Tabanus chrysurus is a potential biological vector of Trypanosoma (Megatrypanum) theileri in Japan.

Several species of horse flies (Diptera: Tabanidae) are known as vectors of Trypanosoma (Megatrypanum) theileri and T. theileri-like trypanosomes; these host-parasite relationships were established based on the developmental stages of these parasites discovered in the hindgut of horse flies. T. theileri and T. theileri-like trypanosomes have been detected in cattle and wild deer in Japan; however, the vector horse fly species remains unidentified. Therefore, in this study, we aimed to identify the potential horse fly species serving as vectors of T. theileri in Japan. A total of 176 horse flies were collected between June to September 2020 and 2021 in Tokachi, Hokkaido, Japan. The T. theileri infection in the captured horse flies was determined by PCR and microscopic analyses of their midgut and hindgut. Additionally, the trypanosome, microscopically detected in a horse fly, was molecularly characterized and phylogenetically analyzed using 18S rRNA and partial cathepsin L-like protein gene (CATL) sequence of the trypanosome. The microscopy and PCR analyses revealed 0.57% and 35.8% prevalence of T. theileri in horse flies, respectively. Epimastigote stages of T. theileri, adhered to the hindgut epithelial cells of Tabanus chrysurus via flagella or actively moving in the lumen of the gut, were detected. Phylogenetic analysis revealed the connection of isolated trypanosomes with T. theileri in the TthI clade. These results suggest that Ta. chrysurus is a potential vector of T. theileri.

Ecological determinants of prevalence of the male-killing bacterium Arsenophonus nasoniae.

Male-killing bacteria are found in a broad range of arthropods. Arsenophonus nasoniae is a male-killing bacterium, causing a 80% reduction of the male progeny in infected Nasonia vitripennis wasps. Although the discovery of A. nasoniae dates from the early 80's, knowledge about the biology and ecology of this endosymbiont is still scarce. One of these poorly studied features is the ecological factors underlying A. nasoniae incidence on its Nasonia spp. hosts in different geographical locations. Here, we studied the prevalence of A. nasoniae in Iberian wild populations of its host N. vitripennis. This wasp species is a common parasitoid of the blowfly Protocalliphora azurea pupae, which in turn is a parasite of hole-nesting birds, such as the blue tit (Cyanistes caeruleus). We also examined the effects of bird rearing conditions on the prevalence of A. nasoniae through a brood size manipulation experiment (creating enlarged, control and reduced broods). Both the wasp and bacterium presence were tested through PCR assays in blowfly pupae. We found A. nasoniae in almost half (47%) of nests containing blowflies parasitized by N. vitripennis. The prevalence of A. nasoniae was similar in the two geographical areas examined (central Portugal and southeastern Spain) and the probability of infection by A. nasoniae was independent of the number of blowfly pupae in the nest. Experimental manipulation of brood size did not affect the prevalence of A. nasoniae nor the prevalence of its host, N. vitripennis. These results suggest that the incidence of A. nasoniae in natural populations of N. vitripennis is high in the Iberian Peninsula, and the infestation frequency of nests by N. vitripennis carrying A. nasoniae is spatially stable in this geographical region independently of bird rearing conditions.

Distribution of phoretic mites and lice in Pseudolynchia canariensis living on pigeons and the relationship with seasonality, carrier sex, plumage coloration and age of definitive hosts.

Among the parasites, some groups that have a limited capacity for locomotion, such as mites and lice, the transmission is challenging to win. These ectoparasites disperse through direct contact between hosts or, in some cases, through phoresy. However, these processes are not well-documented in detail because they are difficult to observe and quantify. In the present study, the patterns of distribution of skin mites and phoretic lice on hippoboscid louse fly Pseudolynchia canariensis sampled from Columba livia were evaluated. The analyzed pigeons were juveniles and adults, with three distinct plumage colors: blue checker, spread, or wild type, and were caught over 24 months. A total of 1,381 hippoboscid flies were collected on 377 hosts. The plumage color did not influence the infestation patterns of louse flies on juvenile and adult pigeons, nor did it influence the infestation patterns of skin mites and phoretic lice on the hippoboscid flies. However, the environmental temperature was directly related to higher prevalence, mean infestation intensity, and phoretic species richness on P. canariensis during the hottest seasons. Furthermore, a higher abundance of phoretic mite eggs, including embryonated eggs, was observed in females of P. canariensis in all seasons.

