Buffer strips have been widely recognized as to promote infiltration, deposition and sorption of contaminants for protecting surface water against agricultural contamination. However, such strips do not intercept all contaminants, particularly soluble ones. Although preferential flow (PF) has been suggested as one factor among several decreasing the efficiency of buffer strips, the mechanisms involved are not well understood. This project examines buffer strip efficiency at intercepting solutes when subsurface PF occurs. Two soluble sorbed tracers, FD&C Blue #1 and rhodamine WT, were applied on an agricultural sandy loam soil to evaluate the ability of a naturally vegetated buffer strip to intercept soluble contaminants. Rhodamine was applied about 15 m from the creek, while the Blue was applied 15 m to 165 m from the creek. Tracer concentration was measured over a two-year period in both the creek and the buffer strip through soil and water samples. Although the tracers traveled via different pathways, they both quickly moved toward the creek, passing beneath the buffer strip through the soil matrix. Our results demonstrate that the risk of water contamination by soluble contaminants is high in such systems, even when a well-vegetated buffer strip is used. The design of buffer strips should be modified to account for underground bypass, either by using plants that have deep, fine roots that do not favour PF or by adding a filter extending deep underground that can be regularly changed. Vegetative buffer strips are considered to be one of the best practices for reducing contaminant transport toward surface water –2. TheirThe effectiveness of such barriers is nevertheless highly variable and less known for below ground movement. This variability is explained in part by the multiplicity of processes and factors involved and by their dynamic and complex nature . Width, The density of the buffer strip vegetation and its resistance to bending are also determining factors of buffer strip efficiency at the soil surface , 9. HoweTransportation time in the subsurface generally decreases with an increase in slope, in the presence of shallow soil, with low permeable soil underneath the surficial one, with artificial drainage , in the presence of large lateral roots or biopores which creates lateral preferential flows called ‘pipe flows’, with other lateral preferential flow processes, and when there is sufficient precipitation to trigger water movement . Within Particles may move below ground at high speed when in natural or artificial pipes or in concentrated flow paths . NaturalThese belowground PF processes and bypass systems have been put forward as potential explanations for buffer strip inefficiencies with regard to various soluble agricultural contaminants. To our knowledge, there have been very few field studies that have tested this hypothesis. The goal of this study was to examine the ability of buffer strips to intercept solutes in a loamy soil where subsurface PF is suspected to occur at the field-scale using soluble tracers.The field experiment was conducted on private land in the Bras d’Henri watershed at 46°28'51.5" N and 71°13'03.9" W in Quebec, Canada. The landowner gave his permission for the study on condition that we not modify his field or soil management regime. The study did not involve endangered or protected species.The field  is a cat-3) is typical of agricultural soils  Beauv, Elymus repens, Taraxacum officinale, and others. During the growing season, vegetation along the creek bank was dense and homogeneous, growing to a height of about 1.4 m high, with a deep rooting system extending at least 0.6 m deep. The flat section of the buffer was also covered with grass, but vegetation did not grow as much in this area because of compaction. During spring 2011, the flat section was seeded with the ‘Cave-in-Rock’ variety of switchgrass (Panicum virgatum L.). Prior to seeding, this section was never worked but used only for machinery circulation.A mixture of indigeneous and invasive plants grows on the creek bank section of the buffer strip. The vegetation included -2 on all plots on 8 October 2010, while rhodamine WT  was applied at a rate of 2.5 g m-2 on the same day using the same solution and method, but only on plots A and B (close to the buffer strip). The tracers were chosen to represent weakly sorbed (Blue) and more sorbed (rhodamine) solutes. The Blue mimics phosphate [Four 2- x 2-m plots were established in the field. Three were aligned along distinct parts of the catena at 15 m (plot B), 60 m (plot C), and 165 m (plot D) from the creek . The fouhosphate  or atrazhosphate , and rhohosphate  in low-shosphate –28. They1 interface), 0.24 (B1 and B2 interface), and 0.85 (B2 and C interface) m depth. One year passed before we started collecting water samples from the lysimeters in order to permit soil settlement. The lysimeter plates worked only when the soil surrounding the plates was saturated, since they only intercepted free water. They were positioned to only capture water moving horizontally or vertically in between two soil layers. The plates reacted independently of each other, since the activated ones were not always the same. When the creek rose too high, the deepest lysimeters were not sampled because the water level was higher than they were.Two series of zero-tension lysimeters were installed in the buffer strip  in May 2i.e., when the creek rose to at least 0.25 m as detected by the water height detector in the creek. Tracer concentrations represent the average concentration recorded by the lysimeters over the 12 hrs during and after precipitation events. Collected water was transported through PVC piping and stored in Nalgene bottles until sampling. The samples were kept at 4°C in darkness until analysed.Lysimeter water collection occurred from May to September in 2011 and 2012 during intense rainfall events, Water was sampled in the creek about 60 m downstream from the lysimeter series. The autosampler collected 50 mL of water every 4 hours. These samples were combined into composite samples, each of which represented about 3.5 days of sampling. Additional samples were taken during heavy rainfall events: the autosampler collected samples hourly for 15 hrs whenever the creek reached a height of 0.25 m. In this case, the composite samples were made from 4 individual samples (average of 4 hrs). All samples were filtered and frozen before tracer analysis. Creek depth was measured daily in between rainfall events and hourly during precipitation. Water was also occasionally sampled from the outlet of a drain situated between both lysimeter series .A meteorological station was installed on site to monitor daily air temperature, relative humidity, vapour pressure, wind speed and direction, direct and reflected solar radiation, and rainfall depth.-3 m, WC-10 m) [-3. Since the soils of the B1 and B2 horizons were very similar, they were often combined for measuring and interpreting their properties.Trenches were dug very close to the lysimeters in November 2011 and May 2012 (2 profiles per date) for soil sampling in the buffer strip. Disturbed soil samples were extracted at a range of depths and distances from the creek. The samples were conserved at 4°C in darkness until tracer concentration analysis. Additional soil was sampled to characterize general soil attributes  such as WC-10 m) , and watWC-10 m)  of the sSoil samples were dried for 24 hrs at 105°C prior to tracer extraction. A ratio 1:5 soil:water was used for tracer extraction . Samples-1. A luminescence spectrophotometer  was used to measure rhodamine concentration. The rhodamine was excited at 560 nm and emissions were measured at 577 nm [−3 mg L-1.A UV-visible spectrophotometer  set at 630 nm was used to measure Blue concentration –37. The t 577 nm . The rho-1; http://www.jagson.com/food-color/brilliantblue.php, consulted on 4 November 2014). By contrast, less than 1∕3 of rhodamine was recovered  were used to measure the natural noise for tracer analyses. Blanks received the same treatment as other samples. Noise levels for Blue and rhodamine were relatively low  and wereecovered . RhodamiLinear sorption isotherms (Kd) were calculated using a range of concentrations slightly larger than that found in the samples , 38. TheDescriptive statistics were calculated using PROC UNIVARIATE in the SAS/STAT software v.9.1 . Data were transformed using the Box–Cox method to increase their normality when required . CorrelaThe tracer budget could not be estimated in this study since tracer concentrations were measured only during the unfrozen period, drains were rarely sampled, neither the concentration nor the flow in surface runoff were measured, and only a very small section of the buffer strip was monitored. In addition, tracer uptake by vegetation was not measured.The fall season of 2010 received more rain than the other seasons. Tracer application took place between two precipitation events in October, enabling the tracers to penetrate the soil surface before freeze-up. Some erosion occurred at the soil surface  followinThe temperature (data not shown) and precipitation of winter 2011 were representative of the 30-year average. The month of May received one large (83 mm) rain event over a short period , represeThe months of May and June 2012 received more than 179 and 159 mm of rain, respectively, with 6 major precipitation events. Limited erosion but not gully formation was observed during these events. The other events of the summer were less important.-1 in 2012. The maximum flows in 2011 and 2012 were 4 532 L s-1 (30 May 2011) and 3 776 L s-1 (30 September 2012). The average flow in 2011 was almost five times that of 2012 . Neither time lag, actual concentration, relative concentration, nor mass flow (creek flow x tracer concentration) improved these correlations. Rhodamine was almost always present in the background, but its concentration did not necessary increase with creek flow.2 = 0.17) with the total precipitation between two measurement events. This may be due in part to spatial and temporal field variability in precipitation. Precipitation amounts greatly varied over very short distances at this site, changing spatial dilution from one rain to another. In addition, tracers already in the creek bank may have been released when the water level rose.Rather than being correlated to creek flow, tracer concentrations were better correlated, although weakly, to the cumulative amount of precipitation between concentration measurements. The Blue concentration tended to increase . Blue concentration ranged across almost three orders of magnitude, while rhodamine ranged across only two.Tracer concentrations in the creek were not significantly correlated with one another during 2011, but were during 2012. Rhodamine concentration tended to increase with Blue in 2012 . No surface runoff containing Blue was apparent at that time, but we could visually detect it in the drain. This happened two weeks before the heavy rainfall event that led to gully formation and important surface runoff. Therefore, very fast tracer movement occurred belowground to reach the drain, with movement in part perpendicular to the surface slope. Since rhodamine could not be observed as easily with the naked eye, we do not know if its movement was associated with surface runoff.2 = 0.87).Drain water samples contained rhodamine most of the time, but Blue was more rarely present. When Blue was detected, its concentration was strongly correlated to that of rhodamine  and rhodamine (4.05 x 10−3 mg L-1) were higher than those in the creek in 2011 (0.15 mg L-1 for the Blue and 0.37 x 10−3 mg L-1 for the rhodamine) and lower in 2012 (0.44 mg L-1 for the Blue and 2.60 x 10−3 mg L-1 for the rhodamine). There was always at least one of the tracers in the water in at least one lysimeter. Rhodamine was more frequently observed in lysimeters  than Blue, which was detected almost exclusively in the deepest lysimeters. When Blue was detected, its concentration was at most ten times higher than that of the rhodamine  with precipitation, although only in 2011. These results indicate that Blue was not carried into the buffer strip by water from the creek, but rather from the upper part of the