BackgroundNeurotropic arboviral infections are an important cause of encephalitis. A zoonotic, vector-borne alphavirus, Madariaga virus (MADV; formerly known as South American eastern equine encephalitis virus), caused its first documented human outbreak in 2010 in Darien, Panama, where the genetically similar Venezuelan equine encephalitis virus (VEEV) is endemic. We report the results of a seroprevalence survey of animals and humans, illustrating contrasting features of MADV and VEEV ecology and epidemiology.MethodsSmall mammals were trapped in 42 sites in Darien, Panama, using Sherman traps, Tomahawk traps, and mist nets for bats. Blood was tested for the presence of neutralizing antibodies to MADV and VEEV. In addition, bird sera collected in 2007 in Chagres, Panama, were tested for MADV and VEEV neutralizing antibodies. Viremia was ascertained by RT-PCR. Human exposure to these two viruses was determined by IgG ELISA, followed by plaque reduction neutralization tests. To identify relevant risk factors for MADV or VEEV exposure, logistic regression analysis was performed, and the most parsimonious model was selected based on the Akaike information criterion.ResultsThe animal survey yielded 32 bats (16 species), 556 rodents (12 species), and 20 opossums (4 species). The short-tailed cane mouse (Zygodontomys brevicauda) found abundantly in pasture and farms, had the highest MADV seroprevalence (8.3%). For VEEV, the shrub and forest-dwelling long-whiskered rice rat (Transandinomys bolivaris) had the highest seroprevalence (19.0%). Viremia was detected in one animal (Z. brevicauda). Of the 159 bird sera (50 species) tested, none were positive for either virus. In humans (n = 770), neutralizing antibodies to MADV and VEEV were present in 4.8% and 31.5%, respectively. MADV seropositivity was positively associated with cattle ranching, farming, and fishing. Having VEEV antibodies and shrubs near the house diminished risk. Age, forest work, farming and fishing were risk factors for VEEV, while having MADV antibodies, glazed windows, waste pick-up and piped water were protective.ConclusionOur findings suggest that the short-tailed cane mouse and the long-whiskered rice rat serve as hosts for MADV and VEEV, respectively. The preferred habitat of these rodent species coincides with areas associated with human infection risk. Our findings also indicate that MADV emerged recently in humans, and that the transmission cycles of these two sympatric alphaviruses differ spatially and in host utilization.

Hantavirus cardiopulmonary syndrome (HCPS), which is caused by infection with Choclo virus, is uncommon in Panama, yet seropositivity among rural residents is as high as 60%. To clarify the environmental risk factors favoring rodent-to-human transmission, we tested serum from 3,067 rodents captured over a five-year period for antibodies against recombinant N protein of hantavirus by enzyme immunoassay and strip immunoblot. Among 220 seropositive rodents, Oligoryzomys fulvescens, the reservoir of Choclo virus, had the highest overall seroprevalence (23.5%); more abundant rodents (Zygodontomys brevicauda and Sigmodon hirsutus) had lower seroprevalences. In the mixed (combined modern and traditional) productive agroecosystem, the highest seroprevalence was among O. fulvescens captured in residences and in crops grown within 40 meters of a residence, with significantly lower seroprevalence in adjacent pasture and non-productive vegetation. Thus, crop habitats may serve as refugia for invasion into adjacent human residences and suggests several interventions to reduce human infection.

Madariaga virus (MADV) has recently been associated with severe human disease in Panama, where the closely related Venezuelan equine encephalitis virus (VEEV) also circulates. In June 2017, a fatal MADV infection was confirmed in a community of Darien Province. We conducted a cross-sectional outbreak investigation with human and mosquito collections in July 2017, where sera were tested for alphavirus antibodies and viral RNA. In addition, by applying a catalytic, force-of-infection (FOI) statistical model to two serosurveys from Darien Province in 2012 and 2017, we investigated whether endemic or epidemic alphavirus transmission occurred historically. In 2017, MADV and VEEV IgM seroprevalences were 1.6% and 4.4%, respectively; IgG antibody prevalences were MADV: 13.2%, VEEV: 16.8%, Una virus (UNAV): 16.0%, and Mayaro virus: 1.1%. Active viral circulation was not detected. Evidence of MADV and UNAV infection was found near households, raising questions about its vectors and enzootic transmission cycles. Insomnia was associated with MADV and VEEV infections, depression symptoms were associated with MADV, and dizziness with VEEV and UNAV. Force-of-infection analyses suggest endemic alphavirus transmission historically, with recent increased human exposure to MADV and VEEV in Aruza and Mercadeo, respectively. The lack of additional neurological cases suggests that severe MADV and VEEV infections occur only rarely. Our results indicate that over the past five decades, alphavirus infections have occurred at low levels in eastern Panama, but that MADV and VEEV infections have recently increased-potentially during the past decade. Endemic infections and outbreaks of MADV and VEEV appear to differ spatially in some locations of eastern Panama.

