An investigation of culicoides (Diptera: Ceratopogonidae) as potential vectors of medically and veterinary important arboviruses in South Africa

Culicoides-borne viruses such as bluetongue, African horse sickness, and Schmallenberg virus cause major economic burdens due to animal outbreaks in Africa and their emergence in Europe and Asia. However, little is known about the role of Culicoides as vectors for zoonotic arboviruses. In this study, we identify both veterinary and zoonotic arboviruses in pools of Culicoides biting midges in South Africa, during 2012–2017. Midges were collected at six surveillance sites in three provinces and screened for Alphavirs, Flavivirus, Orthobunyavirus, and Phlebovirus genera; equine encephalosis virus (EEV); and Rhaboviridae, by reverse transcription polymerase chain reaction. In total, 66/331 (minimum infection rate (MIR) = 0.4) pools tested positive for one or more arbovirus. Orthobunyaviruses, including Shuni virus (MIR = 0.1) and EEV (MIR = 0.2) were more readily detected, while only 2/66 (MIR = 0.1) Middelburg virus and 4/66 unknown Rhabdoviridae viruses (MIR = 0.0) were detected. This study suggests Culicoides as potential vectors of both veterinary and zoonotic arboviruses detected in disease outbreaks in Africa, which may contribute to the emergence of these viruses to new regionsA US CDC Global Disease Detection grant for Zoonotic arboviruses, the NRF, the Meat Industry Trust and the Poliomyelitis Research foundation.https://medicalpapers.cn1699.com/journal/viruses-baselpm2022Medical Virolog

Use of real-time quantitative reverse transcription polymerase chain reaction for the detection of African horse sickness virus replication in Culicoides imicola

Despite its important role as vector for African horse sickness virus (AHSV), very little information is available on the dissemination of this virus in Culicoides (Avaritia) imicola Kieffer (Diptera: Ceratopogonidae). This study reports on the applicability of a real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) to detect AHSV in dissected midges. A total of 96 midges were fed on AHSV-infected blood, after which one test group was dissected into head/thorax and abdomen segments immediately after feeding and the other only after 10 days of incubation. The majority of the midges (96%) ingested the virus successfully and there was no significant difference between the virus concentration in the heads/thoraxes and the abdomens immediately after feeding. After incubation, virus was detected in 51% of the midges and it was confined to the abdomen in the majority of these. The fact that virus was detected only in the heads/thoraxes of four Culicoides midges after incubation suggests the presence of a mesenteronal escape barrier. Replication in the salivary glands was not shown. An increase of the mean virus concentration in the abdomen after incubation indicates localised viral replication. The real-time RT-qPCR is recommended for further studies investigating the replication and dissemination of AHSV in Culicoides midges

Culicoides species abundance and potential overwintering of African horse sickness virus in the Onderstepoort area, Gauteng, South Africa

In South Africa, outbreaks of African horse sickness (AHS) occur in summer; no cases are reported in winter, from July to September. The AHS virus (AHSV) is transmitted almost exclusively by Culicoides midges (Diptera: Ceratopogonidae), of which Culicoides imicola is considered to be the most important vector. The over-wintering mechanism of AHSV is unknown. In this study, more than 500 000 Culicoides midges belonging to at least 26 species were collected in 88 light traps at weekly intervals between July 2010 and September 2011 near horses in the Onderstepoort area of South Africa. The dominant species was C. imicola. Despite relatively low temperatures and frost, at least 17 species, including C. imicola, were collected throughout winter (June–August). Although the mean number of midges per night fell from > 50 000 (March) to < 100 (July and August), no midge-free periods were found. This study,using virus isolation on cell cultures and a reverse transcriptase polymerase chain reaction (RT-PCR) assay, confirmed low infection prevalence in field midges and that the detection of virus correlated to high numbers. Although no virus was detected during this winter period,continuous adult activity indicated that transmission can potentially occur. The absence of AHSV in the midges during winter can be ascribed to the relatively low numbers collected coupled to low infection prevalence, low virus replication rates and low virus titres in the potentially infected midges. Cases of AHS in susceptible animals are likely to start as soon as Culicoides populations reach a critical level

Possible over-wintering of bluetongue virus in Culicoides populations in the Onderstepoort area, Gauteng, South Africa

