Monitoring influenza vaccine effectiveness using the national influenza surveillance system

Background: Flu vaccine composition is reformulated on a yearly basis. As such, the vaccine effectiveness (VE) from previous seasons cannot be considered for subsequent years, and it is necessary to monitor the VE for each season. This study (MonitorEVA- monitoring vaccine effectiveness) intends to evaluate the feasibility of using the national influenza surveillance system (NISS) for monitoring the influenza VE. Material and methods: Data was collected within NISS during 2004 to 2014 seasons. We used a case-control design where laboratory confirmed incident influenza like illness (ILI) patients (cases) were compared to controls (ILI influenza negative). Eligible individuals consisted on all aged individuals that consult a general practitioner or emergency room with ILI symptoms with a swab collected within seven days of symptoms onset. VE was estimated as 1- odds ratio of being vaccinated in cases versus controls adjusted for age and month of onset by logistic regression. Sensitivity analyses were conducted to test possible effect of assumptions on vaccination status, ILI definition and timing of swabs (<3 days after onset). Results: During the 2004-2014 period, a total of 5302 ILI patients were collected but 798 ILI were excluded for not complying with inclusion criteria. After data restriction the sample size in both groups was higher than 148 individuals/ season; minimum sample size needed to detect a VE of at least 50% considering a level of significance of 5% and 80% power. Crude VE point estimates were under 45% in 2004/05, 2005/06, 2011/12 and 2013/14 season; between 50%-70% in 2006/07, 2008/09 and 2010/11 seasons, and above 70% in 2007/08 and 2012/13 season. From season 2006/07 to 2013/14, all crude VE estimates were statistically significant. After adjustment for age group and month of onset, the VE point estimates decreased and only 2008/09, 2012/13 and 2013/14 seasons were significant. Discussion and Conclusions: MonitorEVA was able to provide VE estimates for all seasons, including the pandemic, indicating if the VE was higher than 70% and less than 50%. When comparing with other observational studies, MonitorEVA estimates were comparable but less precise and VE estimates were in accordance with the antigenic match of the circulating virus/ vaccine strains. Given the sensitivity results, we propose a MonitorEVA based on: a) Vaccination status defined independently of number of days between vaccination and symptoms onset; b) use of all ILI data independent of the definition; c) stratification of VE according to time between onset and swab (< 3 and ≥3 days).

The nasal microbiota in infants with cystic fibrosis in the first year of life: a prospective cohort study.

BACKGROUND Respiratory tract infections and subsequent airway inflammation occur early in the life of infants with cystic fibrosis. However, detailed information about the microbial composition of the respiratory tract in infants with this disorder is scarce. We aimed to undertake longitudinal in-depth characterisation of the upper respiratory tract microbiota in infants with cystic fibrosis during the first year of life. METHODS We did this prospective cohort study at seven cystic fibrosis centres in Switzerland. Between Feb 1, 2011, and May 31, 2014, we enrolled 30 infants with a diagnosis of cystic fibrosis. Microbiota characterisation was done with 16S rRNA gene pyrosequencing and oligotyping of nasal swabs collected every 2 weeks from the infants with cystic fibrosis. We compared these data with data for an age-matched cohort of 47 healthy infants. We additionally investigated the effect of antibiotic treatment on the microbiota of infants with cystic fibrosis. Statistical methods included regression analyses with a multivariable multilevel linear model with random effects to correct for clustering on the individual level. FINDINGS We analysed 461 nasal swabs taken from the infants with cystic fibrosis; the cohort of healthy infants comprised 872 samples. The microbiota of infants with cystic fibrosis differed compositionally from that of healthy infants (p=0·001). This difference was also found in exclusively antibiotic-naive samples (p=0·001). The disordering was mainly, but not solely, due to an overall increase in the mean relative abundance of Staphylococcaceae in infants with cystic fibrosis compared with healthy infants (multivariable linear regression model stratified by age and adjusted for season; second month: coefficient 16·2 [95% CI 0·6-31·9]; p=0·04; third month: 17·9 [3·3-32·5]; p=0·02; fourth month: 21·1 [7·8-34·3]; p=0·002). Oligotyping analysis enabled differentiation between Staphylococcus aureus and coagulase-negative Staphylococci. Whereas the analysis showed a decrease in S aureus at and after antibiotic treatment, coagulase-negative Staphylococci increased. INTERPRETATION Our study describes compositional differences in the microbiota of infants with cystic fibrosis compared with healthy controls, and disordering of the microbiota on antibiotic administration. Besides S aureus, coagulase-negative Staphylococci also contributed to the disordering identified in these infants. These findings are clinically important in view of the crucial role that bacterial pathogens have in the disease progression of cystic fibrosis in early life. Our findings could be used to inform future studies of the effect of antibiotic treatment on the microbiota in infants with cystic fibrosis, and could assist in the prevention of early disease progression in infants with this disorder. FUNDING Swiss National Science Foundation, Fondation Botnar, the Swiss Society for Cystic Fibrosis, and the Swiss Lung Association Bern.

