Preparedness for a possible influenza pandemic caused by highly pathogenic avian influenza A subtype H5N1 has become a global priority. The spread of the virus to Europe and continued human infection in Southeast Asia have heightened pandemic concern. It remains unknown from where the pandemic strain may emerge; current attention is directed at Vietnam, Thailand, and, more recently, Indonesia and China. Here, we report that genetically and antigenically distinct sublineages of H5N1 virus have become established in poultry in different geographical regions of Southeast Asia, indicating the long-term endemicity of the virus, and the isolation of H5N1 virus from apparently healthy migratory birds in southern China. Our data show that H5N1 influenza virus, has continued to spread from its established source in southern China to other regions through transport of poultry and bird migration. The identification of regionally distinct sublineages contributes to the understanding of the mechanism for the perpetuation and spread of H5N1, providing information that is directly relevant to control of the source of infection in poultry. It points to the necessity of surveillance that is geographically broader than previously supposed and that includes H5N1 viruses of greater genetic and antigenic diversity. genetics ͉ human ͉ influenza A ͉ virus evolution ͉ avian

Six species of the genus Myxobolus (Myxozoa) from the marine environment were collected from two species of mullet (Mugil cephalus and Liza ramada) in Ichkeul Lake, Tunisia. Four of these species were described previously (Myxobolus bizerti, Myxobolus ichkeulensis, Myxobolus spinacurvatura, and Myxobolus episquamalis) and two (Myxobolus exiguus and Myxobolus muelleri) are redescribed. The small subunit ribosomal (18S rDNA) sequences of these six myxozoans were obtained and compared to traditional criteria used in the identification and taxonomy of myxozoan species (such as spore morphology, host specificity, and tissue tropism). A distance analysis of 1,600-1,700 base pairs of the 18S rDNA of the six species indicates that they formed a monophyletic group separate from Myxobolus spp. found as parasites of freshwater fish. The sequence analyses also confirm that these morphologically different Myxobolus spp. that infect mullet represent different species. Lastly, M. exiguus and M. muelleri, which were found in the same host, exhibit clear differences in spore morphology but sequencing of two different regions of the 18S rDNA show they are closely related. These results demonstrate the utility of DNA sequence data in providing more detailed relationships among the Myxobolus species based upon existing morphological taxonomic approaches. We suggest that future descriptions of Myxobolus spp. provide both careful spore descriptions as part of the traditional criteria but also 18S rDNA sequence data that will aid in situations where morphological details may be absent or misleading.

Four species of the genus

A new Myxosporea, Zschokkella soleae sp. n., was found in the gall bladder and the bile of common sole, Solea solea (L.), from Ghar El Melh Lagoon in north-east Tunisia. This is the first record for the presence of Zschokkella species in Tunisian waters. The parasite's plasmodia are polysporic with variable size and shape. Some plasmodia appeared attached to the gall bladder epithelium while others were found free in bile. Mature spores are ellipsoidal in frontal view 13.8±0.38 μm long and 10.86±0.40 μm wide with two equal size spherical polar capsules 3.6±0.43 μm in size. The prevalence of infection seems to correlate with host size and changes over the year with maximum percentage in summer. Based on the 18S rDNA sequence data, Z. soleae sp. n. is readily distinguishable from other myxozoan DNA sequences in GenBank. Phylogenetically, the new species is placed in the freshwater Myxidium clade including several Zschokkella spp. infecting the gall bladder. Morphology, histology as well as DNA sequence analysis indicate that the examined species differs from all previously described Zschokkella species.

A new marine myxosporean species, Ceratomyxa aegyptiaca n. sp. is described from the gall-bladder of Solea aegyptiaca Chabanaud collected from the Ghar El Melh Lagoon in northeastern Tunisia. Mature spores are elongate and crescent-shaped, measuring 8-11 μm in length and 48-58 μm in width. The polar capsules are spherical, 3.2-4 μm in diameter and equal in size. Trophozoites are polysporous and float free in the bile or are attached on the epithelium of the gall-bladder. Morphological data and molecular analysis based on 18S rDNA sequences are provided. The 18S rDNA of C. aegyptiaca is readily distinguishable from that of other myxozoan species, as the genetically most similar myxozoan parasite, C. seriolae Yokoyama & Fukuda, 2001 (AB530265) collected from Seriola quinqueradiata Temminck & Schlegel in Japanese waters, shares with it only 67.5% identical nucleotides over a 1,680-bp long fragment of 18S rDNA.

Many Pauciconfibula spp. have a long and complicated taxonomic history. The remaining unsolved taxonomic confusion in this genus is impelled by the host range and status of Pauciconfibula spp. from trachinid fishes: Pauciconfibula trachini and Pauciconfibula draconis, from Trachinus radiatus and Trachinus draco (Trachinidae), respectively. Pauciconfibula trachini was reported on Trachinus draco, type host of Pauciconfibula draconis suggesting thus a stenoxenic specificity for the former monogenean and the occurrence of two congeneric polyopisthocotyleans on a single host. Moreover, the validity of Pauciconfibula draconis was repeatedly questioned by several authors, unjustified synonymy between the two species was proposed, and the delimitations between the two species remained unsolved. Original descriptions were also incomplete and poorly illustrated. In this study, we provide a detailed illustrated redescription of both species based on newly collected specimens of Pauciconfibula trachini and Pauciconfibula draconis collected from their type hosts from off three Mediterranean localities: Algeria, Tunisia, and Italy. Integrative taxonomy using COI sequences was applied to resolve the delimitation between Pauciconfibula trachini and P. draconis. This study provides the first DNA barcoding for members of this genus.

Henneguya tunisiensis n. sp., a new myxosporean, is described from the gill-arches of the East Atlantic peacock wrasse Symphodus tinca (L.) collected from off the Kerkennah Islands, Tunisia. It is characterised by the presence of elongate white plasmodia of 1-1.5 x 1.5-2 mm in size. The mature spores are rounded in frontal view and have two identical polar capsules and two caudal appendages which taper considerably at the end. Both light and electron microscopical data show that this species differs in several morphological features from all previously described Henneguya spp. A molecular analysis, based on 18S rDNA sequence data, indicates that H. tunisiensis n. sp. is readily distinguishable from other myxozoan DNA sequences in GenBank. Phylogenetically, the new species is placed in the marine Henneguya clade, which is a sister group of marine Myxobolus spp. from perciform fishes in Tunisian waters.

We describe a new myxosporean species, Zschokkella trachini n. sp., infecting the gallbladder of greater weever Trachinus draco Linnaeus 1758 from Tunisia. This is the first record of Zschokkella species in T. draco. Plasmodia were polysporic producing six to eight mature spores; they were attached to the gallbladder epithelium or free floating in the bile. Mature spores were sub-ovoid in the frontal view, measured 15.2 ± 0.6 (14.4-16.0) μm in length and 9.8 ± 0.7 (9.0-10.8) μm in width. Two equal spherical polar capsules 4.0 ± 0.4 (3.6-4.5) μm in diameter, were located separately at the spore's extremities. The prevalence of infection ranged from 23.5 to 87.7 %. Morphological data and molecular analysis of the small subunit rDNA gene identified this parasite as a new species of Zschokkella. Neighbour joining clustered the parasite in a sub-clade containing other Zschokkella species parasiting the gallbladder of marine fish hosts, located within the coelozoic clade of the major freshwater clade. This is the second Zschokkella species reported from Tunisia.