Open Access Original Research Article

Physico-chemical and Bacteriological Characteristics of the Drinking Water of the Aguégués Lake District in the Lower Ouémé Valley - Benin

Toussaint Cokou Dan, Rodrigue C. Landeou, Boko Dominique, Thierry Azonhe, Bénoît N’ Bessa

Microbiology Research Journal International, Page 1-14
DOI: 10.9734/mrji/2020/v30i830246

Water is an essential resource for life on earth and is the source of several microbiological and toxicological diseases. The Aguégués one of the communes of the lower valley of Ouémé-Benin are very exposed to these type of diseases. Pathogens swarm in a highly polluted environment because of the peculiarity of this predominantly aquatic area. This is what justifies this research, which aims to assess the quality of drinking water for the populations of the Commune des Aguégués. The research technique focused on the field survey which made it possible to identify the different sources of water supply used by the population. Likewise, the physico-chemical and bacteriological quality of these water sources has been determined. These research techniques have made it possible to discover that the population of Aguégués uses several sources of water. Apart from the recommended sources (National Water Company of Benin and Village Water Supply) the populations use other sources of water. These are rainwater consumed in the rainy season by 97% of the households surveyed, surface water used by 78.26% of the population, especially during flood periods for various uses. Physico-chemical and bacteriological analyzes have shown that apart from SONEB waters only available in the districts of Zoungamey and Houédomey, the other sources of water have parameters which do not comply with WHO standards. This justifies the prevalence of diarrheal diseases in the study environment, which represents the second cause of consultation in health centers.

Open Access Original Research Article

Isolation, Characterization, and Molecular Identification of Indigenous Bacteria from Fermented Almonds (Prunus dulcis)

Salwa Nurhasanah, Edy Fachrial, Nyoman Ehrich Lister

Microbiology Research Journal International, Page 15-22
DOI: 10.9734/mrji/2020/v30i830247

Aims: This study aims to isolate and identify the indigenous bacteria of almonds fermentation.

Methods: Characterization of the indigeneous bacteria are using gram staining, biochemical tests, 16SrRNA gene sequencing, and the antimicrobial activity against Escherichia coli bacteria.

Results: Approximately 28 x 106 CFU / mL bacteria were obtained from almonds fermentations with 14 isolates from enrichment results. Three randomly selected isolates were gram-positive rod-shaped with a negative catalase and positive fermentation test. However, one isolate showed positive results on the motility test. The antimicrobial test results from the three randomly selected isolates using the disk diffusion method obtained inhibition zones of 7 mm, 6.7 mm, and 7 mm, respectively. Therefore, by using 16S rRNA gene sequencing, three different microorganisms were found, namely Bacillus subtilis strain IAM 12118, Bacillus Piscis strain 16MFT21, and Bacillus licheniformis strain BaDB27.

Conclusion: It was found that Bacillus subtilis strain IAM 12118, Bacillus Piscis strain 16MFT21, and Bacillus licheniformis strain BaDB27 in almonds fermentation and also can be used as probiotic bacteria.

Open Access Original Research Article

Detection of Antibiotic Resistance Genes in Some Multiple Antibiotic Resistant E. coli from Apparently Healthy Pregnant Women

O. C. Adekunle, A. J. Falade- Fatila, R. Ojedele, G. Odewale

Microbiology Research Journal International, Page 23-31
DOI: 10.9734/mrji/2020/v30i830248

The emerging drug resistance, especially among the Escherichia coli (E.coli) isolates from pregnant women, spread rapidly within the community. Urinary tract infection (UTI) is a well-known bacterial infection posing serious health problem in pregnant women. Also, multi-drug resistance is becoming rampant, and it is of serious public health concern. Treatment of E. coli is now a challenge due to continuous increase in resistance towards commonly prescribed antibiotics, thus posing a threat to treatment. Hence, the aim of the study is to determine antibiotic resistance genes in some multiple antibiotic resistant E.coli from apparently healthy pregnant women in Osun State. A cross-sectional study design was used to collect 150 mid-stream urine samples from apparently healthy pregnant women from March, 2018 to September, 2018. A well structured questionnaire and informed consent were used for data collection. Standard loop technique was used to place 0.001 ml of urine on Cysteine Lactose Electrolyte Deficient (CLED) medium, Blood agar, MacConkey agar and incubated at 37 °C for 24 h. A standard agar disc diffusion method was used to determine antimicrobial susceptibility pattern of the isolates. The molecular detection of the resistant genes was done using PCR techniques. The ages of women enrolled in this study ranges from 22 to 42 years (mean ± standard deviation = 31 ± 4.7 years). Escherichia coli showed high percentage of resistance to ampicillin and low resistance to ciprofloxacin and penicillin. All the E. coli isolates were sensitive to levofloxacin, and most were resistant to Meropenem. Multiple drug resistance was observed in all the isolates. Resistance genes in VIM 390bp, bla ctx-M 585bp and TEM 517bp were detected in some of the representative E. coli isolates profiled. This study identified the presence of Multi-drug resistance genes in E. coli associated UTI among pregnant women in Osogbo.

