Phylogenetic Analysis of Mediterranean Mugilids by Allozymes and 16S mt-rRNA Genes Investigation: Are the Mediterranean Species of Liza Monophyletic?

The family Mugilidae (Pisces, Mugiliformes) includes species which are present in all tropical and temperate regions. Six species, Chelon labrosus, Mugil cephalus, Liza aurata, L. ramada, L. saliens, Oedalechilus labeo, are commonly found in the Mediterranean. These species have been widely studied through morphological, biochemical, and molecular markers. However, their phylogenetic relationships, and therefore the assumed monophyly of Liza species, still remain unclear: To further investigate this topic, gene-enzyme systems and sequences of the partial 16S rRNA mitochondrial gene were analyzed in Italian samples of all six Mediterranean species. The phylogenetic reconstructions indicated M. cephalus as being the most divergent species and the existence of a main cluster including all the Mediterranean species of Liza and C. labrosus. The parametric bootstrap approach adopted to test alternative phylogenetic hypotheses indicated that the Mediterranean species of Liza do not form a monophyletic group exclusive of Chelon.     

5S ribosomal RNA genes in six species of Mediterranean grey mullets: genomic organization and phylogenetic inference.

This paper describes a study of the 5S ribosomal RNA genes (5S rDNA) in a group of 6 species belonging to 4 genera of Mugilidae. In these 6 species, the relatively short 5S rDNA repeat units, generated by PCR and ranging in size from 219 to 257 bp, show a high level of intragenomic homogeneity of both coding and spacer regions (NTS-I). Phylogenetic reconstructions based on this data set highlight the greater phylogenetic and genetic diversity of Mugil cephalus and Oedalechilus labeo compared with the genera Liza and Chelon. Comparative sequence analysis revealed significant conservation of the short 5S rDNA repeat units across Chelon and Liza. Moreover, a second size class of 5S rDNA repeat units, ranging from roughly 800 to 1100 bp, was produced in the Liza and Chelon samples. Only short 5S rDNA repeat units were found in M. cephalus and O. labeo. The sequences of the long 5S rDNA repeat units, obtained in Chelon labrosus and Liza ramada, differ owing to the presence of 2 large insertion/deletions (indels) in the spacers (NTS-II) and show considerable sequence identity with NTS-I spacers. Interspecific sequence variation of NTS-II spacers, excluding the indels, is low. Southern-blot hybridization patterns suggest an intermixed arrangement of short and long repeat units within a single chromosome locus.     

Competition between mullet fry

The fry of Liza ramada, L. aurata, L. saliens, Chelon labrosus and Mugil cephalus in a lagoon in NE Spain fed mainly on zooplanktonic crustacea, such as cyclopoids, calanoids and cladocerans, but adult chironomids were also important. A high trophic and temporal overlap was found between the fry of several of these species. Trophic overlap was asymmetrical for almost all the species pairs and niche breadth expanded for the less competitive species when they were alone in the estuary. In contrast, niche breadth did not expand for the most competitive species ( L. ramada ) when it was alone. These results suggest that competition was occurring among these species, but experimental evidence is needed to prove its existence.     

A revision of the species of <Emphasis Type="Italic">Saturnius</Emphasis> Manter, 1969 (Digenea: Hemiuridae), parasites of mullets (Teleostei: Mugilidae)

The genus Saturnius Manter, 1969 is defined, its species re-examined and a key to the species presented. S. overstreeti n. sp. is described from Mugil soiuy Basilewsky and M. cephalus L. from the Russian coast of the Sea of Japan and distinguished from the morphologically related S. papernai Overstreet, 1977 and S. maurepasi Overstreet, 1977. S. segmentatus Manter, 1969 is redescribed on the basis of the type- and newly collected material from M. cephalus on the Russian coast of the Sea of Japan. The morphometric variation of S. papernai is studied based on newly collected material from Liza aurata (Risso) in the Ebro Delta and off Santa Pola, Spain. The comparisons reveal lower ranges of most metrical features than previously known. A principal component analysis, carried out after adding the new data to those of Blasco-Costa et al. (2006), confirms the species identification. Other valid species recognised are S. mugilis (Yamaguti, 1970), S. maurepasi, S. belizensis Fischthal, 1977, S. dimitrovi Blasco-Costa et al., 2006 and S. minutus Blasco-Costa et al., 2006. Forms considered species inquirendae are S. valamugilis Rekharani & Madhavi, 1984, Bunocotyle constrictus Domnich & Sarabeev, 1999 [=S. papernai of Domnich & Sarabeev (2000a, b, c, d)], B. mugilis Yamaguti, 1970 of Solonchenko (1976) and S. mugilis of Dmitrieva & Gaevskaya (2001). Host and locality information is given in detail for all species. Lisa ramado (Risso) and Chelon labrosus (Cuvier) are new host records for S. papernai (sensu stricto) and S. dimitrovi. L. aurata is a new host record for S. dimitrovi and S. minutus, and L. saliens (Risso) is a new host record for S. minutus.     

