Robert J. Novak

Larval Habitat Diversity and Ecology of Anopheline Larvae in Eritrea

Abstract Studies on the spatial distribution of anopheline mosquito larvae were conducted in 302 villages over two transmission seasons in Eritrea. Additional longitudinal studies were also conducted at eight villages over a 24-mo period to determine the seasonal variation in anopheline larval densities. Eight anopheline species were identified with Anopheles arabiensis predominating in most of the habitats. Other species collected included: An. cinereus, An. pretoriensis, An. d’thali, An. funestus, An. squamosus, An. adenensis, and An. demeilloni. An. arabiensis was found in five of the six aquatic habitats found positive for anopheline larvae during the survey. Anopheles larvae were sampled predominantly from stream edges and streambed pools, with samples from this habitat type representing 91.2% (n = 9481) of the total anopheline larval collection in the spatial distribution survey. Other important anopheline habitats included rain pools, ponds, dams, swamps, and drainage channels at communal water supply points. Anopheline larvae were abundant in habitats that were shallow, slow flowing and had clear water. The presence of vegetation, intensity of shade, and permanence of aquatic habitats were not significant determinants of larval distribution and abundance. Larval density was positively correlated with water temperature. Larval abundance increased during the wet season and decreased in the dry season but the timing of peak densities was variable among habitat types and zones. Anopheline larvae were collected all year round with the dry season larval production restricted mainly to artificial aquatic habitats such as drainage channels at communal water supply points. This study provides important information on seasonal patterns of anopheline larval production and larval habitat diversity on a countrywide scale that will be useful in guiding larval control operations in Eritrea.

Use of the polymerase chain reaction for the sensitive detection of St. Louis encephalitis viral RNA

Polymerase chain reaction (PCR) assays were developed for the detection of RNA from the St. Louis encephalitis (SLE) virus. Using computer-assisted analysis of the MSI-7 strain SLE virus genome, two primer pairs were selected from the capsid-coding and the membrane associated protein-coding genes, and one from the envelope-coding gene. Reverse transcription was primed with either specific oligomers or with random hexamers; these methods were compared for cDNA synthesis and subsequent PCR amplification with the oligomeric pairs. Random hexamers provided more sensitive detection of viral RNA. Each primer pair specifically amplified the expected sized fragment from the Parton SLE strain grown in Aedes albopictus cells, but did not amplify Aedes albopictus cell RNA controls. The technique also detected SLE virus RNA in 1 pg of total cellular RNA added to a background of 1 microgram boiled brain tissue, and in 0.5 pg of total RNA added to homogenized mosquito abdomen. PCR-based assays may be adaptable to detect SLE virus RNA in naturally infected mosquitoes, birds, and human cerebrospinal fluid and brain.

Problems in Estimating Mosquito Infection Rates Using Minimum Infection Rate

Determining the role of various mosquito species in the transmission of arboviruses relies on an accurate estimate of the proportion of infected individuals in an area. Basically, there are two methods for estimating the proportion of infected individuals from pooled samples. One is the widely used minimum infection rate (MIR), which is calculated as the ratio of the number of positive pools to the total number of mosquitoes tested. The underlying assumption of the MIR is that only one infected individual exists in a positive pool. This assumption is valid for arboviruses that are relatively rare, but it becomes problematic when infection rates are high and/or pool sizes are large. The other method of estimating the proportion of infected individuals in pooled samples is the maximum likelihood estimation (MLE), which is defined as the infection rate most likely observed given the testing results and an assumed probabilistic model (i.e., binomial distribution of infected individuals in a positive …

Assessment of arbovirus vector infection rates using variable size pooling.

Abstract.  Pool testing of vector samples for arboviruses is widely used in surveillance programmes. The proportion of infected mosquitoes (Diptera: Culicidae) is often estimated from the minimum infection rate (MIR), based on the assumption of only one infected mosquito per positive pool. This assumption becomes problematic when pool size is large and/or infection rate is high. By relaxing this constraint, maximum likelihood estimation (MLE) is more useful for a wide range of infection levels that may be encountered in the field. We demonstrate the difference between these two estimation approaches using West Nile virus (WNV) surveillance data from vectors collected by gravid traps in Chicago during 2002. MLE of infection rates of Culex mosquitoes was as high as 60 per 1000 at the peak of transmission in August, whereas MIR was less than 30 per 1000. More importantly, we demonstrate roles of various pooling strategies for better estimation of infection rates based on simulation studies with hypothetical mosquito samples of 18 pools. Variable size pooling (with a serial pool sizes of 5, 10, 20, 30, 40 and 50 individuals) performed consistently better than a constant size pooling of 50 individuals. We conclude that variable pool size coupled with MLE is critical for accurate estimates of mosquito infection rates in WNV epidemic seasons.

