Cemeteries as sources of Aedes aegypti and other mosquito species in southeastern Puerto Rico.

OBJECTIVE: To investigate the presence and abundance of mosquito species in containers found in different types of cemeteries in Puerto Rico to assess their importance and make control recommendations. METHODS: We conducted surveys of containers with water in 16 cemeteries in southeastern Puerto Rico to detect the presence of larvae and pupae of Aedes aegypti and other mosquitoes; to identify the most common and productive containers and to study their variation in relation to the type of cemetery. RESULTS: The most common containers with water were flowerpots, followed in abundance by a variety of discarded containers and open tombs. We found a positive relationship between density of containers with water and rainfall. There was a rich community of mosquito species developing in containers of the inspected cemeteries: nine mosquito species belonging to four genera with Ae. aegypti and Ae. mediovittatus being the most frequent and abundant. We sampled 13 cement-type cemeteries, 2 mixed and only 1 lawn cemetery, consequently, we could not draw any conclusion regarding container productivity and cemetery type. CONCLUSIONS: Surveyed cemeteries were important sources of Ae. aegypti and other mosquitoes in flowerpots, discarded containers and open tombs. We recommend conducting further studies to establish how frequently inspections should occur; and mosquito control by emptying aquatic habitats and larviciding to reduce mosquito productivity and protect workers and visitors from mosquito bites and possible transmission of arboviruses.

Continuation of Mosquito Surveillance and Control During Public Health Emergencies and Natural Disasters.

Mosquitoborne disease outbreaks occur every year in the United States from one or more of the arboviral diseases dengue, West Nile, LaCrosse, Eastern equine encephalitis, and Zika (1). Public opinion communicated through traditional and social media and the Internet, competing public health and resource priorities, and local conditions can impede the ability of vector control organizations to prevent and respond to outbreaks of mosquitoborne disease. The Environmental Protection Agency (EPA) and CDC performed a coordinated review of the concerns and challenges associated with continuation of mosquito surveillance and control during public health emergencies and disasters. This report highlights the first joint recommendation from EPA and CDC. Mosquito surveillance and control should be maintained by state and local mosquito control organizations to the extent that local conditions and resources will allow during public health emergencies and natural disasters. Integrated pest management (IPM) is the best approach for mosquito control (2). IPM uses a combination of methods, including both physical and chemical means of control (3). For chemical means of control, CDC and EPA recommend the use of larvicides and adulticides following the EPA label. It is imperative that public health recommendations be followed to ensure the safety of the pesticide applicator and the public.

EphrinB2/EphB4 signaling regulates non-sprouting angiogenesis by VEGF.

Vascular endothelial growth factor (VEGF) is the master regulator of angiogenesis, whose best-understood mechanism is sprouting. However, therapeutic VEGF delivery to ischemic muscle induces angiogenesis by the alternative process of intussusception, or vascular splitting, whose molecular regulation is essentially unknown. Here, we identify ephrinB2/EphB4 signaling as a key regulator of intussusceptive angiogenesis and its outcome under therapeutically relevant conditions. EphB4 signaling fine-tunes the degree of endothelial proliferation induced by specific VEGF doses during the initial stage of circumferential enlargement of vessels, thereby limiting their size and subsequently enabling successful splitting into normal capillary networks. Mechanistically, EphB4 neither inhibits VEGF-R2 activation by VEGF nor its internalization, but it modulates VEGF-R2 downstream signaling through phospho-ERK1/2. In vivo inhibitor experiments show that ERK1/2 activity is required for EphB4 regulation of VEGF-induced intussusceptive angiogenesis. Lastly, after clinically relevant VEGF gene delivery with adenoviral vectors, pharmacological stimulation of EphB4 normalizes dysfunctional vascular growth in both normoxic and ischemic muscle. These results identify EphB4 as a druggable target to modulate the outcome of VEGF gene delivery and support further investigation of its therapeutic potential.

