Trends in cocirculation of oncogenic HPV genotypes in single and multiple infections among the unvaccinated community.

Cocirculation of multiple human papillomavirus (HPV) infections with low, probably high, and high-risk genotypes are to be associated with various grades of infections and cancer progression. The oncogenic high-risk HPVs are distributed and cocirculated throughout the world. This study was investigated to identify HPV genotypes related to genital disorders in unvaccinated women. The subjects were referred from clinics to a molecular lab for HPV testing in Iran as a low-coverage vaccinated country. HPVs DNAs of cervical scrapping and genital tissue specimens of 1,133 un-vaccinated women were genotyped using an in vitro diagnostic line probe (reverse hybridization) assay. In addition, phylogenetic trees were constructed on 100 MY09/MY11 polymerase chain reaction (PCR) amplicons of common genotypes of HPV L1 gene by Sanger sequencing. The mean age of the population study was 32.7 ± 8.0 and the mean age of HPV-positive cases was 31.6 ± 7.8. HPV DNA was detected in 57.8% (655/1133) of women subjects and 42.2% (478/1133) of cases were undetected. Among 655 HPV-positive cases, 639 subjects (56.4%) were related to defined genotypes and 16 subjects (1.4%) were untypeable. The highest prevalence rate of HPV genotypes was identified in the 25-34 years. The top 6 dominant HPVs in single and multiple genotypes were HPV6 (284/655 [43.4%]), HPV16 (111/655 [16.9%]), HPV31 (72/655 [11%]), HPV53 (67/655 [10.2%]), HPV11 (62/655 [9.5%]), and HPV52 (62/655 [9.5%]). Moreover, single, multiple and untypeable HPV genotypes were diagnosed as follows: 1 type (318/655 [48.5%]), 2 types (162/655 [24.8%]), 3 types (83/655 [12.7%]), 4 types (42/655 [6.5%]), more than 5 types (34/655 [5.3%]), and 1.4% un-typeable subjects. The sequenced partial L1 gene of HPV genotypes (GenBank databases under the accession numbers: MH253467-MH253566) confirmed and determined the cocirculated HPV genotypes' origins and addressed helpful insights into the future viral epidemiology investigations. Multiple HPV infections and cocirculation of various oncogenic HPV genotypes among the normal population (women and men) with asymptomatic forms are still challenging in unvaccinated communities. The preventive and organized surveillance programs for HPV screening are needed to be considered and compiled by health policy makers of low or unvaccinated countries.

The point mutation analysis of Cyp2C9*2 (Arg144Cys C>T), Cyp2C9*3 (Ile359Leu A>C) and VKORC1 (1639G>A) in women with cervical cancer related to HPV: A case-control study.

Human papillomavirus (HPV) is the most common sexually transmitted viral infection worldwide. HPV tumorigenesis genotypes are the causative agents of cervical cancer and genital malignancies. The scientific literature has demonstrated that life style, environmental, epigenetic accompanied with HR-HPV genotypes are potential risk factors for cervical cancer progression. The frequencies of the Cyp2C9*2 , Cyp2C9*3 , and vitamin K epoxide reductase complex subunit 1 (VKORC1) genotypes as potential molecular biomarkers have been investigated on Iranian women with cervical malignancy related to HPV genotypes. As a case-control study, the mutations were appraised using a polymerase chain reaction-restriction fragment length polymorphism procedure on women suffering from HPV infection (60 cases), CC (46 cases), and 40 subjects of as healthy control. The outcomes demonstrated that Cyp2C9*3  showed a meaningful relationship between women diagnosed with cervical cancer and the healthy population (AA vs. AC; OR, 7.15; 95% CI, 1.94-26.3; p = .003). It was also observed that the Cyp2C9*3 mutation in women with cervical cancer and VKORC1 in healthy population with HPV (+), did not follow the Hardy-Weinberg equilibrium. Our findings aid understanding the genetic polymorphism distribution of Cyp2C9*2 , Cyp2C9*3 , and VKORC1 in women with genital malignancies. This can also be useful in predicting the susceptibility risk factors for developing cervical cancer. However, allelic discrimination as a molecular biomarker requires further research.

