Laboratory and field evaluations of industrial formulations of Bacillus thuringiensis serovar. israelensis against some mosquito species of central Italy

Two wettable powder formulations (Bactimos and Vectobac) and a flowable concentrate (Teknar) formulation of Bacillus thuringiensis serovar. israelensis were evaluated as larvicides of Culex pipiens, Aedes caspius, and Aedes detritus. In the laboratory, the levels of susceptibility of these species to the test formulations were essentially similar and corresponded to their relative potencies; the LC₉₀ values ranged from 0.042 to 0.37 ppm. C. pipiens, A. caspius, and A. detritus, in that order, were susceptible to the biocide. Under field conditions in central Italy. Bactimos at 0.5 kg/ha gave 98% control of C. pipiens in an irrigation canal. Teknar at 1.25 liters/ha gave 86–100%, and at 2.5 liters/ha gave 90–100% control of C. pipiens in two natural ponds. Against A. caspius in salt marsh habitats, Bactimos at 0.5 kg/ha and Teknar at 2.5 liters/ha yielded complete control of the larvae, while a lower rate (0.2 kg/ha) of Bactimos, and Vectobac at 0.5 kg/ha resulted in 82–94% and 67–91% control, respectively. Higher rates (0.75 and 1.0 kg/ha) of Vectobac gave 76–100% and 98–100% control of A. caspius. Bactimos at 0.15 kg/ha gave 93–98% control of A. detritus in two salt marsh ponds. B. thuringiensis serovar. israelensis is practically economical for the control of C. pipiens, A. caspius, and A. detritus in the various biotopes in central Italy.     

The effect of infection with Dirofilaria immitis (dog heartworm) on fluid secretion rates in the Malpighian tubules of the mosquitoes Aedes taeniorhynchus and Anopheles quadrimaculatus

Dirofilaria immitis, the parasitic nematode causing the disease dog heartworm, is transmitted by female mosquitoes. During their development, larval nematodes reside in the cells of the Malpighian tubules of these mosquitoes for approx 13 days. We have examined the effect of the presence of these large intracellular parasites on the main physiological function of the Malpighian tubules, i.e. fluid secretion. Rates of fluid secretion were examined in vitro using both normal and infected tubules of the mosquito species Aedes taeniorhynchus and Anopheles quadrimaculatus. Tubules of A. quadrimaculatus show changes in trasport rate during the reproductive cycle. Those of A. taeniorhynchus do not. Infection with larvae of D. immitis had no effect on the rate of fluid secretion in tubules of A. quadrimaculatus. In A. taeniorhynchus by contrast, the tubules show decline in transport with time following infection. The reduction in transport capacity is proportional to the number of worms infecting the tubule. The present paper and separate ultrastructural studies demonstrate that parallel changes in microvillar ultrastructure and epithelial transport rates occur in response to infection by the parasite. In both species examined, the survival of the mosquitoes and their vector potential are determined by factors other than the transport capacities of the infected Malpighian tubules.     

Consequences of the introduction of the planarian Girardia anceps (Tricladida: Dugesiidae) in artificial containers with larvae of the mosquitoes Aedes aegypti and Culex pipiens (Diptera: Culicidae) from Argentina

Aedes aegypti and Culex pipiens are container-dwelling mosquito species that are vectors of important diseases to man, such as dengue and lymphatic filariasis, respectively. Predators of these pests are an interesting alternative to be incorporated to biological control measures. We tested the consequences of introducing individuals of Girardia anceps, a native freshwater flatworm species, within artificial water containers where larvae of these mosquitoes thrive. Our goals were to ascertain if mosquito species, density of larvae (high or low), type of water container (tires or ovitraps), and presence or absence of planarians affected mosquito survivorship (measured as number of individuals reaching the pupa stage) in manipulated artificial containers. Furthermore, we monitored ovitraps in the field along several months in order to explore the long-term effect of the presence of planarian on the colonization of these containers by feral mosquitoes under natural conditions. We found that the presence of planarians reduced the number of mosquitoes reaching pupation and that such reduction depends on the initial density of larvae. Reduction of populations of A. aegypti was high along the breeding season of this mosquito, being the effect less evident in C. pipiens. G. anceps could be an agent of control against container-breeding mosquitoes if its release in small water containers is complemented with other suitable management strategies.     