Open Access Original Research Article

Analyses of the Spike Proteins of Severe Acute Respiratory Syndrome-Related Coronaviruses

Peramachi Palanivelu

Microbiology Research Journal International, Page 32-50
DOI: 10.9734/mrji/2020/v30i830249

Aim: To analyze spike proteins of Severe Acute Respiratory Syndrome (SARS)-related coronaviruses (CoVs) for their conserved motifs, Receptor-Binding  Domain (RBD), Receptor Binding Motif (RBM) of SARS-CoV (CoV-1), SARS-CoV-2, Middle East Respiratory Syndrome (MERS)-CoV and their relationship to the bat, pangolin and palm civet-CoVs as possible intermediate hosts.

Study Design: Multiple sequence analysis (MSA) of spike proteins of different SARS-CoVs were studied using Clustal Omega and ExPASy tools.

Methodology: Bioinformatics, SDM and X-ray crystallographic data of the spike proteins from different CoVs including the current epidemic causing SARS-CoV-2 were analyzed. The advanced version of Clustal Omega was used for protein sequence analysis of different spike proteins from various CoVs and ExPASy tool was used for pI analysis.

Results: Spike proteins in coronaviruses play important roles in mediating receptor binding, membrane fusion, and viral entry into human cells. Furthermore, nowadays all the vaccine development programmes are mainly focused on the SARS-CoV-2 spike protein only, as it plays the crucial, first step in the infection process. Therefore, the spike proteins of the SARS-related coronaviruses, the main determinant of coronavirus host specificity, are analyzed for their conserved motifs, RBD, RBM, etc. The recent epidemic causing strain, SARS-CoV-2, showed 2 dipeptide deletions and 4 peptide insertions ranging from tetra- to hepta-peptides in its spike protein as compared to its predecessor CoV-1. Most of the insertions are also found in the bat and pangolin CoVs except one unique tetrapeptide. The RBM region shows that the bats, pangolins and CoV-2 exhibit very similar to identical sequences. The overall analyses show that the latest SARS-CoV-2 is related to bats and more to pangolin-CoVs suggesting that the pangolins could be possibly the intermediate host. On the other hand, it is found that palm civet RBM sequences are highly related to CoV-1 and not CoV-2. Possibly the novel CoV-2 would have taken three insertions from bats and/or pangolins and the fourth insertion –PRRA- which is unique to SARS-CoV-2 is critically placed just in the S1/S2 cleavage region. The recently discovered G614 mutation (D614→G) in CoV-2, the most prevalent form in the global pandemic now, is found near the RBD towards the C-terminal. Placement of the unique tetrapeptide in the S1/S2 loop region and replacement with more positive charges on the spike protein which resulted in marked increase in the basicity of the SARS-CoV-2 spike protein may, possibly result in significant effects on the structure and function of the protein, possibly leading to rapid transmission.

Conclusions: RBD and RBM regions of the spike proteins of SARS-CoV-1 and palm civet show very close identity to each other whereas the SARS-CoV-2, pangolin- and bat-CoVs exhibit very close identities in their RBD and RBM regions. The two crucial modifications in the spike protein of SARS-CoV-2, viz. a marked increase in the basicity of the protein and the insertion of a dibasic tetrapeptide (–PRRA-) at the critical S1/S2 cleavage point possibly make it to bind to the ACE2 receptor with higher affinity and get it cleaved by the host proteases more efficiently with subsequent effective internalization of the viral genome.