Phylogenetic relationships of nine mullet species (Mugilidae) in the Mediterranean Sea

Phylogenetic relationships among four genera and nine species (Mugil cephalus, Mugil soiuy, Liza ramada, Liza aurata, Liza abu, Liza saliens, Liza carinata, Chelon labrosus, Oedalechilus labeo) of the Mediterranean mullets (Mugilidae) were investigated by means of allozyme electrophoresis using a seven-enzyme system (CK*, GAPDH*, G3PDH*, IDHP*, ME*, MDH*, PGM*) comprising eleven putative loci. The highest genetic divergence was 1.299, detected between M. cephalus and L. aurata and the lowest (0.280) was found between L. carinata and L. saliens. The amount of genetic divergence between the genera Chelon and Oedalechilus did not appear to be high (0.285). In a UPGMA tree, all nine species were grouped in two main branchings. In the first branch, C. labrosus and O. labeo clustered as closest taxa and were sister group to L. ramada. The other four Liza species produced two sub-branching in this group; L. carinata branched together with L. saliens, and L. aurata branched together with L. abu. In the second branch the two species of the genus Mugil, M. cephalus and M. soiuy, clustered together and were clearly isolated from the other three genera.     

Parasitic diseases of marine fish: epidemiological and sanitary considerations

Over recent decades, parasitic diseases have been increasingly considered a sanitary and economic threat to Mediterranean aquaculture. In order to monitor the distribution of parasites in cultured marine fish from Italy and study their pathogenic effects on the host, a three-year survey based on parasitological and histopathological exams was carried out on 2141 subjects from eleven fish species and coming from different farming systems (extensive, intensive inland farms, inshore floating cages, offshore floating cages and submersible cages). A number of parasitic species was detected, mostly in European sea bass (Dicentrarchus labrax), gilthead sea bream (Sparus aurata), mullets (Chelon labrosus, Mugil cephalus, Liza ramada) and sharpsnout sea bream (Diplodus puntazzo), with distribution patterns and prevalence values varying in relation to the farming system, in-season period and size category. The epidemiology and pathological effects of the parasites found during the survey are discussed.     

Genetic Divergence and Phylogenetic Relationships in Grey Mullets (Teleostei: Mugilidae) Using Allozyme Data

Genetic divergence and phylogenetic relationships among five species of the Mugilidae family (Liza saliens, Liza aurata, Liza ramada, Chelon labrosus, and Mugil cephalus) were investigated, in the present study, on samples taken from Messolongi lagoon in Greece, using allozyme electrophoresis. Ten enzymic systems corresponding to 22 genetic loci were assayed, among which, four were found to be polymorphic in Liza saliens, Liza aurata, and Chelon labrosus, seven in Liza ramada, while only two were polymorphic in Mugil cephalus. Several loci showed different electrophoretic patterns among the species and thus, they can be useful in species taxonomy as diagnostic markers, as well as for further evolutionary studies. It must be underlined that among them the aGPD-2* locus proved to be species-specific, while the other ones can be also used in various combinations for the same purpose. The observed heterozygosity was found to range from 0.020 to 0.051. Allele frequencies of all loci were used to estimate Nei's (1972) genetic distance, which was found to range between 0.249 and 1.171 among the five species studied. UPGMA and NJ trees, obtained by genetic distance matrix methods, as well as, a tree based on the discrete character parsimony analysis were found to exhibit the same topology. Our result show that the three species of the genus Liza are clustered together, Chelon labrosus being closer to the previous clade, while Mugil cephalus being more distinct.     

Phenetic relationships of six species of mullets (<Emphasis Type="Italic">Mugilidae</Emphasis>) from the south Adriatic,as inferred from the study of the visceral and dermal skeleton

This study examines the interspecific variability and phenetic relationships in six southern Adriatic mullet species: Mugil cephalus (Linnaeus, 1758), Chelon labrosus (Risso, 1826), Liza aurata (Risso, 1810), Liza ramada (Risso, 1826), Liza saliens (Risso, 1810), and Oedalechilus labeo (Cuvier, 1829). Twenty-three morphometric characteristics of the visceral skeleton and the dermatocranium of mullets were analyzed. The major distinguishing characters were those of the jaws. M. cephalus was clearly separated from the other species; O. labeo differed to a somewhat smaller extent; and C. labrosus was the least distinguished species. The species of the genus Liza were phenetically most similar, while the other species of mullet differed significantly from them. The greatest distances were between M. cephalus and C. labrosus, as well as between M. cephalus and O. labeo. This text was submitted by the authors in English.     