Development of a novel molecular tool to study molecular ecology of Ornithomya (Hippoboscidae) avian louse flies.

Louse flies (Diptera: Hippoboscidae) are obligatory hematophagous ectoparasites of birds and mammals. These widely distributed parasitic flies may have a significant impact on wild and farm animals by feeding on their blood and transmitting bloodborne pathogens. However, despite their ecological importance, louse flies are clearly underrepresented in host-parasite research and implementation of genetic approaches in this group is generally hampered by lacking molecular tools. In addition, louse flies that parasitize long-distance migrants can travel long distances with their avian hosts, facilitating the large-scale spread of pathogens across landscapes and geographic regions. Given the wide diversity of louse flies that parasitize a variety of avian hosts, their direct negative impact on host survival, and their high potential to transmit bloodborne pathogens even along avian migration routes, it is surprising that our knowledge of louse fly ecology is rather modest and incomplete. Here, we aimed to develop a novel molecular tool for polyxenous avian louse flies from the genus Ornithomya, which are among the most common and widely distributed representatives of Hippoboscidae family, to improve research of their genetic population structure and molecular ecology. Using the Illumina Mi-seq sequencing, we conducted a genome-wide scan in Ornithomya avicularia to identify putative microsatellite markers. A panel of 26 markers was selected to develop amplification protocols and assess polymorphism in the Central European population of O. avicularia, as well as to test for cross-amplification in a congeneric species (O. chloropus). A genome-scan in O. avicularia identified over 12 thousand putative microsatellite markers. Among 26 markers selected for a population-wide screening; one did not amplify successfully and three were monomorphic. 22 markers were polymorphic with at least two alleles detected. Two markers showed presence of null alleles. A cross-amplification of microsatellite markers in O. chloropus revealed allelic polymorphism at 14 loci, with the mean allelic richness of 3.78 alleles per locus (range: 2-8). Our genome-wide scan in O. avicularia provides a novel and powerful tool for molecular research in Ornithomya louse flies. Our panel of polymorphic microsatellite loci should allow genotyping of louse flies from geographically distinct populations and from a wide spectrum of avian hosts, enhancing population genetic and phylogeographic research in Ornithomya.

An Atypical Presentation Of Cutaneous Myiasis Of The Scalp In North Punjab: A Case Report.

Myiasis is an ectoparasitic infestation caused by larvae of arthropods in the group Diptera. Cutaneous myiasis is the infection of skin and includes the following types: furuncular, migratory and wound myiasis. Out of all the organisms responsible for this disease, Wohlfahrtia vigil is the most common causative agent in Pakistan, usually seen from June to September with nearly all presentations occurring at a young age. In our case, a patient without a history of trauma, skin erosion or animal exposure has a unique presentation of cutaneous myiasis. Here, an effort was made to treat a 15- year-old otherwise healthy female with a multisegmented treatment modality. A follow up visit was conducted every three days for three months at the end of which a 60% decrease in the size of the open wound was observed.

Molecular investigation on infection by haemosporidians in three Western Palearctic species of swift (Apodidae) and their ectoparasitic louse flies.

Swifts (Apodidae) are an unusual group of birds that spend most of their lives in flight, landing only when breeding. Although this aerial lifestyle greatly reduces their likelihood of being bitten by vectors and infected by vector-born parasites, swifts can still be heavily infested during breeding by nest-based vectors such as louse flies (Hippoboscidae). Here, we investigated host, vector, and vector-borne parasite relationships in the three most widespread swift species in the Western Palearctic (WP): common swifts (Apus apus), pallid swifts (A. pallidus), and alpine swifts (Tachymarptis melba), their nest-based louse flies (Crataerina pallida and C. melbae) and avian haemosporidians (genera Haemoproteus, Plasmodium, and Leucocytozoon). Studies of haemosporidian infections in Apodidae remain limited, with clear evidence of infection found to date in just four Neotropical and one Australasian species. The possible role of louse flies in transmitting haemosporidian infections has never been tested in swifts. We assessed the occurrence of haemosporidian infection by PCR screenings of DNA from blood samples from 34 common swifts and 44 pallid swifts from Italy, and 45 alpine swifts from Switzerland. We also screened 20 ectoparasitic louse flies present on 20 birds and identified them by both morphological features and cytochrome oxidase subunit 1 (COI) barcodes. Our results provide no evidence of haemosporidian infection in the 123 swifts tested or in the two louse fly species we identified. Our findings are consistent with available knowledge showing no haemosporidian occurrence in WP swift species and that the most likely infection route for these highly aerial species (via louse fly ectoparasites during nesting) is unlikely.

