A distinct isoform of lymphoid enhancer binding factor 1 (LEF1) epigenetically restricts EBV reactivation to maintain viral latency.

As a human tumor virus, EBV is present as a latent infection in its associated malignancies where genetic and epigenetic changes have been shown to impede cellular differentiation and viral reactivation. We reported previously that levels of the Wnt signaling effector, lymphoid enhancer binding factor 1 (LEF1) increased following EBV epithelial infection and an epigenetic reprogramming event was maintained even after loss of the viral genome. Elevated LEF1 levels are also observed in nasopharyngeal carcinoma and Burkitt lymphoma. To determine the role played by LEF1 in the EBV life cycle, we used in silico analysis of EBV type 1 and 2 genomes to identify over 20 Wnt-response elements, which suggests that LEF1 may bind directly to the EBV genome and regulate the viral life cycle. Using CUT&RUN-seq, LEF1 was shown to bind the latent EBV genome at various sites encoding viral lytic products that included the immediate early transactivator BZLF1 and viral primase BSLF1 genes. The LEF1 gene encodes various long and short protein isoforms. siRNA depletion of specific LEF1 isoforms revealed that the alternative-promoter derived isoform with an N-terminal truncation (ΔN LEF1) transcriptionally repressed lytic genes associated with LEF1 binding. In addition, forced expression of the ΔN LEF1 isoform antagonized EBV reactivation. As LEF1 repression requires histone deacetylase activity through either recruitment of or direct intrinsic histone deacetylase activity, siRNA depletion of LEF1 resulted in increased histone 3 lysine 9 and lysine 27 acetylation at LEF1 binding sites and across the EBV genome. Taken together, these results indicate a novel role for LEF1 in maintaining EBV latency and restriction viral reactivation via repressive chromatin remodeling of critical lytic cycle factors.

Retinoblastoma Protein Is Required for Epstein-Barr Virus Replication in Differentiated Epithelia.

Coinfection of human papillomavirus (HPV) and Epstein-Barr virus (EBV) has been detected in oropharyngeal squamous cell carcinoma. Although HPV and EBV replicate in differentiated epithelial cells, we previously reported that HPV epithelial immortalization reduces EBV replication within organotypic raft culture and that the HPV16 oncoprotein E7 was sufficient to inhibit EBV replication. A well-established function of HPV E7 is the degradation of the retinoblastoma (Rb) family of pocket proteins (pRb, p107, and p130). Here, we show that pRb knockdown in differentiated epithelia and EBV-positive Burkitt lymphoma (BL) reduces EBV lytic replication following de novo infection and reactivation, respectively. In differentiated epithelia, EBV immediate early (IE) transactivators were expressed, but loss of pRb blocked expression of the early gene product, EA-D. Although no alterations were observed in markers of epithelial differentiation, DNA damage, and p16, increased markers of S-phase progression and altered p107 and p130 levels were observed in suprabasal keratinocytes after pRb knockdown. In contrast, pRb interference in Akata BX1 Burkitt lymphoma cells showed a distinct phenotype from differentiated epithelia with no significant effect on EBV IE or EA-D expression. Instead, pRb knockdown reduced the levels of the plasmablast differentiation marker PRDM1/Blimp1 and increased the abundance of c-Myc protein in reactivated Akata BL with pRb knockdown. c-Myc RNA levels also increased following the loss of pRb in epithelial rafts. These results suggest that pRb is required to suppress c-Myc for efficient EBV replication in BL cells and identifies a mechanism for how HPV immortalization, through degradation of the retinoblastoma pocket proteins, interferes with EBV replication in coinfected epithelia. IMPORTANCE Terminally differentiated epithelium is known to support EBV genome amplification and virion morphogenesis following infection. The contribution of the cell cycle in differentiated tissues to efficient EBV replication is not understood. Using organotypic epithelial raft cultures and genetic interference, we can identify factors required for EBV replication in quiescent cells. Here, we phenocopied HPV16 E7 inhibition of EBV replication through knockdown of pRb. Loss of pRb was found to reduce EBV early gene expression and viral replication. Interruption of the viral life cycle was accompanied by increased S-phase gene expression in postmitotic keratinocytes, a process also observed in E7-positive epithelia, and deregulation of other pocket proteins. Together, these findings provide evidence of a global requirement for pRb in EBV lytic replication and provide a mechanistic framework for how HPV E7 may facilitate a latent EBV infection through its mediated degradation of pRb in copositive epithelia.

