Genetic diversity and patterns of demographic expansion in natural populations of milkfish, Chanos chanos (Forsskål, 1775), in the Philippines.

The milkfish, Chanos chanos (Forsskål, 1775) is an important aquaculture species produced in the tropical Pacific region. Milkfish aquaculture is heavily reliant on seedstock; thus, there is a need to assess the genetic diversity of wild populations to identify areas that could be sources of high quality seedstock and broodstock. In this study, 631 milkfish specimens from 11 Philippine and, for comparison, two Indonesian wild populations were analysed for their genetic diversities using mitochondrial control region and cytochrome b. High genetic diversity indices were found for Sarangani, Claveria, and Camarines Sur populations in the Philippines. Philippine samples had either comparable or higher genetic diversity values than Indonesian samples from Medan and Sulawesi. Based on neutrality tests and mismatch distribution analysis, Philippine samples exhibited population expansion. Such pattern was not observed among Indonesian specimens. Genetic structure among populations was weak based on analysis of molecular variance (AMOVA). Correlation between genetic distance and geographic distance was not significant. Differentiation among marine biogeographic regions was small based on hierarchical AMOVA. High gene flow due to natural dispersal and influx of haplotypes brought about by importation may have contributed to the patterns observed, but diversification events within the Philippines were still evident.

Exposure to Aeromonas hydrophila induces inflammation and increases expression of the gene encoding for a putative dual CTLD-containing lectin in milkfish liver.

Milkfish (Chanos chanos Forsskal) is an important aquaculture product and is the sole extant species of the family Chanidae (order Gonorynchiformes). While there are already several reports regarding milkfish aquaculture, studies on milkfish immunity and gene expression are very limited. In this study, we showed that Aeromonas hydrophila induces inflammation in milkfish liver. We identified a milkfish C-type lectin-like domain containing proteins (CTLDcps) gene, designated as CcClec, which was upregulated in respond to A. hydrophila. Full-length sequencing was performed using Rapid Amplification of cDNA Ends (RACE PCR) to produce a complete Coding DNA Sequence (CDS) of the gene. The CcClec gene encoded a predicted protein of 340 amino acids containing two CTLDs that may potentially bind carbohydrates, especially sucrose and cellobiose. The CcClec mRNA transcript was expressed highest in the liver, followed by head kidney, brain, heart, gills, spleen, and midgut. CcClec transcripts were upregulated in damaged liver upon exposure to A. hydrophila. Overall, these findings demonstrated that CcClec is implicated in milkfish innate immunity, and is most highly expressed in the liver, suggesting a role of the liver in the milkfish immune system.

DNA barcoding of feral tilapias in Philippine lakes.

Tilapia (Oreochromis mossambicus) was first introduced to the Philippines in 1950 for aquaculture. Since then, other species of tilapia have been introduced to the country and some of them (mainly Oreochromis niloticus) have become established in lakes and other water bodies. In this study, DNA barcoding using the mitochondrial cytochrome c oxidase subunit I (COI) gene was done to assess the reliability of morphological identification and the degree of introgression among feral tilapias (Oreochromis spp.) in seven major Philippine lakes, namely Laguna de Bay, Lake Lanao, Taal Lake, Lake Mainit, Lake Naujan, Lake Bato, and Lake Buhi. Specimens were also collected from a private hatchery in Sual, Pangasinan to serve as reference. Morphological traits, Nucleotide BLAST (BLASTn), and Translated BLAST (BLASTx) analyses were used to classify the specimens. A Neighbor-Joining tree was constructed using the Kimura 2-Parameter method, incorporating 66 COI sequences generated from the study and 20 additional reference sequences obtained from GenBank. Three Oreochromis clusters were obtained and were classified as the O. niloticus group, O. mossambicus group, and O. aureus group, with bootstrap support values of 99%, 74%, and 99%, respectively. The mean K2P genetic distances within each group were 0.008%, 0.959%, and 0.086%, respectively. The clustering of COI sequences generated from this study corresponded with the results of the BLASTn analysis. Oreochromis hybrids were also found in all the lakes. The study highlights the usefulness of DNA barcoding for molecular identification and detection of introgressed individuals, with potential applications in management of feral stocks.

DNA barcoding of fishes of Laguna de Bay, Philippines.

Laguna de Bay, the largest lake in the Philippines, is an important part of the country's fisheries industry. It is also home to a number of endemic fishes including Gobiopterus lacustris (Herre 1927) of family Gobiidae, Leiopotherapon plumbeus (Kner 1864) of family Terapontidae, Zenarchopterus philippinus (Peters 1868) of family Hemiramphidae and Arius manillensis Valenciennes 1840 of family Ariidae. Over the years, a steady decline has been observed in the abundance and diversity of native fishes in the lake due to anthropogenic disturbances. In this study, a total of 71 specimens of 18 different species belonging to 18 genera, 16 families, and seven orders were DNA barcoded using the mitochondrial cytochrome c oxidase subunit I (COI) gene. All of the fish species were discriminated by their COI sequences and one endemic species G. lacustris, showing deep genetic divergence, was highlighted for further taxonomic investigation. Average Kimura 2-parameter genetic distances within species, family, and order were 1.33%, 18.91%, and 24.22%, respectively. These values show that COI divergence increases as taxa become less exclusive. All of the COI sequences obtained were grouped together according to their species designation in the Neighbor-joining tree that was constructed. This study demonstrated that DNA barcoding has great potential as a tool for fast and accurate species identification and also for highlighting species that warrant further taxonomic investigation.

DNA barcoding of the ichthyofauna of Taal Lake, Philippines.

This study represents the first molecular survey of the ichthyofauna of Taal Lake and the first DNA barcoding attempt in Philippine fishes. Taal Lake, the third largest lake in the Philippines, is considered a very important fisheries resource and is home to the world's only freshwater sardine, Sardinella tawilis. However, overexploitation and introduction of exotic fishes have caused a massive decline in the diversity of native species as well as in overall productivity of the lake. In this study, 118 individuals of 23 native, endemic and introduced fishes of Taal Lake were barcoded using the partial DNA sequence of the mitochondrial cytochrome c oxidase subunit I (COI) gene. These species belong to 21 genera, 17 families and 9 orders. Divergence of sequences within and between species was determined using Kimura 2-parameter (K2P) distance model, and a neighbour-joining tree was generated with 1000 bootstrap replications using the K2P model. All COI sequences for each of the 23 species were clearly discriminated among genera. The average within species, within genus, within family and within order percent genetic divergence was 0.60%, 11.07%, 17.67% and 24.08%, respectively. Our results provide evidence that COI DNA barcodes are effective for the rapid and accurate identification of fishes and for identifying certain species that need further taxonomic investigation.