Species-Specific Detection and Identification of Fusarium Species Complex,the Causal Agent of Sugarcane Pokkah Boeng in China

Background

Pokkah boeng disease caused by the Fusarium species complex results in significant yield losses in sugarcane. Thus, the rapid and accurate detection and identification of the pathogen is urgently required to manage and prevent the spreading of sugarcane pokkah boeng.

Methods

A total of 101 isolates were recovered from the pokkah boeng samples collected from five major sugarcane production areas in China throughout 2012 and 2013. The causal pathogen was identified by morphological observation, pathogenicity test, and phylogenetic analysis based on the fungus-conserved rDNA-ITS. Species-specific TaqMan real-time PCR and conventional PCR methods were developed for rapid and accurate detection of the causal agent of sugarcane pokkah boeng. The specificity and sensitivity of PCR assay were also evaluated on a total of 84 isolates of Fusarium from China and several isolates from other fungal pathogens of Sporisorium scitamineum and Phoma sp. and sugarcane endophyte of Acremonium sp.

Result

Two Fusarium species (F. verticillioides and F. proliferatum) that caused sugarcane pokahh boeng were identified by morphological observation, pathogenicity test, and phylogenetic analysis. Species-specific TaqMan PCR and conventional PCR were designed and optimized to target their rDNA-ITS regions. The sensitivity of the TaqMan PCR was approximately 10 pg of fungal DNA input, which was 1,000-fold over conventional PCR, and successfully detected pokkah boeng in the field-grown sugarcane.

Conclusions/Significance

This study was the first to identify two species, F. verticillioides and F. proliferatum, that were causal pathogens of sugarcane pokkah boeng in China. It also described the development of a species-specific PCR assay to detect and confirm these pathogens in sugarcane plants from mainland China. This method will be very useful for a broad range of research endeavors as well as the regulatory response and management of sugarcane pokkah boeng.     

Technical and Regulatory Shortcomings of the TaqMan Version 1 HIV Viral Load Assay

Background

The lower limit of detection of the original Roche Amplicor HIV plasma viral load (pVL) assay (50 copies/mL) has defined HIV treatment success. The Amplicor assay, however, has been replaced by the Roche TaqMan assay(s). Changes to the limits of detection and calibration have not been validated for clinical utility. Sudden increases in the number of patients with detectable pVL have been reported following the introduction of the TaqMan version 1 assay.

Methods

Between October 2009 and April 2010 all routine pVL samples from British Columbia, Canada, with 40–250 copies/mL by TaqMan were re-tested by Amplicor (N = 1198). Subsequent short-term virological and resistance outcomes were followed in patients with unchanged therapy (N = 279; median 3.2 months follow-up).

Results

TaqMan and Amplicor values correlated poorly at low pVL values. Low-level pVL by TaqMan was not associated with impending short-term virological failure; only 17% of patients with 40–250 copies/mL by TaqMan had detectable pVL by Amplicor at follow-up. During the follow-up period only 20% of patients had an increase in pVL by TaqMan (median [IQR]: 80 [36–283] copies/mL). In addition, in ∼2.4% of samples pVL was dramatically underestimated by TaqMan due to poor binding of the proprietary TaqMan primers.

Conclusions

The replacement of Amplicor with the TaqMan assay has altered the previously accepted definition of HIV treatment failure without any evidence to support the clinical relevance of the new definition. Given the systematic differences in measurement in the low pVL range the British Columbia HIV treatment guidelines now use a threshold of >250 copies/mL by TaqMan to define treatment failure.     

Cobas Ampliprep/Cobas TaqMan HIV-1 v2.0 Assay: Consequences at the Cohort Level

Background

High-sensitive real-time PCR assays are routinely used to monitor HIV-1 infected subjects. Inter-assay discrepancies have been described at the low viral load (VL) end, where clinical decisions regarding possible virological rebound are based.