Particle size as a driver of dewatering performance and its relationship to stabilization in fecal sludge.

Poor and unpredictable dewatering performance of fecal sludge is a major barrier to sanitation provision in urban areas not served by sewers. Fecal sludge comprises everything that accumulates in onsite containments, and its characteristics are distinct from wastewater sludges and from feces. There is little fundamental understanding of what causes poor dewatering in fecal sludge. For the first time, we demonstrate that particle size distribution is a driver of dewatering performance in fecal sludge, and is associated with level of stabilization. Higher concentrations of small particles (<10 µm) and smaller median aggregate size (D50) corresponded to poor dewatering performance (measured by capillary suction time (CST) and supernatant turbidity) in field samples from Kenya and Uganda and in controlled laboratory anaerobic storage experiments. More stabilized fecal sludge (higher C/N, lower VSS/TSS) had better dewatering performance, corresponding to lower concentrations of small particles. Samples with the largest aggregates (D50 > 90 µm) had higher abundance of Gammaproteobacteria Pseudomonas, and samples with the smallest aggregates (D50 ≤ 50 µm) were characterized by higher abundance of Bacteroidetes Vadin HA17 and Rikenellaceae. Contrary to common perceptions, stabilization, particle size distribution, and dewatering performance were not dependent on time intervals between emptying of onsite containments or on time in controlled anaerobic storage experiments. Our results suggest that the stabilization process in onsite containments, and hence the dewaterability of sludge arriving at treatment facilities, is not dependent on time in containment but is more likely associated with specific microbial populations and the in-situ environmental conditions which promote or discourage their growth.

EBV Association with Lymphomas and Carcinomas in the Oral Compartment.

Epstein-Barr virus (EBV) is an oncogenic human herpesvirus infecting approximately 90% of the world's population. The oral cavity serves a central role in the life cycle, transmission, and pathogenesis of EBV. Transmitted to a new host via saliva, EBV circulates between cellular compartments within oral lymphoid tissues. Epithelial cells primarily support productive viral replication, while B lymphocytes support viral latency and reactivation. EBV infections are typically asymptomatic and benign; however, the latent virus is associated with multiple lymphomas and carcinomas arising in the oral cavity. EBV association with cancer is complex as histologically similar cancers often test negative for the virus. However, the presence of EBV is associated with distinct features in certain cancers. The intrinsic ability of EBV to immortalize B-lymphocytes, via manipulation of survival and growth signaling, further implicates the virus as an oncogenic cofactor. A distinct mutational profile and burden have been observed in EBV-positive compared to EBV-negative tumors, suggesting that viral infection can drive alternative pathways that converge on oncogenesis. Taken together, EBV is also an important prognostic biomarker that can direct alternative therapeutic approaches. Here, we discuss the prevalence of EBV in oral malignancies and the EBV-dependent mechanisms associated with tumorigenesis.

Travellers and influenza: risks and prevention.

BACKGROUND: Influenza viruses are among the major causes of serious human respiratory tract infection worldwide. In line with the high disease burden attributable to influenza, these viruses play an important, but often neglected, role in travel medicine. Guidelines and recommendations regarding prevention and management of influenza in travellers are scarce. Of special interest for travel medicine are risk populations and also circumstances that facilitate influenza virus transmission and spread, like travel by airplane or cruise ship and mass gatherings. METHODS: We conducted a PUBMED/MEDLINE search for a combination of the MeSH terms Influenza virus, travel, mass gathering, large scale events and cruise ship. In addition we gathered guidelines and recommendations from selected countries and regarding influenza prevention and management in travellers. By reviewing these search results in the light of published knowledge in the fields of influenza prevention and management, we present best practice advice for the prevention and management of influenza in travel medicine. RESULTS: Seasonal influenza is among the most prevalent infectious diseases in travellers. Known host-associated risk factors include extremes of age and being immune-compromised, while the most relevant environmental factors are associated with holiday cruises and mass gatherings. CONCLUSIONS: Pre-travel advice should address influenza and its prevention for travellers, whenever appropriate on the basis of the epidemiological situation concerned. Preventative measures should be strongly recommended for travellers at high-risk for developing complications. In addition, seasonal influenza vaccination should be considered for any traveller wishing to reduce the risk of incapacitation, particularly cruise ship crew and passengers, as well as those participating in mass gatherings. Besides advice concerning preventive measures and vaccination, advice on the use of antivirals may be considered for some travellers.