Polysaccharide and protein degradation,germination, and virulence against mosquitoes in the entomopathogenic fungus Metarhizium anisopliae

From a genetically uniform wild-type strain of Metarhizium anisopliae pathogenic to mosquitoes, mutants were selected which were altered in the ability to degrade starch, gelatin, or milk. The mutants with enhanced starch degradation (dep), when grown on starch-containing media, proved hypervirulent toward the mosquito Culex pipiens pipiens in standard laboratory tests. Alterations in protein (gelatin or milk) degradation did not correlate with changes in virulence. The dep mutants appear to belong to the same class as mutants selected previously as hypervirulent and characterized by early spore germination. The relationship among polysaccharide degradation, early germination, and virulence is discussed.     

Further study on Ascogregarina culicis in temperate Argentina: Prevalence and intensity in Aedes aegypti larvae and pupae

The bionomics of South American strains of Ascogregarina spp. are poorly known and the first studies were performed a few years ago. Our main objective was to characterize Ascogregarina culicis population in Aedes aegypti immatures in temperate Argentina. A total of 1800 water-filled containers were inspected within a cemetery of Buenos Aires City through a reproductive period of the host (October 2006-June 2007). The parasite was detected in 16.7% (329/1974) of the immatures and 8.5% (15/177) of the breeding sites. The prevalence decreased from 19.9% in larvae to 6.5% in pupae. In those infected breeding sites, about 85% of the immature mosquitoes harbor the parasite with a median intensity of nine trophozoites per larva and six gametocysts per pupa. The prevalence in shaded containers was higher than in sun exposed ones but the intensity of the infection was quite similar between both lighting conditions. Sun-exposed containers recorded water temperatures significantly higher than those under shade throughout the study period. Parasite trophozoites were only found from January to May with a clear seasonal pattern of prevalence. Monthly values of parasite prevalence and mosquito host (percentage of breeding sites and number of immatures) were significantly correlated at p < 0.05 when a temporal delay of two months was considered. Our results suggest that parasite prevalence is spatially and temporally heterogeneous in temperate urban Argentina, and these variations are associated with the host abundance.     

Infectivity of a microsporidium of mosquitoes (Nosema algerae) to larval stages of Schistosoma mansoni in Biomphalaria glabrata

Lai P. F. and Canning E. U. 1980. Infectivity of a microsporidium of mosquitoes (Nosema algerae) to larval stages of Schistosoma mansoni in Biomphalaria glabrata. International Journal for Parasitology10: 293–301. Nosema algerae derived from a closed colony of Anopheles stephensi was fed to Biomphalaria glabrata infected with Schistosoma mansoni. Mother and daughter sporocysts became hyperinfected but the snail tissues remained free of the microsporidia except for rare small aggregates of spores. These lay close to the sites occupied by mother or daughter sporocysts and were probably liberated from them. Irrespective of dose, fewer snails contained infected sporocysts when spores were given at 7 days post-miracidial infection than when given at 14 days. These periods corresponded respectively to stages when mother sporocysts only or daughter sporocysts as well were present in the snails. Infection of the sporocysts began in the tegumental cells, spread to the brood chamber and ultimately to the cercariae themselves. Heavily infected sporocysts contained fewer developing embryos. Doses of 10⁶ and 10⁷ spores/snail caused significant depression of cercaria output when given at 14 days but not at 7 days.     

Interaction between Neoaplectana carpocapsae (Nematoda: Steinernematidae) and Apanteles militaris (Hymenoptera: Bracondiae), a parasitoid of the armyworm, Pseudaletia unipuncta

The DD-136 strain of Neoaplectana carpocapsae adversely affected the development of immature stages of Apanteles militaris, a gregarious internal parasitoid of the armyworm, Pseudaletia unipuncta. The adverse effect of the nematode-bacterium complex was indirect, i.e., the infection by the nematode killed the host before A. militaris could complete its development. However, if armyworms containing 10- or 11-day-old A. militaris were exposed to dauer juveniles in Petri dishes, 48.1 and 94.4%, respectively, produced normal cocoons. If hosts containing 9- or 10-day-old A. militaris were fed dauer juveniles, 42.6 and 73.4% of the armyworms, respectively, produced A. militaris which formed normal cocoons. Cocoon-spinning A. militaris larvae were infected by the nematode. After cocoon formation was completed, the dauer juveniles could not penetrate the cocoon and infect the pupa. However, pupae in cocoons which had been deliberately cut open at one end became infected. A. militaris adults were infected when exposed to dauer juveniles in Petri dishes. After 3 days of exposure to dauer juveniles, 25.0, 44.2, and 7.0% of the adults in three trials were alive, whereas 100, 100, and 96.7% of the control adults were alive. Examination of dead adults in the nematode treatment showed that 67.6% contained nematodes. N. carpocapsae developed and reproduced in unparasitized armyworms, in armyworms containing 9-day-old A. militaris, and in those from which A. militaris had emerged. Production of dauer juveniles was significantly higher in unparasitized armyworms and in armyworms containing 9-day-old A. militaris than in those from which A. militaris had emerged.     

