Shrimp aquaculture plays a pivotal role in global seafood production, particularly in Southeast Asia, where it significantly contributes to the economy and food security. The sustainability of this industry, however, is threatened by disease outbreaks or associated mass mortality events, including those caused by harmful dinoflagellates. Despite their significant impact, there remains a significant gap in research regarding the occurrence and diversity of harmful dinoflagellates species in tropical shrimp aquaculture ponds. A new heterotrophic harmful dinoflagellate Malayana penaeicida was discovered from this system. The thecal morphology of the cells resembled Caladoa but differs by having two anterior intercalary plates and a broader cell shape. Cells were 18.1–29 µm (22.1 ± 2.4 µm) long and 15.2–26.8 µm (18.4 ± 2.3 µm) wide. To further evaluate this lineage, molecular characterization was assessed by applying high-throughput sequencing (HTS) using Illumina MiSeq system (short-read) and Nanopore (long-read) metabarcoding approach. A concatenated phylogenetic tree of 18S-ITS-28S rDNA was obtained from independent 18S and 28S rDNA sequences from single-cell PCR and Sanger sequencing, alongside high-throughput short- and long-read sequencing. Notably, the phylogenetic tree revealed a monophyletic novel lineage, forming a close relationship with Durinskia, and nested within the Kryptoperidiniaceae. These findings emphasize the critical role of advanced high-throughput sequencing and single-cell genomics in uncovering cryptic, potentially pathogenic microalgae, and providing a powerful diagnostic baseline for aquaculture biosecurity. 

Keywords: dinophyte; harmful algal bloom; heterotrophic; metabarcoding; shrimp culture

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