Comparison of In-House Multiplex Real Time PCR, Diagcor GenoFlow HPV Array Test and INNO-LiPA HPV Genotyping Extra Assays with LCD- Array Kit for Human Papillomavirus Genotyping in Cervical Liquid Based Cytology Specimens and Genital Lesions in Tehran, Iran.

BACKGROUND: Human papillomavirus is a major etiologic agent for some human common cancers. Cervical precancer and cancer is the most prevalent dysplasia by HPV genotypes. Various rapid and sensitive methods have been developed into readily HPV genotyping. METHODS: In the present study, we compared the performance of Real Time PCR, GenoFlow HPV Array, and INNO-LiPA HPV Genotyping Extra Assays with LCD- Array. RESULTS: From 108 cervical samples, HPV was detected in 33 women (30.55%). Among detected HPV genotypes, HPV 6 and 11 were dominant genotypes. Comparing these methods revealed that for Real Time PCR, Genoflow, and INNO-LiPA in comparison with LCD Array, sensitivity and specificity were 94.2%, 93%; 76.7%, 93%; 64%, 96.5%, respectively. Overall, accuracy and precision of these methods were more than 80% and 90%, respectively. CONCLUSIONS: It seems that these methods are reliable and suitable for detection and genotyping of HPVs in cervical disorders and other dysplasia associated with human papillomaviruses.

Genotyping of human parechoviruses in Iranian young children with aseptic meningitis and sepsis-like illness.

Human parechoviruses (HPeV) are classified into 14 genotypes. HPeV1 and HPeV2 are the most prevalent genotypes in young children, which have been associated with mild to severe diseases. This study was conducted to investigate the involvement of HPeVs in aseptic meningitis and sepsis-like illness in Iran. Viral RNA was extracted from 148 cerebrospinal fluid samples from children <8 years old with primary diagnosis of aseptic meningitis and/or sepsis-like illness. Specific HPeV, HEV real-time PCR and HPeV typing were done to identify the infection rate of these viruses. HPeV and HEV were detected in 64 (43.24 %), 31 (20.94 %) of 148 patients with 10 (6.75 %) coinfection. VP1/VP3 junction region was successfully sequenced from 12 of the HPeV-positive specimens, and all of them were identified as HPeV1. HPeV was more prevalent than HEV in both aseptic meningitis and sepsis-like illness, so further studies are needed to understand the disease burden of HPeV infections, and clinical manifestations especially in specific illnesses of possible viral etiology. Direct detection of these viruses leads to reduce hospitalization and use of antibiotic, which are often followed by other complications in neonates and young children.

Frequency of human enterovirus 71 in children under 8 years old with aseptic menengitis in Tehran.

BACKGROUND: Human enterovirus 71 (HEV71) was isolated for the first time from an infant with encephalitis in California in 1969 and then spread through the world. It has emerged as a major cause of a vast variety of diseases such as epidemics of hand, foot and mouth disease (HFMD), aseptic meningitis (AM), acute flaccid paralysis, and encephalitis. The aim of this study was to determine the prevalence of enterovirus 71 in children < 8 years old who were hospitalized due to primary diagnosis of AM in Tehran. METHODS: One hundred cerebrospinal fluid samples (CSF) were collected by physicians from children with a diagnosis of AM and transported on ice to the Pasteur Institute of Iran for further processing. Viral RNA was extracted and EV71 infection was detected by RT-PCR method using the specific primers. RESULTS: EV71 infection was detected in 14 patients (14%). Eight (57.14%) patients were younger than 2 years old, 11 (78.57%) were male and 3 (21.43%) were female. The seasonal peaks of EV71 were observed during autumn and winter with 6 (42.86%) and 5 (35.71%) cases, respectively. CONCLUSIONS: EV71 should be considered as a causative agent of AM in Iran with the epidemiological pattern similar to that of other enteroviruses as males are more susceptible to be affected by these viruses. Further studies on this virus are needed to improve our knowledge about them in our country.

Lead (Pb) exposure from outdoor air pollution: a potential risk factor for cervical intraepithelial neoplasia related to HPV genotypes.

