A survey of resistance mutations to reverse transcriptase inhibitors (RTIs) among HIV-1 patients in northeast of Iran.

The use of a combination of three-drug regimen has improved HIV-1 infected patients' life span and quality; however the emergence of drug-resistant strains remains a main problem. Reverse transcriptase inhibitors (RTIs) consist of a main part of highly active anti-retroviral therapy (HAART) regimen. The present study aimed to investigate resistant mutations to RTI drugs in both treatment naïve and under treatment HIV patients in Mashhad city, north-eastern Iran. RNA was extracted from sera of 22 treatment naïve and 22 under treatment patients. The mean age of under treated and treatment naive groups were 38.5±6.7 and 40.8±7.9 respectively. cDNA was synthesized and amplified with Nested PCR assay targeting specific sequences of RT gene. The PCR products were sent for sequencing. Bidirectional sequencing results were analysed using HIV drug resistance database supplied by Stanford University (HIV Drug Resistance Database, https://hivdb.stanford.edu). Among under treatment patients 10 out of 22 (45%) had at least one high-level resistance mutation which was higher than high level resistance mutation rate among treatment naive cases (P<0.01). Detected resistance mutations were as follows: K101E, K103N, K103E, V106M, V108I, E138A, V179T, Y181C, M184V, Y188L, Y188H, Y188F, G190A, L210W, T215F, T215Y, K219Q, and P225H. A high level of resistance mutations to RT inhibitors was observed that causes drug resistance especially against lamivudine (3TC). Such mutations should be considered as probable responsible for therapeutic failure. Serial surveillance studies of circulating drug resistance mutations are recommended.

Human BKV large T genome detection in prostate cancer and benign prostatic hyperplasia tissue samples by nested PCR: A case-control study.

Human BK polyomavirus (BKPyV) is a latent infectious agent in the genitourinary tract associated with hemorrhagic cystitis and nephropathy. This virus can be a risk factor for various human malignancies, including prostate cancer (PCa). It may contribute to prostate cancer development, as it demonstrates oncogenic properties by encoding oncoproteins. This study assessed the prevalence of this virus in benign and malignant prostate tissues. Between 2009 and 2019, 49 formalin-fixed paraffin-embedded (FFPE) PCa and 49 benign prostatic hyperplasia (BPH) samples were gathered from the pathology department of a tertiary care university hospital. They were used as cases and controls, respectively. After deparaffinization and DNA extraction, nested PCR was applied to identify the BKPyVgp5 gene (LTAg) using inner and outer primers. The nested PCR showed a 278-bp bond corresponding to the BKPyVgp5 genome (LTAg) in 53.1% (26/49) of PCa and 14.3% (7/49) of BPH (p<0.001). The presence of BKV was significantly associated with an increased risk of PCa development (OR=6.78, 95% CI=2.55-18.02, p<0.001). The BKV LTAg gene was significantly more prevalent in PCa samples than in BPH samples. These results demonstrate the presence of the virus in prostate cancer tissues.

Characterization of carbapenem-resistant Escherichia coli and Klebsiella: a role for AmpC-producing isolates.

Aim: This study aimed to investigate the role of AmpC enzymes in carbapenem resistance among AmpC/extended-spectrum ß-lactamase (ESBL)-producing clinical isolates of Escherichia coli and Klebsiella spp. Methods: Fifty-six bacterial strains that were AmpC producers were examined. The antibiotic susceptibility test was performed by the disk diffusion and E-test. The prevalence of the plasmid carbapenemase was determined using PCR. Results: The resistance to meropenem in the AmpC+/ESBL+ group was 64%, higher than that reported for the AmpC-/ESBL+ group. Ten isolates of the carbapenem-resistant AmpC producers were negative for carbapenemase-encoding genes. Conclusion: Carbapenem resistance among AmpC-producing isolates with negative results for carbapenemase-encoding genes potentially demonstrates the role of AmpC enzymes among these isolates.

Seroprevalence of VZV and HSV-2 Antibodies among Women of Childbearing Age Referring to Health Centres of Mashhad, Iran: The Need for Consideration of VZV Vaccination Program.

