Interruption of the transmission of Onchocerca volvulus in the Kashoya-Kitomi focus, western Uganda by long-term ivermectin treatment and elimination of the vector Simulium neavei by larviciding.

Uganda is the only country in sub-Saharan Africa whose onchocerciasis elimination programme extensively uses vector control and biannual treatment with ivermectin. The purpose of this study was to assess the impact of combined strategies on interrupting onchocerciasis transmission in the Kashoya-Kitomi focus. Mass Drug Administration annually (13 years) followed by biannual treatments (6 years) and ground larviciding (36 cycles in 3 years) with temephos (Abate®, EC500) against Simulium neavei were conducted. Routine fly catches were conducted for over seven years in six catching sites and freshwater crabs Potamonautes aloysiisabaudiae were examined for immature stages of Simulium neavei. Epidemiological assessments by skin snip were performed in 2004 and 2013. Collection of dry blood spots (DBS) from children <10 years for IgG4 antibodies analysis were done in 2010 and 2013. Treatment coverage with ivermectin improved with introduction of biannual treatment strategy. Microfilaria prevalence reduced from 85% in 1991 to 62% in 2004; and to only 0.5% in 2013. Crab infestation reduced from 59% in 2007 to 0% in 2013 following ground larviciding. Comparison of total fly catches before and after ground larviciding revealed a drop from 5334 flies in 2007 to 0 flies in 2009. Serological assays conducted among 1,362 children in 2010 revealed 11 positive cases (0.8%; 95% CI: 0.4%-1.2%). However, assessment conducted on 3246 children in 2013 revealed five positives, giving point prevalence of 0.15%; 95% CI: 0.02%-0.28%. Four of the five children subjected to O-150 PCR proved negative. The data show that transmission of onchocerciasis has been interrupted based on national and WHO Guidelines of 2012 and 2016, respectively.

Inverse density dependence of parity rates in the onchocerciasis vector Simulium damnosum s.l.

A correlation between parity rates and an index of adult numbers of Simulium damnosum s.l. (Diptera: Simuliidae) indicates an association, but does not prove causality or show the direction of any causal relationship. The question of whether adult numbers affect parity rates or vice versa is reminiscent of the age-old query of which of the chicken and the egg came first. A method for resolving such issues based on analyses of pairs of time series was proposed by Granger in 1969. When Granger's method was applied to monthly numbers of adult female S. damnosum s.l. caught attempting to bite humans at Asubende, Ghana, and their parity rates, a significant relationship (P = 0.005) emerged, clearly showing that parity rates were dependent on adult numbers. Implications of this inverse density dependence and the results of analyses of other similar time series are presented and discussed.

Assessments of the transmission of Onchocerca volvulus by Simulium sanctipauli in the Upper Denkyira District, Ghana, and the intermittent disappearance of the vector.

Following studies on the transmission of Onchocerca volvulus (Leuckart) by Simulium sanctipauli Vajime & Dunbar (Diptera, Simuliidae) in Upper Denkyira District in Ghana in 2001 and 2002 (Kutin et al., Med Vet Ent 18:167-173, 2004), further assessments were carried out in 2006 and 2013/2014 to determine whether transmission parameters had changed since community-directed ivermectin treatment (CDTI) began in 1999. There were no marked changes of the transmission intensities in 2006. Only slight, but non-significant, reductions were observed in infection rates of parous flies with larval stages (L1-L3) of O. volvulus from 44.1 % (of 1672 parous flies) in 2001/2002 to 42.1 % (506) in 2006 and from 6.5 to 5.9 % of flies carrying infective larvae in their heads. This suggested that there was an ongoing transmission in the area and the parasite reservoir in the human population was still high. Unexpectedly, further assessments conducted in October 2013 and March and October 2014 revealed that the vector S. sanctipauli had apparently disappeared and transmission had ceased, probably as a result of intensified gold mining activities along the rivers Ofin and Pra. The water of both rivers was extremely turbid, heavily loaded with suspended solids, probably preventing the development of blackfly larvae. Some breeding and biting of Simulium yahense Vajime & Dunbar was observed in a small tributary of the Pra, the Okumayemfuo, which is not affected by gold mining. However, the infection rate of flies was low, only 3.7 % of 163 parous flies were infected with first stage (L1) larvae of O. volvulus.

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.

The disappearance of onchocerciasis from the Itwara focus, western Uganda after elimination of the vector Simulium neavei and 19 years of annual ivermectin treatments.

