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Please cite this paper as: Jeremy Sueker et al. (2010) Influenza and respiratory disease surveillance: the US military’s global laboratory‐based network. Influenza and Other Respiratory Viruses 4(3), 155–161. The US Department of Defense influenza surveillance system now spans nearly 500 sites in 75 countries, including active duty US military and dependent populations as well as host‐country civilian and military personnel. This system represents a major part of the US Government’s contributions to the World Health Organization’s Global Influenza Surveillance Network and addresses Presidential Directive NSTC‐7 to expand global surveillance, training, research and response to emerging infectious disease threats. Since 2006, the system has expanded significantly in response to rising pandemic influenza concerns. The expanded system has played a critical role in the detection and monitoring of ongoing H5N1 outbreaks worldwide as well as in the initial detection of, and response to, the current (H1N1) 2009 influenza pandemic. This article describes the system, details its contributions and the critical gaps that it is filling, and discusses future plans.

In 2001, we began a prospective longitudinal study in a cohort of schoolchildren 5-13 years of age residing in Maracay, Venezuela, to determine the cumulative incidence of dengue virus (DENV) infections by virus serotype. This report presents serological data from 710 schoolchildren who were tested during the first 2 years of the study. Serological evaluations were conducted by plaque reduction neutralization test (PRNT). At study initiation, 51% of children had PRNT antibody titers against one (30.1% = 13.4% DENV-1, 14.2% DENV-2, 0.6% DENV-3, and 2% DENV-4) or multiple DENV serotypes (20.9%). By the end of the first year, 89 of 348 (25.6%) PRNT-negative children seroconverted, and 94 of 362 (26%) who were PRNT-positive in their baseline sera tested positive for additional serotypes, for an overall cumulative incidence of DENV infections of 25.8%. By serotype, the percentages found were 1.4% DENV-1, 1.4% DENV-2, 19% DENV-3, and 1.2% DENV-4. In the second year, 37 of 259 (14.3%) PRNT-negative children seroconverted, and 83 of 451 (18.4%) who had monotypic and multitypic PRNT patterns in their baseline sera exhibited additional serotype seroconversions, for an overall cumulative incidence of DENV infections of 16.9%. By serotype, the percentages found were 0.8% DENV-1, 1.5% DENV-2, 8.5% DENV-3, and 2.3% DENV-4. Overall, these results suggest a high cumulative incidence of DENV infections among 5-13-year-old school children in Maracay, Venezuela.

More than 35 years ago, population-based studies of viral respiratory illnesses among US families were conducted in Ohio [1], Kansas [2], Louisiana [3], New York [4], and Washington [5]. As a result of these investigations, scientists concluded that adenovirus infection was quite common among children. Approximately 50% of infections were asymptomatic, and symptomatic infections were typically mild and resolved without sequelae. In contrast, military populations experienced severe epidemics of acute respiratory disease, including pneumonia and encephalitis, especially involving adenovirus types 4, 7, and 21. For example, in 1958, adenoviral infection was reported to have caused hospitalization of an estimated 10% of military recruits [6] and to be the etiology of most respiratory disease during winter months. Subsequently, vaccines for adenovirus types 4 and 7 were developed and effectively used among US military trainees from the 1970s until the late 1990s [7, 8]. Thus, until recently, adenovirus infection was considered to have little consequence, except for causing morbidity among military trainees. However, much has changed since these early epidemiological studies were conducted. In contrast to the modest number of adenovirus types recognized 35 years ago, 51 unique serotypes are now recognized. Different serotypes have been found to have different tissue tropisms that correlate with different clinical manifestations of infection. Limited epidemiological investigations have revealed that, among some specific serotypes, multiple genetic variants exist that often have quite different geographical distributions and associated virulence [9-11]. In addition, largely because of molecular diagnostics, adenovirus infection has been associated with a number of acute and chronic diseases, including chronic airway obstruction [12] and pulmonary dysplasia [13], myocarditis and dilated cardiomyopathy [14], mononucleosis-like syndromes [15], intussusception [16], sudden infant perinatal death [17], and obesity [18, 19]. Among some population groups, adenoviral infection is common and, often, severe. For example, the incidence of adenoviral disease among bone marrow transplant recipients varies from 3% to 20% [20], and mortality can exceed 50% [21]. Similarly, multiple recent outbreaks of adenoviral infection in the United States have frequently occurred among institutionalized children and in medical settings, resulting in significant morbidity and mortality among patients and medical staff [22]. In vitro studies suggest that specific adenovirus types are more likely to respond to certain antiviral therapies [23]. Finally, since the military lost its manufacturer of adenovirus types 4 and 7 vaccines in the late 1990s, numerous outbreaks of adenovirus infection have occurred among military trainees, resulting in great morbidity [24, 25]. Subsequently, the US Department of Defense identified a new vaccine manufacturer, and restoration of adenovirus types 4 and 7 vaccines is projected for 2008 [25]. It seems prudent to investigate whether previously very effective and safe vaccines may also benefit some nonmilitary populations. Therefore, an updated look at the epidemiology of human adenovirus infection is warranted. In this report, we present 25 months of viral and patient epidemiological data on clinical adenovirus infection detected through a nationwide network of 22 US military and civilian medical facilities. We used a recently described molecular adenovirus typing technique to characterize the strains of adenovirus [26].

Methicillin-resistant Staphylococcus aureus is now a common isolate of community-acquired staphylococcal infections. We present the first case of concomitant mycotic pseudoaneurysm and purulent pericarditis caused by methicillin-resistant S. aureus. The isolate was found to be SCCmec type I, sequence type 8, and to carry the PVL gene. The patient was successfully treated with a combined surgical and medical approach.

Sexual transmission represents the principal mode of transmission for human immunodeficiency virus type 1 (HIV-1) worldwide. We examined the HIV-1 seroprevalence and risk factors for infection among 613 female commercial sex workers (FCSW) in Isla Margarita, Venezuela. Recruitment was conducted in street venues and working locations. None of the FCSW tested positive for HIV; this correlated with the low self-reported rates of sexually transmitted infections (6%), drug use (20%), and alcohol abuse (12%). Condom use and safe-sex practices were found to be practiced regularly (80% of time) with clients; however, such practices were found to be very uncommon in nonclient relations (20% of the time). Understanding the sexual risk behaviors, beliefs, and drug use patterns of FCSW is important for future development of effective public prevention policies and educational campaigns aimed at decreasing the risk of infection with HIV-1 and other sexually transmitted infections among FCSW.

