Ten Years of Mediterranean Monk Seal Stranding Records in Greece under the Microscope: What Do the Data Suggest?

This paper presents the results of an analysis of stranding events of the Mediterranean monk seal Monachus monachus over a decade. The analysis involved categorization according to the cause of stranding and seasonality, the identification of hotspot stranding areas and an assessment of possible correlations between stranding events and environmental/climatic patterns using time series analysis. Moreover, Generalized Additive Models (GAMs) were applied to explore the effects of the size of small-scale fishing grounds, the number of species sightings, and the occurrence of reproduction sites on "human-related" strandings. Finally, special focus was put on the central part of the eastern Ionian Sea for the assessment of stranding hotspot areas by means of the Multi-Criteria Decision Analysis (MCDA) approach, based on different kinds of spatial information such as anthropogenic pressures and the location of breeding sites and feeding grounds. Time series analysis results revealed that oscillation indices, during the first half of the year, and sea surface temperature (SST) in the Mediterranean from October to December were positively correlated with monk seal stranding events. GAMs underlined that areas combining extended small-scale fishery grounds and a higher number of sightings were more likely to cause more strandings. Regarding spatial analyses, the central Aegean Sea was highlighted as a hotspot for "human-related strandings", while the MCDA approach emphasized that the southern coasts of Cephalonia and the gulf between Lefkada and mainland Greece were susceptible to subadult strandings.

Where does Aedes albopictus (Diptera: Culicidae) really breed in a Mediterranean residential area? Results from a field study in Valencia, Eastern Spain.

Since its introduction in Spain in 2004, Aedes albopictus has rapidly spread across the country. Its aggressive biting behaviour causes nuisance, limiting outdoor activities. Also, its role as a vector of several arboviruses implies a major public health risk, with several cases of autochthonous dengue having been reported nationwide over the past few years. Control strategies usually focus on interventions in breeding sites. As such, accurate knowledge of the main larval habitats becomes a major priority in infested areas. A detailed identification of breeding sites of Ae. albopictus was carried out in the outdoors of 60 residential properties during July-August 2022 in El Vedat de Torrent (Valencia, Eastern Spain), an area recently colonised by this species. A total of 1444 real and potential breeding sites were examined. The most abundant potential larval habitat were plant pot plates (6.48 units/house), although a low infestation level was found, both for larvae (2.06% positivity, x̄ = 30.5 larvae/container), and pupae (0.51%, x̄ = 2.5 pupae/container). A total of 7715 larvae and 205 pupae were found in a disused flooded water pool depuration system. Animal drinkers, buckets and irrigation water containers were found to be the most common positive containers. No statistical difference was observed among the different container materials. A general statistical increase of 1 larva per 11.7 ml of water in breeding sites was detected. Breeding sites of other species such as Culex pipiens (n = 2) and Culex modestus (n = 1) were also rarely found in this residential area. To our knowledge, this is the first aedic index study carried out in Europe, and it provides valuable information about the main domestic breeding habitats of Ae. albopictus, which can greatly improve control programmes.

Physico-chemical characterization of Anopheles gambiae s.l. breeding sites and kdr mutations in urban areas of Cotonou and Natitingou, Benin.

