Epidemiological exploration of fleas and molecular identification of flea-borne viruses in Egyptian small ruminants.

The study aimed to investigate molecularly the presence of flea-borne viruses in infested small ruminants with fleas. It was carried out in Egypt's Northern West Coast (NWC) and South Sinai Governorate (SSG). Three specific primers were used targeting genes, ORF103 (for Capripoxvirus and Lumpy skin disease virus), NS3 (for Bluetongue virus), and Rdrp (for Coronavirus), followed by gene sequencing and phylogenetic analyses. The results revealed that 78.94% of sheep and 65.63% of goats were infested in the NWC area, whereas 49.76% of sheep and 77.8% of goats were infested in the SSG region. Sheep were preferable hosts for flea infestations (58.9%) to goats (41.1%) in the two studied areas. Sex and age of the animals had no effects on the infestation rate (p > 0.05). The season and site of infestation on animals were significantly different between the two areas (p < 0.05). Ctenocephalides felis predominated in NWC and Ctenocephalides canis in SSG, and males of both flea species were more prevalent than females. Molecular analysis of flea DNA revealed the presence of Capripoxvirus in all tested samples, while other viral infections were absent. Gene sequencing identified three isolates as sheeppox viruses, and one as goatpox virus. The findings suggest that Capripoxvirus is adapted to fleas and may be transmitted to animals through infestation. This underscores the need for ongoing surveillance of other pathogens in different regions of Egypt.

Viruses for Landscape-Scale Therapy: Biological Control of Rabbits in Australia.

Viral diseases, whether of animals or humans, are normally considered as problems to be managed. However, in Australia, two viruses have been used as landscape-scale therapeutics to control European rabbits (Oryctolagus cuniculus), the preeminent invasive vertebrate pest species. Rabbits have caused major environmental and agricultural losses and contributed to extinction of native species. It was not until the introduction of Myxoma virus that effective control of this pest was obtained at a continental scale. Subsequent coevolution of rabbit and virus saw a gradual reduction in the effectiveness of biological control that was partially ameliorated by the introduction of the European rabbit flea to act as an additional vector for the virus. In 1995, a completely different virus, Rabbit hemorrhagic disease virus (RHDV), escaped from testing and spread through the Australian rabbit population and again significantly reduced rabbit numbers and environmental impacts. The evolutionary pressures on this virus appear to be producing quite different outcomes to those that occurred with myxoma virus and the emergence and invasion of a novel genotype of RHDV in 2014 have further augmented control. Molecular studies on myxoma virus have demonstrated multiple proteins that manipulate the host innate and adaptive immune response; however the molecular basis of virus attenuation and reversion to virulence are not yet understood.

Comparative Analysis of RNA Virome Composition in Rabbits and Associated Ectoparasites.

Ectoparasites play an important role in virus transmission among vertebrates. Little, however, is known about the nature of those viruses that pass between invertebrates and vertebrates. In Australia, flies and fleas support the mechanical transmission of two viral biological controls against wild rabbits-rabbit hemorrhagic disease virus (RHDV) and myxoma virus. We compared virome compositions in rabbits and these ectoparasites, sequencing total RNA from multiple tissues and gut contents of wild rabbits, fleas collected from these rabbits, and flies trapped sympatrically. Meta-transcriptomic analyses identified 50 novel viruses from multiple RNA virus families. Rabbits and their ectoparasites were characterized by markedly different viromes, with virus abundance greatest in flies. Although viral contigs from six virus families/groups were found in both rabbits and ectoparasites, they clustered in distinct host-dependent lineages. A novel calicivirus and a picornavirus detected in rabbit cecal content were vertebrate specific; the newly detected calicivirus was distinct from known rabbit caliciviruses, while the picornavirus clustered with sapeloviruses. Several picobirnaviruses were also identified that fell in diverse phylogenetic positions, compatible with the idea that they are associated with bacteria. Further comparative analysis revealed that the remaining viruses found in rabbits, and all those from ectoparasites, were likely associated with invertebrates, plants, and coinfecting endosymbionts. While no full genomes of vertebrate-associated viruses were detected in ectoparasites, small numbers of reads from rabbit astrovirus, RHDV, and other lagoviruses were present in flies. This supports a role for flies in the mechanical transmission of RHDV, while their involvement in astrovirus transmission merits additional exploration.IMPORTANCE Ectoparasites play an important role in the transmission of many vertebrate-infecting viruses, including Zika and dengue viruses. Although it is becoming increasingly clear that invertebrate species harbor substantial virus diversity, it is unclear how many of the viruses carried by invertebrates have the potential to infect vertebrate species. We used the European rabbit (Oryctolagus cuniculus) as a model species to compare virome compositions in a vertebrate host and known associated ectoparasite mechanical vectors, in this case, fleas and blowflies. In particular, we aimed to infer the extent of viral transfer between these distinct types of host. Our analysis revealed that despite extensive viral diversity in both rabbits and associated ectoparasites, and the close interaction of these vertebrate and invertebrate species, biological viral transmission from ectoparasites to vertebrate species is rare. We did, however, find evidence to support the idea of a role of blowflies in transmitting viruses without active replication in the insect.