Objectives: To determine the antimicrobial susceptibility and serotype distribution of 205 isolates of Streptococcus pneumoniae, collected from the CSF of meningitis patients identified between 1998‐2003, during sentinel meningitis surveillance in Egypt. Methods: Antimicrobial susceptibility was evaluated against six antibiotics using disc diffusion and Etest methods. Serotyping was performed by latex agglutination and the Quellung test. Results: Forty-nine percent of all isolates were found to be non-susceptible to penicillin (46% intermediate, MIC range 0.12‐1.0mg/L; 3% resistant, MIC 5 2.0mg/L), and 6% of the isolates were nonsusceptible to ceftriaxone (5% intermediate, MIC 5 1.0mg/L; 1.3% resistant, MIC>_ 2mg/L). Resistance rates for tetracycline and trimethoprim/sulfamethoxazole were high (52 and 59.7%, respectively), but those for erythromycin and chloramphenicol were lower (11 and 9%, respectively). Five serotypes (6B, 1, 19A, 23F and 6A) accounted for 37% of the total isolates. Ten isolates (5%) were non-typeable. Overall, 29 and 42% of serotypes were represented in the 7- and 11-valent conjugate vaccines, respectively. However, vaccine coverage for children

Background. Although group A streptococci (GAS) infections are a major cause of morbidity and mortality, outbreaks of associated pneumonia are rare. We report an outbreak of GAS pneumonia that occurred at a US military training camp. Methods. Standard epidemiologic and laboratory procedures were used to characterize the outbreak and causative organism(s). A case-control study and determination of the prevalence of GAS infection among camp personnel were also performed. Results. A total of 162 of 4500 Marine Corps personnel were hospitalized for respiratory symptoms during the period of 1 November and 20 December 2002, and 127 (78%) had radiographically confirmed pneumonia. The attack rate was 1.6 cases per 100 person-months. Thirty-four (27%) of 127 patients with pneumonitis had definite or probable GAS pneumonia; an additional 22 (17.3%) were coinfected with GAS and another pathogen. Pathogens, in addition to GAS, included Chlamydia pneumoniae (27 patients), Mycoplasma pneumoniae (19), adenovirus (5), and Streptococcus pneumoniae (2). A survey revealed that the pharyngeal carriage rate of GAS among camp personnel was 16%. Molecular characterization of the GAS isolates found emm type 3, multilocus sequence type 15. The epidemic ended after administration of additional prophylaxis with a single dose of intramuscular benzathine penicillin (1.2 million U) or azithromycin (1 g orally). Because the number of days from the last penicillin injection was correlated with a positive throat culture result and the occurrence of pneumonia, the dosing interval of benzathine penicillin was shortened from every 28-35 days to every 21 days. Conclusions. This is the largest outbreak of GAS pneumonia reported in >30 years. This outbreak emphasizes the potential for GAS to cause epidemics of severe infection and demonstrates the need for surveillance and consideration of appropriate antibiotic prophylaxis among particularly high-risk populations.

We present 3 cases of Neisseria meningitidis with varied clinical presentations, including meningococcemia, meningitis, and pneumonia. Our cases suggest an association between concomitant bacterial and viral infections, including group A streptococcus and adenovirus, and an increased risk of invasive Neisseria infections. PCR data are a useful adjunct to bacterial cultures for the diagnosis. Our case series exemplify the continued high mortality of N. meningitides and highlights the need for more immunogenic vaccines for prevention.

Infections with hepatitis C virus, (HCV), hepatitis B virus (HBV), and human T lymphotropic type I/II (HTLV-I/II) virus are commonly found in patients infected with human immunodeficiency virus type 1 (HIV-1). We conducted a seroepidemiologic study among 174 HIV-positive heterosexuals in Buenos Aires, Argentina in 1999. Evidence of exposure to HCV, HBV, and HTLV-I/II was found in 32%, 17%, and 5%, respectively. A higher prevalence of HBV infection was observed among males (33%) compared with females (12%; P < 0.05). Among women, a prior history of a sexually transmitted infection, injecting drug use (IDU), having had more than five lifetime sex partners, and having exchanged sex-for-goods were significantly associated with HCV infection, whereas an IDU history, syringe sharing, and having exchanged sex-for-goods were found to be associated with HBV infection. Among men, an IDU history and syringe/needle sharing were significantly associated with HCV infection. The IDU-related and sexual transmission of hepatitis viruses constitute a significant problem among young, HIV-infected, heterosexuals in Argentina.

Since Venezuelan equine encephalitis virus (VEEV) was isolated in Peru in 1942, >70 isolates have been obtained from mosquitoes, humans, and sylvatic mammals primarily in the Amazon region. To investigate genetic relationships among the Peru VEEV isolates and between the Peru isolates and other VEEV strains, a fragment of the PE2 gene was amplified and analyzed by single-stranded conformation polymorphism. Representatives of seven genotypes underwent sequencing and phylogenetic analysis. The results identified four VEE complex lineages that cocirculate in the Amazon region: subtypes ID (Panama and Colombia/Venezuela genotypes), IIIC, and a new, proposed subtype IIID, which was isolated from a febrile human, mosquitoes, and spiny rats. Both ID lineages and the IIID subtype are associated with febrile human illness. Most of the subtype ID isolates belonged to the Panama genotype, but the Colombia/Venezuela genotype, which is phylogenetically related to epizootic strains, also continues to circulate in the Amazon basin.

Prostitution may constitute a risk behavior for infection with hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus (HIV). We conducted a seroepidemiologic study among 200 male trans- vestite commercial sex workers (CSWs) in Montevideo, Uruguay in 1999. Evidence of exposure to HBV, HCV, and HIV was found in 101 (50.5%), 13 (6.5%), and 43 (21.5%) individuals, respectively. Positivity for HIV was correlated with both HBV (odds ratio (OR) 2.15, 95% confidence interval (CI) 1.01−4.67) and HCV (OR 3.47, 95% CI 0.90−12.79) infection. Predictors of infection were older age ( 26 years; P < 0.01) for all 3 viruses and time in CSW (5 years; P < 0.05) for HBV and HIV. Prior history of use of drugs (OR 3.54, 95% CI 1.09−11.52) and sexual contact with foreigners (OR 9.2, 95% CI 1.16−73.12) were found to be associated only with HCV infection. Sexual transmission of these viruses constitutes a significant problem among male transvestite CSWs.

Background Streptococcus pneumoniae infections have periodically caused significant morbidity and outbreaks among military personnel, especially trainees. This study evaluated the effectiveness of the 23-valent polysaccharide pneumococcal vaccine (PPV23) in reducing pneumonia in healthy military trainees. Methods From 2000–2003, 152 723 military trainees from 5 US training camps were enrolled in a double-blind, placebo-controlled trial of PPV23. Participants were closely monitored during basic training for radiographically confirmed pneumonia etiology and loss-of-training days. Participants were also followed using electronic medical encounter data until 1 June 2007 for three additional outcomes: any-cause pneumonia, any acute respiratory disease, and meningitis. Results Comparison of demographic data by study arm suggested the randomization procedures were sound. During basic training, 371 study participants developed radiographically confirmed pneumonia. None had evidence of S. pneumoniae infection, but other etiologies included adenovirus (38%), Chlamydophila pneumoniae (9%), and Mycoplasma pneumoniae (8%). During the follow-up period, many study participants, in both the vaccine and placebo groups, had clinical encounters for the medical outcomes of interest. However, Cox's proportional hazard modeling revealed no evidence of a protective vaccine effect during recruit training (radiographically confirmed pneumonia) or up to 6.7 years after enrollment (any-cause pneumonia, any acute respiratory disease, or meningitis). Conclusions Data from this large, double-blind, placebo controlled trial do not support routine use of PPV23 among healthy new military trainees. This clinical trial was registered at clinicaltrials.gov (registration number NCT02079701, http://www.clinicaltrials.gov/ct2/show/NCT02079701?term=NCT02079701&rank=1 ).