BACKGROUND: This study aimed to investigate the relationship between the physicochemical characteristics of An. gambiae s.s. and An. coluzzii breeding sites, the susceptibility profiles to commonly used insecticides in public health, and the underlying insecticide resistance mechanisms. METHODS: Anopheles breeding sites surveys were conducted in Cotonou and Natitingou in September 2020, January and August 2021. Physicochemical properties and bacterial loads were determined in individual breeding sites. The WHO susceptibility assays were carried out using the female of the emerging adult mosquitoes. Anopheles species were identified through PCR techniques. Kdr L1014F/S, N1575Y and G119S mutations were investigated using TaqMan genotyping assays. RESULTS: Molecular analysis showed that all mosquitoes analyzed in Cotonou were Anopheles coluzzii, while those of Natitingou were Anopheles gambiae s.s. Fecal coliforms were identified as playing a role in this distribution through their significant influence on the presence of An. coluzzii larvae. WHO susceptibility assay indicated a high level of resistance to deltamethrin in the two cities. The resistance levels to deltamethrin were higher in Cotonou (X2 = 31.689; DF = 1; P < 0.0001). There was a suspected resistance to bendiocarb in Cotonou, whereas the mosquito population in Natitingou was resistant. The kdr L1014F mutation was highly observed in both mosquito populations (frequence: 86-91%), while the Ace-1 mutation was found in a small proportion of mosquitoes. In Cotonou, salinity was the only recorded physicochemical parameter that significantly correlated with the resistance of Anopheles mosquitoes to deltamethrin (P < 0.05). In Natitingou, significant correlations were observed between the allelic frequencies of the kdr L1014F mutation and pH, conductivity, and TDS. CONCLUSION: These results indicate a high level of pyrethroid resistance in the anopheles populations of both Cotonou and Natitingou. Moreover, this study report the involvement of abiotic factors influencing Anopheles susceptibility profile.

Diversity and Distribution of Culicidian Fauna in Urban and Rural Areas of Setif Region (North-East Algeria).

Objective: The study explored larval mosquito breeding sites in urban and rural areas from March 2021 to February 2022 in the North-East region of Algeria to provide information on larval biology and ecology of different mosquito species. Methodology: It focused on the effect of the physicochemical characteristics of the breeding sites (pH and water temperature), area, and months on the larval mosquito abundance. Diversity index and equitability were calculated for the registered mosquito species in both areas. Results: Months significantly affected the larval mosquito abundances, with a clear abundance increase from March to August 2021, and then, a clear decrease to February 2022. The interactions between rural and urban areas, water pH, and water temperature as well affected the mosquito abundance. Twelve species of Culicidae were identified and distributed in three genera: Culex, Culiseta, and Aedes. Culex pipiens was the most dominant species in rural and urban sites, with a rate of 69% and 48%, respectively. In rural site, Aedes caspius and Aedes vexans occupied the second position with value of 7-8% each. In urban sites, the second position was occupied by Culiseta longiareolata (36%) and then by Culex theileri (10%). In terms of site diversity, only one site was highly diversified in rural sites, El Hchichia (H' = 1.12 bits). In urban sites, Setif Center, Guellel an Ain Trick, had almost similar diversity with values between 1.03 and 1.04 bits. Equitability values were 0.48 in rural sites and 0.55 in urban sites. Therefore, the Culicidae species presented in these study sites are moderately balanced. Conclusion: The present study provides key tools for planning a better vector control through the information on the effect of some factors on the proliferation of mosquitoes in the urban and rural sites of the Setif region and on the Culicidae diversity and abundance.

Profiles of bacterial communities and environmental factors associated with proliferation of malaria vector mosquitoes within the Kenyan Coast.

Background: Since Anopheles mosquitoes which transmit and maintain the malaria parasite breed in the outdoor environment, there is an urgent need to manage these mosquito breeding sites. In order to elaborate more on the ecological landscape of mosquito breeding sites, the bacterial community structure and their interactions with physicochemical factors in mosquito larval habitats was characterised in Kwale County (Kenya), where malaria is endemic. Methods: The physical characteristics and water physicochemical parameters of the habitats were determined and recorded. Water samples were also collected from the identified sites for total metagenomic DNA extraction in order to characterise the bacterial communities within the breeding sites. Results and Discussion: Sites where mosquito larvae were found were described as positive and those without mosquito larvae as negative. Electrical conductivity, total dissolved solids, salinity and ammonia were lower in the rainy season than in the dry season, which also coincided with a high proportion of positive sites. Pseudomonadota was the most common phyla recovered in all samples followed by Bacteroidota and then Actinomycetota. The presence or absence of mosquito larvae in a potential proliferation site was not related to the bacterial community structure in the sampled sites, but was positively correlated with bacterial richness and evenness. Conclusion: Generally, the presence of Anopheles mosquito larvae was found to be positively correlated with rainy season, bacterial richness and evenness, and negatively correlated with electrical conductivity, total dissolved solids, salinity and ammonia. The findings of this study have implications for predicting the potential of environmental water samples to become mosquito proliferation sites.