Open Access Original Research Article

Case Fatality in COVID-19; a District Perspective

Hamzullah Khan, Faridullah Shah, Khalid Khan, . Abuzar

Microbiology Research Journal International, Page 51-58
DOI: 10.9734/mrji/2020/v30i830250

Objectives: To determine the case fatality rate of COVID-19 by evaluating the data of Patients died due to COVID-19.

Materials and Methods: A total of 243 patients with PCR done from the government designated Public health research laboratory of Khyber Medical University were included in a cross sectional comparative study. Chi-square test, risk analysis, probability testing and survival analysis using Kaplan Meir test was done on data sheet prepared in SPPS version 25 in accordance with the objectives of the study.

Results: Out of total 243 patients, 165 (67.90%) were negative by PCR testing and 78 (32.09%) were COVID-19 positive. The Mean age with SD was 36+17 years. Out of total 178 (73.3%) were males and 65 (26.7%) were females. 34 (13.99%) had age more than 55 years, where 4/5 (80%) of the deaths were recorded. The relationship of an increase in age with rate of mortality was statistically significant (p=0.001). The over all probability of death in our population in age more than 55 years is 24 times higher (OR=24,95CI: 2.6-221.24) with relative risk of 1.1 (rr=1.11, 95CI:1-1.24). Mortality rate was 6.41%. A significant correlation of mortality with case positivity (p=0.003) with relative risk of 1.06 (rr=1.06, 95CI: 1.008-1.13). Three out of 5 of the COVID-19 positive deceased had cardiac diseases and 2/5 had respiratory disease/viral pneumonia. Using Kaplan Meir test, the survival graphs of COVID positive vs COVID negative cases had similar pattern that shows the mortality rate in the positive cases was not solely due to COVID-19, but has aggravated the pre-existing illness to cause death.

Conclusion: The mortality rate was 6.41%, more in age>55 years (80%) and almost all the deceased had chronic co-morbid conditions like CAD, CCF and COPD at time of presentation. COVID-19 is not the sole killer, it contributes killing in immune-deficient patients.

Open Access Original Research Article

Distribution of COVID-19 by Sex and Age Groups and Its Association with Travel and Contact History: An Overview of Early Epidemic

Hamzullah Khan, Zahid Khan, Saadullah Afridi, Fazli Bari

Microbiology Research Journal International, Page 59-66
DOI: 10.9734/mrji/2020/v30i830251

Objectives: To determine the frequency of COVID-19 and its association with age, gender, travel and exposure.

Methods: This cross-sectional study was conducted in COVID-19 clinic, Qazi Hussain Ahmed Medical Complex, Nowshera, from February 2020 to April 2020. A total of 260 suspects were included under strict criteria. Patients received in general OPD and emergencies were excluded. Data was entered in a format in SPSS version 25 for analysis and reporting. Descriptive, inferential and correlation statistics were used.

Results: Out of a total 260 of suspects/patients, 66(25.4%) females and 194(74.6%) were males. The nasopharyngeal swabs of 63(24.23%) cases were sent for viral detection selected for PCR testing on the bases of a scoring system. Out of 63 cases 18(6.92%) were COVID-19 positive, 32(12.30%) negative and results of 12(4.61%) were still awaited. The estimated relative risk in male gender was rr=1.08, 95%CI, 0.77-1.55 and in age group>60 years was rr=2.27, 95%CI, 1.35-5.14. There was a positive statistically significant correlation of disease in patients with contact and travel history (p=0.05, rho=0.12; p=0.001, rho=0.34), respectively.

Conclusion: The results suggest that the age>50 years has higher susceptibility for COVID-19. Similarly, infectivity with 2019-nCoV has a statistically significant correlation with travel history, contact history and age. 

Open Access Original Research Article

Optimization of Mycological Media Using Agro Waste for the Production of Antimicrobial Substance

Evangeline Ogonna Okpalauwaekwe, Chinelo Ursula Umedum, Ikechukwu Harmony Iheukwumere, Leona Chisara Akakuru

Microbiology Research Journal International, Page 67-73
DOI: 10.9734/mrji/2020/v30i830252

Aim: This present study was conducted to optimize mycological media using agro waste for the production of antimicrobial substance.

