Lack of independence, or pseudoreplication, in samples from ecological studies of insects reflects the complexity of working with living organisms: the finite and limited input of individuals, their relatedness (ecological and/or genetic), and the need to group organisms into functional experimental units to estimate population parameters (e.g., cohort replicates). Several decades ago, when the issue of pseudoreplication was first recognized, it was highlighted that mainstream statistical tools were unable to account for the lack of independence. For example, the variability as a result of differences across individuals would be confounded with that of the experimental units where they were observed (e.g., pans for mosquito larvae), whereas both sources of variability now can be separated using modern statistical techniques, such as the linear mixed effects model, that explicitly consider the different scales of variability in a dataset (e.g., mosquitoes and pans). However, the perception of pseudoreplication as a problem without solution remains. This study presents concepts to critically appraise pseudoreplication and the linear mixed effects model as a statistical solution for analyzing data with pseudoreplication, by separating the different sources of variability and thereby generating correct inferences from data gathered in studies with constraints in randomization.

Sergentomyia (Sergentomyia) pondicheriiensis is a new species of phlebotomine sand fly belonging to the genus Sergentomyia and subgenus Sergentomyia is described with illustrations of adult females and males. Other six species under the subgenus Sergentomyia are Se. (Ser.) punjabensis (Sinton), Se. (Ser.) dentata (Sinton), Se. (Ser.) theodori (Parrot), Se. (Ser.) murgabiensis (Perfiliew), Se. (Ser.) mervynae Pringle, and Se. (Ser.) fallax afghanica Artemiev. Sergentomyia pondicherriensis sp. nov was collected from termite mounds, in one of the villages of Puducherry Union Territory, Southern India. A revised key to the species of the subgenus Sergentomyia is also included.

This study examined the effects of different life stages (first, second, third, fourth, and five instars; adult females and adult males) and feeding regimes (starved and blood fed) on the active movement activity of the tropical bed bug, Cimex hemipterus (F.), under mixed-stage conditions, We used an extended arena made from Tygon tube coils and observed the movement frequency and movement distance at selected time intervals up to 120 h. The fifth instars and adult males and females showed significantly (P < 0.01) greater movement frequency compared with the other stages. The first and second instars showed limited movement (<8 m) over the experimental period. Starved bed bugs showed greater movement frequency compared with blood-fed bed bugs, with the exception of adult females. Blood-fed adult females exhibited significantly (P < 0.01) greater movement frequency and distance compared with starved females. Blood-fed females moved up to 42.3 m over 120 h. Regression analysis between movement distance of the fifth instars and adults and the time intervals revealed a positive relationship (r² = 0.6583; P < 0.01), suggesting that delays in bed bug control efforts will increase the risk of the greater infestation. During bed bug inspection, the presence of only late instars and adults in premises would indicate a new infestation, whereas an established infestation likely would consist of mixed stages.

In the framework of a bluetongue surveillance program including clinical, serological, and entomological activities, Culicoides biting midges were light trapped weekly in two regions of central Italy, Lazio and Tuscany. In the period January 2002 through December 2005, 3,944 collections were carried out in 189 trap sites distributed in all the provinces of the two regions. Abundance data of C. obsoletus group were analyzed in relation to trap site altitude, distance from the sea, land use, and number of farmed animals. Species seasonality and overall temporal trend were also described. C. obsoletus was distributed over the whole study area, almost in all trapping sites and with high abundances. The species group was dominant among all captured Culicoides, with higher abundances recorded inland and in areas where land cover was partially or completely natural-wooded. Adults on the wing were caught all year round, with peaks in May–June and middle October. The observed trend through years recorded a peak during autumn 2002, in concomitance with a local epidemic of bluetongue.

