrophoresis and offering a superior alternative to conventional methods. In conclusion, the developed in-house blue LED transilluminator demonstrates high sensitivity in detecting DNA fragments and holds potential for advancing low-cost research facilities.