Identification of diverse arthropod associated viruses in native Australian fleas.

Fleas are important vectors of zoonotic disease. However, little is known about the natural diversity and abundance of flea viruses, particularly in the absence of disease associations, nor the evolutionary relationships among those viruses found in different parasitic vector species. Herein, we present the first virome scale study of fleas, based on the meta-transcriptomic analysis of 52 fleas collected along the eastern coast of Australia. Our analysis revealed 18 novel RNA viruses belonging to nine viral families with diverse genome organizations, although the majority (72%) possessed single-stranded positive-sense genomes. Notably, a number of the viruses identified belonged to the same phylogenetic groups as those observed in ticks sampled at the same locations, although none were likely associated with mammalian infection. Overall, we identified high levels of genomic diversity and abundance of viruses in the flea species studied, and established that fleas harbor viruses similar to those seen to other vectors.

Viral Metagenomics on Blood-Feeding Arthropods as a Tool for Human Disease Surveillance.

Surveillance and monitoring of viral pathogens circulating in humans and wildlife, together with the identification of emerging infectious diseases (EIDs), are critical for the prediction of future disease outbreaks and epidemics at an early stage. It is advisable to sample a broad range of vertebrates and invertebrates at different temporospatial levels on a regular basis to detect possible candidate viruses at their natural source. However, virus surveillance systems can be expensive, costly in terms of finances and resources and inadequate for sampling sufficient numbers of different host species over space and time. Recent publications have presented the concept of a new virus surveillance system, coining the terms "flying biological syringes", "xenosurveillance" and "vector-enabled metagenomics". According to these novel and promising surveillance approaches, viral metagenomics on engorged mosquitoes might reflect the viral diversity of numerous mammals, birds and humans, combined in the mosquitoes' blood meal during feeding on the host. In this review article, we summarize the literature on vector-enabled metagenomics (VEM) techniques and its application in disease surveillance in humans. Furthermore, we highlight the combination of VEM and "invertebrate-derived DNA" (iDNA) analysis to identify the host DNA within the mosquito midgut.

It shouldn't happen to a dog or a veterinarian: clinical paradigms for canine vector-borne diseases.

Canine vector-borne diseases (CVBDs) comprise a diverse group of viral, bacterial, protozoal, and helminth pathogens, transmitted predominantly by ticks and fleas, and cause significant health problems for dogs worldwide. Growing numbers of reports indicate that CVBDs are emerging in regions where they previously did not exist and this, combined with pathogens that are inherently difficult to detect, is providing companion animal veterinarians with some significant diagnostic challenges. This review discusses six paradigms concerning the diagnosis, treatment, prevention, and zoonotic implications of CVBDs from a veterinary clinical perspective.

Understanding the persistence of plague foci in Madagascar.

Plague, a zoonosis caused by Yersinia pestis, is still found in Africa, Asia, and the Americas. Madagascar reports almost one third of the cases worldwide. Y. pestis can be encountered in three very different types of foci: urban, rural, and sylvatic. Flea vector and wild rodent host population dynamics are tightly correlated with modulation of climatic conditions, an association that could be crucial for both the maintenance of foci and human plague epidemics. The black rat Rattus rattus, the main host of Y. pestis in Madagascar, is found to exhibit high resistance to plague in endemic areas, opposing the concept of high mortality rates among rats exposed to the infection. Also, endemic fleas could play an essential role in maintenance of the foci. This review discusses recent advances in the understanding of the role of these factors as well as human behavior in the persistence of plague in Madagascar.

Avian pox in Magellanic Penguins (Spheniscus magellanicus).