Rapid Screening of Aedes aegypti Mosquitoes for Susceptibility to Insecticides as Part of Zika Emergency Response, Puerto Rico.

In response to the 2016 Zika outbreak, Aedes aegypti mosquitoes from 38 locations across Puerto Rico were screened using Centers for Disease Control and Prevention bottle bioassays for sensitivity to insecticides used for mosquito control. All populations were resistant to pyrethroids. Naled, an organophosphate, was the most effective insecticide, killing all mosquitoes tested.

Differences in Prevalence of Symptomatic Zika Virus Infection, by Age and Sex-Puerto Rico, 2016.

Background: During the outbreak of Zika virus (ZIKV) disease in Puerto Rico in 2016, nonpregnant women aged 20-39 years were disproportionately identified with ZIKV disease. We used household-based cluster investigations to determine whether this disparity was associated with age- or sex-dependent differences in the rate of ZIKV infection or reported symptoms. Methods: Participation was offered to residents of households within a 100-m radius of the residences of a convenience sample of 19 laboratory-confirmed ZIKV disease cases. Participants answered a questionnaire and provided specimens for diagnostic testing by reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). Results: Among 367 study participants, 114 (31.1%) were laboratory positive for ZIKV infection, of whom 30% reported a recent illness (defined as self-reported rash or arthralgia) attributable to ZIKV infection. Age and sex were not associated with ZIKV infection. Female sex (adjusted prevalence ratio [aPR], 2.28; 95% confidence interval [CI], 1.40, 3.67), age <40 years (aPR, 2.39; 95% CI, 1.55, 3.70), and asthma (aPR, 1.63; 95% CI, 1.12, 2.37) were independently associated with symptomatic infection. Conclusions: Although neither female sex nor age were associated with an increased prevalence of ZIKV infection, both were associated with symptomatic infection. Further investigation to identify a potential mechanism of age- and sex-dependent differences in reporting symptomatic ZIKV infection is warranted.

Long-lasting fibrin matrices ensure stable and functional angiogenesis by highly tunable, sustained delivery of recombinant VEGF164.

Clinical trials of therapeutic angiogenesis by vascular endothelial growth factor (VEGF) gene delivery failed to show efficacy. Major challenges include the need to precisely control in vivo distribution of growth factor dose and duration of expression. Recombinant VEGF protein delivery could overcome these issues, but rapid in vivo clearance prevents the stabilization of induced angiogenesis. Here, we developed an optimized fibrin platform for controlled delivery of recombinant VEGF, to robustly induce normal, stable, and functional angiogenesis. Murine VEGF164 was fused to a sequence derived from α2-plasmin inhibitor (α2-PI1-8) that is a substrate for the coagulation factor fXIIIa, to allow its covalent cross-linking into fibrin hydrogels and release only by enzymatic cleavage. An α2-PI1-8-fused variant of the fibrinolysis inhibitor aprotinin was used to control the hydrogel degradation rate, which determines both the duration and effective dose of factor release. An optimized aprotinin-α2-PI1-8 concentration ensured ideal degradation over 4 wk. Under these conditions, fibrin-α2-PI1-8-VEGF164 allowed exquisitely dose-dependent angiogenesis: concentrations ≥25 µg/mL caused widespread aberrant vascular structures, but a 500-fold concentration range (0.01-5.0 µg/mL) induced exclusively normal, mature, nonleaky, and perfused capillaries, which were stable after 3 mo. Optimized delivery of fibrin-α2-PI1-8-VEGF164 was therapeutically effective both in ischemic hind limb and wound-healing models, significantly improving angiogenesis, tissue perfusion, and healing rate. In conclusion, this optimized platform ensured (i) controlled and highly tunable delivery of VEGF protein in ischemic tissue and (ii) stable and functional angiogenesis without introducing genetic material and with a limited and controllable duration of treatment. These findings suggest a strategy to improve safety and efficacy of therapeutic angiogenesis.

