Seasonal Dynamics of Mosquito-Borne Viruses in the Southwestern Florida Everglades, 2016, 2017.

Mosquitoes were collected for 12 consecutive months beginning June 2016, from 11 locations in the Florida Everglades, Collier County, and tested for viruses by isolation in Vero cells and subsequent identification. One species complex and 31 species of mosquitoes were identified from 668,809 specimens. Ochlerotatus taeniorhynchus comprised 72.2% of the collection. Other notable species were Anopheles crucians complex, Culex nigripalpus, Cx. erraticus, and Cx. cedecei. Seven species of virus were identified from 110 isolations: Everglades, Gumbo Limbo, Mahogany Hammock, Pahayokee, Shark River, Tensaw, and West Nile viruses. Everglades, West Nile, Tensaw, and Mahogany Hammock viruses were most frequently isolated. Largest numbers of viruses were identified from Cx. cedecei, Cx. nigripalpus, and An. crucians complex. Five species of virus were isolated from Cx. cedecei. Viruses were isolated from mangrove, cypress swamp, hardwood hammock, and sawgrass habitats. West Nile virus was isolated August through October when Cx. nigripalpus was most abundant. Everglades virus was the most frequently isolated virus from nine species of mosquitoes collected from June through August. Tensaw virus was isolated primarily from Anopheles species. Isolations were made in July, August, January, February, and April, suggesting that this virus may be present in host-seeking mosquitoes throughout the year. Mahogany Hammock, Shark River, Gumbo Limbo, and Pahayokee viruses were isolated primarily from Cx. cedecei from June through December. Shotgun metagenomic sequencing was used to document that seven pools of Cx. cedecei were infected with two arboviruses. As communities expand into the Everglades, more humans will become exposed to arboviruses.

Horizontal and Vertical Transmission of West Nile Virus by Aedes vexans (Diptera: Culicidae).

West Nile virus (family Flaviviridae, genus Flavivirus) first caused human and veterinary disease, and was isolated from Culex pipiens pipiens L. and Aedes vexans (Meigen) (Diptera: Culicidae) in the United States in 1999. We report that a Connecticut strain of Ae. vexans was competent to transmit West Nile virus both horizontally to suckling mice and vertically to its progeny in the laboratory. Horizontal transmission was first observed on day 6 post-exposure (pe). Daily horizontal transmission rates generally increased with the day post-virus exposure with highest rates of 67-100% recorded on days 28-30 pe. One female vertically transmitted West Nile virus on day 21 pe, but only after it had taken its third bloodmeal. Horizontal and vertical transmission may contribute to West Nile virus infection rates in Ae. vexans in summer, and vertical transmission provides a means of survival of West Nile virus during winter.

Repellency of Naturally Occurring or Related Compounds, DEET, and Para-Menthane-3,8-Diol to Bed Bugs (Hemiptera: Cimicidae).

Bed bugs, Cimex lectularius L. (Hemiptera: Cimicidae), have become a major health nuisance in the past 20 ysin cities and elsewhere throughout many areas of the world. Few studies have reported on repellent compounds that could reduce their transport in luggage. We evaluated the repellency of six naturally occurring or related compounds used in flavor/fragrance applications or structurally related compounds, para-menthane-3,8-diol, and N,N-diethyl-meta-toluamide (DEET) to bed bugs in a 183 × 183-cm arena . Repellency was assessed using soft-sided polyester lunch bags serving as surrogates of luggage and barrier cloth towels upon which rested untreated lunch bags. We report for the first time repellency of delta dodecalactone, 2-(3, 7-dimethyl-2, 6-nonadien-1-yl)-cyclopentanone (a.k.a. 'methyl apritone'), gamma dodecalactone, and para-menthane-3,8-diol to bed bugs. Propyl dihydrojasmonate, 3-methyl-5-hexyl-2-cyclohexenone, gamma methyl tridecalactone, and DEET are also documented to be repellent to bed bugs. These compounds provided relatively long-term protection. Propyl dihydrojasmonate prevented bed bugs from seeking refuge in treated lunch bags 27 d after treatment, and when applied to cloth towels repelled harborage-seeking bed bugs for 146 d. Methyl apritone blended with 3-methyl-5-hexyl-2-cyclohexenone and delta dodecalactone as an individual compound applied to cloth towels repelled bed bugs for 190 and 276 d, respectively. The above-mentioned compounds, either individually or as blends, may reduce risk of bed bugs seeking harborage in treated suitcases or towels upon which untreated luggage is placed.