Several studies have demonstrated the ability of certain viruses to overwinter in arthropod vectors. The over-wintering mechanism of bluetongue virus (BTV) is unknown. One hypothesis is over-wintering within adult Culicoides midges (Diptera; Ceratopogonidae) that survive mild winters where temperatures seldom drop below 10 °C. The reduced activity of midges and the absence of outbreaks during winter may create the impression that the virus has disappeared from an area. Light traps were used in close association with horses to collect Culicoides midges from July 2010 to September 2011 in the Onderstepoort area, in Gauteng Province, South Africa. More than 500 000 Culicoides midges were collected from 88 collections and sorted to species level, revealing 26 different Culicoides species. Culicoides midges were present throughout the 15 month study. Nine Culicoides species potentially capable of transmitting BTV were present during the winter months. Midges were screened for the presence of BTV ribonucleic acid (RNA) with the aid of a real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay. In total 91.2% of midge pools tested positive for BTV RNA. PCR results were compared with previous virus isolation results (VI) that demonstrated the presence of viruses in summer and autumn months. The results indicate that BTV-infected Culicoides vectors are present throughout the year in the study area. Viral RNA-positive midges were also found throughout the year with VI positive midge pools only in summer and early autumn. Midges that survive mild winter temperatures could therefore harbour BTV but with a decreased vector capacity. When the population size, biting rate and viral replication decrease, it could stop BTV transmission. Over-wintering of BTV in the Onderstepoort region could therefore result in re-emergence because of increased vector activity rather than reintroduction from outside the region

Thermal limits of two biting midges, Culicoides imicola Kieffer and C. bolitinos Meiswinkel (Diptera: Ceratopogonidae)

BACKGROUND : Culicoides imicola Kieffer and Culicoides bolitinos Meiswinkel (Diptera: Ceratopogonidae) are both of veterinary importance, being vectors of Schmallenberg, bluetongue and African horse sickness (AHS) viruses. Within South Africa, these Culicoides species show a marked difference in their abundances according to altitude, with C. imicola highly abundant in lower altitudes, but being replaced as the dominant species by C. bolitinos in cooler, high-altitude regions. METHODS : The thermal physiology of field collected adults of each species was determined to evaluate whether it could account for differences in their distribution and abundance. Critical thermal maxima (CTmax) and minima (CTmin), as well as upper and lower lethal temperatures (ULT and LLT) were assessed after acclimation temperatures of 19°C, 24°C and 29°C. Critical thermal limits were determined using an ecologically relevant rate of temperature change of 0.06°C.min−1. RESULTS : Significant differences in CTmin and CTmax were found between acclimation temperatures for C. imicola and C. bolitinos. In C. bolitinos, the LLT of individuals acclimated at 24°C was significantly improved (LLT50 = −6.01°C) compared with those acclimated at the other temperatures (LLT50 = −4°C). Acclimation had a weak (difference in LLT50 of only 1°C) but significant effect on the LLT of C. imicola. When CTmin, CTmax, LLT and ULT were superimposed on daily maximum and minimum temperature records from locations where each tested Culicoides species is dominant, it was found that temperatures frequently declined below the CTmin and LLT of C. imicola at the location where C. bolitinos was dominant. CONCLUSIONS : The distribution and abundance of C. imicola is likely directly constrained by their relatively poor tolerance of lower temperatures. Results for C. bolitinos suggest that the adult phase is hardy, and it is hypothesised that the thermal biology of other life stages could determine their range.http://www.parasite-journal.org/hb201

Improving the diet for the rearing of Glossina brevipalpis Newstead and Glossina austeni Newstead : blood source and collection - processing - feeding procedures

One of the challenges to maintain tsetse fly (Diptera: Glossinidae) colonies is the sustainable supply of high quality blood meals. The effect of using anticoagulants during collection of the blood, the addition of phagostimulants to the blood meals as well as using mixtures of bovine and porcine blood in different proportions for feeding on colony productivity was assessed. Defibrinated bovine blood was found to be suitable to maintain both the Glossina brevipalpis Newstead and Glossina austeni Newstead colonies. Blood collected with the anticoagulants sodium citrate, citric sodium combination, citrate phosphate dextrose adenine and citric acid did not affect colony performance of both species. Defibrinated bovine and porcine blood in a 1:1 ratio or the feeding of either bovine or porcine blood on alternating days improved pupae production of G. austeni and can be used to enhance colony growth. Bovine blood is appropriate to maintain G. brevipalpis colonies, however, feeding either bovine or porcine blood on alternating days did improve productivity. Adding the phagostimulants inosine tri-phosphate, cytosine mono-phosphate and guanosine mono-phosphate to the blood at a concentration of 10−4 M improved pupae production of the G. brevipalpis colony. The addition of adenosine tri-phosphate and inosine tri-phosphate improved the performance of the G. austeni colony. Decisions on the most suitable rearing diet and feeding protocols will not only depend on the biological requirements of the species but also on the continuous supply of a suitable blood source that can be collected and processed in a costeffective way.The National Assets (000773) at the Agricultural Research Council-Onderstepoort Veterinary Institute and the Joint Food and Agriculture Organization / International Atomic Energy Agency Division of Nuclear Techniques in Food and Agriculture under the coordinated research project (CRP) 12618/R0/RBF Department of Technical Cooperation of the International Atomic Energy Agency under project RAF 5069.http://www.plosone.orgam2017Veterinary Tropical Disease