Human papillomavirus genotypes (HPVs) have been confirmed to be the major cause of cervical intraepithelial neoplasia (CIN) that remains to be one of the most common women cancers around the world. It seems other risk factors have synergistic effects on cervical cancer occurrence including smoking, dietary pattern, sexual behavior, ethnicity, epigenetics, and environmental hazardous materials. Our study characterized the potential cancerous role of lead (Pb) as a common toxic environmental pollutant agent on CIN outcomes. Lead concentration was quantified using an atomic absorption spectrometer in liquid-based cytology specimens of 40 CIN-HPV positive subjects, 50 HPV infected non-cancerous cases, and 43 non-HPV infected/non-cancerous women. Pb concentration was 5.5 (4.7-6.4) µg/dL, 4.7 (4.2-8.7) µg/dL, and 4.7 (4.5-5.4) µg/dL in the CIN-HPV positive group, HPV infected non-cancerous cases, and non-HPV infected/non-cancerous group, respectively. The results showed higher Pb concentration is associated with higher risk for cervical malignancy in comparison with non-HPV infected/non-cancerous subjects, after controlling for age effect (aOR = 4.55, 95% CI: 1.55-15.07, P < 0.01). Our finding suggested a direct significant association between Pb accumulation and CIN existence. The consequences need to be further validated by including more relevant risk factors and controlling the confounders for better understating of Pb impact from outdoor air pollution on cervical cancer progression.

Characteristics of HSV-2, M. genitalium and C. trachomatis in HPV Genotypes Associated with Cervical Intraepithelial Neoplasia and Genital Infections.

BACKGROUND: Cervical cancer is the most common cancer in women. High-Risk HPV types are known as the main agents involved in genital and cervical malignancies. There may be co pathogens like STIs that are involved in enhancing the susceptibility and progression to cervical neoplasia. This study was conducted to detect C. trachomatis, HSV-2 and M. genitalium using qPCR in women suffering from cervical intraepithelial neoplasia, HPV infection and non cancerous- non HPV subjects for the association of burden of genital disorders. MATERIALS AND METHODS: This descriptive study was performed on 195 Liquid Based Cytology (LBCs) specimens collected from women referred to private laboratories. Fifty, 98 and 47 samples were from women with known CIN, HPV positive and non-cancerous/non-HPV, respectively. HSV-2, C. trachomatis, M. genitalium and HPV genotypes have been detected using multiplex TaqMan Real Time PCR and PCR hybridization. RESULTS: A total of 148 HPV positive samples were included. HPVs 6 (35.13%), 16 (32.43%), 18 (21.62%), 11 (9.46%), 31 (9.46%), and 51 (9.46%) were the most common genotypes. Single, 2, 3, and more than 4 multiple HPV genotypes were detected in 46%, 29.7%, 14.2%, 10.1% cases, respectively. The prevalence of M. genitalium, C. trachomatis and HSV2 was 3 (1.54%), 24 (12.3%) and 1(0.5%), respectively. There were no statistically significant differences between these pathogens and cervical intraepithelial neoplasia (p≥ 0.05). CONCLUSION: HR-HPV genotypes were more prevalent in genital infections and cervical cancer. It would seem early detection of dominant STI pathogens such as Chlamydia spp. gains due to effective prevention. Here, further research is needed to understand the co-infections burden of HPV genotypes with STIs in clinical manifestations.

Efficacy of Loop-Mediated Isothermal Amplification for H. pylori Detection as Point-of-Care Testing by Noninvasive Sampling.

For targeted eradication of Helicobacter pylori (H. pylori) to reduce gastric cancer burden, a convenient approach is definitely needed. The purpose of this study was to evaluate the LAMP assay for H. pylori detection using samples collected by noninvasive and self-sampling methods. The available LAMP assay for H. pylori detection was appraised and verified using reference and clinically isolated H. pylori strains. In addition, a clinical study was conducted to assess the LAMP assay on 51 patients, from whom saliva, oral brushing samples, feces, corpus, and antrum specimens were available. Clarithromycin resistance was also analysed through detection of A2143G mutation using the LAMP-RFLP method. The validation and verification analysis demonstrated that the LAMP assay had an acceptable result in terms of specificity, sensitivity, reproducibility, and accuracy for clinical settings. The LAMP assay showed a detection limit for H. pylori down to 0.25 fg/µL of genomic DNA. An acceptable consensus was observed using saliva samples (sensitivity 58.1%, specificity 84.2%, PPV 85.7%, NPV 55.2%, accuracy 68%) in comparison to biopsy sampling as the gold standard. The performance testing of different combinations of noninvasive sampling methods demonstrated that a combination of saliva and oral brushing could achieve a sensitivity of 74.2% and a specificity of 57.9%. A2143G mutation detection by LAMP-RFLP showed perfect consensus with Sanger sequencing results. It appears that the LAMP assay in combination with noninvasive and self-sampling as a point-of-care testing (POCT) approach has potential usefulness to detect H. pylori infection in clinic settings and screening programs.