field through deep percolation [No significant regressions or correlations were found between creek and lysimeter concentrations, with or without time lag analyses, for either tracer at any depth (data not shown). The only significant (P = 0.05) correlation found between lysimeter concentrations and other parameters concerned Blue concentration in the deepest lysimeter, which was weakly correlated  . The higWe observed strong resurgences of water along the buffer strip at very specific narrow points. Some of these points corresponded to holes  associated with old, decayed stumps, some of which were large enough to accommodate the hand of an adult person. These resurgence points in the buffer strip, visible with the naked eye, flowed directly into the creek and were active mainly during rainfall events, but also between them. Any PF pathways that passed through the application plots are likely to have transported tracers at higher concentration. Otherwise, such paths would dilute the tracers. Thus, depending upon the paths during each rainfall event, these pipes may have dilute or carried more tracers through the buffer strip. One of these resurgence points was located close to a lysimeter series and the trench used to measured tracer concentration in the buffer strip soil but did not pass through it, but not directly on the measurement points. We therefore did not measure the direct effect of these resurgence points on tracer transport through the buffer strip.Another narrow resurgence was identified at the footslope slightly uphill from plots A and B. Blue was present at this location, visible with the naked eye. This resurgence point seems to be stable over time as it rewetted after rainfalls and more Blue appeared as time went on. No other such resurgence was observed in the field. Because of its uphill position compared to plots A and B, the Blue could not come from these plots but must have come from higher in the catena (plots C and/or D) and this, through underground preferential lateral flow (LF) since its appearance was very localised.In addition, scarce, localised, narrow points of seeping were observed when trenches were dug in the buffer strip for soil sampling. Some of the samples also contained the Blue in concentration high enough to be visible with the naked eye.The generally accepted way to describe solute transport in soil is the standard advection–dispersion equation , which cCl is the liquid phase solute concentration (mg L-1), t is the time (day), De is the effective coefficient of dispersion (day-1), z is the distance (m), V is the water velocity (m day-1), and rs is the solute appearance or disappearance. In this case, the tracers are the solute.R is the retardation factor (unitless), i.e., by matrix flow, occurs during saturated conditions. If diffusion, appearance, and disappearance are assumed to be negligible compared to the hydrodynamic dispersion , the time (Time) to reach the creek can be estimated as follows [The shortest time required for the tracer to reach the creek from the application plots when moving by the standard advection–dispersion process,  follows :Time=LRv is the volumetric soil water content (m3 m-3). WCv = P at saturation. The q is the flux (m s-1). Given the distance from the application plots to the creek (L = 15 m), the water flux is:θ dH/dz is the hydraulic gradient and equal to 1.0 under saturated conditions.Using these equations, the time for tracers to reach the creek from the application plots is easily estimated from the P, Kd, Ks, and R values given in Tables The predicted values suggest that the Blue could have reached the B horizon of the buffer strip within a few months while rhodamine should not have reached the buffer strip before the third year . ComparaAccording to the predictions, it would take between 2.6 and 15 days, depending upon the horizon, for the water to reach the creek , comparai.e., they moved at about the same speed, then they moved almost exclusively by PF.Since the movement of both tracers was comparable to those of water and they were applied at the same time, position, and depth, we can use their Kd or R ratios to evaluate whether the flows occurred through the soil matrix or PF. If the ratio of the time required to cover a certain distance (slower tracer ∕ faster one) is equal to their Kd or R ratios, then the movement occurred by advection–dispersion within the soil matrix (matrix flow). If the ratio of the time they took to cover a distance is smaller than their Kd or R ratios, then they move not only through the soil matrix but also through PF paths. If the ratio of their time tends toward 1.0, The Kd and R ratios of rhodamine/Blue are about 35 Tables  and 5. BConsidering that (1) the Blue would take at least a couple of months and rhodamine more than 5 years to reach the creek under matrix flow at saturation; (2) it would take at least 35 times longer for rhodamine than Blue to reach the creek with advection-dispersion transport; (2) Blue and rhodamine were observed in the creek only a few days after their application in the field; (3) pipe flows were observed along the buffer strip; (4) a narrow point of resurgence containing the Blue occurred at the footslope; (5) narrow points of seeping containing the tracers were observed within the buffer strip soil; and (6) drain flows were observed to transport the tracers both early in the experiment and thereafter, then multiple belowground PF processes have occurred in the field.In addition, (1) the soil changes along the catena; (2) the Blue was applied on all 4 plots rather than close to the creek as for rhodamine; (3) the Blue concentration was null at the soil surface but significant in the subsurface; (4) the Blue concentration deep in the buffer strip was maintained over time; and (5) its concentration increased at specific downhill at narrow spots over time. Thus, we can conclude that, among different belowground PF processes, lateral PF from uphill was important.e.g., water distribution), intensity and duration of precipitation events, co-existing types of PF processes, and remaining active flow paths change the active network from one rain to another. Thus, the occurrence of different PF processes, the fact that they did not take place at the same time or at the same place during every rainfall in addition to the fact the PF paths interact with one another enhance the spatial and temporal variability of tracer movement [Because Blue concentration was not strongly correlated to precipitation, the active PF paths may not always have been the same. Some PF paths may be activated very rapidly, but may require specific conditions to be activated . These mmovement . It makeBlue concentration in the different compartments  showed higher spatial and temporal variability than that of rhodamine. Rhodamine was consistently measured in the creek, in the lysimeters, and in the drain, while the presence of Blue was more intermittent. Peaks of Blue and rhodamine concentration did not occur at the same time. In addition, rhodamine was present at the soil surface, despite being sensitive to light and degradation processes. By contrast, the Blue was observed at the soil surface only at the resurgence spot and directly at the application spots. Otherwise, it was observed only in the subsurface.i.e., less precise). This was not often to the case. The difference in temporal variation between both tracers was therefore not due to detection limits.These results are counterintuitive, since Blue was applied at the same time as rhodamine, at a higher concentration, and on four rather than on two plots. It is also less sensitive to degradation. We would thus expect the Blue to be observed in the different compartments more frequently than rhodamine. If these observations were due to the detection limits of our apparatus, we would expect high rhodamine concentrations to be detected whenever high Blue concentrations were detected, since the Blue detection limit is 100 times higher than that of rhodamine . In addition, it is known that PF paths that crossed a location where a tracer was present carried the tracer further downhill, while those that did not pass through a tracer location diluted the tracers along their way [The higher variability of Blue concentrations was due in part to its higher solubility, decreasing its response time to rainfall events. Rhodamine moved not only in the subsurface, but probably also at the soil surface in runoff because of its higher sorption on soil particles. The distribution of the two tracers differed at the footslope, with both being present at some spots (the application plots) but only Blue being present at some spots  and only rhodamine being present at others  The tracers passed beneath the buffer strip after having reached it partly through PF paths throughout the field. (2) The tracers bypassed the buffer strip along PF paths, such as pipe flows, within the buffer strip itself, decreasing its underground efficiency. In addition, gullies occurred at the soil surface in the field and in the buffer strip, bypassing it and decreasing its efficiency at the soil surface . (3) Another portion of the tracers moved through drains, further decreasing the efficiency of the buffer strip once the soluble tracers had infiltrated into the soil from different points in the field. Thus, drains, pipe flow, gullies, and lateral PF, which occurred throughout the catena, all decreased the effectiveness of the buffer strip, particularly in the case of more retarded solutes (e.g. rhodamine comparatively to Blue).et al. [Many governments encourage farmers to establish buffer strips in order to decrease the transport of contaminants such as nitrate, phosphorus, and pesticides toward surface water. Based on the combined criteria of the FIHOQ , the MDDet al. , the bufet al.  suggestset al. , 47.In addition to the above new information, we know that problems can occur after buffer strips accumulate contaminants . When the.g., where subsurface bypass (PF) controls most of the water movement toward surface water.It is important to consider how the design of buffer strips might be modified  to improet al. [et al. [It has also been suggested that the addition of perennial plants with deep root systems would be an efficient way to extract some proportion of contaminants in the subsurface . But laret al.  and Robe [et al. , might a-2, the equivalent of 5 x 10−5 kg ha-1, within just a couple of weeks after its application. Such concentrations in the creek would be considered dangerously high in the case of highly toxic chemicals, such as certain pesticides, and for contaminants that, even at small concentrations, affect the environment . We thus expect these observations to be true for soluble contaminants applied at high concentrations, such as nitrates, but also for highly toxic contaminants applied at low concentrations.We were able to detect the presence of Blue in the creek with the naked eye with an applied mass of 125 g mThe concentration of two soluble sorbed tracers was measured in a creek, in lysimeter plates installed in the buffer strip, and in the soil of a riparian buffer strip naturally vegetated with grass during a two-year period. These data and other field observations indicated the existence of important preferential flows (PF) in the field and tracer movement both beneath the buffer strip and through it via lateral PF, drains, and pipe flow. A proportion of both tracers may have used the same pathways to reach the creek, while another portion used different pathways. The Blue movement demonstrated that the entire catena was involved in the long-range transport of this tracer toward surface water, through both very fast and slow paths, although field slope was only 4%. Consequently, although the buffer strip was managed according to recommended methods the transport occurred through subsurface lateral PF processes, reducing the ability of the buffer strip to intercept contaminants. Buffer strips may be upgraded either by the addition of filtering agents installed in the subsurface at hot spots, which could be replaced at a specific interval or by the use of plants with deep, fine root systems to take up the contaminants, after which they could be removed by harvesting the plants.