Experimental hut trials of permethrin-impregnated mosquito nets and eave curtains against malaria vectors in Tanzania

Permethrin impregnated netting was tested against Tanzanian populations of Anopheles arabiensis Patton, An.gambiae Giles and An.funestus Giles in experimental huts fitted with traps to catch samples of the mosquitoes existing during the night. Treated bednets killed some mosquitoes and increased the tendency of survivors to exit during the night. Treated cotton did not perform so well as treated nylon bednets. An impregnated bednet in which holes had been cut, to simulate a torn net, reduced the number of mosquitoes which fed and survived approximately as well as an intact untreated net. Treated curtains around the eaves of experimental huts did not perform so well as bednets but caused considerable reductions in the number of mosquitoes which fed and survived. However, there was no such effect when treated netting was placed around the eaves of a dwelling house. When one child slept under a treated net and another slept outside the net in the same hut, the number of bites on the latter child was less than if neither child had been under a net. Various aspects of the applicability of permethrin impregnated nets on a community basis are discussed.     

Immunisation against a serine protease inhibitor reduces intensity of Plasmodium berghei infection in mosquitoes

The mosquito innate immune response is able to clear the majority of Plasmodium parasites. This immune clearance is controlled by a number of regulatory molecules including serine protease inhibitors (serpins). To determine whether such molecules could represent a novel target for a malaria transmission-blocking vaccine, we vaccinated mice with Anopheles gambiae serpin-2. Antibodies against Anopheles gambiae serpin-2 significantly reduced the infection of a heterologous Anopheles species (Anopheles stephensi) by Plasmodium berghei, however this effect was not observed with Plasmodium falciparum. Therefore, this approach of targeting regulatory molecules of the mosquito immune system may represent a novel approach to transmission-blocking malaria vaccines.     

Development and use of quantitative competitive PCR assays for relative quantifying rumen anaerobic fungal populations in both in vitro and in vivo systems

This paper describes the use of a quantitative competitive polymerase chain reaction (QC-PCR) assay; using PCR primers to the rRNA locus of rumen fungi and a standard-control DNA including design and validation. In order to test the efficiency of this method for quantifying anaerobic rumen fungi, it has been attempted to evaluate this method in in vitro conditions by comparing with an assay based on measuring cell wall chitin. The changes in fungal growth have been studied when they are grown in in vitro on either untreated (US) or sodium hydroxide treated wheat straw (TS). Results showed that rumen fungi growth was significantly higher in treated samples compared with untreated during the 12 d incubation (P < 0.05) and plotting the chitin assay's results against the competitive PCR's showed high positive correlation (R² ≥ 0.87). The low mean values of the coefficients of variance in repeatability in the QC-PCR method against the chitin assay demonstrated more reliability of this new approach. And finally, the efficiency of this method was investigated in in vivo conditions. Samples of rumen fluid were collected from four fistulated Holstein steers which were fed four different diets (basal diet, high starch, high sucrose and starch plus sucrose) in rotation. The results of QC-PCR showed that addition of these non-structural carbohydrates to the basal diets caused a significant decrease in rumen anaerobic fungi biomass. The QC-PCR method appears to be a reliable and can be used for rumen samples.     

Innervation and control of tension in abdominal muscles of Blaberus discoidalis and Gromphadorhina portentosa

In Blaberus discoidalis and Gromphadorhina portentosa, the distribution of motor axons to the muscles which control movements of the spiracular valves at both respiratory and non-respiratory spiracles is identical. Both fast and slowly contracting heads of the opener muscles are innervated by an excitatory motor axon. Physiological properties of the opener excitor axon correlate with valve function. The slowly contracting head of the opener muscle is, in addition, innervated by a common inhibitor which also occasionally innervates closer muscle fibers. Activation of the common inhibitor terminates contraction of slowly contracting opener muscle fibres and initiates a rapid relaxation of these fibres.     