Background: Infections with herpes simplex virus type 2 (HSV-2) and varicella-zoster virus (VZV) are associated with serious maternal and neonatal health consequences. The literature review reveals a research gap regarding the seroprevalence of HSV-2 and VZV among women of reproductive age in Mashhad, Northeast of Iran. The present study aims to evaluate the seroprevalence of these viruses among a group of women in Mashhad, Iran. Methods: Sera were collected by health center personnel using a cluster sampling method from healthy women with specific age characteristics residing in three distinct socioeconomic regions of the city. The participants, aged 20-35, were divided into three groups (20-25, 26-30, and 31-35 years). The levels of VZV and HSV-2 IgG antibodies were evaluated using commercial ELISA kits. Subsequently, the results were analyzed using SPSS software. Results: A total of 93 women were included in the study. Anti-HSV-2 IgG antibody was detected in 3 out of 93 participants (7.5%), while anti-VZV IgG antibody was found positive in 80 out of 93 individuals (83.3%). The HSV-2 positive cases were concurrently positive for the VZV antibody. There was no significant difference in the positivity of anti-HSV-2 and anti-VZV antibody positivity within age groups or socioeconomic status (P > 0.05). Conclusions: The high seroprevalence of VZV among nonvaccinated participants indicates a widespread presence of the virus and underscores its potentially serious impact on community health. Therefore, it is recommended that a VZV vaccination program be considered by the health system. Furthermore, the reactivation of latent HSV-2, whether symptomatic or asymptomatic, during pregnancy should not be disregarded as a life-threatening threat.

Glutathione reductase system changes in HTLV-1 infected patients.

During chronic HTLV-1 infections oxidative stress occurs and contributes in viral pathogenesis. Glutaredoxin (Grx) system is one of the most effective antioxidant components. The system maintains the cellular redox and scavenges reactive oxygen species through the function of glutathione reductase (GR) enzyme, NADPH and reduced glutathione (GSH). This study was performed to investigate potential changes in GR gene expression and activity as well as GSH level, and their association with the viral load in HTLV-1 infection. Forty HTLV-1 seropositive patients divided into two groups: asymptomatic carriers (N = 20) and HAM/TSP (N = 20) with the same number of age and sex-matched healthy controls were recruited in this study. GR cellular gene expression and viral load in PBMCs were determined using Real-time PCR Technique. Enzyme activity and GSH level in sera were measured by commercial kits based on manufacturer's provided protocols. GR gene expression and GR enzyme activity, as well as GSH level, were significantly lower in HTLV-1 patients. A negative correlation between viral load and GR gene expression/enzyme activity was observed in HAM/TSP group. Similarly, a negative relationship between viral load and GSH levels was observed in both carrier and HAM/TSP groups. We also found that in profound complicated condition of HTLV-1 infection, HAM/TSP, Grx system components activity was significantly decreased compared to the controls. Such observation was not the case in clinically healthy HTLV-1 carriers. These findings may shed a light on the conditions contributing in pathogenesis of the complications and exacerbation of the disease in the HAM/TSP cases. Supplementary Information: The online version contains supplementary material available at 10.1007/s13337-022-00758-y.

Evaluation of antioxidant status and oxidative stress markers in HTLV-1 infected individuals: correlation with the severity of virus-induced complications.

Introduction. Human T-cell lymphotropic virus type 1 (HTLV-1), a well-known member of the retroviridae family, potentially causes serious outcomes including adult T-cell leukaemia/lymphoma (ATLL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM-TSP). Oxidative stress plays a key role in progression and clinical exacerbation of several chronic infections. We have previously shown a reduction in serum total antioxidant capacity (TAC) during HTLV-1 infection and this study was set out to investigate the reasons for TAC reduction.Hypothesis/Gap Statement. Oxidant/antioxidant imbalance during HTLV-1 infection may result from disruptions in oxidant levels or antioxidant defence system.Aim. This study aimed to analyse the key enzymes and oxidant molecules playing important roles in virus-induced oxidative stress.Methodology. We measured serum activities of the major antioxidant enzymes; superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX) as well as serum concentrations of the main oxidant markers: nitric oxide (NO) and malondialdehyde (MDA). Totally 40 HTLV-1 infected patients and 40 healthy controls were enrolled in this study. The patient group consisted of chronic carriers and patients with HAM-TSP (N=20).Results. The current study found that serum levels of MDA and NO were significantly higher in patient groups particularly in HAM-TSP patients (P<0.05). In addition, a reductive trend was observed in the serum activities of CAT, SOD, and GPX in HTLV-1 infected patients compared with healthy controls (P<0.05).Conclusion. Reduced activities of CAT, SOD, and GPX antioxidant enzymes along with the observed elevated concentrations of oxidant molecules may contribute to oxidative stress and worse outcomes during HTLV-1 infection.