Over 45,000 Americans are estimated to be newly diagnosed with cancer of the oral cavity and pharynx in 2015, and an estimated 8,650 will die of the disease . It is well known that the combination of smoking and alcohol use increases the risk for developing head and neck squamous cell carcinoma (HNSCC). Head and neck squamous cell carcinoma is a general term that encompasses cancers of the nasal cavity, sinuses, lips, mouth, salivary glands, throat, and larynx. HNSCC has been considered to be a cancer predominantly seen in men of older age .The current trends, deaths, and survival rates demonstrate a modest decrease in HNSCC from 1975 to 2006 . However, despite this general decrease, research demonstrates that oropharyngeal-specific cancers are rising ., 2012.Currently, there is evidence that tissue collected from patients diagnosed with head and neck cancer may be positive for human papillomavirus (HPV) DNA, particularly cancers of the oropharyngeal site ., 2013. According to the Centers for Disease Control and Prevention , 79 million people have HPV, and over 14 million cases are newly diagnosed each year. This virus is associated with cancer of the cervix, vagina, penis, and anus. The exact incidence of HPV-positive HNSCC is unknown and could be largely misrepresented because current practice does not support testing all HNSCC for HPV status.Currently, the Surveillance, Epidemiology, and End Results Program (SEER) does not specify HPV status in statistics on oral cavity and pharynx cancers. However, according to the CDC (2014a), an estimated 72% of oropharyngeal cancers may be attributed to HPV infections, with more than 2,370 new cases diagnosed in women and nearly 9,356 new cases diagnosed in men every year in the United States.Over 100 different subtypes of HPV have been discovered . Most subtypes cause common skin warts. These types are categorized as low or high risk. Low-risk types, such as HPV-6 and HPV-11, can cause benign papillomas . Generally, infections with these types tend to resolve themselves spontaneously within 12 months.However, high-risk types such as HPV-16 and HPV-18 can lead to chronic infection, cellular abnormalities, and then cancer. HPV-16 is linked to HNSCC of the oropharynx . ApproxiInfection with high-risk types of HPV can cause cells to make two proteins: E6 and E7. These proteins can turn off certain genes that control cell proliferation. Uncontrolled cell proliferation or cell growth can lead to cancer .Sexual BehaviorHuman papillomavirus infection can spread from one person to another through sexual contact, including vaginal, anal, and oral sex. Oral sexual behavior and open-mouth kissing have been linked to an increased risk for oropharyngeal infection ., 2014. Current data indicate that oropharyngeal cancer is more prevalent in men than in women .SmokingSmoking is a known carcinogenic that can cause changes in the DNA of cells and contribute to the increased risk for many types of cancers. Several studies demonstrate that smoking status can contribute to the persistence of HPV infection ., 2014b.Precisely how smoking contributes to chronic HPV infection and its progression to malignancy remains elusive. Castle (2008) discussed some plausible theories. First, smoking decreases immunity, which can reduce the body’s ability to effectively clear HPV infection. This, in turn, causes chronic infection and inflammation. Second, infection with high-risk HPV subtypes can cause uncontrolled cellular proliferation. These molecular changes combined with the carcinogens in tobacco smoke can work synergistically to encourage cancer development.In addition, studies have indicated that smoking can adversely affect response to treatment and prognosis ., 2015. Oral HygieneThere may be an association with poor oral hygiene that leads to an increased risk of developing HPV-positive HNSCC ., 2012. Chronic inflammation plays several key roles in the development of cancer. According to In 2014, the National Comprehensive Cancer Network (NCCN) recommendations included testing tumors for HPV in the initial workup of oropharyngeal cancers . Recommendations include either an immunohistochemistry for analysis of p16 expression or HPV in situ hybridization for detection of HPV DNA in tumor cell nuclei . Testing is not considered in management decisions, however, and is used for prognosis purposes.Current treatment guidelines, depending on stage, consist mainly of surgery, radiation therapy, chemoradiation therapy, and targeted therapies . In a majority of studies, HPV-positive patients showed better prognosis and 5-year survival than HPV-negative patients with HNSCC ., 2014. VaccinesThere are currently two HPV vaccines approved by the US Food and Drug Administration (FDA): Gardasil and Cervarix. Gardasil is quadrivalent and protects against HPV subtypes 6, 11, 16, and 18. Subtypes 6 and 11 are associated with genital warts. Cervarix is bivalent and protects against oncogenic types 16 and 18. These vaccines are currently approved to prevent cervical cancer and precancers in women.The FDA has approved both vaccines to be given to girls between the ages of 9 and 26 and has currently approved Gardasil for boys between the ages of 9 and 21 to prevent genital warts. For women older than 26 and men older than 21 who did not receive the vaccine or did not receive all three doses, the vaccine can still be given. The CDC currently recommends that girls receive three doses of either vaccine at the age of 11 or 12. The future impact these vaccines may have on the prevalence of HPV-positive tumors of the head and neck is thus far undetermined.Psychosocial ImpactHuman papillomavirus has only newly been associated with cancers of the head and neck. The impact of discussing HPV results with patients and their partners has not been well studied ., 2013. Patients newly diagnosed with HPV-positive HNSCC can experience misconceptions and uncertainty regarding its transmission, latency, and future spread ., 2013. There are some commonalities between these suggestions and the CDC’s current key messages for women receiving cervical cancer screening and HPV testing (2011). For instance, it is important to deliver information in both verbal and written formats within a neutral, nonstigmatizing context. Teaching should focus on prevalence, signs and symptoms, and risk to partners. To reduce possible accusations between couples, providers must emphasize that HPV can remain dormant for many years and that having HPV does not imply infidelity, nor should it necessarily raise concerns about a partner’s health . It is reasonable to consider that similar topics will be useful in patients newly diagnosed with HPV-positive HNSCC. All clinical providers should aim to decrease any feelings of stigmatism or guilt that patients frequently experience with the diagnosis of a sexually transmitted infection (STI).Moreover, what makes this even more challenging for these patients is being told that not only do they have cancer but that it may be related to an STI they contracted. This can add additional distress and confusion. Some possible strategies and key messages to convey to patients might include the prevalence of HPV, its transmission and prognosis, partner risk, and limitations in our present knowledge  to identify key messages pertinent to patients and the most effective approaches to successful communication. It is imperative that the AP understand the impact of HPV on cancer development in all areas of medicine. Understanding the impact of HPV-related cancers and associated risk factors should guide the physical exam and informative discussions during patient visits. Identifying risk factors can aid in prevention and early detection in the general public. Educating the public and patients regarding risk factors for HPV can help reduce infection rates and thus prevent cancer from developing.The oncology AP, in particular, may be the initial provider to inform patients about HPV testing, diagnosis, and prognosis as it relates to their cancer. It is important that these providers appreciate the common challenges and concerns patients may experience to enhance effective communication, reduce misconceptions, and ensure compliance with treatment.Thus, it is essential for the AP to keep up with the developments in HPV diagnosis as they relate to specific cancers. The AP is in the unique position of having the ability to care for and foster trusting relationships with their patients and their families, as well as possessing advanced knowledge in medical practice. Consequently, the AP is the most appropriate provider to lead the way for future research exploring the psychological impact in the diagnosis of HPV-positive HNSCC.


To assess the demographic and attitudinal factors associated with HPV vaccine initiation and completion among 18–26 year old women in Utah.Between January 2013 and December 2013, we surveyed 325 women from the University of Utah Community Clinics about their HPV vaccine related beliefs and behaviors. Odds ratios (ORs) were estimated from logistic regression models to identify variables related to HPV vaccine initiation and series completion.Of the 325 participants, 204 (62.8 %) had initiated the vaccine and 159 (48.9 %) had completed the 3-dose series. The variables associated with HPV vaccine initiation were lower age (OR = 1.18 per year); being unmarried (OR = 3.62); not practicing organized religion (OR = 2.40); knowing how HPV spreads (OR = 6.29); knowing the connection between HPV and cervical cancer (OR = 3.90); a belief in the importance of preventive vaccination ; strength of doctor recommendation ; and whether a doctor’s recommendation was influential . These variables were also significantly associated with HPV vaccine completion.The implications of these findings may help inform policies and interventions focused on increasing HPV vaccination rates among young women. For example, without this information, programs might focus on HPV awareness; however, the results of this study illustrate that awareness is already high (near saturation) in target populations and other factors, such as strong and consistent physician recommendations, are more pivotal in increasing likelihood of vaccination. Additionally, our findings indicate the need for discussions of risk assessment be tailored to the young adult population.The online version of this article (doi:10.1186/s12905-016-0323-5) contains supplementary material, which is available to authorized users. The American Cancer Society estimates that in 2014, there will be over 12,000 new cases of invasive cervical cancer diagnosed in the United States and over 4,000 women will die from this disease , has beeIn 2006, the Centers for Disease Control (CDC) Advisory Committee on Immunization Practices (ACIP) recommended a 3-dose HPV vaccination series as a routine vaccine for girls age 11–12 years old , 12–14.. State-specific data for this age group is unavailable. In spite of this low HPV vaccine coverage, recommendations from physicians remain suboptimal for all age groups ; not practicing vs. practicing organized religion: OR = 2.4 [95 % CI: 1.49–4.0]; knowledge of HPV transmission: OR = 6.29 [95 % CI: 3.46–11.44]; known connection between HPV and cervical cancer: OR = 3.90 [95 % CI: 2.21–6.89]; known importance of vaccine : OR = 2.45 [95 % CI: 1.79–3.36]; strength of doctor recommendation: OR = 1.86 per Likert scale unit [95 % CI: 1.27–2.70]; and a binary indicator of whether a physician’s recommendation is influential: OR = 1.70 [95 % CI: 1.38–2.09]. These variables were all also significant predictors of completion of the 3-dose series , married or monogamous relationship (n = 