Risk factors and patterns of household clusters of respiratory viruses in rural Nepal.

Viral pneumonia is an important cause of death and morbidity among infants worldwide. Transmission of non-influenza respiratory viruses in households can inform preventative interventions and has not been well-characterised in South Asia. From April 2011 to April 2012, household members of pregnant women enrolled in a randomised trial of influenza vaccine in rural Nepal were surveyed weekly for respiratory illness until 180 days after birth. Nasal swabs were tested by polymerase chain reaction for respiratory viruses in symptomatic individuals. A transmission event was defined as a secondary case of the same virus within 14 days of initial infection within a household. From 555 households, 825 initial viral illness episodes occurred, resulting in 79 transmission events. The overall incidence of transmission was 1.14 events per 100 person-weeks. Risk of transmission incidence was associated with an index case age 1-4 years (incidence rate ratio (IRR) 2.35; 95% confidence interval (CI) 1.40-3.96), coinfection as initial infection (IRR 1.94; 95% CI 1.05-3.61) and no electricity in household (IRR 2.70; 95% CI 1.41-5.00). Preventive interventions targeting preschool-age children in households in resource-limited settings may decrease the risk of transmission to vulnerable household members, such as young infants.

Early emergence of an H275Y mutation in a hematopoietic cell transplant recipient treated with intravenous peramivir.

Oseltamivir resistance in pandemic 2009 influenza A/H1N1 is caused by the neuraminidase mutation H275Y. This mutation has also been associated with in vitro resistance to peramivir, but few clinical cases have been described to date. Using allele-specific real-time reverse transcriptase polymerase chain reaction assay for the H275Y mutation, we were able to identify resistant H1N1 in a hematopoietic cell transplant recipient receiving intravenous peramivir therapy, and through serial testing we determined the molecular evolution of resistance. This case demonstrates that an H275Y mutant population can emerge early and replicate in vivo under peramivir antiviral pressure to become the major viral population.

Diagnostic accuracy of an allele-specific reverse transcriptase-PCR assay targeting the H275Y oseltamivir resistant mutation in 2009 pandemic influenza A/H1N1 virus.

BACKGROUND: Oseltamivir resistant 2009 pandemic influenza A/H1N1 viruses (pH1N1) are emerging and rapid molecular assays identifying these strains are needed for clinical management. OBJECTIVE: Development and evaluation of an allele-specific, real-time reverse transcriptase-PCR assay (ASPCR) targeting the H275Y oseltamivir resistant mutation in pH1N1 virus. STUDY DESIGN: ASPCR uses two allele-specific forward primers (wild-type and mutant) and a common reverse primer and probe. Wild-type and mutant genotypes were defined by the difference in PCR Ct values (DeltaCt(mut-wt)) between the mutant primer and wild-type primer amplification curves for the same sample. Mixtures of wild-type and mutant genotypes were analyzed to evaluate sensitivity and determine assay cut-off values. ASPCR results were confirmed using an allelic discrimination assay (AD) and pyrosequencing. RESULTS: Mixtures containing 5-95% mutant genotype could be detected. A DeltaCt(mut-wt)>or=3.5 identified wild-type genotype (<10% mutant); between 3.5 and -3.5 identified mixed genotypes (10-90% mutant); and 90% mutant). Among 264 clinical samples, 171 were wild-type, 10 were mixed, and 29 were fully mutant. The 39 samples with mixed or mutant results were from 11 patients. Of 107 samples with sufficient volume tested by ASPCR and AD, 12 were indeterminate by AD due to low viral load, 86 were wild-type by both assays, and 9 were mutant by both assays. Thirteen samples were confirmed by pyrosequencing and one discrepant sample was mixed by ASPCR and fully mutant by pyrosequencing. CONCLUSIONS: ASPCR is sensitive, quantitative and specific for H275Y mutation analysis and provides an accurate approach for detecting pH1N1 oseltamivir resistance in clinical samples.

Comparison of conventional and molecular detection of respiratory viruses in hematopoietic cell transplant recipients.