Genotype × Environment Interaction and Stability Analysis of Selected Cassava Cultivars in South Africa.

Cassava (Manihot esculenta Crantz) is an important root crop worldwide. It is adapted to a wide range of environmental conditions, exhibiting differential genotypic responses to varying environmental conditions. The objectives of this study were: (1) to examine the effect of genotype, environment and genotype × environment interaction (GEI) on fresh root yield (FRY) and dry matter content (DMC); and (2) to identify superior genotypes that exhibit high performance for the traits of interest using the genetic tools of additive main effects and multiplicative interaction (AMMI) and genotype stability index (GSI) analysis. Eleven cassava genotypes were evaluated in a randomized complete block design at six trial sites in South Africa. The combined analysis of variance based on AMMI revealed significant genotype, environment and GEI for the traits. The percentage variation due to GEI was higher than the percentage variation due to genotype for FRY, reflecting differential genotypic responses across the experimental sites. The proportion of variance due to genotype variation was larger for DMC. Genotype stability index (GSI) showed that UKF3 (G6), 98/0002 (G2) and P4/10 (G5) were the highest yielding and most stable genotypes for FRY, and 98/0002 (G1), UKF3 (G6) and UKF9 (G11) were the highest yielding and most stable genotypes for DMC. Cultivars 98/0002 and UKF3 were identified as providing high stability with superior fresh root yield and DMC. These genotypes could be recommended to farmers for food, feed and industrial applications without the need for further breeding. The AMMI-2 model clustered the testing environments into three mega-environments based on the winning genotypes for FRY and DMC. Mabuyeni (KwaZulu-Natal), Shatale (Mpumalanga) and Mandlakazi (Limpopo) would be the best testing sites in future cassava-genotype evaluation and breeding programs. This study provides a baseline for a future study on the GEI of cassava varieties, using a larger set of genotypes, factoring in seasonal variation.

Public health impact of the spread of Anopheles stephensi in the WHO Eastern Mediterranean Region countries in Horn of Africa and Yemen: need for integrated vector surveillance and control.

BACKGROUND: Anopheles stephensi is an efficient vector of both Plasmodium falciparum and Plasmodium vivax in South Asia and the Middle East. The spread of An. stephensi to countries within the Horn of Africa threatens progress in malaria control in this region as well as the rest of sub-Saharan Africa. METHODS: The available malaria data and the timeline for the detection of An. stephensi was reviewed to analyse the role of An. stephensi in malaria transmission in Horn of Africa of the Eastern Mediterranean Region (EMR) in Djibouti, Somalia, Sudan and Yemen. RESULTS: Malaria incidence in Horn of Africa of EMR and Yemen, increased from 41.6 in 2015 to 61.5 cases per 1000 in 2020. The four countries from this region, Djibouti, Somalia, Sudan and Yemen had reported the detection of An. stephensi as of 2021. In Djibouti City, following its detection in 2012, the estimated incidence increased from 2.5 cases per 1000 in 2013 to 97.6 cases per 1000 in 2020. However, its contribution to malaria transmission in other major cities and in other countries, is unclear because of other factors, quality of the urban malaria data, human mobility, uncertainty about the actual arrival time of An. stephensi and poor entomological surveillance. CONCLUSIONS: While An. stephensi may explain a resurgence of malaria in Djibouti, further investigations are needed to understand its interpretation trends in urban malaria across the greater region. More investment for multisectoral approach and integrated surveillance and control should target all vectors particularly malaria and dengue vectors to guide interventions in urban areas.

Comparative assessment of the bacterial communities associated with Anopheles darlingi immature stages and their breeding sites in the Brazilian Amazon.