36), cost of vaccine or unsure if insurance covers vaccine (n = 23), concern of side effects (n = 18), not sexually active (n = 14), and vaccine inconvenience (n = 7)  have received just one dose of the 3-dose series , 15, 16.Resources are currently available to help increase HPV vaccination in Utah. In April 2007, Jon Huntsman, Sr. donated $1 million to the Utah Department of Health to educate Utahans about cervical cancer and provide low cost HPV vaccines to eligible women . An addiOur study of young women age 18–26 years old found several correlates of vaccine initiation and completion that may be useful for future public health interventions for this population. Among these correlates were lower age, awareness of HPV transmission, knowledge of its connection to cervical cancer, belief in the importance of the HPV vaccine, and a physician recommendation  , had access to healthcare, and response rates were higher in the vaccinated vs. the unvaccinated, which may have introduced unmeasured response bias. Furthermore, cross-sectional study design prohibits assessment of causal relationships.There are limitations to the generalizability of the current study. One of the primary limitations was the low survey return rate. Even after incentivizing a second wave of survey dissemination, response rates totaled well under 30 %. Therefore, our sample size was limited. Additionally, those who vaccinated were more likely to respond than those who did not vaccinate, leaving the potential for voluntary response bias. Furthermore, cross-sectional study design prohibits assessment of causal relationships.The majority of participants were White/Caucasian (>80 %), had access to healthcare, and response rates were higher in the vaccinated vs. the unvaccinated, which may have introduced unmeasured response bias.The implications of these findings may help inform policies regarding HPV vaccination education among young women. For example, without this information programs might focus on awareness , 42–44, ACIP, advisory committee on immunization practices; CDC, centers for disease control; DAG, directed acyclic graph; HBM, health belief model; HPV, human papillomavirus; IRB, institutional review board; OR, odds ratio; SCM, social cognitive model


Globally, cervical cancer is one of the most deadly forms of cancer affecting women—killing more than 270,000 people every year. The majority of these deaths  occur in developing countries. A global health priority, therefore, is to provide low-cost, highly effective cervical cancer prevention, detection, and control options to all women who need them.The human papillomavirus (HPV) family is a group of about 150 related viruses, a subset of which is thought to be responsible for the majority of cervical cancer cases. One of the most successful translational achievements in cervical cancer research in recent years is the development of highly effective preventative vaccines which directly target these oncogenic HPV strains. The bivalent vaccine Cervarix(r) (specific for HPV-16 and -18 strains) and the quadrivalent vaccine Gardasil(r)  contain viral L1 proteins from these strains, which are able to self-assemble into virus-like particles (VLPs). Recently, Gardasil(r) 9 has been approved by the United States FDA which will contain an additional 5 strains of HPV not currently covered by available vaccines. These vaccines induce antibody responses against HPV, but do not contain any of the viral genomic DNA present in natural viruses and, therefore, are not able to establish an infection. Available data indicate that these vaccines are highly effective at preventing precancerous cell induction in women, and since their introduction, global HPV infection rates appear to be declining.Although these prophylactic vaccines are highly effective at preventing new infections, neither vaccine is capable of inducing immune responses against established HPV infections. A major healthcare goal, currently, is to develop therapeutic approaches to resolving potentially cancerous HPV lesions , or cervical dysplasia). Several approaches are underway, including a therapeutic vaccine which is likely to work by inducing an adaptive T cell-mediated response against the viral E6 and E7 proteins, which are expressed in these lesions. Several drugs are also being explored, including an antibody (bevacizumab) against vascular endothelial growth factor (VEGF), which may help patients by limiting the growth of new blood vessels to the cancer. Clinical trials are underway to test whether chemotherapy drugs or immune checkpoint inhibitors such as ipilimumab may work well in combination with radiation therapy. Autologous T cell transfers, where the patients own immune cells are “trained” outside of the body to attack cancerous cells infected with HPV are also being explored. Armed with a basic scientific understanding of how HPV replicates and may contribute to cervical cancer, and how the patient's own immune system may be “kicked into gear” in order to attack those cancer cells has given researchers several potential avenues which may already be explored as treatment options.In addition to established principles about viral replication and immune system function, scientists are also using systems approaches to identify new genetic markers which are common in transformed cervical lesions, and ongoing studies are looking at the basic molecular mechanisms of how HPV-infected cervical cells become cancerous. These basic approaches may help researchers identify and manipulate new molecular pathways that are biologically relevant for cervical cancer transformation, and which may be amenable to therapeutic manipulation.Although several promising treatment and prevention options are available or under development, early detection remains one of our most effective tools for combating cervical cancer. The Pap test, introduced in the 1950s, is a simple means of screening cervical cells for abnormalities and may now be combined with HPV DNA screening to identify those patients at greatest risk for developing cervical cancer. Although highly effective, the Pap test may not be a practical screening tool for low-resource countries, and there is an urgent need for development and effective implementation of inexpensive, easily-administered screening options for these women—such as HPV DNA screening kits that may not necessarily need expensive refrigeration or a well-equipped laboratory to administer. Visual inspection with acetic acid (VIA) is an inexpensive option currently in practice, but this method may not be as accurate as DNA testing.We have come a long way since the introduction of the Pap test. Identification of HPV as the major etiological agent of cervical cancer, for example, has allowed rational vaccine design and the development of targeted immunotherapies. However, there is still much work to do to improve practical screening options in developing countries, and to add to our arsenal of treatment options for established HPV infections.EBioMedicine


Over the last two decades it has become clear that distinct types of human papillomavirus (HPV), the so-called high-risk types (hrHPV), are the major cause of cervical cancer. The hrHPV-DNA testing has shown excellent performance in several clinical applications from screening to the follow-up of conservatively treated patients.in situ, and microinvasive carcinoma compared to Pap smear cytology.We conducted a systematic review of the recent literature on the performance of HPV DNA testing in follow-up after treatment of high-grade cervical lesions, adenocarcinoma in situ, the hrHPV-DNA test is the most significant independent predictor of recurrent disease or progression to invasive cancer, and the combination of viral DNA testing and cytology reaches 90% sensitivity in detecting persistent lesions at the first follow-up visit and 100% at the second follow-up visit. The cause of microinvasive squamous cervical carcinoma is increasingly treated with conservative therapies in order to preserve fertility, and an effective strategy allowing early detection of residual or progressive disease has become more and more important in post-treatment follow-up. Primary results seem to indicate that the median time for viral clearance is relatively longer compared with patients treated for CIN and suggest a prolonged surveillance for these patients. However, the potential clinical value of HPV-DNA testing in this clinical setting needs to be confirmed by further observations.Observational studies have demonstrated that the high risk hrHPV-DNA test is significantly more sensitive (95%) compared to follow-up cytology(70%) in detecting post-treatment squamous intraepithelial high-grade lesions. Moreover, in patients treated conservatively for cervical adenocarcinoma in situ adenocarcinoma.The excellent sensitivity, negative predictive value, and optimal reproducibility of the hrHPV DNA testing, currently is considered a powerful tool in the clinicians’ hands to better manage post-treatment follow-up either in cervical squamous lesion or  Neverth–3 and inld-knife . A recenld-knife , 7. The ld-knife .Currently, despite it’s low sensitivity , 2, the A large amount of published data over recent decades on HPV DNA testing by means of pooled-based or type-specific methods has definitely demonstrated that persistent positivity of viral infection is considered a prognostic index of persistent/recurrent disease in patients treated for CIN2–3/ AIS. Observational studies demonstrated that post-treatment positive HPV-DNA test predicts residual/recurrent CIN with a significantly higher sensitivity and a non-significantly lower specificity than conventional cytology-based follow-up, 95% versus 70% and 75% versus 78% respectively. Overall, combined hrHPV and cytology testing yielded the best performance; the combination hrHPV-DNA testing and cytology demonstrated a 96% sensitivity, 81% specificity  with 99%Moreover, it has been observed that during post-treatment follow-up, hrHPV-DNA positivity is reduced from 90% to 20% (p < 0.01) at six months and positive endocervical margin of the cone, lesion grade in the surgical sample and age above 35 years have been indicated as significant independent factors predicting HPV persistence . FurtherDespite the higher sensitivity, hrHPV testing shows a lower specificity than cytology   in identAs mentioned before, it is noted that among the women resulting HPV-positive during surveillance, certain genotypes confer a higher risk of post-therapy recurrence , 14. HenAs HPV genotyping assays become increasingly used in clinical practice, further research will be necessary to shed light on this controversial topic to determine whether type-specific HPV DNA tests can better define the risk stratification of disease persistence or recurrence and the length of the follow-up period in women harbouring specific hrHPV genotype .Moreover recent studies suggest that the oncogenic activity of hrHPV-mRNA transcripts  may be aIn conclusion, the role of hr-HPV-testing has been confirmed as an accurate index of disease clearance and is accepted along with Pap smear, in the routine post-treatment workup of CIN- treated patients. However, it is essential to make the distinction between individuals carrying the virus and those having a persistent clinical HPV lesion. The former are women in whom the only evidence for virus persistence can be obtained by the molecular diagnostic tools, but who do not have any clinically detectable HPV lesion whereas the latter group have a clinical lesion detectable by Pap smear, colposcopy, and biopsy. These are the cases that make the Pap test more specific and give it a