BACKGROUND: Sensitive detection of respiratory viruses is important for early diagnosis of infection in patients following hematopoietic cell transplantation (HCT). To evaluate the relative sensitivity of respiratory virus detection in specimens from HCT recipients, we compared the results of conventional and quantitative molecular methods. METHODS: We tested 688 nasal wash samples collected prospectively from 131 patients during the first 100 days after HCT by viral culture, fluorescent antibody staining (FA), and real-time quantitative reverse transcription-polymerase chain reaction (PCR) assay for detection of respiratory syncytial virus (RSV), influenza virus types A (FluA) and B (FluB), and parainfluenza virus types 1 (PIV1) and 3 (PIV3). Testing for human metapneumovirus (MPV) was performed only by PCR. Data regarding 10 respiratory symptoms were collected with each sample. RESULTS: By any method 37 specimens were positive for a respiratory virus; 34 were positive by PCR, 15 by culture, and 6 by FA. Four specimens were positive by all 3 methods (3 RSV, 1 FluA). One specimen was positive for PIV1, and 2 were positive for rhinovirus by culture alone. Specimens positive by PCR alone included 2 RSV, 2 PIV1, 8 PIV3, and 8 MPV. In 10 specimens positive for RSV, PIV, or influenza virus collected from patients reporting no respiratory symptoms, 9, 4, and 1 specimen were positive by PCR, culture, and FA, respectively. Overall, specimens positive only by PCR had significantly fewer viral copies/mL (mean log(10)=4.32) than specimens positive by both PCR and culture (mean log(10)=5.75; P=0.002) or PCR and FA (mean log(10)=6.83; P<0.001). CONCLUSIONS: FA testing alone did not detect a significant proportion of respiratory virus-positive samples in HCT recipients, especially in patients with no respiratory symptoms and patients with PIV detection. PCR increased the yield of positive specimens 2 times relative to culture and more than 4 times relative to FA. Detection of respiratory viruses by PCR alone was associated with lower virus quantities and with fewer reported respiratory symptoms compared with concomitant detection by both PCR and conventional methods, indicating that PCR may be important to detect asymptomatic or mildly symptomatic stages of respiratory viral infections.

Maximum accuracy evaluation scheme for wireless saw delay-line sensors.

This paper describes an evaluation scheme that prevents phase ambiguity of surface acoustic wave (SAW) delay-line sensors. Although it is well-known that phase evaluation yields accuracies of 150~1500 times higher than time-delay evaluation, the problem of phase ambiguity has prevented phase evaluation of sensors operating over a range larger than 2 pi. This paper addresses this unsolved problem with a complete strategy. Furthermore, the existence of an optimum choice of the relative reflector positions on the sensor is shown. The presented relations enable the design of maximum accuracy SAW delay-line sensors.

Excellence in sexually transmitted infection (STI) diagnostics: recognition of past successes and strategies for the future.

Diagnostic advances do not generally receive the recognition given to prevention and treatment contributions, for the control and management of infectious diseases including sexually transmitted infections (STIs). In order to identify seminal diagnostic contributions over a half century (1950-2000), the Editorial Board of the WHO Sexually Transmitted Diseases Diagnostics Initiative (SDI) Publication Review or "electronic journal club" were asked to nominate their choices of peer-reviewed publications for special recognition. From 43 nominations, 13 were voted by a panel of 25 "experts" as having made the most significant contributions. The 1964 article by Thayer and Martin, which identified a selective media for gonococcal culture, was chosen unanimously by all panel members and is identified as the classic STI diagnostic article for this era.

Disseminating sexually transmitted infections diagnostics information: the SDI web publication review series.