BACKGROUND: The neotropical anopheline mosquito Anopheles darlingi is a major malaria vector in the Americas. Studies on mosquito-associated microbiota have shown that symbiotic bacteria play a major role in host biology. Mosquitoes acquire and transmit microorganisms over their life cycle. Specifically, the microbiota of immature forms is largely acquired from their aquatic environment. Therefore, our study aimed to describe the microbial communities associated with An. darlingi immature forms and their breeding sites in the Coari municipality, Brazilian Amazon. METHODS: Larvae, pupae, and breeding water were collected in two different geographical locations. Samples were submitted for DNA extraction and high-throughput 16S rRNA gene sequencing was conducted. Microbial ecology analyses were performed to explore and compare the bacterial profiles of An. darlingi and their aquatic habitats. RESULTS: We found lower richness and diversity in An. darlingi microbiota than in water samples, which suggests that larvae are colonized by a subset of the bacterial community present in their breeding sites. Moreover, the bacterial community composition of the immature mosquitoes and their breeding water differed according to their collection sites, i.e., the microbiota associated with An. darlingi reflected that in the aquatic habitats where they developed. The three most abundant bacterial classes across the An. darlingi samples were Betaproteobacteria, Clostridia, and Gammaproteobacteria, while across the water samples they were Gammaproteobacteria, Bacilli, and Alphaproteobacteria. CONCLUSIONS: Our findings reinforce the current evidence that the environment strongly shapes the composition and diversity of mosquito microbiota. A better understanding of mosquito-microbe interactions will contribute to identifying microbial candidates impacting host fitness and disease transmission.

Egg-laying by female Aedes aegypti shapes the bacterial communities of breeding sites.

BACKGROUND: Aedes aegypti, the main arboviral mosquito vector, is attracted to human dwellings and makes use of human-generated breeding sites. Past research has shown that bacterial communities associated with such sites undergo compositional shifts as larvae develop and that exposure to different bacteria during larval stages can have an impact on mosquito development and life-history traits. Based on these facts, we hypothesized that female Ae. aegypti shape the bacteria communities of breeding sites during oviposition as a form of niche construction to favor offspring fitness. RESULTS: To test this hypothesis, we first verified that gravid females can act as mechanical vectors of bacteria. We then elaborated an experimental scheme to test the impact of oviposition on breeding site microbiota. Five different groups of experimental breeding sites were set up with a sterile aqueous solution of larval food, and subsequently exposed to (1) the environment alone, (2) surface-sterilized eggs, (3) unsterilized eggs, (4) a non-egg laying female, or (5) oviposition by a gravid female. The microbiota of these differently treated sites was assessed by amplicon-oriented DNA sequencing once the larvae from the sites with eggs had completed development and formed pupae. Microbial ecology analyses revealed significant differences between the five treatments in terms of diversity. In particular, between-treatment shifts in abundance profiles were detected, showing that females induce a significant decrease in microbial alpha diversity through oviposition. In addition, indicator species analysis pinpointed bacterial taxa with significant predicting values and fidelity coefficients for the samples in which single females laid eggs. Furthermore, we provide evidence regarding how one of these indicator taxa, Elizabethkingia, exerts a positive effect on the development and fitness of mosquito larvae. CONCLUSIONS: Ovipositing females impact the composition of the microbial community associated with a breeding site, promoting certain bacterial taxa over those prevailing in the environment. Among these bacteria, we found known mosquito symbionts and showed that they can improve offspring fitness if present in the water where eggs are laid. We deem this oviposition-mediated bacterial community shaping as a form of niche construction initiated by the gravid female.

Impact of physicochemical parameters of Aedes aegypti breeding habitats on mosquito productivity and the size of emerged adult mosquitoes in Ouagadougou City, Burkina Faso.