higher positive predictive value (PPV) in detecting significant pathology compared with molecular tests .Hence, it is clear that evidence-based post-treatment follow-up should include both conventional cytology and hrHPV-DNA testing to identify patients at increased risk for disease recurrence.in situ (AIS) continues to be a controversial issue [Over the past 50 years the incidence of invasive and pre-invasive glandular lesions of the uterine cervix has increased in western countries. Prior to 1970 adenocarcinomas accounted for approximately 5% of all cervical cancer cases, while by 1980s they had increased to 20–25% of all cervical carcinomas [al issue , 22. In al issue . The conal issue . More real issue , 25.Moreover, testing HR-HPV positive at any time during follow-up is the most significant independent predictor of recurrent disease. Finally, hrHPV detection was the single most powerful predictor of progression to invasive cancer. The status of the cone margins is also a good predictor and free cone margins had a significant protective effect against progressive disease .in situ or microinvasive AdCa, these women should be kept under close observation for at least the first three years after treatment.These results suggest that testing hrHPV positive at any time point during follow-up is the most significant independent predictor of progressive disease, while showing free margins in cone has a significant protective effect against progression of invasive adenocarcinoma (AdCa). However, as a small percentage of conservatively treated AIS patients with persistent, recurrent, or progressive disease experienced a late (fifth and sixth FU visit) diagnosis of Microinvasive squamous cervical carcinoma (MIC), which most commonly occurs in young women who are of childbearing age is increasingly treated with conservative therapies in order to preserve fertility. As a consequence, an effective strategy allowing early detection of residual or progressive disease has become more and more important in post-treatment follow-up. To date, however, there are no standard practice guidelines on the optimal methods of surveillance of patients. Studies on long term follow-up after a primary conservative treatment have demonstrated that about 20% of patients experienced early or late diagnosis of recurrent disease; the majority (15%) of cases had an intraepithelial lesion while in 5% a MIC or invasive carcinoma was identified –28.Follow-up protocols have so far consisted of repeat cytology and possibly colposcopy. The Pap smear has some inherent flaws, in particular a relatively high false negative rate, and the colposcopy has been shown to add very little to the detection rate of residual/recurrent CIN . Despitein situ adenocarcinoma.A large number of studies have definitely demonstrated the excellent performance of hrHPV-DNA testing in the follow-up of conservative CIN 2-3/AIS treated patients. Possible options in the management of patients are indicated in


During its February 2015 meeting, the Advisory Committee on Immunization Practices (ACIP) recommended 9-valent human papillomavirus (HPV) vaccine (9vHPV)  as one of three HPV vaccines that can be used for routine vaccination . HPV vacRecommendations for routine use of vaccines in children, adolescents and adults are developed by the Advisory Committee on Immunization Practices (ACIP). ACIP is chartered as a federal advisory committee to provide expert external advice and guidance to the Director of the Centers for Disease Control and Prevention (CDC) on use of vaccines and related agents for the control of vaccine-preventable diseases in the civilian population of the United States. Recommendations for routine use of vaccines in children and adolescents are harmonized to the greatest extent possible with recommendations made by the American Academy of Pediatrics (AAP), the American Academy of Family Physicians (AAFP), and the American College of Obstetricians and Gynecologists (ACOG). Recommendations for routine use of vaccines in adults are harmonized with recommendations of AAFP, ACOG, and the American College of Physicians (ACP). ACIP recommendations approved by the CDC Director become agency guidelines on the date published in the Morbidity and Mortality Weekly Report (MMWRhttp://www.cdc.gov/vaccines/acip/.). Additional information about ACIP is available at From October 2013 to February 2015, the ACIP HPV Vaccine Work Group reviewed clinical trial data assessing the efficacy, immunogenicity, and safety of 9vHPV, modeling data on cost-effectiveness of 9vHPV, and data on burden of type-specific HPV-associated disease in the United States. Summaries of reviewed evidence and Work Group discussions were presented to ACIP before recommendations were proposed. Recommendations were approved by ACIP in February 2015. Evidence supporting 9vHPV use was evaluated using the Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) framework .HPV is associated with cervical, vulvar, and vaginal cancer in females, penile cancer in males, and anal cancer and oropharyngeal cancer in both females and males –10. The In a phase III efficacy trial comparing 9vHPV with 4vHPV among approximately 14,000 females aged 16 through 26 years, 9vHPV efficacy for prevention of ≥CIN2, vulvar intraepithelial neoplasia grade 2 or 3, and vaginal intraepithelial neoplasia grade 2 or 3 caused by HPV 31, 33, 45, 52, or 58 was 96.7% in the per protocol populationTwo immunobridging trials were conducted. One compared 9vHPV in approximately 2,400 females and males aged 9 through 15 years with approximately 400 females aged 16 through 26 years. Over 99% seroconverted to all nine HPV vaccine types; GMTs were significantly higher in adolescents aged 9 through 15 years compared with females aged 16 through 26 years. In a comparison of 4vHPV with 9vHPV in approximately 600 adolescent females aged 9 through 15 years, 100% seroconverted to HPV 6, 11, 16, and 18 in both groups, and GMTs were noninferior in the 9vHPV group compared with the 4vHPV group.Immunogenicity in males aged 16 through 26 years was compared with females of the same age group in a separate study. In both females and males, >99% seroconverted to all nine HPV vaccine types, and GMTs in males were noninferior to those in females .The immunogenicity of concomitant and nonconcomitant administration of 9vHPV with quadrivalent meningococcal conjugate vaccine  and tetanus, diphtheria, acellular pertussis vaccine  was evaluated. The GMTs were noninferior for all nine HPV vaccine types in the co-administered group . For Menactra, the noninferiority criterion was met for all four serogroups, and for Adacel, for diphtheria, tetanus, and all four pertussis antigens.Safety has been evaluated in approximately 15,000 subjects in the 9vHPV clinical development program; approximately 13,000 subjects in six studies were included in the initial application submitted to FDA . The vacIntroduction of 9vHPV in both males and females was cost-saving when compared with 4vHPV for both sexes in a cost-effectiveness model that assumed 9vHPV cost $13 more per dose than 4vHPV. Cost-effectiveness ratios for 9vHPV remained favorable compared with 4vHPV  when varying assumptions about HPV natural history, cervical cancer screening, vaccine coverage, vaccine duration of protection, and health care costs, but were sensitive to 9vHPV cost assumptions . BecauseACIP recommends that routine HPV vaccination be initiated at age 11 or 12 years. The vaccination series can be started beginning at age 9 years. Vaccination is also recommended for females aged 13 through 26 years and for males aged 13 through 21 years who have not been vaccinated previously or who have not completed the 3-dose series  recommends routine HPV vaccination at age 11 or 12 years. The vaccination series can be started beginning at age 9 years. Vaccination is also recommended for females aged 13 through 26 years and for males aged 13 through 21 years who have not been vaccinated previously or who have not completed the 3-dose series. Males aged 22 through 26 years may be vaccinated. ACIP recommends vaccination of men who have sex with men and immunocompromised persons through age 26 years if not vaccinated previously.Why are the recommendations being updated now?9-valent HPV vaccine (9vHPV) was approved by the Food and Drug Administration on December 10, 2014. This vaccine targets HPV types 6, 11, 16, and 18, the types targeted by the quadrivalent HPV vaccine (4vHPV), as well as five additional types, HPV types 31, 33, 45, 52, and 58. ACIP reviewed results of a randomized trial among approximately 14,000 females aged 16 through 26 years that showed noninferior immunogenicity for the types shared by 4vHPV and 9vHPV and high efficacy for the five additional types. Other trials in the 9vHPV clinical development program included studies that compared antibody responses across age groups and females and males and concomitant vaccination studies. The evidence supporting 9vHPV vaccination was evaluated using the Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) framework and determined to be type 2 (moderate level of evidence) among females and 3 (low level of evidence) among males; the recommendation was designated as a Category A recommendation .What are the new recommendations?9vHPV, 4vHPV or 2vHPV can be used for routine vaccination of females aged 11 or 12 years and females through age 26 years who have not been vaccinated previously or who have not completed the 3-dose series. 9vHPV or 4vHPV can be used for routine vaccination of males aged 11 or 12 years and males through age 21 years who have not been vaccinated previously or who have not completed the 3-dose series. ACIP recommends either 9vHPV or 4vHPV vaccination for men who have sex with men and immunocompromised persons (including those with HIV infection) through age 26 years if not vaccinated previously.2vHPV, 4vHPV, and 9vHPV are each administered in a 3-dose schedule. The second dose is administered at least 1 to 2 months after the first dose, and the third dose at least 6 months after the first doseIf vaccination providers do not know or do not have available the HPV vaccine product previously administered, or are in settings transitioning to 9vHPV, any available HPV vaccine product may be used to continue or complete the series for females for protection against HPV 16 and 18; 9vHPV or 4vHPV may be used to continue or complete the series for males. There are no data on efficacy of fewer than 3 doses of 9vHPV.HPV vaccination is recommended through age 26 years for men who have sex with men and for immunocompromised persons (including those with HIV infection) who have not been vaccinated previously or have not completed the 3-dose series.HPV vaccines are contraindicated for persons with a history of immediate hypersensitivity to any vaccine component. 4vHPV and 9vHPV are contraindicated for persons with a history of immediate hypersensitivity to yeast. 2vHPV should not be used in persons with anaphylactic latex allergy.HPV vaccines are not recommended for use in pregnant women . If a wohttp://vaers.hhs.gov.Adverse events occurring after administration of any vaccine should be reported to VAERS. Additional information about VAERS is available by telephone (1–800–822–7967) or online at Cervical cancer screening is recommended beginning at age 21 years and continuing through age 65 years for both vaccinated and unvaccinated women ,20. RecoA clinical trial is ongoing to assess alternative dosing schedules of 9vHPV. ACIP will formally review the results as data become available. HPV vaccination should not be delayed pending availability of 9vHPV or of future clinical trial data.