The ecology of Phlebotomus sand flies in cutaneous leishmaniasis foci as a result of Leishmania tropica in the Judean Desert was studied. Between 2005 and 2007, > 265,000 specimens were trapped outdoors and 1,233 specimens were collected indoors. The catches included Phlebotomus sergenti Parrot, Phlebotomus papatasi (Scopoli), Phlebotomus syriacus Adler & Theodor, and Phlebotomus tobbi Adler & Theodor. P. sergenti, the local vector of Leishmania tropica, comprised 90% of outdoor catches, and relatively few were caught indoors. Conversely, P. papatasi were >90% of the indoor collections, and only a few were caught outdoors. The efficiency of trapping methods varied, but species composition and sex ratio remained constant irrespective of method. Sand flies were abundant on slopes facing east where wind velocity was low, and scarce on slopes facing west and residential areas. Large numbers and high proportion of males that occur near breeding sites were found in man-made rock walls and in rock crevices on slopes of uncultivated hills. Population increase began in April, was more intensive between May and November, peaked in August–September, and significantly decreased in December. Indoors, most of the P. sergenti (<80%) were collected from September to November. A few sand flies were found between January and March. The effects of climatic factors and human activities on sand fly populations and the risk of Leishmania infections are discussed.

Understanding on the chemical ecology of mosquito behavior is of paramount importance in developing control programs employing attractants and repellents, Several workers focused on topical repellents and oviposition attractants of mosquitoes, however, only limited work has been accomplished on mosquito oviposition repellents, The present systematic investigation provides evidence on the effectiveness of a C₂₁ fatty acid ester- hexadecyl pentanoate, to stimulate antennal olfactory receptors of Aedes aegypti (L.), Ae. albopictus (Skuse), and Anopheles stephensi (Liston) that mediate their long-range olfaction guided flight orientation behavior by repelling the gravid females of these mosquito vectors in the olfactometer. The compound loaded onto an effervescent tablet retained its repellent property in the treated substrates for up to 1 wk at 10 mg/L. In places, where the mosquito breeding habitats are near to human habitations, could be treated with hexadecyl pentanoate to repel the ovipositing gravid females as a component of the integrated approach for mosquito management by disrupting the mosquito life cycle and population growth.

Females of the African malaria vector, Anopheles gambiae Giles sensu stricto, use human volatiles to find their blood-host. Previous work has shown that ammonia, lactic acid, and aliphatic carboxylic acids significantly affect host orientation and attraction of this species, In the current study, these compounds were tested for their attractiveness relative to human emanations in vivo and in vitro. Emanations from a human hand, incubated sweat, and foot skin residues on a nylon sock were significantly attractive when tested against clean air. In a dual-choice test, foot skin residues were significantly more attractive than emanations from a human hand in vivo. Ammonia alone attracted more mosquitoes than fresh or incubated sweat, However, the odor of a human hand or of foot skin residues were more attractive than ammonia. A known attractive blend of ammonia with lactic acid and carboxylic acids was less effective than natural foot odorants, The results demonstrate that the synthetic blend based on skin odor is attractive for An. gambiae, but that in a choice situation in vitro natural skin odors are still preferred by the mosquito. Differences in volatile organic compound abundances between a worn sock and the synthetic blend may have resulted in stronger attraction to the sock. This suggests that candidate attractants should be evaluated with consideration of the strength of natural odorant sources, The data furthermore suggest that additional unidentified compounds from the human foot are involved in the host-seeking behavior of this mosquito species.

The Southern house mosquito, Culex quinquefasciatus, is an important human health pest as a vector of several pathogens, including agents of lymphatic filariasis and arboviruses like West Nile virus. We conducted preliminary experiments in Recife, Brazil, to explore applications of Culex oviposition attractants in combination with Bacillus thuringiensis variety israelensis (Bti) in an attract-and-kill approach. Simple, cost-effective oviposition traps, BR-OVT, loaded with Bti and baited with or without attractant, were deployed in 10 homes for 30 d in 2 consecutive yr. Significantly higher numbers of egg rafts were deposited in traps baited with skatole or infusion than the control water traps. In the first year, 2006, significantly higher numbers of eggs were deposited in infusion-baited traps, particularly in the first 15 d of the experiment, than in skatole traps, but in the following year no significant difference was observed between synthetic and natural attractants. The tests strongly demonstrate that skatole or infusion can be used to enhance the number of egg rafts deposited on Bti-treated oviposition traps.

