Reemergence of yellow fever virus in southeastern Brazil, 2017–2018: What sparked the spread?

BackgroundThe 2017–2018 yellow fever virus (YFV) outbreak in southeastern Brazil marked a reemergence of YFV in urban states that had been YFV-free for nearly a century. Unlike earlier urban YFV transmission, this epidemic was driven by forest mosquitoes. The objective of this study was to evaluate environmental drivers of this outbreak.Methodology/Principal findingsUsing surveillance data from the Brazilian Ministry of Health on human and non-human primate (NHP) cases of YFV, we traced the spatiotemporal progression of the outbreak. We then assessed the epidemic timing in relation to drought using a monthly Standardized Precipitation Evapotranspiration Index (SPEI) and evaluated demographic risk factors for rural or outdoor exposure amongst YFV cases. Finally, we developed a mechanistic framework to map the relationship between drought and YFV. Both human and NHP cases were first identified in a hot, dry, rural area in northern Minas Gerais before spreading southeast into the more cool, wet urban states. Outbreaks coincided with drought in all four southeastern states of Brazil and an extreme drought in Minas Gerais. Confirmed YFV cases had an increased odds of being male (OR 2.6; 95% CI 2.2–3.0), working age (OR: 1.8; 95% CI: 1.5–2.1), and reporting any recent travel (OR: 2.8; 95% CI: 2.3–3.3). Based on this data as well as mosquito and non-human primate biology, we created the “Mono-DrY” mechanistic framework showing how an unusual drought in this region could have amplified YFV transmission at the rural-urban interface and sparked the spread of this epidemic.Conclusions/SignificanceThe 2017–2018 YFV epidemic in Brazil originated in hot, dry rural areas of Minas Gerais before expanding south into urban centers. An unusually severe drought in this region may have created environmental pressures that sparked the reemergence of YFV in Brazil’s southeastern cities.     

GPX5 orthologs of the mouse epididymis-restricted and sperm-bound selenium-independent glutathione peroxidase are not expressed with the same quantitative and spatial characteristics in large domestic animals

We report here on the cloning of cDNAs coding bovine and equine orthologs of mouse epididymis-restricted and sperm-bound glutathione peroxidase 5 (GPX5), a selenium-independent member of the multigenic GPX family in mammals. The complete sequence of bovine GPX5 as well as a partial sequence of the equine GPX5 were characterized, conceptually translated and aligned with other known mammalian GPX5 proteins. Using Northern blotting assays, we show that the level of expression of GPX5 is high in bovine but low in equine and that in both species the regionalization of GPX5 expression in epididymis is not totally identical to what was reported for rodent mouse GPX5. An antibody was produced against GPX5 and used in Western blot assays as well as in immunohistochemistry assays on bovine epididymis sections. It shows that the protein is essentially present in the cytoplasmic compartment of the caput segment 2 epithelium of the bovine epididymis. Unlike in the mouse model, bovine GPX5 seems to be poorly secreted and does not seem to be present on cauda epididymal spermatozoa.     

Expression of a truncated form of inositol 1,4,5-trisphosphate receptor type III in the cytosol of DT40 triple inositol 1,4,5-trisphosphate receptor-knockout cells

In non-excitable cells, the inositol 1,4,5-trisphosphate receptor (IP3R) is an intracellular Ca2+ channel playing a major role in Ca2+ signaling. Three isoforms of IP3R have been identified and most cell types express different proportions of each isoform. The DT40 B lymphocyte cell line lacking all three IP3R isoforms (DT40IP3R-KO cells) represents an excellent model to re-express any recombinant IP3R and analyze its specific properties. In the study presented here, we confirmed that DT40IP3R-KO cells do not express any IP3-sensitive Ca2+ release channel. However, with an immunoblot approach and a [3H]IP3 binding approach we demonstrated the presence of a C-terminally truncated form of IP3R type III in the cytosolic fraction of DT40IP3R-KO cells. We further showed that this truncated IP3R retained the ability to couple to the Ca2+ entry channel TRPC6. Therefore, a word of caution is offered about the interpretation of results obtained in using DT40IP3R-KO cells to study the cellular mechanisms of Ca2+ entry.     