On the life cycle of Brachymeria podagrica (Fabricius, 1787) (Hymenoptera: Chalcididae) - a parasitoid of sacrophagid, calliphorid, and muscid flies.

The lifecycle of Brachymeria podagrica, a parasitic wasp with a worldwide distribution, was studied under laboratory conditions using the flesh fly, Sarcophaga dux, as a host. Two hundred parasite-free 3rd instars of S. dux were exposed for 24 h to 20 female B. podagrica. In daily intervals, maggots and later pupae were examined for developmental stages of the parasitoid. The whole pre-imaginal development at a temperature of 26 °C lasted 21 to 26 days. Three morphologically different instars, followed by a prepupal and a pupal stage, were described using light and scanning electron microscopy. In a second experiment with 100 3rd stage Sarcophaga larvae and 10 parasitoids, a total of 70 wasps emerged 20 to 25 days after exposure. Two fly larvae did not pupate and dried out, while 28 pupae contained a dry or caseous content, dead wasp imagos, or their larval stages. No fly imagines emerged from exposed groups, while all 100 unexposed larvae pupated and adults eclosed between day 12 and day 14 after the start of the experiment, while the imagoes of the parasitoids appeared 8 to 12 days later.

The global prevalence of parasites in non-biting flies as vectors: a systematic review and meta-analysis.

BACKGROUND: Non-biting flies such as the house fly (Musca domestica), the Australian sheep blowfly (Lucilia cuprina) and the oriental latrine fly (Chrysomya megacephala) may carry many parasites. In the present study, we performed a systematic overview of the different species of parasites carried by non-biting flies, as well as of isolation methods, different geographical distribution, seasonality and risk assessment. METHODS: A meta-analysis was carried out with the aim to review the global prevalence of parasite transmission in non-biting flies. A total sample size of 28,718 non-biting flies reported in studies worldwide satisfied the predetermined selection criteria and was included in the quantitative analysis. RESULTS: The global prevalence of parasites in non-biting flies was 42.5% (95% confidence interval [CI] 31.9-53.2%; n = 15,888/28,718), with the highest prevalence found for non-biting flies in Africa (58.3%; 95% CI 47.4-69.3%; n = 9144/13,366). A total of 43% (95% CI 32.1-54.4%; n = 7234/15,282) of house flies (M. domestica), the fly species considered to be the most closely associated with humans and animals, were found with parasites. The prevalence of parasites in the intestine of non-biting flies was 37.1% (95% CI 22.7-51.5%; n = 1045/3817), which was significantly higher than the prevalence of parasites isolated from the body surface (35.1%; 95% CI 20.8-49.4%; n = 1199/3649; P < 0.01). Of the 27 reported parasites, a total of 20 known zoonotic parasites were identified, with an infection rate of 38.1% (95% CI 28.2-48.0%; n = 13,572/28,494). CONCLUSIONS: This study provides a theoretical basis for the public health and ecological significance of parasites transmitted by non-biting flies.

Possible role of Lipoptena fortisetosa (Diptera: Hippoboscidae) as a potential vector for Theileria spp. in captive Eld's deer in Khao Kheow open zoo, Thailand.