Bovine blood inoculated with bacteria isolated from screwworm [Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae) ]-infested animal wounds was tested as an attractant for oviposition for gravid screwworms. Eight species of gram-negative coliform (Enterobacteriaceae) bacteria mixed with bovine blood singly or all species combined and incubated for various times produced volatiles that attracted gravid flies in a cage bioassay in varying numbers. In 15-min duration tests, volatiles from five species of bacteria (Klebsiella oxytoca, Proteus mirabilis, Proteus vulgaris, Providencia rettgeri, and Providencia stuartii) attracted more females than volatiles of the three species (Enterobacter cloacae, Enterobacter sakazakii, and Serratia liquefaciens). In 1-h duration oviposition tests, volatiles from the substrate using the same five species of bacteria attracted more females to oviposit than the other three species. Volatiles from 24-h incubation period elicited least attraction and oviposition whereas volatiles from the 48- and 72-h incubation period resulted in significantly more attraction and oviposition. Attraction and oviposition decreased significantly when the substrates were incubated for 96 h. Volatiles from substrate with all species of bacteria combined attracted a significantly higher percentage of flies to land and oviposit than those from substrates prepared with single species. It is possible that multiple active chemicals present in volatiles of the all-species substrate may act as synergists resulting in greater response than those observed with volatiles from single-species substrate. Before oviposition flies took a bloodmeal from the oviposition substrate. It is possible that the oviposition is moderated by two different factors in screwworm—first, by using a chemical cue to land on a potential oviposition site and second, by using a bloodmeal to stimulate oviposition.

The increasing usage of long-lasting insecticide-treated nets allows protection of millions of people from malaria infection. Monitoring studies should be planned during any wide-scale malaria control program integrating insecticide-treated materials, to evaluate their effects and effectiveness on epidemiologically relevant parameters. Such operational control interventions may be challenged by insecticide resistance spread within vector populations, as a result of wide insecticide pressure. A nationwide distribution of long-lasting insecticidal nets was implemented throughout Niger in 2005. We studied the population genetic structure of major malaria vectors across Nigerien Sahel, and investigated potential effects of this large malaria control intervention. Wild-caught Anopheles gambiae sensu lato females from seven villages and two wet seasons were genotyped at 12 microsatellite loci. The genetic diversity within both species appeared homogenous between villages and years. The estimated genetic differentiation among samples was very low within both species, indicating high gene flow across the area. An absence of differentiation was also found between 2005 and 2006 wet seasons, for all samples but one, showing that the net distribution did not impact significantly the genetic diversity and structure of vector populations in a single year. We provide valuable results participating to document effects of large malaria control programs, to maximize the efficiency of available tools in future interventions.

Immature mosquito development and survival of adults are highly sensitive to environmental temperature, which can alter gene expression during the mosquito life-cycle. To further understand how heat shock proteins are developmentally expressed in mosquitoes, we subjected first instar larvae, 16-h old pupae and female of Aedes aegypti (L.) (Diptera: Culicidae) to heat shock treatment for 0, 15, 30, 60, and 180 min at 23 and 42°C. The heat shock protein genes AeaHsp26, AeaHsp83, and AeaHsc70 were examined by comparing relative transcript expression levels at 42°C compared with 23°C. Upregulated transcripts from heat shock treatment at 42°C and control were further confirmed and quantified by quantitative real-time polymerase chain reaction. Data revealed that first instar larvae were more sensitive to heat shock treatment than pupae and adults (i.e., relative AeaHsp26 expression levels in larvae were 10-fold greater than in the females. AeaHsp83 expression levels in larvae, pupae and adults were upregulated 2- to 50-fold greater by heat shock treatment at 42°C compared with 23°C. AeaHsc70 expression levels in larvae, pupae and adults, however, were upregulated less than AeaHsp26 and AeaHsp83 at the higher temperature. Statistical analysis indicated that AeaHsp26 and AeaHsp83 genes were significantly upregulated in Ae. aegypti larvae and pupae after 15, 30, 60, and 180 min exposure to high temperature (42°C). The current study has shown that AeaHsp26 and AeaHsp83 are important markers of stress and may function as critical proteins to protect and enhance survival of Ae. aegypti larvae and pupae.

