Online ISSN: 2515-8260

Keywords : PCR

Molecular Analysis of Chlamydia trachomatis in Infertile Women in Basrah

Maysoon Sharief; Mayada T. Abdulrahman; Hanadi A. Jasim

European Journal of Molecular & Clinical Medicine, 2021, Volume 8, Issue 3, Pages 530-539

ABSTRACTBackground Chlamydia trachomatis is a gram negative bacteria, which involve in sexually transmitted disease. This study was aimed to detect C. trachomatis by molecular methods and to evaluate Chlamydial infections in women suffering from primary and secondary infertility with special emphasis. Method 200 endocervical cytobrush were obtained from 200 infertile women having primary and secondary infertility. The work has been carried out at Basra Maternity and Child Hospital for molecular analysis of C. trachomatis . The primer CT1& CT4 w used to investigate the 144bp of MOMP of C. trachomatis in the endocervical brush samples. Results Out of 200 infertile women 96(48%) were positive for C. trachomatis by PCR. Conclusion The percentage of C.trachomatis in primary infertile women with blocked tubes was higher than women with patent tubes and also higher than C. trachomatis in secondary infertile women with both blocked and patent tubes.

Identification of cultivable and non-cultivable organisms causing intraarticular and bone infections using molecular diagnostic techniques

Deepali J Shetty; Dr Shalini Shenoy Mulki; Dr B N Jagannath Kamath; Dr Sevitha Bhat; Dr Archana Bhat

European Journal of Molecular & Clinical Medicine, 2021, Volume 8, Issue 3, Pages 643-651

Introduction: Osteomyelitis is pathologically defined as inflammation of bone and bone marrow due to infection with a microbial pathogen. Septic arthritis is pyogenic infection of the joint either due to direct extension from local tissue infection or more commonly as a result of bacteremia. Microbiological diagnosis in these cases can be made by culture on enriched media and PCR. As PCR detects bacterial DNA, it does not rely on the presence of viable bacteria in the sample for identification. The rapidity of PCR along with its higher sensitivity and specificity enables faster and more accurate diagnosis and treatment

Study Of Some Virulence Factors And Virulence Genes Produced By Gram (+Ve) And Gram (-Ve) Cocci Isolated From Clinical Samples In Al-Anbar Province/Iraq

Rawaa Turki Abdul Ghafoor; Asst. prof. Hana Abdel Latif Yassin

European Journal of Molecular & Clinical Medicine, 2020, Volume 7, Issue 9, Pages 535-547

This study aimed to isolate the G (+ve) and G(-ve) cocci from different clinical samples and study some virulence factors and virulence genes produced by them .
In this study, (177) samples were collected from different clinical sources, including (49) blood samples, (43) urine samples, (30) pus swabs, (40) cervical swabs and (15) sputum samples from patients attending Ramadi Teaching Hospital for Maternity and Children, Ramadi Teaching Hospital in Ramadi in Al-Anbar province/Iraq during the period from 19/12/2018 to 12/5/2019, and (203) isolates were identified as Gram (+ve) and Gram (-ve) bacteria, then (166) Gram (+ve) and Gram (-ve) cocci were chosen as study samples.
After diagnosis of the isolates, results of Gram staining showed that the highest number and percentage of Gram positive and Gram negative bacteria were from urine samples 48(28.9%) followed by blood samples 44(26.5%) then pus swab and cervical isolates 37(22.3%) and 23(13.9%) respectively, while the lowest number and percentage of isolates were from sputum 14(8.4%). Staphylococcus aureus showed the highest number and percentage of isolates 48(28.9%). For gram (-ve) cocci, only (10.4%) of Neisseria gonorrhea was isolated from urine samples and (14.2%) of Moraxella catarrhalis was isolated from sputum samples
Regarding virulence factor production, the hemolysin producer isolates were the highest 103(62.04%), and results of hemolysin producing bacteria was S.aureus, S.hominis, S.haemolyticus, S.chromogenes, S.xylosu and S.warneri showed (100%) beta hemolysis, while only one isolate of the genus Micrococcus was beta-hemolytic all isolates of Streptococcus pyogenes and Streptococcus agalactiae showed (100%) beta hemolysis (β), whereas the isolates of Strept. pneumoniae, Aerococcus urinae and Aerococcus viridans showed (100%) partial or alpha (α) type hemolysis, and (8) isolates of E.faecalis showed complete hemolysis (β-haemolysis).
All S. aureus isolates were (100%) gelatinase producers Kocuria kristinae was (100%) producer, all Strep. pyogenes, Strep. Agalactiae, Strep. Anginosus, Strep. Lc. lactis lactis and Strep.mitis isolates showed (100%) ability to produce this enzyme, The (24) isolates of the genus Enterococcus bacteria were (100%) able to produce the enzyme gelatinize, including (19) E.faecalis isolates and (5) E. faecium. As for the genus Gram (-ve) cocci, two isolates of Moraxella catarrhalis showed (100%) ability to produce the gelatinase enzyme.
The number and percentage of urease enzyme producers were 49(29.51%), where 38(79.16%) of S.aureus isolates were able to produce it. Also, 3(17.64%) of S.hominis isolates were able to produce it, while only one isolate of S.lentus (20%) was able to produce it. Moreover, both S.xylosus and S.warneri isolates were (100%) urease-producers.
From the total (166) isolates, 20(12.04%) isolates formed biofilms. E.faecalis bacteria was the most biofilm-producing isolate 9(47.36%), followed by S.aureus bacteria 3(6.25%) then
S. lentus isolates (40%), while Kocuria rosea bacteria produced biofilm as 1(100%), Strep.pneumoniae and Strep.pyogenes produced as 1( 25%) for each isolate, and 1(50%) for Strep. anginosus isolate, whereas Strep.salivarius and Strep.mitis produced biofilm as 1(100%) isolate for each of them.
Also (100%) of the virulence genes icaC, srtC, and Emm were detected in S.aureus, E.faecalis and Strep.pyogenes isolates, which produced strong, moderate and weak biofilm types by using the PCR technology.