NIH grants AI 43336 and AI 10984 The University of Texas Medical Branch. Departments of Microbiology and Immunology. Galveston, Texas / The University of Texas Medical Branch. Departments of Microbiology and Pathology. Galveston, Texas. Minist?rio da Sa?de. Funda??o Nacional de Sa?de. Instituto Evandro Chagas. Bel?m, PA, Brasil. Minist?rio da Sa?de. Funda??o Nacional de Sa?de. Instituto Evandro Chagas. Bel?m, PA, Brasil. Minist?rio da Sa?de. Funda??o Nacional de Sa?de. Instituto Evandro Chagas. Bel?m, PA, Brasil. U.S. Naval Medical Research Center. Detachment, Lima, Peru. U.S. Naval Medical Research Center. Detachment, Lima, Peru. The University of Texas Medical Branch. Departments of Microbiology and Immunology. Galveston, Texas / The University of Texas Medical Branch. Departments of Microbiology and Pathology. Galveston, Texas / The University of Texas Medical Branch. Center for Tropical Diseases. Galveston, Texas. The University of Texas Medical Branch. Departments of Microbiology and Immunology. Galveston, Texas / The University of Texas Medical Branch. Departments of Microbiology and Pathology. Galveston, Texas / The University of Texas Medical Branch. Center for Tropical Diseases. Galveston, Texas. Oropouche (ORO) virus is an emerging infectious agent that has caused numerous outbreaks of an acute febrile (dengue-like) illness among humans in Brazil, Peru, and Panama. Diagnosis of ORO virus infection is based mainly on serology. Two different antigens, hamster serum antigen (HSA) and Vero cell lysate antigen (VCLA), are currently used in enzyme immunoassays (EIAs) in Brazil and Peru, respectively, to investigate the epidemiology of ORO virus infection. Both antigens involve use of infectious virus, and for this reason their use is restricted. Consequently, the frequency and distribution of ORO virus infection are largely unexplored in other countries of South America. This report describes the use of a bacterially expressed recombinant nucleocapsid (rN) protein of ORO virus in EIAs for the diagnosis of ORO virus infection. The data revealed that the purified rN protein is comparable to the authentic viral N protein in its antigenic characteristics and is highly sensitive and specific in EIAs. Among 183 serum samples tested, a high degree of concordance was found between rN protein-based EIA and HSA- and VCLA-based EIAs for the detection of both ORO virus-specific immunoglobulin M (IgM) and IgG antibodies. The high sensitivity, specificity, and safety of the rN protein-based EIA make it a useful diagnostic technique that can be widely used to detect ORO virus infection in South America.

An outbreak of delta hepatitis occurred during 1998 among the Waorani of the Amazon basin of Ecuador. Among 58 people identified with jaundice, 79% lived in four of 22 Waorani communities. Serum hepatitis B surface antigen (HBsAg) was found in the sera of 54% of the jaundiced persons, and 14% of asymptomatic persons. Ninety-five percent of 105 asymptomatic Waorani had hepatitis B core (HBc) IgG antibody, versus 98% of 51 with jaundice. These data confirm that hepatitis B virus (HBV) infection is highly endemic among the Waorani. Sixteen of 23 (70%) HBsAg carriers identified at the onset of the epidemic had serologic markers for hepatitis D virus (HDV) infection. All 16 were jaundiced, where as only two of seven (29%) with negative HDV serology were jaundiced (P = .0006). The delta cases clustered in families, 69% were children and most involved superinfection of people chronically infected with HBV. The data suggest that HDV spread rapidly by a horizontal mode of transmission other than by the sexual route.

The Naval Health Research Center conducted laboratory-based surveillance for febrile respiratory infections at the 2003 Cobra Gold Exercise in Thailand. Seventeen individuals met the case definition for febrile respiratory illness, and diagnostic specimens were obtained from 16. Laboratory testing identified influenza A for 44%; sequence analysis demonstrated that these were Fujian-like influenza strains, which represented the predominant strain found globally in 2003/2004. Other pathogens identified included coronavirus OC43, respiratory syncytial virus, and rhinovirus. Logistical challenges were overcome as laboratory-supported febrile respiratory illness surveillance was conducted during a military training exercise. With heightened concern over the potential for another global influenza pandemic, such surveillance could prove critical for the detection of emerging influenza and respiratory pathogen strains with potential for importation to the United States.

IntroductionFollowing the 2009 influenza A/H1N1 (pH1N1) pandemic, both seasonal and pH1N1 viruses circulated in the US during the 2010-2011 influenza season; influenza vaccine effectiveness (VE) may vary between live attenuated (LAIV) and trivalent inactivated (TIV) vaccines as well as by virus subtype.Materials and methodsVaccine type and virus subtype-specific VE were determined for US military active component personnel for the period of September 1, 2010 through April 30, 2011. Laboratory-confirmed influenza-related medical encounters were compared to matched individuals with a non-respiratory illness (healthy controls), and unmatched individuals who experienced a non-influenza respiratory illness (test-negative controls). Odds ratios (OR) and VE estimates were calculated overall, by vaccine type and influenza subtype.ResultsA total of 603 influenza cases were identified. Overall VE was relatively low and similar regardless of whether healthy controls (VE = 26%, 95% CI: -1 to 45) or test-negative controls (VE = 29%, 95% CI: -6 to 53) were used as comparison groups. Using test-negative controls, vaccine type-specific VE was found to be higher for TIV (53%, 95% CI: 25 to 71) than for LAIV (VE = -13%, 95% CI: -77 to 27). Influenza subtype-specific analyses revealed moderate protection against A/H3 (VE = 58%, 95% CI: 21 to 78), but not against A/H1 (VE = -38%, 95% CI: -211 to 39) or B (VE = 34%, 95% CI: -122 to 80).ConclusionOverall, a low level of protection against clinically-apparent, laboratory-confirmed, influenza was found for the 2010-11 seasonal influenza vaccines. TIV immunization was associated with higher protection than LAIV, however, no protection against A/H1 was noted, despite inclusion of a pandemic influenza strain as a vaccine component for two consecutive years. Vaccine virus mismatch or lower immunogenicity may have contributed to these findings and deserve further examination in controlled studies. Continued assessment of VE in military personnel is essential in order to better inform vaccination policy decisions.

When introduced in the 1950s, benzathine penicillin G (BPG) was shown to be effective in eradicating group A beta-hemolytic streptococcus (GAS) for at least 3 weeks after administration. Several studies since the 1990s suggest that at 3-4 weeks serum penicillin G levels are less than adequate (below MIC(90) of 0.016 µg/ml). We studied these levels for 4 weeks after the recommended dose of BPG in military recruits, for whom it is used as prophylaxis against GAS. The 329 subjects (mean age 20 years) each received 1.2 million units BPG IM and gave sera 1 day post injection and twice more at staggered time points over 4 weeks. Serum penicillin G levels were measured by liquid chromatography/tandem mass spectometry. The half-life of serum penicillin G was 4.1 days. By day 11, mean levels were