Methods

A retrospective study was performed to analyze frequencies of viral blips after transition to the COBAS Ampliprep/COBAS TaqMan v2.0 HIV-1 assay (Taqman v2.0) in patients with prior undetectable VLs as measured with the Roche Cobas Ampliprep Amplicor HIV-1 Monitor Test, v1.5 (Amplicor) and was evaluated in comparison to a group of patients monitored with the Abbott Real-time HIV-1 assay (Abbott RT) during the same period of time.

Results

85 of 373 patients with VLs below the limit of quantification with Amplicor had VLs >50 copies/mL after transition to the TaqMan v2.0 assay. Among these 74.1% had VLs ranging from 50–499 copies/mL, 22.9% had VLs >500 copies/mL. From 22 patients with initial Taqman v2.0 based VLs exceeding 500 copies/mL, 6 patients had VLs <20 copies/mL after novel VL measurement on a next visit. In our control group with VL quantification using the Abbott RT assay, only 1 patient became detectable and showed a VL of <40 copies/mL after new measurement.

Conclusions

Transition to the Taqman v2.0 assay was accompanied by an increase of quantifiable HIV-1 VLs in patients with long term viral suppression under antiretroviral therapy that might be attributed to technical shortcomings of the Taqman v2.0 assay. A high test variability at the low VL end but also beyond was observed, making meaningful clinical interpretation of viral blips derived from different assays difficult.     

HPV Testing by cobas HPV Test in a Population from Catalonia

Background

HPV testing in cervical cancer screening has been proposed as an alternative or complementary to cytology in women older than 30 years. However, adequate clinical sensitivity and specificity are crucial for a new test to be implemented. Hybrid Capture 2 (HC2) has proved good clinical performance in selecting women at risk for high-grade intraepithelial lesions with a high sensitivity and specificity. cobas HPV Test has been recently launched and its performance in different clinical settings needs to be determined.

Objectives

The aim of this study was to evaluate the cobas HPV Test for the detection of cervical HPV infection in a population of women in Catalonia (Spain) using HC2 as a reference.

Materials and Methods

Cervical liquid cytology samples from 958 women have been studied. Sensitivity was analyzed in 60 samples from patients with a high-grade intraepithelial lesion (≥CIN2) on histology and specificity was determined in 898 samples from women with no ≥CIN2. All cases had HC2 and cobas HPV Test performed. Statistical analyses of sensitivity, specificity and comparison between HC2 and cobas HPV Test by a non-inferiority test were applied.

Results

Sensitivity of HC2 and cobas HPV Test for detecting ≥CIN2 proved identical (98.3%) while specificity was 85.3% and 86.2% respectively. The non-inferiority test demonstrated that cobas HPV Test surpassed 90% sensitivity and 98% specificity of HC2.

Conclusion

The cobas HPV Test results fulfilled sensitivity and specificity requirements for HPV based cervical cancer screening and for the triage of minor cytological abnormalities, allowing its introduction in clinical settings.     

Prevalence of Human Papillomavirus (HPV) in Oesophageal Squamous Cell Carcinoma in Relation to Anatomical Site of the Tumour

Background

The prevalence and role of human papillomavirus (HPV) in the aetiology of oesophageal squamous cell carcinoma is uncertain. Based on the presence of HPV in the oral cavity and its causal association with squamous cell carcinoma of the oropharynx, we hypothesised that HPV is more strongly associated with proximal than distal oesophageal squamous cell carcinoma.

Methods

A population-based study comparing HPV infection in relation to tumour site in patients diagnosed with oesophageal squamous cell carcinomas in the Stockholm County in 1999–2006. Multiplex polymerase chain reaction genotyping (PCR) with Luminex was conducted on pre-treatment endoscopic biopsies to identify type specify HPV. Carcinogenic activity of HPV was assessed by p16^INK4a expression. Multivariable logistic regression was used to calculate odds ratios and 95% confidence intervals.

Results

Among 204 patients, 20 (10%) had tumours harbouring HPV DNA, almost all (90%) of HPV high-risk type, mainly HPV16. Tumours containing HPV were not overrepresented in the upper compared to the middle or lower third of the oesophagus (odds ratio 0.6, 95% confidence interval 0.2–1.9). P16^INK4a expression was similarly common (24% and 16%) in the HPV-positive and HPV-negative groups.

