International Journal of Advanced Biological and Biomedical Research

A study of Mycobacterium tuberculosis Zopf, 1883 (Mycobacteriaceae) in Iraq

Haidar Khalid Mostafa; Mohsen Hashim Risan; Mohammed Al-Faham

Volume 10, Issue 1 , February 2022, , Pages 1-17

https://doi.org/10.22034/ijabbr.2021.534391.1362

Abstract

Background: Tuberculosis (TB) is considered to be one of the deadliest bacterial infections in the world and it can have a large impact on global health, drawing international attention ... Read More Background: Tuberculosis (TB) is considered to be one of the deadliest bacterial infections in the world and it can have a large impact on global health, drawing international attention with increasing number of cases worldwide in developed and developing countries. Mycobacterium tuberculosis (MTB) is the main causative agent of TB which is an aerobic pathogenic bacillus that establishes infection in the lungs. It is believed that two billion people are carrying non-eradicated intra-granulomatous Tuberculosis bacilli as LTBI (short for latent tuberculosis infection) and around 10% of those people will be infected with active tuberculosis during their lifetime. Therefore, this study aimed to study Mycobacterium tuberculosis in Iraq.Methods: All bacteriological tests were done in biological safety cabinet class II. Gloves and masks (N 95) were used during the laboratory work. Phenol 5% and ethanol alcohol 70% were used to sterilize and disinfect the benches and hood (IUATLD, 1996). Autoclave was used at 121 °C at 1.5 bar for 20-30 minutes for sterilizing all culture media. The study was conducted at National Reference Lab (NRL) of tuberculosis/Baghdad from July 2016 to December 2016. A total of 188 sputum samples were collected from suspected patient. In addition, 80 samples were collected from healthy subjects Specimens were amassed within the outdoor and as away as feasible from different human beings. Each subject was advised to inhale deeply 2-3 instances, cough out deep from the chest and spit the sputum into sterile prevalent container. We needed to ensure that the specimen was of enough extent (3 to 5 mL) and that it incorporated strong or purulent material. Two samples were gathered from the subject. The first was taken when they reached the institute and the second was taken early morning, earlier than breakfast. The early morning collection represents the pulmonary secretions accrued in a single day, and consequently it typically has a better positivity. Gathered specimens were stored at –20 °C until use.Results: Totally, 118 Tuberculosis suspected samples were used throughout this study; 103 (54.78%) cases were positive using direct assay (AFB smear microscopy) and 85 (45.21%) were negative cases. From the 103 TB patients, 88 (85.43%) represented as new TB cases; the others 15 (14.57%) patients appeared to follow up assessment of the directly observed treatment (DOT) program. The percentage of infected patients was 73 (70.87%) for males to 30 (29.12 %) for females, with over all male to female ratio of 2.43 (73/30) with a highly significant difference (P≤0.01). The age of the study patients ranged from 14 years to 75 years. The mean age was 36 and the median was 30 with highly significant difference between age group. As in many developing countries, direct sputum microscopy is the widely used method for the diagnosis of pulmonary TB in Iraq. Only 103 from 188 patients (suspected) were positive by using ZN smears (Ziehl-Neelsen stain technique). This study for those 188 suspected showed that 119 (63.29%) cases were positive by culture (Lowenstein–Jensen medium) and 69 (36.7%) cases were negative by L.J Culture. This study showed that the percentage of sensitivity, specificity, positive predictive value, and negative predictive value was 74.78%, 79.7%, 86.4% and 64.7%, respectively, using AFB method. The results revealed out of the total 188 specimens, 49 (26.06%) of the specimens were positive by DiaSpot (Tuberculosis Rapid Test on serum) and 139 (73.93%) specimens were negative. This study reported the sensitivity of 33.6% and the specificity of 86.95 % PPV 81.6 and NPV 43.16 for DiaSpot TB Rapid test.Conclusion: Males aged 15 to 45 years are at high risk for TB infection. M. tuberculosis plays a major role for causing tuberculosis in human in Baghdad.