Green synthesis of selenium nanoparticles using Rosmarinus officinalis and investigated their antimicrobial activity.

The interest of many has been attracted by plant-mediated synthesizing procedures for nanoparticles since they provide certain qualities including being cost-effective, quick, and compatible with the environment. In this regard, this work introduces the production of selenium-nanoparticles (Se-NPs) in a biological manner utilizing aqueous extracts of Rosmarinus officinalis (R. officinalis). Production of Se-NPs was confirmed using UV-visible (UV-Vis) spectrophotometry. Also, dynamic light scattering (DLS) analysis was used for determination particle size distribution, while we distinguished the identification of crystalline construction of nanoparticles through the means of X-ray diffraction (XRD) pattern, DLS, and transmission electron microscopy (TEM) examination indicated that Se-NPs are often spherical with a size about 20 to 40 nm. The minimum inhibitory concentration (MIC) of the synthesized Se-NPs by R. officinalis extract against Mycobacterium tuberculosis (M. tuberculosis), Staphylococcus aureus (S. aureus), Streptococcus mutans (S. mutans), Escherichia coli (E. coli), and Pseudomonas aeruginosa (P. aeruginosa) was 256, 16, 32, 128, and 64 µg/mL, respectively. The synthesized Se-NPs had no significant effect on Mycobacterium simiae (M. simiae) and had exhibited a strong antimicrobial functionality towards the gram-positive and gram-negative bacteria and can stand as a potent antibacterial agent.

Total antioxidant capacity as a marker of severity of COVID-19 infection: Possible prognostic and therapeutic clinical application.

The pathogenesis of SARS-CoV-2 infection, causative pathogen of the known COVID-19 pandemic is not well clarified. In this regard oxidative stress is one of the topics that need to be investigated. Therefore, the present research was performed to explore the relationship between the oxidant/antioxidant system and COVID-19 exacerbation. Sera were collected from 120 patients with COVID-19 infection and 60 healthy volunteers as the control group. The patient group consisted of 60 cases with mild disease and 60 severely ill patients. Serum levels of total antioxidant capacity (TAC) and nitric oxide (NO) as well as serum activities of the two main antioxidant defense enzymes, superoxide dismutase (SOD) and catalase (CAT), were measured. TAC levels were considerably lower in patients compared with healthy individuals (p < 0.05) and also between patients with mild and severe diseases (p < 0.05). A rather decreasing trend was also found in NO concentration as well as SOD and CAT activity, though, the observed differences were not statistically significant (p > 0.05). These findings suggest that COVID-19 patients may be susceptible to depleted total antioxidant capacity. Moreover, showing such variations in blood samples of infected individuals could be considered as a predictive marker of COVID-19 severity.

Higher detection of JC polyomavirus in colorectal cancerous tissue after pretreatment with topoisomerase I enzyme; colorectal tissue serves as a JCPyV persistence site.

BACKGROUND: The JC polyomavirus has been blamed to contribute in colorectal cancer (CRC), however, the topic is still controversial. Varying detection rate of JCPyV genome has been reported mainly due to technical reasons. Here, we provide summative data on the topic, with emphasize on technical issues. METHODS: Formalin-fixed paraffin-embedded tissue samples from 50 patients with CRC, consisting of tumoral and non-cancerous marginal tissue (totally 100 samples) were included in the study. After DNA extraction, specific JCPyV T-Ag sequences were targeted using Real-time PCR. To unwind the supercoiled JCPyV genome, pretreatment with topoisomerase I, was applied. Immunohistochemical (IHC) staining was performed using an anti-T-Ag monoclonal antibody. RESULTS: In the first attempts, no samples were found to be positive in Real-time PCR assays. However, JCPyV sequences were found in 60% of CRC tissues and 38% of non-cancerous colorectal mucosa after application of pre-treatment step with topoisomerase I enzyme (P = 0.028). T-Ag protein was found in the nuclear compartment of the stained cells in IHC assays. CONCLUSIONS: The presence of JCPyV in CRC tissues, as well as T-Ag localization in the nucleolus, where its oncogenic effect takes place, may provide supporting evidence for JCPyV involvement in CRC development. The study highlights the importance of using topoisomerase I to enhance JCPyV genome detection. Also, colorectal tissue is one of the permissive human tissue for JC resistance after preliminary infection.