The Itwara onchocerciasis focus is located around the Itwara forest reserve in western Uganda. In 1991, annual treatments with ivermectin started in the focus. They were supplemented in 1995 by the control of the vector Simulium neavei, which was subsequently eliminated from the focus. The impact of the two interventions on the disease was assessed in 2010 by nodule palpations, examinations of skin snips by microscopy and PCR, and Ov16 recombinant ELISA. There was no evidence of any microfilaria in 688 skin snips and only 2 (0.06%) of 3316 children examined for IgG4 were slightly above the arbitrary cut off of 40. A follow up of the same children 21 months later in 2012 confirmed that both were negative for diagnostic antigen Ov-16, skin snip microscopy and PCR. Based on the World Health Organization (WHO) elimination criteria of 2001 and the Uganda onchocerciasis certification guidelines, it was concluded that the disease has disappeared from the Itwara focus after 19 years of ivermectin treatments and the elimination of the vector around 2001. Ivermectin treatments were recommended to be halted.

The elimination of the vector Simulium neavei from the Itwara onchocerciasis focus in Uganda by ground larviciding.

The Itwara focus of onchocerciasis covers an area of approximately 600 km(2) in western Uganda about 20 km north of Fort Portal. The vector is Simulium neavei, whose larvae and pupae live in a phoretic association on freshwater crabs. The phoretic host in the Itwara focus is the crab Potamonautes aloysiisabaudiae. Before any onchocerciasis control, ATPs were estimated to reach between 4500 and 6500 infective larvae per person per year. S. neavei was found to be a very efficient vector with 40% of parous flies harbouring developing larvae of Onchocerca volvulus. After 4 years of community-based distribution of ivermectin transmission was still considerable and in 1995 monthly treatment of streams with the larvicide temephos commenced in the first of three sub-foci, and was gradually extended to the whole focus. Biting S. neavei disappeared from the first sub-focus (Itwara main) in June 1996, and the last infested crab was caught in November 1996. In the second sub-focus (Siisa) treatment commenced towards the end of 1995, and the last biting fly was caught in March 1997, but a deterioration in the security situation interrupted the programme (after only three treatments in the third sub-focus). Monthly treatments restarted in the second and third sub-foci (Aswa) in September 1998, and when the situation was reassessed in 2003 no biting flies were found anywhere, and the flies had not reinvaded the first sub-focus, but infected crabs were found in the second and third sub-foci. The last treatments were carried out in April-June 2003, and since then no infested crabs have been found. In summary, no S. neavei-infested crabs have been found anywhere in the focus since June 2003 and the vector is considered eliminated from that date. However, transmission had already been halted since February 2001, when the last biting flies had been collected. The parasite reservoir should die out in the human population by 2016.

Efficiency of Simulium sanctipauli as a vector of Onchocerca volvulus in the forest zone of Ghana.

The role of Simulium sanctipauli Vajime & Dunbar (Diptera: Simuliidae) as a vector of Onchocerca volvulus (Leuckart) (Spirurida: Onchocercidae) in the forest zone of central Ghana was studied in the Upper Denkyira district, where onchocerciasis is hyperendemic. Simulium sanctipauli was found to be a highly efficient vector, with a mean of 377 infective (L3) larvae in the heads of 1000 parous and 122 in the heads of 1000 biting flies. The overall infection rate of 44% of the parous flies with L1, L2 and L3 stages of O. volvulus (identity confirmed by polymerase chain reaction) demonstrates marked anthropophily. Female flies dispersed over a wide area and can transmit onchocerciasis up to at least 10 km away from their breeding sites. Annual community-directed treatments with ivermectin did not have a noticeable effect on the infection rates and parasitic loads of fly populations, which were as high 2 months after as 3 months before the distribution of ivermectin. This failure can be attributed to poor coverage, with treatment taken by only 24.4% of the population of the six study villages.

Dispersal range of Simulium neavei in an onchocerciasis focus of western Uganda.

The dispersal range of the vector Simulium neavei Roubaud (Diptera: Simuliidae) was investigated in an onchocerciasis focus in Kabarole district, western Uganda, adjacent to the Itwara Forest Reserve, where the species breeds in phoretic association with the river crab Potamonautes aloysiisabaudiae (Nobili) (Decapoda: Potamonautidae). Biting rates were highest at the edge of the forest, but blood-seeking flies were collected up to a distance of about 4 km from the forest. Entomological data did not agree with the geographical distribution of onchocerciasis, which is hyperendemic in larger distances from the forest. The possible reasons are discussed.

Molecular identification and phylogeny of East African Simulium damnosum s.l. and their relationship with west african species of the complex (Diptera: Simuliidae).

The phylogenetic relationships of East and West African members of the Simulium damnosum complex were studied by sequence analyses of the mitochondrial 16s ribosomal RNA (rRNA) gene. Results suggest that: (i) the S. damnosum complex is divided into an East and a West African clade, and (ii) S. pandanophilum and the cytoform 'Kiwira' form an East African subbranch distinct from the 'Sanje' group. In contrast to former assumptions from cytogenetic analyses, our molecular data do not support a direct relationship between the East African S. kilibanum and the West African S. squamosum. Length differences of the rDNA internal transcribed spacer 1 (ITS-1) turned out to be useful to distinguish between cytoforms.