Reduced Incidence of Chikungunya Virus Infection in Communities with Ongoing Aedes Aegypti Mosquito Trap Intervention Studies - Salinas and Guayama, Puerto Rico, November 2015-February 2016.

Aedes species mosquitoes transmit chikungunya virus, as well as dengue and Zika viruses, and bite most often during the day.* Infectious mosquito bites frequently occur in and around homes (1,2). Caribbean countries first reported local transmission of chikungunya virus in December 2013, and soon after, chikungunya virus spread throughout the Americas (3). Puerto Rico reported its first laboratory-positive chikungunya case in May 2014 (4), and subsequently identified approximately 29,000 suspected cases throughout the island by the end of 2015.(†) Because conventional vector control approaches often fail to result in effective and sustainable prevention of infection with viruses transmitted by Aedes mosquitoes (5), and to improve surveillance of mosquito population densities, CDC developed an Autocidal Gravid Ovitrap (AGO) (6) to attract and capture the female Aedes aegypti mosquitoes responsible for transmission of infectious agents to humans (Figure). The AGO trap is a simple, low-cost device that requires no use of pesticides and no servicing for an extended period of time (6).

Operational Aspects of the Centers for Disease Control and Prevention Autocidal Gravid Ovitrap.

Dengue viruses cause hundreds of millions of infections every year in tropical and subtropical countries. Unfortunately, there is not a single universal vector control method capable of suppressing Aedes aegypti (L.) populations. Amongst novel control tools or approaches are various types of traps targeting gravid females or their eggs. Here, we provide details of the operational use of the Centers for Disease Control and Prevention autocidal gravid ovitrap (CDC-AGO trap) for the surveillance and control of Ae. aegypti. Adult mosquitoes were monitored every week in 2 isolated neighborhoods treated with 3 AGO traps per house in 85% of houses and in 2 reference neighborhoods without control traps. Between March 2013 and April 2015 we serviced the AGO traps 14 times in each community (every 2 months). Common trap problems were absent or broken trap tops (1-1.5%), flooded (0.1-0.7%) or dry (0.5-1.3%) traps, and missing (0.3-0.8%) or vandalized (0.5-1.4%) traps. Most traps kept a volume of infusion between 45% and 97% of their original volume (10 liters). Nontarget organisms captured in AGO traps were mostly small flies, and to a lesser extent ants, cockroaches, grasshoppers, butterflies, dragonflies, and lizards. Trap coverage ranged between 83% and 87% of houses in both communities throughout the study. We interpret such high levels of trap retention over time as an expression of acceptance by the community.

Evaluation of Household Bleach as an Ovicide for the Control of Aedes aegypti.

Accumulations of dormant eggs in container habitats allow Aedes aegypti populations to survive harsh environmental conditions and may frustrate control interventions directed at larval and adult life stages. While sodium hypochlorite solutions (NaOCl) have long been recognized as ovicides for use against dengue vectors, the susceptibility of eggs to spray applications has not been robustly evaluated on substrate materials representative of the most frequently utilized artificial container habitats. Experiments were performed under controlled and natural conditions by applying dilutions of household bleach (52.5 ppt NaOCl) as a spray to eggs on plastic, rubber, and concrete surfaces, with and without a smectite clay thickener. Laboratory assays identified the minimum NaOCl concentrations required to eliminate eggs on plastic (10 ppt), rubber (20 ppt) and concrete (20 ppt) surfaces. Addition of smectite clay reduced the minimum effective concentration to 10 ppt NaOCl for all 3 substrates. A minimum exposure period of 24 h was required to completely eliminate egg viability on concrete surfaces, even at the highest NaOCl concentration (52.5 ppt). Field experiments verified that spray application of a 1∶3 dilution of household bleach mixed with smectite clay can reduce egg hatching by ≥ 99% in shaded and sun-exposed plastic containers. Similarly, 4∶1 dilution of household bleach (with or without smectite clay) eliminated ≥ 98% of eggs from concrete surfaces in outdoor, water-filled drums. In this study, we propose a practical, effective and safe strategy for using household bleach to eliminate Ae. aegypti eggs in a range of artificial container habitats.