Oropouche (ORO) virus, a member of the Simbu serogroup, is one of the few human pathogens in the Orthobunyavirus genus in the family Bunyaviridae. Genetic analyses of ORO-like strains from Iquitos, Peru, identified a novel reassortant containing the S and L segments of ORO virus and the M segment of a novel Simbu serogroup virus. This new pathogen, which we named Iquitos (IQT) virus, was first isolated during 1999 from a febrile patient in Iquitos, an Amazonian city in Peru. Subsequently, the virus was identified as the cause of outbreaks of “Oropouche fever” during 2005 and 2006 in Iquitos. In addition to the identification of 17 isolates of IQT virus between 1999 and 2006, surveys for neutralizing antibody among Iquitos residents revealed prevalence rates of 14.9% for ORO virus and 15.4% for IQT virus. Limited studies indicate that prior infection with ORO virus does not seem to protect against disease caused with the IQT virus infection. Identification of a new Orthobunyavirus human pathogen in the Amazon region of Peru highlights the need for strengthening surveillance activities and laboratory capabilities, and investigating the emergence of new pathogens in tropical regions of South America., Author Summary Oropouche (ORO) virus is one of the few human pathogens in the Orthobunyavirus genus in the family Bunyaviridae. Phylogenetic analyses of ORO-like strains isolated from febrile patients in Iquitos, Peru, identified a novel ORO reassortant virus, which we named Iquitos (IQT) virus based on the location of the isolation of the virus. This novel pathogen was first isolated during 1999 from a 13-year-old boy who had an illness that included symptoms of fever, headache, eye pain, body pain, arthralgias, diarrhea, and chills. Subsequently, the virus was identified as the cause of outbreaks of “Oropouche fever” during 2005 and 2006 in Iquitos. Limited serological studies indicate that prior infection with ORO virus does not seem to protect against disease caused with the IQT virus infection. In summary, we identified a new Orthobunyavirus human pathogen in the Amazon region of Peru; these results highlight the need for strengthening surveillance activities and investigating the emergence of new pathogens in tropical regions of South America.

This study addresses the effects of dynamic strain turnover and antibiotic prophylaxis on rates of group A Streptococcus (GAS) antibiotic resistance and disease. The authors analyzed the strain distributions, disease rates, and patterns of antibiotic resistance of 802 GAS isolates collected from 2002 through 2007. These samples were collected from patients with GAS infection symptoms at 10 military facilities. Macrolide resistance peaked at 25% during 2004, due to the geographically widespread dominance of a single resistant strain (M75). The resistant strain was not retained regardless of local patterns of macrolide use, and resistance rates decreased upon replacement of M75 with macrolide-susceptible strains. Disease rates were similarly correlated with dominance of specific M types. Statistical analysis revealed temporal correlations between strain distributions at multiple locations. Only the most common strains yielded enough data at multiple sites for statistically significant comparison of temporal fluctuations in dominance, but these (including M44, M3, M18, M118, and M6) all yielded highly significant temporal correlations of 90% or greater on yearly scales. As expected given the complexity and variability of strain distributions on shorter time scales, analysis on a monthly scale yielded lower degrees of positive correlation (31-62%), but in this case all significant correlations were still positive. Shifts in antibiotic resistance profiles and disease rates at specific sites appear to be associated with strain replacements happening on larger scales, independent of antibiotic use at individual sites.

Background Comprehensive, longitudinal field studies that monitor both disease and vector populations for dengue viruses are urgently needed as a pre-requisite for developing locally adaptable prevention programs or to appropriately test and license new vaccines. Methodology and Principal Findings We report the results from such a study spanning 5 years in the Amazonian city of Iquitos, Peru where DENV infection was monitored serologically among ∼2,400 members of a neighborhood-based cohort and through school-based absenteeism surveillance for active febrile illness among a subset of this cohort. At baseline, 80% of the study population had DENV antibodies, seroprevalence increased with age, and significant geographic variation was observed, with neighborhood-specific age-adjusted rates ranging from 67.1 to 89.9%. During the first 15 months, when DENV-1 and DENV-2 were co-circulating, population-based incidence rates ranged from 2–3 infections/100 person-years (p-years). The introduction of DENV-3 during the last half of 2001 was characterized by 3 distinct periods: amplification over at least 5–6 months, replacement of previously circulating serotypes, and epidemic transmission when incidence peaked at 89 infections/100 p-years. Conclusions/Significance Neighborhood-specific baseline seroprevalence rates were not predictive of geographic incidence patterns prior to the DENV-3 introduction, but were closely mirrored during the invasion of this serotype. Transmission varied geographically, with peak incidence occurring at different times among the 8 geographic zones in ∼16 km2 of the city. The lag from novel serotype introduction to epidemic transmission and knowledge of spatially explicit areas of elevated risk should be considered for more effective application of limited resources for dengue prevention., Author Summary To develop prevention (including vaccines) and control programs for dengue fever, a significant mosquito-borne disease in the tropics, there is an urgent need for comprehensive long term field epidemiological studies. We report results from a study that monitored ∼2,400 school children and some adult family members for dengue infection at 6 month intervals from 1999 to 2005, in the Amazonian city of Iquitos, Peru. At enrollment, ∼80% of the participants had a previous infection with DENV serotypes 1 and 2 or both. During the first 15 months, about 3 new infections for every 100 participants were observed among the study participants. In 2001, DENV-3, a serotype not previously observed in the region, invaded Iquitos in a process characterized by 3 distinct periods: amplification over at least a 5–6 month period, replacement of previously circulating serotypes, and epidemic transmission when incidence peaked. Incidence patterns of new infections were geographically distinct from baseline prevalence rates prior to arrival of DENV-3, but closely mirrored them during the invasion. DENV transmission varied geographically corresponding to elevated mosquito densities. The invasion of a novel serotype is often characterized by 5–6 months of silent transmission before traditional surveillance programs detect the virus. This article sets the stage for subsequent publications on dengue epidemiology.

INTRODUCTION: A novel A/H1N1 virus is the cause of the present influenza pandemic; vaccination is a key countermeasure, however, few data assessing prior seasonal vaccine effectiveness (VE) against the pandemic strain of H1N1 (pH1N1) virus are available. MATERIALS AND METHODS: Surveillance of influenza-related medical encounter data of active duty military service members stationed in the United States during the period of April-October 2009 with comparison of pH1N1-confirmed cases and location and date-matched controls. Crude odds ratios (OR) and VE estimates for immunized versus non-immunized were calculated as well as adjusted OR (AOR) controlling for sex, age group, and history of prior influenza vaccination. Separate stratified VE analyses by vaccine type (trivalent inactivated [TIV] or live attenuated [LAIV]), age groups and hospitalization status were also performed. For the period of April 20 to October 15, 2009, a total of 1,205 cases of pH1N1-confirmed cases were reported, 966 (80%) among males and over one-half (58%) under 25 years of age. Overall VE for service members was found to be 45% (95% CI, 33 to 55%). Immunization with prior season's TIV (VE = 44%, 95% CI, 32 to 54%) as well as LAIV (VE = 24%, 95% CI, 6 to 38%) were both found to be associated with protection. Of significance, VE against a severe disease outcome was higher (VE = 62%, 95% CI, 14 to 84%) than against milder outcomes (VE = 42%, 95% CI, 29 to 53%). CONCLUSION: A moderate association with protection against clinically apparent, laboratory-confirmed Pandemic (H1N1) 2009-associated illness was found for immunization with either TIV or LAIV 2008-09 seasonal influenza vaccines. This association with protection was found to be especially apparent for severe disease as compared to milder outcome, as well as in the youngest and older populations. Prior vaccination with seasonal influenza vaccines in 2004-08 was also independently associated with protection.