OBJECTIVES: The World Health Organization Sexually Transmitted Diseases Diagnostics Initiative (SDI) website publication review seeks to provide health care providers in all geographic and economic settings with timely, critical, and concise information concerning new developments in laboratory and field diagnosis of sexually transmitted infections (STI). METHODS: Since 2003, the website (www.who.int/std_diagnostics/literature_reviews) has disseminated information in the form of annotated abstracts and commentaries on articles covering studies of STI laboratory-based and rapid assays that are commercially available or under development. Articles identified through searches of PubMed, specific journals, and by referrals from Editorial Board members are selected for inclusion if they meet pre-specified criteria. The objectives, methods, results, and conclusions for each article are summarised and board members are invited to prepare commentaries addressing study design and applicability of findings to end users. RESULTS: Currently, 91 STI diagnostics experts from 17 countries on six continents serve on the Editorial Board. Twelve quarterly issues have been posted that include summaries of 214 original and 17 review articles published from January 2002 through March 2005, with expert commentaries on 153 articles. Interest in the site has increased every year. In 2005, over 36 700 unique visitors from more than 100 countries viewed over 75,000 pages of information. CONCLUSIONS: The SDI Publication Review series has the potential to contribute to SDI's goal of improving care for patients with STI by increasing knowledge and awareness of STI diagnostics. Given the proliferation of internet-based STI testing services, this website may be broadened to meet the needs of a wider range of users.

[No major difference between population screening for cervical carcinoma at the present screening interval of 5 years and the former interval of 3 years]. / Geen groot verschil tussen bevolkingsonderzoek op cervixcarcinoom bij het huidige screeningsinterval van 5 en het vorige van 3 jaar.

OBJECTIVE: To assess the effect of extending the screening interval from 3 to 5 years on the detection of premalignant changes and invasive cervical carcinoma in the restructured population screening programme. DESIGN: Retrospective follow-up study. METHOD: The results were collected of the 1st round (1996-2000; 277, 377 women) and a part of the 2nd round (2001; 49,622 women; screening interval: 5 years) of the screening programme in Region West, the Netherlands. Histoscores for cervical intraepithelial neoplasia (CIN) 3 and squamous cell carcinoma (n/100 women investigated) and the hit count (sum of the histoscores for CIN 3, adenocarcinoma in situ and (micro)invasive cervical carcinoma) were calculated. Data of women with adenocarcinoma in situ and endocervical (adeno)carcinoma were recorded separately. The results of the 1st and 2nd round of the current screening programme (commenced in 1996) were compared with those of the historical screening programme that commenced in 1976 (screening interval: 3 years). RESULTS: From the 1st to the 2nd round of the historical screening programme that commenced in 1976, the histoscores for CIN 3 (3.33, 1.88) and squamous cell carcinoma (0.53, 0.19) and the hit count (3.92, 2.15) all diminished significantly. The current restructured programme, which commenced in 1996, showed low starting values for all three parameters, comparable to those in the 2nd round of the 1976 programme; a further reduction (0.16, 0.08; p < 0.01) was seen only in the histoscore for squamous cell carcinoma. In both rounds of both programmes, the histoscores for adenocarcinoma in situ (0.02, 0.02, 0.05, 0.04, respectively) and endocervical adenocarcinoma (0.04, 0.06, 0.05, 0.04) remained stable. CONCLUSION: In the current cervical carcinoma screening programme, with a screening interval of 5 years, the hit count of serious abnormalities remained constant while the incidence of squamous cell carcinoma decreased; this is in contrast to the historical screening programme (commenced in 1976), when both the hit count and the histoscore for CIN 3 diminished significantly. There were indications that cervical screening has no beneficial effect on the prevention of cervical adenocarcinoma.

Papanicolaou test screening and prevalence of genital human papillomavirus among women who have sex with women.

OBJECTIVES: The purpose of this study was to examine frequency of and attitudes toward Papanicolaou (Pap) test screening in women who have sex with women (WSW) and to determine prevalence of genital human papillomavirus (HPV). METHODS: Women were eligible if they reported having engaged in sex with another woman in the preceding year Medical and sexual histories were obtained. Cervical specimens for Pap tests and cervical and vaginal specimens for HPV DNA testing were collected. RESULTS: HPV DNA was detected in 31 of 248 WSW (13%). Women who had never had sex with men were less likely to have undergone pelvic examinations and had fewer recent Pap tests. Reasons for not undergoing Pap tests included lack of insurance, previous adverse experiences, and belief that Pap tests were unnecessary. CONCLUSIONS: Despite the occurrence of genital HPV, WSW do not receive adequate Pap test screening. Pap test screening recommendations should not differ for WSW, regardless of sexual history with men.