Plant-derived H7 VLP vaccine elicits protective immune response against H7N9 influenza virus in mice and ferrets.

In March 2013, the Chinese Centre for Disease Control and Prevention confirmed the first reported case of human infection with an avian influenza A H7N9 virus. Infection with this virus often caused severe pneumonia and acute respiratory distress syndrome resulting in a case fatality rate >35%. The risk of pandemic highlighted, once again, the need for a more rapid and scalable vaccine response capability. Here, we describe the rapid (19 days) development of a plant-derived VLP vaccine based on the hemagglutinin sequence of influenza H7N9 A/Hangzhou/1/2013. The immunogenicity of the H7 VLP vaccine was assessed in mice and ferrets after one or two intramuscular dose(s) with and without adjuvant (alum or GLA-SE™). In ferrets, we also measured H7-specific cell-mediated immunity. The mice and ferrets were then challenged with H7N9 A/Anhui/1/2013 influenza virus. A single immunization with the adjuvanted vaccine elicited a strong humoral response and protected mice against an otherwise lethal challenge. Two doses of unadjuvanted vaccine significantly increased humoral response and resulted in 100% protection with significant reduction of clinical signs leading to nearly asymptomatic infections. In ferrets, a single immunization with the alum-adjuvanted H7 VLP vaccine induced strong humoral and CMI responses with antigen-specific activation of CD3(+) T cells. Compared to animals injected with placebo, ferrets vaccinated with alum-adjuvanted vaccine displayed no weight loss during the challenge. Moreover, the vaccination significantly reduced the viral load in lungs and nasal washes 3 days after the infection. This candidate plant-made H7 vaccine therefore induced protective responses after either one adjuvanted or two unadjuvanted doses. Studies are currently ongoing to better characterize the immune response elicited by the plant-derived VLP vaccines. Regardless, these data are very promising for the rapid production of an immunogenic and protective vaccine against this potentially pandemic virus.

Travel-acquired infections in Canada: CanTravNet 2011-2012.

BACKGROUND: Important gaps remain in our knowledge of the infectious diseases people acquire while travelling and the impact of pathogens imported by Canadian travellers. OBJECTIVE: To provide a surveillance update of illness in a cohort of returned Canadian travellers and new immigrants. METHODS: Data on returning Canadian travellers and new immigrants presenting to a CanTravNet site between September 2011 and September 2012 were extracted and analyzed by destination, presenting symptoms, common and emerging infectious diseases and disease severity. RESULTS: During the study period, 2283 travellers and immigrants presented to a CanTravNet site, 88% (N=2004) of whom were assigned a travel-related diagnosis. Top three destinations for non-immigrant travellers were India (N=132), Mexico (N=103) and Cuba (N=89). Fifty-one cases of malaria were imported by ill returned travellers during the study period, 60% (N=30) of which were Plasmodium falciparum infections. Individuals travelling to visit friends and relatives accounted for 83% of enteric fever cases (15/18) and 41% of malaria cases (21/51). The requirement for inpatient management was over-represented among those with malaria compared to those without malaria (25% versus 2.8%; p<0.0001) and those travelling to visit friends and relatives versus those travelling for other reasons (12.1% versus 2.4%; p<0.0001). Nine new cases of HIV were diagnosed among the cohort, as well as one case of acute hepatitis B. Emerging infections among travellers included hepatitis E virus (N=6), chikungunya fever (N=4) and cutaneous leishmaniasis (N=16). Common chief complaints included gastrointestinal (N=804), dermatologic (N=440) and fever (N=287). Common specific causes of chief complaint of fever in the cohort were malaria (N=47/51 total cases), dengue fever (14/18 total cases), enteric fever (14/17 total cases) and influenza and influenza-like illness (15/21 total cases). Animal bites were the tenth most common diagnosis among tourist travellers. INTERPRETATION: Our analysis of surveillance data on ill returned Canadian travellers provides a recent update to the spectrum of imported illness among travelling Canadians. Preventable travel-acquired illnesses and injuries in the cohort include malaria, enteric fever, HIV, hepatitis B, hepatitis A, influenza and animal bites. Strategies to improve uptake of preventive interventions such as malaria chemoprophylaxis, immunizations and arthropod/animal avoidance may be warranted.