The identification of mothers in whose buccal smears HPV was detected can help develop a group of children who run a relatively significant risk of being infected.The impact of human papillomavirus (HPV) infection on pregnancy is a major problem of medicine. The transmission of the virus from mother to fetus is a process yet unresolved. The immune response and changed hormonal status of pregnant women might facilitate infection. A research on the prevalence of HPV infection was conducted at the Clinic of Obstetrics, Medical University of Lublin (Poland). The studied group included 152 randomly selected women. The material was tested for the presence of HPV DNA by means of polymerase chain reaction (PCR). The aim of the research was to assess the relation between HPV infections detected in the buccal smears of the neonates and the incidence of such infections in the cervical/buccal smears of their mothers. In the group of 152 infants HPV was found in 16 (10.53%). Among the cervical/buccal smears, HPV was isolated, respectively, in 24 (15.79%) and in 19 (12.5%) pregnant women. Statistically significant differences in the prevalence of HPV swabs from the newborns and the cervical/buccal smears of their mothers were found ( The influence of human papillomavirus infection on pregnancy remains an unresolved matter. HPV infection is commonly included in the group of sexually transmitted diseases (STDs) and is one of the most frequently transmitted among them , 2. The n = 7), (2) abnormal cervical smear (n = 4), and (3) multiple pregnancy (n = 6). The subjects gave their written, informed consent before the start of any procedure. The protocol of this study was approved by the Local Bioethical Committee at the Medical University of Lublin. Prior to delivery, smears were obtained from the cervix and buccal mucosa of each pregnant woman. The cervical HPV DNA specimen was collected with a cytobrush inserted into the vagina. The cervix and ectocervix were swabbed using a circular motion. The oral specimens were collected during the same time. Immediately after the birth, the neonatal oral mucosa smears were obtained in similar proceedings with a cytobrush. All samples were collected by the same, properly trained person. The material was placed in sterile tubes for HPV DNA testing , frozen, and stored at 70°C until further analyses. The material was tested for the presence of HPV DNA by means of PCR method. Identification of HPV types was performed using consensus PCR primers for L1:MY09: 5′-CGTCCMARRGGAWACTGATC-3′ and MY11: 5′-GCMCAAGGWCATAAYAATGG-3′, where M = A + C, R = A + G, W = A + T, and Y = C + T. This set of primers amplifies DNA from at least 33 different HPV genotypes. HPV types 16 and/or 18 were identified using the following type specific PCR primers: HPV16/L1A/HPV16/L1B, 5′-GCCTGTGTAGGTGTTGAGGT-3′, and 5′-TGGATTTACTCCAACATTGG-3′ product size: 264 bp; HPV18/L1A/HPV18/L1B, 5′-GTGGACCAGCAAATACAGGA-3′, and 5′-TGCAACGACCACGTGTTGGA-3′, product site: 162 bp; HPV18ME12/HPV18ME50/E6, 5′-CACGGCGACCCTACAAGCTACCTG-3′, and 5′-TGCAGCACGAATTGGCACTGGCCTC-3′, product site: 404 bp. The reaction was performed according to Tucker et al. [μL of PCR mixture contained 1 μM of primers, 200 μM of deoxynucleotide triphosphates, 1x PCR buffer , the investigated DNA (10 ng/μL), and Tag polymerase at a final concentration of 40 U/mL. After preliminary denaturation (15 min at 94°C), samples were amplified for 31 cycles in a thermal cycler. Each cycle consisted of the following steps: denaturation at 94°C for 30 seconds and annealing at 59°C for the same amount of time, followed by primer extension at 72°C for 1 min. In the last PCR cycle, the stage of complementary DNA synthesis at 72°C was extended to 420 seconds. PCR products were analysed using agarose gel electrophoresis in the presence of pBluescript DNA digested with HindI. The presence of HPV DNA was additionally verified, and HPV genotypes were identified by direct sequencing from CR reaction tube (without purification from MY09 and MY11 oligonucleotides). The results of sequencing were analysed using the BLAST database (http://blast.ncbi.nlm.nih.gov/).This study was conducted between March 2009 and June 2011 and included 152 pregnant women who delivered at the Clinic of Obstetrics and Pathology of Pregnancy, Medical University of Lublin (Poland). The studied group included random obstetric population recruited prior to the delivery. The routine medical examination was followed by previous history of HPV-related genital diseases and treatments. The study had the following inclusion criteria: (1) singleton pregnancy, (2) lack of clinical symptoms suggesting the possibility of HPV infection, (3) normal cervical smear, and (4) term delivery. The number of women who refused was 17. The exclusion criteria included (1) history of HPV infection  package, with the level of significance set at p ≤ 0.05.Normal distribution of continuous variables was verified with the Kolmogorov-Smirnoff test. Statistical characteristics of these variables were presented as arithmetic means and standard deviations (SD), and their intergroup comparisons were performed with Student's p < 0.001). There is an eleven times greater relative risk (OR = 11.363 CI: 3.536–36.509) of discovering HPV DNA in buccal smears of newborn of a HPV-positive mother than of the mother with negative results of HPV. The correlation of the frequency of HPV infections was presented in As the result of the conducted research HPV DNA was isolated in 12.5% (19/152) of swabs from the oral mucosa of the mothers and in 10.53% (16/152) of the children. In the group of nineteen mothers with HPV infection we found 8 (42.11%) HPV-positive children and 11 (57.89%) HPV-negative children. In the mothers group without HPV infection 133/152 (87.5%) we observed 8/133 (6.02%) HPV (+) newborns and 125/133 (93.98%) HPV (−) newborns. A statistically significant difference in the prevalence of HPV DNA in the swabs from the oral mucosa of the newborns and the swabs from the oral mucosa of their mothers was found  HPV DNA was isolated in 24 (15.79%) swabs from the mothers' cervix, and in 16 (10.52%) from oral mucosa of newborns. Out of 24 HPV-positive mothers, HPV DNA was isolated in 11 (45.83%) of their children. In group of 24 HPV-positive mothers we found 13 (54.17%) HPV-negative children. In the mothers group without HPV infection 128/152 (84.21%) we observed 5/128 (3.91%) HPV (+) newborns and 123/128 (96.09%) HPV (−) newborns. A statistically significant difference in the prevalence of HPV DNA in the swabs from the oral mucosa of the newborns and the swabs from the cervix of their mothers was found (p < 0.001). Based on the analysis of the results the relative risk of HPV DNA existing in swabs from the oral mucosa of the children whose mothers were tested positive for HPV in cervical smears is almost 21 times greater than in children whose mothers were diagnosed as healthy . The correlation between HPV infection in oral mucosa of children and the cervix of mother was presented in In the studied group . Among the swabs of the cervical smears HPV DNA was isolated in 24/152 (15.79%) women and among the swabs from the oral mucosa HPV DNA was observed in 19/152 (12.5%). Both variables are separable and analysed as such. Simultaneous HPV infection in both swabs from the oral mucosa and the cervix was found in 14/152 (9.21%) of the total population. No HPV DNA was found in any of the assessed swabs taken from 123/152 (80.92%) women giving birth. As a result of the research a statistically significant difference in HPV DNA incidence in cervical smears in relation to HPV DNA occurrence in buccal smears of women was found (p < 0.001). Out of 19 women in whose swabs from the oral mucosa HPV DNA was identified the virus was also found in 14 swabs from cervix. There is a 34 times greater relative risk  of HPV DNA incidence in swabs from cervix if HPV DNA is found in oral mucosa smears of the patient. The correlation between HPV infections in the cervical/buccal smears of woman giving birth was presented in HPV DNA was identified in 29 women giving birth which was 19.08% of the total population (p < 0.0005). Place of residence of the patients might be another factor that determines the frequency of HPV infections. In pregnant and nonpregnant women in Uganda HPV DNA is detected in not less than 60% of cases, whereas it is estimated that 6.5% of pregnant women in Spain are infected [A well-documented relation between the frequency of HPV infections, young age, contraceptives intake, smoking, number of sexual partners, marital status, and level of education obtained exists , 21, 22.infected , 25. A rinfected . Furtherinfected . Similarinfected . What isinfected . Presentinfected , 30, 31.infected , 33. Higinfected , 18, 27.infected . One of infected . Similarinfected , 37. Smainfected –40. TherA connection between buccal HPV incidence in mothers and newborns was proved in the research. A statistically significant difference in HPV DNA incidence in cervical smears in relation to HPV DNA occurrence in mothers' buccal smears was found. A thorough examination and follow-up of HPV (+) mothers' newborns may be vital for the selection of a group of children prone to HPV-related entities.


HPV type 16 leads to altered biological functions with possible clinical significance in the different geographical locations. Sixteen major variants (V1-V16) in full length L1 gene of HPV-16 were identified following analysis of 250 prospectively collected cervical cancer tissue biopsies and their effect on immunogenicity was studied. The effect of these major variations on the epitopes were predicted by in silico methods and the immunogenicity of variants and respective reference DNA vaccine constructs were evaluated by administration of prepared DNA vaccine constructs in female BALB/c mice to evaluate antibody titer. In the present study, L500F (V16) variation showed a significant ~2.7 fold (p < 0.002) increase in antibody titer, whereas T379P (V8) showed ~0.4 fold (p < 0.328) decrease after final injection. These results showed a promising roadmap for the development of DNA based vaccine and for the generation of effective response, though there is a need to study more prevalent variants of HPV in the Indian population.Cervical cancer is one of the most common gynecological cancers in the world but in India, it is the top most cancer among women. Persistent infection with high-risk human papillomaviruses (HR-HPVs) is the most important risk factor. The sequence variation(s) in the most common HR-HPV  Cervical cancer (CaCx) is the third most common cancer among women worldwide with an estimate of 527,624 new cases diagnosed annually and is the most common gynecological cancer in developing counties like India78Identification of HPV as a major causative agent for cervical cancer gives an opportunity to prevent it by vaccine development. The major capsid (L1) and minor capsid (L2) proteins of HPV are attractive candidates and are extensively used for prophylactic vaccine development as they induce virus-specific immune response and have highly immunogenic repetitive epitopes on the surface of virions and have no oncogenic activity. Earlier studies have reported that variations in L1 gene can affect the viral assembly, immunological recognition by the host and immortalization activity which ultimately affect the protein structure or conformation and lead to altered biological functions with clinical significance11The role of intra-type variants among HPVs cannot be ruled out; therefore, intratype genomic diversity of HPV sequence is important for the development of efficient diagnostic/prognostic tools and vaccine development. For efficient vaccine, the recognition of correct epitope sequence is important for the generation of efficient immune response13in-silico) as well as on potential immunogenicity in-vivo (BALB/c mice).It is, therefore, imperative to understand the geographical variants of HPV for better targeting the vaccines against it. In India, very limited studies have been carried out on molecular variant analysis of full length L1 of HPV-161617Out of 250 tumor biopsies, 231 showed HPV infection (92.4%) of which 221/231 (95.6%) samples harbored HPV-16; 4/231 (1.7%) was infected with HPV-18, 2/231 (0.8%) showed co-infection of both HPV-16/HPV-18 and the remaining 4/231 (1.7%) had infection with other HPV sub types.We observed 16 major variations (V1-V16) in full length L1 ; 13 bialOn further analysis, it was observed that variations V3, V12 and V13 were observed in all HPV 16 positive samples (100%), which correspond to amino acid change from histidine to aspartate at position H228D, an insertion of serine residue at 448 and deletion of aspartate residue at 465 position respectively . V6 corrin-silico analysis showed the replacement of threonine by proline at 379 causing