Place and Duration of Study: Agro waste (sugarcane and sweet potato, sugarcane and jack fruit) collected within Anambra state between February- August 2019.

Methodology: Sugarcane and sweet potato (AMSSP), sugarcane and jack fruit (AMSJ) were peeled and the peels were air-dried and then ground into powdered form. 10 g each of the agro waste samples was weighed into 400 ml of distilled water in 1000 ml Erlenmeyer flask and allowed for 7 days, after which the mixture was filtered.

Then 200 ml of the filtrate was used.

The experimental conditions were optimized by using agro wastes (20/80 and 50/50 concentrations) as a culture medium, altering the temperature (30ºC and 37ºC), pH (5, 6, 7, 8, and 9), as well as the carbon and nitrogen source (glucose and NaNO3). The fungi used were Aspergillus niger, Aspergillus fischeri, Aspergillus aculeatus and Aspergillus fumigatus.

Results:  Various agro wastes medium AMSSP and AMSJ were formulated as mycological media and the growth and nutritional conditions were optimized to ascertain antimicrobial substance production using some fungal isolates. Based on different concentrations Aspergillus fumigatus showed a promising zone of inhibition on AMSSP at a concentration of 20/80 while in AMSJ the concentration the 50/50 showed a maximum zone of inhibition on Aspergillus fumigatus ascertaining the presence of antimicrobial substance. AMSSP was able to produce maximum antimicrobial substance when supplemented with 1.0% glucose, 1.0% NaNO3 at pH 7 and at temperature of 30 ± 2ºC.

Conclusion: Agro wastes from AMSSP as well as from AMSJ contain nutrients that may support fungal growth. Maximum antimicrobial substance production is enhanced when supplemented with 1.0% of the carbon and nitrogen source at a pH of 7 and at a temperature of 30 ± 2ºC.

Open Access Original Research Article

Biofilm Forming Ability and Antibiotic Susceptibility of Food-borne Pathogens Isolated from Common Dairy Products: Madara and Nono Vended in Makurdi Metropolis

Amina Ojochide Hassan, Innocent Okonkwo Ogbonna, Victor Ugochukwu Obisike

Microbiology Research Journal International, Page 74-83
DOI: 10.9734/mrji/2020/v30i830253

Microbial resistance to antibiotics and biofilm formation ability of food-borne pathogens are major global health challenges. Most milk and milk products (Madara and Nono) could be vehicles for the transmission of multidrug resistant genes among any community. This study was aimed at determining the antibiotic susceptibility patterns and biofilm forming ability of some food-borne pathogens isolated from common dairy products: Madara and Nono in Makurdi metropolis. Two hundred and forty (240) samples comprising of one hundred and twenty (120) each of Madara (fresh raw milk from cow “FRM”)) and Nono (chance fermented cow milk “CFM”) were examined for the presence of pathogens. Antibiogram of bacterial isolates (Staphylococcus aureus, Escherichia coli, Shigella spp., Salmonella spp. and Klebsiella spp.) using the disc diffusion method revealed that susceptibility for Ampicillin (86.9%), Streptomycin (83.9%) and Ciprofloxacin (75.0%). Resistance was shown (26.7%) to Nalidixic acid, a commonly used antibiotic reflecting a public health concern. Most resistant isolates had a multiple antibiotics index of 0.3 (27.54%) with a least multiple antibiotics resistance index of 0.6 (0.85%). Detection of biofilm formation of isolates was done by Tube method. The study also revealed that out the total of 236 isolates tested for biofilm formation, 67 (28.4%) isolates were non or weak biofilm producers, 77 (32.6%) isolates were moderate biofilm producers and 92 (39%) isolates were strong biofilm producers. Findings of this research show high presence of a wide range of microorganisms, particularly enteric pathogens and enterotoxigenic strains of S. aureus which portrayed multidrug resistance and biofilm formation suggesting that FRM (Madara) and CRM (Nono) products might be important sources of food-borne infections and intoxication.

Open Access Original Research Article

Genotypic Detection and Characterization of Adhesins in Clinical Escherichia coli Isolates

M. Y. Iliyasu, I. Mustapha, H. Yakubu, H. M. Shuaibu, A. F. Umar, E. B. Agbo, A. Uba, Y. Y. Deeni

Microbiology Research Journal International, Page 84-93
DOI: 10.9734/mrji/2020/v30i830254

Background of Study: Many virulence determinants contribute to the pathogenicity of Gram negative bacteria, like Escherichia coli, which is the most common cause of many infections worldwide such as urinary tract infection (UTI), profuse diarrhoea and septicaemia.