Biochemical Characteristics and Antibiotics Susceptibility of Streptococcus Mutans Isolates from Dental Caries in Baghdad City

Aya Raad Salh; Mohsen Hashim Risan; Hameed Majeed Jasim

Volume 10, Issue 1 , February 2022, , Pages 32-43

https://doi.org/10.22034/ijabbr.2022.534492.1363

Abstract

Background: Genus Streptococcus comprises important pathogens that have a severe impact on human health. According to the classification in Bergey's Manual of Determinative Bacteriology, ... Read More Background: Genus Streptococcus comprises important pathogens that have a severe impact on human health. According to the classification in Bergey's Manual of Determinative Bacteriology, the genus Streptococcus includes the pyogenic, oral and anaerobic groups of streptococci, as well as a group of other streptococci. S. mutans is a Gram-positive bacterium, which plays a key role in the formation of the dental plaque biofilm as an early coloniser and is the most important bacterium in the formation of dental caries. S. mutans do not have flagella, but do have pili and are non-motile facultative anaerobes that grow optimally at 37 °C. Methods: Eighty samples were collected from patients with different dental caries (pit, fissure and dental roots). The patients had referred to Al-dora Health center and Al-Zewiya Health center in Baghdad city. Bacterial isolates obtained from dental caries samples were streaked on selective medium (Mitis Salivaris Agar) for isolation of S. mutans, then the plates were incubated at 37 °C for 48 hours under anaerobic conditions. Bacterial isolates were identified according to their morphological and cultural characteristics and biochemical tests. The disk diffusion method was used to test the antibiotic susceptibility of the selected isolate to different antibiotics. A sterile cotton swab was dipped in fresh culture of S. mutans and streaked on the surface of Muller-Hinton agar plates by rotating the plate approximately 60 between streaking to ensure even distribution. The inoculated plates were incubated at room temperature for 10 minutes to allow absorption of excess moisture, then antibiotic disks were fixed by sterile forceps on the surface of plates and incubated at 37 °C for 18 hrs.Results: Totally, 98 bacterial isolates were obtained. The results indicated that antibiotic susceptibility pattern of the locally isolated S. mutans S2 was resistant to two antibiotics (bacitracin and erythromycin), while it was sensitive to the other eight antibiotics, consisting amoxicillin, ampicillin, chloramphenicol, vancomycin, clindamycin, imipenem, gentamycin and cephalothin.Conclusion: Locally isolated S. mutans S2 was sensitive to different antibiotics, while it resisted only against bacitracin and erythromycin.

Screening of bacteria Streptomyces Waksman and Henrici 1943 (Streptomycetaceae) Isolates from Soil Samples in Iraq

Bassam Qasim; Mohsen Hashim Risan

Volume 9, Issue 4 , December 2021, , Pages 340-351

https://doi.org/10.22034/ijabbr.2021.534456.1364

Abstract

Background: The genus Streptomyces Waksman & Henrici 1943 includes aerobic, gram-positive, and filamentous bacteria which produce well developed vegetative hyphae with branches. ... Read More Background: The genus Streptomyces Waksman & Henrici 1943 includes aerobic, gram-positive, and filamentous bacteria which produce well developed vegetative hyphae with branches. The wall consists of a large mixture of different compounds, including peptidoglycan, teichuronic acid, teichoic, and polysaccharides. The peptidoglycan components consist of glycan as a chains of irregular N-acetyl- d-muramic acid (NAM), diaminopimelic acid, and N-acetyl-d-glucosamine (NAG) and DAP, which is unique in the cell walls of prokaryotic microorganisms. The teichoic and teichuronic acid are chemically bonded to peptidoglycan.Methods: One gram of soil samples was used to make suspension, by adding 99 mL of sterile distilled water (stock suspension) into it and shaking it in a shaker at 160 rpm for 30 minutes at room temperature. Serial dilutions from 0.1-0.001 were made from the stock suspension, and left for 10 minutes. After shaking, 0.1 mL of each dilution was cultured on Yeast Extract and Malt Extract agar (YEME) with Streptomycin 50 ug/mL. The inoculated plates were incubated at 28 °C for 7 to 10 days. Based on cultural characteristics, suspected colonies of Streptomyces were selected, which are characterized as small, white, pin-point, rough, chalky, and a clear zone of inhibition around them. These colonies were confirmed their identification by types of Gram’s stain, aerial and substrate mycelium color, pigment production, and pigment color. Streptomyces were re-streaked on International Streptomyces project (ISP) to obtain pure colonies used for identification. Results: The current study aimed to screen bacteria Streptomyces isolates. Only 21 samples of soil were suspected to contain Streptomyces, and 45 isolates were obtained with different morphology types per samples of soil. The colonies suspects were selected basis on color that ranged from gray, white and creamy. The microscopic examination of local Streptomyces spp. after Gram-staining method was conducted. The observations revealed that local Streptomyces is gram positive and rod shaped similar to features of fungal in possessing branched mycelium. The Streptomyces produced extra cellular enzymes like amylase, urease, catalase, protease, Gelatinase, cellulase and phosphatase. Utilization of citrate was positive, with no Melanine reaction production and soluble pigmented, and negative for indole production.Conclusion: The identification of the Streptomyces is a very complex process. Morphological and biochemical characteristics are two important aspects for the classification in the Streptomycetaceae family. By studying the morphological, cultural, and biochemical characteristics, it is observed that the local isolates are belonging to the genus of Streptomyces.