Split for the cure: VEGF, PDGF-BB and intussusception in therapeutic angiogenesis.

Therapeutic angiogenesis is an attractive strategy to treat patients suffering from ischaemic conditions and vascular endothelial growth factor-A (VEGF) is the master regulator of blood vessel growth. However, VEGF can induce either normal or aberrant angiogenesis depending on its dose localized in the microenvironment around each producing cell in vivo and on the balanced stimulation of platelet-derived growth factor-BB (PDGF-BB) signalling, responsible for pericyte recruitment. At the doses required to induce therapeutic benefit, VEGF causes new vascular growth essentially without sprouting, but rather through the alternative process of intussusception, or vascular splitting. In the present article, we briefly review the therapeutic implications of controlling VEGF dose on one hand and pericyte recruitment on the other, as well as the key features of intussusceptive angiogenesis and its regulation.

Use of the CDC autocidal gravid ovitrap to control and prevent outbreaks of Aedes aegypti (Diptera: Culicidae).

Populations ofAedes aegypti (L.) can be managed through reductions in adult mosquito survival, number of offspring produced, or both. Direct adult mortality can be caused by the use of space sprays or residual insecticides to mosquito resting sites, and with a variety of residual insecticide-impregnated surfaces that are being tested, such as curtains, covers for water-storage vessels, bednets, and ovitraps. The fertility ofAe. aegypti populations can be reduced by the use of autocidal oviposition cups that prevent the development of mosquitoes inside the trap by mechanical means or larvicides, as well as by releasing sterile, transgenic, and para-transgenic mosquitoes. Survival and fertility can be simultaneously reduced by capturing gravid female Ae. aegypti with sticky gravid traps. We tested the effectiveness of the novel Centers for Disease Control and Prevention autocidal gravid ovitrap (CDC-AGO trap) to control natural populations ofAe. aegypti under field conditions in two isolated urban areas (reference vs. intervention areas) in southern Puerto Rico for 1 yr. There were significant reductions in the captures of female Ae. aegypti (53-70%) in the intervention area The presence of three to four AGO control traps per home in 81% of the houses prevented outbreaks of Ae. aegypti, which would be expected after rains. Mosquito captures in BG-Sentinel and AGO traps were significantly and positively correlated, showing that AGO traps are useful and inexpensive mosquito surveillance devices. The use of AGO traps to manage Ae. aegypti populations is compatible with other control means such as source reduction, larviciding, adulticiding, sterile insect techniques, induced cytoplasmic incompatibility, and dominant lethal gene systems.

Biological control of mosquitoes in scrap tires in Brownsville, Texas, USA and Matamoros, Tamaulipas, Mexico.

Dengue periodically circulates in southern Texas and neighboring Tamaulipas, Mexico; thus, a closer examination of human and vector ecology at the northern limits of North American transmission may improve prevention activities. Scrap tires produce large mosquito populations and increase the risk of dengue transmission. Some households choose not to pay tire disposal fees, and many tires are illegally dumped in residential areas. Biological control may provide low-cost and environmentally friendly mosquito control. This pilot study evaluated the ability of Mesocyclops longisetus to reduce mosquito populations in existing residential scrap tire piles. Mosquito populations were measured by the number of all mosquito pupae within tires or adult Aedes aegypti and Ae. albopictus near piles. Mesocyclops longisetus treated piles did not significantly reduce total mosquito pupae (P = 0.07) in Matamoros, Mexico. The study also evaluated the efficacy of native Toxorhynchites moctezuma which preferentially colonized tire piles under vegetation cover in Brownsville, TX. Toxorhynchites moctezuma larvae significantly reduced total mosquito pupae, but the strength of control diminished over time.