Conclusion

This study found a limited presence of HPV in oesophageal squamous cell carcinoma of uncertain oncogenic relevance and did not demonstrate that HPV was more strongly associated with proximal than distal tumours.     

High-resolution melting molecular signatures for rapid identification of human papillomavirus genotypes

Background

Genotyping of human papillomarvirus (HPV) is crucial for patient management in a clinical setting. This study accesses the combined use of broad-range real-time PCR and high-resolution melting (HRM) analysis for rapid identification of HPV genotypes.

Methods

Genomic DNA sequences of 8 high-risk genotypes (HPV16/18/39/45/52/56/58/68) were subject to bioinformatic analysis to select for appropriate PCR amplicon. Asymmetric broad-range real-time PCR in the presence of HRM dye and two unlabeled probes specific to HPV16 and 18 was employed to generate HRM molecular signatures for HPV genotyping. The method was validated via assessment of 119 clinical HPV isolates.

Results

A DNA fragment within the L1 region was selected as the PCR amplicon ranging from 215–221 bp for different HPV genotypes. Each genotype displayed a distinct HRM molecular signature with minimal inter-assay variability. According to the HRM molecular signatures, HPV genotypes can be determined with one PCR within 3 h from the time of viral DNA isolation. In the validation assay, a 91% accuracy rate was achieved when the genotypes were in the database. Concomitantly, the HRM molecular signatures for additional 6 low-risk genotypes were established.

Conclusions

This assay provides a novel approach for HPV genotyping in a rapid and cost-effective manner.     

Ambient Stable Quantitative PCR Reagents for the Detection of Yersinia pestis

Background

Although assays for detecting Yersinia pestis using TaqMan probe-based real-time PCR have been developed for years, little is reported on room-temperature-stable PCR reagents, which will be invaluable for field epidemic surveillance, immediate response to public health emergencies, counter-bioterrorism investigation, etc. In this work, a set of real-time PCR reagents for rapid detection of Y. pestis was developed with extraordinary stability at 37°C.

Methods/Principal Findings

TaqMan-based real-time PCR assays were developed using the primers and probes targeting the 3a sequence in the chromosome and the F1 antigen gene caf1 in the plasmid pMT1of Y. pestis, respectively. Then, carbohydrate mixtures were added to the PCR reagents, which were later vacuum-dried for stability evaluation. The vacuum-dried reagents were stable at 37°C for at least 49 days for a lower concentration of template DNA (10 copies/µl), and up to 79 days for higher concentrations (≥10² copies/µl). The reagents were used subsequently to detect soil samples spiked with Y. pestis vaccine strain EV76, and 5×10⁴ CFU per gram of soil could be detected by both 3a- and caf1-based PCR reagents. In addition, a simple and efficient method for soil sample processing is presented here.

Conclusions/Significance

The vacuum-dried reagents for real-time PCR maintain accuracy and reproducibility for at least 49 days at 37°C, indicating that they can be easily transported at room temperature for field application if the machine for performing real-time PCR is available. This dry reagent is of great significance for routine plague surveillance.     

High viral load of human bocavirus correlates with duration of wheezing in children with severe lower respiratory tract infection

Background

Human bocavirus (HBoV) is a newly discovered parvovirus and increasing evidences are available to support its role as an etiologic agent in lower respiratory tract infection (LRTI). The objective of this study is to assess the impact of HBoV viral load on clinical characteristics in children who were HBoV positive and suffered severe LRTI.

Methods

Lower respiratory tract aspirates from 186 hospitalized children with severe LRTI were obtained by bronchoscopy. HBoVs were detected by real-time PCR and other 10 infectious agents were examined using PCR and/or direct fluorescent assay.

Results

Thirty-one patients (24.6%) were tested positive for HBoV in the respiratory tract aspirates. Fifteen samples had a high viral load (>10⁴ copies/mL) and the other sixteen samples had a low viral load (<10⁴ copies/mL). The duration of presented wheezing and hospitalization was longer in children with high viral load of HBoV than that in children with low viral load. The days of wheezing showed a correlation with viral load of HBoV.