Relatively Small Quantities of CO2, Ammonium Bicarbonate, and a Blend of (E)-2-Hexenal Plus (E)-2-Octenal Attract Bed Bugs (Hemiptera: Cimicidae).

Bed bugs, Cimex lectularius L., feed on humans, have increased exponentially in the past two decades in many major cities throughout the world, have caused intense infestations, and have become a significant health concern. Improved traps are needed to detect early infestations, to assess control programs, and for control of bed bugs. Carbon dioxide released alone or simultaneously with other attractants into three types of traps at the relatively low rate of 1 ml/min caught significantly more bed bugs than untreated controls in a 183- by 183-cm arena. This finding may enable CO2 to be used more economically in traps. Three percent ammonium bicarbonate released at a rate of ≤0.03 ml/h also caught significantly more bed bugs than untreated controls. A blend of (E)-2-hexenal and (E)-2-octenal at concentrations of 0.025% or 0.1% each and released at 0.02 ml/h attracted significantly more bed bugs than untreated controls. These findings ought to improve detection of bed bugs.

Arboviruses in North Dakota, 2003-2006.

To investigate arbovirus transmission in North Dakota, we collected and screened mosquitoes for viral infection by Vero cell culture assay. Seven viruses were isolated from 13 mosquito species. Spatial and temporal distributions of the important vectors of West Nile virus (WNV), Cache Valley virus, Jamestown Canyon virus (JCV), and trivittatus virus are reported. Snowshoe hare virus, Potosi virus, and western equine encephalomyelitis virus were also isolated. The risks of Culex tarsalis and Aedes vexans transmitting WNV to humans were 61.4% and 34.0% in 2003-2006, respectively, but in 2003 when the largest epidemic was reported, risks for Ae. vexans and Cx. tarsalis in Cass County were 73.6% and 23.9%, respectively. Risk of humans acquiring an infectious bite was greatest from about the second week of July through most of August. West Nile virus sequences were of the WN02 genotype. Most JCV strains belonged to a single clade of genetically related strains. Cache Valley virus and JCV were prevalent during August and early September and during July and August, respectively.

Relating the progeny production curve to the speed of an epidemic.

The dependence of the initial infection rate, r, on the basic reproductive number, R(0), and the temporal moments of the progeny production curve are examined. A solution to the linearized Kermack-McKendrick equation is presented and used to analyze a variety of theoretical models of pathogen reproduction. The solution yields a relation between r and the basic reproductive number, R(0); the mean time between pathogen generations, µ; and the standard deviation about this mean, σ. A transformation using the dimensionless variables rµ and rσ is introduced, which maps the solution onto a one-dimensional curve. An approximation for the value of r in terms of R(0) and the first four temporal moments of the reproductive curve is derived. This allows direct comparison of epidemics resulting from theoretical models with those generated using experimentally obtained reproduction curves. For epidemics characterized by a value of rµ < 5, the value of r is well determined (<2%) by this fourth-order expansion regardless of the functional form of the reproduction curve.

Horizontal and vertical transmission of West Nile virus genotype NY99 by Culex salinarius and genotypes NY99 and WN02 by Culex tarsalis.

Culex tarsalis is a superior horizontal and vertical vector of West Nile virus (WNV) compared with Culex salinarius. Culex salinarius transmitted WNV genotype NY99 (CT 2741-99 strain) horizontally to suckling mice at significantly lower rates than Cx. tarsalis on Days 8, 9, 10, and 12 post-infection, and Cx. salinarius transmitted WNV genotype NY99 to offspring at a lower vertical transmission infection rate than Cx. tarsalis. Culex tarsalis transmitted WNV genotypes NY99 and WN02 (CT S0084-08 strain) with equal efficiency. Daily percent horizontal transmission of genotype NY99 by Cx. tarsalis-infected per os and by intra-thoracic infection was not significantly different from daily transmission of genotype WN02 from Days 5-23 and Days 2-9 post-infection, respectively. Our findings do not support the previously published hypothesis that genotype NY99 was replaced in the New World by WN02 because of a shorter extrinsic incubation of WN02.

