**2. Amphibian viruses**

#### **2.1 Ranaviruses**

Amphibian ranaviruses are enveloped icosahedral DNA viruses, in the family Iridoviridae, with variable size ranging, depending on the species [5, 7, 13]. Isolates causing disease have been found in wild and cultured amphibians in Australia, the

#### *Frog Virology: Biosafety in an Experimental Farm DOI: http://dx.doi.org/10.5772/intechopen.96605*

Americas, Asia and Europe [8–10, 15]. These include Frog virus 3, Tadpole edema virus, *Rana catesbeiana* virus Z, *Bohle iridovirus*, and UK ranavirus. Other ranavirus-like were found in captive frogs (*Rana esculenta*) in Croatia, causing lethargy, edema, hemorrhages, and skin necrosis, and also in wild-caught frogs (*Bufo marinus* and *Hopodactylus* sp.) in Venezuela. In this case infected animals had no external lesions or internal symptoms [8].

The trade of amphibians for food, research [13] and as pets contributed to the dissemination of pathogens such as ranaviruses, within and among continents [8, 9]. In North America, ranaviruses are responsible for massive mortality in amphibian larvae and recent metamorphs, while die-offs rarely occur in adults. These events often occur during summer and involve hundreds to thousands of moribund and dead larvae within a few days [8].

Ranavirus epidemics seem to occur in late spring and in summer, what can be explained by the seasonal amphibian's vulnerability to ranaviral infection when the larvae of many species begin to metamorphose. In fact, many components of the amphibian immune system are down-regulated just prior to metamorphosis [8].

Infections occur mostly in amphibians that breed in standing-water habitats [3, 24], and frog farms are associated with permanent water which may increase the exposure to the pathogen, considering that water is an effective transmission route. Animals can be sublethally infected and contain the virus over a period of at least 1 year [7]. Ranaviruses can cause asymptomatic infections in resistant animals, facilitating the spread of disease with the movement of infected animals, and contributing to the prevalence of the infection in the population [8, 10].

Frog virus 3 (Ranavirus type I), was first isolated from aclinically infected leopard frogs (*Rana pipiens*) collected in the United States in 1962 [7, 8, 10, 22]. Since then, FV3-like viruses, such as Tadpole edema virus (TEV), *Rana catesbeiana* virus Z (RCVZ), and UK Ranaviruses have been study.

In laboratory, Ranavirus was shown to cause edema, necrosis, hemorrhage, and death in embryos, tadpoles, and recent metamorphs. During experimental infections, metamorphic toads developed hemorrhages and edema in the ventral skeletal musculature, stomach, and intestines [8, 10, 24]. Mortality in embryos can occur 3 to 12 days post-exposure and clinical signs include depigmentation, skin sloughing, and spinal curvature [8, 25]. Generally, the lesions caused by FV3 appear to be milder than those caused by TEV [8].

Tadpole edema virus (Ranavirus type III) is the first acutely fatal viral infection of wild tadpoles, such as the bullfrog, *Rana catesbeiana*, bufonids (*Bufo americanus*, *Bufo woodhousei fowleri*), and pelobatids (*Spea intermontana*) [22, 25]. Present gross lesions include marked edema, erythema and hemorrhages of the skin and subcutis of the body and proximal hind limbs, hydro coelom, and petechial hemorrhages in the stomach, intestines and skeletal muscles [8, 25].

*Rana catesbeiana* virus Z was isolated from cultured *R. catesbeiana* tadpoles in the USA. RCVZ appears to be much more pathogenic than FV3, causing massive mortality of exposed tadpoles. Similar to other ranaviruses, symptoms included edema in the abdomen, hemorrhaging in ventral regions, and lethargy [8].

The contemporary strains in the United Kingdom, in common frogs (*Rana temporaria*) and in captive-breeding facilities [12, 26] worldwide, may had origin in North America [8]. Four clinical syndromes were associated with ranaviruslike particles, in English populations of the European common frog: "ulcerative syndrome", "hemorrhagic syndrome", "ulcerative and hemorrhagic syndrome" [7, 12, 26], and "reddened skin syndrome" [25]. The ulcerative form of the disease is characterized by ulcers of the skin and the skeletal muscle, and sometimes digits necrosis, while the hemorrhagic form is described with internal hemorrhages, commonly involving the gastrointestinal and reproductive tracts [12].