Dispersal range of Simulium neavei in an onchocerciasis focus of western Uganda

Dispersal range of the vector Simulium neavei Roubaud (Diptera: Simuliidae) was investigated in an onchocerciasis focus in Kabarole district; western Uganda; adjacent to the Itwara Forest Reserve; where there species breeds in phoretic association with the river crab Potamonautes aloysiisabaudiae (Nobili) (Decapoda: Potamonautidae). Biting rates were highes at the edge of the forest; but blood-seeking flies were collected to a distance of about 4 km from the forest. Entomological data did not agree with the geographical distribution of onchocerciasis; which is hyperendemic in larger distances from the forest. The possible reasons are discussed

The Simulium damnosum complex in western Uganda and its role as a vector of Onchocerca volvulus.

The status of onchocerciasis vectors in the former Ruwenzori focus in western Uganda was re-examined some 15 years after control measures against Simulium damnosum s.l. were suspended. The four cytoforms S. kilibanum, 'Sebwe', 'Nkusi' and S. pandanophilum were found. While the nonanthropophilic 'Sebwe' was still widely distributed in rivers north, east and south of the Ruwenzori, the only anthropophilic species and vector, S. kilibanum, had disappeared from most of its former habitats and was now restricted to two limited foci, where high biting densities were encountered. It was still a vector south of the Ruwenzori (Kasese focus), where 15.4% of the parous flies were infected with larval stages of Onchocerca volvulus and 34 infective larvae were found in the heads of 1000 parous flies. In the second focus along the Mahoma and Nsonge rivers, a chromosomally highly polymorphic population of S. kilibanum had replaced the former vector S. neavei, but does not act as a vector. Only 2.3% of the parous females were infected and just 1 infective larva was found in the heads of 1000 parous flies.

Morphometric characterization of members of the Simulium damnosum Theobald complex (Diptera: Simuliidae) from East and West Africa.

Morphometric multivariate analyses were carried out in order to separate the females of the closely related anthropophilic and non-anthropophilic members of the Simulium damnosum s.l. complex occurring in western Uganda. Simulium kilibanum (= 'Nyamagasani'), 'Nkusi', 'Sebwe' and S. pandanophilum were compared with the West African S. damnosum subcomplex, S. soubrense and S. yahense to correlate the results with those of previous studies. Simulium pandanophilum could be clearly discriminated. However, only 72%-88% of identifications among the closely related S. kilibanum, 'Sebwe' and 'Nkusi', which all belong to the 'Sanje' subcomplex, were correct. The two forest species S. soubrense and S. yahense slightly overlapped but were distinct from all others.

A new species of the Simulium damnosum complex from Uganda, and comparative morphology of the tarsal claws in females of this complex.

The female, male, pupa and larva of Simulium (Edwardsellum) pandanophilum sp. nov. (Diptera: Simuliidae) from western Uganda are described. Diagnostic features are the shape of the tarsal claws of females and, cytogenetically, new intraspecific inversions on chromosomes I and II. Simulium pandanophilum is assigned to the Simulium damnosum complex. Different tarsal claw shapes of West African members of the S. damnosum complex are also described.

PCR and DNA hybridization indicate the absence of animal filariae from vectors of Onchocerca volvulus in Uganda.

In order to identify Onchocerca volvulus larvae from vectors, DNA of filaria larvae from dissected blackflies was isolated, and a 150-bp long tandemly repeated DNA sequence (0-150), which occurs in many Onchocerca species, was amplified using polymerase chain reaction (PCR). Subsequently, the PCR product was blotted onto a nylon membrane and hybridized with DNA probes specific for O. volvulus or Onchocerca ochengi. Filaria larvae from 395 infected Simulium neavei were examined and 259 samples produced detectable PCR products. Among these samples, 239 (92%) reacted with an O. volvulus-specific oligonucleotide. A sample of 69 PCR products was tested using an O. ochengi DNA probe, but all failed to hybridize. Filaria larvae from 64 infected Simulium damnosum, presumably of the cytotypes "Nyamagasani" and "Nkusi" were studied and 0-150 was amplified from 38 samples. From these samples, 35 (92%) hybridized specifically with an O. volvulus probe but none with the O. ochengi-specific DNA sequence. Nonamplified samples were obtained mainly from blackflies that contained only 1 or 2 filaria larvae, and therefore, an insufficient DNA extraction was assumed. It can be concluded that few, if any, filaria species of animal origin were transmitted by S. neavei and S. damnosum s.l. in Kabarole and Kasese districts in Uganda.

Reduced prevalence of onchocerciasis in Uganda following either deforestation or vector control with DDT.