Background Group A Streptococcus pyogenes (GAS) exhibits a high degree of clinically relevant phenotypic diversity. Strains vary widely in terms of antibiotic resistance (AbR), clinical severity, and transmission rate. Currently, strain identification is achieved by emm typing (direct sequencing of the genomic segment coding for the antigenic portion of the M protein) or by multilocus genotyping methods. Phenotype analysis, including critical AbR typing, is generally achieved by much slower and more laborious direct culture-based methods. Methodology/Principal Findings We compare genotype identification (by emm typing and PCR/ESI-MS) with directly measured phenotypes (AbR and outbreak associations) for 802 clinical isolates of GAS collected from symptomatic patients over a period of 6 years at 10 military facilities in the United States. All independent strain characterization methods are highly correlated. This shows that recombination, horizontal transfer, and other forms of reassortment are rare in GAS insofar as housekeeping genes, primary virulence and antibiotic resistance determinants, and the emm gene are concerned. Therefore, genotyping methods offer an efficient way to predict emm type and the associated AbR and virulence phenotypes. Conclusions/Significance The data presented here, combined with much historical data, suggest that emm typing assays and faster molecular methods that infer emm type from genomic signatures could be used to efficiently infer critical phenotypic characteristics based on robust genotype: phenotype correlations. This, in turn, would enable faster and better-targeted responses during identified outbreaks of constitutively resistant or particularly virulent emm types.

Background Military personnel are highly susceptible to febrile respiratory illnesses (FRI), likely due to crowding, stress and other risk factors present in the military environment. Objective Our objective was to investigate the viral etiological agents responsible for FRI among military recruits training in a tropical climate in Singapore. Study design From March 2006 through April 2007, a total of 1354 oropharyngeal (throat) swabs were collected from military recruits who reported sick with an oral temperature of ≥38 °C and a cough and/or sore throat. Real-time polymerase chain reaction (PCR) was used to assay for the presence of influenza A and B viruses and adenoviruses (H-AdV), and conventional PCR used for the remaining respiratory viruses in all specimens. Results Influenza A virus was the dominant infection with a laboratory-confirmed incidence of 24% (326/1354) and a predominance of the H3N2 subtype. The temporal pattern for influenza A virus infections coincided with the nation-wide pattern in the civilian community. Detection rates of 12% (159/1354) and 2.7% (5/1354) were obtained for influenza B virus and other respiratory viruses, respectively. Conclusions The laboratory findings identified influenza A virus as the primary causative viral agent for FRI in the Singapore military, in strong contrast to findings from temperate countries and countries where recruits are often vaccinated for influenza. Our results suggest that influenza vaccination should be considered as a requirement to reduce the incidence of influenza infections. This is the first report describing respiratory infections in a tropical military setting, in a developed country in Asia.

Military recruits experience a high incidence of febrile respiratory illness (FRI), leading to significant morbidity and lost training time. Adenoviruses, group A Streptococcus pyogenes, and influenza virus are implicated in over half of the FRI cases reported at recruit training center clinics, while the etiology of the remaining cases is unclear. In this study, we explore the carriage rates and disease associations of adenovirus, enterovirus, rhinovirus, Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis in military recruits using high-density resequencing microarrays. The results showed that rhinoviruses, adenoviruses, S. pneumoniae, H. influenzae, and N. meningitidis were widely distributed in recruits. Of these five agents, only adenovirus showed significant correlation with illness. Among the samples tested, only pathogens associated with FRI, such as adenovirus 4 and enterovirus 68, revealed strong temporal and spatial clustering of specific strains, indicating that they are transmitted primarily within sites. The results showed a strong negative association between adenoviral FRI and the presence of rhinoviruses in recruits, suggesting some form of viral interference.

Background. Effective influenza surveillance requires new methods capable of rapid and inexpensive genomic analysis of evolving viral species for pandemic preparedness, to understand the evolution of circulating viral species, and for vaccine strain selection. We have developed one such approach based on previously described broad-range reverse transcription PCR/ electrospray ionization mass spectrometry (RT-PCR/ESI-MS) technology. Methods and Principal Findings. Analysis of base compositions of RT-PCR amplicons from influenza core gene segments (PB1, PB2, PA, M, NS, NP) are used to provide subspecies identification and infer influenza virus H and N subtypes. Using this approach, we detected and correctly identified 92 mammalian and avian influenza isolates, representing 30 different H and N types, including 29 avian H5N1 isolates. Further, direct analysis of 656 human clinical respiratory specimens collected over a seven-year period (1999–2006) showed correct identification of the viral species and subtypes with .97% sensitivity and specificity. Base composition derived clusters inferred from this analysis showed 100% concordance to previously established clades. Ongoing surveillance of samples from the recent influenza virus seasons (2005–2006) showed evidence for emergence and establishment of new genotypes of circulating H3N2 strains worldwide. Mixed viral quasispecies were found in approximately 1% of these recent samples providing a view into viral evolution. Conclusion/Significance. Thus, rapid RT-PCR/ESI-MS analysis can be used to simultaneously identify all species of influenza viruses with clade-level resolution, identify mixed viral populations and monitor global spread and emergence of novel viral genotypes. This high-throughput method promises to become an integral component of influenza surveillance.

Background. High levels of morbidity caused by adenovirus among US military recruits have returned since the loss of adenovirus vaccines in 1999. The transmission dynamics of adenovirus have never been well understood, which complicates prevention efforts. Methods. Enrollment and end-of-study samples were obtained and active surveillance for febrile respiratory illnesses (FRIs) was performed for 341 recruits and support personnel. Environmental samples were collected simultaneously. Classic and advanced diagnostic techniques were used. Results. Seventy-nine percent (213/271) of new recruits were seronegative for either adenovirus serotype 4 (Ad-4) or adenovirus serotype 7 (Ad-7). FRI caused by Ad-4 was observed in 25% (67/271) of enrolled recruits, with 100% of them occurring in individuals with enrollment titers

The study's purpose was to elucidate the evolutionary, microbiologic, and clinical characteristics of methicillin-resistant Staphylococcus aureus (MRSA) infections.MRSA cases from military medical facilities in San Diego, from 1990 to 2004, were evaluated and categorized as community-acquired or nosocomial. Sequence type, staphylococcal chromosomal cassette gene type, and Panton-Valentine leukocidin gene status were determined for a subset of isolates.Over the 15-year period, 1888 cases of MRSA were identified; 65% were community acquired. The incidence (155 infections/100000 person-year in 2004) and household-associated cases rapidly increased since 2002. Among persons with community-acquired MRSA, 16% were hospitalized and only 17% were initially given an effective antibiotic. Community-acquired MRSA cases compared with nosocomial MRSA cases were more often soft-tissue and less often urinary, lung, or bloodstream infections (P.001). Patients with community-acquired MRSA were younger (22 vs 64 years, P.001) and less likely to have concurrent medical conditions (9% vs 98%, P.001). Clindamycin resistance increased among community-acquired MRSA isolates during 2003 and 2004 compared with previous years (79% vs 13%, P.001). Genetically, nosocomial MRSA isolates were significantly different than those acquired in the community. Although community-acquired MRSA isolates were initially diverse by 2004, one strain (staphylococcal chromosomal cassette type IV, sequence type 8, Panton-Valentine leukocidin gene positive) became the predominant isolate.Community-acquired and intrafamilial MRSA infections have increased rapidly since 2002. Our 15 years of surveillance revealed the emergence of distinct community-acquired MRSA strains that were genetically unrelated to nosocomial MRSA isolates from the same community.