Seroprevalence of seven zoonotic infections in Nunavik, Quebec (Canada).

In Nunavik, common practices and food habits such as consumption of raw meat and untreated water place the Inuit at risk for contracting zoonotic diseases. The aim of this study was to determine the seroprevalence of seven zoonotic infections among the permanent residents of Nunavik. The study was conducted in the fall 2004 as part of the Nunavik Health Survey. Blood samples from adults aged 18-74 years (n = 917) were collected and analysed for the presence of antibodies against Trichinella spp., Toxocara canis, Echinococcus granulosus, Brucella spp., Coxiella burnetii, Leptospira spp. and Francisella tularensis. Information on sociodemographic characteristics, traditional activities, drinking water supply and nutrition was gathered using english/inuktitut bilingual questionnaires. The chi-squared test was used to evaluate associations between seropositivity and other measured variables. Statistically significant variables were included in a multivariate logistic regression model to control for confounding factors. Estimated seroprevalences were 8.3% for E. granulosus, 3.9% for T. canis, 5.9% for Leptospira spp. and 18.9% for F. tularensis. Seroprevalence was ≤ 1% for Trichinella spiralis, Brucella spp. and C. burnetii. For most infections, seropositivity tended to increase with age. In multivariate analyses, seroprevalence was positively (i.e. directly) associated with age and residence in the Ungava coast area for F. tularensis; age and residence in the Hudson coast area for T. canis; female gender, lower level of schooling and frequent cleaning of water reservoirs for E. granulosus. No risk factor for Leptospira spp. infection was identified. No associations were detected with regards to food habits or environmental exposures. A small but significant portion of the Nunavik population has serologic evidence of exposure to at least one of the pathogenic microorganisms investigated. Further studies are needed to better understand the mechanisms for transmission of zoonotic infections and their potential reservoirs in Nunavik.

Smart sensor systems for human health breath monitoring applications.

Breath analysis techniques offer a potential revolution in health care diagnostics, especially if these techniques can be brought into standard use in the clinic and at home. The advent of microsensors combined with smart sensor system technology enables a new generation of sensor systems with significantly enhanced capabilities and minimal size, weight and power consumption. This paper discusses the microsensor/smart sensor system approach and provides a summary of efforts to migrate this technology into human health breath monitoring applications. First, the basic capability of this approach to measure exhaled breath associated with exercise physiology is demonstrated. Building from this foundation, the development of a system for a portable asthma home health care system is described. A solid-state nitric oxide (NO) sensor for asthma monitoring has been identified, and efforts are underway to miniaturize this NO sensor technology and integrate it into a smart sensor system. It is concluded that base platform microsensor technology combined with smart sensor systems can address the needs of a range of breath monitoring applications and enable new capabilities for healthcare.

Spatio-temporal variations and age effect on Toxoplasma gondii seroprevalence in seals from the Canadian Arctic.

Toxoplasmosis is a significant public health threat for Inuit in the Canadian Arctic. This study aimed to investigate arctic seals as a possible food-borne source of infection. Blood samples collected from 828 seals in 7 Canadian Arctic communities from 1999 to 2006 were tested for Toxoplasma gondii antibodies using a direct agglutination test. Polymerase chain reaction (PCR) was used to detect T. gondii DNA in tissues of a subsample of seals. Associations between seal age, sex, species, diet, community and year of capture, and serological test results were investigated by logistic regression. Overall seroprevalence was 10·4% (86/828). All tissues tested were negative by PCR. In ringed seals, seroprevalence was significantly higher in juveniles than in adults (odds ratio=2·44). Overall, seroprevalence varied amongst communities (P=0·0119) and by capture year (P=0·0001). Our study supports the hypothesis that consumption of raw seal meat is a significant source of infection for Inuit. This work raises many questions about the mechanism of transfer of this terrestrial parasite to the marine environment, the preponderance of infection in younger animals and the natural course of infection in seals. Further studies to address these questions are essential to fully understand the health risks for Inuit communities.