Isoelectric focusing of lateral muscle myogen and haemoglobins of two species of Mugilidae

1. The myogen protein patterns of lateral muscle of Liza ramada (Risso) and Chelon labrosus (Risso), revealed by isoelectric focusing, are reported. Specific differences are noted both in the white and in the red muscle (where they are more evident). 2. Red muscle shows the presence of a chromoprotein, found to be myoglobin, with a pI characteristic for the species. 3. Blood haemoglobins were examined with the same technique and also found to be species specific.     

Two new species of Pseudohapladena yamaguti, 1952 (Digenea: Waretrematidae) from mugilid fish from the Taiwan Straits,China

Two new species of Pseudohapladena Yamaguti, 1952 (Digenea: Waretrematidae) are described. Pseudohapladena megoorchis n. sp. was obtained from the intestine of Mugil engeli (Bleeker) (Mugilidae), and Pseudohapladena lizae n. sp. from the intestine of Liza carinatus (Cuvier and Valenciennes) (Mugilidae). Pseudohapladena megaorchis is the most closely related in general morphology to Pseudohapladena scatophagi Yamaguti, 1952, but differs from the latter in the position of the hermaphroditic sac, and in the nature and distribution of the vitellaria. Features of P. lizae distinguishing it from other species of the genus include the shorter uterus, which contains only 1 egg, and the lesser extent of the vitellaria.     

A new natural intergeneric cyprinid hybrid from the Jordan River drainage, with a key to the large barbine cyprinids of the southern Levant

In Ain al-Qunaiya. an isolated source within the Jordan River drainage basin of Jordan, several fishes were found to have characters intermediate between two cyprinid species. Barbus canis Valenciennes in Cuvier & Valenciennes. 1842 and Capoeta damascina (Valenciennes in Cuvier & Valenciennes. 1842). The hypothesis of their hybrid origin is advanced. A key to the large barbine cyprinids of the Jordan River system and their hybrids is given.     

Morphometric variability between and within species of Ligophorus Euzet & Suriano, 1977 (Monogenea: Ancyrocephalidae) in the Mediterranean Sea

Monogeneans belonging to the genus Ligophorus Euzet & Suriano, 1977 are parasitic on the gills of several fish species of the family Mugilidae (mullets). At present, 12 species belonging to this genus have been described in the Mediterranean Sea and each host species is infected by a number of Ligophorus species that are not found on other mullets. A multivariate study using principal component and canonical variate analyses was carried out, with the aim of investigating the extent of morphometric variability within and between the 12 Mediterranean species of Ligophorus. Significant intraspecific morphometric variation was detected between two allopatric populations of L. angustus Euzet & Suriano, 1977 parasitic on Chelon labrosus from Sardinia; however, the existence of a new species of Ligophorus needs further evidence. Analysis of interspecific variation confirms the validity of the morphological keys used at present for the identification of Ligophorus at the species level and provides a picture of the relationships between species, indicating that the two species parasitising Mugil cephalus, L. chabaudi Euzet & Suriano, 1977 and L. mugilinus (Hargis, 1955) are the most differentiated. The hypothesis that strict host-parasite ecological relationships may be the result of co-speciation processes is briefly discussed.     

Molecular Phylogeny of Grey Mullets (Teleostei: Mugilidae) in Greece: Evidence from Sequence Analysis of mtDNA Segments

Mitochondrial DNA sequence analysis has been used to explore genetic differentiation and phylogenetic relationships among five species of the Mugilidae family, Mugil cephalus, Chelon labrosus, Liza aurata, Liza ramada, and Liza saliens. DNA was isolated from samples originating from the Messolongi Lagoon in Greece. Three mtDNA segments (12s rRNA, 16s rRNA, and CO I) were PCR amplified and sequenced. Sequencing analysis revealed that the greatest genetic differentiation was observed between M. cephalus and all the other species studied, while C. labrosus and L. aurata were the closest taxa. Dendrograms obtained by the neighbor-joining method and Bayesian inference analysis exhibited the same topology. According to this topology, M. cephalus is the most distinct species and the remaining taxa are clustered together, with C. labrosus and L. aurata forming a single group. The latter result brings into question the monophyletic origin of the genus Liza.     

