Introduction

Kudoa spp. (Myxozoa, Multivalvulida) are frequent parasites of marine and estuarine fish. A recent synopsis of the species reported the existence of 95 nominal species (Eiras, Saraiva, & Cruz, 2014b) infecting a wide range of hosts all over the world. Since then, at least 6 new species were described, one of them infecting a freshwater host (Azevedo et al., 2015).

A little more than half of the parasites locate within the skeletal muscle fibers, and a number of species are especially important once they cause liquefaction of the host muscle within a short period after the death of the host which becomes useless for consumption (Cruz, Silva, & Saraiva, 2011; Gilman & Eiras 1998; Henning, Hoffman, & Manley, 2013; Moran, Whitaker, & Kent, 1999; Romero & Burgos 1996; Yokoyama & Itoh 2005; Yokoyama, Whipps, Kent, Mizuno, & Kawakami, 2004). This can occur in both wild and cultured fish, and the infection may be decisive for the economic success of fisheries and farming.

In Brazil there are only a few records of these parasites: K. aequidens infecting Aequidens plagiozonatus (Casal, Matos, Matos, & Azevedo, 2008), K. sciaenae from Stellifer minor (Oliva, Luque, Terán, & Llicán, 1992), K. crumena from Thunnus albacares, K. orbicularis described from the freshwater host Chaetobranchopsis orbicularis (Azevedo et al. 2015), and several not identified species parasitizing M. liza, Trichiurus lepturus, Plagioscion squamosissimus, M. platanus, and Scomberomorus brasiliensis (Eiras, Lima, Cruz, & Saraiva, 2014a). From the above mentioned species, only K. crumena and K. orbibularis were subjected to molecular studies. K. aequidens was described by light and transmission electron microscopic observations, and the remaining forms were characterized by light microscopy.

In this paper we report Kudoa sp. infecting O. bonariensis (Valenciennes, 1835), M. furnieri (Desmarest, 1823) and M. liza Valenciennes, 1836, captured off Rio Grande, Rio Grande do Sul State, Brazil.

Material and methods

Fresh fish specimens were acquired from fishermen at Rio Grande fish market. In the laboratory they were identified, measured (total length) and inspected for Kudoa infection. Small samples of muscle, collected from both sides of the specimens, were crashed between two glass slides and inspected at the stereomicroscope. When detected, the plasmodia were preserved in formalin at 4%, and latter 15 to 25 spores were examined at the microscope and measured according to Burger and Adlard (2010). The spores dimensions were compared using Kruskall-Wallis test, followed by multiple comparisons when significant differences (p < 0.05) were detected. The eventual myoliquefactive action of the parasites was assessed by visual inspection of the muscle to evaluate the integrity of the muscle fibers, and by palpation of the fish to detect softening of the muscle.

Results

The fish species examined, number and total length of the specimens are indicated in Table 1.

Three fish species were infected by Kudoa sp.: M. liza (2 specimens), M. furnieri (2 specimens), and O. bonariensis (4 specimens, 23.7 - 29.0 cm in total length). The intensity of the infection was low –several muscle samples were required to detect a plasmodium.

The parasites were located inside the skeletal muscles fibers in the anterior part of both sides of the body forming small intracellular elongate plasmodia, with both extremities round or slightly tapering. The dimensions of the plasmodia were similar in all the hosts, measuring 0.72 - 1.0 mm in length by about 0.07 mm wide. All the plasmodia were in the same developmental stage containing only mature spores.

Table 1.
Fish species examined for Kudoa infection, number and total length (cm) of the specimens.

The spores were also morphologically similar in all the hosts. They were bell-like shaped in lateral view. In apical view they were stellate, quadrate, having four equal radiating shell valves with margins smoothly curved, without projections or extensions. The four polar capsules were slightly pyriform, elongated, equally sized with pointed anterior extremities. The number of coils of the polar filament was not visible in optic microscopy.