BACKGROUND: Outbreaks of dengue fever caused by viruses transmitted by Aedes aegypti mosquitoes are repeated occurrences in West Africa. In recent years, Burkina Faso has experienced major dengue outbreaks, most notably in 2016 and 2017 when 80% of cases were recorded in Ouagadougou City (Central Health Region). In order to better understand the ecology of this vector and to provide information for use in developing control measures, a study on the characteristics of Aedes container breeding sites and the productivity of such sites, as measured by the abundance of immature stages and resultant adult body size, was undertaken in three health districts (Baskuy, Bogodogo and Nongremassom) of Ouagadougou. METHODS: Adult mosquitoes were collected indoors and outdoors in 643 households during the rainy season from August to October 2018. The presence of water containers was systematically recorded and the containers examined for the presence or absence of larvae. Characteristics of the container breeding sites, including size of the container and temperature, pH and conductivity of the water contained within, were recorded as well as the volume of water. Traditional Stegomyia indices were calculated as quantitative indicators of the risk of dengue outbreaks; generalised mixed models were fitted to larval and pupal densities, and the contribution of each covariate to the model was evaluated by the Z-value and associated P-value. RESULTS: A total of 1061 container breeding sites were inspected, of which 760 contained immature stages of Ae. aegypti ('positive' containers). The most frequent container breeding sites found in each health district were tyres and both medium (buckets/cans/pots) and large (bins/barrels/drums) containers; these containers were also the most productive larval habitats and the types that most frequently tested positive. Of the Stegomyia indices, the Breteau, House and Container indices exceeded WHO dengue risk thresholds. Generalised linear mixed models showed that larval and pupal abundances were associated with container type, physicochemical characteristics of the water and collection month, but there were significant differences among container types and among health districts. Aedes aegypti body size was positively associated with type and diameter of the container, as well as with electrical conductivity of the water, and negatively associated with pH and temperature of the water and with the level of exposure of the container to sunlight. CONCLUSION: This study provides data on putative determinants of the productivity of habitats regarding Ae. aegypti immature stages. These data are useful to better understand Ae. aegypti proliferation. The results suggest that identifying and targeting the most productive container breeding sites could contribute to dengue vector control strategies in Burkina Faso.

Report of Mosquito Vectors of Arboviruses from a Federal Conservation Unit in the Atlantic Forest, Rio de Janeiro State, Brazil.

Arbovirus infections, such as dengue, zika, chikungunya, and yellow fever, are a major public health problem worldwide. As the main vectors, mosquitoes have been classified by the Center for Disease Control and Prevention as one of the deadliest animals alive. In this ecological study, we analyzed the population dynamics of important genera and species of mosquito vectors. Mosquito immatures were collected using ovitraps and at natural breeding sites: bamboos and bromeliads. Adult mosquitoes were captured using CDC traps with CO2, Shannon traps, and manual suction tubes. Collections took place during the rainy and dry seasons from 2019 to 2020 in the Serra dos Órgãos National Park, Rio de Janeiro state, Brazil. The highest number of species was recorded in the ovitraps, followed by CDC and bromeliads. The breeding site with the lowest diversity was bamboo, though it showed the highest level of evenness compared to the other breeding sites. The medically important genera reported were Haemagogus spp., Aedes spp., Culex spp., and Wyeomyia spp. Culicid eggs increased in the rainy season, with a peak in November 2019 and January and February 2020, and lower abundance in the dry season, from September to October 2019. Mosquito eggs had a strong positive correlation (ρ = 0.755) with temperature and a moderate positive correlation (ρ = 0.625) with rainfall. This study shows how environmental variables can influence the ecology of disease-vector mosquitoes, which are critical in the maintenance of arbovirus circulation in a threatened biome within the most densely populated region of Brazil.

Anopheles stephensi: a guest to watch in urban Africa.

Malaria vector control programs in Sub-Saharan Africa have invested many efforts and resources in the control of eight-sibling species of Anopheles gambiae complex and An. funestus group. The behaviour of sibling species of these vectors is well known and used for implementing the current intervention tools. The reports of An. stephensi in urban Africa with different habitats breeding behaviour is an alert on the success of malaria vector control efforts achieved so far. This communication intends to give an insight on what should be considered as a challenge for the management of An. stephensi in urban Africa to retain the achievement attained in malaria control.