Mosquito Species Succession and Physicochemical Factors Affecting Their Abundance in Rice Fields in Mwea, Kenya

Abstract The succession of mosquito species and abiotic factors affecting their distribution and abundance in rice (Oryza spp.) fields was investigated over a 16-wk rice growing cycle covering the period between January and May 2006. Fifteen experimental rice plots were sampled for mosquito larvae and characterized based on rice height, number of tillers, floating vegetation cover, water depth, water temperature, turbidity, salinity, pH, dissolved oxygen, total dissolved solids, and conductivity. Microscopic identification of 3,025 larvae yielded nine mosquito species predominated by Anopheles arabiensis Patton (45.0%), Culex quinquefasciatus Say (35.8%), Anopheles pharoensis Theobald (9.0%) and Ficalbia splendens Theobald (7.1%). Other species, including Anopheles rufipes Gough, Anopheles coustani Laveran, Anonopheles maculipalpis Giles, Culex annulioris Theobald, and Culex poicilipes Theobald made up 3.1% of the total collection. Anopheles gambiae s.l., Cx. quinquefasciatus, and An. pharoensis occurred throughout the cycle, but they were more abundant up to 4 wk posttransplanting with peaks after fertilizer application. As rice plants became established, three groups of mosquitoes were recognized: the first groups included An. rufipes, Fl. splendens, and Cx. annulioris, which occurred throughout much of the second half of the rice cycle, whereas the second group included Cx. poicilipes, which was found in the middle of the rice cycle. An. coustani and An. maculipalpis formed the third group occurring toward the end of the cycle. Dissolved oxygen, number of tillers, and rice height were negatively associated with the abundance of An. arabiensis and Cx. quinquefasciatus larvae. In addition, Cx. quinquefasciatus also was associated with water depth (−ve) and turbidity (+ve). Abundance of An. pharoensis larvae was significantly associated with water temperature (+ve), the number of tillers (−ve), and rice height (−ve), whereas Fl. splendens was significantly associated with the number of tillers (+ve). The results demonstrate a complex nature of the interactions between some of the factors in the ecosystem and mosquito species abundance and calls for time-dependent and species-specific mosquito control operations.

Identification of two genetic markers that distinguish pathogenic and nonpathogenic strains of Acanthamoeba spp.

Abstract Species-level identification of Acanthamoeba isolates is difficult and gives little or no indication of the isolates pathogenicity. We identified two amplification-based genetic markers that were highly correlated with pathogenicity in Acanthamoeba spp. One marker, designed to amplify a 485-bp fragment of the small-subunit ribosomal RNA gene (ssrDNA), was preferentially amplified from the nonpathogenic strains; amplifications from the pathogenic strains yielded anomalous fragments of 650 and 900 bp. A second marker was developed on the basis of the anomalous 650-bp fragment. Primers to this sequence preferentially amplified a noncoding locus (called Ac6) only from the pathogenic strains. These two genetic markers may be useful for identification of pathogenic Acanthamoeba spp. strains.

Culex Population Dynamics and West Nile Virus Transmission in East-Central Illinois

ABSTRACT Temporal changes in the abundance Culex restuans and Culex pipiens were monitored in east-central Illinois for over a decade using infusion-baited oviposition traps. The 2 species typically exhibited a seasonal shift in relative abundance with a mean crossover date (when the proportion of egg rafts from both species is equal) of August 10 or 11, depending on leap year, with a 95% confidence interval of ±10.7 days. The date of crossover was linearly related to the date of last spring frost and occurred on average about 123 days after the last spring frost. Despite the predictability of crossover, the weekly pattern in the proportion of Cx. pipiens before and after crossover varied considerably, even between years with similar crossover dates. After West Nile virus became established in our area, we found that transmission based on Culex from gravid traps did not increase until Cx. pipiens abundance increased in oviposition traps. Infection rates peaked within the half-month period after crossover. The peak in Cx. pipiens abundance in oviposition traps during this 3-year period was between the 2nd half of August and the end of September. A higher magnitude of transmission in 2002 coincided with warmer temperatures during July and August and an extended period in which the 2 Culex species were in relatively equal abundance.

Evaluation of Four Sampling Techniques for Surveillance of Culex quinquefasciatus (Diptera: Culicidae) and Other Mosquitoes in African Rice Agroecosystems

Abstract Field studies were conducted in a rice, Oryza sativa L., agroecosystem in Mwea Kenya to compare the efficiency of CO2-baited Centers for Disease Control (CDC) light traps against nonbaited CDC light traps and gravid traps against oviposition traps in outdoor collection of Culex quinquefasciatus Say (Diptera: Culicidae) and other mosquitoes. Collectively, 21 mosquito species from the genera Culex, Anopheles, Mansonia, Ficalbia, and Aedes were captured during the 10-wk study period. Cx. quinquefasciatus was the predominant species in all trap types with proportions ranging from 57% in the nonbaited CDC light traps to 95% in the gravid traps. Significantly higher numbers of Cx. quinquefasciatus and Culex annulioris Theobald were collected in the CO2-baited CDC light traps than in the nonbaited CDC light traps, but the numbers of other mosquito species, including malaria vectors Anopheles arabiensis Patton and Anopheles funestus Giles did not differ significantly between the two trap types. More Cx. quinquefasciatus females were collected in grass infusion-baited gravid traps than egg rafts of this species in oviposition traps containing the same infusion. Although most mosquitoes captured in CO2-baited and nonbaited CDC light traps were unfed, most of those collected in gravid traps were gravid. From these findings, it is concluded that at least in the rice-growing area of Mwea Kenya, CO2-baited CDC light traps in conjunction with gravid traps can be used in monitoring of Cx. quinquefasciatus both for control and disease surveillance.