Historical Hot Spots of Dengue and Zika Viruses to Guide Targeted Vector Control in San Juan, Puerto Rico (2010-2022).

Dengue viruses (DENV) continue to cause large outbreaks in tropical countries, while chikungunya and Zika (ZIKV) viruses have added complexity to Aedes-borne disease prevention and control efforts. Because these viruses are transmitted by the same vectors in urban areas, it is useful to understand if sequential outbreaks caused by these viruses have commonalities, such as similar seasonal and spatial patterns, that would help anticipate and perhaps prevent future outbreaks. We explored and analyzed the heterogeneity of confirmed cases of DENV (2010-2014 and 2015-2022) and ZIKV (2016-2017) during outbreaks in the San Juan metropolitan area of Puerto Rico to explore their degree of overlap and prioritize areas for Aedes aegypti control. Deidentified, georeferenced case data were aggregated into grid cells (500 × 500 m) within a geographical information system of the study area and analyzed to calculate the degree of overlap between outbreaks. Spatial autocorrelations using local indicators of spatial associations were conducted to identify significant disease case hot spots and correlations between outbreaks. We found that 75% of cases during the three transmission periods were concentrated in 25% of the total number of grid cells covering the study area. We also found significant clustering of cases during each outbreak, enabling identification of consistent disease hot spots. Our results showed 85% spatial overlap between cases of ZIKV in 2015-2017 and DENV in 2010-2014 and 97% overlap between DENV cases in 2010-2014 and 2015-2022. These results reveal urban areas at greater risk of future arbovirus outbreaks that should be prioritized for vector control.

VEGF over-expression in skeletal muscle induces angiogenesis by intussusception rather than sprouting.

Therapeutic over-expression of vascular endothelial growth factor (VEGF) can be used to treat ischemic conditions. However, VEGF can induce either normal or aberrant angiogenesis depending on its dose in the microenvironment around each producing cell in vivo, which limits its clinical usefulness. The goal herein was to determine the cellular mechanisms by which physiologic and aberrant vessels are induced by over-expression of different VEGF doses in adult skeletal muscle. We took advantage of a well-characterized cell-based platform for controlled gene expression in skeletal muscle. Clonal populations of retrovirally transduced myoblasts were implanted in limb muscles of immunodeficient mice to homogeneously over-express two specific VEGF(164) levels, previously shown to induce physiologic and therapeutic or aberrant angiogenesis, respectively. Three independent and complementary methods (confocal microscopy, vascular casting and 3D-reconstruction of serial semi-thin sections) showed that, at both VEGF doses, angiogenesis took place without sprouting, but rather by intussusception, or vascular splitting. VEGF-induced endothelial proliferation without tip-cell formation caused an initial homogeneous enlargement of pre-existing microvessels, followed by the formation of intravascular transluminal pillars, hallmarks of intussusception. This was associated with increased flow and shear stress, which are potent triggers of intussusception. A similar process of enlargement without sprouting, followed by intussusception, was also induced by VEGF over-expression through a clinically relevant adenoviral gene therapy vector, without the use of transduced cells. Our findings indicate that VEGF over-expression, at doses that have been shown to induce functional benefit, induces vascular growth in skeletal muscle by intussusception rather than sprouting.

Therapeutic angiogenesis due to balanced single-vector delivery of VEGF and PDGF-BB.