A nasally administered trivalent inactivated influenza vaccine is well tolerated, stimulates both mucosal and systemic immunity, and potentially protects against influenza illness.

A randomized placebo-controlled double-blind trial of a nasally administered inactivated trivalent influenza vaccine formulated with partially purified meningococcal outer membrane proteins (OMP-TIV) was conducted in 1349 healthy adults aged 18-64 years. Subjects received either vaccine containing 15 µg of haemagglutinin (HA) of each of three influenza strains for the 2003-2004 season on days 0 and 14, or 30 µg on day 0 and saline placebo on day 14, or placebo on days 0 and 14. Vaccination was well tolerated, with similar reactogenicity as placebo. Compared to placebo, statistically significant increases in mean serum haemagglutinin inhibition reciprocal titers and salivary secretory IgA to all 3 antigens were seen on day 28 for both vaccine dose groups. The incidence of culture-positive influenza and fever >37.8°C and cough and one or more of sore throat, runny nose or nasal congestion, muscle or joint ache, headache, fatigue, or chills or culture positive influenza and at least two of these symptoms was low (16/1349; 1.2%). In the intent-to-immunize population too few febrile culture-confirmed illness events (n=4) occurred to perform analysis. Fever occurred infrequently, even in the presence of positive cultures and disabling multi-symptom disease. In participants receiving all doses of either vaccine regimen the incidence of culture-confirmed influenza with respiratory symptoms and with or without fever was 0.77% (7/904) vs. 2.03% (9/443) in placebo recipients (p=0.045, Fisher's exact test; relative risk reduction 62%), despite circulation of a drift variant A/H3N2 that was poorly matched to vaccine. An OMP-TIV vaccine was well tolerated and reduced risk of symptomatic culture confirmed influenza. Vaccine efficacy will need to be validated in a season with a higher attack rate.

Seroprevalence of Toxoplasma gondii among Nunavik Inuit (Canada).

As a result of their intimate contact with the land and their nutritional habits, the Inuit of Nunavik are considered to be at risk from zoonotic infections. To better understand the risk factors for Toxoplasma gondii infection, a serosurvey was conducted in Nunavik, Québec, in September 2004. A representative sample of the Inuit adult population of Nunavik participated in this cross-sectional study (n = 917). Antibodies (IgG) against T. gondii were detected by immunoassay. Information on sociodemographic characteristics, traditional activities, domestic environment and nutrition was gathered by questionnaire and explored as variables explanatory of seropositive results. Associations found to be statistically significant in univariate analyses were assessed by multivariable logistic regression to control for confounding factors. Almost two thirds (59.8%) of the Inuit of Nunavik were found to be seropositive for T. gondii. In multivariate analyses, risk factors for seropositivity were: increasing age, gender (women > men), lower level of education, consumption of potentially contaminated water (determined by an index of risk from waterborne infections), frequent cleaning of water reservoirs, and consumption of seal meat and feathered game. There was some variation in seroprevalence between the Ungava Bay coast (52.3%) and the Hudson Bay coast (65.6%), the two main regions of Nunavik, but this variation was not significant in the multivariable logistic regression model. This cross-sectional study demonstrated high T. gondii seroprevalence in the Inuit population and revealed that age, gender, schooling and community of residence all influence serostatus in this population. Variables related to drinking water and food choices may also influence the risk of infection. These results raise important questions about T. gondii transmission in Nunavik including possible links between terrestrial and marine cycles.

High-yield amplification of Cryptosporidium parvum in interferon gamma receptor knockout mice.