Saliva of blood-sucking arthropods contains a complex mixture of peptides that affect their host's hemostasis, inflammation, and immunity. These activities can also modify the site of pathogen delivery and increase disease transmission. Saliva also induces hosts to mount an antisaliva immune response that can lead to skin allergies or even anaphylaxis. Accordingly, knowledge of the salivary repertoire, or sialome, of a mosquito is useful to provide a knowledge platform to mine for novel pharmacological activities, to develop novel vaccine targets for vector-borne diseases, and to develop epidemiological markers of vector exposure and candidate desensitization vaccines. The mosquito Ochlerotatus triseriatus is a vector of La Crosse virus and produces allergy in humans. In this work, a total of 1,575 clones randomly selected from an adult female O. triseriatus salivary gland cDNA library was sequenced and used to assemble a database that yielded 731 clusters of related sequences, 560 of which were singletons. Primer extension experiments were performed in selected clones to further extend sequence coverage, allowing for the identification of 159 protein sequences, 66 of which code for putative secreted proteins. Supplemental spreadsheets containing these data are available at  http://exon.niaid.nih.gov/transcriptome/Ochlerotatus_triseriatus/S1/Ot-S1.xls and  http://exon.niaid.nih.gov/transcriptome/Ochlerotatus_triseriatus/S2/Ot-S2.xls.

The control efficacy of clove, Eugenia caryophyllata, and eucalyptus, Eucalyptus globulus, essential oils and 15 formulations containing these essential oils alone (8, 12, and 15% sprays) and their binary mixtures (7:3, 5:5, and 3:7 by weight) against adult females of insecticide-susceptible KR-HL and dual malathion - and permethrin-resistant BR-HL strains of head louse, Pediculus humanus capitis (De Geer), was examined by using contact plus fumigant and human hair wig (placed over the head of mannequin) mortality bioassays. In contact plus fumigant mortality bioassay, essential oils from eucalyptus (0.225 mg/cm²) and clove (1.149 mg/cm²) were less effective than either d-phenothrin (0.0029 mg/cm²) or pyrethrum (0.0025 mg/cm²) based on 6-h median lethal concentration values. However, the efficacies of eucalyptus and clove oils were almost identical against females from both strains, despite high levels of resistance of the BR-HL females to d-phenothrin (resistance ratio, 667) and pyrethrum (resistance ratio, 754). In human hair wig mortality bioassay, eucalyptus oil spray treatment gave better control efficacy than either spray treatment with clove oil alone or their binary mixtures, Thus, eucalyptus applied as 8% sprays (15 or 20 ml) appears to provide effective protection against pediculosis even to insecticide-resistant head louse populations. Once the safety issues resolved, covering the treated hair and scalp with bath shower cap or hat would ensure the fumigant action of the essential oil.

Aedes aegypti (L.) (Diptera: Culicidae) the primary vector of dengue viruses (DENV1–4), oviposit in and around human dwellings, including sites difficult to locate, making control of this mosquito challenging. We explored the efficacy and sustainability of Aedes Densonucleosis Virus (AeDNV) as a biocontrol agent for Ae. aegypti in and among oviposition sites in large laboratory cages (>92 m³) as a prelude to field trials. Select cages were seeded with AeDNV in a single oviposition site (OPS) with unseeded OPSs established at varied distances. Quantitative real-time polymerase chain reaction was used to track dispersal and accumulation of AeDNV among OPSs. All eggs were collected weekly from each cage and counted. We asked: (1) Is AeDNV dispersed over varying distances and can it accumulate and persist in novel OPSs? (2) Are egg densities reduced in AeDNV treated populations? AeDNV was dispersed to and sustained in novel OPSs. Virus accumulation in OPSs was positively correlated with egg densities and proximity to the initial infection source affected the timing of dispersal and maintenance of viral titers. AeDNV did not significantly reduce Ae. aegypti egg densities. The current study documents that adult female Ae. aegypti oviposition behavior leads to successful viral dispersal from treated to novel containers in large-scale cages; however, the AeDNV titers reached were not sufficient to reduce egg densities.