Evaluation of light emitting diode suction traps for the collection of livestock-associated Culicoides species in South Africa

Risk analysis of pathogens transmitted by Culicoides (Diptera; Ceratopogonidae) depends on the ability to detect all potential vectors attacking livestock in an area. Onderstepoort 220-V ultraviolet (UV) down-draught light traps are considered the gold standard for this purpose. To improve the flexibility of this trap in the field, in the absence of 220-V power, the possibility of using low-energy light emitting diodes (LEDs) was assessed. The efficiency of a standard 220-V Onderstepoort trap (30 cm 8 W fluorescent UV light tube) was compared to that of 220-V Onderstepoort traps fitted with either two, four or eight individual white LEDs. The Onderstepoort 220-V trap was also compared to a 12-V Onderstepoort trap fitted with an 8 W fluorescent UV light tube, a 12-V Onderstepoort trap with 12 individual white LEDs and 12-V and 220-V Onderstepoort traps fitted with 12 individual UV LEDs. Higher numbers of Culicoides as well as species diversity were collected with a brighter light source. The use of UV LEDs in both the 12-V and 220-V combinations was comparable to the Onderstepoort 220-V light trap with ration to species diversity collected. The Onderstepoort 220-V light trap is recommended if large numbers of Culicoides need to be collected.The Agricultural Research Council—Onderstepoort Veterinary Research, The South African Department of Science and Technology and the National Research Foundation of South Africa.https://onlinelibrary.wiley.com/journal/13652915hj2022Veterinary Tropical Disease

An update of the tsetse fly (Diptera: Glossinidae) distribution and African animal trypanosomosis prevalence in north-eastern KwaZulu-Natal, South Africa

An unpredicted outbreak of African animal trypanosomosis or nagana in 1990 in north-eastern KwaZulu-Natal necessitated an emergency control programme, utilising the extensive cattledipping system in the area, as well as a reassessment of the tsetse and trypanosomosis problem in the province. Since 1990, sporadic blood sampling of cattle at the dip tanks in the naganainfested areas were undertaken to identify trypanosome species involved and to determine the infection prevalence in cattle. The distribution and species composition of the tsetse populations in the area were also investigated. From November 2005 to November 2007 selected dip tanks were surveyed for trypanosome infection prevalence. During April 2005 to August 2009 the distribution and abundance of tsetse populations were assessed with odour-baited H traps. The tsetse and trypanosome distribution maps were updated and potential correlations between tsetse apparent densities (ADs) and the prevalence of trypanosomosis were assessed. Glossina brevipalpis Newstead and Glossina austeni Newstead were recorded in locations where they have not previously been collected. No significant correlation between tsetse relative abundance and nagana prevalence was found, which indicated complex interactions between tsetse fly presence and disease prevalence. This was epitomised by data that indicated that despite large differences in the ADs of G. austeni and G. brevipalpis, trypanosome infection prevalence was similar in all three districts in the area. This study clearly indicated that both tsetse species play significant roles in trypanosome transmission and that it will be essential that any control strategy, which aims at sustainable management of the disease, should target both species

Phylogenetic relationships and molecular delimitation of Culicoides Latreille (Diptera: Ceratopogonidae) species in the Afrotropical region: interest for the subgenus Avaritia

Phylogenetic relationships of Culicoides species of the Afrotropical region are problematic as different authors disagree on the placement of species into specific subgenera or groups. In this study we sequenced two mitochondrial (COI and 16S rDNA) and two nuclear (CAD and 28S rDNA) gene fragments to reconstruct phylogenetic relationships within the Avaritia, Remmia and Synhelea subgenera and the Milnei, Neavei and Similis groups of Culicoides using both Bayesian inference and maximum-likelihood approaches. Based on phylogenetic trees, we used the bGMYC (Bayesian General Mixed Yule Coalescent model) and the PTP (Bayesian Poisson Tree Processes) to investigate species boundaries. All species relationships within the studied subgenera and groups were well-supported by using morphological characters and molecular analyses. The subgenus Avaritia includes (i) all of the species of the Imicola group, as well as the putative new species, C. sp. #22, and we confirmed the monophyly of this group; (ii) the Dasyops group includes C. kanagai and C. sp. #54 Meiswinkel (new species), shown to be monophyletic; (iii) the C. sp. #20 belongs to the Orientalis group; (iv) C. grahamii, C. gulbenkiani and C. kibatiensis. Our results also show that subgenus Remmia is monophyletic. Relationships of species of the Milnei group were well-supported and demonstrate the monophyly of this group. Borkent's classification for Similis group is confirmed. In addition, C. neavei and C. ovalis (Neavei group) are placed in the subgenus Synhelea. (Résumé d'auteur