Aim: To determine the genotypic characteristics of adhesin-producing E. coli isolates from clinical specimens.

Place and Duration of Study: Conducted at the Infectious diseases hospital Bayara, Bauchi state, Nigeria, between February to March, 2019.

Methods: A total of twelve (12) Gram negative bacterial isolates were selected based on the ability to grow on Luria-Bertani (LB) agar medium containing 100 µg/ml ampicillin. The isolates were from urine, stool, and blood specimens. The isolates were screened for multidrug resistant pattern according to Kirby-Bauer disc diffusion method. Adhesion factors, Fimbrial adhesin (fimH) and Invasive plasmid adhesin (ipaH) was genotyped by conventional PCR and sequenced.

Results: All the isolates were resistant to Ampicillin, Cephalothin, Erythromycin, Fusidic acid, Novobiocin and Oxacillin, but sensitive to Augmentin, Colistin sulphate and Imipenem. Presence of fimH and ipaH genes were observed in nine isolates that expressed strong relationship with. Multidrug resistance (MDR). The fimH was the most prevalent found in urine, stool and blood isolates. Most of the adhesion genes sequence (61.8%) in this study had significant alignment (95 to 100% homology) with E.coli genome in the NCBI database.

Conclusion: This study revealed the role of adhesin as virulence markers in MDR Gram negative bacteria and FimH is one of the commonest gene in MDR E.coli pathotypes.

Open Access Original Research Article

Laboratory Findings from 100 Patients with COVID-19 Pneumonia in Madinah, Saudi Arabia

Al-ahmadey Ziab

Microbiology Research Journal International, Page 94-103
DOI: 10.9734/mrji/2020/v30i830255

The coronavirus disease 2019 (COVID-19) virus, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is spreading rapidly. The purpose of this study is to explore high risk patients and guiding future management and summarize the results of routine laboratory testing of asymptomatic, mild to moderate, severe and critical COVID-19 in order to define practical indicators for the diagnosis and treatment of COVID-19. A total of 100 confirmed COVID-19 patients from Madinah city, Saudi Arabia, hospitalized between March to June, 2020 were included, and categorized into asymptomatic, mild to moderate, severe and critically ill patients. Fasting blood samples were withdrawn from all patients for estimation of complete blood count, coagulation profile, biochemistry and serology. Patients were grouped on the basis of the interval between symptom onset: group 1 (asymptomatic), group 2 (mild to moderate), group 3 (severe), and group 4 (critical). Laboratory features and their distribution were analysed and compared across the four groups. Combining assessment of clinical and laboratory findings could facilitate early diagnosis of COVID-19 pneumonia. Median age was 51 years old and 85% of the patients were men. Overall, all patients were admitted to hospitals and 42% required ICU treatment. The majority of patients (29%) were diagnosed with mild to moderate disease, 28% of patients were critical, 26% of patients were severe and 17% of patients were asymptomatic. The comparison COVID-19 patients’ four parameters, using Kusakal-Wallis test, showed a significant difference in the levels of lymphocytes, ESR, PT, INR, d-dimer, CK, BUN and ferritin (P <0.05). C-reactive protein greater than 2.97 (0.84-9.18; p<0.0001). The potential risk factors of older age, lymphopenia, d-dimer greater than 1µg/mL and ferritin greater than 500ng/mL could help physicians to identify patients with a poor prognosis at an early stage.

Open Access Original Research Article

Bacteriological and Physicochemical Profiles of Soils in Selected Oil-Contaminated Sites in Yorla, Ogoni Land

Onyinyechi Stainless Uzor, Gideon Chijioke Okpokwasili, Ejiro Obakpororo Agbagwa

Microbiology Research Journal International, Page 104-117
DOI: 10.9734/mrji/2020/v30i830256