Hematophagous parasites such as leeches, ticks, mites, lice, bedbugs, mosquitoes, and myiasis-producing fly larvae are common health problems in Lao People's Democratic Republic. Several arthropod-borne infections, e.g., malaria, dengue fever, and Japanese encephalitis, are endemic there. Effective vector control methods including the use of pesticides, insecticide-treated bed nets, and synthetic and plant-based repellents are important means of control against such invertebrates and the pathogens they may transmit or directly cause. In this study, we documented traditional knowledge on plants that are used to repel or kill hematophagous arthropods, including mosquitoes, bedbugs, human lice, mites and ticks, fly larvae, and blood-sucking leeches. Structured interviews were carried out in 66 villages comprising 17 ethnic groups, covering a range of cultures, throughout Lao People's Democratic Republic. A total of 92 plant species was recorded as traditional repellents (including plants for pesticidal usages) in 123 different plant-ectoparasite combinations. The number and species of plants, and animal taxa repelled (or killed) per plant species differed per region, village, and ethnic group. Traditional use was confirmed in the scientific literature for 74 of these plant species, and for an additional 13 species using literature on closely related species. The use of botanical repellents and pesticides from many plant species is common and widespread in the Lao countryside. In the future, the identification of the active components in certain plants to develop more optimal, inexpensive repellents, insecticides, acaricides, or antileech compounds as alternatives to synthetic repellents/pesticides against blood-feeding insects, ticks, mites, and leeches is warranted.

From April 2007 to September 2008,1,793 adult and nymphal ixodid ticks were collected from 49 counties in Tennessee. Six species were identified, including Dermacentor variabilis (Say), Amblyomma americanum (L.), Ixodes texanus (Banks), Ixodes cookei Packard, Ixodes scapularis (Say), and Amblyomma maculatum Koch, from 13 medium- to large-sized mammalian hosts and dragging through vegetation. Raccoons were the most common vertebrate source (198 captures), accounting for 60% of ticks collected. Dermacentor variabilis was the predominant species from raccoons with a prevalence of 92% and mean intensity of 5.3. A. americanum was predominated in white-tailed deer and drags with respective mean intensities of 3.1 and 14.1 and prevalence values of 94%. All tick species were identified between April and August, coinciding with the majority of animal captures. Only A. americanum, I. texanus, and I. cookei were identified from 22 animal captures from November to March. I. texanus and I. cookei were more common in the eastern portions of the state, but this may be a result of higher raccoon captures in those areas. Only four specimens of I. scapularis were collected in this study, which may reflect the absence of small mammal or reptile captures. Two A. maculatum were collected, and we report new distribution records in Tennessee for this species. Despite unequal sampling among ecoregions, the large numbers of D. variabilis and A. americanum from multiple host species suggest their widespread distribution throughout the state. These species of ticks can transmit multiple pathogens, including spotted fever group rickettsiae and ehrlichiae.

Persistent West Nile virus (WNV) infection in the mosquito Culex quinquefasciatus Say (Diptera: Culicidae) is associated with pathological changes in the salivary glands, including apoptotic cell death and a corresponding reduction in virus transmission over time. The vector host response to WNV infection and the molecular basis of WNV pathogenesis in Cx. quinquefasciatus was investigated using oligonucleotide microarrays designed to detect differences in the salivary gland transcriptome between WNV-infected mosquitoes and uninfected controls. Transcripts with increased abundance in infected salivary glands included those related to immunity, transcription, protein transport and degradation, amino acid and nucleotide metabolism, signal transduction, and cellular detoxification. Microarray-based analysis detected a decrease in transcript levels of a Culex inhibitor of apoptosis gene (IAP-1) and a decrease in abundance of 11 transcripts encoding salivary gland proteins, Transcript levels for an endonuclease, a proline-rich mucin, and several D7 protein family members also decreased. Transcripts with the greatest change in abundance during infection had either no similarity to sequences found in GenBank, VectorBase, and FlyBase, or were similar to sequences with uncharacterized protein products. These transcripts represent exciting targets for future analysis. Results from this study suggest that WNV infection influences transcriptional changes in an invertebrate host target tissue that may confer an advantage to the replicating virus, induce a host defense response, and alter the composition of vector saliva. The ramifications of these changes are discussed in terms of mosquito vector competence and WNV pathogenesis.

