Publications

4852

A 59-year-old female living in Rayong Province, eastern Thailand, presented with painless, right upper eyelid nodule for 3 months. Upon removal of the eyelid mass, a well-circumscribed, firm globular mass with diameter about 1 cm was found. Histopathological examination revealed an immature female dirofilarial worm reminiscent of Dirofilaria repens, characterized by prominent sharp longitudinal ridges at external surface of the cuticle. Analysis of the mitochondrial 12S rRNA sequence showed that the worm belongs to Candidatus Dirofilaria hongkongensis. It is likely that some infections previously reported as D. repens based on histological examination may have actually been due to Candidatus D. hongkongensis.

Citrus huanglongbing (HLB) disease associated with the ‘Candidatus Liberibacter asiaticus’ (CLas) bacterium has caused significant financial damage to many citrus industries. Large-scale pathogen surveys are routinely conducted in California to detect CLas early in the disease cycle by lab-based qPCR assays. We have developed an improved reference gene for the sensitive detection of CLas from plants in diagnostic duplex qPCR and analytical digital droplet PCR (ddPCR) assays. The mitochondrial cytochrome oxidase gene (COX), widely used as a reference, is not ideal because its high copy number can inhibit amplification of small quantities of target genes. In ddPCRs, oversaturation of droplets complicates data normalization and quantification. The variable copy numbers of COX gene in metabolically active young tissue, greenhouse plants, and citrus relatives suggest the need for a non-variable, nuclear, low copy, universal reference gene for analysis of HLB hosts. The single-copy nuclear gene, malate dehydrogenase (MDH), developed here as a reference gene, is amenable to data normalization, suitable for duplex qPCR and ddPCR assays. The sequence of MDH fragment selected is conserved in most HLB hosts in the taxonomic group Aurantioideae. This study emphasizes the need to develop standard guidelines for reference genes in DNA-based PCR assays.

Abstract Background Hemotropic mycoplasmas, previously classified in the genus Eperythrozoon, have been reported as causing human infections in Brazil, China, Japan, and Spain. Methods In 2017, we detected DNA from Candidatus Mycoplasma haemohominis in the blood of a Melanesian patient from New Caledonia presenting with febrile splenomegaly, weight loss, life-threatening autoimmune hemolytic anemia, and hemophagocytosis. The full genome of the bacterium was sequenced from a blood isolate. Subsequently, we retrospectively (2011–2017) and prospectively (2018–2019) tested patients who had been hospitalized with a similar clinico-biological picture. In addition, as these patients had been in contact with frugivorous bats (authorized under conditions for hunting and eating in New Caledonia), we investigated the role of these animals and their biting flies by testing them for hemotropic mycoplasmas. Results There were 15 patients found to be infected by this hemotropic mycoplasma. Among them, 4 (27%) died following splenectomy performed either for spontaneous spleen rupture or to cure refractory autoimmune hemolytic anemia. The bacterium was cultivated from the patient’s blood. The full genome of the Neocaledonian Candidatus M. haemohominis strain differed from that of a recently identified Japanese strain. Of 40 tested Pteropus bats, 40% were positive; 100% of collected bat flies Cyclopodia horsfieldi (Nycteribiidae, Diptera) were positive. Human, bat, and dipteran strains were highly similar. Conclusions The bacterium being widely distributed in bats, Candidatus M. haemohominis, should be regarded as a potential cause of severe infections in humans.

Abstract Background Candidatus (Ca.) Neoehrlichia (N.) mikurensis is an emerging tick-borne pathogen of humans that is closely related to Ehrlichia and Anaplasma species. This strict intracellular bacterium escapes detection by routine microbiologic diagnostic methods such as blood culture, leading to considerable under-diagnosis of the infectious disease it causes, neoehrlichiosis. Methods Here, we describe the vascular and thromboembolic events afflicting a series of 40 patients diagnosed with neoehrlichiosis in Sweden during a 10-year period (2009–2019). Results The majority of the patients (60%) developed vascular events ranging from repeated thrombophlebitis, deep vein thrombosis, pulmonary embolism, transitory ischemic attacks, to arteritis. Younger age was a risk factor for vascular complications. In contrast, there was no difference in the incidence of vascular events between immunosuppressed and immunocompetent patients. However, there were qualitative differences, such that deep vein thrombosis exclusively afflicted the immunosuppressed patients, whereas arteritis was restricted to the immunocompetent persons. We also present the case histories of two patients who developed vasculitis mimicking polyarteritis nodosa and giant cell arteritis. Both were cured by doxycycline treatment. Conclusions Ca. N. mikurensis infection should be considered in patients living in tick-endemic areas of Europe and northern Asia who present with atypical vascular and/or thromboembolic events. Early diagnosis and antibiotics targeting this emerging infectious agent can eradicate the infection and prevent the development of new vascular events.

Candidatus Liberibacter solanacearum (CaLso) is associated with diseases in tomato crops and transmitted by the tomato psyllid Bactericera cockerelli. A polymeric water-dispersible nanobactericide (PNB) was evaluated against CaLso as a different alternative. PNB is a well-defined polycationic diblock copolymer designed to permeate into the vascular system of plants. Its assessment under greenhouse conditions was carried out with tomato plants previously infected with CaLso. Using a concentration as low as 1.0 mg L−1, a small but significant reduction in the bacterial load was observed by real-time qPCR. Thus, to achieve an ecologically friendly dosage and set an optimum treatment protocol, we performed experiments to determine the effective concentration of PNB to reduce ~65% of the initial bacterial load. In a first bioassay, a 40- or 70-fold increase was used to reach that objective. At this concentration level, other bioassays were explored to determine the effect as a function of time. Surprisingly, a real reduction in the symptoms was observed after three weeks, and there was a significant decrease in the bacterial load level (~98%) compared to the untreated control plants. During this period, flowering and formation of tomato fruits were observed in plants treated with PNB.