Eld's deer (Rucervus eldii thamin) is an endangered species endemic to South Asia. Various ectoparasites (hematophagous insects and ticks) and blood parasites (e.g., piroplasms such as Babesia and Theileria) have been reported in this deer. Deer keds of the genus Lipoptena (L.) are wingless hematophagous insects acting as ectoparasites and potential vectors, thereby transmitting diseases to animals and humans. Many Lipoptena species have been reported, including L. fortisetosa; the latter may be a potential vector of several pathogens such as Babesia spp. and Theileria spp. However, the available data regarding Lipoptena in domestic animals and wildlife in Thailand is limited. The aim of this study was to investigate the presence of L. fortisetosa in Eld's deer as well as the role of this insect as a disease vector in Thailand by employing molecular analysis. A total of 91 wingless insects were collected and morphologically identified as L. fortisetosa. A partial fragment of the cytochrome c oxidase subunit I gene (COI) was amplified and successfully sequenced from twelve insects, and the COI nucleotide Basic Local Alignment Search Tool results revealed a 94.28%-94.45% identity to L. fortisetosa (accession number: OL850869/China). The undertaken phylogenetic analysis revealed that the L. fortisetosa samples from Thailand belong to a clade that is distinct from the previously deposited (in GenBank®) L. fortisetosa. As far as the pathogen detection is concerned, 46.2% (42/91) of the deer keds were positive for Theileria, while no Lipoptena was found to be positive for Babesia. Twenty-one sequences of Theileria were obtained and exhibited a 98.84%-100% identity to the Theileria sp. from several hosts. The phylogenetic analysis of Theileria revealed that Theileria capreoli and Theileria cervi were present in our L. fortisetosa samples. It can be implied that L. fortisetosa may serve as a vector of Theileria spp. in the Eld's deers of Thailand. We believe that the particular open zoo (from where the sampling took place) should implement preventive and control strategies for deer keds, other vectors, and vector-borne diseases.

The genomic basis of host and vector specificity in non-pathogenic trypanosomatids.

Trypanosoma theileri, a non-pathogenic parasite of bovines, has a predicted surface protein architecture that likely aids survival in its mammalian host. Their surface proteins are encoded by genes which account for ∼10% of their genome. A non-pathogenic parasite of sheep, Trypanosoma melophagium, is transmitted by the sheep ked and is closely related to T. theileri. To explore host and vector specificity between these species, we sequenced the T. melophagium genome and transcriptome and an annotated draft genome was assembled. T. melophagium was compared to 43 kinetoplastid genomes, including T. theileri. T. melophagium and T. theileri have an AT biased genome, the greatest bias of publicly available trypanosomatids. This trend may result from selection acting to decrease the genomic nucleotide cost. The T. melophagium genome is 6.3Mb smaller than T. theileri and large families of proteins, characteristic of the predicted surface of T. theileri, were found to be absent or greatly reduced in T. melophagium. Instead, T. melophagium has modestly expanded protein families associated with the avoidance of complement-mediated lysis. We propose that the contrasting genomic features of these species is linked to their mode of transmission from their insect vector to their mammalian host. This article has an associated First Person interview with the first author of the paper.

Development of two species of the Trypanosoma theileri complex in tabanids.

BACKGROUND: Trypanosoma theileri species complex includes parasites of Bovidae (cattle, sheep, goat, etc.) and Cervidae (deer) transmitted mainly by Tabanidae (horse flies and deerflies) and keds (Hippoboscidae). While morphological discrimination of species is challenging, two big clades, TthI and TthII, each containing parasites isolated from bovids and cervids, have been identified phylogenetically. To date, the development in the vector has been studied in detail only for the ked-transmitted sheep parasite T. melophagium (TthII), while the fate of trypanosomes in tabanids was described only briefly by light microscopy. METHODS: We collected infected tabanids of various species and identified trypanosomes by molecular phylogenetic analysis. The morphology and development of trypanosomes was studied using the combination of statistical analyses as well as light and electron microscopy. RESULTS: Two trypanosome species belonging to both TthI and TthII clades of the T. theileri complex were identified. The phylogenetic position of these two trypanosomes suggests that they parasitize deer. Both species were indiscernible by morphology in the vector and showed the same development in its intestine. In contrast to the previously described development of T. melophagium, both trypanosomes of tabanids only transiently infected midgut and settled mainly in the ileum, while pylorus and rectum were neglected. Meanwhile, the flagellates developing in the tabanid ileum (pyriform epimastigotes and metacyclic trypomastigotes) showed similarities to the corresponding stages in T. melophagium by morphology, mode of attachment to the host cuticle and formation of the fibrillar matrix surrounding the mass of developing parasites. In addition, for the first time to our knowledge we documented extraintestinal stages in these trypanosomes, located in the space between the epithelium and circular muscles. CONCLUSIONS: The development of different species of flagellates of the T. theileri complex in their insect vectors shows many similarities, which can be explained not only by their common origin, but also the same transmission mode, i.e. contamination of the oral mucosa with the gut content released after squashing the insect either by tongue or teeth. The observed differences (concerning primarily the distribution of developmental stages in the intestine) are associated rather with the identity of vectors than the phylogenetic position of parasites.