Epidemic respiratory infections are responsible for extensive morbidity and mortality within both military and civilian populations. We describe a high-throughput method to simultaneously identify and genotype species of bacteria from complex mixtures in respiratory samples. The process uses electrospray ionization mass spectrometry and base composition analysis of PCR amplification products from highly conserved genomic regions to identify and determine the relative quantity of pathogenic bacteria present in the sample. High-resolution genotyping of specific species is achieved by using additional primers targeted to highly variable regions of specific bacterial genomes. This method was used to examine samples taken from military recruits during respiratory disease outbreaks and for follow up surveillance at several military training facilities. Analysis of respiratory samples revealed high concentrations of pathogenic respiratory species, including Haemophilus influenzae , Neisseria meningitidis , and Streptococcus pyogenes . When S. pyogenes was identified in samples from the epidemic site, the identical genotype was found in almost all recruits. This analysis method will provide information fundamental to understanding the polymicrobial nature of explosive epidemics of respiratory disease.

A multiplex PCR was developed that is capable of detecting four of the most important bacterial agents of atypical pneumonia, Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila, and Bordetella pertussis in uncultured patient specimens. These organisms cause similar symptomologies and are often not diagnosed because they are difficult to identify with classical methods such as culture and serology. Given this, the overall impact of these pathogens on public health may be grossly underestimated. The molecular test presented here provides a simple method for identification of four common, yet diagnostically challenging, pathogens.

Viral culture isolation has been widely accepted as the “gold standard” for laboratory confirmation of viral infection; however, it requires ultralow temperature specimen storage. Storage of specimens in ethanol at room temperature could expand our ability to conduct active surveillance and retrospective screenings of viruses with rapid and inexpensive real-time PCR tests, including isolates from remote regions where freezing specimens for culture is not feasible. Molecular methods allow for rapid identification of viral pathogens without the need to maintain viability. We hypothesized that ethanol, while inactivating viruses, can preserve DNA and RNA for PCR-based methods. To evaluate the use of ethanol-stored specimens for augmenting surveillance for detection of influenza viruses A and B and adenoviruses (AdV), paired nasal swab specimens were collected from 384 recruits with febrile respiratory illness at Fort Jackson, S.C., in a 2-year study. One swab was stored at ambient temperature in 100% ethanol for up to 6 months, and the other swab was stored at −70°C in viral medium. For viral detection, frozen specimens were cultured for a variety of respiratory viruses, and ethanol-fixed specimens were tested with TaqMan (TM) probe and LightCycler SYBR green (SG) melting curve assays with at least two different PCR targets for each virus. The sensitivities of the TM and SG assays on specimens stored in ethanol for 1 month were 75% and 58% for influenza A, 89% and 67% for influenza B, and 93 to 98% and 57% for AdV, respectively. Lower specificities of the real-time assays corresponded to the increased detection of PCR-positive but culture-negative specimens. Influenza virus RNA was detected as well or better after 6 months of storage in ethanol.

An outbreak of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) skin infections was observed in a population of U.S. military trainees in the summer of 2002. A questionnaire was developed and administered to 206 trainees, 22 of whom had MRSA infections. Factors associated with infection were described by multivariable logistic regression modeling and included having a roommate in training with a prior skin infection (odds ratio [OR] = 3.44) or having a family member or friend who worked in a health care setting (OR = 2.79). Previous antibiotic use, hospitalization, or health problems were not associated with MRSA infection. This outbreak of MRSA skin infections in an otherwise-healthy, well-defined, military population provided an opportunity to describe risk factors for CA-MRSA which may help focus prevention efforts in this and other communities.

Background Bacterial conjunctivitis usually occurs as sporadic cases; outbreaks are uncommon and usually are associated with school campuses. We report an outbreak of Streptococcus pneumoniae conjunctivitis at a military training facility. Methods An outbreak investigation was done. Each case of conjunctivitis was evaluated with an assessment tool including demographic and clinical data. Conjunctival swabs were obtained. Pneumococci underwent standard testing, including serotyping with the Quellung reaction, capsular staining, and multilocus sequence typing. Sequence types were compared with previous reported outbreak strains by construction of dendrograms. Carriage rates of S. pneumoniae were determined among previously undiagnosed case patients with conjunctivitis, and a case-control study was performed. Control measures included education to increase hand washing, distribution of alcohol-based hand gel, and prompt treatment of patients with conjunctivitis. Results During a 6-week period, 92 cases of conjunctivitis occurred among 3500 persons, with an attack rate of 1.75 cases per 100 person-months. Eighty cases (87%) were due to S. pneumoniae; 45 (49%) were confirmed, and 35 (38%) were probable. Ten percent of recruits surveyed carried the outbreak strain. Twenty-two percent self-reported symptoms consistent with conjunctivitis during the outbreak period; sharing washcloths was associated with conjunctivitis (odds ratio, 11.7; P=.03). The causative organism was resistant to azithromycin but susceptible to telithromycin. The outbreak strain was an unencapsulated S. pneumoniae that has not been previously described; it was most closely related to the sequence type causing the Dartmouth College (Hanover, NH) outbreak of conjunctivitis in 2002. Conclusions We report a conjunctivitis outbreak among military trainees caused by a novel, unencapsulated strain of S. pneumoniae.

Previously, we observed that serum from humans immune to dengue serotype 1 (dengue-1) neutralized the American genotype of dengue serotype 2 (American-2) to a greater extent than it neutralized the Asian genotype of dengue serotype 2 (Asian-2). To determine if this activity is protective, Aotus nancymae monkeys were infected with dengue-1 followed by either American-2 or Asian-2. Dengue-1-infected animals produced antibody with neutralizing titers of 2656 antibodies against dengue-1, 409 against American-2, and

Infections of adenovirus type 4 (Ad4) and Ad7 were discovered among previously vaccinated individuals through febrile respiratory illness surveillance at military recruit camps. Genetic analysis was performed on these isolates and a sample of adenovirus isolates from unvaccinated patients. Antigenic regions of the adenovirus hexon gene from 21 vaccinated and 31 unvaccinated patients were sequenced and compared to homologous regions of Ad4 and Ad7 vaccine strains and of other representative hexon sequences archived in GenBank. The phylogenetic distribution of sequences from vaccinated individuals closely resembled those from unvaccinated individuals. The most common Ad7 strain was the Ad7d2 hexon genotype, and the most common Ad4 strain was a genotype nearly identical to the recently discovered Z-G 95-873 Ad4 variant. Near exclusive isolation of Ad4 since 1999 indicates that the Ad4 variant is currently responsible for the vast majority of adenovirus morbidity in military recruit camps. Different ratios of nonsynonymous to synonymous nucleotide substitution rates in known antigenic regions compared to nonantigenic regions indicated positive selection for diversity in the antigenic regions and purifying selection in the nonantigenic regions.

In December 2001, a fatal case of pneumococcal meningitis in a Marine Corps recruit was identified. The pneumococcal isolate was serotype 38, a serotype not covered by current pneumococcal vaccine formulations, This is the first report in North America of invasive disease due to this potentially emerging pathogen.

Antibiotic resistance and emm gene types were examined from 692 Group A streptococci isolates from eight United States military basic training sites between 1998 and 2001. Macrolide resistance was associated with geographic sites and emm type. These data are useful for vaccine development initiatives and antimicrobial treatment considerations.