Aedes japonicus japonicus and associated woodland species attracted to Centers for Disease Control and Prevention miniature light traps baited with carbon dioxide and the Traptech mosquito lure.

Twelve reported mosquito attractants, alone or in combination, and 3 different types of traps were evaluated under field conditions for their attractiveness to host-seeking and oviposition-seeking female Aedes japonicus japonicus and associated woodland species in Windsor, CT, in 2010 and 2011. This study highlights the effectiveness of combining CO2 with the TrapTech Mosquito Lure in a Centers for Disease Control and Prevention (CDC) miniature light trap for collection of Ae. j. japonicus and associated woodland mammalian-feeding mosquitoes. The TrapTech Mosquito Lure is a proprietary blend of Bedoukian Research, Inc. It contained 250 mg of R-1-octen-3-ol and 1900 mg of ammonium bicarbonate, which were slowly released from a plastic disperser. On average, 567 Ae. j. japonicus individuals were collected per trap per night in the CDC miniature light traps baited with CO2 plus TrapTech Mosquito Lure. The numbers collected in this trap were 28 times and 100 times greater than the numbers of Ae. j. japonicus collected in the CDC miniature light trap baited only with CO2 and the gravid trap baited with hay infusion, 2 commonly used traps to assess abundance of Ae. j. japonicus. The average catches of other mammalian-biting species, Ae. cinereus, Ae. triseriatus, Ae. trivittatus, Ae. vexans, Anopheles punctipennis, An. quadrimaculatus, Coquillettidia perturbans, and Culex salinarius, were all significantly greater in the CDC miniature light trap baited with CO2 plus TrapTech Mosquito Lure than in traps with CO2 alone, but their average numbers were not as large as were those of Ae. j. japonicus. These data demonstrate that the TrapTech Mosquito Lure used in combination with CO2 in a CDC miniature light trap has potential to be a versatile and simple surveillance method for Ae. j. japonicus and other species.

Control of mosquitoes in catch basins in Connecticut with Bacillus thuringiensis israelensis, Bacillus sphaericus, [corrected] and spinosad.

Catch basins are a major source of Culex pipiens pipiens, Cx. restuans, and Aedes japonicus in northeastern USA. VectoBac CG (Bacillus thuringiensis israelensis [Bti]), VectoLex CG (Bacillus sphaericus [Bs]), and VectoBac 12AS (Bti), each applied at maximum label rate of 1.8 g, 1.8 g, and 0.193 ml per catch basin, respectively, significantly reduced the numbers of larvae for 1 wk. The dosages on the labels for treatment of mosquito larvae in catch basins, where mosquito breeding is continuous, are not adequate for providing long-term control in the northeastern USA without the need for frequent retreatment. When applied at 3 times the maximum label rate, VectoLex CG, VectoBac 12AS, and VectoBac CG significantly reduced the numbers of larvae for 5, 4, and 2 wk, respectively. A single application of VectoMax WSP (Bti + Bs) (1 pouch containing 10 g) per catch basin significantly reduced the numbers of 3rd and 4th instars and healthy pupae in catch basins in 2008, but numbers of 3rd and 4th instars in treated catch basins at 21 days after treatment had increased to 40% of the numbers in untreated catch basins. A 2nd treatment of 1 pouch per catch basin reduced the numbers of 3rd and 4th instars and healthy pupae to near zero for the next 4 wk, into the middle of September 2008. In 2009, VectoMax applied as 1 pouch per catch basin on July 1 and again on August 18 significantly reduced the numbers of healthy pupae throughout the summer until the end of September. A 2nd application of VectoMax to catch basins is likely needed during summer, when rainfall averages 13.7 in. (approximately 34.25 cm) during June through September, to keep the numbers of Culex and Ae. japonicus significantly reduced to lower risk of human exposure to West Nile virus. The application of 1 Natular XRT tablet, each weighing approximately 40.5 g (6.25% spinosad), to individual catch basins in 2009 significantly reduced the total numbers of larvae for 5 wk.