Alphaviruses (Togaviridae) have rarely been found to persist for long in the adult insects that serve as their vectors. The ectoparasitic swallow bug (Hemiptera: Cimicidae: Oeciacus vicarius Horvath), the vector for Buggy Creek virus (BCRV; Togaviridae, Alphavirus), lives year-round in the mud nests of its host, the cliff swallow (Petrochelidon pyrrhonota Vieillot). We measured the prevalence of BCRV in swallow bugs at sites with cliff swallows present and at the same sites after cliff swallows had been absent for 2 yr. We collected bugs directly from cliff swallow nests in the field and screened bug pools with BCRV-specific real-time-polymerase chain reaction (RT-PCR) and plaque assay. At two colony sites last occupied by birds 2 yr earlier, we found 12.5 and 55.6% of bug pools positive for BCRV RNA by RT-PCR, Infection rates (per 1,000 bugs) for these sites were 1.32 and 7.39. RNA prevalence in the unfed bugs was not significantly different from that in fed bugs 2 yr earlier at the same sites. The RNA-positive samples from unfed bugs failed to yield cytopathic BCRV by Vero-cell plaque assay. However, viral RNA concentrations did not differ between unfed bugs and bugs at active sites, and over 84% of positive bug pools were cytopathic to Vero cells 4–5 wk later, after cliff swallows moved into one of the colony sites. These data demonstrate the persistence of potentially infectious BCRV in unfed swallow bugs for at least 2 yr in nature.

To evaluate the vectorial capacity of mosquitoes for viruses in Japan, the host-feeding habits of the mosquitoes were analyzed by sequencing polymerase chain reaction-amplified fragments of the cytochrome b and 16S ribosomal RNA regions of the mitochondrial DNA of 516 mosquitoes of 15 species from seven genera that were collected from residential areas during 2003–2006. Culex pipiens L. and Aedes albopictus Skuse were the most commonly collected species in urban and suburban residential areas. Anautogenous Culex pipiens pallens Coquillett was distinguished from the autogenous Cx. pipiens form molestus Forskal using a polymerase chain reaction-based identification method. Both Cx. p. pollens and Cx. p. form molestus exhibited similar host-feeding habits, broadly preferring avian (50.0 and 42.5% of avian, respectively) and mammalian (38.6 and 45.0% of avian, respectively) hosts, such as tree sparrows, ducks, and humans. Conversely, Ae. albopictus exhibited a highly mammalophilic and anthropophilic feeding pattern, with 84.2% feeding on mammalian hosts and 68.5% of these on humans. We concluded that in Japan, Cx. pipiens might play a significant role in the avian-to-mammal transmission of viruses, such as West Nile virus, whereas Ae. albopictus might play a role in the human-human transmission of dengue and Chikungunya viruses.

West Nile virus is a pathogen of concern for both human and wildlife health, Although many aspects of the ecology of West Nile virus are well understood, the mechanisms by which this and similar mosquito-borne viruses overwinter and become reinitiated each spring in temperate regions is not known. A thorough understanding of this mechanism is crucial to risk assessment and development of control strategies. One of the hypotheses to explain the mechanism by which this virus persists from year to year is the spring recrudescence of latent virus in avian reservoir hosts. Stress-related immunosuppression is implicated in the recrudescence of latent viruses in birds. We tested the spring recrudescence hypothesis in a controlled laboratory experiment using hatching-year gray catbirds (Dumatella carolinensis) captured in northern Ohio (July–August 2006), Catbirds (n = 60) were experimentally infected (September 2006) and later examined for the effects of immuno-suppression through exogenous hormones and artificially induced migratory disposition, We found no effect of either testosterone or migratory behavior on infection status in any of the treatment birds. Moreover, we detected no viral RNA in the kidney, spleen, brain, or liver upon necropsy at 24 wk postinfection.

