Biotechnology

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Browsing Biotechnology by Author "Alemayehu Godana"

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    Antimicrobial Resistance Profile of Escherichia Coli Isolated from Hospital Sewage and Polluted River: the Cases of Adama Hospital Medical College in Adama and Yerer River in Dukem
    (Addis Ababa University, 2024-11) Kajelcha Fikadu; Alemayehu Godana
    The antimicrobial resistance profile of Escherichia coli is dramatically increasing across the world, particularly in low- and middle-income countries where untreated wastewater can disseminate resistant bacteria into the environment. Untreated wastewater discharged from hospitals and industries is the most known vehicle for the emergence and spread of antimicrobial resistance genes. The study aimed to evaluate the antimicrobial resistance profile of E. coli isolates from hospital sewage and polluted river samples in Adama and Dukem, respectively. The hospital sewage and polluted water samples were collected from Adama Hospital Medical College and two rivers in Dukem. The samples were transported to Addis Ababa University, Institute of Biotechnology Laboratory. For the purpose of isolating E. coli, the samples were cultivated on Eosin Methylene Blue agar, MacKonkey agar, and Nutrient agar media. Biochemical tests, including Gram staining, IMViC, TSI, and SCA, were used for the identification of E. coli isolates. The researchers evaluated the antimicrobial susceptibility patterns of the E. coli isolates using the Kirby-Bauer’s disc diffusion method. The identified E. coli isolates were then cultured on Mueller Hinton agar media at 37°C for 24 hours, after which the inhibition zone was measured by a digital ruler. Polymerase chain reaction (PCR) was used to confirm the presence of resistance-associated genes in the E. coli isolates. A total of 75 presumed E. coli isolates were obtained via culture. Among these isolates, 50 isolates were subjected to AST. The isolates from Yerer River before the industrial waste entry site (YRBI) did not show multiple antimicrobial resistance (MAR), while the isolates from Xadacha River (XR) and Yerer River After Industrial waste entry site (YRAI) showed resistance profiles of 66.67% and 90%, respectively. This study also revealed that 86.67% of hospital sewage isolates and 65% of river water isolates showed MAR. PCR amplification confirmed the presence of tetA and blaTEM genes in 83.33% and 57.14% of the AMR isolates, respectively. In conclusion, the study showed that untreated hospital sewage and pollute river water are considered major reservoirs for the emergence of antimicrobial-resistant E. coli among humans, animals, and the environment. The study advocates for improved wastewater treatment, stringent regulations on antimicrobial use, and regular monitoring to curb the emergence and spread of antimicrobial resistance.
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    Chloroquine Resistance Transporter (Pfcrt) and Multidrug Resistance 1 (Pfmdr1) Genes Mutation in Plasmodium Falciparum Population Under Varying Level of Endemicity with Plasmodium Vivax in Selected Parts of Ethiopia
    (Addis Ababa University, 2023-02) Tajudin Abdurahman; Alemayehu Godana
    The global controlling of Plasmodium falciparum infections faces significant challenges due to the spread of parasites resistant to antimalarial drugs. In Ethiopia, where both P. vivax and P. falciparum coexist, the treatment for uncomplicated falciparum malaria shifted from chloroquine (CQ) to sulfadoxine-pyrimethamine (SP) in 1998 and then to Coartem (artemether-lumefantrine (AL)) in 2004. AL has been the standard treatment for over two decades for P. falciparum, while P. vivax is still treated with CQ. The coexistence of these two species and the accessibility of CQ for P. vivax treatment raise questions about whether switching from CQ to AL for P. falciparum treatment might lead to the resurgence of CQ -susceptible P. falciparum strains due to reversal mutations or the efficacy of AL. The sudy aimed to assess the prevalence of pfcrt-76 and pfmdr1-86 gene mutations in the P. falciparum population under varying levels of endemicity with P. vivax in selected parts of Ethiopia. In this study the frequency of gene mutations of P. falciparum chloroquine resistance transporter 76 (pfcrt-76) and P. falciparum multidrug resistance 1-86 (pfmdr1-86) in P. falciparum collected from