Helicobacter pylori infection and autoimmune diseases; Is there an association with systemic lupus erythematosus, rheumatoid arthritis, autoimmune atrophy gastritis and autoimmune pancreatitis? A systematic review and meta-analysis study.

Autoimmune diseases are considered as one of the most important disorders of the immune system, in which the prolonged and chronic processes eliminate self-tolerance to the auto-antigens. The prevalence of autoimmune diseases has been increasing worldwide in the recent years. According to the literature, biological processes such as the host genome, epigenetic events, environmental condition, drug consumption, and infectious agents are the most important risk factors that make the host susceptible to the development of autoimmune diseases. In the recent years, the role of Helicobacter pylori in the induction of autoimmune diseases has attracted extensive attention. Via molecular mimicry, epitope spreading, bystander activation, polyclonal activation, dysregulation in immune response, and highly immune-dominant virulence, such as cagA, H. pylori causes tissue damage, polarity, and proliferation of the host cells leading to the modulation of host immune responses. Moreover, given the large population worldwide infected with H. pylori, it seems likely that the bacterium may develop into autoimmune diseases through dysregulation of the immune response. The frequency and relationship between H. pylori infection and systemic lupus erythematosus, rheumatoid arthritis, autoimmune atrophy gastritis, and autoimmune pancreatitis were evaluated using the data from 43 studies involving 5052 patients. According to statistical analysis it is probable that infection with more virulent strains of H. pylori (such as H. pylori cagA positive) can increase the risk of autoimmune diseases. In addition, it was shown that infection with H. pylori can prevent the development of atrophic gastritis by stimulating inflammation in the gastric antrum. However, future studies should confirm the validity of this study.

The case report of Mycobacterium arupense wound infection in diabetes mellitus patients; the first report and literature review.

Mycobacterium arupense is among the opportunist pathogens of atypical mycobacteria emergence (atypical mycobacteria) that is one of the isolated and reported environmental and clinical specimens. Numerous cases of osteo-articular infections of this bacterium are reported nowadays, while the pulmonary infection is rare. We identified Mycobacterium arupense in non-healing wound infection of an elderly woman with history of diabetes mellitus. She has negative tests for HIV, HBV and HCV, but was positive for HTLV-1. The patient was referred according to mild-fever, non-healing, destructive, and swelled lesion on her left foot. The mycobacterial wounds infection was suspected due to her non-conclusive previous treatment. The pathology, acid-fast staining, conventional and 16S rRNA sequencing confirmed the micro-organism to be M. arupense . Finally, the patient recovered following two-week consumption of clarithromycin, ethambutol and rifabutin. The results of this study provide evidence on the potential pathogenicity, clinical outcomes and treatment of infections caused by this bacterium.

Expression changes of cytotoxicity and apoptosis genes in HTLV-1-associated myelopathy/tropical spastic paraparesis patients from the perspective of system virology.

Although human T-cell lymphotropic virus type-1 (HTLV-1) was the first retrovirus among human pathogens to be identified, insufficient information on the pathogenesis of HTLV-1 infection means that no precise mechanism has yet been provided for HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Based on previous studies, it was found that apoptosis and inflammation stimulation were among the most important mechanisms underlying HAM/TSP. The present study provides an in-silico analysis of the microarray data related to HAM/TSP patients. Expression changes of the genes responsible for cytotoxicity and apoptosis processes of HAM/TSP patients and asymptomatic carriers were investigated. Expression of the genes involved in cytotoxicity and apoptosis in HAM/TSP patients was decreased; hence, a model was proposed indicating that the spread of immune responses in HAM/TSP may be due to expression of HTLV-1 virulence factors and the resistance of HTLV-1-infected cells to apoptosis.

What is the role of the thioredoxin antioxidant complex in relation to HAM/TSP?

We have little information about the definite role of the thioredoxin antioxidant complex system during viral infection, particularly during human T-cell lymphotropic virus type 1 (HTLV-1) infection and the HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) state. Therefore, we conducted comprehensive next-generation sequencing (NGS) analysis to determine Trx system expression changes in three categories of subjects: sero-negative HTLV-1 individuals, asymptomatic HTLV-1 people and HAM/TSP patients. We found that Trx capacity is reduced in the HAM/TSP state compared to healthy individuals, which indicates increasing inflammation and reduction of apoptosis, which might contribute to the progression of inflammation in the spinal cord, which in turn may develop into the HAM/TSP state.