Coordinate regulation of the mother centriole component nlp by nek2 and plk1 protein kinases

Mitotic entry requires a major reorganization of the microtubule cytoskeleton. Nlp, a centrosomal protein that binds gamma-tubulin, is a G(2)/M target of the Plk1 protein kinase. Here, we show that human Nlp and its Xenopus homologue, X-Nlp, are also phosphorylated by the cell cycle-regulated Nek2 kinase. X-Nlp is a 213-kDa mother centriole-specific protein, implicating it in microtubule anchoring. Although constant in abundance throughout the cell cycle, it is displaced from centrosomes upon mitotic entry. Overexpression of active Nek2 or Plk1 causes premature displacement of Nlp from interphase centrosomes. Active Nek2 is also capable of phosphorylating and displacing a mutant form of Nlp that lacks Plk1 phosphorylation sites. Importantly, kinase-inactive Nek2 interferes with Plk1-induced displacement of Nlp from interphase centrosomes and displacement of endogenous Nlp from mitotic spindle poles, while active Nek2 stimulates Plk1 phosphorylation of Nlp in vitro. Unlike Plk1, Nek2 does not prevent association of Nlp with gamma-tubulin. Together, these results provide the first example of a protein involved in microtubule organization that is coordinately regulated at the G(2)/M transition by two centrosomal kinases. We also propose that phosphorylation by Nek2 may prime Nlp for phosphorylation by Plk1.     

Semi-rational approaches to engineering enzyme activity: combining the benefits of directed evolution and rational design

Many research groups successfully rely on whole-gene random mutagenesis and recombination approaches for the directed evolution of enzymes. Recent advances in enzyme engineering have used a combination of these random methods of directed evolution with elements of rational enzyme modification to successfully by-pass certain limitations of both directed evolution and rational design. Semi-rational approaches that target multiple, specific residues to mutate on the basis of prior structural or functional knowledge create 'smart' libraries that are more likely to yield positive results. Efficient sampling of mutations likely to affect enzyme function has been conducted both experimentally and, on a much greater scale, computationally, with remarkable improvements in substrate selectivity and specificity and in the de novo design of enzyme activities within scaffolds of known structure.     

Murine glycosyltransferases responsible for the expression of globo-series glycolipids: cDNA structures, mRNA expression, and distribution of their products

Biological functions of globo-series glycosphingolipids are not well understood. In this study, murine cDNAs of two glycosyltransferases responsible for the synthesis of globo-series glycolipids and mRNA expression of those genes were analyzed. Distribution of their products was also analyzed. Murine cDNAs for Gb3/CD77 synthase and Gb4 synthase predicted that both of them are type II membrane proteins with 348 and 331 amino acids, respectively. In northern blotting, Gb3/CD77 synthase gene was mainly expressed in kidney and lung but also detected in many other tissues. Gb4 synthase was expressed in brain, heart, kidney, liver, skin, and testis. In the immunohistological analysis, Gb3/CD77 was mainly expressed in the proximal tubules as revealed with coincidental expression with angiotensin-converting enzyme (ACE). In spleen, it was detected in pre-B cells in the peripheral region of the white pulp, as suggested with coincidental expression with CD10. It was also expressed on the endothelia of the alveolar capillaries in lung and on the sebaceous ducts aside of the hair follicles. Gb4 was also detected mainly on the proximal tubules in kidney and on the endothelia of the alveolar capillaries in lung as Gb3/CD77. But it was also detected on the epithelium of the bronchus, seminiferous tubules and tails of spermatozoa in testis, blood vessels of choroids plexus and endothelial cells in brain, and central and hepatoportal veins in liver. The expression patterns of two genes and their products almost corresponded with some exception. The results would provide essential information for the functional studies of globo-series glycolipids.     