'A flying start': Wildlife trypanosomes in tissues of Australian tabanids (Diptera: Tabanidae).

Tabanids (syn. horse flies) are biting-flies of medical and veterinary significance because of their ability to transmit a range of pathogens including trypanosomes - some species of which carry a combined health and biosecurity risk. Invertebrate vectors responsible for transmitting species of Trypanosoma between Australian wildlife remains unknown, thus establishing the role of potential vector candidates such as tabanids is of utmost importance. The current study aimed to investigate the presence of indigenous trypanosomes in tabanids from an endemic area of south-west Australia. A total of 148 tabanids were collected, with morphological analysis revealing two subgenera: Scaptia (Pseudoscione) and S. (Scaptia) among collected flies. A parasitological survey using an HRM-qPCR and sequencing approach revealed a high (105/148; 71%) prevalence of trypanosomatid DNA within collected tabanids. Individual tissues - proboscis (labrum, labium and mandibles, hypopharynx), salivary glands, proventriculus, midgut, and hindgut and rectum - were also tested from a subset of 20 tabanids (n = 140 tissues), confirming the presence of Trypanosoma noyesi in 31% of screened tissues, accompanied by T. copemani (3%) and T. vegrandis/T.gilletti (5%). An unconfirmed trypanosomatid sp. was also detected (9%) within tissues. The difference between tissues infected with T. noyesi compared with tissues infected with other trypanosome species was statistically significant (p < 0.05), revealing T. noyesi as the more frequent species detected in the tabanids examined. Fluorescence in situ hybridisation (FISH) and scanning electron microscopy (SEM) confirmed intact parasites within salivary glands and the proboscis respectively, suggesting that both biological and mechanical modes of transmission could occur. This study reveals the presence of Australian Trypanosoma across tabanid tissues and confirms intact parasites within tabanid salivary glands and the proboscis for the first time. Further investigations are required to determine whether tabanids have the vectorial competence to transmit Australian trypanosomes between wildlife.

Relationship among bats, parasitic bat flies, and associated pathogens in Korea.

BACKGROUND: Bats are hosts for many ectoparasites and act as reservoirs for several infectious agents, some of which exhibit zoonotic potential. Here, species of bats and bat flies were identified and screened for microorganisms that could be mediated by bat flies. METHODS: Bat species were identified on the basis of their morphological characteristics. Bat flies associated with bat species were initially morphologically identified and further identified at the genus level by analyzing the cytochrome c oxidase subunit I gene. Different vector-borne pathogens and endosymbionts were screened using PCR to assess all possible relationships among bats, parasitic bat flies, and their associated organisms. RESULTS: Seventy-four bat flies were collected from 198 bats; 66 of these belonged to Nycteribiidae and eight to Streblidae families. All Streblidae bat flies were hosted by Rhinolophus ferrumequinum, known as the most common Korean bat. Among the 74 tested bat flies, PCR and nucleotide sequencing data showed that 35 (47.3%) and 20 (27.0%) carried Wolbachia and Bartonella bacteria, respectively, whereas tests for Anaplasma, Borrelia, Hepatozoon, Babesia, Theileria, and Coxiella were negative. Phylogenetic analysis revealed that Wolbachia endosymbionts belonged to two different supergroups, A and F. One sequence of Bartonella was identical to that of Bartonella isolated from Taiwanese bats. CONCLUSIONS: The vectorial role of bat flies should be checked by testing the same pathogen and bacterial organisms by collecting blood from host bats. This study is of great interest in the fields of disease ecology and public health owing to the bats' potential to transmit pathogens to humans and/or livestock.