Predator species identification from saliva at kill sites with limited remains

Thesis (Master's)--University of Washington, 2016-06

Function and Regulation of Cytochrome P450 4V2 and the Implications in Bietti’s Crystalline Dystrophy

Thesis (Ph.D.)--University of Washington, 2016-06

Parvovirus B19 infection presenting as 'bathing trunk' erythema with pustules

A 7-year-old girl presented with acute vulval erythema and pustules, associated with a petechial eruption in her flexures and over her feet. There was a mild prodromal illness and the patient was afebrile. There were minimal symptoms associated with the rash. Skin and throat swabs were negative and blood examination showed mild neutrophilia and lymphopaenia. Parvovirus B19 IgM was detected on serology and cutaneous features resolved within 4 days. This is a further case of parvovirus B19 infection presenting as a 'bathing trunk' exanthem that has unique dermatologic features, including the presence of pustules and distant petechiae. © 2006 The Australasian College of Dermatologists.

Real-time PCR assays for detection of bocavirus in human specimens

The recently discovered human bocavirus (HBoV) is the first member of the family Parpoviridae, genus Bocavirus, to be potentially associated with human disease. Several studies have identified HBoV in respiratory specimens from children with acute respiratory disease, but the full spectrum of clinical disease and the epidemiology of HBoV infection remain unclear. The availability of rapid and reliable molecular diagnostics would therefore aid future studies of this novel virus. To address this, we developed two sensitive and specific real-time TaqMan PCR assays that target the HBoV NS1 and NP-1 genes. Both assays could reproducibly detect 10 copies of a recombinant DNA plasmid containing a partial region of the HBoV genome, with a dynamic range of 8 log units (10(1) to 10(8) copies). Eight blinded clinical specimen extracts positive for HBoV by an independent PCR assay were positive by both real-time assays. Among 1,178 NP swabs collected from hospitalized pneumonia patients in Sa Kaeo Province, Thailand, 53 (4.5%) were reproducibly positive for HBoV by one or both targets. Our data confirm the possible association of HBoV infection with pneumonia and demonstrate the utility of these real-time PCR assays for HBoV detection.

Rapid testing for group B streptococcus during labour: a test accuracy study with evaluation of acceptability and cost-effectiveness