distortion of a sheet structure (disappeared), that may be due to the unusual structure of proline , leucine (L) and proline (P) were the most prominent amino acids with threonine being the most variable amino acid noted. There is an increase in frequency of asparagine, phenyl alanine and serine while there is a decrease in frequency of histidine, threonine in the mutant as compared to reference . The pre proline . Figure  proline . RamachaWe identified India specific major variations of L1, which may play an important role in immunogenicity. Previously, it was showed that amino acids from 494 to 518 of L1 were known as hypervariable epitope constructs (HEC) regions19Structure of the epitope was predicted and docked with antibody (1JRH) using Patchdock and best model refinement was done by Firedock. The best docked model having lowest global energy for V16R, V16V, 8R and 8V were selected and visualized in chimera. The comparison of respective reference and the variant peptide showed a new hydrogen bond in case of V16V and loss of hydrogen bond in case of V8V, which also causes change in global energy i.e. −61.67 for V8R and −54.60 for V8V, where as in for V16R and V16V were −49.55 and 50.86, respectively . In V8V,in-silico results was evaluated. Around 100 μg of prepared plasmid constructs of reference and variants i.e. pV8R & pV8V and pV16R & pV16V, respectively were injected in BALB/c mice quadriceps muscles at four weeks interval for three times. After two weeks of final injection, isolated serum of mice were proceeded for evaluation of anti-HPV-16 L1 antibody titer by ELISA. An induction of circulating IgG class anti-HPV16 L1 antibodies was observed in vaccinating mice  in comparison to the reference construct  to form VLP of HPV shows a lot of promise for the prevention of papillomavirus-associated cancer and two available VLP based vaccines Gardasil and Cervarix currently in use. However, their high cost of production and ineffectiveness against other strains of HR-HPV as well as not having therapeutic value3435in-vivo experimental results also confirmed that the variant construct had ~0.4 fold less antibody titer than reference DNA vaccine construct. Probably, this may be due to the structure constraint of proline having pyrrolidine ring which gives unusual shape resulting into conformational constraints or secondary structural preferences -immunized mice showed breath of reactivity and represent the immunodominant B-cell epitopes of the major capsid proteins of HPVferences . On the in-silico results. So, this study shows promising results which appear to be useful for the development of India specific cost effective DNA based vaccines.Recently, several reports showed that naked DNA vaccination with HPV-16 L1 in mice/or pig could produce increased level of serum IgG, anti-L1antibodies38In conclusion, there is no study in Indian population that shows the effect of different variations of HR-HPV, and their effect on pathogenicity/antigencity. Our results showed the major variations like T379P of full length L1 gene which may play important role in the immunogenicity against HPV by affecting the binding affinity of immunogenic peptide (epitope). On the other hand, L500F (V16) was found to be more immunogenic than the reference DNA vaccine constructs. T379P (V8) variation causes the reduction of immunogenicity of epitope present in their vicinity. Therefore, these variations may be important for the oncogenic potential of HPV-16 bearing these mutations. So, we need to target potent epitopic sites to evoke the effective immune response. These results show a roadmap for the development of epitope based vaccine and for the generation of effective response, there is a need to study the more prevalent variants of HPV in specific geographical populations. In future, on the basis of this study priority may be given on the development of multi-epitope DNA vaccine, which may be a better alternative for the development of effective immune response against different variants. There was a study showing the effect of different variants using series of VLP prepared, showed the structural importance of it to induce effective immune response (IgG) and viral assembly for HPV mediated carcinogenesis(ICPO-ICMR/IEC/2007/P-001). Written informed consent from all the patients was obtained and the study was carried out in accordance with the principles of Helsinki declaration.A total of 250 CaCx tissue biopsy samples were collected from the Cancer Clinic of Department of Obstetrics and Gynecology, Lok Nayak Jai Prakash Hospital, New Delhi and control tissue from Department of Obstetrics and Gynecology, Safdarjung Hospital, New Delhi. The samples comprised of 102 biopsies of tumor stage I + II and 148 biopsies of stage (III + IV). Standard International Federation of Obstetrics and gynecology (FIGO) criteria was used to determine the tumor stage. All the tissue biopsies were collected in sterile sample collection vials  containing chilled phosphate-buffered saline (PBS) and stored at −80 °C in a deep freezer for further processing. The study was approved by the ethics committee of the institute Genomic DNA was extracted by standard method using Proteinase-K digestion followed by phenol/chloroform treatment as described earlier28xl). For variant analysis, Multiple Sequence alignment (MSA) of sequenced sample was done by Mega 5.1 softwareThe full length L1 gene of HPV-16 positive cases was amplified using forward primer 5′-AGCATCGATACCATGGCTCTTTGGCTGCCTAGTG AGG-3′ and reverse primer 5′-GCGGCCGCTTACAGCTTACGTTTTTGCGTTTAGCA GTTGTAG-3′for 1.5 kbp product. The amplified products were purified by Exonuclease-Shrimp Alkaline Phosphatase (Exo-SAP) treatment and directly sequenced according to the manufacturer’s protocol (ABI-3103http://web.expasy.org/translate/) for determination of amino acid change. Computation of various physical and chemical parameters of protein sequences was calculated by Protparam, an online server (http://web.expasy.org/protparam/)http://bioinf.cs.ucl.ac.uk/psipred/) was used, which provides a simple and accurate secondary structure prediction method. Using a very stringent cross validation method to evaluate the method’s performance, PSIPRED3.2 achieves an average Q3 score of 81.6%http://www.iedb.org/).Nucleotide sequences were translated by translate tool of ExPASy server (http://bioinfo3d.cs.tau.ac.il/PatchDock/index.html) and the refinement of the protein-protein docking solutions was done using FireDock (http://bioinfo3d.cs.tau.ac.il/FireDock/index.html) and the best model was selected on the basis of the lowest global energy.The structure of variant and reference sequence was modeled by taking HPV L1 structure (PDB: 1DZL A) as template with Modeller 9.10 of Sali labThe segment of L1 gene having V8 and V16 variants and their reference was amplified by using specific primers . The amp(ACBR/12/IAEC/2856(a)).Plasmid DNA was dissolved in 0.9% saline without adjuvant at a concentration of 1 mg/ml. A total of 36 BALB/c mice (6 ± 8-week-old) were grouped into control, pcDNA, 16R, 16V, 8R and 8V (n = 6/group) and immunized intramuscularly with 50 μg of plasmid into each quadriceps muscles , three times at 4 weeks interval. Blood was taken before each immunization and the mice were sacrificed two weeks after the third injection, serum was separated and stored at −80 °C. The animal study was approved by animal ethics committee at Dr. B. R. Ambedkar Center for Biomedical Research, University of Delhi, Delhi The antibody titer in response to injected constructs has been detected by direct Enzyme-linked immunosorbent assay (ELISA) according to the method described previouslyAll the statistical analysis was calculated using GraphPad InStat version3.0 software. One way ANOVA was used to determine the statistical significance of experimental groups with control. For statistical significance of reference and their respective variant were analyzed by applying Mann Whitney test (non parametric test). The p-values less than 0.05 were considered significant.How to cite this article: Kumar, A. et al. Identification and validation of immunogenic potential of India specific HPV-16 variant constructs: In-silico & in-vivo insight to vaccine development. Sci. Rep.5, 15751; doi: 10.1038/srep15751 (2015).


Australia commenced human papillomavirus (HPV) vaccination in 2007, with a two-year catch-up to the age of 26; catch-up cohorts are thus now entering their thirties. Plans for monitoring vaccine impact involve pre- and post-vaccination assessment of cervical HPV DNA in the general population and in high grade abnormalities. Although HPV serology is less sensitive than DNA genotyping, it assesses lifetime exposure and may be easier to measure in the general population. However, benchmark pre-vaccination seroprevalence of vaccine-included types in unvaccinated women with high grade abnormalities has not previously been reported.We assessed seroprevalence for HPV16/18 from a population-based sample of 3,729 women with normal cytology and 971 women with confirmed high grade abnormalities (CIN2/3), aged 30–64 years, unvaccinated, and recruited in New South Wales in 2006–2010. We examined the variation in HPV16/18 seropositivity by age and in relation to a range of reproductive and behavioural characteristics in the subgroup of normal cytology women with no recent history of high grade cervical disease.The HPV 16, 18 and combined seroprevalence was 19%, 7% and 24% among women with normal cytology, and 39%, 13% and 44% among women with CIN2/3, respectively. For both groups, HPV16/18 seroprevalence was highest at age 30–39 years and decreased with age. In multivariable analysis for women with normal cytology, HPV16 and HPV18 seropositivity were each associated with the number of lifetime sexual partners  and for HPV16 this was also associated with age (p-trend <0.001) and prior diagnosis of Chlamydia .The findings of this study inform pre-vaccination estimates of HPV seropositivity in women with normal cytology and women with high grade abnormalities. Almost a quarter of unvaccinated women aged over 30 years with normal cytology, and more than 40% of those with CIN2/3, had seroconverted to HPV 16 or 18. These findings provide a potential additional benchmark for assessing the effects of HPV vaccination.The online version of this article (doi:10.1186/s12879-014-0676-z) contains supplementary material, which is available to authorized users. Human papillomavirus (HPV) types 16 and 18 are the most frequent oncogenic HPV types detected in invasive cervical cancers and precancerous cervical intraepithelial neoplasia grade 3 (CIN3) worldwide . In 2013HPV surveillance is an important mechanism of monitoring HPV prevalence at the population level, to assess the potential effects of cross-protection against non-vaccine-included types, and to exclude potential type-replacement of the vaccine-included types. HPV genotyping of normal cervical smears and for those with high grade abnormalities pre- and post-vaccination, has been proposed as part of such a program in Australia  and moreFor serology-based surveillance, benchmark data on seroprevalence in unvaccinated women are required. Large studies assessing age-stratified, pre-vaccination seroprevalence data in women with and without cervical dysplasia are sparse. In Australia, we identified only one study , using aData and samples for this analysis have been obtained from the Cervical Health Study, a large population-based case–control study conducted in New South Wales (NSW), which was designed primarily to assess the role of exogenous hormonal co-factors in the development of cervical pre-cancerous lesions. Recruitment was conducted between December 2006 and July 2011 in women aged 20–64 years, via invitation letters issued from the centralised NSW Pap Test Register (PTR) which records all cervical cytology and histology test results, as well as past disease, except for those associated with <1% of women who opt-out. General Practices were first approached to consent for invitations to be sent to women under their care. Invitation letters were then issued to eligible women with a new record of a cytological/histological test. Cases had a new registry record of a low or high grade precancerous cytological or histological abnormality while controls had a new record of a normal cytological result in the same week as the abnormal smear for the matched case. Screening histories for up to 5 years prior to study participation were obtained from the PTR. Following informed consent, participants completed a self-administered questionnaire assessing demographic and medical details, and lifestyle factors. In the first phase of the study (to 2010) participants optionally provided a blood sample (women aged 30–64 years only). The Cervical Health Study commenced prior to the vaccination catch-up program implementation (covering ages of 12–26 years) but continued throughout the duration of the program. Therefore, vaccination status was confirmed by self-report.For the current analysis, HPV16/18 seropositivity was investigated in women with histologically confirmed CIN2/3 and in women with normal cytology. Data has been obtained from the Cervical Health Study, however, analyses were restricted to women aged 30 years of age and above, who provided both a questionnaire and a blood sample. Women who indicated in their questionnaire that they had been vaccinated against HPV were excluded (n = 4). Analyses were conducted on 4700 women including 3,729 women with normal cytology and 971 with histologically confirmed CIN2/3.Blood samples were collected by phlebotomists from December 2006 to August 2010 at several laboratories in NSW. Blood was collected in serum separating tubes, left to clot at room temperature for 30 mins, centrifuged, aliquoted into 2 ml aliquots within 48 hours of collection and stored at −80°C until analysis. Serum samples were pre-randomised by study number prior to aliquoting to ELISA plates. 