Conclusion

We confirmed that HBoV was frequently detected in patients with severe LRTI. Wheezing was one of the most common symptoms presented by patients with positive HBoV. A high HBoV viral load could be an etiologic agent for LRTI, which led to more severe lower respiratory tract symptom, longer duration of wheezing and hospitalization.     

Prevalence, genotype distribution and persistence of human papillomavirus in oral mucosa of women: a six-year follow-up study

Background

Human papillomavirus (HPV) infections have been linked to a subset of oral and oropharyngeal cancers. However, little is known on the natural history of oral HPV infections. We designed the prospective Finnish HPV Family Study to assess the dynamics of HPV infections in parents and their infants. This study reports HPV genotype distribution and virus persistence in oral mucosa of the mothers.

Materials and Methods

Totally, 324 pregnant women were enrolled at the 3^rd trimester of pregnancy and followed-up for 6 years. Oral scrapings taken with a brush were collected and HPV-genotyping was performed with nested PCR and Multimetrix® test (Progen, Heidelberg, Germany). The predictors of persistent oral HPV species 7/9 infections were analyzed using generalized estimating equation models.

Results

The point prevalence of oral HPV varied from 15% to 24% during the 6-year follow-up. Altogether, 18 HPV genotypes were identified either as single or multiple-type oral infections. HPV16 was the most prevalent type at 9.7%–18.4%, followed by HPV18, HPV6, and multiple infections. Altogether, 74 women had persistent oral HPV infection determined as at least two consecutive samples positive with the same HPV genotype. HPV16 and HPV6 were the two most frequent types to persist (76% and 9%) for a mean of 18.6 and 20.2 months, respectively, followed by multiple infections (8%) for 18.3 months. An increased risk for persistent oral HPV infection with species 7/9 was associated with being seropositive for low-risk (LR)-HPV-types at baseline, whereas the use of oral contraceptives and a second pregnancy during follow-up were protective. Clinical oral lesions were detected in 17% of these women, one-third of whom had persistent oral HPV-infections.

Conclusion

HPV16 and HPV6 were the most common genotypes in oral HPV-infections and were also most likely to persist. Use of oral contraceptives and a second pregnancy protected against oral HPV persistence.     

mRNA and DNA Detection of Human Papillomaviruses in Women of All Ages Attending Two Colposcopy Clinics

Objective

HPV infection is a common finding, especially in young women while the majority of infections are cleared within a short time interval. The aim of this study was to examine the efficacy of HPV DNA and mRNA testing in a population attending colposcopy units of two University hospitals.

Methods

1173 liquid based cervical samples from two colposcopy clinics were tested for HPV DNA positivity using a commercial typing kit and HPV E6/E7 mRNA positivity with a flow cytometry based commercial kit. Statistic measures were calculated for both molecular tests and morphological cytology and colposcopy diagnosis according to histology results.

Results

HPV DNA, high-risk HPV DNA, HPV16 or 18 DNA and HPV mRNA was detected in 55.5%, 50.6%, 20.1% and 29.7% of the cervical smears respectively. Concordance between the DNA and the mRNA test was 71.6% with their differences being statistically significant. Both tests’ positivity increased significantly as lesion grade progressed and both displayed higher positivity rates in samples from women under 30 years old. mRNA testing displayed similar NPV, slightly lower sensitivity but significantly higher specificity and PPV than DNA testing, except only when DNA positivity for either HPV16 or 18 was used.

Conclusions

Overall mRNA testing displayed higher clinical efficacy than DNA testing, either when used as a reflex test or as an ancillary test combined with morphology. Due to enhanced specificity of mRNA testing and its comparable sensitivity in ages under 25 or 30 years old, induction of mRNA testing in young women could be feasible if a randomized trial verifies these results.     