Despite the similar morphology of the spores of the three different hosts there were consistent differences concerning the morphometry of the spores. Considering the three forms observed the specimens from O. bonariensis presented the highest dimensions and those of M. furnieri were smaller than M. liza for spore width and spore thickness. M. furnieri spores were clearly smaller than those from O. bonariensis and M. liza, and those from M. liza presented intermediate values - (Table 2).

Table 2.
Kudoa sp. spores measures in μm, from different hosts (mean ± s.d; (range) and N). Significant differences detected among spores measures by Kruskal-Wallis test are indicated (similar letters mean no significant differences). a.v. - apical view; l.v.- lateral view.

Discussion

The morphometry of the spores and statistical data lead us to the conclusion that the parasites from the three hosts correspond at least to two different species, one infecting M. furnieri, and another infecting O. bonariensis and M. liza. However, the existence of three different species can not be ruled out, and a molecular study of the specimens would be necessary to clarify this question.

Our specimens were compared with all the Kudoa spp. described for Brazilian hosts, muscle parasites from hosts from other sampling places, and parasites with other location than the fish muscles. From the comparison with species from Brazilian hosts it is obvious that our specimens belong to different species due to differences in the size of the spores and polar capsules.

From the about 100 nominal species described so far a little more than half of the species parasitize the muscles of the hosts. The comparison of the present material with all the species strongly suggests they correspond also to different species.

The specimens from O. bonariensis are similar to K. petala from Sillago sihama in China, though a Little smaller in length and a little bigger in width. The polar capsules of K. petala are drop-like, while in our material they are elongated, and K. petala is coelozoic in the gall bladder (Zhou & Zhao, 2008). Therefore the identity of both species is unlikely. Our specimens are also similar to K. scomberi in the somatic muscles of Scomber japonicus in Japan, forming plasmodia seen as minute white cysts, having spores longer (6.4) and thicker (8.1) than our specimens and larger drop-like polar capsules (Li et al. 2013). For these reasons the two species are considered different. Some similarities exist with K. shiomitsui detected in the pericardial cavity and heart of Takifugu rubripes (Egusa & Shiomitsu 1983). However, the spores from Brazilian hosts are not so wide, and the different location within the hosts separates both forms. Interestingly, they are very similar in dimensions to K. permulticapsula (6.0 x 9.1 x 7.7) infecting the somatic muscles of S. commerson in Australia (Whipps, Adlard, Bryant, & Kent, 2003). However, K. permulticapsula has 13 polar capsules (occasionally 14, rarely 15), which distinguishes immediately the two species without further considerations.

Concerning the specimens infecting M. liza, they do not match any species, namely those infecting Mugil spp.: the dimensions of the spores and polar capsules are higher than those of K. bora infecting the muscles of M. japonicus (Nigrelli, 1946). They are smaller than K. haridase in the gall bladder of M. persina (Sarkar & Gosh 1991), and larger tan K. intestinalis from the muscular layer of intestine of M. cephalus (Maeno, Nagasawa, & Sorimachi, 1993), and smaller than K. tetraspora located around the optic lobes of M. cephalus (Narasimhamurti & Kalavati 1979). Other differences relate to the size and shape of the polar capsules and the shape and dimensions of the plasmodia. As a consequence our forms can not be identified with any of the species infecting Mugil spp., conclusion which is re-inforced by the different location of the parasites within the hosts, excepting K. bora which infects also the muscles. Comparison with other species failed to show forms identical to the present material.

The parasites of M. furnieri are distinct from all the other species. The most identical one is K. azoni infecting Pleurogrammus azonus in the Sea of Japan (Amur Bay and Ussuri Bay). However it locates between, and not inside the muscle fibres, and causes post-mortem myoliquefaction (Aseeva, 2004), and can not be identified to the parasites from M. furnieri.

It is concluded that the Kudoa spp. Observed infecting O. bonariensis, M. liza and M. furnieri are most probably species not described hitherto, and correspond to two or three different species. However, and due to lack of molecular studies, we refrain for the moment further considerations about their identity.