Bacteriological and physicochemical properties of oil–polluted sites in Yorla, Ogoniland was studied. Soil samples were collected from three different points and analyzed for bacteriological and physicochemical characterization using standard methods. The total culturable heterotrophic bacteria (THC) counts from the polluted sites ranged from 7.2. *107to9.5*107cfu/g while the THC of the unpolluted sites ranged from 11.9 *107 to 12.9 *107cfu/g. Hydrocarbon utilizing bacterial (HUB) counts from the polluted sites ranged from 2.8 *106 to 3.3 * 106cfu/g while the hydrocarbon utilizing bacterial (HUB) counts of the unpolluted sites ranged from 1.1 *106to 1.5 *106.Bacteria obtained from the various sites were isolated using standard microbiological methods. Identification of isolates was carried out using 16S rRNA (Ribonucleic acid) gene amplification and sequencing. Electropherograms of generated sequences were inspected with Chromas Lite 2.0.1 software. Sequence identification was carried out using GenBank’s Basic Local Alignment Search Tool (BLAST) algorithm of National Centre for Biotechnology and Information (NCBI). Sequence analysis revealed the presence of Acinetobacter venetianus , Bacillus cereus, Bacillus flexus, Bacillus thuringiensis, Acinetobacter junii, Shewanella sp., Lysinibacillus xylanilyticus and Pseudomonas pseudoalcaligenes,  The physicochemical properties of the soil samples analyzed showed that the pH values of the polluted sites ranged from 7.86 to 9.16 while those of  the unpolluted sites were 6.19 and 7.10. Total organic carbon (TOC) contents of the polluted sites ranged from 3.18% to 3.96% while the unpolluted sites TOC ranged from 3.00% -3.06%. Statistical analyses of the results were carried out using analysis of variance (ANOVA) and t-test analysis. This work revealed the presence of reasonable population of indigenous hydrocarbon-utilizing bacteria in oil-polluted sites in Yorla community which can be monitored and enhanced to improve their bioremediation abilities in the oil-polluted sites.

Open Access Original Research Article

GC-MS Analysis and Antimicrobial Spectrum of Stem Bark Extracts of Ficus sycomorus

Yunusa Saheed, Mustapha Usman Nasir, Baballe Abbas, Rabi Yakubu Bello

Microbiology Research Journal International, Page 118-128
DOI: 10.9734/mrji/2020/v30i830257

Aim: This work investigated the gas chromatography-mass spectroscopy of methanolic extract and antimicrobial spectrum of acetone and aqueous crude extracts of Ficus sycomorus stem bark.

Place and Duration of the Study: Department chemistry research laboratory and microbiology laboratory, Yobe State University, Damaturu between April 2019 and August, 2019.

Methodology: Pure isolates of Staphylococcus aureus, Salmonella typhi, Shigella dysentariae, proteus spp, Pseudomonas auruginosa, Escherichia coli, Bacillus subtilis, Klebsiella pneumonia and a fungi, Candida albicans were collected from National Veterinary Research Institute (NVRI), Vom, Plateau State. The plant was collected and the extract was used in preliminary phytochemical screening, GC-MS analysis and reconstituted for antimicrobial activities testing.

Results: The preliminary phytochemical screening revealed presence of saponin, flavonoids, alkaloids, carbohydrates, cardiac glycoside and tannin. GC-MS analysis revealed the presence of heterocyclic members of pyrazole, imidazole, triazole, thiazole and 1, 3-Benzenedicarboxylic acid, bis (2-hethylhexyl) ester. Acetone extract showed the greatest antimicrobial activity against Bacillus subtilis, Pseudomonas aeruginosa, Proteus mirabilis and Shigella spp while the aqueous extract produced antibacterial activity against Bacillus subtilis, Staphylococcus aureus, Klebsiella pneumonia and Candida albicans. Both acetone and aqueous extracts of both stem bark extracts did not exhibit any activity against Escherichia coli. All the extracts were tested at 100mg/ml concentration. The Minimum Inhibitory Concentration (MIC) and the Minimum Bactericidal Concentration (MBC) (125 – 250 mg/ml) of both extracts were higher than those of ciprofloxacin (5.0 mg/ml).

Conclusion: There is scientific basis for the uses of F. sycomorus in traditional medicine for the treatment of various diseases as prescribed earlier on and stronger basis for recommendation for its cautious use for the presence of 2-ethylhexyl which could be Carcinogenic. This is first study to identify 2-ethylhexyl from F. sycomorus stem bark extract and recommend cautious use.