Length–weight relationships of five mullet fish species from the Southern Taiwan coast

The length–weight relationships (LWRs) are described for five fish species, Liza affinis (Gunther, 1861), Liza alata (Steindachner, 1892), Chelon subviridis (Valenciennes, 1836), Chelon macrolepis (Smith, 1846) and Oedalechilus labiosus (Valenciennes, 1836), obtained from Southern Taiwan coast during July 2013 to October 2016.The length–weight parameter b for these species ranged from 2.734 to 2.974, with regression coefficients (r²) ranging from 0.966 to 0.979.     

Mitochondrial DNA divergence and phylogenetic relationships in mullets (Pisces: Mugilidae) of the Sea of Japan and the Sea of Azov revealed by PCR-RFLP-analysis

Three Mugilid species: Mugil cephalus (Linnaeus, 1758) and Liza haematocheila (Temminck et Schlegel, 1845; syn. Mugil soiuy, M. haematocheilus, L. soiuy, Chelon haematocheilus) from the Sea of Japan, as well as M. cephalus and Liza aurata (Risso, 1810) from the Sea of Azov were investigated on the basis of PCR-RFLP analysis of mitochondrial DNA (mtDNA) fragments, which included 12S/16S rRNA, and ND3/ND4L/ND4 genes. Among 61 individuals of three Mugilid species thirteen different haplotypes were detected. Eight and thirteen restriction endonucleases were found to be species-specific in 12S/16SrRNA and ND3/ND4L/ND4 respectively. This method may be useful for species identification. M. cephalus showed the largest genetic divergence while L. haematocheila and L. aurata were closely related and clustered together. The level of mtDNA differentiation between the two M. cephalus samples from the Sea of Japan and the Sea of Azov, i.e., nucleotide substitutions of approximately 3%, appeared to be relatively high.     

Critical thermal maxima of three species of Hawaiian estuarine fish: a comparative study*

Crticial thermal maxima (CTM) of three species of Hawaiian estuarine fishes were determined in relation to acclimation temperature and time of day. The species studied were Mugil cephalus L., Chelon engeli (Bleeker) and Kuhlia sandvicensis (Steindachner). Critical thermal maxima of all three species were significantly affected by acclimation temperature and time. In relation to acclimation temperature, Mugil cephalus and Chelon engeli have similar critical thermal maxima with Kuhlia sandvicensis having a critical thermal maxima about 1° C below that of the other two species studied. Mugil cephalus and Chelon engeli showed an increasing critical thermal maxima toward midday which decreased therafter. Critical thermal maximum of Kuhlia sandvicensis increased throughout the day.     

Genetic divergence and phylogenetic relationships in grey mullets (Teleostei: Mugilidae) based on PCR-RFLP analysis of mtDNA segments

The genetic differentiation and phylogenetic relationships among five species of the Mugilidae family (Mugil cephalus, Chelon labrosus, Liza aurata, Liza ramada, and Liza saliens) were investigated at the mtDNA level, on samples taken from Messolongi lagoon-Greece. RFLP analysis of three PCR-amplified mtDNA gene segments (12s rRNA, 16s rRNA, and CO I) was used. Ten, eight, and nine restriction enzymes were found to have at least one recognition site at 12s rRNA, 16s rRNA, and CO I genes, respectively. Several fragment patterns were revealed to be species-specific, and thus they could be useful in species taxonomy as diagnostic markers, as well as for further evolutionary studies. Seven different haplotypes were detected. The greatest amount of genetic differentiation was observed at the interspecific level, while little variation was revealed at the intraspecific level. The highest values of nucleotide sequence divergence were observed between M. cephalus and all the other species, while the lowest was found between C. labrosus and L. saliens. Dendrograms obtained by the three different methods (UPGMA, Neighbor-Joining, and Dollo parsimony), were found to exhibit in all cases the same topology. According to this, the most distinct species is M. cephalus, while the other species are clustered in two separate groups, the first one containing L. aurata and L. ramada, the other L. saliens and C. labrosus. This last clustering makes the monophyletic origin of the genus Liza questionable.     

The use of carp, Cyprinus carpio (Linnaeus, 1758), instead of rabbit as antibody donor in immunological research on fishes

Antisera produced by carp were tested in the differentiation between two cyprinid fishes, bream and roach, and between the lipovitellins of two grey mullets, Mugil cephalus and Liza ramada. These antisera were more specific in the recognition o! different fish species than those produced by rabbits.
An antiserum against carp lipovitellin was produced with male carp as antibody donor.     

The use of carp, Cyprinus carpio (Linnaeus, 1758), instead of rabbit as antibody donor in immunological research on fishes.

Antisera produced by carp were tested in the differentiation between two cyprinid fishes, bream and roach, and between the lipovitellins of two grey mullets, Mugil cephalus and Liza ramada. These antisera were more specific in the recognition of different fish species than those produced by rabbits. An antiserum against carp lipovitellin was produced with male carp as antibody donor.