Emergence of African swine fever virus, northwestern Iran.

In 2008, African swine fever was introduced into Georgia, after which it spread to neighboring Armenia, Azerbaijan, and the Russian Federation. That same year, PCR and sequence analysis identified African swine fever virus in samples from 3 dead female wild boars in northwestern Iran. Wild boars may serve as a reservoir.

Brucellosis: The Current Situation in Iran and Neighboring Countries.

It is frequently discussed that Brucella spp is an endemic infection in Iran and the Middle Eastern countries. The average incidence of brucellosis rate has been increasing and the zoonotic infections like brucellosis are being easily transmitted in these lands through the trafficking of livestock and chattels. A survey has been conducted to evaluate the pitfalls surrounding the diagnostic and clinical exposures of brucellosis in developing and undeveloped countries.

miR-21 and miR29-a: Potential Molecular Biomarkers for HPV Genotypes and Cervical Cancer Detection.

BACKGROUND AND OBJECTIVE: This study aimed to assess miR21 and miR-29a diagnostic usefulness for HPV infection and Cervical Cancer (CC) detection. METHODS: miRs extraction has been performed on HPV-positive, CC and negative control of 43, 50 and 46 individuals, respectively. The expression level of miRs has been analyzed using Real-Time RT-PCR. RESULTS: There was a correlation between the expression of miRs and the Receiver Operating Characteristic (ROC) analysis in CC and HPV-positive patients. CONCLUSION: The correlation between expressions of miRs is notable. ROC curve analysis could be applied in molecular diagnostic exposure for CC related to HPV infections.

The Impact of MTHFR 1298 A > C and 677 C > T Gene Polymorphisms as Susceptibility Risk Factors in Cervical Intraepithelial Neoplasia Related to HPV and Sexually Transmitted Infections.

BACKGROUND: HPV genotypes are the most common etiological factor for genital neoplasia. It would appear that sexually transmitted infections accompanied with HPV genotypes might have synergistic interactions in cancer progression. The genetic polymorphisms are involved in metabolizing carcinogens which may contribute to the susceptibility of developing genital cancers by less efficient or overly down metabolic pathways and cell signaling. MTHFR polymorphisms are related to several metabolic disorders and human cancers. We investigated the contribution of MTHFR 1298 and MTHFR 677 polymorphisms as potential risk factors for outcomes with HPV genotypes and STIs in Iranian population. MATERIALS AND METHODS: As a case-control study, MTHFR A1298C and C677T were assessed for SNPs analysis using a PCR-RFLP assay in 50 cervical intraepithelial neoplasia (CIN) cases, 98 HPV-positive subjects and 47 non-cancerous/non-HPV patients as healthy controls. RESULTS: Finding suggested a significant association between the MTHFR 1298 CC polymorphisms (OR = 3.5, 95% CI = 1.13-10.82, P ≤ 0.05) in women with CIN as compared to non-cancerous/non-HPV subjects. There was not a significant difference of MTHFR 677 between outcomes. DISCUSSION: It would seem MTHFR 1298 CC is more likely to be a potential risk factor for HPV-cervical cancer progression. Consequences support further attempts to understand the clinical manifestations of neoplasia related to genital infections and gene mutations.

In silico characteristics for re-emerging possibility of Vibrio cholerae genotypes in Iran.