OBJECTIVE: To determine the accuracy, acceptability and cost-effectiveness of polymerase chain reaction (PCR) and optical immunoassay (OIA) rapid tests for maternal group B streptococcal (GBS) colonisation at labour. DESIGN: A test accuracy study was used to determine the accuracy of rapid tests for GBS colonisation of women in labour. Acceptability of testing to participants was evaluated through a questionnaire administered after delivery, and acceptability to staff through focus groups. A decision-analytic model was constructed to assess the cost-effectiveness of various screening strategies. SETTING: Two large obstetric units in the UK. PARTICIPANTS: Women booked for delivery at the participating units other than those electing for a Caesarean delivery. INTERVENTIONS: Vaginal and rectal swabs were obtained at the onset of labour and the results of vaginal and rectal PCR and OIA (index) tests were compared with the reference standard of enriched culture of combined vaginal and rectal swabs. MAIN OUTCOME MEASURES: The accuracy of the index tests, the relative accuracies of tests on vaginal and rectal swabs and whether test accuracy varied according to the presence or absence of maternal risk factors. RESULTS: PCR was significantly more accurate than OIA for the detection of maternal GBS colonisation. Combined vaginal or rectal swab index tests were more sensitive than either test considered individually [combined swab sensitivity for PCR 84% (95% CI 79-88%); vaginal swab 58% (52-64%); rectal swab 71% (66-76%)]. The highest sensitivity for PCR came at the cost of lower specificity [combined specificity 87% (95% CI 85-89%); vaginal swab 92% (90-94%); rectal swab 92% (90-93%)]. The sensitivity and specificity of rapid tests varied according to the presence or absence of maternal risk factors, but not consistently. PCR results were determinants of neonatal GBS colonisation, but maternal risk factors were not. Overall levels of acceptability for rapid testing amongst participants were high. Vaginal swabs were more acceptable than rectal swabs. South Asian women were least likely to have participated in the study and were less happy with the sampling procedure and with the prospect of rapid testing as part of routine care. Midwives were generally positive towards rapid testing but had concerns that it might lead to overtreatment and unnecessary interference in births. Modelling analysis revealed that the most cost-effective strategy was to provide routine intravenous antibiotic prophylaxis (IAP) to all women without screening. Removing this strategy, which is unlikely to be acceptable to most women and midwives, resulted in screening, based on a culture test at 35-37 weeks' gestation, with the provision of antibiotics to all women who screened positive being most cost-effective, assuming that all women in premature labour would receive IAP. The results were sensitive to very small increases in costs and changes in other assumptions. Screening using a rapid test was not cost-effective based on its current sensitivity, specificity and cost. CONCLUSIONS: Neither rapid test was sufficiently accurate to recommend it for routine use in clinical practice. IAP directed by screening with enriched culture at 35-37 weeks' gestation is likely to be the most acceptable cost-effective strategy, although it is premature to suggest the implementation of this strategy at present.

Intrapartum tests for group B streptococcus:accuracy and acceptability of screening

Objective: To assess the accuracy and acceptability of polymerase chain reaction (PCR) and optical immunoassay (OIA) tests for the detection of maternal group B streptococcus (GBS) colonisation during labour, comparing their performance with the current UK policy of risk factor-based screening. Design Diagnostic test accuracy study. Setting and population Fourteen hundred women in labour at two large UK maternity units provided vaginal and rectal swabs for testing. Methods The PCR and OIA index tests were compared with the reference standard of selective enriched culture, assessed blind to index tests. Factors influencing neonatal GBS colonisation were assessed using multiple logistic regression, adjusting for antibiotic use. The acceptability of testing to participants was evaluated through a structured questionnaire administered after delivery. Main outcome measures The sensitivity and specificity of PCR, OIA and risk factor-based screening. Results Maternal GBS colonisation was 21% (19-24%) by combined vaginal and rectal swab enriched culture. PCR test of either vaginal or rectal swabs was more sensitive (84% [79-88%] versus 72% [65-77%]) and specific (87% [85-89%] versus 57% [53-60%]) than OIA (P <0.001), and far more sensitive (84 versus 30% [25-35%]) and specific (87 versus 80% [77-82%]) than risk factor-based screening (P <0.001). Maternal antibiotics (odds ratio, 0.22 [0.07-0.62]; P = 0.004) and a positive PCR test (odds ratio, 29.4 [15.8-54.8]; P <0.001) were strongly related to neonatal GBS colonisation, whereas risk factors were not (odds ratio, 1.44 [0.80-2.62]; P = 0.2). Conclusion Intrapartum PCR screening is a more accurate predictor of maternal and neonatal GBS colonisation than is OIA or risk factor-based screening, and is acceptable to women. © RCOG 2010 BJOG An International Journal of Obstetrics and Gynaecology.