Considering the high diversity of marine fish from Brazil, estimated at around 1,300 species, it is surprising that so few species of Kudoa were reported so far. The same situation occurs generally in South America. As far as we are aware only 6 species of Kudoa were reported for South American countries besides Brazil (Argentina, Chile, Peru and Uruguai), and Kudoa sp. were found in Brazil, Chile and Peru, infecting in all only a total of 14 different host species. Clearly more research is needed on these parasites. It is important to emphasize that, besides the scientific interest of the parasites, the study of its biology is important due to the host myoliquefaction caused by a number of species which may impair the commercialization and the farming of some host species.

Conclusion

The Kudoa spp. observed in fish caught off Rio Grande increase the knowledge about these parasites infecting marine fish in Brazil. Taking into account the low number of species described for the country it is emphasized the importance of the study of these parasites. Furthermore, the fish infection by Kudoa may have important economic consequences concerning the exploitation of wild and farmed fish due to the dramatic changes which some species induce in the host muscle. This is one more and important reason for the study of these parasites in Brazil.

Acknowledgement

Participation of J. C. Eiras, A. Saraiva and C. Cruz on this research was partially supported by the European Regional Development Fund (ERDF) through the Compete - Operational Competitiveness Programme, and national funds through FCT – Foundation for Science and Technology, under the project “Pest-C/MAR/LA0015/2013”. Cobselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq- 300753 / 2012-8) and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (Capes - Basic Parasitology Program - 1272/2011) supported J. Pereira Jr.

References

Aseeva, N. L. (2008). New species of Myxosporea from genus Kudoa (Myxosporea, Multivalvulida) found in muscles of some fishes of the Sea of Japan. Vestnik Zoologii, 38(2), 75-77.

Azevedo, C., Rocha, S., Matos, E., Oliveiras, E., Matos, P., Al-Quraish, S., & Casal G. (2015). Ultrastructural and phylogenetic description of Kudoa orbicularis n. sp. (Myxosporea: Multivalvulida): a parasite infecting the muscle of the fish Chaetobranchopsis orbicularis (Teleostei: Cichlidae) in the Amazon Region. Journal of Eukaryotic Microbiology, 63(1), 27-36.

Burger, M. A. A., & Adlard, R. D. (2010). Four new species of Kudoa Meglitsch, 1947 (Myxosporea: Multivalvulida) from Australia with recommendations for species descriptions in the Kudoidae. Parasitology, 137(5), 793-814.

Casal, G., Matos, E., Matos, P., & Azevedo, C. (2008). Ultrastructural description of a new myxosporean parasite Kudoa aequidens sp. n. (Myxozoa, Myxosporea), found in the sub-opercular musculature of Aequidens plagiozonatus (Teleostei) from the Amazon River. Acta Protozoologica, 47(2), 135-141.

Cruz, C., Silva, A., & Saraiva, A. (2011). Intensity of Kudoa sp. infection and alterations of muscle texture in Sardina pilchardus in Portugal. Bulletin of the European Association of Fish Pathologists, 31(6), 205-210.

Egusa, E., & Shiomitsu, T. (1983). Two new species of the genus Kudoa (Myxosporea: Multivalvulida) from marine cultured fishes in Japan. Fish Pathology, 18(3), 163-171.

Eiras, J. C., Lima, J. T. A. X., Cruz, C., & Saraiva, A. (2014a). A note on the infection of Scomberomorus brasiliensis (Osteichthyes, Scombridae) by Kudoa sp. (Myxozoa: Multivalvulida). Brazilian Journal of Biolology, 74(3, Suppl.1), 164-166.

Eiras, J. C., Saraiva, A., & Cruz, C. (2014b). Synopsis of the species of Kudoa Meglitsch, 1947 (Myxozoa: Myxosporea: Multivalvulida). Systematic Parasitology, 87(2), 153-180.

Gilman, M. M., & Eiras, J. C. (1998). Kudoa sp. (Myxosporea, Multivalvulida) infecting Sardina pilchardus (Walb.,1792) off the Portuguese coast. Research and Reviews in Parasitology, 58(2), 135-137.