Human papillomaviruses and cervical cancer in Bangkok. I. Risk factors for invasive cervical carcinomas with human papillomavirus types 16 and 18 DNA.

Personal interviews, tests for antibodies to herpes simplex virus type 2, Treponema pallidum, and hepatitis B, tests for hepatitis B surface antigen (HBsAg), and polymerase chain reaction-based assays for human papillomavirus (HPV) DNA in cervical scrapings were obtained from 190 women with squamous cell and 42 women with adenomatous cervical carcinoma and from 291 hospitalized controls diagnosed in Bangkok, Thailand, between September 1991 and September 1993. Risk was strongly associated with oncogenic HPV types, with types 16 and 18 predominating in squamous and adenomatous lesions, respectively. The 126 cases with HPV-16 and the 42 cases with HPV-18 were compared with 250 controls with no evidence of any HPV. The risk of both viral tumor types increased with decreasing age at first intercourse in this predominantly monogamous population, which may be explained by more visits to prostitutes by the husbands of cases with early than late age at first intercourse. HPV-16 tumors were weakly associated with HBsAg carrier state and smoking. The risk of tumors of both viral types increased with parity and use of oral contraceptives but not with injectable progestogens. Factors that may predispose to persistent, oncogenic HPV-16 or -18 infection may include estrogens or progestins in the presence of estrogens, immunosuppression, and smoking, but other factors related to low socioeconomic status are also involved.

Human papillomaviruses and cervical cancer in Bangkok. II. Risk factors for in situ and invasive squamous cell cervical carcinomas.

To identify risk factors for progression of intraepithelial cervical lesions, 190 women with invasive cervical cancer were compared with 75 women with in situ disease diagnosed in Bangkok, Thailand, between September 1991 and September 1993. Polymerase chain reaction-based assays for type-specific human papillomavirus (HPV) DNA in cervical scrapings revealed oncogenic types in 79% of invasive and 57% of intraepithelial tumors. Types 16 and 18, but not types 31/33/35/39, were more common in invasive than intraepithelial tumors, and untyped HPV DNA was found more commonly in the in situ lesions, suggesting that in situ disease is four times more likely to become invasive if due to type 16 or 18 than to other causes, and that tumors with only untyped HPV are not at increased risk of progression. After controlling for HPV type, the risk of developing invasive diseases, compared with the risk of developing intraepithelial lesions, was not related to any of a large number of sexual and hormonal factors considered or to smoking, suggesting that any cofactors these variables represent act before the development of in situ carcinoma. Two indices of socioeconomic status were associated with a reduced risk of only invasive disease, suggesting the existence of unknown protective factors that operate after intraepithelial lesions develop.

Human papillomaviruses and cervical cancer in Bangkok. III. The role of husbands and commercial sex workers.

Between September 1991 and September 1993, husbands of women with and without cervical neoplasia and commercial sex workers in one brothel and one massage parlor in Bangkok, Thailand, were interviewed; serologic tests for sexually transmitted infections were performed; and cervical and penile scrapings were tested for human papillomavirus (HPV) DNA. The risks of cervical carcinoma in monogamous women and of oncogenic HPV in their husbands were associated with the men's having unprotected intercourse with prostitutes. The prevalence of oncogenic HPV was higher in commercial sex workers than in women attending gynecologic and family planning clinics. Oncogenic HPV prevalence declined with age in human immunodeficiency virus (HIV)-negative, but not in healthy HIV-positive, commercial sex workers and was weakly associated with hepatitis B antigenemia, suggesting that persistence of HPV infection is due to subtle changes in immunity. Associations of HPV with recent pregnancy and oral contraceptive use suggest that hormonal factors may increase the risk of cervical neoplasia by enhancing persistence of HPV infection. The prevalence of high-grade squamous intraepithelial lesions was strongly related to oncogenic HPV types and weakly to HIV infection only in their presence. Commercial sex workers in Bangkok are reservoirs of oncogenic HPV, and cervical cancer in monogamous Thai women develops in part as a result of transmission of these viruses to them by their husbands from prostitutes.