Progesterone release from a silicone intravaginal ring in pigtailed macaques

Background: Heterosexual HIV transmission continues to spread worldwide. Intravaginal rings (IVRs) formulated with antiretroviral drugs hold great promise for HIV prevention in women. IVRs provide the benefit of being coitally-independent and coitally-covert for an extended period. As a proof-of-concept, we tested the in vivo release of progesterone from a silicone elastomer vaginal ring device. Methods: Six female pig-tailed macaques were treated with a GnRH agonist (Lupron) prior to ring placement. Four macaques received a progesterone-loaded silicone ring, and two macaques received a blank silicone ring. Blood, vaginal swabs, CVL, and/or biopsies were collected during ring placement, and after ring removal. Results: The median plasma progesterone levels for macaques with a progesterone IVR were 13,973 pg/ml (day 3), 12,342 pg/ml (day 7), 10,112 pg/ml (day 14), 8445 pg/ml (day 21) and 8061 pg/ml (day 28), with a significant decrease from day 14 to day 21 (P = 0.0286). The median plasma progesterone levels for macaques with a blank IVR were 221±±± ±±88 pg/ml. Macaques with a progesterone IVR had CVL progesterone levels of 20,935 pg/ml (day 7), 6892 pg/ml (day 21) and 11,515 pg/ml (day 28). Macaques with a blank IVR had CVL progesterone levels of 29 �± 13 pg/ml. There were no disturbances to the normal vaginal microflora, and plasma and CVL cytokine analysis did not indicate a proinflammatory response due to ring placement. The vaginal biopsies did not display any pathology following ring removal. Overall, the IVRs were well tolerated without any indication of inflammation or significant changes in the vaginal compartment.

Perceptions of self-testing for chlamydia:understanding and predicting self-test use

Background: Self-testing technology allows people to test themselves for chlamydia without professional support. This may result in reassurance and wider access to chlamydia testing, but anxiety could occur on receipt of positive results. This study aimed to identify factors important in understanding self-testing for chlamydia outside formal screening contexts, to explore the potential impacts of self-testing on individuals, and to identify theoretical constructs to form a Framework for future research and intervention development. Methods: Eighteen university students participated in semi-structured interviews; eleven had self-tested for chlamydia. Data were analysed thematically using a Framework approach. Results: Perceived benefits of self-testing included its being convenient, anonymous and not requiring physical examination. There was concern about test accuracy and some participants lacked confidence in using vulvo-vaginal swabs. While some participants expressed concern about the absence of professional support, all said they would seek help on receiving a positive result. Factors identified in Protection Motivation Theory and the Theory of Planned Behaviour, such as response efficacy and self-efficacy, were found to be highly salient to participants in thinking about self-testing. Conclusions: These exploratory findings suggest that self-testing independently of formal health care systems may no more negatively impact people than being tested by health care professionals. Participants’ perceptions about self-testing behaviour were consistent with psychological theories. Findings suggest that interventions which increase confidence in using self-tests and that provide reassurance of test accuracy may increase self-test intentions.

The development of direct ultra-fast PCR for forensic genotyping using short channel microfluidic systems with enhanced sieving matrices

There are situations in which it is very important to quickly and positively identify an individual. Examples include suspects detained in the neighborhood of a bombing or terrorist incident, individuals detained attempting to enter or leave the country, and victims of mass disasters. Systems utilized for these purposes must be fast, portable, and easy to maintain. The goal of this project was to develop an ultra fast, direct PCR method for forensic genotyping of oral swabs. The procedure developed eliminates the need for cellular digestion and extraction of the sample by performing those steps in the PCR tube itself. Then, special high-speed polymerases are added which are capable of amplifying a newly developed 7 loci multiplex in under 16 minutes. Following the amplification, a postage stamp sized microfluidic device equipped with specially designed entangled polymer separation matrix, yields a complete genotype in 80 seconds. The entire process is rapid and reliable, reducing the time from sample to genotype from 1-2 days to under 20 minutes. Operation requires minimal equipment and can be easily performed with a small high-speed thermal-cycler, reagents, and a microfluidic device with a laptop. The system was optimized and validated using a number of test parameters and a small test population. The overall precision was better than 0.17 bp and provided a power of discrimination greater than 1 in 106. The small footprint, and ease of use will permit this system to be an effective tool to quickly screen and identify individuals detained at ports of entry, police stations and remote locations. The system is robust, portable and demonstrates to the forensic community a simple solution to the problem of rapid determination of genetic identity.