Field evaluation of a lethal ovitrap against dengue vectors in Brazil

Field evaluation of a "lethal ovitrap" (LO) to control dengue vector Aedes mosquitoes (Diptera: Culicidae), was undertaken in two Brazilian municipalities, Areia Branca and Nilopolis, in the State of Rio de Janeiro. The LO is designed to kill Aedes via an insecticide-treated ovistrip (impregnated with deltamethrin). In each municipality, the intervention was applied to a group of 30 houses (10 LOs/house) and compared to 30 houses without LOs in the same neighbourhood. Five LOs were put outside and five LOs inside each treated house. Three methods of monitoring Aedes density were employed: (i) percentage of containers positive for larvae and/or pupae; (ii) total pupae/house; (iii) total adult females/house collected by aspirator indoors. Weekly mosquito surveys began during the month before LO placement, by sampling from different groups of 10 houses/week for 3 weeks pre-intervention (i.e. 30 houses/month) and for 3 months post-intervention in both treated and untreated areas. Prior to LO placement at the end of February 2001, Aedes aegypti (L) densities were similar among houses scheduled for LO treatment and comparison (untreated control) at each municipality. Very few Ae. albopictus (Skuse) were found and this species was excluded from the assessment. Post-intervention densities of Ae. aegypti were significantly reduced for most comparators (P < 0.01), as shown by fewer positive containers (4-5 vs. 10-18) and pupae/house (0.3-0.7 vs. 8-10) at LO-treated vs. untreated houses, 3 months post-treatment at both municipalities. Numbers of adult Ae. aegypti females indoors were consistently reduced in LO-treated houses at Areia Branca (3.6 vs. 6.8/house 3 months post-intervention) but not at Niloplis (approximately 3/house, attributed to immigration). These results demonstrate sustained impact of LOs on dengue vector population densities in housing conditions of Brazilian municipalities.     

Nematicidal activity of Haplophyllum tuberculatum and Plectranthus cylindraceus oils against Meloidogyne javanica

The potentials of Haplophyllum tuberculatum and Plectranthus cylindraceus oils to control Meloidogyne javanica were investigated in vitro and in a greenhouse. A mixture of Haplophyllum and Plectranthus oils (1:1) was highly toxic to M. javanica in vitro, as it killed all nematode juveniles and inhibited hatching of eggs at 12.5 μg/ml concentration after 24 h exposure time, as did carbofuran at the same concentration. In the green-house, tomatoes grown in soil treated with a combination (1:1) of the two oils developed fewer root galls than those grown in soil treated with higher doses of either oil. The oil mixture, at 2.5 and 5.0 μg/ml of soil, was not phytotoxic to tomato plants as evident from the appearance and height of plants after 12 weeks exposure time, compared to treatment over the same period at lower effective doses. The nematicidal activity of the combined essential oils was suggested by the presence of C₁₀ dienes, C₁₀ trienes and C₁₀ phenol.     

Prevalence and seasonality of Ascogregarina culicis (Apicomplexa: Lecudinidae) in natural populations of Aedes aegypti (Diptera: Culicidae) from temperate Argentina

Ascogregarina infections from South America were recently documented in Brazil and Argentina. The aim of this study was to report our recent findings on the prevalence and seasonality of Ascogregarina culicis in Aedes aegypti adults from temperate Argentina. Between December 2003 and May 2005, 391 females of Ae. aegypti were captured in two areas of Greater Buenos Aires. Overall prevalence of A. culicis was 21.2% (83/391), and a significant difference was observed between both areas (28.4% vs. 8%). Infected Ae. aegypti were found from November to May, with highest values in March (24-37%). Parasite prevalence and host abundance showed similar seasonal patterns. Our observations suggest a widespread infestation of A. culicis among Ae. aegypti populations from temperate Argentina.     

Optimal control of Aedes aegypti mosquitoes by the sterile insect technique and insecticide

We present a mathematical model to describe the dynamics of mosquito population when sterile male mosquitoes (produced by irradiation) are introduced as a biological control, besides the application of insecticide. In order to analyze the minimal effort to reduce the fertile female mosquitoes, we search for the optimal control considering the cost of insecticide application, the cost of the production of irradiated mosquitoes and their delivery as well as the social cost (proportional to the number of fertilized females mosquitoes). The optimal control is obtained by applying the Pontryagin’s Maximum Principle.     