Switching virally suppressed, treatment-experienced patients to a raltegravir-containing regimen does not alter levels of HIV-1 DNA

Background

Current HIV-1 antiretroviral therapy (ART) greatly reduces virus replication but does not significantly affect the viral reservoir. Raltegravir, a recently introduced integrase inhibitor, could, at least theoretically, reduce residual viremia in patients on ART and affect the viral reservoir size. The aim of this study was to assess whether switching therapy in treatment-experienced patients that were virally suppressed to a raltegravir-containing regimen reduces the size of the viral reservoir, and if such treatment leads to a change in levels of HIV 2-LTR circles in this patient group.

Methods

14 ART experienced individuals with a suppressed viral load (<50 HIV-1 RNA copies/mL plasma) at baseline (for at least 2 months) were switched to a raltegravir-containing regimen. Blood samples were taken at baseline and at ≥2 timepoints up to 48±6 weeks. Levels of total HIV-1 DNA and 2-LTR circles in peripheral blood mononuclear cells (PBMCs) were measured using real-time PCR assays.

Results

There was no significant change in HIV-1 total DNA levels over the study duration (p = 0.808), median slope 0.24 (conservative nonparametric 95% CI: −11.78, 26.23). Low levels of 2-LTR circles were detected in 2 patients. One had 16 copies/10⁶ PBMCs at baseline and the other had 34 copies/10⁶ PBMCs at week 51.

Conclusions

The switch to a raltegravir containing regimen was not associated with a significant change in HIV-1 total DNA levels in this cohort. There were no observed changes in the levels of HIV-1 2-LTR circles associated with raltegravir treatment initiation.     

Rapid qualitative urinary tract infection pathogen identification by SeptiFast real-time PCR

Background

Urinary tract infections (UTI) are frequent in outpatients. Fast pathogen identification is mandatory for shortening the time of discomfort and preventing serious complications. Urine culture needs up to 48 hours until pathogen identification. Consequently, the initial antibiotic regimen is empirical.

Aim

To evaluate the feasibility of qualitative urine pathogen identification by a commercially available real-time PCR blood pathogen test (SeptiFast^®) and to compare the results with dipslide and microbiological culture.

Design of study

Pilot study with prospectively collected urine samples.

Setting

University hospital.

Methods

82 prospectively collected urine samples from 81 patients with suspected UTI were included. Dipslide urine culture was followed by microbiological pathogen identification in dipslide positive samples. In parallel, qualitative DNA based pathogen identification (SeptiFast^®) was performed in all samples.

Results

61 samples were SeptiFast^® positive, whereas 67 samples were dipslide culture positive. The inter-methodological concordance of positive and negative findings in the gram+, gram- and fungi sector was 371/410 (90%), 477/492 (97%) and 238/246 (97%), respectively. Sensitivity and specificity of the SeptiFast^® test for the detection of an infection was 0.82 and 0.60, respectively. SeptiFast^® pathogen identifications were available at least 43 hours prior to culture results.

Conclusion

The SeptiFast^® platform identified bacterial DNA in urine specimens considerably faster compared to conventional culture. For UTI diagnosis sensitivity and specificity is limited by its present qualitative setup which does not allow pathogen quantification. Future quantitative assays may hold promise for PCR based UTI pathogen identification as a supplementation of conventional culture methods.     

Abbott RealTime HBV Assay Is More Sensitive in Detection of Low Viral Load and Little Impacted by Drug Resistant Mutation in Chronic Hepatitis B Patients under Nucleot(s)ide Analogues Therapy

Background

Selection of drug-resistant strains may lead to failure of HBV antiviral therapy. There is little information whether there is detection difference in drug resistant mutations between different viral load assays of HBV.

Objectives

This study is aimed to investigate whether there is drug-resistant strains related detection difference between Abbott RealTime HBV (RealTime) and CobasAmpliPrep/CobasTaqMan HBV assays 2.0 (TaqMan).

Study Design

One hundred and thirty-four CHB patients who received HBV anti-viral therapy were enrolled. HBV virological markers were tested 3 months apart regularly. Serum HBV DNA levels were determined using the TaqMan and RealTime. YMDD (rt180M and rt204V) mutation was checked in patients who experienced virologic breakthrough (VBT).

