Cryptic species Anopheles daciae (Diptera: Culicidae) found in the Czech Republic and Slovakia

Parasitology Research - We report the distribution of mosquitoes of the maculipennis complex in two distinct areas of the Czech Republic (Bohemia and South Moravia) and in one locality of...     

Species composition and distribution of adult Anopheles (Diptera: Culicidae) in Panama

Anopheles (Diptera: Culicidae) species composition and distribution were studied using human landing catch data over a 35-yr period in Panama. Mosquitoes were collected from 77 sites during 228 field trips carried out by members of the National Malaria Eradication Service. Fourteen Anopheles species were identified. The highest average human biting rates were recorded from Anopheles (Nyssorhynchus) albimanus (Wiedemann) (9.8 bites/person/night) and Anopheles (Anopheles) punctimacula (Dyar and Knab) (6.2 bites/person/night). These two species were also the most common, present in 99.1 and 74.9%, respectively, of the sites. Anopheles (Nyssorhynchus) aquasalis (Curry) was encountered mostly in the indigenous Kuna Yala Comarca along the eastern Atlantic coast, where malaria case history and average human biting rate (9.3 bites/person/night) suggest a local role in malaria transmission. An. albimanus, An. punctimacula, and Anopheles (Anopheles) vestitipennis (Dyar and Knab) were more abundant during the rainy season (May-December), whereas An. aquasalis was more abundant in the dry season (January-April). Other vector species collected in this study were Anopheles (Kerteszia) neivai (Howard, Dyar, and Knab) and Anopheles (Anopheles) pseudopunctipennis s.l. (Theobald). High diversity of Anopheles species and six confirmed malaria vectors in endemic areas of Panama emphasize the need for more detailed studies to better understand malaria transmission dynamics.     

Spatial and habitat distribution of Anopheles gambiae and Anopheles arabiensis (Diptera: Culicidae) in Banambani village, Mali

We studied the larval distribution and composition of Anopheles arabiensis Patton, An. gambiae s.s. Giles, and its forms, among local habitats; and their association with the adults between these habitats in Banambani village, Mali during the mid-rainy seasons of 1997-1999. For species and form identification we used polymerase chain reaction (PCR) and PCR-restriction fragment-length polymorphism (RFLP). Differences among species in the distribution of larvae were observed in 1998, but not in 1997 or 1999, although they were on the borderline of statistical significance. Differences among the M and S molecular forms were statistically significant in 1999 when rainfall was high, but not in the two prior, drier sampling periods. Combining all information into the Fisher multiple comparisons test, there were statistically significant differences between species and molecular forms during the 3-yr study period. Hybrid larvae between the M and S forms were observed (0.57%), the first such observation to our knowledge. In spite of differences among larval distribution, no differences of adult species composition were observed among habitats. Factors that influence the distributions of An. gambiae larval populations are discussed.     

Reproduction-longevity trade-off in Anopheles gambiae (Diptera: Culicidae)

Reduced survival and future reproduction due to of current reproduction is a trade-off known as the cost of reproduction. Surprisingly, only a few studies have assessed the cost of reproduction in arthropod disease vectors, despite its effect on longevity, and thus on vectorial capacity. We evaluated the cost of reproduction on survival of Anopheles gambiae Giles by comparing mosquitoes that were denied exposure to the other sex, hereafter named virgins, and those that were allowed exposure to the other sex and mating, hereafter named mated. Merely 6 d of exposure to females with mating activity reduced male survival from a median of 17 d in virgins to 15 d in mated, indicating that male mating cost is substantial. The increase in mortality of mated males began several days after the exposure to females ended, indicating that mating is not associated with immediate mortality risk. Notably, body size was negatively correlated with male mortality in mated males, but not in virgins. The rate of insemination declined after 4 d of exposure to females, indicating that male mating capacity is limited and further supporting the hypothesis that mating is costly for males. Consistent with previous studies, female survival on sugar alone (median=16 d) was shorter than on blood and sugar (median=19 d), regardless if she was mated or virgin. Overall, survival of mated females was lower than that of virgins on a diet of blood and sugar, but no difference was found on a diet of sugar only. However, the cost of reproduction in females remains ambiguous because the difference in survival between virgin and mated females was driven by the difference between virgin (median=19 d) and uninseminated females exposed to males (median=17 d), rather than between virgin and inseminated females (median=19 d). Accordingly, sperm and seminal fluid, egg development, and oviposition have negligible cost in terms of female survival. Only exposure to males without insemination decreased female survival. Nonetheless, if exposure to males under natural conditions is also associated with reduced survival, it might explain why females remain monogamous.     