BackgroundChikungunya virus (CHIKV) typically causes explosive epidemics of fever, rash and polyarthralgia after its introduction into naïve populations. Since its introduction in Panama in May of 2014, few autochthonous cases have been reported; most of them were found within limited outbreaks in Panama City in 2014 and Puerto Obaldia town, near the Caribbean border with Colombia in 2015. In order to confirm that Panama had few CHIKV cases compared with neighboring countries, we perform an epidemiological analysis of chikungunya cases reported from May 2014 to July 2015. Moreover, to understand this paucity of confirmed CHIKV cases, a vectorial analysis in the counties where these cases were reported was performed.MethodsChikungunya cases were identified at medical centers and notified to health authorities. Sera samples were analyzed at Gorgas Memorial Institute for viral RNA and CHIKV-specific antibody detection.ResultsA total of 413 suspected cases of CHIKV infections were reported, with incidence rates of 0.5 and 0.7 per 100,000 inhabitants in 2014 and 2015, respectively. During this period, 38.6% of CHIKV cases were autochthonous with rash and polyarthralgia as predominant symptoms. CHIKV and DENV incidence ratios were 1:306 and 1:34, respectively. A phylogenetic analysis of E1/E2 genomic segment indicates that the outbreak strains belong to the Asian genotype and cluster together with CHIKV isolates from other American countries during the same period. Statistical analysis of the National Vector Control program at the district level shows low and medium vector infestation level for most of the counties with CHIKV cases. This index was lower than for neighboring countries.ConclusionsPrevious training of clinical, laboratory and vector workers allowed a good caption and detection of the chikungunya cases and fast intervention. It is possible that low/medium vector infestation level could explain in part the paucity of chikungunya infections in Panama.

BackgroundHotspot detection and characterization has played an increasing role in understanding the maintenance and transmission of zoonotic pathogens. Identifying the specific environmental factors (or their correlates) that influence reservoir host abundance help increase understanding of how pathogens are maintained in natural systems and are crucial to identifying disease risk. However, most recent studies are performed at macro-scale and describe broad temporal patterns of population abundances. Few have been conducted at a microscale over short time periods that better capture the dynamical patterns of key populations. These finer resolution studies may better define the likelihood of local pathogen persistence. This study characterizes the landscape distribution and spatio-temporal dynamics of Oligoryzomys fulvescens (O. fulvescens), an important mammalian reservoir in Central America.MethodsInformation collected in a longitudinal study of rodent populations in the community of Agua Buena in Tonosí, Panama, between April 2006 and December 2009 was analyzed using non-spatial analyses (box plots) and explicit spatial statistical tests (correlograms, SADIE and LISA). A 90 node grid was built (raster format) to design a base map. The area between the nodes was 0.09 km2 and the total study area was 6.43 km2 (2.39 x 2.69 km). The temporal assessment dataset was divided into four periods for each year studied: the dry season, rainy season, and two months-long transitions between seasons (the months of April and December).ResultsThere were heterogeneous patterns in the population densities and degrees of dispersion of O. fulvescens that varied across seasons and among years. The species typically was locally absent during the late transitional months of the season, and re-established locally in subsequent years. These populations re-occurred in the same area during the first three years but subsequently re-established further south in the final year of the study. Spatial autocorrelation analyses indicated local populations encompassed approximately 300–600 m. The borders between suitable and unsuitable habitats were sharply demarcated over short distances.ConclusionOligoryzomys fulvescens showed a well-defined spatial pattern that evolved over time, and led to a pattern of changing aggregation. Thus, hot spots of abundance showed a general shifting pattern that helps explain the intermittent risk from pathogens transmitted by this species. This variation was associated with seasonality, as well as anthropogenic pressures that occurred with agricultural activities. These factors help define the characteristics of the occurrence, timing, intensity and duration of synanthropic populations affected by human populations and, consequently, possible exposure that local human populations experience.