Epidemic cholera has been registered several times within recent years in Iran. The dominant genotype was Ogawa until 2011, but this gradually changed to Inaba. However, in 2015, the re-appearance of a previous Ogawa genotype was detected by the Iranian CDC. This raised worries because no evidence was found for its origin abroad. The aim of the present study was to identify clearly the source of this outbreak. Pulsed field gel electrophoresis (PFGE) was used to compare the recently detected Vibrio cholerae strains with those isolated from 2011 to 2015. We selected one strain per PFGE pattern, and compared the distinct patterns. BioNumerics software was applied, which enables interpretation of phenotypic and genotypic differences. In total, we studied 33 V. cholerae Ogawa strains. Analysis showed that strains could be discriminated on the basis of annual clusters but with a similarity of more than 80%. The highest homology was observed among those isolated each year from 2011 to 2014. In contrast, strains isolated in 2015 also exhibited close correlation with each other but were located in distinct clusters. The analysis also proved genetic variations among some strains. All 2015 strains showed differences with regard to previous genotypes despite some similarities. The new genotypes were probably imported into Iran from neighbouring countries such as Iraq by travellers or contaminated food sources since 2015. However, more investigations are required to identify the exact source of the 2015 outbreak.

Deregulation of miR-21 and miR-29a in Cervical Cancer Related to HPV Infection.

BACKGROUND: Early diagnosis is an important factor to improve the survival of Invasive Cervical Cancer (ICC) patients. Molecular biomarkers such as micro RNA (miRNA) can be used in the early detection of ICC. The expression of miR-21 and miR-29a are deregulated in many types of human cancers. OBJECTIVE: The aim of this study was to investigate the differences in miR-21 and miR-29a expression patterns in the Human Papilloma Virus (HPV) infection and various grades of cervical cancer among Iranian women. METHODS: Small RNAs were extracted from positive for HPV, cervical cancer and healthy samples from 43, 50 and 46 individuals, respectively. Expression levels of miR-21 and miR-29a were analyzed by SYBR Green real-time RT-PCR using specific primers, and 5s rRNA as the internal reference gene. RESULTS: Results have shown a significant increase in miR-21 and decrease in miR-29 in cancerous samples in comparison with the control groups (P < 0.0001). CONCLUSION: This study illustrated that miR-21 and miR-29a could be operated as an oncogene and tumor-suppressor in cervical cancer progression. More studies are needed to demonstrate the role of miR-21 and miR-29a as potential biomarkers for the diagnosis of cervical cancer in future investigations.

Possible Synergistic Interactions Among Multiple HPV Genotypes in Women Suffering from Genital Neoplasia

Objective: Persistence of HPV infection is the true cause of cervical disorders. It is reported that competition may exist among HPV genotypes for colonization. This survey was designed to establish the multiple HPV genotype status in our community and the probability of multiple HPV infections involvement. Methods: All multiple HPV infections were selected for investigation in women suffering from genital infections referred to private laboratories in Tehran, Iran. A total of 160 multi HPV positive specimens from cervical scraping were identified by the HPV genotyping methods, "INNO-LiPA and Geno Array". Result: In present study, HPV 6 (LR), 16 (HR), 53 (pHR), 31 (HR) and 11 (LR) were included in 48.8% of detected infections as the most five dominant genotypes. HPV 16 was detected at the highest rate with genotypes 53, 31 and 52, while HPV 53 appeared linked with HPV 16, 51 and 56 in concurrent infections. It appears that HPV 16 and 53 may have significant tendencies to associate with each other rather than with other genotypes. Analysis of the data revealed there may be some synergistic interactions with a few particular genotypes such as "HPV 53". Conclusion: Multiple HPV genotypes appear more likely to be linked with development of cervical abnormalities especially in patients with genital infections. Since, there are various patterns of dominant HPV genotypes in different regions of world, more investigations of this type should be performed for careHPV programs in individual countries.

Characterization of JAK2 V617F (1849 G > T) Mutation in Cervical Cancer Related to Human Papillomavirus and Sexually Transmitted Infections.