Protease-dependent uncoating of a complex retrovirus

Although retrovirus egress and budding have been partly unraveled, little is known about early stages of the replication cycle. In particular, retroviral uncoating, a process during which incoming retroviral cores are altered to allow the integration of the viral genome into host chromosomes, is poorly understood. To get insights into these early events of the retroviral cycle, we have used foamy complex retroviruses as a model. In this report, we show that a protease-defective foamy retrovirus is noninfectious, although it is still able to bud and enter target cells efficiently. Similarly, a retrovirus mutated in an essential viral protease-dependent cleavage site in the central part of Gag is noninfectious. Following entry, wild-type and mutant retroviruses are able to traffic along microtubules towards the microtubule-organizing center (MTOC). However, whereas nuclear import of Gag and of the viral genome was observed for the wild-type virus as early as 8 hours postinfection, incoming capsids and genome from mutant viruses remained at the MTOC. Interestingly, a specific viral protease-dependent Gag cleavage product was detected only for the wild-type retrovirus early after infection, demonstrating that cleavage of Gag by the viral protease at this stage of the virus life cycle is absolutely required for productive infection, an unprecedented observation among retroviruses.     

Ducts of the rat pancreas in agarose matrix culture

Summary Interlobular and intralobular ducts isolated from the pancreas of the rat by digestion with collagenase and chymotrypsin were cultured in an agarose matrix containing CMRL-1066 supplemented with insulin, dexamethasone,l-glutamine, soybean trypsin inhibitor, antibiotics, and fetal bovine serum. The cut ends of most interlobular ducts sealed to create encolosed lumina. Some ducts retained their original cylindrical organization; others enlarged to varying degrees, resulting in structures that ranged from cylindrical to spherical in shape. The duct walls consisted of viable epithelium and connective tissue, although the amount of connective tissue declined with age. Both epithelial and connective tissue cells became flattened in the enlarged ducts. Intralobular and small interlobular ducts often remained associated with the larger interlobular ducts. These duct fragments have been cultured for as long as 6 weeks. This study was supported by National Cancer Institute Grant CA 19177 through the National Pancreatic Cancer Project and by Biomedical Research Support Grant RR 07196 from the Division of Research Resources, National Institutes of Health.     

Environmental conditions terminate reproductive diapause and influence pheromone perception in the long‐lived moth Caloptilia fraxinella

Reproductive diapause enables long‐lived insects to time mating with environmental conditions suitable for offspring development. Plasticity in the perception of pheromones used in sexual communication may enable mate‐finding at the appropriate time of year. The moth Caloptilia fraxinella (Ely) (Lepidoptera: Gracillariidae) undergoes a 9‐month reproductive diapause, during which the male response to pheromone is plastic and is highest during the period of reproductive activity. The mechanisms controlling this pheromone response plasticity are not well‐understood, and the aim of the present study is to determine the main factors involved. In the present study, the impact of temperature, photoperiod, juvenile hormone analogue (JHA) and adult nutrition on diapause termination are tested using electroantennogram (EAG) and behavioural response to pheromone in male C. fraxinella. Eclosion in a state of reproductive diapause occurs in most males; diapause is maintained under short‐day or cool conditions indoors, or under natural conditions outdoors. Exposure to long‐day, warm conditions over a period of 4 weeks causes a small number of males to become behaviourally responsive to pheromone; a larger number of males become behaviourally responsive over a period of 3 months of post‐eclosion. Treatment with a JHA impacts male EAG and the behavioural response to pheromone during the period of reproductive diapause. A carbohydrate food source is not required by reproductively active adult male C. fraxinella to respond to pheromone and express mate location behaviours. The main factors involved in controlling male pheromone response plasticity and the implications of these factors for the C. fraxinella population in its expanded range are discussed.