Establishment of Nested PCR for the Detection of Pseudomonas plecoglossicida and Epidemiological Survey of Larimichthys crocea in the Southeast Coastal Region.

The visceral white nodules disease in the internal organs of Larimichthys crocea has caused significant harm in the aquaculture of this species, with Pseudomonas plecoglossicida considered one of the core pathogens causing this disease. In this study, we designed three pairs of specific nested PCR primers targeting the sctU gene of P. plecoglossicida, a crucial component of the Type III secretion system (T3SS), which is instrumental in bacterial pathogenesis and virulence. Through the optimization of PCR reaction conditions, specificity testing, and sensitivity determination, a method was established for the accurate detection of P. plecoglossicida. This method yielded single amplification products, exhibited a false positive rate of zero for reference bacteria, and achieved a detection sensitivity of a minimum of 2.62 copies/reaction for the target sequence. Using the detection method, we conducted analyses on the diseased populations of L. crocea, involving a total of 64 screened fishes along the southeast coast of China from 2021 to 2023. The results revealed that the infection rate of P. plecoglossicida in diseased L. crocea exceeded over 90% in March and April, while in other months, the maximum recorded infection rate was merely 10%. The detection method developed in this study shows potential for early warning and routine monitoring of visceral white nodules disease in the internal organs of species such as L. crocea.

The Complete Mitochondrial Genome of Pilumnopeus Makianus (Brachyura: Pilumnidae), Novel Gene Rearrangements, and Phylogenetic Relationships of Brachyura.

Pilumnopeus makianus is a crab that belongs to Pilumnidae, Brachyura. Although many recent studies have focused on the phylogeny of Brachyura, the internal relationships in this clade are far from settled. In this study, the complete mitogenome of P. makianus was sequenced and annotated for the first time. The length of the mitogenome is 15,863 bp, and includes 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNA), and 2 ribosomal RNA genes (rRNA). The mitogenome exhibits a high AT content (72.26%), with a negative AT-skew (-0.01) and a GC-skew (-0.256). In the mitogenome of P. makianus, all the tRNA genes are folded into the typical cloverleaf secondary structure, except trnS1 (TCT). A comparison with the ancestors of Brachyura reveals that gene rearrangement occurred in P. makianus. In addition, phylogenetic analyses based on thirteen PCGs indicated that P. makianus, Pilumnus vespertilio, and Echinoecus nipponicus clustered into a well-supported clade that supports the monophyly of the family Pilumnidae. These findings enabled a better understanding of phylogenetic relationships within Brachyura.