Identification of Anopheles daciae in Germany through ITS2 sequencing

Until the middle of the twentieth century, malaria was frequently endemic in parts of Germany; Anopheles maculipennis complex species were considered the primary vectors. Three species of this complex have been identified in Germany: A. maculipennis s.s., Anopheles messeae and Anopheles atroparvus; the last predominantly from the coastal regions of Northern Germany. Anopheles daciae is a recently described member of the A . maculipennis complex and resembles the well-characterised species A. messeae, although the two species can be distinguished through their egg morphology and sequencing of the internal transcribed spacer 2 (ITS2) region of their nuclear rDNA. In this study, we harvested larval and adult mosquito samples from five breeding sites and ten CO₂ trap collection sites in the Upper Rhine Valley of Southwestern Germany to analyse the complement of anopheline species present. Mosquito ITS2 DNA was extracted and polymerase chain reaction (PCR)-amplified using established protocols. Genomic analysis was performed by a species-diagnostic restriction fragment length polymorphism assay as well as by sequencing of PCR products; the data obtained were aligned against nucleic acid sequences from English mosquitoes retrieved from GenBank. Additionally, the larval breeding sites of A. messeae were characterised through water quality measurement. Forty-seven samples were successfully processed, of which 6 were identified as A. daciae and 41 as A. messeae. All samples of A. daciae, which has not previously been found in Central Europe, originated from one CO₂ trap collection site in Dettenheim, close to Karlsruhe, Southwestern Germany. The identification of this malarial vector in a novel area may have implications for the re-emergence of disease subsequent to climatic changes.     

Integration of Anopheles beklemishevi (Diptera: Culicidae) in a PCR assay diagnostic for palaearctic Anopheles maculipennis sibling species

A few years ago a PCR-based assay for a quick and reliable identification of six palaearctic sibling species of the Anopheles maculipennis complex was presented making use of differences in the nucleotide sequence of the ITS2 ribosomal mosquito DNA. An. beklemishevi, which is distributed in Scandinavia and Russia only, has now been integrated into this test after analysis of its ITS2 region which turned out to be much longer than those of the other sibling species. Three oligonucleotides putatively specific for An. beklemishevi were constructed and tested in combination with a universal genus-specific primer for the amplification of an An. beklemishevi-specific ITS2 DNA-fragment. Two of the three oligos generated accurate and specific PCR products, even when used in a multiplex PCR together with the specific primers for the other six sibling species. Cross-hybridization of the primers to heterologous culicid DNA was never observed. The amplicons that identify An. beklemishevi consist of 554 and 735 bp, respectively, and are easily distinguished from those specific for the other sibling species after gel electrophoresis.     

Rubidium marking of Anopheles stephensi (Diptera: Culicidae)

Immature Anopheles stephensi Liston were reared in untreated water and water containing eight 2-fold dilutions of rubidium (Rb) from 1,000 to 7.8 ppm to determine the concentration that allowed reliable detection and produced the least toxic effects as measured by adult emergence, weight, and survival. The amount of Rb detected in mosquitoes increased positively with increasing concentrations in the rearing water. Concentrations > or = 31.2 ppm Rb in the rearing water provided high and consistent detection levels of > or = 3,500 ppm Rb/mg of adult mosquito. There were no adverse effects of Rb on the weights of mosquitoes. However, increased Rb concentrations in the rearing water were associated with decreased emergence and survival. After 8 d, percentage emergence from Rb concentrations of 0-31.2 ppm was > or = 50%. At day 21, Rb concentrations of 0-31.2 ppm showed < or = 29% reduction of female survival compared with controls. The recommended concentration for reliable Rb detection with minimal toxic effects in An. stephensi was 32 ppm.     

Factors associated with distribution of Anopheles aquasalis and Anopheles oswaldoi (Diptera: Culicidae) in a malarious area, northeastern Venezuela

Spatial and temporal abundance patterns of anopheline larvae and their relationships with wetland conditions were studied in an endemic malaria area in northeastern Venezuela, where Anopheles aquasalis Curry is the main vector. Larvae were sampled over a 2-yr period in 7 wetland types (brackish and freshwater herbaceous swamps, mangrove swamps, freshwater ponds, clear-cut marsh forests, small irrigation canals, and swamp forests), covering 3 environmental gradients (salinity, aquatic vegetation, and habitat permanence). Twelve variable were quantified to describe each habitat. Two species of anophelines were collected. An. aquasalis was the species with the widest distribution, and its highest abundance was in the seasonal brackish mangrove habitat during the rainy season. An. oswaldoi Peryassu was rarely encountered, but was mainly associated with the dry season and with the permanent fresh water wetlands (such as ponds). Principal components and correlation analyses revealed that the physicochemical (salinity, dissolved oxygen) variables of the wetland were associated most strongly with the spatial distribution of both species. Variations in salinity were strongly associated with the abundance of An. aquasalis. Both the occurrence and abundance of An. oswaldoi were most closely correlated with dissolved oxygen. Changes in seasonal abundance of both species were associated with rainfall. The relevance of these results to vector control in northern Venezuela is discussed.     