ABSTRACT. The identity of two insular populations of rodents of the nominal species, Zygodontomys brevicauda (Allen & Chapman, 1893), from the Veragua Archipelago was examined. The mitochondrial cytochrome-b gene was sequenced from specimens collected on Isla Coiba (n = 10), Isla Cébaco (n = 3) and on the nearby Peninsula Azuero (n = 3) in Panama and compared with sequences of Z. brevicauda and a number of other related species from GenBank. For Panama, phylogenetic analyses identified two clades within Zygodontomys Allen, 1897; one representing Isla Coiba and another clade composed of rats from Isla Cébaco and from the mainland on the Peninsula Azuero, as well as a GenBank sequence of Z. brevicauda from Venezuela. We suggest that the population from Isla Coiba may represent a previously undescribed species of sigmodontine rodent that is endemic to this Pacific Island.

43 44 ¶ These authors contributed equally 1 Author summary. 2 3 Prior to 2010, it was believed that the Madariaga virus (MADV) was primarily associated 4 with equine disease. However, an outbreak reported in Panama, in an endemic area where 5 Venezuelan equine encephalitis virus (VEEV) also circulates, suggested a change in its 6 epidemiological profile. We aimed to reconstruct the epidemiological dynamics of MADV 7 and VEEV, as well as additional alphaviruses known to circulate in the region in order to 8 understand MADV emergence. For this, cross-sectional serosurveys were used to 9 demonstrate that the Alphaviruses MADV, VEEV and Una virus have repeatedly infected 10 humans in eastern Panama over the past five decades. Whilst their historical transmission 11 has been low, we confirm that the transmission has recently increased for both MADV and 12 VEEV. 13 14 15 Abstract 16 Background. Madariaga virus (MADV), has recently been associated with severe human 17 disease in Panama, where the closely related Venezuelan equine encephalitis virus (VEEV) 18 also circulates. In June, 2017, a fatal MADV infection was confirmed in a community of 19 Darien province. 20 Methods. We conducted a cross-sectional outbreak investigation with human and 21 mosquito collections in July 2017, where sera were tested for alphavirus antibodies and 22 viral RNA. Additionally, by applying a catalytic, force-of-infection statistical model to two 23 serosurveys from Darien province in 2012 and 2017, we investigated whether endemic or 24 epidemic alphavirus transmission occurred historically. 25 Results. In 2017, MADV and VEEV IgM seroprevalence was 1.6% and 4.4%, respectively; IgG 26 antibody prevalences were MADV: 13.2%; VEEV: 16.8%; Una virus (UNAV): 16.0%; and 27 Mayaro virus (MAYV): 1.1%. Active viral circulation was not detected. Evidence of MADV 28 and UNAV infection was found near households -raising questions about its vectors and 29 enzootic transmission cycles. Insomnia was associated with MADV and VEEV infection, 30 depression symptoms were associated with MADV, and dizziness with VEEV and UNAV. 31 Force-of-infection analyses suggest endemic alphavirus transmission historically, with 32 recent increased human exposure to MADV and VEEV in some regions. 33 Conclusions. The lack of additional neurological cases suggest that severe MADV and VEEV 34 infections occur only rarely. Our results indicate that, over the past five decades, 35 alphavirus infections have occurred at low levels in eastern Panama, but that MADV and 36 VEEV infections have recently increased -potentially during the past decade. Endemic 37 infections and outbreaks of MADV and VEEV appear to differ spatially. 38 39

The Costa Rican pygmy rice rat (Oligoryzomys costaricensis) is the primary reservoir of Choclo orthohantavirus (CHOV), the causal agent of hantavirus disease, pulmonary syndrome, and fever in humans in Panama. Since the emergence of CHOV in early 2000, we have systematically sampled and archived rodents from >150 sites across Panama to establish a baseline understanding of the host and virus, producing a permanent archive of holistic specimens that we are now probing in greater detail. We summarize these collections and explore preliminary habitat/virus associations to guide future wildlife surveillance and public health efforts related to CHOV and other zoonotic pathogens. Host sequences of the mitochondrial cytochrome b gene form a single monophyletic clade in Panama, despite wide distribution across Panama. Seropositive samples were concentrated in the central region of western Panama, consistent with the ecology of this agricultural commensal and the higher incidence of CHOV in humans in that region. Hantavirus seroprevalence in the pygmy rice rat was >15% overall, with the highest prevalence in agricultural areas (21%) and the lowest prevalence in shrublands (11%). Host–pathogen distribution, transmission dynamics, genomic evolution, and habitat affinities can be derived from the preserved samples, which include frozen tissues, and now provide a foundation for expanded investigations of orthohantaviruses in Panama.