Suppression of Verticillium Wilt of Eggplant by Earthworms.

The role of earthworms in plant disease has received little attention. To address whether earthworms would affect the severity of Verticillium wilt of eggplant (Solanum melongena) in the field, we grew eggplants in experimental field plots that were naturally infested with Verticillium dahliae in 2005, 2006, and 2007. Three earthworm treatments were compared: (i) no treatment (untreated control), (ii) earthworm populations reduced via chemical eradicants (carbaryl or hot mustard) (reduced treatment), and (iii) earthworm populations increased by addition of adult Canadian nightcrawlers (Lumbricus terrestris, 11 earthworms per m2) (augmented treatment). Compared to the untreated control, the estimates of the area under the disease progress curve (AUDPC) were reduced while estimates of the canopy growth curve (CGC) and the final plant weights were increased in plots augmented with earthworms in all 3 years. In 2 out of 3 years, eggplant yield (weight and number of fruit) was increased in plots augmented with earthworms. When a carbaryl drench was used to reduce earthworm numbers, the treatment resulted in plants with more disease than in the untreated controls in 2005. However, in 2005 and 2006, carbaryl-treated plants had larger CGC values and higher yield than in the untreated controls and were not significantly different from the augmented plots. When a hot mustard extraction procedure was used to reduce earthworm densities in 2007, plant growth, yield, and disease variables did not differ from the untreated control. Although the effects of reducing earthworms were variable and difficult to explain, our findings suggest that augmenting earthworm populations can suppress Verticillium wilt of eggplant, and strategies that increase earthworm numbers may contribute to disease suppression.

Nocturnal activity of mosquitoes (Diptera: Culicidae) in a West Nile virus focus in Connecticut.

Six species of mosquitoes (Diptera: Culicidae) were collected in sufficient numbers for analysis in segregating traps set at 2-h intervals by using CO2 and light as attractants in a West Nile virus (family Flaviviridae, genus Flavivirus, WNV) focus in Stratford, CT. The Kolmogorov-Smirnov one-sided test for two samples was used to analyze the data. Mosquito activity began shortly before sunset and continued until shortly after sunrise the next morning. All species had geometric means that were significantly higher during the 2-h period shortly after sunset compared with the 2-h collection before sunset. Species, known to be naturally infected with WNV, were often attracted to these traps in about equal numbers at 2-h intervals during an 8- to 10-h period commencing shortly after sunset. Differences of geometric means were not significant among the four or five 2-h collection periods commencing at sunset for Aedes vexans (Meigen), Culex salinarius Coquillett, and Aedes cinereus Meigen. Aedes cantator (Coquillett) had a significantly higher geometric mean for the 2-h period commencing at sunset, and Coquillettidia perturbans (Walker) was captured in significantly greater numbers during the 2-h period starting at sunset compared with periods commencing 6 h after sunset. Culex pipiens L. tended to have an activity pattern that was primarily nocturnal. Time of night, not meteorological conditions, was the most important factor in determining the nightly variation in the number of trapped mosquitoes. Parity rates of Cx. pipiens collected during specific periods of the night were not significant. In total, 39 isolations of WNV were made from seven species collected primarily during periods of total darkness. Humans are at risk of being bitten by infected mosquitoes throughout the night.

West Nile virus from female and male mosquitoes (Diptera: Culicidae) in subterranean, ground, and canopy habitats in Connecticut.