Therapeutic angiogenesis by delivery of vascular growth factors is an attractive strategy for treating debilitating occlusive vascular diseases, yet clinical trials have thus far failed to show efficacy. As a result, limb amputation remains a common outcome for muscle ischemia due to severe atherosclerotic disease, with an overall incidence of 100 per million people in the United States per year. A challenge has been that the angiogenic master regulator vascular endothelial growth factor (VEGF) induces dysfunctional vessels, if expressed outside of a narrow dosage window. We tested the hypothesis that codelivery of platelet-derived growth factor-BB (PDGF-BB), which recruits pericytes, could induce normal angiogenesis in skeletal muscle irrespective of VEGF levels. Coexpression of VEGF and PDGF-BB encoded by separate vectors in different cells or in the same cells only partially corrected aberrant angiogenesis. In marked contrast, coexpression of both factors in every cell at a fixed relative level via a single bicistronic vector led to robust, uniformly normal angiogenesis, even when VEGF expression was high and heterogeneous. Notably, in an ischemic hindlimb model, single-vector expression led to efficient growth of collateral arteries, revascularization, increased blood flow, and reduced tissue damage. Furthermore, these results were confirmed in a clinically applicable gene therapy approach by adenoviral-mediated delivery of the bicistronic vector. We conclude that coordinated expression of VEGF and PDGF-BB via a single vector constitutes a novel strategy for harnessing the potency of VEGF to induce safe and efficacious angiogenesis.

A novel autocidal ovitrap for the surveillance and control of Aedes aegypti.

We describe an inexpensive autocidal ovitrap for Aedes aegypti that uses cross-linked polyacrylamide (PAM) gel as the oviposition substrate. Aedes aegypti females readily laid eggs on PAM gel that had been hydrated with either hay infusion or water. Aedes aegypti larvae that hatched from their eggs desiccated on the surface of the PAM gel. We tested the effects of gel hydration, texture, and type of attractant on trap performance, and compared the capture rates of standard ovitraps with those of PAM gel ovitraps in the field. The results showed that the number of eggs did not vary over a range of gel hydration levels (40-100%) and that more eggs were recovered from ovitraps containing coarse gel than from those containing homogenized gel. The PAM gel hydrated with hay infusion was more attractive to gravid female mosquitoes than gel hydrated with water. In the field, the number of eggs recovered from autocidal ovitraps with PAM gel was similar to that recovered from standard ovitraps with hay infusion.

An improved trap to capture adult container-inhabiting mosquitoes.

Although dengue viruses are thought to be transmitted by Aedes aegypti in Puerto Rico, Aedes mediovittatus, the Caribbean tree hole mosquito, is also a potential vector. This species is native to the Greater Antilles and has been shown to be a competent vector of dengue viruses in the laboratory. Consequently, it has been suggested that Ae. mediovittatus could be acting as a secondary vector or virus reservoir. This study was part of an ongoing investigation into this, and it aimed to determine whether BG-Sentinel traps (BGS traps) could be used to collect adults of this mosquito and could be modified to increase the number of captures of this species in the field. We conducted experiments to test the relative attractiveness of BGS traps to Ae. mediovittatus and Ae. aegypti and explored the effects of chemical lures (BG-Lure, CO2, octenol) and optical properties (color, size) on the capture rates of BGS traps in a large, outdoor cage in San Juan city, Puerto Rico. We also conducted field tests to compare modified BGS traps with the original traps in a rural community in Patillas municipality, Puerto Rico. Results obtained from the large, outdoor cage experiments indicated that trap captures of both mosquito species could be significantly enhanced by using black instead of white BGS traps combined with BG-Lure. Field experiments revealed that the modified traps captured a significantly greater number of Ae. aegypti, Ae. mediovittatus, and Culex quinquefasciatus, with greater sensitivity for the latter 2 species, and also captured a larger number of mosquito species and a smaller ratio of Ae. aegypti to Ae. mediovittatus, with greater than expected species co-occurrences.

El Niño Southern Oscillation (ENSO) effects on local weather, arboviral diseases, and dynamics of managed and unmanaged populations of Aedes aegypti (Diptera: Culicidae) in Puerto Rico.