Factors affecting the thermal behavior of the sand lizard Liolaemus wiegmannii in natural and modified grasslands of temperate coastal dunes from Argentina

The present study describes and compares the thermal behavior of individuals of Liolaemus wiegmannii, inhabiting patches of natural grassland and modified patches with exotic trees of Acacia longifolia, by analyzing the factors affecting its behavior. Thermal behavior of L. wiegmannii was assessed by radiotracking 22 adult individuals in a coastal dune area from Argentina. In order to account for individual responses we analyzed the factors affecting thermal behavior using Generalized Linear Mixed Models. Thermal behavior of L. wiegmannii was mainly affected by the time of day and the substrate temperature. The individuals basking on bare sand in the morning shifted to filtered sunlight during midday and to the shade of clump and erect stems and herbs during the afternoon. The individuals buried in sand when the substrate temperature was low and the wind speed was high. The use of sub-shrubs and shrubs could not be explained solely by thermal factors. In natural L. wiegmannii regulated its body temperature by shuttling between microhabitats that provided them with different levels of exposure to sunlight during the day. However, in modified patches individuals varied in the choice of plants used as shaded places to mitigate high microenvironmental temperatures and they were also more exposed to full shaded sites and less to warm sand to bask, which could have negative consequences for its thermal biology.     

The occurrence of Melissococcus plutonius in healthy colonies of Apis mellifera and the efficacy of European foulbrood control measures

European foulbrood (EFB) persists in England and Wales despite current treatment methods, all of which include feeding honey bee colonies with the antibiotic oxytetracycline (OTC). A large-scale field experiment was conducted to monitor a husbandry-based method, using comb replacement (known as Shook swarm), as a drug free EFB control option. The understanding of EFB epidemiology is limited, with little information on the presence of Melissococcus plutonius in disease free colonies. Additional samples were collected from diseased and disease free apiaries to identify symptomless infection. EFB reoccurrence was not significantly different between OTC and husbandry methods and real-time PCR data demonstrated that fewer Shook swarm treated colonies contained M. plutonius carryover to the Spring following treatment. Asymptomatic colonies from diseased apiaries showed an increased risk of testing positive for M. plutonius compared to asymptomatic colonies from disease free apiaries. The probability of a sample being symptomatic increased when a greater quantity of M. plutonius was detected in adult bees and larvae. The possibility of treating EFB as an apiary disease rather than a colony disease and the implications of a control strategy without antibiotics are discussed.     

One-pot,multicomponent synthesis of 2,3-disubstituted quinazolin-ones with potent and selective activity against Toxoplasma gondii

The discovery of two quinazolinones with selective, single-digit micromolar activity (IC₅₀?=?6–7?µM) against the tachyzoites of the apicomplexan parasite Toxoplasma gondii is reported. These potent and selective third generation derivatives contain a benzyloxybenzyl substituent at C2 and a bulky aliphatic moiety at N3. Here we show that these quinazolinones inhibit T. gondii tachyzoite replication in an established infection, but do not significantly affect host cell invasion by the tachyzoites.     

Biological control of container-breeding mosquitoes, Aedes albopictus and Culex quinquefasciatus, in a Japanese island by release of Toxorhynchites splendens adults

To control container-breeding mosquitoes in the small island of Minnajima (0.56 km2), northern Okinawa, Japan, laboratory-reared adults (aged 7-10 days) of Toxorhynchites splendens (Palawan strain), a mosquito with predatory larvae, were released repeatedly during 1984, 1986 and 1987. Thirteen species of mosquitoes (Diptera: Culicidae) occurred in artificial containers, ground pools or crab-holes on the island, the predominant species being Aedes (Stegomyia) albopictus and Culex (Culex) quinquefasciatus. Predatory mosquito larvae of Culex (Lutzia) fuscanus and Cx (Lt.) halifaxii were found commonly in wet containers. In the first year of study, during a period of 54 days from 13 May to 5 July 1984, totals of 879 female and 806 male adults of Tx.splendens were released on six occasions. Similarly, between 29 April and 30 August 1986, totals of 2920 female and 2878 male adult Tx.splendens were released. In the third study year, totals of 2041 female and 1783 male Tx.splendens were released on eight occasions during 199 days from 23 April to 7 November 1987. After adult releases at two sites, the immature stages of Tx.splendens were found in 164 out of 502 traps in 1984, 421 out of 933 traps in 1986, and 151 out of 502 traps in 1987. The number of immatures of Tx.splendens present in each trap varied from 1 to 40 in 1984, 1 to 29 in 1986 and 1 to 9 in 1987. Numbers of immatures of the target species found in the traps during August-September averaged 71.9/trap/month in 1984, 114.7/trap/month in 1986 and 36.0/trap/month in 1987, significantly less in the traps with Tx.splendens than in those without them. The present field studies indicated that, in this small island, approximately 250 adult female and 200 male Tx.splendens per month should be released from April to November, and the releases should be carried out every year, in order to control effectively the target mosquitoes Ae.albopictus and Cx quinquefasciatus breeding in artificial containers in Minnajima.