The second distinct amphibian ranavirus species discovered was *Bohle Iridovirus* (BIV), isolated from metamorphosed ornate burrowing frogs (*Limnodynastes ornatus*) in Australia [8, 22]. Experimentally, BIV is highly pathogenic to tadpoles and metamorphs of *L. ornatus*, and also, to tadpoles, metamorphs and adults of the giant toad, *Bufo marinus*. Lesions produced by BIV are multifocal necroses of the liver, mesonephroid, and lungs [25].

Species within the anuran family Ranidae were generally more susceptible to ranavirus infection than other family's species (Hylidae, Bufonidae, Scaphiopodidae), as shown by phylogenetic comparative methods [5, 24].

#### **2.2 Herpesviruses**

Other viral infection of the North American leopard frog is caused by the herpesvirus, and induces a form of renal adenocarcinoma, known as Lucke's renal tumor. The tumor grows during the spring and summer, with the virus being shed in the spring to infect other frogs. Renal failure occurs with weight loss and death. There is no treatment for this disorder [27].

A herpesvirus-like dermatitis with numerous dorsal and lateral epidermal vesicle, was also detected in specimens of the spring frog, *Rana dalmatina*, in a north Italy region [24]. These enveloped viruses tend to be less stable in the environment, and transmission, from one enclosure to another by human vectors, is feasible but could be prevent by good hygiene practices [18].

#### **2.3 Arboviruses**

Arboviruses are known to infect hosts by infected arthropods. Amphibians and reptiles have been studied as potential reservoir hosts of Chikungunya virus. The possible role of ectothermic vertebrates as reservoirs or overwintering hosts has been evaluated for several arboviruses, and numerous species of mosquitoes have been described to feed on a variety of reptiles and amphibians, including mosquitoes such as *Aedes aegypti* [28].

Amphibians are infected with virus through physical contact, skin exposure to contaminated water or direct ingestion of viruses [3, 29].

In one study, a frog (*Rana ridibunda*) was found to be viremic and was able to transmit the virus to *Culex pipiens*, a bloodsucker [30]. Therefore, a frogmosquito-frog cycle also appears to be possible under certain ecological conditions.

Since necrophagy and cannibalism are considered important forms in direct transmission of viruses in amphibians, both in the tadpole and metamorphosed phases, the ingestion of virus-carrying insects can also be a form of infection. Transmission by necrophagy and cannibalism is common in host species such as *Arnbystollla tigrinum*, *Rana sylvatica* and *R. latastei* [29, 31] and infections acquired by these routes appear to be more lethal.

Thus, the problem of transmission of arboviruses between amphibians and, as they may be carriers, must be taken in the spread of this class of viruses to insects and their transmission to other vertebrates (including humans).

## **3. Transmission**

Ranavirus horizontal transmission can occur via direct (necrophagy, cannibalism, [13, 17] touching, biting, [8] scavenging, virus particles persisting in the environment), or indirect routes (fomites, soil, contaminated water) [8, 12, 15]. The potential for human involvement in transmission and spread of diseases, within

#### *Frog Virology: Biosafety in an Experimental Farm DOI: http://dx.doi.org/10.5772/intechopen.96605*

and among amphibian populations, is very significant [16]. Three is no evidence of ranavirus vertical transmission [32].

Rate and infection outcome vary with the route of exposure [12, 15]. Due to nutritional and energetic limitations and physiological trade-offs, host life history characteristics such as fast development, short life span, and high fecundity can be associated with increased susceptibility to pathogens [24]. Concerning nonenveloped viruses, like iridoviruses, with a tendence to be stable in the environment, prevent spread presents a greater challenge [33].

Spread of ranaviruses may be due to water movement, via fomites sedimentation, or by sublethally/aclinically infected animals [8]. Supplying several tanks with water from a single source, and allowing the water to run through successive tanks, may contribute to a serious outbreak of diseases [32]. Larvae could become infected with ranavirus when exposed to water that previously housed infected larvae [8]. Habitats, with optimal conditions for the pathogen's persistence, may form "reservoirs" [1].

Under laboratory conditions, test animals may not be exposed to the normal array of environmental conditions (diel temperature fluctuations, exposure to proper ultraviolet-B radiation), microbial communities, or other environmental elements that could influence transmission [3]. Transmission through indirect routes had been demonstrated in the laboratory [8] and, previous laboratory studies shown that ranaviruses can persist from days to years, depending on the environmental conditions [8, 34].