The objective of this study was to assess whether the distribution of Borrelia burgdorferi, the causative agent of Lyme disease, and its vector tick Ixodes scapularis Say (Acari: Ixodidae) across Indiana is influenced by large-scale landscape features, specifically the proportion of forest within the surrounding landscape and the distance to water features such as lakes and major streams, Hunter-killed deer were checked for ticks in designated check-stations in the opening firearm hunting season between 2005 and 2007. Hunting locations for ≈3,600 deer were used in ArcGIS (ESRI, Redlands, CA) to examine the influence of forest and water features in the surrounding area on the occurrence of the tick and bacteria. In total, 82 of the 92 (89%) Indiana counties were sampled from 2005 to 2007. The proportion of tick-infested deer was 13.6, 15.8, and 25.5% in these years, respectively, There was a significant nonlinear response for I. scapularis to forest cover in 1 yr that indicated a greater probability of this tick presence at intermediate levels of forest area, Infested does were harvested in significantly more forested areas than bucks. No significant correlation was found neither between levels of forest area and B. burgdorferi nor between water bodies and both organisms.

Six viral isolates were obtained from 23,243 female mosquitoes (examined in 513 pools) belonging to 16 species and collected along the lower reaches of the Dyje River in South Moravia (Czech Republic, central Europe) during 2006–2008: five isolates of Orthobunyavirus Ťahyňa (TAHV, California group, family Bunyaviridae: three isolations from Aedes vexans (Meigen), one from Ae. sticticus (Meigen), one from Culex modestus Ficalbi); and one isolation of Flavivirus West Nile (WNV, Japanese encephalitis group, family Flaviviridae)-strain Rabensburg (proposed lineage 3 of WNV) from Ae. rossicus (Dolbeshkin et al). All viral isolates were recovered from mosquitoes collected in 2006 (15,882 mosquitoes examined). while no virus was isolated from mosquitoes trapped in 2007 and 2008, when 1,555 and 5,806 mosquitoes were examined, respectively. The population density of local mosquitoes was very low in 2007 and 2008 because of warm and dry summer including a considerably low water table, compared with environmental conditions favorable for mosquito development in 2006. The virus isolation procedure was based on intracerebral inoculation of newborn mice. In parallel, more than one-third of the samples (183 pools consisting of 8,470 individual mosquitoes) were also examined by inoculating Vero cell cultures in Leighton tubes. However, the latter method detected only three of the six virus isolates (including WNV-Rabensburg). Ae. rossicus is a new potential vector for WNV-Rabensburg. This species feeds mostly on mammals including man; this raises the question whether this virus lineage is not adapted to an alternative mosquito-mammal cycle in the South-Moravian natural focus.

In this study, we evaluated Amblyomma americanum (lone star tick) in Mississippi for the presence of Ehrlichia chaffeensis, causative agent of human monocytic ehrlichiosis; Ehrlichia ewingii, causative agent of human and canine granulocytic ehrlichiosis; Borrelia lonestari, putative agent of southern tick-associated rash illness; Francisella tularensis, the agent of tularemia; and Rickettsia spp., particularly R. amblyommii, a suspected pathogen, We collected adult A. americanum from four regions of Mississippi: Northeast, Northwest, Southeast, and East. Of the ticks collected, 192 were dissected and DNA was extracted for nested polymerase chain reaction (PCR) assays to detect the above bacteria. In all, 3% of tick extracts had evidence of Borrelia sp., 4% for E. chaffeensis, 6% for E. ewingii, and 44% for a Rickettsia species. As determined by sequencing, most Rickettsia spp, were R. amblyommii. In addition, extracts from 42 pools (total of 950) of larval A. americanum collected in Southwest Mississippi were tested for the presence of E. chaffeensis and Rickettsia species. Of these extracts from pools, nine of 37 (24%) were PCR positive for a Rickettsia sp., most often, R. amblyommii; none had evidence of E. chaffeensis, supporting the ability of lone star ticks to transovarially transmit R. amblyommii, but not E. chaffeensis. This study demonstrates E. chaffeensis, E. ewingii, “B. lonestari”, and R. amblyommii in A. americanum by PCR for the first time in Mississippi. Understanding the prevalence and epidemiology of these agents in Mississippi should increase awareness of tick-borne disease in the medical community.