Henning, S. S., Hoffman, L. C., & Manley, M. (2013). A review of Kudoa-induced myoliquefaction of marine fish species in South Africa and other countries. South African Journal of Science, 109(11/12), 1-5.

Li, Y.-C., Sato, H., Tanaka, S., Ohnishi, T., Kamata, Y., & Sugita-Konishi, Y. (2013). Characterization of the ribosomal RNA gene of Kudoa neothunni (Myxosporea: Multivalvulida) in tunas (Thunnus spp.) and Kudoa scomberi n. sp. in a chub mackerel (Scomber japonicus). Parasitology Research, 112(5), 1991-2003.

Maeno, Y., Nagasawa, K., & Sorimachi, M. (1993). Kudoa intestinalis n. sp. (Myxosporea: Multivalvulida) from the intestinal musculature of the stripped mullet, Mugil cephalus, from Japan. Journal of Parasitolology, 79(2), 190-192.

Moran, J. D. W., Whitaker, D. J., & Kent, M.L. (1999). A review of the myxosporean genus Kudoa Meglitsch, 1947, and its impact on the international industry and commercial fisheries. Aquaculture, 17(2), 163-196.

Narasimhamurti, C. C., & Kalavati, C. (1979). Kudoa tetraspora n. sp. (Myxosporidea: Protozoa) parasitic in the brain tissue of Mugil cephalus. Proceedings of the Indian Academy of Sciences, Section B, 88(1), 85-89.

Nigrelli, R. (1946). Parasites and diseases of the Ocean pout, Macrozoarces americanus (Bloch and Schneider). Bulletin of the Bingham Oceanographic Collection, 9, 198-221.

Oliva, M. E., Luque, J. L., Terán, L., & Llicán, L. (1992). Kudoa sciaenae (Myxozoa: Multivalvulidae) cysts distribution in the somatic muscles of Stellifer minor (Tschudi, 1844) (Pisces: Sciaenidae). Memórias do Instituto Oswaldo Cruz, 87(1), 33-35.

Romero, R. C., & Burgos, R. (1996). Kudoa thyrsithes (Myxozoa, Multivalvulida) causing milky condition in the musculature of Paralichthys adspersus (Neopterygii, Pleuronectiformes, Paralichthyidae) from Chile. Memórias do Instituto Oswaldo Cruz, 91(2), 163-164.

Sarkar, N. K., & Ghosh, S. (1991). Two new coelozoic myxosporida (Myxozoa: Myxosporea) from estuarine teleost fishes (Mugilidae) of West Bengal, India. Proceedings of the Zoological Society of Calcutta, 44(2), 131-135.

Whipps, C. M., Adlard, R. D., Bryant, M. S., & Kent, M. L. (2003). Two unusual myxozoans, Kudoa quadricornis n. sp. (Multivalvulida) from the muscle of goldspotted trevally (Carangoides fulvoguttatus) and Kudoa permulticapsula n. sp. (Multivalvulida) from the muscle of Spanish mackerel (Scomberomorus commerson) from the Great Barrier reef, Australia. Journal of Parasitology, 89(1), 168-173.

Yokoyama, H., & Itoh, N. (2005). Two multivalvulid myxozoans causing postmortem myoliquefaction: Kudoa megacapsula n. sp. from red barracuda (Sphyraena pinguis) and Kudoa thyrsites from splendid alfonso (Beryx splendens). Journal of Parasitology, 91(5), 1132-1137.

Yokoyama, H., Whipps, C. M., Kent, M. L., Mizuno, K. &, Kawakami H. (2004). Kudoa thyrsites from Japanese flounder and Kudoa lateolabracis n. sp. from Chinese sea bass: causative myxozoans of post-mortem myoliquefaction. Fish Pathology, 39(2), 79-85.

Zhou, Y., & Zhao, Y.-J. (2008). Description of three new species of Kudoa (Myxosporea, Multivalvulida, Kudoidae) parasitic in marine fishes from the East China Sea. Acta Hydrobiologica Sinica, 32(1), 51-56.

Notes