Results

The correlation of HBV DNA observed between the RealTime and TaqMan was good for all 571 samples (R² = 0.797; P<0.001). However, the correlation in the 434 samples with HBV DNA level <3 log₁₀ IU/ml was not as good as in all samples (R² = 0.457). Overall, 21.5% of samples had a detection difference of ≥1 log₁₀ IU/ml with 91.9% of these having HBV DNA level <3 log₁₀ IU/ml. Twenty-four patients experiencedVBT. Three of these patients had acquired the YMDD mutation and exhibited discordant viral load results between the two methods tested. In each case, persistent HBV DNA was detected by RealTime and undetectable with TaqMan. Of the patients who experienced a VBT and had acquired YMDD mutation, 4.7% had undetectable HBV DNA by TaqMan while all were detectable with RealTime.

Conclusions

RealTime assay is more sensitive and is little impacted by the development of drug resistant mutation.     

Genotype-Specific Incidence and Clearance of Human Papillomavirus in Oral Mucosa of Women: A Six-Year Follow-Up Study

Background

There are no previous longitudinal studies on genotype-specific natural history of human papillomavirus (HPV) infections in oral mucosa of women.

Methods

In the Finnish Family HPV Study, 329 pregnant women were enrolled and followed up. HPV-genotyping of oral scrapings was performed with nested PCR and Multimetrix® test (Progen, Heidelberg, Germany). Incidence and clearance times and rates for each HPV-genotype identified in oral mucosa were determined. Predictors for incident and cleared HPV infections for species 7/9 genotypes were analyzed using Poisson regression model.

Results

Altogether, 115 baseline HPV-negative women acquired incident oral HPV infection, and 79 women cleared their infection. HPV16 and multiple HPVs most frequently caused incident infections (65% and 12%) in 13.3 and 17.1 months respectively, followed by HPV58, HPV18 and HPV6 (close to 5% each) in 11–24 months. HPV58, HPV18 and HPV66 were the most common to clear. HPV6 and HPV11 had the shortest clearance times, 4.6 months and 2.5 months, and the highest clearance rates, 225.5/1000 wmr and 400/1000 wmr, respectively. The protective factors for incident oral HPV-species 7/9 infections were 1) new pregnancy during follow-up and 2) having the same sexual partner during FU. Increased clearance was related with older age and a history of atopic reactions, whereas previous sexually transmitted disease and new pregnancy were associated with decreased clearance.

Conclusions

HPV16 was the most frequent genotype to cause an incident oral HPV-infection. Low risk HPV genotypes cleared from oral mucosa more quickly than high risk HPV genotypes. Pregnancy affected the outcome of oral HPV infection.     

HPV Infections among MSM in Shenzhen,China

Background

An increasing incidence of anal cancer among men, especially men who have sex with men (MSM) suggests a need to better understand anal human papillomavirus (HPV) infection among this group.

Methods

A cross-sectional study was conducted among MSM in Shenzhen, China. Blood was collected for HIV serological testing and syphilis serological screening, and anal swabs were collected for HPV genotyping. Difference of HPV prevalence between HIV seropositive and HIV seronegative MSM was assessed by chi-square test. Factors associated with anal canal HPV infection were assessed by univariate and multivariate logistic regression.

Results

A total of 408 MSM were recruited. HIV and HPV prevalence were 6.9% and 36.4%, respectively. HPV was detected in the anal canal in 71.4% of the HIV-positive MSM and in 33.8% of the HIV-negative MSM (P<0.001). Oncogenic types were seen more often in anal specimens of HIV-positive MSM than in specimens of HIV-negative MSM (P = 0.001). The HPV genotypes detected most frequently were HPV06 (8.2%), HPV16 (7.2%), HPV11 (6.4%), HPV18 (4.7%), HPV58 (4.7%), and HPV52 (4.2%).

Conclusions

In this study, HIV positive MSM had a higher burden of HPV infection, especially oncogenic HPV infection. HPV types 52 and 58 were as popular as those types designed for the currently available vaccine (HPV6, 11, 16, 18).     

Human Papillomavirus in Head and Neck Squamous Cell Carcinoma of Unknown Primary Is a Common Event and a Strong Predictor of Survival

Background

The purpose of this study was to examine the prevalence of human papillomavirus (HPV) in patients with head and neck squamous cell carcinoma of unknown primary (CUP).