To date, large-scale production of Cryptosporidium parvum oocysts has only been achieved by amplification in neonatal calves and sheep. Many laboratories currently depend on supplies from external sources and store oocysts for prolonged periods which results in progressive loss of viability. Six to 8-week-old interferon gamma receptor knockout (IFN gamma R-KO) mice on a C57BL/6 background were inoculated by gavage (2000 oocysts/animal). Fecal pellets were collected daily from 7 days post-infection (p.i.) up to 2 weeks p.i. Intestinal oocyst yield was assessed at days 11, 12 and 14 p.i. by homogenization of intestinal tissues. Ether extraction and one or more NaCl flotations were used to purify oocysts. Total recoveries averaged 2.6 x 10(6) oocysts/mouse from fecal material and 3.8 x 10(7) oocysts/mouse from intestinal tissues. Overall, 2.3 x 10(9) purified oocysts were obtained from 60 mice. Recovered oocysts were capable of sporulation and were shown to be infectious both in vitro and in vivo. Oocyst amplification was achieved in only 11-14 days with minimal expense. The simplicity of this method presents a practical alternative for the routine passage, maintenance and storage of C. parvum in biomedical laboratories.

Heterologous prime-boost strategy to immunize very young infants against measles: pre-clinical studies in rhesus macaques.

Infants in developing countries are at high risk of developing severe clinical measles if they become infected during the "window of vulnerability" (age 4-9 months), when declining maternal antibodies do not protect against wild virus, yet impede successful immunization by attenuated measles vaccine. We developed two Sindbis replicon-based DNA vaccines expressing measles virus hemagglutinin and fusion protein with the goal of priming young infants to respond safely and effectively to subsequent boosting with attenuated measles vaccine. Intradermal prime with DNA vaccines by needle-free injection followed by aerosol or parenteral boost with licensed measles vaccine was well tolerated by juvenile and young infant rhesus macaques, and protected against clinical measles and viremia on wild-type virus challenge. A proteosome-measles vaccine administered alone (three doses) or as a boost following DNA vaccine priming was also safe and protective. These promising results pave the way for clinical trials to assess this prime-boost strategy.

HIV-1 and HIV-2 prevalence and associated risk factors among postnatal women in Harare, Zimbabwe.

Studies of antenatal women form the predominant source of data on HIV-1 prevalence in Africa. Identifying factors associated with prevalent HIV is important in targeting diagnostic services and care. Between November 1997 and January 2000, 14,110 postnatal women from Harare, Zimbabwe were tested by ELISAs reactive to both HIV-1 and HIV-2; a subset of positive samples was confirmed with assays specific for HIV-1 and HIV-2. Baseline characteristics were elicited and modelled to identify risk factors for prevalent HIV infection. HIV-1 and HIV-2 prevalences were 32.0% (95% CI 31.2-32.8) and 1.3% (95% CI 1.1-1.5), respectively; 4% of HIV-1-positive and 99% of HIV-2-positive women were co-infected. HIV-1 prevalence increased from 0% among 14-year-olds to >45% among women aged 29-31 years, then fell to <20% among those aged>40 years. In multivariate analyses, prevalence increased with parity, was lower in married women than in single women, divorcees and widows, and higher in women with the lowest incomes and those professing no religion. Adjusted HIV-1 prevalence increased during 1998 and decreased during 1999. Age modified the effects of parity, home ownership and parental education. Among older women, prevalence was greater for women who were not homeowners. Among younger women, prevalence increased with parity and low parental education. None of these factors distinguished women co-infected with HIV-2 from those infected with HIV-1 alone. Prevalent HIV-1 infection is associated with financial insecurity and weak psychosocial support. The ZVITAMBO study apparently spanned the peak of the HIV-1 epidemic among reproductive women in Harare.

Field evaluation of four novel enzyme immunoassays for Chagas' disease in Venezuela blood banks: comparison of assays using fixed-epimastigotes, fixed-trypomastigotes or trypomastigote excreted-secreted antigens from two Trypanosoma cruzi strains.