Host Plant Association and Distribution of the Onion Thrips, Thrips tabaci Cryptic Species Complex.

Onion thrips, Thrips tabaci Lindeman, 1889 (Thysanoptera: Thripidae) is a pest of economic importance traditionally treated as a polyphagous, cosmopolitan single species. Recent genetic evidence, however, suggests that it is rather a cryptic species complex of three lineages referred to by their host association and displaying different biological and ecological characteristics: leek-associated 1, leek-associated 2 and tobacco-associated. This study reviews host plant associations and distribution of the lineages of this cryptic species complex and discusses its consequences from an agronomical perspective. Overall, leek-associated 2 lineage has the broadest host range, including major crops from different plant families, and it is the only lineage with a confirmed worldwide distribution. Leek-associated 1 lineage shares some host plants with leek-associated 2. It is often found in Allium crops and its geographic distribution is limited to a few dozen countries. Finally, tobacco-associated lineage has only been collected from tobacco and their associated weeds in central and east Europe, and the Middle East. Additionally, this work presents a list of 391 plant species on which breeding and development of T. tabaci occurs, regardless of lineage. These host plant species belong to 64 different families, most importantly Asteraceae, Fabaceae, Brassicaceae, Poaceae, and Solanaceae.

A Survey on Native and Invasive Mosquitoes and Other Biting Dipterans in Northern Spain.

PURPOSE: Haematophagous Diptera, such as mosquitoes (Culicidae), biting midges (Ceratopogonidae), and black flies (Simuliidae), are important insects for public and animal health due to their capacity to bite and transmit pathogens. Outdoor recreation areas are usually affected by biting species and provide suitable habitats to both adult and immature stages. This study aimed to determine the species diversity and larval sites of these Diptera groups in two golf courses. METHODS: A multi-method collection approach using ultraviolet-CDC traps, human landing catches, collection in breeding sites, and ovitraps was implemented during summer 2020 in northern Spain. Insects were determined by morphological features accompanied by DNA barcoding. RESULTS: A total of ten native mosquito species were recorded either as adults or as larval stages. The invasive species Aedes japonicus was collected only at egg or pupa stage in ovitraps. Culex pipiens s.l. and Culex torrentium were both common mosquito species accounting for 47.9% of the total larval site collections and their larvae might be found in a wide range of natural and artificial sites. Culiseta longiareolata specimens were also prominent (30.1% of the total) and occurred exclusively in man-made water-filled containers. A total of 13 Culicoides species were identified, 10 of which were captured by ultraviolet-CDC traps, particularly members of the Obsoletus complex (Culicoides obsoletus/Culicoides scoticus, 74.9%) and seven species by emergence traps, being the two most abundant C. kibunensis (44.8%) and C. festivipennis (34.9%). Simulium cryophilum was also collected hovering around the operator under field sampling. CONCLUSION: A comprehensive representation of the blood-sucking Diptera fauna and their larval sites was obtained by the multi-method approach in two Spanish golf courses.

[Anopheles gambiae s.l. larval habitats and resistance mechanisms in Kribi, Cameroon]. / Habitats larvaires d'Anopheles gambiae s.l. et mécanismes de résistance à Kribi (Cameroun).