210 samples were analysed in duplicate. Samples were tested ‘blind’ at the University of Queensland for circulating antibodies to the HPV16/18 late capsid protein L1 using a HPV-neutralising assay based on pseudoviruses carrying a secreted alkaline phosphatase reporter gene as described by Pastrana et al. . A samplThe seroprevalences of HPV16, HPV18 and HPV16/18 combined, in the two groups of women were estimated and 95% confidence intervals (CIs) were calculated using the exact binomial method. Subjects were categorised into age groups .trachomatis). Associations between seropositivity and use of hormone contraceptives or use hormone replacement therapy were not assessed because the numbers of HPV16 or HPV18 seropositive users  were too low to provide sufficient statistical power for sub-group analysis to be conducted. A complete case approach was adopted for the analysis. Dose response associations (p-trend) were examined for ordinal variables which could be treated as continuous assuming a linear trend in the models and p-nominal values were estimated for all variables. All analyses were conducted using SAS software, version 9.3.Factors associated with seropositivity to HPV16 only or HPV18 only were investigated in a subset of 3,462 of the control women with an index negative smear and a history of normal cytology in the last five years. To evaluate the characteristics associated with each HPV type separately, women (N = 267) who were seropositive for both HPV types were excluded from these analyses. In doing so we followed the same analytical strategy as a major prior National Cancer Institute (NCI) USA study . As in tThe study was approved in 2004 by the Cancer Institute NSW Population Ethics Committee; reference number Ref 2004/05/073.The overall HPV 16, 18 and combined HPV16 or 18 seroprevalence was 19.4% , 7.1%  and 23.5%  among women with normal cytology, and 38.5% (374/971), 12.7% (123/971) and 43.8% (425/971) among women with CIN2/3, respectively. Table trachomatis was associated with an increased risk of being HPV16 seropositive . After multivariable adjustment no other significant associations were seen.Table Results for HPV18 are presented in Table trachomatis were significantly associated with HPV16 seropositivity.To our knowledge this is the first study to provide population-based benchmarking information on HPV16 and 18 serology in unvaccinated women with normal cytology and with high grade abnormalities in Australia. The seroprevalence measurements obtained in unvaccinated women in this study indicate high levels of exposure to HPV 16/18, most notably in women in their thirties. As expected, seroprevalence for both HPV types was higher in women with CIN 2/3 than those with normal cytology. The highest seroprevalence in both groups was observed in 30–39 year old women and it decreased with age. We also found that the number of lifetime sexual partners was associated with the detection of HPV16 and 18 antibodies in the subgroup of women with a history of normal cytology and that this association was due to past exposure. In this subgroup, younger age and ever having a diagnosis of C.Population cervical HPV DNA prevalence for oncogenic types tends to peak earlier than seroprevalence, with the DNA prevalence pattern reflecting a high risk of current infection around the time that young women first initiate sexual activity. Age-specific patterns of infection for whole-of-population cervical HPV DNA prevalence tend to be comparable but slightly higher than those for women in the population with normal cytology , and most studies of population DNA prevalence, including the IARC HPV surveys , report In terms of seroprevalence, our finding of decreasing HPV16 and 18 seroprevalence with increasing age in unvaccinated mid-adult and older women with normal cytology is consistent with other Australian data, although, studies done with different assays and at different laboratories complicate comparisons across studies. For the age groups for which comparative data were available, we observed a close correspondence between our findings and the general population findings of Newall et al. , which iThe underlying reasons for declining seroprevalence of these HPV types at older ages are uncertain, but may include waning of naturally-acquired immunity over time and/or changes in the risk of HPV infection within successive birth cohorts over the past several decades. Waning anti-HPV16 antibody levels could in part explain our observation of a decreasing prevalence with age  althoughtrachomatis infection and HPV16 seropositivity; having a previous C. trachomatis infection may be a marker of additional risk beyond that associated with the lifetime number of partners per se, which could occur, for example, if it acted as a marker for contact with higher risk partners. A similar association has been observed previously between herpes simplex virus-2 seroprevalence and HPV seropositivity ,, and reflsitivity . No evidA number of studies have reported on pre-vaccination cervical HPV DNA prevalence in women with confirmed high grade abnormalities in various populations. Our recent study in New Zealand reported that the HPV16/18 DNA prevalence in women with CIN2/3, peaked among women 20–29 years of age and decreased with older age , indicatThis study is subject to potential limitations. Participants were recruited from the NSW Pap Test Register and therefore results may not be representative of the general female population of NSW. Although few women opt-out of the register, recent national figures indicate that only 57% of women 20–69 years of age attended cervical screening . FurtherBy providing information on seroprevalence of HPV types 16 and 18 in unvaccinated women with normal cytology and with high grade abnormalities, this study establishes the possibility of using serology for monitoring changes in population level exposure over time. However, as HPV seropositivity due to natural infection is based on repeated and persistent exposure rather than incident infection, serology results have to be interpreted with caution; furthermore vaccine exposure will lead to complex serological patterns for vaccine-included types in cohorts offered vaccination. We also note that when used in combination with individual-level information on vaccination status , serology could be used to measure natural HPV 16 and 18 exposure in populations where vaccinated women mostly received the vaccine at a young enough age to be protected from HPV 16 and 18 infection. This is the expectation for Australia, in years to come, given that successive cohorts of 12–13 year old schoolgirls will receive the vaccine through the vaccination program and given that the government no longer subsidises the vaccination of older women. In such populations, vaccinated women can be reasonably assumed to be unexposed to natural infection with HPV 16 and 18 because of the protection provided by the vaccine. Serology may then have a role to estimate HPV 16/18 exposure in the unvaccinated remainder of the population.


Human population density in the coastal zone and potential impacts of climate change underscore a growing conflict between coastal development and an encroaching shoreline. Rising sea-levels and increased storminess threaten to accelerate coastal erosion, while growing demand for coastal real estate encourages more spending to hold back the sea in spite of the shrinking federal budget for beach nourishment. As climatic drivers and federal policies for beach nourishment change, the evolution of coastline mitigation and property values is uncertain. We develop an empirically grounded, stochastic dynamic model coupling coastal property markets and shoreline evolution, including beach nourishment, and show that a large share of coastal property value reflects capitalized erosion control. The model is parameterized for coastal properties and physical forcing in North Carolina, U.S.A. and we conduct sensitivity analyses using property values spanning a wide range of sandy coastlines along the U.S. East Coast. The model shows that a sudden removal of federal nourishment subsidies, as has been proposed, could trigger a dramatic downward adjustment in coastal real estate, analogous to the bursting of a bubble. We find that the policy-induced inflation of property value grows with increased erosion from sea level rise or increased storminess, but the effect of background erosion is larger due to human behavioral feedbacks. Our results suggest that if nourishment is not a long-run strategy to manage eroding coastlines, a gradual removal is more likely to smooth the transition to more climate-resilient coastal communities. In the United States, the coastline is now the most densely populated region in the country, and population density continues to increase . The coaTo manage this vulnerability along sandy coastlines, humans nourish beaches—placing sand, typically from offshore sources, onto the beach to widen the beach and combat erosion . EmpiricPredictions of increased rates of sea level rise and changing storminess imply that coastal vulnerability will increase –9. HowevWe use a stochastic dynamic optimization model to explore the influence of storms, rising sea level, and changing management approaches on coastal property values. The optimal nourishment frequency for the beach determined by the model reflects a property owner’s point of view, taking physical forcing and property values into consideration. We represent the evolution of the beach as three components: chronic background erosion , erosion that results as beaches and shorelines adjust to nourishment events, and erosion from punctuated storm events. The erosion from stochastic storm events is represented as a loss of the entire nourished portion of the beach that tends to occur during a hurricane . Optimalo). Beach width at any given time (t) is characterized by:We explore how the coupled physical-economic coastline depends upon changes in policy and physical forcing through a positive analysis in which managers follow optimal beach nourishment practice. While there is a large body of work on normative analysis and valuation studies of coastal economies –17, our Benefits from wide beaches are capitalized in to coastal property values and total benefits for an interval T between two nourishment projects are:(x(t)β)dtwhere A Nourishment costs include fixed costs (c) and variable costs (Φ) that depend on the amount of nourishment sand required to return the beach to its initial width:1, τ2, τ3, represent times between loss of nourished beach width (times between nourish events or storms). If storms are Poisson distributed and planned nourishment interval is denoted as T then the cumulative distribution function for the independent random variable τi is written as:We extend the optimal rotation model above, which is rooted in forest economics, to include punctuated storm events. This extension follows the introduction of stochastic forest fires into optimal forest management . We assuk) from the kth nourishment episode are:With the possibility of a storm, the net benefits π (τk). After a storm, the nourished portion of the beach is lost and benefits accrue only from the remaining beach, which faces background linear erosion [If a storm occurs before the next nourishment (T), then benefits from replenishment (nourished portion of the beach) accrue only to the moment the storm occurs Click here for additional data file.S1 FigHistorical (black dots) and modeled nourishment (red circles) intervals for four coastal locations: Wrightsville Beach (WB), Carolina Beach (CB), Emerald Isle (EI) and Kure Beach (KB). Error bars correspond to the 95% confidence intervals.(EPS)Click here for additional data file.