Many serological tests have been developed for the diagnosis of Chagas' disease, but few have been subjected to a rigorous field evaluation. We have recently described several novel enzyme immunoassays (EIAs) based on fixed-whole organisms or trypomastigote excretory-secretory antigens (TESA) from different Trypanosoma cruzi strains (Tulahuen or Brazil). This study evaluated the most promising of these novel assays (e.g. fixed-epimastigotes, fixed-trypomastigotes, TESA Brazil and TESA Tulahuen antigens) in a field study of Venezuelan blood bank specimens. The assays were tested in an operator-blinded fashion using 2038 blood bank samples obtained from low and high T.cruzi prevalence regions of Venezuela (n= 1050 and n= 988 from Bolivar and Portuguesa states, respectively). Based on National Laboratory for Chagas Immunodiagnosis (NLCI) 'gold standard' results, all novel EIAs were superior to the commercial kit currently used in Venezuela, achieving 100% sensitivity and >99% specificity at optimal cut-off values. The novel assays identified seven false-negative samples compared with the routine screening performed by the Venezuelan blood bank although two samples were also misclassified as positive. Minor differences in the performance of the four novel assays were observed at lower arbitrary cut-off values. This study confirms the potential utility of both the fixed-organism and the TESA-based assays in the diagnosis of T.cruzi infection.

Randomized, double-blind clinical trial of topical imiquimod 5% with parenteral meglumine antimoniate in the treatment of cutaneous leishmaniasis in Peru.

BACKGROUND: Current treatments for cutaneous leishmaniasis are limited by their toxicity, high cost, and discomfort and the emergence of drug resistance. New approaches, including combination therapies, are urgently needed. We performed a double-blind, randomized trial of therapy with parenteral antimony plus topical imiquimod, an innate immune-response modulator, versus therapy with antimony alone, in subjects with cutaneous leishmaniasis for whom an initial course of antimony therapy had failed. METHODS: Forty subjects with clinical resistance to antimony were recruited in Lima, Peru, between February 2001 and December 2002. All subjects received meglumine antimoniate (20 mg/kg/day im or iv) and were randomized to receive either topical imiquimod 5% cream (Aldara; 3M Pharmaceuticals) or vehicle control every other day for 20 days. Lesions and adverse events were evaluated during treatment and at 1, 2, 3, 6, and 12 months after the treatment period. RESULTS: The mean number of lesions was 1.2 per person; 71% of the lesions were facial and 76% were ulcerative. There were no major differences between the groups, and all but 2 subjects completed therapy. Mild adverse events were reported by 73% of the subjects, but only erythema occurred more commonly in the imiquimod group (P < or = .02). Lesions resolved more rapidly in the imiquimod group: 50% of the imiquimod group achieved cure at 1 month after the treatment period versus 15% of the vehicle cream group (P < or = .02); 61% of the imiquimod group at 2 months versus 25% of the vehicle cream group (P < or = .03); and 72% of the imiquimod group at 3 months versus 35% of the vehicle cream group (P < or = .02). Residual scarring in the imiquimod group was less prominent than in the vehicle cream group. CONCLUSIONS: Combined antimony plus imiquimod treatment was well tolerated, accelerated healing of lesions, and improved scar quality. This therapy may have particular advantages for subjects with facial lesions.

Measles vaccination in the presence of maternal antibodies primes for a balanced humoral and cellular response to revaccination.