Comparison of Microscopy and Polymerase Chain Reaction Examination Results in the Detection of Malaria Parasites

Nailatul Hana; Lia Faridah; Hesti Lina Wiraswati; Jontari Hutagalung

European Journal of Molecular & Clinical Medicine, 2020, Volume 7, Issue 10, Pages 2871-2881

Background: Microscopy remains the mainstay method for malaria diagnosis worldwide, although species misidentifications have been detected in practices due to various limitations, such as hypnozoites detection and lower parasitemia in asymptomatic malaria. Polymerase Chain Reaction (PCR) is a molecular diagnostic method with high accuracy in detecting species of organisms. This study was aimed to evaluate the diagnostic performance of microscopy compared to nested PCR in detecting malaria parasites.
Methods: A cross-sectional study was conducted with previous data on malaria assessment in East Nusa Tenggara. More than 500 asymptomatic respondents were included by the systematic random sampling method from 5 sub-districts area in the region based on API. Microscopic assessment by thick and thin blood smears was made following protocols from the Ministry of Health, while DNA isolation was done using 200 μl fresh blood sample and nested PCR amplification protocol with specific primers of the malaria parasites species Plasmodium sp.
Results: A total of 555 specimens were collected, and 1.6% (9/555) of those were microscopy-positive and 32.6% (181/555) were detected positive by nested PCR. Of microscopy-positive samples, 33.3% (3/9) were P. falciparum and 66.7% (6/9) were P. vivax, whereas among PCR-positive samples, 31.5% (57/181) were P. falciparum, 52.5% (95/181) were P. vivax, and 16.0% (29/181) were mixed infection of both species. From this study, microscopy was found to had a slight measure of agreement (κ = 0.055) compared to nested PCR.
Conclusion: In lower parasitemia and asymptomatic malaria, the microscopic assessment may not be sensitive. Thus, this increases the need of using PCR assessment to confirm the identification of malaria parasites.

Isolation and Molecular detection of Capnocytophagacanimorsusfromcanine oral samples

Mustafa Salah Hasan; Omar Attalla Fahad

European Journal of Molecular & Clinical Medicine, 2020, Volume 7, Issue 2, Pages 3600-3606

A fifty oral swabs (25 from Alanbar and 25 from Salah Aldein)of different aged dogs,
different global strains and both sex were collected.A designed gene was used to
diagnoseCapnocytophaga canimorsus by PCR. Collectedsamples were cultivated
primarily onto nutrient, blood and macConkey agars, then the colonies were
subcultured on brain heart infusion and blood agars supplemented with 2.5 μg/ ml of
trimethoprim and 2.5 μg/ ml of amphotericin B. A gram stain and biochemical tests
were done including catalase, oxidase and fermentation of glucose, maltose, sucrose,
sorbitol and mannitol. The results of isolation and identification showed that twenty
two (9 from Alanbar and 13 from Salah Aldein) isolates suspected that were
Capnocytophaga spp. these suspected isolates of Capnocytophaga spp. were
accompanied to assay of PCR, the viewing results presented thatfourteen (6 from
Alanbar and 8 from Salah Aldein) isolates were positive for the designed gene. Also,
the results of percentage for C. canimorsus showed that 24% in Alanbar and
32%from Salah Aldein provinces, while the other 36 dogs showed negative results for
C. canimorsus, 19(76%) from Alanbar and 17(68%) from Salah Aldein.

Comparative Analysis Of Rt-Pcr And Immunohistochemistry Methods For Determining Her2 Status In Breast Cancer Samples

Shokhista Rustamova; Yodgormirza Nurmatov; Muazzam Bakiyeva; Tokhirjon Rakhmanov

European Journal of Molecular & Clinical Medicine, 2020, Volume 7, Issue 3, Pages 3924-3929

This study aimed to compare two methods of quantitative polymerase chain reaction (qPCR) using the intercalating SYBR Green dye and TaqMan hybridization probes to determine the amount of the HER2 gene (human epidermal receptor) in breast tumours.
Methods--- The experiments were carried out with 32 validated samples of breast cancer and two gastric cancer cell lines. An immunohistochemical (IHC) assay was used to evaluate the accuracy of the RT-PCR methods.
Results--- The obtained results show that real-time PCR with the TaqMan probes allows the use of a small amount of DNA (≥0.4 ng/μl) to determine the overexpression of the HER2 gene. Real-time PCR with SYBR Green allowed us to determine the minimum number of false negatives resulting from the absence of marker expression in tumour tissue.
Conclusions--- The full correspondence of the results of the RTPCR and immunohistochemistry methods obtained for the cell line samples makes it possible to introduce the qPCR method into clinical practice for use in detecting the HER2/neu gene content.