Marine leech Ozobranchus margoi parasitizing loggerhead turtle ( Caretta caretta ) in Rio Grande do Sul , Brazil

This paper reports the finding of several Ozobranchus margoi (Annelida: Hirudinea) parasitizing a loggerhead turtle (Caretta caretta) that was found in the municipality of Tavares, state of Rio Grande do Sul, southern Brazil. Since this parasite is considered to be a vector of chelonid herpesvirus 5 (ChHV-5), the leeches collected were tested for the presence of this virus. All the specimens were negative on PCR analysis. Although O. margoi is considered to be a common sea turtle parasite, this is the first official record describing collection of this parasite from a loggerhead turtle in southern Brazil, within the country’s subtropical zone. This finding draws attention to the presence of this parasite and to the risk of leech-borne infectious diseases among turtles found along the coast of southern Brazil.


Introduction
Leeches are members of phylum Annelida, class Hirudinida. Most leeches are blood-suckers and can be found as ectoparasites of several terrestrial or aquatic vertebrates (DAVIES; GOVEDICH, 2001). Ozobranchus spp. belongs to the family Ozobranchidae, characterized by the presence of typical lateral digitiform branchiae (MacCALLUM;MacCALLUM, 1918). This family includes two main species, which are permanent and exclusive ectoparasites of sea turtles: O. branchiatus and O. margoi (CHRISTOFFERSEN, 2008).
Although O. branchiatus and O. margoi can parasitize several species of sea turtles, these organisms demonstrate some degree of host preference. In general, O. branchiatus is commonly found parasitizing the green turtle (Chelonia mydas). On the other hand, O. margoi is more frequently associated with parasitism on the loggerhead turtle (Caretta caretta) (BUNKLEY-WILLIAMS et al., 2008).
A single turtle can carry more than one hundred leeches. Parasitism by Ozobranchus spp. may cause severe skin lesions, deep cutaneous erosion, eye injuries and even host death (DAVIES; CHAPMAN, 1974;SCHWARTZ, 1974;BUNKLEY-WILLIAMS et al., 2008). Another potential hazard associated with Ozobranchus spp. parasitism is the risk of chelonid herpesvirus 5 (ChHV-5) transmission. ChHV-5 is associated with the development of fibropapillomatosis (FP), a pathological condition of sea turtles characterized by occurrences of debilitating tumors in the skin and internal organs, which can progress to the animal's death (GREENBLATT et al., 2004).
Ozobranchus spp. is considered to be a common ectoparasite of sea turtles, and it has been reported to parasitize turtles in  (RICHARDSON, 1969;SCHWARTZ, 1974;DAVIES;CHAPMAN, 1974). O. margoi has also been identified parasitizing C. caretta (DAVIES; CHAPMAN, 1974). In Brazil, a great deal of anecdotal information circulates, especially among marine biologists and wild-animal veterinarians, regarding the presence of these parasites in turtles found on the coastline of tropical areas of the country. However, no official records reporting on collection and identification of O. margoi on sea turtles along the Brazilian coastland have been published, particularly in relation to the subtropical zone. Here, we report the finding of several O. margoi parasitizing a loggerhead turtle (C. caretta) that was found in the state of Rio Grande do Sul, Brazil. Also, the leeches collected were tested for the presence of ChHV-5 by means of PCR analysis.

Report and Discussion
A loggerhead sea turtle C. caretta (Figure 1a The O. margoi specimens collected ( Figure 2) were identified based on morphological characteristics (DAVIES, 1978). Adult O. margoi specimens are usually 10-15 mm long. They are white/ whitish and often show large dark spots, usually after a blood meal. The body is segmented, and divided into the trachelosome and urosome. They typically have five pairs of gills. These are thin bristle-like structures that branch out as smaller parts. The first gill pair occurs on segment XIII and is bigger and more complex than the others. From the second to the fifth pair, the gills become sequentially smaller and less complex. The mouth is terminal-ventral (DAVIES, 1978).
Some of the leeches collected were subjected to PCR analysis to detect ChHV-5. The leeches were macerated in sterile phosphatebuffered saline (PBS) solution and DNA extraction was conducted as previously established (CHOMCZYNSKI, 1993). PCR for ChHV-5 detection was carried out as described by Quackenbusch et al. (2001). The DNA samples from the leeches collected did not yield any amplicons through the PCR test, and therefore these samples were considered to be negative for ChHV-5. Although the leeches collected were negative for ChHV-5, we cannot dismiss the possibility that Ozobranchus spp. may be a vector for ChHV-5 along the Brazilian coastline. Greenblatt et al. (2004) reported that Ozobranchus spp. may carry a high viral load, reaching 10 million copies per leech. Moreover, considering the lack of reports on this subject, further studies searching for ChHV-5 and also for FP in sea turtles in southern Brazil should be conducted.
The loggerhead turtle is one of the most common sea turtles on the Brazilian coast, including in the country's subtropical zone. Nevertheless, this turtle is currently threatened with extinction, and is classified as an endangered species (IUCN, 2011). Wyneken et al. (1988) mentioned that the overall decline in sea turtle populations can be directly or indirectly attributed to the destruction of their habitats, and to anthropic action on the nesting beaches. Moreover, strong evidence suggests that predation of juveniles and eggs, predatory fishing and water pollution contribute towards this risk. In order to avoid decreases in turtle populations, conservation programs are currently in progress in Brazil (MARCOVALDI;MARCOVALDI, 1999). It is important to note that, as a further important conservation procedure, the diseases that affect sea turtles should be identified. Occurrences of parasitic and infectious diseases can impair animal health and also cause death, thus reducing sea turtle numbers. In this regard, Ozobranchus spp. represents a health hazard to sea turtles, since it can cause direct damage and also be a vector for infectious agents.
As far as we are aware, this is the first official record describing collection of O. margoi on a loggerhead turtle (C. caretta) in Southern Brazil, within the country's subtropical zone. These findings draw the attention of marine biologists, veterinarians and parasitologists to the presence of this parasite and to the risk of leech-borne infectious diseases in sea turtles on the southern Brazilian coast.