The analysis of immunoglobulin (Ig) M and IgG serum isotypes as indicators of the presence of Oestrus ovis developmental stages was carried out. A serological survey to discover the seasonal variations in the prevalence of oestrosis in sheep from an oceanic climate area was developed. Six hundred and sixty-nine blood samples were analyzed for the presence of IgM and IgG using an enzyme-linked immunosorbent assay and O. ovis second-stage larvae excretory/ secretory antigens. In addition, the effect of an ivermectin-based treatment on the humoral immune response was measured. The percentage of positive animals was 54% for IgM and 55% for IgG. The highest percentages of sheep positive to IgM were recorded in the winter and to IgG in the summer. No animals positive to IgM were detected in the ivermectin-treated sheep. These results seem to underline that oestrosis is a parasitic disease to keep watch for in regions with an oceanic climate. A noninvasive evaluation of the success of chemotherapy seems possible by analyzing the serum IgM response.

For the carrion-feeding blow flies, a common source of vitellogenic protein are the carcasses that also serve as larval substrates. This attraction continues well after the blow flies have ceased to oviposit on the carrion, and it has been assumed that a decomposed cadaver remains a suitable source for vitellogenic protein; however, this assumption has never been tested. To test this hypothesis, blow flies (Lucilia sericata Meigen) were supplied with either fresh or decomposed beef liver as protein sources for vitellogenesis for 5 d. Both treatments produced identical ovarian development, indicating that decomposed flesh remains a suitable protein source for carrion-feeding blow flies. These results support the theory that virgin flies are attracted to cadavers of advanced decomposition as a source of protein.

We report a rare case of myiasis caused simultaneously by three dipterous species. A 54 yr-old indigent patient was admitted to Andaraí Hospital with painful eruptions on the scalp. The parieto-occipital sulcus showed two lesions caused by scratching associated with deep, odoriferous and exudative pediculosis. Larvae removed with the help of forceps and vaseline produced 153 adults, identified in the laboratory as 114 specimens of Chrysomya megacephala (F., 1794), 38 of Sarcophaga (Liopygia) ruficornis (F., 1794), and one of Musca domestica (L., 1758).

The calliphorid fly, Lucilia cuprina (Wiedemann), is known to cause serious malign myiasis in animals, whereas its sibling species Lucilia sericata (Meigen) is commonly a carrion breeder and is used in maggot debridement therapy (MDT). The current study reports an accidental involvement of L. cuprina in MDT in Alexandria, Egypt, that has proved to be safe and effective. In November 2008, the laboratory colonies of L. sericata (the species regularly used in MDT) at the Faculty of Science, Alexandria University were renewed by Lucilia flies collected as third instar larvae on exposed rabbit carcasses. Flies from the new colonies were successfully used to heal the diabetic foot wounds of two patients at Alexandria Main University Hospital. Analysis of DNA sequences and adult and larval morphology then revealed that these flies were and still are L. cuprina. Breeding of this species in carrion in Alexandria is a new record. Despite the safety of this strain of L. cuprina in MDT, entomologists rearing blow flies for the purpose of wound debridement should regularly maintain high quality assurance of their species' identity to avoid possible clinical complications that may result from the introduction of an unexpected and invasive species to their laboratory colonies.