Development and Validation of a Protein Array for Detection of Antibodies against the Tick-Borne Pathogen Borrelia miyamotoi.

Current serological tests for the emerging tick-borne pathogen Borrelia miyamotoi lack diagnostic accuracy. To improve serodiagnosis, we investigated a protein array simultaneously screening for IgM and IgG reactivity against multiple recombinant B. miyamotoi antigens. The array included six B. miyamotoi antigens: glycerophosphodiester phosphodiesterase (GlpQ), multiple variable major proteins (Vmps), and flagellin. Sera included samples from cases of PCR-proven Borrelia miyamotoi disease (BMD), multiple potentially cross-reactive control groups (including patients with culture-proven Lyme borreliosis, confirmed Epstein-Barr virus, cytomegalovirus, or other spirochetal infections), and several healthy control groups from regions where Ixodes is endemic and regions where it is nonendemic. Based on receiver operating characteristic (ROC) analyses, the cutoff for reactivity per antigen was set at 5 µg/mL for IgM and IgG. The individual antigens demonstrated high sensitivity but relatively low specificity for both IgM and IgG. The best-performing single antigen (GlpQ) showed a sensitivity of 88.0% (95% confidence interval [CI], 78.9 to 93.5) and a specificity of 94.2% (95% CI, 92.7 to 95.6) for IgM/IgG. Applying the previous published diagnostic algorithm-defining seroreactivity as reactivity against GlpQ and any Vmp-revealed a significantly higher specificity of 98.5% (95% CI, 97.6 to 99.2) but a significantly lower sensitivity of 79.5% (95% CI, 69.3 to 87.0) for IgM/IgG compared to GlpQ alone. Therefore, we propose to define seroreactivity as reactivity against GlpQ and any Vmp or flagellin which resulted in a comparable sensitivity of 84.3% (95% CI, 74.7 to 90.8) and a significantly higher specificity of 97.9% (95% CI, 96.9 to 98.7) for IgM/IgG compared to GlpQ alone. In conclusion, we have developed and validated a novel serological tool to diagnose BMD that could be implemented in clinical practice and epidemiological studies. IMPORTANCE This paper describes the protein array as a novel serological test for the diagnosis of Borrelia miyamotoi disease (BMD), by reporting the methodology, the development of a diagnostic algorithm, and its extensive validation. With rising numbers of ticks and tick bites, tick-borne diseases, such as BMD, urgently deserve further societal and medical attention. B. miyamotoi is prevalent in Ixodes ticks across the northern hemisphere. Humans are exposed to, and infected by, B. miyamotoi and develop BMD in Asia, in North America, and to a lesser extent in Europe. However, the burden of infection and disease remains largely unknown, due to the noncharacteristic clinical presentation, together with the lack of awareness and availability of diagnostic tools. With this paper, we offer a novel diagnostic tool which will assist in assessing the burden of disease and could be implemented in clinical care.

Ontogenetic consequences of dysgenic crosses in Drosophila virilis.

Hybrid dysgenesis (HD) syndrome in Drosophila virilis presumably results from the mobilization of several unrelated mobile genetic elements in dysgenic hybrids. Morphogenetic events during oogenesis and spermatogenesis were investigated in detail in the progeny of D. virilis dysgenic crosses. Using germ-cell specific anti-Vasa staining, we monitored the fate of germline cells at different ontogenetic stages in strains of D. virilis and their hybrids. Anti-Vasa staining indicated that the major loss of pole cells occurs in dysgenic embryos at stage 11-14 after primordial germ cells (PGC) pass the midgut wall. At later ontogenetic stages, including larvae, pupae and imagoes, we often observed an abnormal development of gonads in dysgenic individuals with a frequent occurrence of unilateral and bilateral gonadal atrophy. Dysgenic females were characterized by the presence of various sterile ovarian phenotypes that predominantly include agametic ovarioles, while other atypical forms such as tumor-like ovarioles and dorsalized ovariolar follicles may also be present. Testis abnormalities were also frequently observed in dysgenic males. The sterility manifestations depended on the strain, the growing temperature and the age of the flies used in crosses. The observed gonadal sterility and other HD manifestations correlated with the absence of maternal piRNAs homologous to Penelope and other transposons in the early dysgenic embryos. We speculate that gonadal abnormalities mimicking several known sterility mutations probably result from the disturbance of developmental gene expression machinery due to the activation of unrelated families of transposons in early dysgenic embryos.

Bacteria in ovarioles of females from maleless families of ladybird beetles Adalia bipunctata L. (Coleoptera: Coccinellidae) naturally infected with Rickettsia, Wolbachia, and Spiroplasma.

Ovarioles were found to be infected with Spiroplasma, Wolbachia, and Rickettsia in Adalia bipunctata females with maleless progeny in different natural populations. Ooplasm was infected with few Wolbachia bacteria. In ooplasm infected by Rickettsia, bacteria were present in small foci. Spiroplasmas were found encapsulated into ooplasm from the wider intercellular spaces between epithelial and oocyte cells. The cytoplasm of follicular epithelia infected with Rickettsia was heavily destroyed, but the nucleus was intact and free from bacteria. The essential feature of follicular epithelium cells from Spiroplasma and Wolbachia infected A. bipunctata females was inclusions of three types: crystalline, filaments, and concentric myelin-like lamellae. Observations of smears prepared from ovaries of A. bipunctata from natural populations revealed a low concentration of bacteria within a microscopy field (less 10 bacteria) in more than 90% of specimens, and only a few ovaries were heavily infected. Two different ways of bacterial invasion of the oocyte are suggested: Spiroplasma-like, through the intercellular spaces in the epithelium and Rickettsia-like, through the cytoplasm of follicular epithelium cells. Bacteria were not found in germarium zones and we suggest that each follicle is infected from haemolymph.