To determine the prevalence of onchocerciasis in western Uganda following deforestation and vector control, three foci were re-examined 20 years after previous surveys. In the Ruteete focus Simulium neavei had apparently disappeared and the prevalence of onchocerciasis declined in adults from about 70% in 1971 to a standardised prevalence of 12% in 1992. An increase of population density together with extended deforestation was assumed as cause of this strong reduction. In Bugoye, a S. damnosum s.l. focus, the standardised prevalence of microfilaria carriers declined from 62% in 1972 to 4.7% in 1992. Entomological data indicated the absence of man biting blackflies in the nineties. It can be suggested that the vector control using DDT performed during the seventies had lead to a change of the species composition from anthropophilic to non-anthropophilic S. damnosum s.l. In the focus Kicheche environmental changes were insignificant, deforestation was not progressive and S. neavei was abundant. Here the standardised prevalence of microfilaria carriers was still high (61%).

The vectorial efficiency of Simulium yahense populations in south-eastern Ghana.

Adult female Simulium yahense were identified amongst catches of S. damnosum s. 1. caught at Tsatsadu Falls and Wegbe in south-eastern Ghana. Dissections of stained flies to assess the species' role as a vector of onchocerciasis in the region revealed as many as 357 L3 per 1000 biting flies in a sample from Wegbe. When the data from the two sites were combined 154 L3 per 1000 biting flies and 315 L3 per 1000 parous flies were recorded. The values for wet season samples were about 12% higher than those for the dry season. The results are discussed in relation to other studies of S. yahense populations and regarding the species' recent spread in Ghana and Togo.

Seasonal variation in onchocerciasis transmission by Simulium squamosum at perennial breeding sites in Togo.

Analyses of data collected routinely by the World Health Organization Onchocerciasis Control Programme in West Africa showed that the transmission of onchocerciasis by Simulium squamosum at Amou-Oblo, in Togo, was maximal in the dry season. Mean monthly transmission potentials ranged from 10.4 in August to 519.6 in February. Infectivity in terms of numbers of L3 larvae in the head per 1000 parous flies was highest in March. Additional, more detailed, studies involving the staining of flies from Amou-Oblo, Djodji and Tinkiro confirmed these seasonal trends but also revealed significantly higher rates with L1/L2 larvae in the dry season. The results are discussed in relation to seasonal changes in fly numbers, fly longevity, fly size and man-fly contact rates.

A temporary focus of savanna species of the Simulium damnosum complex in the forest zone of Liberia.

Savanna species of the Simulium damnosum complex, the major vectors of the blinding savanna type of onchocerciasis, were considered to be rare in Liberia, until the dry season of 1988. In 1988 they became a serious nuisance, biting people at the Bong iron ore mine within the rain forest zone. S. damnosum s.str. and S. sirbanum were found breeding, in association with S. adersi a non man-biting savanna species, in a stream emerging from the mine's tailings pond. The local forest species were extremely rare in this stream. The water of the stream was characterized by an increased hardness and higher temperatures in comparison with those of natural watercourses in the area. The mass occurrence of flies was probably related to expansions of the tailings ponds, when rich nutrition was provided for the blackfly larvae by the decaying forests submerged by the rising water levels. The phenomenon did not recur in 1989 when only a few savanna flies were caught, and none were seen in the dry season of 1990. Although it is not known why the artificial environment of the tailings ponds was so attractive for the savanna species the events clearly demonstrated that savanna flies seasonally invading the area, possibly aided by the northeasterly harmattan winds, can become established in the rain forest zone if suitable conditions are met. No infections with Onchocerca volvulus were found in more than 1000 flies caught by vector collectors but, after experimental infection with the local forest strain, a few parasites developed to the infective stage. As yet there is no evidence that the occurrence of savanna flies in the rain forest zone of Liberia was of epidemiological significance.

The relationship between innate susceptibility to Onchocerca, and haemolymph attenuation of microfilarial motility in vitro using British and west African blackflies.

The effect of haemolymph from individual flies of differing species of Simulium were compared in their ability to attenuate Onchocerca microfilariae in vitro. Small maintenance systems, each of 3-4 microliters total volume, were set up in Terasaki plates. Three British species of greatly differing susceptibilities to Onchocerca lienalis (Simulium ornatum, S. equinum and S. vernum) were compared in their effect on O. lienalis. Three forms of the S. damnosum complex from Liberia (S. yahense and 2 geographical variants of S. soubrense) were similarly compared in their effects on O. volvulus microfilariae. The more refractory species in both sets of experiments were found to significantly reduce the in vitro motility of the microfilariae more than the susceptible species. It is proposed that this innate variation in in vitro attenuation, may be caused be humoral haemolymph proteins. Such an assay, perhaps in a modified form could be used in predicting vector susceptibility. It could also be used to analyse mechanisms responsible for variation in susceptibility, of different S. damnosum complex members, to the geographical forms of O. volvulus.