High prevalence of blaCMY AmpC beta-lactamase in ESBL co-producing Escherichia coli and Klebsiella spp. clinical isolates in the northeast of Iran.

OBJECTIVE: The production of ß-lactamase enzymes such as AmpC ß-lactamases and extended-spectrum ß-lactamases (ESBLs) is among the main mechanisms for resistance to expanded-spectrum cephalosporins. The present study was conducted to investigate the prevalence and molecular epidemiology of plasmid-mediated AmpC beta (ß)-lactamase in ESBL co-producing Escherichia coli (E. coli) and Klebsiella spp. (Klebsiella pneumoniae and Klebsiella oxytoca) clinical isolates in the northeast of Iran. METHODS: A total of 602 E. coli and Klebsiella spp. clinical isolates were collected from three hospitals in Mashhad (northeast of Iran). A combination disk test (CDT) was performed for the phenotypic detection of ESBLs. Screening for the detection of AmpC ß-lactamases was performed by a susceptibility test to a cefoxitin disc among ESBL producing isolates. A confirmatory test for AmpC ß-lactamases was performed using the Mast® D68C test. Identification of plasmid-mediated AmpC cluster genes was done by multiplex polymerase chain reaction (PCR). RESULTS: Among 336 ESBL-producing strains, 230 (68.4%) isolates were resistant to cefoxitin. Results of the Mast® D68C test showed that 30% (69/230) of cefoxitin-resistant isolates simultaneously exhibited ESBL and AmpC activity and 22% (51/230) of isolates probably showed multi-drug resistant (MDR) phenotype. Results of multiplex PCR among ESBL-positive isolates showed that, 16.7% (56/336) of isolates were positive for plasmid-borneampC cluster genes, and CMY (38%) was the most frequent genotype of plasmid mediated AmpC. CONCLUSION: Findings of the study revealed that an increase in the prevalence of ESBL and AmpC co-producer in E. coli and Klebsiella spp. strains may become an important public health issue. Therefore, there is a vital need for surveillance of spread of these clinical isolates.

Thioredoxin reductase gene expression and activity among human T-cell lymphotropic virus type 1-infected patients.

BACKGROUND: The thioredoxin (Trx) system is a reducing complex, consisting of Trx, Trx reductase (TrxR), and NADPH, that scavenges reactive oxygen species. The system is a natural protective mechanism to prevent apoptosis and progression of oxidative stress-related diseases. The present study was conducted to explore possible changes in TrxR activity and gene expression as a response to the oxidative stress during HTLV-1 infection. MATERIALS AND METHODS: Blood samples were collected from 40 HTLV-1-infected patients and 40 age- and sex-matched healthy controls. The patient group consisted of chronic asymptomatic carriers and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM-TSP) patients. A commercial kit was used to measure the TrxR enzyme activity, and real-time polymerase chain reaction was performed to evaluate TrxR gene expression in extracted peripheral blood mononuclear cells (PBMCs). RESULTS: A decreasing pattern of TrxR enzyme activity was observed among control, carrier, and HAM-TSP groups (mean ± SD; controls, 0.1734 ± 0.056; carriers, 0.134 ± 0.065; and HAM-TSP, 0.0928 ± 0.047 µmol/min/mL). Cellular TrxR gene expression showed the same decreasing trend. The fold differences of gene expression in carriers and HAM-TSP groups compared with healthy controls were 0.8 and 0.7 vs 1, respectively. CONCLUSION: We found a reduction in TrxR expression as well as serum enzyme activity in HTLV-1-infected individuals, particularly in HAM-TSP patients. The reduced TrxR activity during HTLV-1 infection may hamper the natural protective mechanisms, thereby contributes to virus-induced complications.

Detection of JC Polyomavirus tumor antigen in gastric carcinoma: a report from Iran.