Mosquitoes (Diptera: Culicidae) of metropolitan Hamburg,Germany

In Europe, mosquito-related public health concerns are growing due to the increasing spread of invasive mosquito species and the recent emergence of mosquito-borne arboviruses. A vital backbone in the assessment of these issues is detailed knowledge of the mosquito fauna, i.e. regional mosquito inventories. It was therefore decided to intensify nationwide investigations on the occurrence and distribution of mosquitoes in Germany in order to update old records and to detect possible faunal changes. This paper is focussing on a densely populated metropolitan region, the federal state of Hamburg and its adjacent environs, taking two historical baseline inventories into consideration, spanning almost 100 years of mosquito research in Hamburg. In the period between 2010 and 2014, more than 10,000 juvenile, neonate and adult mosquito specimens were sampled and trapped at 105 sites in Hamburg and its environs, of which about 60 % have been identified to species level, resulting in a total of 33 recorded species. Of these, Anopheles algeriensis, Culex modestus, Ochlerotatus caspius, Ochlerotatus nigrinus and Ochlerotatus sticticus are new to the area. The most common species in Hamburg are Culex pipiens/torrentium and Ochlerotatus annulipes/cantans. In contrast, two previously common species, Anopheles atroparvus and Ochlerotatus excrucians, were not detected. Despite substantial environmental changes due to reconstruction, urbanisation and renaturation in the Hamburg metropolitan region in recent decades, there has been remarkably little change within the mosquito fauna during the last century.     

Diving ability of Anopheles gambiae (Diptera: Culicidae) larvae

Anopheles gambiae Giles larvae usually live near the surface of shallow and temporary aquatic habitats. How deep the larvae can dive and how long they can submerge may be related to feeding efficiency and predator avoidance. This study examined diving behavior of An. gambiae larvae in the laboratory. We recorded diving depths and larval mortality of second and fourth instars in clean water and muddy water by using deep water (32-cm) and shallow water (20-cm) columns. In deep water columns with clean water, we found that 2% of second instars and 6% of fourth instars died from diving, whereas 3% of second instars and 11% of fourth instars died in muddy water. The fourth instars dived deeper in muddy water than in clean water. The mortality rates of the fourth instars subjected to diving stimulations were significantly higher than those in the shallow water columns. Therefore, larval diving behavior may offer the benefits of predator avoidance and food acquisition but also incur energetic costs and increased mortality.     

Further reports of Anopheles algeriensis Theobald, 1903 (Diptera: Culicidae) in Germany,with evidence of local mass development

Parasitology Research - Anopheles algeriensis, a thermophilic mosquito species widely distributed in the Mediterranean, is supposed to be extremely rare and to occur in very low abundances in...     

Occurrence of Anopheles hermsi (Diptera: Culicidae) in Arizona and Colorado

Historically, malaria was a significant cause of morbidity and mortality throughout the western United States, and Anopheles freeborni Aitken was thought to be the vector west of the Continental Divide. In 1989, Anopheles hermsi Barr & Guptavanij was described and subsequently found to be an effective laboratory vector of Plasmodium. The adults of these two species are morphologically indistinguishable, and therefore polymerase chain reaction was used to analyze the DNA from 48 mosquitoes collected in Arizona and Colorado (identified morphologically as An. freeborni). All specimens were identified as An. hermsi. This was the first report of An. hermsi in Arizona and Colorado and indicated that this Anopheles species historically may have been a malaria vector in these two western states.     