A nationwide survey of the tabanid fauna of Cameroon.

BACKGROUND: Tabanids are a neglected group of haematophagous dipterans despite containing 4434 species, regrouped in > 144 genera. They are mechanical vectors of important pathogens, including viruses, bacteria and protozoa of humans and domesticated and wild animals. As it is > 50 years since the publication of a preliminary nationwide record of the tabanids of Cameroon identified 84 species, updated information is needed. The aim of this study was to provide current data on the species composition, abundance and distribution of tabanids in the five main agro-ecological zones (AEZs) of Cameroon. METHODS: From 2015 to 2017, a systematic entomological study using Nzi, Vavoua, Biconical and Sevi traps (n = 106) was conducted in 604 trapping points over 11,448 trap-days in the five main AEZs of Cameroon. RESULTS: A total of 25,280 tabanids belonging to 25 species were collected, including eight species not previously documented in Cameroon, namely Tabanus latipes (1 female), Tabanus ricardae (1 female), Tabanus fasciatus (32 females and 6 males), Haematopota pluvialis (18 females), Haematopota decora (19 females and 3 males), Haematopota nigripennis (18 females), Chrysops distinctipennis (47 females and 5 males) and Ancala fasciata (34 females and 7 males). The distribution maps of the newly identified tabanids differed between AEZs, with most tabanids collected from the Guinean savanna. The highest apparent density of tabanids was recorded in the Sudan Savanna region, and the mean apparent densities of species with sites was statistically significantly different (Student t-test: 2.519, df = 24, P = 0.019). The highest species diversity was found in the rainforest. CONCLUSIONS: This study increased the list of tabanids recorded in Cameroon from 84 species in the preliminary record to 92 species, with most of the newly identified species occurring in the Guinea Savanna AEZ. The high diversity and abundance of tabanids in the livestock/wildlife interface areas of the rain forests and Sudan Savanna AEZs, respectively, suggest risk of mechanical transmission of pathogens. Investigations of the microbiota of tabanids in the different AEZs to define their role as disease vectors are proposed.

Non-lethal effects of entomopathogenic nematode infection.

Entomopathogenic nematodes are typically considered lethal parasites of insect hosts. Indeed they are employed as such for biological control of insect pests. The effects of exposure to entomopathogenic nematodes are not strictly limited to mortality, however. Here we explore non-lethal effects of exposure to entomopathogenic nematodes by introducing the relatively non-susceptible pupal stage of Delia antiqua to thirteen different strains. We specifically chose to inoculate the pupal stage because it tends to be more resistant to infection, yet resides in the soil where it could come into contact with EPN biological control agents. We find that there is no significant mortality at the pupal stage, but that there are a host of strain-dependent non-lethal effects during and after the transition to adulthood including altered developmental times and changes in risk of death compared to controls. We also find that exposure to specific strains can reduce risk of mortality. These results emphasize the strain-dependent nature of entomopathogenic nematode infection and highlight the positive and negative ramifications for non-lethal effects for biological control of insect pests. Our work emphasizes the need for strain-specific screening of biological control agents before wide-spread adoption.

The role of sheep ked (Melophagus ovinus) as potential vector of protozoa and bacterial pathogens.

The sheep ked (Melophagus ovinus) hematophagous insect may act as a potential vector of vector-borne pathogens. The aim of this study was to detect the presence of Trypanosoma spp., Bartonella spp., Anaplasma phagocytophilum and Borrelia burgdorferi sensu lato in sheep ked collected from sheep in Poland. In total, Trypanosoma spp. was detected in 58.91% of M. ovinus, whereas Bartonella spp. and B. burgdorferi s.l. were found in 86.82% and 1.55% of the studied insects, respectively. A. phagocytophilum was not detected in the studied material. In turn, co-infection by Trypanosoma spp. and Bartonella spp. was detected in 50.39%, while co-infection with Trypanosoma spp. and Bartonella spp. and B. burgdorferi s.l. was found in 1.55% of the studied insects. The conducted study showed for the first time the presence of B. burgdorferi s. l. in M. ovinus, as well as for the first time in Poland the presence of Trypanosoma spp. and Bartonella spp. The obtained results suggest that these insects may be a potential vector for these pathogens, but further-more detailed studies are required.