BACKGROUND: Human papillomavirus (HPV) genotypes that infect the genital tract play a main etiologic role in cervical cancer progression. Other environmental factors, such as sexually transmitted diseases and the host genetic pattern, contribute to infection persistence of the uterus and cervical epithelium in sustaining their malignancy. The Janus kinase 2 is a non-receptor tyrosine kinase in cell signaling process of tumor genesis. In the present study, JAK2 V167F mutation was distinguished in women with sexually transmitted infections, such as Herpes simplex virus 2, Chlamydia trachomatis and Mycoplasma genitalium and cervical cancer. METHODS: This case-control survey was performed on 195 liquid based cytology of women specimens. Fifty, 98, and 47 samples were from women with known cervical cancer, HPV positive and HPV negative, respectively. Single nucleotide polymorphism analysis, sexually transmitted infections detection and HPV genotyping were carried out using approved PCR- RFLP, in-house multiplex TaqMan Real Time PCR and the reverse dot blot hybridization assay. RESULTS: HPVs 6, 16, 18, 11, 31, and 51 were the most common genotypes. The prevalence rate of multiple HPV genotypes was 46.0% to 10.1%. Analysis of JAK2 V617F (1849 G > T) showed that prevalence of mutation was GG (65.1%), GA (34.9%), and TT (0%), respectively. There were no statistically significant differences between this mutation and variables of population survey (P ≥ 0.05). CONCLUSIONS: The molecular epidemiology study on the genetic polymorphisms, i.e., JAK2 V617F and other single nucleotide polymorphisms as a diagnostic tool is necessary for cancer screening and prophylactic programs.

Glutathione S-transferase omega gene polymorphism as a biomarker for human papilloma virus and cervical cancer in Iranian women

Objective: Human papillomavirus (HPV) infection is an important sexually-transmitted infection worldwide. Persistent infections with different high-risk HPV genotypes may cause cervical intraepithelial neoplasia and cervical cancer. Single nucleotide polymorphisms of glutathione S-transferase omega (GSTO) 1 and 2 play an important role in cancer progression. To evaluate GSTO gene polymorphism influence on women's susceptibility to low-risk or high-risk HPV infections and also risk of cervical cancer development. Material and Methods: We examined 50 patients with cervical cancer, 43 patients who were positive for HPV, and 43 healthy individuals as negative controls. We used polymerase chain reaction-restriction fragment length polymorphism to determine GSTO1 A140D and GSTO2 N142D variants in study participants. Results: We found a significant association between the GSTO1 A140D gene polymorphism and HPV 6, 16, 18, 16/18 infections and cervical cancer in Iranian women. We noted a significant difference for the 140AD/142NN combination genotype between patients in the cervical cancer group and healthy controls. There were no significant differences for the GSTO2 N142D genotype and allele frequencies between the patient (i.e., cervical cancer and HPV-positive) groups and controls. Conclusion: The 140AD genotype, 140D allele, and 140AD/142NN combination genotype seem to confer a protective property in women's susceptibility to HPV 6, 16, 18, 16/18 infections and cervical cancer. However, the GSTO2 N142D polymorphism is not associated with HPV infections and cervical cancer. It would appear that GSTO1 A140D SNPs likely play a role in the level of susceptibility to HPV-related cervical cancer.

Cervical Cancer and Genital Infections: Assessment of Performance and Validation in Human Papillomavirus Genotyping Assays in Iran, its Neighbouring Countries and Persian Gulf Area.

BACKGROUND: The accuracy of diagnostic assays in Human Papillomavirus (HPV) genital infection and cervical cancer has remained a clinical challenge in diagnosis. Evidence indicates that a large proportion of cervical cancer can be prevented through organized care for HPV and testing. Countries with low per capita income, such as Iran and its neighbours, have no national organized program for cervical cancer screening and vaccination. The aim of this study was to review recent published papers in this region for evaluating the efficacy of released data regarding HPV genotyping system in genital infections and cervical cancer. METHODS: Investigating various medical search engines retrieved 46 reports, mostly after 2010, consisting of either home brew protocols or commercial technologies in this field. RESULTS: Summarized results demonstrated that except a few cases, all reports were limited studies performed in confined populations focusing on attending patients at clinics for regular checkups. In the present study, 52.8% of papers were from Iran and the rest belonged to other countries. The rate of HPV infection was reported in the range of 0.62% to 25% in the normal population, while it varied from 18.75% to 100% in females with cervical cancer. In HPV genotyping surveys, only 26.1 % (12/46) of reports had validated and World Health Organization (WHO) proficient procedures. Also, multiple infections were not mentioned in 56.52% (25/46) of researches. CONCLUSIONS: Employing reliable genotyping methods is the best way for regular screening of cervical cancer related to HPV and precancerous diseases in females of these areas. The focus of most surveys was to come up with the best national policies for establishing a preventive program in Iran and Persian Gulf area.