Molecular Pathogens 2024, Vol.15, No.2, 72-82 http://microbescipublisher.com/index.php/mp 73 2 Isolation of Kelp Pathogens 2.1 Sampling methods Sampling methods for isolating kelp pathogens involve collecting samples from various parts of the kelp and its surrounding environment. For instance, bacterial strains have been isolated from coastal sediment samples and fresh kelp samples collected from kelp culture areas (Ye et al., 2022). Additionally, biofilm samples from brown macro-algae, such as Macrocystis pyrifera, have been collected for metagenomic analysis (Vollmers et al., 2017). Sampling also includes collecting kelp from specific locations, such as Li Island in Rongcheng, China, to isolate difficult-to-cultivate bacterial strains. These methods ensure a diverse collection of potential pathogens and associated microorganisms for further analysis. 2.2 Laboratory Isolation Techniques Laboratory isolation techniques for kelp pathogens include a variety of methods to culture and identify bacterial strains. For example, strains can be isolated using standard microbiological techniques, such as growing them on specific media under controlled conditions (Ye et al., 2022). Metagenomic approaches, including shotgun and amplicon sequencing, are also employed to analyze biofilm samples and identify novel microbial species (Vollmers et al., 2017). Additionally, innovative binning approaches are used to untangle genomes of novel species from metagenomic data (Vollmers et al., 2017). Techniques such as the use of Taxoblast for detecting bacterial contaminants in kelp genomes are also crucial for ensuring the accuracy of genomicdata (Dittami and Corre, 2017). 2.3 Challenges in Isolation Isolating kelp pathogens presents several challenges. One significant challenge is the presence of bacterial contaminants and hybrid sequences in kelp genomes, which can complicate the identification of true pathogens (Dittami and Corre, 2017). Another challenge is the difficulty in cultivating certain bacterial strains, such as those from the phylum Verrucomicrobiota, which require specific conditions for growth (Ye et al., 2022). Additionally, the complex interactions between kelp and its associated microbiome can make it difficult to isolate and identify specific pathogenic organisms (Weigel et al., 2022). The need for advanced genomic and bioinformatics tools to accurately identify and characterize these pathogens further adds to the complexity of the isolation process (Dittami and Corre, 2017; Vollmers et al., 2017; Weigel et al., 2022). By addressing these challenges through meticulous sampling, advanced laboratory techniques, and robust bioinformatics tools, researchers can improve the isolation and identification of kelp pathogens, contributing to a better understanding of their impact on kelp health and ecosystem dynamics. 3 Identification of Kelp Pathogens 3.1 Morphological identification Morphological identification of kelp pathogens involves the examination of physical characteristics of the pathogens under a microscope. This method is often the first step in identifying pathogens and can provide immediate information about the presence and type of infection. For instance, fluorescence in situ hybridization (FISH) can be used to identify microbial pathogens by binding fluorescence-labeled probes to ribosomes of infectious agents, allowing for the visualization and differentiation of pathogens based on their morphology (Frickmann et al., 2017). This technique is particularly useful for identifying key pathogens in mixed species samples and provides spatial resolution that is crucial for understanding the distribution and interaction of pathogens within the kelp tissue. 3.2 Biochemical tests Biochemical tests are essential for the further characterization and identification of kelp pathogens. These tests involve assessing the metabolic and enzymatic activities of the pathogens. For example, the API20E biochemical identification system can be used to enhance the discrimination of environmental bacteria isolated from kelp. This system evaluates various biochemical reactions, such as the production of specific enzymes or the utilization of
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