Potential distribution of two species in the medically important Anopheles minimus complex (Diptera: Culicidae)

Anopheles minimus Theobald (=An. minimus A) and possibly Anopheles harrisoni Harbach & Manguin (=An. minimus C) are important malaria vector species in the Minimus Complex in Southeast Asia. The distributions of these species are poorly known, although detailed information could benefit malaria vector incrimination and control. We used published collection records of these species and environmental geospatial data to construct consensus ecological niche models (ENM) of each species' potential geographic distribution. The status of the Indian taxon An. fluviatilis S as a species distinct from An. harrisoni has been debated in the literature, so we tested for differentiation in ecological niche characteristics. The predicted potential distribution of An. minimus is more southerly than that of An. harrisoni: Southeast Asia is predicted to be more suitable for An. minimus, and China and India are predicted more suitable for An. harrisoni, so An. harrisoni seems to dominate under cooler conditions. The distribution of An. minimus is more continuous than that of An. harrisoni: disjunction in the potential distribution of the latter is suggested between India and Southeast Asia Anopheles fluviatilis S occurrences are predicted within the An. harrisoni ecological potential, so we do not document ecological differentiation that might reject conspecificity. Overall, model predictions offer a synthetic view of the distribution of this species complex across the landscapes of southern and eastern Asia.     

Molecular evidence for similarity between Anopheles hyrcanus (Diptera: Culicidae) and Anopheles pseudopictus (Diptera: Culicidae), sympatric potential vectors of malaria in France

Malaria was a former public health problem in the Camargue, southeastern France, where members of the Hyrcanus group were recently described as the main malaria potential vectors. However, the systematic status in this group, which includes at least two sympatric sibling species, Anopheles hyrcanus (Pallas) and Anopheles pseudopictus Grassi as well as a morphologically intermediate form in the Camargue, is unclear. Indeed, both species have been alternatively considered as separated or synonymous species. We examined sequence variation of the internal transcribed spacer (ITS) 2 and domain-3 (D3) of 28S ribosomal DNA and the cytochrome oxidase subunit I and II (COI and COII) genes of mitochondrial DNA of the Hyrcanus group mosquitoes from the Camargue and Turkey to infer the taxonomic status of the members of this group. DNA sequence analysis of ITS2 and D3 showed no difference between either species or geographical origin (mean pairwise genetic distances d = 0.000-0.003). The COI and COII sequences between French specimens also were nearly identical (d = 0.001-0.002), whereas French and Turkish Anopheles were genetically distinct (d = 0.009-0.014). The distinction between populations of the two areas, supported, respectively, by four and five fixed mutations, attested the differentiation by the distance. Finally, the high degree of genetic similarity, despite morphological differences between An. hyrcanus, An. pseudopictus, and an intermediate form, suggests that these three taxa may belong to a single species in the Camargue.     

Insecticide resistance in Anopheles funestus (Diptera: Culicidae) from Mozambique

Malaria control in southern Mozambique is currently by indoor residual carbamate insecticide treatment, with pyrethroid-treated bed-nets distributed to pregnant women and children under five in northern Mozambique. The susceptibility of Anopheles funestus s.s. to pyrethroid, carbamate, organochlorine, and organophosphorus insecticides was determined by World Health Organization adult mosquito susceptibility tests at 19 localities in Mozambique, from March 2000 to July 2002. Biochemical assays were carried out on mosquitoes from the same families to detect shifts in the quantity or activity of enzyme families involved in insecticide detoxification. An. funestus from all localities remained fully susceptible to DDT and the organophosphorus insecticide malathion. A high level of pyrethroid resistance was detected in An. funestus populations in southern Mozambique. An. funestus outside Maputo province were still susceptible to pyrethroids. An. funestus from six localities also were resistant to carbamate insecticides propoxur and bendiocarb. Both pyrethroid and carbamate resistance occurred in five of these six localities. Mosquitoes from five of the localities with elevated p450 estimates, compared with the insecticide-susceptible Durban strain, were pyrethroid-resistant. The only exception to this trend was Mozal, which had elevated p450 estimates but full pyrethroid susceptibility by bioassay. The lack of cross-resistance between pyrethroids and DDT in Mozambican An. funestus suggests that a kdr-type target site resistance mechanism has not been selected. Low levels of insecticide-insensitive acetylcholinesterase, the target site for carbamates and organophosphates, were found in all populations tested. The high level of metabolically based pyrethroid resistance has implications for current malaria control programs in Mozambique.     

Reassociation kinetics of Anopheles gambiae (Diptera: Culicidae) DNA.

The renaturation kinetics of nuclear DNA from the G3 colony of Anopheles gambiae Giles was studied to estimate the genome size and to determine the proportion of repeated sequences. An. gambiae has a haploid DNA content of 0.27 picograms or 2.6 x 10(8) basepairs. Analysis of reassociation kinetics indicated that the genome is composed of 61% single-copy and 33% repetitive sequences, with 6% foldback sequences.