Avian pox is an enveloped double-stranded DNA virus that is mechanically transmitted via arthropod vectors or mucosal membrane contact with infectious particles or birds. Magellanic Penguins (Spheniscus magellanicus) from two colonies (Punta Tombo and Cabo Dos Bahías) in Argentina showed sporadic, nonepidemic signs of avian pox during five and two of 29 breeding seasons (1982-2010), respectively. In Magellanic Penguins, avian pox expresses externally as wart-like lesions around the beak, flippers, cloaca, feet, and eyes. Fleas (Parapsyllus longicornis) are the most likely arthropod vectors at these colonies. Three chicks with cutaneous pox-like lesions were positive for Avipoxvirus and revealed phylogenetic proximity with an Avipoxvirus found in Black-browed Albatross (Thalassarche melanophrys) from the Falkland Islands in 1987. This proximity suggests a long-term circulation of seabird Avipoxviruses in the southwest Atlantic. Avian pox outbreaks in these colonies primarily affected chicks, often resulted in death, and were not associated with handling, rainfall, or temperature.

Transmission of feline calicivirus via the cat flea (Ctenocephalides felis).

In this study, a possible role of the cat flea (Ctenocephalides felis) in transmitting feline calicivirus (FCV) was examined. Fleas were fed via artificial membranes with FCV-spiked bovine blood, free of anti-FCV antibodies. Flea feces were collected daily for 10 days and incubated at room temperature. Infectivity of the feces was tested in vitro using Crandell-Reese Feline Kidney (CRFK) cells. FCV remained infectious for 8 days. These flea feces were also used to oronasally inoculate four specific pathogen-free (SPF) kittens. All kittens were successfully infected as demonstrated by virus isolation from pharyngeal swabs and seroconversion. Two of the cats showed, in addition, clinical signs. Besides the infection of cats with flea feces containing FCV, four SPF kittens were exposed to fleas that were fed with FCV-spiked bovine blood. One of the kittens was successfully infected via this route as demonstrated by virus isolation from pharyngeal swabs and virus isolation. The results of this study show that fleas can spread infectious virus through their feces or by stitch and must be considered a source of infection for uninfected cats.

[Molecular genetic variety of rickettsiae].

In this review modern data on the study of the genome of the representatives of the family Rickettsiaceae and, as follows from the accumulated information, on changes in the nomination and hierarchical place of its pathogenic and avirulent representatives are presented.

Experimental quantification of the feline leukaemia virus in the cat flea (Ctenocephalides felis) and its faeces.

Cat fleas (Ctenocephalides felis) were fed via artificial membranes and infected with the feline leukaemia virus (FeLV) from cell cultures. After removing the fleas from the blood source, the quantity of virus in the flea and its faeces was measured over a defined period of time. The virus was detectable in the fleas for up to 30 h at room temperature and up to 115 h at 4 degrees C. In the faeces, the amount of virus decreased much more slowly--after 2 weeks half of the initial amount of virus could still be detected. Thus the faeces might be a source of further infections, e.g. for the flea larvae or the cat itself.

Flea-borne Bartonella grahamii and Bartonella taylorii in bank voles.

Bartonella species are increasingly associated with a range of human and animal diseases. Despite this, we have a poor understanding of the ecology and epidemiology of many species, especially those circulating in wild populations. Previous studies have demonstrated that a diverse range of Bartonella species are abundant in wild rodent populations; little is known regarding their modes of transmission, although both direct and indirect routes have been suggested. In this study, with bank voles (Clethrionomys glareolus) as the host species, we demonstrate that the rodent flea Ctenophthalmus nobilis is a competent vector of at least two Bartonella species, B. grahamii, which has previously been associated with human infection, and B. taylorii. In contrast, no evidence of either horizontal or vertical transmission was seen in bank voles inoculated with B. taylorii maintained in an arthropod-free environment; this finding suggests that fleas may be essential for transmitting some Bartonella species.

Evidence of horizontal transmission of feline leukemia virus by the cat flea ( Ctenocephalides felis).

The feline leukemia virus (FeLV) is a naturally occurring and widespread retrovirus among domestic cats. The virus is mainly transmitted horizontally through saliva, blood and other body fluids by close contact between cats. Vectors other than cats, e.g. blood-sucking parasites, have not been reported. This study tested the vector potential of the cat flea ( Ctenocephalides felis) for FeLV. In a first feeding, fleas were fed for 24 h with blood from a FeLV-infected cat with persistent viremia. FeLV could be detected in the fleas, as well as in their feces. Fleas were then divided in two populations and fed in a second feeding for 5 h or 24 h with non-infected non-viremic blood. FeLV was again detected in the fleas and their feces. In addition, the two resulting blood samples of the second feeding were subsequently tested for FeLV and both samples were positive for FeLV RNA. The cat flea transmitted the FeLV from one blood sample to another. In a third feeding, the same populations of fleas were fed again with non-infected blood for 5 h or 24 h. This time FeLV was not detected in the fleas, or in the feces or blood samples. Results show that cat fleas are potential vectors for FeLV RNA in vitro and probably also in vivo.