We investigated the effects of interannual El Niño Southern Oscillation (ENSO) events on local weather, Aedes aegypti populations, and combined cases of dengue (DENV), chikungunya (CHIKV), and Zika (ZIKV) viruses in 2 communities with mass mosquito trapping and 2 communities without mosquito control in southern Puerto Rico (2013-2019). Gravid adult Ae. aegypti populations were monitored weekly using Autocidal Gravid Ovitraps (AGO traps). Managing Ae. aegypti populations was done using 3 AGO traps per home in most homes. There were drought conditions in 2014-2015 concurrent with the emergence of a strong El Niño (2014-2016), wetter conditions during La Niña (2016-2018), a major hurricane (2017), and a weaker El Niño (2018-2019). The main factor explaining differences in Ae. aegypti abundance across sites was mass trapping. Populations of Ae. aegypti reached maximum seasonal values during the wetter and warmer months of the year when arbovirus epidemics occurred. El Niño was significantly associated with severe droughts that did not impact the populations of Ae. aegypti. Arbovirus cases at the municipality level were positively correlated with lagged values (5-12 mo.) of the Oceanic El Niño Index (ONI), droughts, and abundance of Ae. aegypti. The onset of strong El Niño conditions in Puerto Rico may be useful as an early warning signal for arboviral epidemics in areas where the abundance of Ae. aegypti exceeds the mosquito density threshold value.

FACS-purified myoblasts producing controlled VEGF levels induce safe and stable angiogenesis in chronic hind limb ischemia.

We recently developed a method to control the in vivo distribution of vascular endothelial growth factor (VEGF) by high throughput Fluorescence-Activated Cell Sorting (FACS) purification of transduced progenitors such that they homogeneously express specific VEGF levels. Here we investigated the long-term safety of this method in chronic hind limb ischemia in nude rats. Primary myoblasts were transduced to co-express rat VEGF-A(164) (rVEGF) and truncated ratCD8a, the latter serving as a FACS-quantifiable surface marker. Based on the CD8 fluorescence of a reference clonal population, which expressed the desired VEGF level, cells producing similar VEGF levels were sorted from the primary population, which contained cells with very heterogeneous VEGF levels. One week after ischemia induction, 12 × 10(6) cells were implanted in the thigh muscles. Unsorted myoblasts caused angioma-like structures, whereas purified cells only induced normal capillaries that were stable after 3 months. Vessel density was doubled in engrafted areas, but only approximately 0.1% of muscle volume showed cell engraftment, explaining why no increase in total blood flow was observed. In conclusion, the use of FACS-purified myoblasts granted the cell-by-cell control of VEGF expression levels, which ensured long-term safety in a model of chronic ischemia. Based on these results, the total number of implanted cells required to achieve efficacy will need to be determined before a clinical application.

Vertebrate hosts of Aedes aegypti and Aedes mediovittatus (Diptera: Culicidae) in rural Puerto Rico.

The distribution of Aedes (Stegomyia) aegypti (L.), the main vector of dengue viruses (DENV) worldwide, overlaps with Aedes (Gymnometopa) mediovittatus (Coquillett), the Caribbean treehole mosquito, in urban, suburban, and rural areas. Ae. mediovittatus is a competent vector of DENV with high rates of vertical DENV transmission in the laboratory. This study determined whether Ae. mediovittatus feeds on humans and compared its feeding patterns with co-occurring Ae. aegypti in two rural communities of Puerto Rico. Adult mosquitoes were captured for three consecutive days every week from July 2009 to May 2010 using BG-Sentinel traps with skin lures that were placed in the front yard of houses in both communities. Three methods were used to identify the 756 bloodmeals obtained in this study: a multiplex polymerase chain reaction (PCR) for humans and dogs targeting cytochrome b; a PCR targeting the 16S rRNA; and a nested PCR targeting cytochrome b. Ae. mediovittatus fed mostly on humans (45-52%) and dogs (28-32%) but also on cats, cows, horses, rats, pigs, goats, sheep, and chickens. Ae. aegypti fed mostly on humans (76-79%) and dogs (18-21%) but also on cats, horses, and chickens. Our results indicate that Ae. mediovittatus may have a relatively high rate of vector-human contact, which might facilitate virus transmission or harborage in rural areas of Puerto Rico.