Taxonomy and comparative genomics of bacterial species associated with onion bulb rot focusing on Rahnella species

Abstract

Onions (Allium cepa L.) are one of the most widely consumed vegetables globally, thus playing a significant role in food security. However, onion bulbs are highly susceptible to bacterial pathogens, and emerging opportunistic pathogens such as Rahnella species, which have become notable threats to onion production. Identifying these pathogens is essential for managing disease outbreaks. In Chapter 1, we conducted a literature review focusing on the cultivation of onion bulbs and then explored onion production in South Africa and the United States. In addition, a number of onion bacterial pathogens were discussed that included Burkholderia, Rahnella, Erwinia, Serratia, Enterobacter species, the Pantoea complex, and Klebsiella. The management strategies currently implemented to help combat onion diseases, both pre- and post-harvest, were also reviewed. To understand the underlying issues related to bacterial diseases in onions, we analysed the pathogenicity factors commonly utilised by these pathogens. This analysis enhanced our understanding of the bacterial taxonomy of onion pathogens, pathogenicity mechanisms, and genomics, which is crucial for identification of pathogenicity traits used by pathogenic bacteria. In Chapter 2, bacterial pathogens responsible for onion bulb rot in South Africa during the 2021–2022 period were characterised and identified. Diseased onion samples were collected from fresh produce markets and storage facilities in the Western Cape, Free State, and Gauteng provinces. Symptoms observed included softened necks, scale dryness, internal discolouration, slippery, and sour skin. Pathogen identification revealed the presence of Burkholderia gladioli, Klebsiella grimontii, Klebsiella michiganensis, Klebsiella pneumoniae, Pantoea eucalypti, and a Pectobacterium? sp. Koch’s postulates confirmed that all these bacteria were causal agents of onion bulb rot. This study emphasised the importance of identifying bacterial pathogens to reduce pre- and post-harvest losses and sustain onion production in South Africa. In Chapter 3, Rahnella species isolated from onion bulb rot in South Africa (SA) and the United States (USA) were identified using multilocus sequence analysis (MLSA) based on four housekeeping genes (atpD, gyrB, infB, and rpoB. The isolates clustered into five species, viz. R. perminowiae, R. aquatilis, R. aceris, R. variigena, and R. victoriana. Among them, R. perminowiae, R. aquatilis, and R. aceris were the the most frequently isolated. Pathogenicity testing revealed that Rahnella isolates caused symptoms on onion bulbs but did not affect onion leaves and were negative in the red scale necrosis (RSN) assays. Geographical location did not appear to influence the diversity or pathogenicity of these isolates. This analysis highlights the growing importance of Rahnella species as emerging pathogens in agriculture and there is a need to understand their opportunistic tendencies in onion bulbs. In Chapter 4, comparative genomic analyses were used to investigate pathogenicity-related traits in five Rahnella strains isolated from onion bulbs: R. aquatilis (strains 20CA0197 and 20CA0198), R. aceris (samples 20WA0051 and 20WA0057), and R. perminowiae FS4. The pathogenicity-related genes were categorized into metabolic functions, secretion systems, motility, adherence, and lipopolysaccharide biosynthesis, in an attempt to highlight their potential roles in plant interactions. The presence of the thiosulfinate gene clusters in R. aquatilis (strains 20CA0197 and 20CA0198) and R. perminowiae FS4 suggests an adaptation to thiosulfinate-rich environments, in contrast, R. aceris (strains 20WA0051 and 20WA0057) did not possess thiosulfinate gene clusters. The copper resistance genes were absent in all five Rahnella strains. Additionally, biosynthetic gene clusters related to siderophores, quorum sensing, and antimicrobial compounds were identified. Further functional validation is necessary to determine the precise roles of these genes in plant-bacteria interactions and to assess whether they contribute to Rahnella's opportunistic behaviour in onion bulbs. This study reports on the growing threat of opportunistic bacterial pathogens to onion production and highlights the need for comprehensive disease monitoring strategies for opportunistic bacteria. The identification of diverse bacterial species, including Rahnella, Burkholderia, Pantoea, Klebsiella, and Ewingella allii, provides new insights into the complexity of onion bulb rot and emerging opportunistic pathogens. Comparative genomic analyses revealed critical pathogenicity-related traits, indicating potential adaptation mechanisms to onion-associated environments. While Rahnella species exhibited mild pathogenicity, their increasing prevalence necessitates further research to assess their long-term impact on onion crops. Future efforts should focus on pathogen surveillance, functional validation of virulence factors, and the development of sustainable control measures to safeguard onion production and food security.