Evaluation of Methods for Sampling the Malaria Vector Anopheles darlingi (Diptera, Culicidae) in Suriname and the Relation With Its Biting Behavior

The effectiveness of CO2-baited and human-baited mosquito traps for the sampling of Anopheles darlingi Root was evaluated and compared with human landing collections in Suriname. Biting preferences of this mosquito on a human host were studied and related to trapping data. Traps used were the Centers for Disease Control and Prevention Miniature Light trap, the BG Sentinel mosquito trap, the Mosquito Magnet Liberty Plus mosquito trap (MM-Plus), and a custom-designed trap. Carbon dioxide and humans protected by a bed net were used as bait in the studies. The number of An. darlingi collected was greater with human landing collections than with all other collection methods. An. darlingi did not show a preference for protected humans over CO2 bait. The BG Sentinel mosquito trap with CO2 or human odor as bait and the MM-Plus proved the best alternative sampling tools for An. darlingi. The BG Sentinel mosquito trap with CO2 or human odor as bait was also very efficient at collecting Culex spp. In a field study on biting preferences of wild An. darlingi, the females showed directional biting behavior (P <0.001), with a majority of females (93.3%) biting the lower legs and feet when approaching a seated human host. Higher efficiency of the closer-to-the-ground collecting MM-Plus and BG Sentinel mosquito trap when compared with the other trapping methods may be a result of a possible preference of this mosquito species for low-level biting. It is concluded that odor-baited sampling systems can reliably collect An. darlingi, but the odor bait needs to be improved, for instance, by including host-specific volatiles, to match live human baits

Carbon dioxide baited trap catches do not correlate with human landing collections of Anopheles aquasalis in Suriname

Three types of carbon dioxide-baited traps, i.e., the Centers for Disease Control Miniature Light Trap without light, the BioGents (BG) Sentinel Mosquito Trap (BG-Sentinel) and the Mosquito Magnet® Liberty Plus were compared with human landing collections in their efficiency in collecting Anopheles (Nyssorhynchus) aquasalis mosquitoes. Of 13,549 total mosquitoes collected, 1,019 (7.52%) were An. aquasalis. Large numbers of Culex spp were also collected, in particular with the (BG-Sentinel). The majority of An. aquasalis (83.8%) were collected by the human landing collection (HLC). None of the trap catches correlated with HLC in the number of An. aquasalis captured over time. The high efficiency of the HLC method indicates that this malaria vector was anthropophilic at this site, especially as carbon dioxide was insufficiently attractive as stand-alone bait. Traps using carbon dioxide in combination with human odorants may provide better results

Levend of dood, dat is de vraag!

In de literatuur zijn voor de detectie van plantenpathogenen diverse methodieken beschreven. De biologische methodieken detecteren alleen levende organismen. Morfologische, serologische en moleculaire technieken maken mmestal geen onderscheid tussen dood en levend of infectieus en niet infectieus. Met name voor quarantaineorganismen is het onderscheid tussen levende en dode pathogenen van essentieel belang. Binnen het FES-programma 'Versterking infrastructuur plantgezondheid' is binnen werkpakket 3 'Ontwikkeling van methoden voor het aantonen van vitaliteit van plantenpathogenen' gewerkt aan de detectie van vitaliteit in nematoden, schimmels en bacteriën

Spatial and temporal clustering of dengue virus transmission in Thai villages.

BackgroundTransmission of dengue viruses (DENV), the leading cause of arboviral disease worldwide, is known to vary through time and space, likely owing to a combination of factors related to the human host, virus, mosquito vector, and environment. An improved understanding of variation in transmission patterns is fundamental to conducting surveillance and implementing disease prevention strategies. To test the hypothesis that DENV transmission is spatially and temporally focal, we compared geographic and temporal characteristics within Thai villages where DENV are and are not being actively transmitted.Methods and findingsCluster investigations were conducted within 100 m of homes where febrile index children with (positive clusters) and without (negative clusters) acute dengue lived during two seasons of peak DENV transmission. Data on human infection and mosquito infection/density were examined to precisely (1) define the spatial and temporal dimensions of DENV transmission, (2) correlate these factors with variation in DENV transmission, and (3) determine the burden of inapparent and symptomatic infections. Among 556 village children enrolled as neighbors of 12 dengue-positive and 22 dengue-negative index cases, all 27 DENV infections (4.9% of enrollees) occurred in positive clusters (p &lt; 0.01; attributable risk [AR] = 10.4 per 100; 95% confidence interval 1-19.8 per 100]. In positive clusters, 12.4% of enrollees became infected in a 15-d period and DENV infections were aggregated centrally near homes of index cases. As only 1 of 217 pairs of serologic specimens tested in positive clusters revealed a recent DENV infection that occurred prior to cluster initiation, we attribute the observed DENV transmission subsequent to cluster investigation to recent DENV transmission activity. Of the 1,022 female adult Ae. aegypti collected, all eight (0.8%) dengue-infected mosquitoes came from houses in positive clusters; none from control clusters or schools. Distinguishing features between positive and negative clusters were greater availability of piped water in negative clusters (p &lt; 0.01) and greater number of Ae. aegypti pupae per person in positive clusters (p = 0.04). During primarily DENV-4 transmission seasons, the ratio of inapparent to symptomatic infections was nearly 1:1 among child enrollees. Study limitations included inability to sample all children and mosquitoes within each cluster and our reliance on serologic rather than virologic evidence of interval infections in enrollees given restrictions on the frequency of blood collections in children.ConclusionsOur data reveal the remarkably focal nature of DENV transmission within a hyperendemic rural area of Thailand. These data suggest that active school-based dengue case detection prompting local spraying could contain recent virus introductions and reduce the longitudinal risk of virus spread within rural areas. Our results should prompt future cluster studies to explore how host immune and behavioral aspects may impact DENV transmission and prevention strategies. Cluster methodology could serve as a useful research tool for investigation of other temporally and spatially clustered infectious diseases