In total, 93,532 female mosquitoes (Diptera: Culicidae) were captured in traps placed in subterranean (catch basin), ground (approximately 1.5 m above ground), and canopy (approximately 7.0 m above ground) habitats in Stamford and Stratford, CT, during 2003-2005. Culex pipiens L. was the most abundant (64.8%) of the 31 species identified. Significantly greater numbers of Cx. pipiens were captured in canopy-placed mosquito magnet experimental traps, and significantly greater numbers were collected in catch basin-placed (Centers for Disease Control) CDC traps than in CDC traps placed elsewhere. Culex restuans Theobald was captured in significantly greater numbers in traps placed in catch basins. Aedes vexans (Meigen), Aedes cinereus Meigen, and Aedes cantator (Coquillett) were significantly more abundant in ground traps. In total, 429 isolations of West Nile virus (WNV) were made from seven species of mosquitoes from late June through the end of October during 2003 through 2005. Three hundred ninety-eight (92.8%) isolates were from Cx. pipiens. Others were from Cx. restuans (n = 16), Culex salinarius Coquillett (n = 5), Ae. vexans (n = 4), Ae. cantator (n = 3), Aedes triseriatus (Say) (n = 2), and Ae. cinereus (n = 1). Multiple isolates from Cx. pipiens were made each week, primarily during the later part of July through the end of September. Weekly minimum infection rates (MIRs) were lower in 2004 (highest weekly MIR = 7.1) when no human cases were reported in Connecticut in comparison with 2003 and 2005 (highest weekly MIR = 83.9) when human cases were documented. Frequencies of infected pools were significantly higher in Cx. pipiens captured in traps in the canopy and significantly higher in catch basin placed traps than in traps at ground level. The physiological age structure of Cx. pipiens captured in the canopy was significantly different from that of Cx. pipiens collected in catch basins. Invariably, Cx. pipiens captured in the canopy were nulliparous or parous with ovaries in Christophers' stage 2, whereas 58.7% of the females captured in catch basins possessed ovaries filled with mature oocytes in Christophers' stage 5. Our results suggest that females in the canopy are seeking hosts, and after digestion of the bloodmeal and development of mature oocytes, they descend to catch basins for shelter and deposition of eggs. WNV was isolated from three, one, and two pools of male Cx. pipiens captured in catch basin-, ground-, and canopy-placed traps, respectively, and from six nulliparous Cx. pipiens females collected in the canopy. Weekly MIR ranged from 1.2 to 31.1 per 1,000 male specimens. These data show that mosquitoes become infected by means other than by blood feeding, possibly by transovarial transmission. The placement of traps in tree canopies and in catch basins can be used to augment current practices of placement of traps near the ground for surveillance of mosquitoes infected with WNV and for studies of the ecology of WNV.

The explicit dependence of quadrat variance on the ratio of clump size to quadrat size.

ABSTRACT In the past decade, it has become common practice to pool mapped binary epidemic data into quadrats. The resultant "quadrat counts" can then be analyzed by fitting them to a probability distribution (i.e., betabinomial). Often a binary form of Taylor's power law is used to relate the quadrat variance to the quadrat mean. The fact that there is an intrinsic dependence of such analyses on quadrat size and shape is well known. However, a clear-cut exposition of the direct connection between the spatial properties of the two-dimensional pattern of infected plants in terms of the geometry of the quadrat and the results of quadrat-based analyses is lacking. This problem was examined both empirically and analytically. The empirical approach is based on a set of stochastically generated "mock epidemics" using a Neyman-Scott cluster process. The resultant spatial point-patterns of infected plants have a fixed number of disease foci characterized by a known length scale (monodisperse) and saturated to a known disease level. When quadrat samples of these epidemics are fit to a beta-binomial distribution, the resulting measures of aggregation are totally independent of disease incidence and most strongly dependent on the ratio of the length scale of the quadrat to the length scale of spatial aggregation and to a lesser degree on disease saturation within individual foci. For the analytical approach, the mathematical form for the variation in the sum of random variates is coupled to the geometry of a quadrat through an assumed exponential autocorrelation function. The net result is an explicit equation expressing the intraquadrat correlation, quadrat variance, and the index of dispersion in terms of the ratio of the quadrat length scale to the correlative length scale.

Measuring spatial aggregation in binary epidemics: correlative analysis and the advantage of fractal-based sampling.