Background: The effectiveness of Long-Lasting Insecticidal Nets (LLINs) and indoor residual spraying (IRS) in controlling malaria vectors is hampered by the resistance of anopheles to insecticides. A good knowledge of the breeding sites and of the resistance profile of the vectors could facilitate the development of an appropriate control strategy. This study looks at the larval ecology and the susceptibility profile of An. gambiae s.l. to insecticides in urban and rural areas in Kribi, South Region of Cameroon. Methods: Mosquito breeding sites were categorized and geo-referenced. For each site, larvae were collected and reared and physicochemical parameters were measured in situ. The susceptibility of anopheles to dichlorodiphenyltrichloroethane (DDT), deltamethrin and permethrin, after pre-exposure to piperonyl butoxide (PBO) or not, was evaluated on the reared larvae. The Kdr mutation was detected using the Hot Oligonucleotide Ligation Assay (HOLA). Results: Natural breeding sites of An. gambiae s.l. were tyre tracks (12%, n=10), unbuilt wells (5%, n=4), pools of residual water (57%, n=48), foot and hoof prints, gullies, streams and the banks of the Kienké River (15%, n=13). Artificial breeding sites were abandoned dugouts (11%, n=9). Breeding sites in urban areas were characterized by higher mean values of temperature, conductivity, salinity and turbidity compared to the breeding sites in the rural area. The breeding sites of An. gambiae s.l. in urban Kribi were found to be sunnier than those in rural Kribi. A total of 4320 adults were used for testing, 1 440 mosquitoes from rural Kribi, 1 440 from urban Kribi and 1 440 specimens from the laboratory Kisumu strain. For DDT and deltamethrin, susceptibility tests showed that mortality was lower in a situation of no pre-exposure to PBO than in a situation of pre-exposure to PBO in the two study areas. The frequency of the resistant allele (R) was high for the Kdr West mutation in both urban (0.94) and rural areas in Kribi (0.93). Conclusion: An. gambiae s.l. colonizes a wide range of breeding sites and develops metabolic and mutation resistance to recommended insecticides. The search of alternative molecules for vector control is a necessity.

Characterization of sand fly breeding sites in district Malakand, Khyber Pakhtunkhwa, Pakistan, and evaluation of risk factors for cutaneous leishmaniasis in the region.

Present study was carried to determine the sand fly species composition, breeding sites ecology, seasonal abundance, and spatial distribution in district Malakand, Khyber Pakhtunkhwa, Pakistan. In addition, risk factors associated with cutaneous leishmaniasis (CL) were also evaluated. Survey of indoor and outdoor habitats was carried out using sticky traps in 31 villages of Dargai and Batkhela tehsils of Malakand. Soil from habitats of adult and immature sand flies was analysed. Questionnaire-based household survey was also performed in these villages to assess risk factors associated with CL. Soil samples from selected CL positive households were analysed for its contents. Additionally, clinicoepidemiological data from local health centres was examined for the year 2019. Total of 3,140 sand flies belonging to 18 species were collected. Phlebotomus sergenti was the most abundant species (38.16%). Its abundance had a strong positive correlation with mean monthly relative humidity and negative correlation with average temperature. Phlebotomus sergenti and Phlebotomus papatasi were abundant at an elevation ranging from 320 to 1,120 m above sea level and in agricultural lands near human settlements. Flight height preference apparatus collected maximum sand flies at 30 cm (1ft) above the ground and all species associated negatively with height. Soil analysis from habitats of adult and immature flies showed that highest mean number of adults and immatures were recorded from silt loam which carried highest concentrations of K2 O, Mg, Ca, and Zn. Number of immature sand flies correlated moderately (r = .7, p < .05) with K2 O soil concentrations. There was significant similarity between organic matter contents in soil samples from positive breeding sites and CL households (Wilcoxon rank-sum test, p = .1976). In multivariate analysis model for CL risk factors, age (26-35 and >35 years), knowledge of leishmaniasis, living in a middle and upper class, preachers visit to villages, and assumption that Afghan refugees are more prone to CL were significant. CL patient's archived data from health centres showed that majority of patients had lesions on face and hands. Patient's influx was highest in February and March.

First Report of Culicoides Biting Midges (Diptera: Ceratopogonidae) Attacking People in Italy, With the Description of Extreme Larval Breeding Sites and Diurnal Activity of Culicoides riethi.