Impact of Gut Bacteria on the Infection and Transmission of Pathogenic Arboviruses by Biting Midges and Mosquitoes

Tripartite interactions among insect vectors, midgut bacteria, and viruses may determine the ability of insects to transmit pathogenic arboviruses. Here, we investigated the impact of gut bacteria on the susceptibility of Culicoides nubeculosus and Culicoides sonorensis biting midges for Schmallenberg virus, and of Aedes aegypti mosquitoes for Zika and chikungunya viruses. Gut bacteria were manipulated by treating the adult insects with antibiotics. The gut bacterial communities were investigated using Illumina MiSeq sequencing of 16S rRNA, and susceptibility to arbovirus infection was tested by feeding insects with an infectious blood meal. Antibiotic treatment led to changes in gut bacteria for all insects. Interestingly, the gut bacterial composition of untreated Ae. aegypti and C. nubeculosus showed Asaia as the dominant genus, which was drastically reduced after antibiotic treatment. Furthermore, antibiotic treatment resulted in relatively more Delftia bacteria in both biting midge species, but not in mosquitoes. Antibiotic treatment and subsequent changes in gut bacterial communities were associated with a significant, 1.8-fold increased infection rate of C. nubeculosus with Schmallenberg virus, but not for C. sonorensis. We did not find any changes in infection rates for Ae. aegypti mosquitoes with Zika or chikungunya virus. We conclude that resident gut bacteria may dampen arbovirus transmission in biting midges, but not so in mosquitoes. Use of antimicrobial compounds at livestock farms might therefore have an unexpected contradictory effect on the health of animals, by increasing the transmission of viral pathogens by biting midges.</p

Fine Scale Spatiotemporal Clustering of Dengue Virus Transmission in Children and Aedes aegypti in Rural Thai Villages

Background: Based on spatiotemporal clustering of human dengue virus (DENV) infections, transmission is thought to occur at fine spatiotemporal scales by horizontal transfer of virus between humans and mosquito vectors. To define the dimensions of local transmission and quantify the factors that support it, we examined relationships between infected humans and Aedes aegypti in Thai villages. Methodology/Principal Findings: Geographic cluster investigations of 100-meter radius were conducted around DENV-positive and DENV-negative febrile ‘‘index’’ cases (positive and negative clusters, respectively) from a longitudinal cohort study in rural Thailand. Child contacts and Ae. aegypti from cluster houses were assessed for DENV infection. Spatiotemporal, demographic, and entomological parameters were evaluated. In positive clusters, the DENV infection rate among child contacts was 35.3% in index houses, 29.9% in houses within 20 meters, and decreased with distance from the index house to 6.2% in houses 80–100 meters away (pAe. aegypti were DENV-infectious (i.e., DENV-positive in head/thorax) in positive clusters (23/1755; 1.3%) than negative clusters (1/1548; 0.1%). In positive clusters, 8.2% of mosquitoes were DENV-infectious in index houses, 4.2% in other houses with DENV-infected children, and 0.4% in houses without infected children (pAe. aegypti pupae and adult females were more numerous only in houses containing infectious mosquitoes. Conclusions/Significance: Human and mosquito infections are positively associated at the level of individual houses and neighboring residences. Certain houses with high transmission risk contribute disproportionately to DENV spread to neighboring houses. Small groups of houses with elevated transmission risk are consistent with over-dispersion of transmission (i.e., at a given point in time, people/mosquitoes from a small portion of houses are responsible for the majority of transmission)

CRISPR-Cas9 ribonucleoprotein-mediated co-editing and counterselection in the rice blast fungus