BACKGROUND AND OBJECTIVES: Factors contributing to development of gastric cancer are still under investigation. The JC Virus (JCV), as an oncogenic virus, has been indicated to play a possible role in gastric carcinogenesis. Theoretically, tumor antigen (T-Ag), the viral transforming protein, is capable of binding and inactivating tumor suppressor proteins p53 and pRb, there by promoting cancer development although such a role in gastric cancer is still controversial and additional data is needed to reach a definite conclusion. The prevalence of the virus varies in different geographic regions, therefore, we aimed to investigate JCV presence in cancerous gastric tissues of Iranian patients. MATERIALS AND METHODS: Thirty-one paired samples were included in this study (total of 62 samples). T-Ag sequences were investigated using real-time PCR in formalin fixed paraffin embedded (FFPE) tissue samples from the tumor site and relevant adjacent non-cancerous tissues (ANCT). In positive samples, JCV copy number (viral load) was also measured using real-time PCR. To evaluate T-Ag protein expression, immunohistochemistry examination was performed using an anti-T-Ag specific antibody. RESULTS: JCV sequences were detected in 17 out of 31 gastric cancer tissue samples (54.84%) and in 10 out of 31 of the non-cancerous adjacent gastric mucosa (32.25%) (Odds ratio of 2.4). Viral load in tumoral and adjacent tissue samples was not statistically different (p=0.88). Immunohistochemical study confirmed presence of JC T-Ag in the nuclear compartment. CONCLUSION: We showed the presence of the JC virus in gastric carcinoma tissue samples in our geographic region. This finding provides supportive data for a possible contribution of JCV in gastric cell transformation to malignancy. However, we highly recommend additional investigations to further explore JC virus and gastric cancer in order to reach a conclusion.

Detection of Porcine Bocavirus From a Child With Acute Respiratory Tract Infection.

Porcine bocavirus is a recently discovered virus classified within the Bocavirus genus. We present a case of upper respiratory tract infection associated with porcine bocavirus in a 3-year-old child who was in close contact with hogs in northeastern Iran. To the best of our knowledge, this is the first report on the human porcine bocavirus infection.

Prevalence of Toxoplasma and Echinococcus IgG antibodies in slaughterhouse workers, a serosurvey in Northeast Iran

Cystic echinococcosis and toxoplasmosis are two human parasitic diseases. Butchers and slaughterhouse workers are in close contact with body fluids and tissues of ruminants. To investigate the prevalence of Toxoplasma and Echinococcus IgG antibodies in slaughterhouse workers in Northeast of Iran, Mashhad, 2016. This cross-sectional study was performed on all personnel working at the largest industrial slaughterhouse of Khorasan Province. Serum samples were taken and kept frozen until used. IgG against Echinococcus and Toxoplasma were quantified using commercial ELISA kits. A questionnaire addressing possible risk factors of infection acquisition was filled by participants. Out of 91 male participants, 58.2% were positive for toxoplasmosis, 5.5% were positive for cystic echinococcosis and 3.3% were positive for both. Except using gloves, and gown and boots, other personal protective equipment (PPE) were not completely used by all personnel; mask 38%, tool disinfectants: 12%, face and hand disinfection: 14.3%. There was a high risk of Toxoplasma in slaughterhouse workers, however, such finding was not observed in Echinococcus parasite. Importance of PPE and tool disinfectants to reduce the risk of zoonotic infections in the workplace should be emphasised.

Designing and Construction Pcdna3.1 Vector Encoding Cfp10 Gene of Mycobacterium tuberculosis.

BACKGROUND: Pathogenic mycobacteria are a major cause of human morbidity and mortality. Mycobacterium tuberculosis is an etiological agent of human tuberculosis (TB). Designing new vaccines, including DNA vaccines, may be a useful strategy for preventing TB. OBJECTIVES: The purpose of this study was to design and construct an eukaryotic expression vector containing M. tuberculosis. MATERIALS AND METHODS: Genomic DNA of M. tuberculosis H37Rv cultured on Lowenstein Jensen medium was extracted, and cfp10 was amplified by PCR. After digesting the PCR product and the plasmid, the cfp10 fragment was ligated into the vector pcDNA3.1 (+). Correct insertion was confirmed by colony PCR, restriction enzyme digestion, and sequencing. RESULTS: Electrophoresis of the PCR product on gel showed a 303-bp target fragment. Colony PCR, restriction enzyme digestion, and Sequencing methods confirmed the accuracy of the gene cloning. Colony PCR and restriction enzyme digestion confirmed the cloning. CONCLUSIONS: Cloning of cfp10 of M. tuberculosis into an eukaryotic expression vector was performed successfully. We propose this recombinant plasmid for inducing immunity in animal models in future studies. This recombinant vector can also be used in the construction of fusion proteins.