Remote and field level quantification of vegetation covariates for malaria mapping in three rice agro-village complexes in Central Kenya.

BackgroundWe examined algorithms for malaria mapping using the impact of reflectance calibration uncertainties on the accuracies of three vegetation indices (VI)s derived from QuickBird data in three rice agro-village complexes Mwea, Kenya. We also generated inferential statistics from field sampled vegetation covariates for identifying riceland Anopheles arabiensis during the crop season. All aquatic habitats in the study sites were stratified based on levels of rice stages; flooded, land preparation, post-transplanting, tillering, flowering/maturation and post-harvest/fallow. A set of uncertainty propagation equations were designed to model the propagation of calibration uncertainties using the red channel (band 3: 0.63 to 0.69 μm) and the near infra-red (NIR) channel (band 4: 0.76 to 0.90 μm) to generate the Normalized Difference Vegetation Index (NDVI) and the Soil Adjusted Vegetation Index (SAVI). The Atmospheric Resistant Vegetation Index (ARVI) was also evaluated incorporating the QuickBird blue band (Band 1: 0.45 to 0.52 μm) to normalize atmospheric effects. In order to determine local clustering of riceland habitats Gi*(d) statistics were generated from the ground-based and remotely-sensed ecological databases. Additionally, all riceland habitats were visually examined using the spectral reflectance of vegetation land cover for identification of highly productive riceland Anopheles oviposition sites.ResultsThe resultant VI uncertainties did not vary from surface reflectance or atmospheric conditions. Logistic regression analyses of all field sampled covariates revealed emergent vegetation was negatively associated with mosquito larvae at the three study sites. In addition, floating vegetation (-ve) was significantly associated with immature mosquitoes in Rurumi and Kiuria (-ve); while, turbidity was also important in Kiuria. All spatial models exhibit positive autocorrelation; similar numbers of log-counts tend to cluster in geographic space. The spectral reflectance from riceland habitats, examined using the remote and field stratification, revealed post-transplanting and tillering rice stages were most frequently associated with high larval abundance and distribution.ConclusionNDVI, SAVI and ARVI generated from QuickBird data and field sampled vegetation covariates modeled cannot identify highly productive riceland An. arabiensis aquatic habitats. However, combining spectral reflectance of riceland habitats from QuickBird and field sampled data can develop and implement an Integrated Vector Management (IVM) program based on larval productivity.

Spatially targeting Culex quinquefasciatus aquatic habitats on modified land cover for implementing an Integrated Vector Management (IVM) program in three villages within the Mwea Rice Scheme, Kenya

BackgroundContinuous land cover modification is an important part of spatial epidemiology because it can help identify environmental factors and Culex mosquitoes associated with arbovirus transmission and thus guide control intervention. The aim of this study was to determine whether remotely sensed data could be used to identify rice-related Culex quinquefasciatus breeding habitats in three rice-villages within the Mwea Rice Scheme, Kenya. We examined whether a land use land cover (LULC) classification based on two scenes, IKONOS at 4 m and Landsat Thematic Mapper at 30 m could be used to map different land uses and rice planted at different times (cohorts), and to infer which LULC change were correlated to high density Cx. quinquefasciatus aquatic habitats. We performed a maximum likelihood unsupervised classification in Erdas Imagine V8.7® and generated three land cover classifications, rice field, fallow and built environment. Differentially corrected global positioning systems (DGPS) ground coordinates of Cx. quinquefasciatus aquatic habitats were overlaid onto the LULC maps generated in ArcInfo 9.1®. Grid cells were stratified by levels of irrigation (well-irrigated and poorly-irrigated) and varied according to size of the paddy.ResultsTotal LULC change between 1988–2005 was 42.1 % in Kangichiri, 52.8 % in Kiuria and and 50.6 % Rurumi. The most frequent LULC changes was rice field to fallow and fallow to rice field. The proportion of aquatic habitats positive for Culex larvae in LULC change sites was 77.5% in Kangichiri, 72.9% in Kiuria and 73.7% in Rurumi. Poorly – irrigated grid cells displayed 63.3% of aquatic habitats among all LULC change sites.ConclusionWe demonstrate that optical remote sensing can identify rice cultivation LULC sites associated with high Culex oviposition. We argue that the regions of higher Culex abundance based on oviposition surveillance sites reflect underlying differences in abundance of larval habitats which is where limited control resources could be concentrated to reduce vector larval abundance.