The rice blast fungus Magnaporthe oryzae is the most serious pathogen of cultivated rice and a significant threat to global food security. To accelerate targeted mutation and specific genome editing in this species, we have developed a rapid plasmid-free CRISPR-Cas9-based genome editing method. We show that stable expression of Cas9 is highly toxic to M. oryzae. However efficient gene editing can be achieved by transient introduction of purified Cas9 pre-complexed to RNA guides to form ribonucleoproteins (RNPs). When used in combination with oligonucleotide or PCR-generated donor DNAs, generation of strains with specific base pair edits, in-locus gene replacements, or multiple gene edits, is very rapid and straightforward. We demonstrate a co-editing strategy for the creation of single nucleotide changes at specific loci. Additionally, we report a novel counterselection strategy which allows creation of precisely edited fungal strains that contain no foreign DNA and are completely isogenic to the wild type. Together, these developments represent a scalable improvement in the precision and speed of genetic manipulation in M. oryzae and are likely to be broadly applicable to other fungal species

Methods for microbial DNA extraction from soil for PCR amplification

Amplification of DNA from soil is often inhibited by co-purified contaminants. A rapid, inexpensive, large-scale DNA extraction method involving minimal purification has been developed that is applicable to various soil types (1). DNA is also suitable for PCR amplification using various DNA targets. DNA was extracted from 100g of soil using direct lysis with glass beads and SDS followed by potassium acetate precipitation, polyethylene glycol precipitation, phenol extraction and isopropanol precipitation. This method was compared to other DNA extraction methods with regard to DNA purity and size

Prediction of Dengue Disease Severity among Pediatric Thai Patients Using Early Clinical Laboratory Indicators

Patients with severe dengue illness typically develop complications in the later stages of illness, making early clinical management of all patients with suspected dengue infection difficult. An early prediction tool to identify which patients will have a severe dengue illness will improve the utilization of limited hospital resources in dengue endemic regions. We performed classification and regression tree (CART) analysis to establish predictive algorithms of severe dengue illness. Using a Thai hospital pediatric cohort of patients presenting within the first 72 hours of a suspected dengue illness, we developed diagnostic decision algorithms using simple clinical laboratory data obtained on the day of presentation. These algorithms correctly classified near 100% of patients who developed a severe dengue illness while excluding upwards of 50% of patients with mild dengue or other febrile illnesses. Our algorithms utilized white blood cell counts, percent white blood cell differentials, platelet counts, elevated aspartate aminotransferase, hematocrit, and age. If these algorithms can be validated in other regions and age groups, they will help in the clinical management of patients with suspected dengue illness who present within the first three days of fever onset

Effects of co-habitation between Anopheles gambiae s.s. and Culex quinquefasciatus aquatic stages on life history traits

<p>Abstract</p> <p>Background</p> <p>The effective measures for the control of malaria and filariasis vectors can be achieved by targeting immature stages of anopheline and culicine mosquitoes in productive habitat. To design this strategy, the mechanisms (like biotic interactions with conspecifc and heterospecific larvae) regulating mosquito aquatic stages survivorship, development time and the size of emerging adults should be understood. This study explored the effect of co-habitation between <it>An. gambiae </it>s.s. and <it>Cx. quinquefasciatus </it>on different life history traits of both species under different densities and constant food supply in the habitats of the same size under semi-natural conditions.</p> <p>Methods</p> <p>Experiments were set up with three combinations; <it>Cx. quinquefasciatus </it>alone (single species treatment), <it>An. gambiae </it>s.s. alone (single species treatment); and <it>An. gambiae </it>s.s. with <it>Cx. quiquefasciatus </it>(co-habitation treatment) in different densities in semi field situation.</p> <p>Results</p> <p>The effect of co-habitation of <it>An. gambiae </it>s.s. and <it>Cx. quinquefasciatus </it>was found to principally affect three parameters. The wing-lengths (a proxy measure of body size) of <it>An. gambiae </it>s.s. in co-habitation treatments were significantly shorter in both females and males than in <it>An. gambiae </it>s.s single species treatments. In <it>Cx. quinquefasciatus</it>, no significant differences in wing-length were observed between the single species and co-habitation treatments. Daily survival rates were not significantly different between co-habitation and single species treatments for both <it>An. gambiae </it>s.s. and <it>Cx. quinquefasciatus</it>. Developmental time was found to be significantly different with single species treatments developing better than co-habitation treatments. Sex ratio was found to be significantly different from the proportion of 0.5 among single and co-habitation treatments species at different densities. Single species treatments had more males than females emerging while in co-habitation treatments more females emerged than males. In this study, there was no significant competitive survival advantage in co-habitation.</p> <p>Conclusion</p> <p>These results suggest that co-habitation of <it>An. gambiae </it>s.s. and <it>Cx. quinquefasciatus </it>in semi-natural conditions affect mostly <it>An. gambiae </it>s.s. body size. Hence, more has to be understood on the effects of co-habitation of <it>An. gambiae </it>s.s. and <it>Cx. quinquefasciatus </it>in a natural ecology and its possible consequences in malaria and filariasis epidemiology.</p