Candida Berkh. (1923) Species and Their Important Secreted Aspartyl Proteinases (SAP) Genes Isolated from Diabetic Patients

Thamer A Muhsen; Mohsen Hashim Risan; Nawras N Alqaysi

Volume 9, Issue 3 , September 2021, , Pages 286-297

https://doi.org/10.22034/ijabbr.2021.534165.1361

Abstract

Background: Candida Berkh. (1923) occurs naturally in the body. But it becomes opportunistic fungi, meaning that it infects humans when there is any weakening of the immune system, ... Read More Background: Candida Berkh. (1923) occurs naturally in the body. But it becomes opportunistic fungi, meaning that it infects humans when there is any weakening of the immune system, such as exposure to chemotherapy, diabetes, or organ transplantation. Most species of Candida grow at a temperature between 20-40 °C and have a pH of 3-8. Human pathogens of Candida species include C. albicans, C. glabrata, C. lusitaniae, C. parapsilosis, C. tropicalis and C. utili. C. albicans has many virulence factors that facilitate injury process. Virulence factors are considered as a measure of pathogenicity, and it is in the form of fungal toxins, enzymes, or cell structures that facilitate infection, as well as pathogen resistance in different conditions. This study aimed to investigate the frequency of some secreted aspartyl proteinases (SAP) genes from some Candida species isolated from diabetic patientsMethods: Candida spp. were identified on CHROMagar medium by color of each species after incubation at 37 °C for 48 hours such as: C. krusei, C. albicans, C. glabrata, and C. tropicalis. Germ tube formation test was used to distinguish between germ tube-forming species, such as C. albicans and C. dubliniensis , from those that do not produce germ tube. A portion of the isolation was taken and placed in a test tube containing 0.5 ml of serum, and then incubated at 37 °C for 2-4 hours. A drop of serum was taken out and examined under a microscope for the presence of the germ tube. Growth at 45 °C, has the advantage of distinguishing between C. albicans and C. dubliniensis by culturing Candida spp. on SDA medium and, incubation for 48-72 hours at 45 °C.Results: The results of isolation and diagnosis showed that Candida species grew at 37 °C for 48 hours on SDA medium, the shape of the colonies was round or oval-convex, and creamy white, shiny and smooth color. The results of the microscopic examination of the cells were spherical or oval, C. albicans was distinguished by its ability to grow at 45 °C and its ability to form the germ tube. The surface growth test showed the ability of C. tropicalis to grow surface on SDB medium. The medium of CHROMagar showed that C. albicans were light green in color. C. glabrata showed a pale pink color, while C. tropecalis showed a blue color and C. dublineiensis was dark green. DNA extracted samples, including 20 isolates, were used for four species of Candida, 1-9 C. albicans, 10-16 C. glabrata, 17-18 C. dubliniensis and 19-20 C. tropicalis. Parts of the body (mouth, vagina, Urine) detect SAP1 and SAP7 genes by PCR, and after electrophoresis of the PCR product, the results were positive for all Candida species studied as all isolates that contained SAP1 (578bp) and SAP7 (466bp).Conclusion: The SAP genes are among the main virulence factors for the occurrence of candidiasis. SAP1 and SAP7 genes are responsible for digesting and breaking down protein to penetrate the host's tissues.

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Author = Mohsen Hashim Risan

A study of Mycobacterium tuberculosis Zopf, 1883 (Mycobacteriaceae) in Iraq

Abstract

Biochemical Characteristics and Antibiotics Susceptibility of Streptococcus Mutans Isolates from Dental Caries in Baghdad City

Abstract

Screening of bacteria Streptomyces Waksman and Henrici 1943 (Streptomycetaceae) Isolates from Soil Samples in Iraq

Abstract

Candida Berkh. (1923) Species and Their Important Secreted Aspartyl Proteinases (SAP) Genes Isolated from Diabetic Patients

Abstract