ABSTRACT The incomplete sampling of a binary epidemic is nothing more than the overlap of two spatial patterns: the pattern of diseased plants and the pattern of sampled points. Thus, the information on the spatial arrangement of diseased plants obtained from such a sampling explicitly depends on the geometric locations of the sampled points. A number of procedures for sampling disease incidence are examined. These include samples placed on a regular grid, spatially clustered samples, randomly selected samples, and samples specified by a nested fractal design. The performance of these various sampling schemes was examined using simulated binary epidemics with varying degrees of spatial aggregation over different length scales, generated using a Neyman-Scott cluster process. A modification of spatial correlation analysis specifically geared to binary epidemics is derived and shown to be equivalent to a X(2) test comparing the number of infected plant pairs to that expected from a spatially random epidemic. This analysis was applied to the data obtained using the various sampling schemes and the results are compared and contrasted. For the same number of sampling points, the fractal design is most efficient in the detection of contagion and provides spatial information over a larger range of distance scales than other sampling schemes. However, the regular grid sampling scheme consistently yielded an estimate of average disease incidence that had the smallest variance. Sampling patterns consisting of randomly selected points were intermediate in behavior between the two extremes.

Concentration-dependent kinetics of pollutant desorption from soils.

Sorption-desorption kinetics play a major role in transport and bioavailability of pollutants in soils. Contaminant concentration is a potentially important factor controlling kinetics. A previous paper dealt with the effect of solute concentration on fractional uptake rates of phenanthrene and pyrene from a finite aqueous source. In this study we determined the effect of initial phenanthrene sorbed concentration (q(0)) on the fractional mass desorption rates from each of six soils to a zero-concentration solution, approximated by including a polymer adsorbent (Tenax) as a third-phase sink. The soils were preequilibrated with phenanthrene for 180 d. Consistent with theory, the fractional desorption rates determined by empirical curve fitting increased with q(0) provided the isotherm was nonlinear. After 500 to 600 d of desorption at the steepest possible concentration gradient, all soils retained a highly resistant fraction, which ranged from 4 to 31% of q(0), except for one soil at a high q(0). The highly resistant fraction decreased with increasing q(0), for nonlinear isotherm cases, but increased with q(0) for linear or nearly linear isotherm cases. Application of a nonlinear diffusion model, the dual-mode diffusion model (DMDM), to the nonresistant fraction gave reasonably good fits. The DMDM attributes the increase with concentration of the apparent diffusivity to a decrease in the proportion of sorbate occupying immobile sites (holes) in soil organic matter. The concentration-dependent term in the expression for the apparent diffusivity correlated with either of two indices that reflect the linearity of the sorption isotherm. Bunker C oil present in one soil acted as a partition domain. The findings of this study are consistent with heterogeneous models of soil organic matter, and indicate that concentration effects should be taken into account whenever desorption rate is important.

Spatial Analyses of Viral Epidemics in Australian Hop Gardens: Implications for Mechanisms of Spread.

Plant-to-plant spread of Hop mosaic virus (HpMV), Hop latent virus (HpLV), and Prunus necrotic ringspot virus (PNRSV) were monitored in two commercial hop gardens of cv. Victoria in Australia. At site 1, the cultural practice of mowing to remove excess basal growth operated exclusively along rows, while at site 2, mowing occurred both along and across rows. Other cultural practices such as stringing, bine-training, spraying, and harvesting were directed along rows at both sites. Spatial aggregation within and across rows was assessed by ordinary runs analysis, and by radial correlation analysis using the program 2DCORR. The dominant spread mechanism of carlaviruses differed between sites. At site 1, along-row aggregation of both HpLV and HpMV in 1997 reflected the importance of basal-growth intertwining for virus transmission, either due to mechanical transmission or movement of apterous or alatae vectors along rows. At site 2, the random incidence of HpLV and HpMV reflected spread by alatae vectors. The spread of PNRSV at both sites was associated with mowing direction. This study reports the first use of a modified version of radial correlation analysis.

Isolation of Nectria galligena from Cankers on Sweet Birch.

The fungus Nectria galligena infects many species of hardwood trees, causing "perennial canker," "European canker," or "target canker." Cultures from ascospores produced in stromata on the cankers have confirmed the pathogen species, but the fungus has rarely been successfully isolated from inside the cankers. Concern for the sweet birch trees affected by this disease in Connecticut prompted us to try isolations, using a method successful in isolating chestnut tree pathogens. Small pieces of cankered bark were stabbed into Granny Smith apples and, after incubation in boxes in the laboratory, pure cultures of N. galligena were easily removed as the pathogen slowly grew out, unchallenged, into the tissue of the apples. This technique will now be used to sample the N. galligena population in the birch populations being studied.