Biting midges of the genus Culicoides (Diptera: Ceratopogonidae) play a paramount role in medical and veterinary entomology worldwide, particularly as vectors of pathogens which cause animal diseases. Biting midges are also infamous for the nuisance they provoke to people involved in outdoor activities. Nonetheless, attacks to man by midges from any Culicoides species have not been reported in Italy. An entomological investigation was performed following repeated attacks to man in a nature park near Rome (central Italy). The study area is a natural degassing zone, characterized by widespread hazardous gas emissions of CO2 and H2S, with several water bodies including permanent lakes, ponds, and pools. The biting midge C. riethi Kieffer, 1914 was very active during daytime in the period April-June. The species has been identified as responsible for attacks on people in the area. An in-depth analysis of the extreme environmental conditions revealed the ability of larvae to thrive in several water bodies, characterized by an extremely low pH and a high concentration of sulfates.

Mosquito larva distribution and natural Wolbachia infection in campus areas of Nakhon Ratchasima, Thailand

Objective: To determine the prevalence of mosquito larvae in campus areas and the infection rate of endosymbiotic bacteria, Wolbachia in mosquito larvae. Method: The mosquito larvae samples were collected in residential areas and academic buildings of Suranaree University of Technology located in Northeastern Thailand during 2017-2018. Mosquito species identification was performed using GLOBE mosquito protocols and Rattanarithikul & Panthusiri's keys. The gene encoding for the surface protein of Wolbachia was amplified by PCR and confirmed by DNA sequencing. Results: Armigeres sp. is the highest proportion of mosquito larvae followed by Culex spp., Aedes albopictus, Aedes aegypti, and Toxorynchites spp., respectively. Aedes aegypti have breeding sites mostly in the containers found indoors, whereas the main breeding sites of Aedes albopictus were found in both outdoors and indoors. The House Index and Breteau Index for Aedes spp. was more than 5% and 20%, respectively, in both areas, indicating that these areas are dengue sensitive. The highest proportion of Wolbachia infection was found in the larvae of Culex spp. (86.21%), followed by Aedes albopictus (69.23%) and rarely detected in Aedes aegypti (9.09%). Conclusion: The present study reported the first natural infection of Wolbachia in mosquito larvae in Thailand. Our result suggested that the mosquito species containing higher proportion of Wolbachia are less likely to be vectors for dengue. Therefore, Wolbachia transfection in mosquito larvae could be applied as a biocontrol for dengue and other mosquito-borne disease prevention.

Multi-Omic Analysis of Symbiotic Bacteria Associated With Aedes aegypti Breeding Sites.

Mosquito breeding sites are complex aquatic environments with wide microbial diversity and physicochemical parameters that can change over time during the development of immature insect stages. Changes in biotic and abiotic conditions in water can alter life-history traits of adult mosquitos but this area remains understudied. Here, using microbial genomic and metabolomics analyses, we explored the metabolites associated with Aedes aegypti breeding sites as well as the potential contribution of Klebsiella sp., symbiotic bacteria highly associated with mosquitoes. We sought to address whether breeding sites have a signature metabolic profile and understand the metabolite contribution of the bacteria in the aquatic niches where Ae. aegypti larvae develop. An analysis of 32 mosquito-associated bacterial genomes, including Klebsiella, allowed us to identify gene clusters involved in primary metabolic pathways. From them, we inferred metabolites that could impact larval development (e.g., spermidine), as well as influence the quality assessment of a breeding site by a gravid female (e.g., putrescine), if produced by bacteria in the water. We also detected significant variance in metabolite presence profiles between water samples representing a decoupled oviposition event (oviposition by single females and manually deposited eggs) versus a control where no mosquito interactions occurred (PERMANOVA: p < 0.05; R 2 = 24.64% and R 2 = 30.07%). Five Klebsiella metabolites were exclusively linked to water samples where oviposition and development occurred. These data suggest metabolomics can be applied to identify compounds potentially used by female Ae. aegypti to evaluate the quality of a breeding site. Elucidating the physiological mechanisms by which the females could integrate these sensory cues while ovipositing constitutes a growing field of interest, which could benefit from a more depurated list of candidate molecules.