Concurrent and sequential acquisition of different genital human papillomavirus types.

Coinfection with multiple types of genital human papillomavirus (HPV) has been reported, but how frequently it occurs and whether prior infection with specific HPV types inhibits subsequent infection by related types are not known. To address this, 518 women were followed for an average of 2.9 years, and behavioral information and cervical and vulvovaginal swabs for HPV DNA assay were obtained at 4-month intervals. A polymerase chain reaction-based method was used to detect types frequently found in cervical cancers (HPV 16, 18, 31, and 45) and in genital warts (HPV 6 and 11). Concurrent acquisition of multiple types occurred more often than expected by chance. However, no 2 types were more or less likely to be acquired concurrently than any other 2 types. When considering sequential acquisition of HPV types, we found that risk of acquiring a new HPV type was not decreased among those with prior infection by a phylogenetically related or unrelated type (hazard ratio [95% confidence interval], 1.0 [0.4-3.0] and 1.3 [0.8-2.1], respectively).

Comparison of human papillomavirus types 16, 18, and 6 capsid antibody responses following incident infection.

The relationship between human papillomavirus (HPV) DNA in the genital mucosa and serum IgG to HPV-16, -18, and -6 was studied in a cohort of 588 college women. Among women with incident HPV infections, 59.5%, 54.1%, and 68.8% seroconverted for HPV-16, -18, or -6, respectively, within 18 months of detecting the corresponding HPV DNA. Transient HPV DNA was associated with a failure to seroconvert following incident HPV infection; however, some women with persistent HPV DNA never seroconverted. Antibody responses to each type were heterogeneous, but several type-specific differences were found: seroconversion for HPV-16 occurred most frequently between 6 and 12 months of DNA detection, but seroconversion for HPV-6 coincided with DNA detection. Additionally, antibody responses to HPV-16 and -18 were significantly more likely to persist during follow-up than were antibodies to HPV-6.

Detection of cervical antibodies to human papillomavirus type 16 (HPV-16) capsid antigens in relation to detection of HPV-16 DNA and cervical lesions.

A more sensitive luminescence immunoassay (LIA) for human papillomavirus type 16 (HPV-16) was developed and used to measure HPV-16 antibodies in cervical samples from 292 college-aged women who were examined at 4-month intervals. Of the 609 collected samples, IgG, IgA, and secretory piece-associated antibodies to HPV-16 were detected in 12%, 6%, and 8%, respectively, of samples tested. Cervical IgG antibodies were most strongly associated with HPV-16 DNA detected within the previous 12 months (odds ratio, 3.3; 95% confidence interval, 1.4-7.8). Secretory IgA (cervical IgA- and secretory piece-positive) was most strongly associated with detection of a squamous intraepithelial lesions 4-8 months earlier (odds ratio, 6.4; 95% confidence interval, 1.9-21.8). As with serum HPV-16 antibodies, there appears to be a several-month delay between cervical HPV infection and detection of cervical antibodies.

Association of antiretroviral therapy with detection of HIV-1 RNA and DNA in the anorectal mucosa of homosexual men.

OBJECTIVE: To determine whether combination antiretroviral therapy is associated with reduced detection of HIV-1 RNA and DNA in the anorectal mucosa of men who have sex with men (MSM). DESIGN: Cross-sectional study of 233 MSM recruited from community and clinic sites in Seattle, Washington between July 1996 and December 1997. METHODS: HIV-1 RNA and HIV-1 DNA were detected in anorectal swab specimens by polymerase chain reaction amplification assays. RESULTS: HIV-1 RNA was detected significantly less often in anorectal specimens from users of combination antiretroviral therapies, whether a protease inhibitor was received (15/89; 17%) or not (16/53; 30%), than in men not receiving therapy (43/88; 49%) (P < 0.001, P = 0.03, respectively). In contrast, HIV-1 DNA was detected only slightly less frequently in anorectal specimens obtained from men receiving protease inhibitors (35/81; 43%) or reverse transcriptase inhibitors alone (22/48; 46%) than in specimens from men not receiving therapy (45/78; 58%) (P = 0.07, P = 0.20, respectively). Among men with < 50 copies HIV-1 RNA/ml plasma, detection of HIV-1 RNA in anorectal specimens was rare (1/54; 2%) but detection of HIV-1 DNA was common (14/50; 28%). CONCLUSIONS: Combination antiretroviral therapy is associated with reductions in HIV-1 RNA, but HIV-1 DNA remains detectable in the anorectal canal of almost half of MSM receiving such therapy. Condom use during anal intercourse should be encouraged, regardless of plasma viral load response to potent antiretroviral therapy.