malaria-infected patients in Abobo, Dera, Fentale, and Metema districts, Ethiopia, were examined. Confirmed P. falciparum samples (n = 258) with microscope were collected through health facility-targeted cross-sectional surveys in areas with varying levels of P. falciparum and P.vivax prevalence. Genomic DNA was extracted from dried blood spots by using MagMAX DNA Multi-Sample Kit, operated by the Kingfisher Flex Automated Extractor. An 18S rRNA gene-based multiplex real-time PCR assay was employed to confirm P. falciparum mono-infection samples (n = 258). The study analyzed 258 P. falciparum-infected patients using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), examining Pfmdr1-86 and Pfcrt-76 gene mutations. Fisher's exact test determined marker distribution significance. Out of 250 genotyped for Pfcrt K76T, 68.8% had mutant 76T, and 31.2% were wild-type K76. For 257 genotyped for Pfmdr1-N86Y, 98.44% was N86 wild-type, and 1.56% was 86Y mutants. The mutant Pfcrt-76T was more prevalent in areas with higher P. vivax endemicity, including 93.33% (Fentale), 84.71% (Dera), 57.5% (Metema), and 43.75% (Abobo). Pfcrt-76 gene single nucleotide polymorphisms (SNPs) significantly correlated with P. vivax endemicity (P = 0.000). Pfmdr1-N86 was predominantly 100% except in Abobo, where 95.18% were N86, and 4.81% were 86Y. Pfmdr1 gene SNPs were significantly associated with P. vivax endemicity (P = 0.025). Despite CQ discontinuation for over two decades in Ethiopia, a substantial proportion of P. falciparum isolates still carry mutant 76T genotypes, indicating latent CQ pressure. The use of AL for uncomplicated P. falciparum malaria may lead to the return of Pfmdr1 N86 wild-type genes. Further molecular epidemiological investigations in varied endemic regions with different CQ usage histories are recommended to understand chloroquine (CQ) susceptibility recovery and AL therapy efficacy.
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    Plasmodium Vivax Duffy Binding Protein Copy Number Variation and their Effects on Reticulocyte Invasion in Selected Parts of Ethiopia
    (Addis Ababa University, 2024-01) Yasin Nasir; Alemayehu Godana; Fitsum Girma
    The connection between the Duffy binding protein and the Duffy antigen receptor for chemokine is required for Plasmodium vivax penetration into human reticulocytes. A previous analysis of Plasmodium vivax samples in Ethiopia determined that Duffy binding protein duplications are more prevalent than in any other Plasmodium vivax location. However, its prevalence and importance in large samples remain unclear. Duffy blood group genotyping was done by amplifying the GATA1 transcription factor-binding region of DARC geneamong 349 Plasmodium vivax isolates were determined by a real-time PCR technology. In addition,duplications of Duffy binding protein and their relationship with Duffy-negativity and parasite densitywere examined. Duffy binding protein duplications and Duffy-negative antigens were detected in 74% and 3.2% isolates respectively. Most of the Duffy negative participants were found in Gondar Zuria district (72.7%). To know whether Duffy binding protein amplification contributes to the takeover of Duffy-negative reticulocytes, the relationship between Duffy binding protein copy numbers and Duffy status was investigated using Fisher's exact test and post-hoc tests. Fisher's exact test reveals a significant association (p = 0.000313). In addition, a post-hoc test indicates a significant association between the Duffy binding protein copy numbers (single and 2-3 copy) and the DARC status (p = 0.000281). However, the number of Duffy-negative patients infected by multiple Duffy binding protein copy is small. Therefore, we cannot decide that multiple Duffy binding protein copy number increase parasite's invading ability against Duffy-negative patients. The mean parasite burden differs between Duffy negative and positive in a statistically significant way (P <0.0001). Duffy-negative patients aren't resistant to Plasmodium vivax however; the detailed mechanisms of the infection in Duffy-negative patients remain unclear. In reference to the high rate of Duffy-binding protein duplication, further investigation should be explored by extending study site across Ethiopia by using the most sensitive molecular detection tools known as digital PCR. It is also useful to look into additional parasite ligands related to the invasion of Duffy-negative reticulocytes.