The family Bunyaviridae is the most diversified family of RNA viruses. We describe a novel prototypic bunyavirus, tentatively named Gouléako virus, isolated from various mosquito species trapped in Côte d'Ivoire. The S segment comprised 1,087 nucleotides (nt), the M segment 3,188 nt, and the L segment 6,358 nt, constituting the shortest bunyavirus genome known so far. The virus had shorter genome termini than phleboviruses and showed no evidence of encoded NSs and NSm proteins. An uncharacterized 105-amino-acid (aa) putative open reading frame (ORF) was detected in the S segment. Genetic equidistance to other bunyaviruses (74 to 88% aa identity) and absence of serological cross-reactivity with phleboviruses suggested a proposed novel Bunyaviridae genus.

Most viruses of the genus Flavivirus, e.g., YFV and dengue virus, are mosquito borne and transmitted to vertebrates during blood feeding of mosquitoes. Within the last decade, an increasing number of viruses with a host range exclusively restricted to insects in close relationship to the vertebrate-pathogenic flaviviruses were discovered in mosquitoes. To identify barriers that could block the arboviral vertebrate tropism, we set out to identify the steps at which the ISF replication cycle fails in vertebrates. Our studies revealed blocks at several levels, suggesting that flavivirus host range expansion from insects to vertebrates was a complex process that involved overcoming multiple barriers.

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Rapid diagnosis of tuberculosis (TB) and antibiotic resistances are imperative to initiate effective treatment and to stop transmission of the disease. A new generation of more sensitive, automated molecular TB diagnostic tests has been recently launched giving microbiologists more choice between several assays with the potential to detect resistance markers for rifampicin and isoniazid. In this study, we determined analytical sensitivities as 95% limits of detection (LoD 95 ) for Xpert MTB/Rif Ultra (XP-Ultra) and BD-MAX MDR-TB (BD-MAX) as two representatives of the new test generation, in comparison to the conventional Fluoro-Type MTB (FT-MTB). Test matrices used were physiological saline solution, human and a mucin-based artificial sputum (MUCAS) each spiked with Mycobacterium tuberculosis in declining culture-and qPCR-controlled concentrations. With BD-MAX, XP-Ultra, and FT-MTB, we measured LoD 95 TB values of 2.1 cfu/ml (CI 95% : 0.9-23.3), 3.1 cfu/ml (CI 95% : 1.2-88.9), and 52.1 cfu/ml (CI 95% : 16.7-664.4) in human sputum; of 6.3 cfu/ml (CI 95% : 2.9-31.8), 1.5 cfu/ml (CI 95% : 0.7-5.0), and 30.4 cfu/ml (CI 95% : 17.4-60.7) in MUCAS; and of 2.3 cfu/ml (CI 95% : 1.1-12.0), 11.5 cfu/ml (CI 95% : 5.6-47.3), and 129.1 cfu/ml (CI 95% : 82.8-273.8) in saline solution, respectively. LoD 95 of resistance markers were 9 to 48 times higher compared to LoD 95 TB . BD-MAX and XP-Ultra have an equal and significantly increased analytical sensitivity compared to conventional tests. MUCAS resembled human sputum, while both yielded significantly different results than normal saline. MUCAS proved to be suitable for quality control of PCR assays for TB diagnostics.

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