Pigment-producing microorganisms have become a main focus due to the search
of food-grade microbial pigments for natural food colourant industry. Therefore,
this study aimed to isolate and genetically identify bacteria including pigmentproducing
bacterial strains associated with the tentacles of Holothuria
(Lessonothuria) pardalis (Selenka 1867) from Pulau Tinggi, Johor.
Morphological characteristics of bacteria based on Gram staining were observed
using a digital compound microscope. Partial 16S ribosomal RNA (rRNA) gene
sequencing was also incorporated to support the morphological approach. A
number of six partial gene sequences of non-protein coding 16S rRNA were
obtained and the Basic Local Alignment Search Tool (BLAST) analysis
suggested the presence of three known bacteria species from the genera of
Pseudomonas, Chryseobacterium and Klebsiella. The neighbourhood-joining
method and Maximum Likelihood method used in the phylogenetic analysis
supported the identification and classification of the bacteria species. Among the
bacterial isolates, only strain B14 identified as Chryseobacterium sp. produced
pigment i.e. orange pigment. The highest point of cell density of
Chryseobacterium sp. was recorded at 16 hours. However, further analysis is
needed to confirm the pigment characterisation and the potential of this pigment
to be exploited as natural colourant in the food industry.
Nursyamfarhani Akashah, Siti Najihah Solehin, Marsheilla Lia Janing, Nur Aina Yusra Manap, Nor Shahida Ab Rahman, Muhammad Abdul Latiff Abu Bakar, et al.
Sea cucumber or locally known as gamat and timun laut is a well-known attraction
of Pulau Langkawi, Kedah, Malaysia. Not only limited to Pulau Langkawi, sea
cucumber can also be found in other coastal water areas of Malaysia. However,
the species richness and genetic diversity of sea cucumber in Pulau Tinggi, Johor
is still unknown to the marine scientific communities, especially for Holothuria
(Mertensiothuria) leucospilota, the most dominant species in Malaysia.
Therefore, this study aimed to generate DNA barcodes of H. leucospilota from
Pulau Tinggi. Protein-coding cytochrome c oxidase 1 (CO1) mtDNA gene was
used as it is commonly and widely used for molecular species identification via
DNA barcoding. Polymerase chain reaction (PCR) and DNA sequencing were
incorporated to generate the DNA barcodes of CO1 mtDNA gene. The BLAST
program confirmed the species status of the sea cucumber specimens as H.
leuscospilota or locally known as bat puntil or white threads fish. The application
for GenBank, NCBI, U.S. National Library of Medicine registration was
successfully accepted on October 17, 2019 with accession numbers of MN580537
and MN580538). The phylogenetic trees i.e neighbor joining tree (NJ, distancebased
method), maximum parsimony tree (MP, character-based method) and
maximum likehood tree (ML, character-based method) reconstructed using
MEGA X software version 10.0.5 (BETA) further supported the species status of
the sea cucumber specimens from Pulau Tinggi as H. leuscospilota. This is the
first record of CO1 mtDNA barcodes of H. leuscospilota from Pulau Tinggi,
Johor, Malaysia.