[Geographical divisions of rat transmissible diseases].

In order to prevent rat transmissible diseases, it is necessary to know the geographical distribution of rats, their external parasitic arthropod and rat-borne disease in human, and to make a graph of geographical division. Stratified sampling was conducted by county to capture rats, and to sort out flea for identification of their genus and species. A total of 55,064 rats sorting as 6 families, 17 genera and 21 species were captured, and a total of 34,733 flea sorting as 4 families, 25 genera and 53 species were picked. Three kinds of rat-borne diseases, namely plague, leptospirosis and epidemic hemorrhagic fever have been occurred and epidemic in Shanxi Province. Following the general principles for geographical division, Shanxi Province was divided into 4 sub-regions: (1) basin among mountains and prairie in Yanbei; (2) loess plateau, hill and prairie with shrub in the west; (3) hill, shrub and basin, prairie with forest in the middle part; (4) hill, basin with forest and prairie in the southeast. When bubonic plague was epidemic, its transmissible mode was probably Meriones meridianus--Xenopsylla conformis--Mus musculus, Rattus norvegicus--Xenopsylla cheoopis--human. Bubonic plague has been occurred within Xingxian and Linxian conxian counties of the second sub-region. Leptospirosis has been occurred in Xiangfen county of the third sub-region, the source of infection was from pigs instead of rats. Epidemic hemorrhagic fever has been occurred in the third and fourth sub-regions, the source of infection was mainly from Ruttus norvegicus.

Horizontal transmissible protection against myxomatosis and rabbit hemorrhagic disease by using a recombinant myxoma virus.

We have developed a new strategy for immunization of wild rabbit populations against myxomatosis and rabbit hemorrhagic disease (RHD) that uses recombinant viruses based on a naturally attenuated field strain of myxoma virus (MV). The recombinant viruses expressed the RHDV major capsid protein (VP60) including a linear epitope tag from the transmissible gastroenteritis virus (TGEV) nucleoprotein. Following inoculation, the recombinant viruses induced specific antibody responses against MV, RHDV, and the TGEV tag. Immunization of wild rabbits by the subcutaneous and oral routes conferred protection against virulent RHDV and MV challenges. The recombinant viruses showed a limited horizontal transmission capacity, either by direct contact or in a flea-mediated process, promoting immunization of contact uninoculated animals.

The role of deer as a possible reservoir host of potosi virus, a newly recognized arbovirus in the United States.

Potosi (POT) virus (Bunyaviridae) was isolated from Aedes albopictus, an introduced Asian mosquito species, collected at a used tire yard in Potosi, Missouri (USA), in August and September, 1989. In September, 1990, small animals were trapped at the tire yard and six cattle were sampled at an adjacent farm; in November 1990 and 1991, blood samples were collected with filter paper strips from 364 hunter-killed, white-tailed deer (Odocoileus virginianus) in the region to determine the possible reservoir hosts of the virus. Deer specimens from Arkansas (n = 70), Colorado (n = 29), and Iowa (n = 763) (USA) were also analyzed. Specimens from 33 small vertebrates captured at the tire yard were negative for viruses. Only one eastern chipmunk (Tamias striatus) and none of six cattle had neutralizing (N) antibody against POT virus by the plaque-reduction serum neutralization test in Vero cell culture but 45 (25%) of 178 deer specimens in 1990 and 55 (30%) of 186 in 1991 were antibody positive. The 186 deer sera from 1991 were tested further and 29 (16%) were also N antibody positive to Cache Valley (CV) virus. From the 763 deer specimens tested from Iowa in 1993, 114 (15%) had N antibody to POT virus. Of 70 serum specimens from Arkansas deer in 1990, 33 (47%) had N antibody to POT and 15 (21%) to CV viruses; two (7%) of 29 CV negative serum specimens from Colorado deer in 1981 were serologically positive to POT virus. Three eastern chipmunks were experimentally inoculated with POT virus to determine their reservoir potential; none became viremic but all developed N antibody. Thus we propose that POT virus may be another virus regularly infecting wild deer populations but its impact on the health of these animals is unknown.