Early or low dose antigen exposure can prime the immune system for subsequent responses; the so-called "prime-boost" effect. In the context of a Sudanese measles vaccine trial, we assessed whether or not such early exposure could influence the response to revaccination. Children received either Connaught high titer vaccine (CN: n = 53; 10(4.7)pfu) or meningococcal A + C vaccine as a placebo (MEN: n = 58) at 5 months of age. At 9 months of age, all received standard titer Schwarz vaccine (SCH: 10(3.9)pfu). Neutralizing antibodies were measured before initial vaccination and at 9 months of age (plaque reduction neutralization assay (PRN)) and again at 5 years of age (syncytium inhibition assay (SIA)). Lymphoproliferative responses to measles virus (MV) antigens were evaluated at 5 years of age. Eleven of the 53 CN-SCH children (21%) had sub-protective neutralizing antibody titers prior to revaccination (log PRN 1.5 +/- 0.03 versus 2.9 +/- 0.07 in the remaining 42 children; P < 0.004). Maternal antibody titers at the time of initial vaccination in these 11 were high (PRN 2.44 +/- 0.12 versus 1.9 +/- 0.04; P < 0.0001). At 5 years of age, neutralizing antibodies were comparable in the 11 CN-SCH poor responders (log SIA 2.1 +/- 0.09), the remaining CN-SCH children (2.2 +/- 0.06) and the MEN-SCH group vaccinated only once at 9 months of age (2.25 +/- 0.06). In contrast, 7/11 of the CN-SCH poor responders (64%) had stimulation indices (SI) > 3 in response to MV antigens at 5 years of age (SI 3.1 +/- 0.6) compared with only 14% in the remaining children of the CN-SCH group (2.0 +/- 0.3; P = 0.05) and 8% in the MEN-SCH group (1.4 +/- 0.2; P < 0.0003). These data suggest that early measles vaccination in the presence of maternal antibodies can sometimes prime for a balanced humoral and cellular immune response to subsequent revaccination.

Immune response to measles vaccine in Peruvian children.

OBJECTIVE: To evaluate the immune response in Peruvian children following measles vaccination. METHODS: Fifty-five Peruvian children received Schwarz measles vaccine (about 10(3) plaque forming units) at about 9 months of age. Blood samples were taken before vaccination, then twice after vaccination: one sample at between 1 and 4 weeks after vaccination and the final sample 3 months post vaccination for evaluation of immune cell phenotype and lymphoproliferative responses to measles and non-measles antigens. Measles-specific antibodies were measured by plaque reduction neutralization. FINDINGS: The humoral response developed rapidly after vaccination; only 4 of the 55 children (7%) had plaque reduction neutralization titres <200 mlU/ml 3 months after vaccination. However, only 8 out of 35 children tested (23%) had lymphoproliferative responses to measles antigens 3-4 weeks after vaccination. Children with poor lymphoproliferative responses to measles antigens had readily detectable lymphoproliferative responses to other antigens. Flow cytometric analysis of peripheral blood mononuclear cells revealed diffuse immune system activation at the time of vaccination in most children. The capacity to mount a lymphoproliferative response to measles antigens was associated with expression of CD45RO on CD4+ T-cells. CONCLUSION: The 55 Peruvian children had excellent antibody responses after measles vaccination, but only 23% (8 out of 35) generated detectable lymphoproliferative responses to measles antigens (compared with 55-67% in children in the industrialized world). This difference may contribute to the less than uniform success of measles vaccination programmes in the developing world.

Increasing referral of at-risk travelers to travel health clinics: evaluation of a health promotion intervention targeted to travel agents.

BACKGROUND: Increases in travel-related illness require new partnerships to ensure travelers are prepared for health risks abroad. The travel agent is one such partner and efforts to encourage travel agents to refer at-risk travelers to travel health clinics may help in reducing travel-attributable morbidity. METHODS: A health promotion intervention encouraging travel agents to refer at-risk travelers to travel health clinics was evaluated. Information on the knowledge, attitudes, and behaviors of travel agents before and after the intervention was compared using two self-administered questionnaires. The Wilcoxon signed rank test was used to compare the mean difference in overall scores to evaluate the overall impact of the intervention and also subscores for each of the behavioral construct groupings (attitudes, barriers, intent, and subjective norms). Multiple regression techniques were used to evaluate which travel agent characteristics were independently associated with a stronger effect of the intervention. RESULTS: A small improvement in travel agents overall attitudes and beliefs (p =.03) was found, in particular their intention to refer (p =.01). Sixty-five percent of travel agents self-reported an increase in referral behavior; owners or managers of the agency were significantly more likely to do so than other travel agents (OR = 7.25; 95% CI: 1.64 32.06). Older travel agents, those that worked longer hours and those with some past referral experience, had significantly higher post-intervention scores. CONCLUSIONS: Travel agents can be willing partners in referral, and agencies should be encouraged to develop specific referral policies. Future research may be directed toward investigating the role of health education in certification curricula, the effectiveness of different types of health promotion interventions, including Internet-facilitated interventions, and the direct impact that such interventions would have on travelers attending travel health clinics.