Publications

4367

‘ Candidatus Liberibacter asiaticus’, the putative causal agent of citrus greening disease, is transmitted by the Asian citrus psyllid, Diaphorina citri, in a propagative, circulative, and persistent manner. Unfortunately, ‘ Ca. L. asiaticus’ is not yet available in pure culture to carry out Koch's postulates and to confirm its etiology. When a pure culture is available, an assay to test its infectivity in both the insect vector and the plant host will be crucial. Herein, we described a transmission assay based on the use of phloem sap extracted from infected citrus plants and topical feeding to D. citri nymphs. Phloem sap was collected by centrifugation, diluted with 0.1 M phosphate buffer pH 7.4 containing 20% (wt/vol) sucrose and 0.1% ascorbic acid (wt/vol) as an antioxidant, and delivered to third through fifth instar nymphs by placing droplets on the mouthparts. Nymphs unfolded the stylets and acquired the phloem sap containing the bacterial pathogen. Nymphs were then placed onto Citrus macrophylla seedlings (10 nymphs per seedling) for an inoculation period of 2 weeks. A transmission rate of up to 80% was recorded at 6 months postinoculation. The method could be a powerful tool to test the transmissibility of the bacterial pathogen after various treatments to reduce the viability of the bacteria or to block its transmission. In addition, it might be a potent assay to achieve Koch's postulates if a pure culture of ‘ Ca. L. asiaticus’ becomes available.

AbstractSpotted fever group (SFG) rickettsia, the causative agent of SFG rickettsiosis, is predominantly carried by ticks, whereas Orientia tsutusgamushi, the causative agent of scrub typhus, is primarily transmitted by chigger mites in Japan. In this study, we attempted to isolate intracellular eubacteria from Leptotrombidium scutellare, a major vector of O. tsutsugamushi; moreover, we isolated an SFG rickettsia using a mosquito‐derived cell line. Draft genome sequences of this unique isolate, by applying criteria for species delimitation, classified this isolate as a novel strain, proposed as “Rickettsia kedanie.” Further genetic analysis identified conserved virulence factors, and the isolate successfully propagated in mammalian cells, suggesting its ability to cause diseases in humans. The presence of SFG rickettsia in unfed larvae implies potential dual‐pathogen carriage and reflects a symbiotic relationship similar to that between the mites and O. tsutsugamushi, indicating possibility of its transovarial transmission from female adults. Furthermore, conserved genomic similarity of the novel isolate to known SFG rickettsia suggests potential multiple hosts, including chiggers and ticks. In the natural environment, ticks, chigger mites, and wild animals may carry new isolates, complicating the infection cycle and increasing the transmission risks to humans. This discovery challenges the conventional association of SFG rickettsia with ticks, emphasizing its implications for research and disease control. However, this study was confined to a particular species of chigger mites and geographic area, underscoring the necessity for additional studies to comprehend the ecological dynamics, host interactions, and health implications linked to this newly identified SFG rickettsia.

The tomato-potato psyllid, Bactericera cockerelli (Šulc), belonging to the Hemiptera order, is an insect pest of solanaceous crops and vectors a fastidious bacterium, Candidatus Liberibacter solanacearum (CLso), the presumptive causal agent of zebra chip and vein greening diseases in potatoes and tomatoes, respectively. The genome of B. cockerelli has been sequenced recently, providing new avenues to elucidate mechanistic insights into pathogenesis in vegetable crops. In this study, we performed RNA-sequencing of the critical psyllid organs (salivary glands and ovaries) involved in CLso pathology and transmission to host plants. Transcriptome analysis revealed differentially expressed genes and organ-specific enrichment of gene ontology (GO) terms related to metabolic processes, response to stress/stimulus, phagocytosis, proteolysis, endocytosis, and provided candidate genes encoding transcription factors (TFs). To examine gene regulatory networks across the psyllid organs under CLso(-) and CLso(+) conditions, we performed weighted gene co-expression network analysis (WGCNA), and unique modules differentiating the psyllid organs were identified. A comparative GO analysis of the unique gene modules revealed functional terms enriched in response to stress, gene regulation, and cell division processes in the ovaries. In contrast, respiration, transport, and neuronal transmission-related GO terms were enriched in the salivary glands. Altogether, this study reveals new insights into tissue-specific expression of the psyllid organs in the absence or presence of CLso bacterium. This knowledge can be leveraged to develop new pest and disease management strategies by delineating the regulatory networks involved in the psyllid-CLso interaction.

AbstractAnemia is a common problem in South American camelids (SACs). Infections with Candidatus Mycoplasma haemolamae (CMh), a cell-wall free, hemotropic bacterium, are often suspected to be an important cause of anemia, as the pathogen infects the erythrocytes and is found in the blood of up to 30% of SACs. The information on the clinical signs of animals infected with this pathogen vary widely. Most infections are clinically inapparent. Treatment is usually carried out with oxytetracycline. A detailed overview of the clinical and hematological findings in 13 alpacas infected with Candidatus M. haemolamae (CMh+), based on patients from our university clinic and comparing those findings with the results of 22 negative alpacas (CMh−) is provided. Assignment to both groups was based on the PCR result. No relevant clinical or hematological differences between CMh+ and CMh− were found, the clinical signs in CMh+ were usually due to comorbidities. The examination of a blood smear alone proved to be insufficient; a PCR test should be carried out to confirm or rule out an infection. A critical review of the need for antibiotic treatment on the basis of a positive test result alone is recommended.

Four Gram-stain-negative, aerobic, rod-shaped and motile strains (LYT19WT, DXS22WT, DXS29WT and BYS139WT) were isolated from streams in China. All four strains showed highest 16S rRNA gene sequence similarities to the species of genus Ideonella. The calculated average nucleotide identity, digital DNA–DNA hybridization and average amino acid identity values among strains LYT19WT, DXS22WT, DXS29WT, BYS139WT and other closely related strains were less than 79.5, 22.5 and 74.0%, respectively, indicating that each of the four strains should represent an independent novel species. The further reconstructed phylogenomic tree showed that strains LYT19WT and DXS29WT clustered closely with Ideonella strains, but strains DXS22WT and BYS139WT formed an independent clade with Aquabacterium terrae and Aquabacterium pictum. Comparing with other Aquabacterium species, A. terrae and A. pictum harboured distinct physiological and genetic characteristics, so it was reasonable to propose a novel genus Pseudaquabacterium to accommodate A. terrae, A. pictum, DXS22WT and BYS139WT. The major fatty acids of the four strains contained C16 : 0, summed feature 3 (C16 : 1  ω6c and/or C16 : 1  ω7c) and summed feature 8 (C18 : 1  ω6c and/or C18 : 1  ω7c). The predominant polar lipids were phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol and one unidentified aminophospholipid. Combining above descriptions, strains LYT19WT and DXS29WT should represent two novel species of the genus Ideonella, for which the names Ideonella margarita sp. nov. (type strain LYT19WT=GDMCC 1.3205T=KCTC 92545T) and Ideonella lacteola sp. nov. (type strain DXS29WT=GDMCC 1.3207T=KCTC 92547T) are proposed, respectively. Strains DXS22WT and BYS139WT should represent two novel species of the new genus Pseudaquabacterium, for which the names Pseudaquabacterium inlustre sp. nov. (type strain DXS22WT=GDMCC 1.3206T=KCTC 92546T) and Pseudaquabacterium rugosum sp. nov. (type strain BYS139WT=GDMCC 1.3208T=KCTC 92548T) are proposed, respectively.