In 1994-1996, 185 strains of dengue (DEN) virus types 1, 2, and 4 were recovered from febrile United States and other United Nations military personnel in Haiti. We wondered whether risk factors for dengue hemorrhagic fever (DHF) existed and, if so, were DHF cases occurring among Haitian children. Dengue transmission rates were studied in 210 school children (6-13 years old) resident in Carrefour Borough, Port-au-Prince, Haiti. When sera were tested for plaque-reduction neutralizing antibodies to DEN 1-4 viruses, nearly 85% had antibodies to two or more DEN serotypes. The annual transmission rate was estimated at 30%, a rate observed in countries endemic for DHE Haitian DEN 2 isolates were genotype I, which are repeatedly associated with DHF cases in Southeast Asia and American regions. Despite positive virologic pre-conditions, DHF cases were not recorded by experienced Port-au-Prince pediatricians. These observations, which are reminiscent of those in Africa, provide further evidence of a dengue resistance gene in black populations.

The 3' non-coding region (3'NCR) of strains of dengue 1 (DEN 1), DEN 2, DEN 3, and DEN 4 viruses, isolated in different geographical regions, was sequenced and compared to published sequences of the four dengue viruses. A total of 50 DEN 2 strains was compared: 7 West African strains, 3 Indonesian mosquito strains, 1 Indonesian macaque isolate, and 39 human isolates from Southeast Asia, the South Pacific, and the Caribbean and Americas. Nucleotide sequence alignment revealed few deletions and no repeat sequences in the 3' NCR of DEN 2 viruses and showed that much of the 3' NCR was well conserved. The strains could be divided into two groups, sylvatic and human/mosquito/macaque, based on nucleotide sequence homology. A hypervariable region was identified immediately following the NS5 stop codon, which involved a 2-10 nucleotide deletion in human, mosquito, and macaque isolates compared with the sylvatic strains. The DEN 2 3'NCR was also compared with 3'NCR sequences from strains of DEN 1, DEN 3, and DEN 4 viruses. DEN 1 was found to have four copies of an eight nucleotide imperfect repeat following the NS5 stop codon, while DEN 4 virus had a deletion of 75 nucleotides in the 3'NCR. We propose that the variation in nucleotide sequence in the 3'NCR may have evolved as a function of DEN virus transmission and replication in different mosquito and non-human primate/human host cycles. The results from this study are consistent with the hypothesis that DEN viruses arose from sylvatic progenitors and evolved into human epidemic strains. However, the data do not support the hypothesis that variation in the 3'NCR correlates with DEN virus pathogenesis.

A DNA vaccine that expresses the premembrane/membrane (prM) and envelope (E) genes of dengue virus serotype-1 was tested for immunogenicity and protection against dengue-1 virus challenge in Aotus nancymae monkeys. The vaccine, in 1 mg doses, was administered intradermally (i.d.) to three monkeys and intramuscularly (i.m.) to three others. For controls, a 1 mg dose of vector DNA was administered i.d. to two monkeys and i.m. to one. All animals were primed and then boosted at one and five months post priming. Sera were collected monthly and analyzed for dengue-1 antibodies by enzyme linked immunosorbent assay (ELISA) and plaque reduction neutralization test (PRNT). Dengue-1 antibodies were detectable in the sera from i.d. and i.m. vaccine inoculated animals one month after the first boost and peaked one month after the second boost. The antibody levels from sera of animals that received the vaccine via the i.d. route were twice those from sera of animals that received the vaccine via the i.m. route. Six months after the second boost all inoculated and two naive monkeys were challenged with 1.25x10(4) plaque forming units (PFU) of dengue-1 virus. Two vaccine immunized animals were protected from viremia while the others showed a reduction in viremia. The mean days of viremia were 1 and 1.3 for the animals that were immunized with the vaccine via the i.d. or i.m. route, respectively vs 4 and 2 mean days of viremia in the animals inoculated with control DNA. Naive animals were viremic for an average of 4 days. All of the three control monkeys that received control DNA inoculum by either the i.d. or i.m. route had an intermittent viremia pattern with one or more negative days interspersed between the positive days. This pattern was not observed in any of the vaccine recipients or the naïve control monkeys. These results demonstrate that DNA immunization is a promising approach for the development of dengue vaccines and that A. nancymae monkeys are suitable for dengue vaccine trials.

Background Knowledge of spatial patterns of dengue virus (DENV) infection is important for understanding transmission dynamics and guiding effective disease prevention strategies. Because movement of infected humans and mosquito vectors plays a role in the spread and persistence of virus, spatial dimensions of transmission can range from small household foci to large community clusters. Current understanding is limited because past analyses emphasized clinically apparent illness and did not account for the potentially large proportion of inapparent infections. In this study we analyzed both clinically apparent and overall infections to determine the extent of clustering among human DENV infections. Methodology/Principal Findings We conducted spatial analyses at global and local scales, using acute case and seroconversion data from a prospective longitudinal cohort in Iquitos, Peru, from 1999–2003. Our study began during a period of interepidemic DENV-1 and DENV-2 transmission and transitioned to epidemic DENV-3 transmission. Infection status was determined by seroconversion based on plaque neutralization testing of sequential blood samples taken at approximately six-month intervals, with date of infection assigned as the middate between paired samples. Each year was divided into three distinct seasonal periods of DENV transmission. Spatial heterogeneity was detected in baseline seroprevalence for DENV-1 and DENV-2. Cumulative DENV-3 seroprevalence calculated by trimester from 2001–2003 was spatially similar to preexisting DENV-1 and DENV-2 seroprevalence. Global clustering (case-control Ripley's K statistic) appeared at radii of ∼200–800 m. Local analyses (Kuldorf spatial scan statistic) identified eight DENV-1 and 15 DENV-3 clusters from 1999–2003. The number of seroconversions per cluster ranged from 3–34 with radii from zero (a single household) to 750 m; 65% of clusters had radii >100 m. No clustering was detected among clinically apparent infections. Conclusions/Significance Seroprevalence of previously circulating DENV serotypes can be a predictor of transmission risk for a different invading serotype and, thus, identify targets for strategically placed surveillance and intervention. Seroprevalence of a specific serotype is also important, but does not preclude other contributing factors, such as mosquito density, in determining where transmission of that virus will occur. Regardless of the epidemiological context or virus serotype, human movement appears to be an important factor in defining the spatial dimensions of DENV transmission and, thus, should be considered in the design and evaluation of surveillance and intervention strategies., Author Summary To target prevention and control strategies for dengue fever, it is essential to understand how the virus travels through the city. We report spatial analyses of dengue infections from a study monitoring school children and adult family members for dengue infection at six-month intervals from 1999–2003, in the Amazonian city of Iquitos, Peru. At the beginning of the study, only DENV serotypes 1 and 2 were circulating. Clusters of infections of these two viruses were concentrated in the northern region of the city, where mosquito indices and previous DENV infection were both high. In 2002, DENV-3 invaded the city, replacing DENV-1 and -2 as the dominant strain. During the invasion process, the virus spread rapidly across the city, at low levels. After this initial phase, clusters of infection appeared first in the northern region of the city, where clusters of DENV-1 and DENV-2 had occurred in prior years. Most of the clusters we identified had radii >100 meters, indicating that targeted or reactive treatment of these high-risk areas might be an effective proactive intervention strategy. Our results also help explain why vector control within 100 m of a dengue case is often not successful for large-scale disease prevention.