A novel method for rapid and selective extraction of male DNA from rape kits using alkaline lysis and pressure cycling technology (PCT)

There is an increasing demand for DNA analysis because of the sensitivity of the method and the ability to uniquely identify and distinguish individuals with a high degree of certainty. But this demand has led to huge backlogs in evidence lockers since the current DNA extraction protocols require long processing time. The DNA analysis procedure becomes more complicated when analyzing sexual assault casework samples where the evidence contains more than one contributor. Additional processing to separate different cell types in order to simplify the final data interpretation further contributes to the existing cumbersome protocols. The goal of the present project is to develop a rapid and efficient extraction method that permits selective digestion of mixtures. ^ Selective recovery of male DNA was achieved with as little as 15 minutes lysis time upon exposure to high pressure under alkaline conditions. Pressure cycling technology (PCT) is carried out in a barocycler that has a small footprint and is semi-automated. Typically less than 10% male DNA is recovered using the standard extraction protocol for rape kits, almost seven times more male DNA was recovered from swabs using this novel method. Various parameters including instrument setting and buffer composition were optimized to achieve selective recovery of sperm DNA. Some developmental validation studies were also done to determine the efficiency of this method in processing samples exposed to various conditions that can affect the quality of the extraction and the final DNA profile. ^ Easy to use interface, minimal manual interference and the ability to achieve high yields with simple reagents in a relatively short time make this an ideal method for potential application in analyzing sexual assault samples.^

A Novel Method for Rapid and Selective Extraction of Male DNA from Rape Kits using Alkaline Lysis and Pressure Cycling Technology (PCT)

There is an increasing demand for DNA analysis because of the sensitivity of the method and the ability to uniquely identify and distinguish individuals with a high degree of certainty. But this demand has led to huge backlogs in evidence lockers since the current DNA extraction protocols require long processing time. The DNA analysis procedure becomes more complicated when analyzing sexual assault casework samples where the evidence contains more than one contributor. Additional processing to separate different cell types in order to simplify the final data interpretation further contributes to the existing cumbersome protocols. The goal of the present project is to develop a rapid and efficient extraction method that permits selective digestion of mixtures. Selective recovery of male DNA was achieved with as little as 15 minutes lysis time upon exposure to high pressure under alkaline conditions. Pressure cycling technology (PCT) is carried out in a barocycler that has a small footprint and is semi-automated. Typically less than 10% male DNA is recovered using the standard extraction protocol for rape kits, almost seven times more male DNA was recovered from swabs using this novel method. Various parameters including instrument setting and buffer composition were optimized to achieve selective recovery of sperm DNA. Some developmental validation studies were also done to determine the efficiency of this method in processing samples exposed to various conditions that can affect the quality of the extraction and the final DNA profile. Easy to use interface, minimal manual interference and the ability to achieve high yields with simple reagents in a relatively short time make this an ideal method for potential application in analyzing sexual assault samples.

Surface-enhanced Raman spectroscopy of novel psychoactive substances using polymer-stabilized Ag nanoparticle aggregates

A set of seized "legal high'' samples and pure novel psychoactive substances have been examined by surface-enhanced Raman spectroscopy using polymer-stabilized Ag nanoparticle (Poly-SERS) films. The films both quenched fluorescence in bulk samples and allowed identification of mu g quantities of drugs collected with wet swabs from contaminated surfaces.

Non-invasive diagnosis of pediatric tuberculosis

Thesis (Master's)--University of Washington, 2016-08