A Network Perspective on the Vectoring of Human Disease.

Blood-sucking insects are important vectors of disease, with biting Diptera (flies) alone transmitting diseases that cause an estimated 700 000 human deaths a year. Insect vectors also bite nonhuman hosts, linking them into host-biting networks. While the major vectors of prominent diseases, such as malaria, yellow fever, dengue, and Zika, are intensively studied, there has been limited focus on the wider interactions of biting insects with nonhuman hosts. Drawing on network analysis and visualisation approaches from food-web ecology, we discuss the value of a network perspective for understanding host-insect-disease interactions, with a focus on Diptera vectors. Potential applications include highlighting pathways of disease transmission, highlighting reservoirs of infection, and identifying emerging and previously unrecognised vectors.

Validation of loop-mediated isothermal amplification for the detection of Loa loa infection in Chrysops spp in experimental and natural field conditions.

BACKGROUND: The mass drug administration of ivermectin for onchocerciasis control has contributed to a significant drop in Loa loa microfilaria loads in humans that has, in turn, led to reduction of infection levels in Chrysops vectors. Accurate parasite detection is essential for assessing loiasis transmission as it provides a potential alternative or indirect strategy for addressing the problem of co-endemic loiasis and lymphatic filariasis through the Onchocerciasis Elimination Programme and it further reflects the true magnitude of the loiasis problem as excess human mortality has been reported to be associated with the disease. Although microscopy is the gold standard for detecting the infection, the sensitivity of this method is compromised when the intensity of infection is low. The loop-mediated isothermal amplification (LAMP) assay of parasite DNA is an alternative method for detecting infection which offers operational simplicity, rapidity and versatility of visual readout options. The aim of this study was to validate the Loa loa LAMP assay for the detection of infected Chrysops spp. under experimental and natural field conditions. METHODS: Two sets of 18 flies were fed on volunteers with either a low (< 10 mf/ml) or high (> 30,000mf/ml) microfilarial load. The fed flies were maintained under laboratory conditions for 14 days and then analysed using LAMP for the detection of L. loa infection. In addition, a total of 9270 flies were collected from the north-west, east, and south-west regions (SW 1 and 2) of Cameroon using sweep nets and subjected to microscopy (7841 flies) and LAMP (1291 flies plus 138 nulliparous flies) analyses. RESULTS: The LAMP assay successfully detected parasites in Chrysops fed on volunteers with both low and high microfilariaemic loads. Field validation and surveillance studies revealed LAMP-based infection rates ranging from 0.5 to 31.6%, with the lowest levels in SW 2 and the highest infection rates in SW 1. The LAMP assay detected significantly higher infection rates than microscopy in four of the five study sites. CONCLUSION: This study demonstrated the potential of LAMP as a simple surveillance tool. It was found to be more sensitive than microscopy for the detection of experimental and natural L. loa infections in Chrysops vectors.

Parasitylenchus myiophagus n. sp. (Nematoda: Parasitylenchidae), a tylenchid nematode parasite of long-legged flies (Diptera: Dolichopodidae).

A tylenchid nematode parasite of a male long-legged fly, Tachytrechus sanus Osten Sacken (Diptera: Dolichopodidae) from Montana, USA is described as Parasitylenchus myiophagus n. sp. (Nematoda: Parasitylenchidae). The new species is characterized by the presence of an extremely long first generation female and numerous short and wide second generation males and females produced in enclosed clusters in the hosts body cavity. Both generation female nematodes are ovoviviparous, with short stylets lacking knobs and simple tails lacking spikes, palps or mucrons. The second generation males have paired, separate spicules, short stylets, and a bursa but no visible gubernaculum. The fly host shows evidence of demasculinization, which is attributed to nematode parasitism. The gonads of the second generation adults are infected with a microsporidium (Microsporidia), which is a new host record for tylenchid nematodes. A tylenchid-infected Baltic amber dolichopodid shows that associations between these two organisms extend back at least to the Eocene.