Background Arthropod-borne viruses (arboviruses) are among the most common agents of human febrile illness worldwide and the most important emerging pathogens, causing multiple notable epidemics of human disease over recent decades. Despite the public health relevance, little is know about the geographic distribution, relative impact, and risk factors for arbovirus infection in many regions of the world. Our objectives were to describe the arboviruses associated with acute undifferentiated febrile illness in participating clinics in four countries in South America and to provide detailed epidemiological analysis of arbovirus infection in Iquitos, Peru, where more extensive monitoring was conducted. Methodology/Findings A clinic-based syndromic surveillance system was implemented in 13 locations in Ecuador, Peru, Bolivia, and Paraguay. Serum samples and demographic information were collected from febrile participants reporting to local health clinics or hospitals. Acute-phase sera were tested for viral infection by immunofluorescence assay or RT-PCR, while acute- and convalescent-phase sera were tested for pathogen-specific IgM by ELISA. Between May 2000 and December 2007, 20,880 participants were included in the study, with evidence for recent arbovirus infection detected for 6,793 (32.5%). Dengue viruses (Flavivirus) were the most common arbovirus infections, totaling 26.0% of febrile episodes, with DENV-3 as the most common serotype. Alphavirus (Venezuelan equine encephalitis virus [VEEV] and Mayaro virus [MAYV]) and Orthobunyavirus (Oropouche virus [OROV], Group C viruses, and Guaroa virus) infections were both observed in approximately 3% of febrile episodes. In Iquitos, risk factors for VEEV and MAYV infection included being male and reporting to a rural (vs urban) clinic. In contrast, OROV infection was similar between sexes and type of clinic. Conclusions/Significance Our data provide a better understanding of the geographic range of arboviruses in South America and highlight the diversity of pathogens in circulation. These arboviruses are currently significant causes of human illness in endemic regions but also have potential for further expansion. Our data provide a basis for analyzing changes in their ecology and epidemiology., Author Summary Over recent decades, the variety and quantity of diseases caused by viruses transmitted to humans by mosquitoes and other arthropods (also known as arboviruses) have increased around the world. One difficulty in studying these diseases is the fact that the symptoms are often non-descript, with patients reporting such symptoms as low-grade fever and headache. Our goal in this study was to use laboratory tests to determine the causes of such non-descript illnesses in sites in four countries in South America, focusing on arboviruses. We established a surveillance network in 13 locations in Ecuador, Peru, Bolivia, and Paraguay, where patient samples were collected and then sent to a central laboratory for testing. Between May 2000 and December 2007, blood serum samples were collected from more than 20,000 participants with fever, and recent arbovirus infection was detected for nearly one third of them. The most common viruses were dengue viruses (genera Flavivirus). We also detected infection by viruses from other genera, including Alphavirus and Orthobunyavirus. This data is important for understanding how such viruses might emerge as significant human pathogens.

For more than four decades the cause of most type A influenza virus infections of humans has been attributed to only two viral subtypes, A/H1N1 or A/H3N2. In contrast, avian and other vertebrate species are a reservoir of type A influenza virus genome diversity, hosting strains representing at least 120 of 144 combinations of 16 viral hemagglutinin and 9 viral neuraminidase subtypes. Viral genome segment reassortments and mutations emerging within this reservoir may spawn new influenza virus strains as imminent epidemic or pandemic threats to human health and poultry production. Traditional methods to detect and differentiate influenza virus subtypes are either time-consuming and labor-intensive (culture-based) or remarkably insensitive (antibody-based). Molecular diagnostic assays based upon reverse transcriptase-polymerase chain reaction (RT-PCR) have short assay cycle time, and high analytical sensitivity and specificity. However, none of these diagnostic tests determine viral gene nucleotide sequences to distinguish strains and variants of a detected pathogen from one specimen to the next. Decision-quality, strain- and variant-specific pathogen gene sequence information may be critical for public health, infection control, surveillance, epidemiology, or medical/veterinary treatment planning. The Resequencing Pathogen Microarray (RPM-Flu) is a robust, highly multiplexed and target gene sequencing-based alternative to both traditional culture- or biomarker-based diagnostic tests. RPM-Flu is a single, simultaneous differential diagnostic assay for all subtype combinations of type A influenza viruses and for 30 other viral and bacterial pathogens that may cause influenza-like illness. These other pathogen targets of RPM-Flu may co-infect and compound the morbidity and/or mortality of patients with influenza. The informative specificity of a single RPM-Flu test represents specimen-specific viral gene sequences as determinants of virus type, A/HN subtype, virulence, host-range, and resistance to antiviral agents.

Background In December 2001, a fatal case of pneumococcal meningitis in a Marine Corps recruit was identified. As pneumococcal vaccine usage in recruit populations is being considered, an investigation was initiated into the causative serotype. Case presentation Traditional and molecular methods were utilized to determine the serotype of the infecting pneumococcus. The pneumococcal isolate was identified as serotype 38 (PS38), a serotype not covered by current vaccine formulations. The global significance of this serotype was explored in the medical literature, and found to be a rare but recognized cause of carriage and invasive disease. Conclusion The potential of PS38 to cause severe disease is documented in this report. Current literature does not support the hypothesis that this serotype is increasing in incidence. However, as we monitor the changing epidemiology of pneumococcal illness in the US in this conjugate era, PS38 might find a more prominent and concerning niche as a replacement serotype.

The aim of this study was to conduct an epidemiological and laboratory surveillance of Influenza-Like Illnesses (ILI) in Polish Armed Forces, civilian military personnel and their families in 2011/2012 epidemic season, under the United States Department of Defense-Global Emerging Infections Surveillance and Response System (DoD-GEIS). ILI incidence data were analyzed in relation to age, gender, patient category as well as pathogen patterns. Multiple viral, bacterial and viral-bacterial co-infections were identified. Nose and throat swabs of active duty soldiers in the homeland country and in the NATO peacekeeping forces KFOR (Kosovo Force), as well as members of their families were tested for the presence of viral and bacterial pathogens. From October 2011 to May 2012, 416 specimens from ILI symptoms patients were collected and analyzed for the presence of viral and bacterial pathogens. Among viruses, coronavirus was the most commonly detected. In the case of bacterial infections, the most common pathogen was Staphylococcus aureus.

The Armed Forces Health Surveillance Center’s Division of Global Emerging Infections Surveillance and Response System (AFHSC-GEIS) supports and oversees surveillance for emerging infectious diseases, including respiratory diseases, of importance to the U.S. Department of Defense (DoD). AFHSC-GEIS accomplishes this mission by providing funding and oversight to a global network of partners for respiratory disease surveillance. This report details the system’s surveillance activities during 2009, with a focus on efforts in responding to the novel H1N1 Influenza A (A/H1N1) pandemic and contributions to global public health. Active surveillance networks established by AFHSC-GEIS partners resulted in the initial detection of novel A/H1N1 influenza in the U.S. and several other countries, and viruses isolated from these activities were used as seed strains for the 2009 pandemic influenza vaccine. Partners also provided diagnostic laboratory training and capacity building to host nations to assist with the novel A/H1N1 pandemic global response, adapted a Food and Drug Administration-approved assay for use on a ruggedized polymerase chain reaction platform for diagnosing novel A/H1N1 in remote settings, and provided estimates of seasonal vaccine effectiveness against novel A/H1N1 illness. Regular reporting of the system’s worldwide surveillance findings to the global public health community enabled leaders to make informed decisions on disease mitigation measures and controls for the 2009 A/H1N1 influenza pandemic. AFHSC-GEIS’s support of a global network contributes to DoD’s force health protection, while supporting global public health.