Etiology of genital ulcer disease in Dakar, Senegal, and comparison of PCR and serologic assays for detection of Haemophilus ducreyi.

We used PCR assays to determine the etiology of genital ulcers in patients presenting to a sexually transmitted disease clinic in Dakar, Senegal, and evaluated the ability of two PCR tests (groEL and recD) and two serological tests (adsorption enzyme immunoassay [EIA] and lipooligosaccharide [LOS] EIA) to detect current Haemophilus ducreyi infection. We found that in this population, H. ducreyi, T. pallidum, and herpes simplex virus HSV DNA were detected in 56, 15, and 13% of 39 genital ulcer specimens, respectively, and H. ducreyi DNA was detected in 60% (3 of 5) of samples from ulcerated bubos. Among 40 consecutive patients with genital ulcer disease and with sufficient sample for both PCR assays, the recD and groEL H. ducreyi PCR assays were 83% concordant, with the recD PCR assay detecting six (15%) additional positive specimens and the groEL assay detecting one (3%) additional positive specimen. Compared to PCR, the adsorption EIA and LOS EIA tests had sensitivities of 71 and 59% and specificities of 57 and 90%, respectively, for the diagnosis of current H. ducreyi infection. While these differences in specificity could be due either to previous infection with H. ducreyi or to the detection of cross-reacting antibodies, only 6% of patients from a nearby family planning clinic gave a positive reaction in both the adsorption EIA and LOS EIA assays, indicating that cross-reacting antibodies are not prevalent among clinic attendees in this city. Our studies indicate that the adsorption EIA detects both current and past infection, while the LOS EIA assay is more specific for current infection with H. ducreyi in this population.

A matched prospective study of human immunodeficiency virus serostatus, human papillomavirus DNA, and cervical lesions detected by cytology and colposcopy.

OBJECTIVE: To compare the prevalence and type of human papillomavirus (HPV) infections in the genital tract of human-immunodeficiency-virus- (HIV) seropositive and -seronegative women matched for cytology and to examine prospectively the relationship of HPV DNA, colposcopic findings and cervical squamous intraepithelial lesions (SIL) in these matched seropositive and seronegative cohorts. METHODS: A matched prospective study of HIV-seropositive and -seronegative women undergoing cytologic screening, colposcopy, and testing for HPV DNA and other infections at each visit. RESULTS: Twenty-three HIV-seropositive women were matched with 23 seronegative women by cervical cytology reading, lifetime number of sexual partners, age, and follow-up length. Fourteen pairs of these women had follow-up visits every 4 months, for 56 and 53 total visits in seropositive and seronegative women, respectively. After matching, the groups had a similar overall prevalence of HPV DNA and of HPV oncogenic (high risk) types at baseline. On follow up, HIV-seropositive women were more likely than seronegative women to develop SIL (38% vs. 10%), less likely to have negative cytology (34% vs. 60%, overall P = 0.03), more visits with HPV DNA detected (68% vs. 40% P = 0.04), and more visits with multiple HPV DNA types detected (18% vs. 0%, P = 0.02). Colposcopic lesions in the seropositive women were more likely to have sharp borders or mosaicism or to be thick white (P = 0.009). CONCLUSIONS: After matching for baseline Papanicolaou smear readings, these data suggest that over time seropositive women have more visits that yield abnormal cytology, more persistent HPV DNA detection, and more colposcopic abnormalities than seronegative women.