Methods

All patients diagnosed with and treated for CUP between January 1, 2000, and June 1, 2011, at two Danish medical centers were included. All patients received a thorough diagnostic work-up, including FDG-PET, before being diagnosed as CUP. We determined the HPV status in all patients using a combination of HPV DNA PCR and p16 stain. In addition, clinical information on the study patients was retrieved from clinical records.

Results

Of the identified 60 patients with CUP, 13 were shown to be positive for HPV DNA, amounting to 22% of the study population. In addition, we were able to show a clear disease-free and overall-survival benefit in the HPV-positive group, with a hazard ratio of 0.16 (95% CI: 0.038–0.67) for over-all survival. This survival benefit was also apparent when adjusted for advanced age in a multivariate Cox regression analysis.

Conclusion

A fairly large percentage of CUP cases are HPV-related, and because this is related to both the location and prognosis, we recommend HPV testing as part of the diagnostic work-up.     

Epstein-Barr Virus,Human Papillomavirus and Mouse Mammary Tumour Virus as Multiple Viruses in Breast Cancer

Background

The purpose of this investigation is to determine if Epstein Barr virus (EBV), high risk human papillomavirus (HPV), and mouse mammary tumour viruses (MMTV) co-exist in some breast cancers.

Materials and Methods

All the specimens were from women residing in Australia. For investigations based on standard PCR, we used fresh frozen DNA extracts from 50 unselected invasive breast cancers. For normal breast specimens, we used DNA extracts from epithelial cells from milk donated by 40 lactating women. For investigations based on in situ PCR we used 27 unselected archival formalin fixed breast cancer specimens and 18 unselected archival formalin fixed normal breast specimens from women who had breast reduction surgery. Thirteen of these fixed breast cancer specimens were ductal carcinoma in situ (dcis) and 14 were predominantly invasive ductal carcinomas (idc).

Results

EBV sequences were identified in 68%, high risk HPV sequences in 50%, and MMTV sequences in 78% of DNA extracted from 50 invasive breast cancer specimens. These same viruses were identified in selected normal and breast cancer specimens by in situ PCR. Sequences from more than one viral type were identified in 72% of the same breast cancer specimens. Normal controls showed these viruses were also present in epithelial cells in human milk – EBV (35%), HPV, 20%) and MMTV (32%) of 40 milk samples from normal lactating women, with multiple viruses being identified in 13% of the same milk samples.

Conclusions

We conclude that (i) EBV, HPV and MMTV gene sequences are present and co-exist in many human breast cancers, (ii) the presence of these viruses in breast cancer is associated with young age of diagnosis and possibly an increased grade of breast cancer.     

Patterns of Human Papillomavirus DNA and Antibody Positivity in Young Males and Females,Suggesting a Site-Specific Natural Course of Infection

Background

To monitor the impact of human papillomavirus types 16 and 18 vaccine on HPV infection dynamics in the Netherlands, we started an ongoing study in sexually transmitted infection (STI) clinics in 2009. Here, we analyze baseline type-specific HPV DNA and HPV-specific antibody positivity rates.

Methods

We enrolled 3569 men and women, 16–24 years of age, from 14 STI clinics, and estimated genital and anal HPV DNA and antibody positivity rates of 7 main carcinogenic HPV types. Generalized estimating equations regression analyses were applied to determine risk factors for, and associations between, type-specific HPV DNA and antibody positivity.

Results

Genital HPV DNA positivity rates were higher in women than in men; anal HPV DNA was especially high in men who have sex with men (MSM). HPV antibody seropositivity rates were also highest in women and MSM. High-risk sexual behavior was predictive of both HPV DNA and antibody positivity. Despite a strong correlation in serological profiles for multiple HPV types, seropositivity was independently associated with homologous HPV DNA detection.

Conclusions

HPV DNA and antibody positivity rates are higher in women and MSM than in heterosexual men, but their association is similar across gender. This suggests a site-specific natural course of infection.