Deep Mutational Scanning Comprehensively Maps How Zika Envelope Protein Mutations Affect Viral Growth and Antibody Escape.

Functional constraints on viral proteins are often assessed by examining sequence conservation among natural strains, but this approach is relatively ineffective for Zika virus because all known sequences are highly similar. Here, we take an alternative approach to map functional constraints on Zika virus's envelope (E) protein by using deep mutational scanning to measure how all amino acid mutations to the E protein affect viral growth in cell culture. The resulting sequence-function map is consistent with existing knowledge about E protein structure and function but also provides insight into mutation-level constraints in many regions of the protein that have not been well characterized in prior functional work. In addition, we extend our approach to completely map how mutations affect viral neutralization by two monoclonal antibodies, thereby precisely defining their functional epitopes. Overall, our study provides a valuable resource for understanding the effects of mutations to this important viral protein and also offers a roadmap for future work to map functional and antigenic selection to Zika virus at high resolution.IMPORTANCE Zika virus has recently been shown to be associated with severe birth defects. The virus's E protein mediates its ability to infect cells and is also the primary target of the antibodies that are elicited by natural infection and vaccines that are being developed against the virus. Therefore, determining the effects of mutations to this protein is important for understanding its function, its susceptibility to vaccine-mediated immunity, and its potential for future evolution. We completely mapped how amino acid mutations to the E protein affected the virus's ability to grow in cells in the laboratory and escape from several antibodies. The resulting maps relate changes in the E protein's sequence to changes in viral function and therefore provide a valuable complement to existing maps of the physical structure of the protein.

Complexities in understanding antimicrobial resistance across domesticated animal, human, and environmental systems.

Antimicrobial resistance (AMR) is a significant threat to both human and animal health. The spread of AMR bacteria and genes across systems can occur through a myriad of pathways, both related and unrelated to agriculture, including via wastewater, soils, manure applications, direct exchange between humans and animals, and food exposure. Tracing origins and drivers of AMR bacteria and genes is challenging due to the array of contexts and the complexity of interactions overlapping health practice, microbiology, genetics, applied science and engineering, as well as social and human factors. Critically assessing the diverse and sometimes contradictory AMR literature is a valuable step in identifying tractable mitigation options to stem AMR spread. In this article we review research on the nonfoodborne spread of AMR, with a focus on domesticated animals and the environment and possible exposures to humans. Attention is especially placed on delineating possible sources and causes of AMR bacterial phenotypes, including underpinning the genetics important to human and animal health.

Critically important antibiotics: criteria and approaches for measuring and reducing their use in food animal agriculture.

Globally, increasing acquired antimicrobial resistance among pathogenic bacteria presents an urgent challenge to human and animal health. As a result, significant efforts, such as the One Health Initiative, are underway to curtail and optimize the use of critically important antimicrobials for human medicine in all applications, including food animal production. This review discusses the rationale behind multiple and competing "critically important antimicrobial" lists and their contexts as created by international, regional, and national organizations; identifies discrepancies among these lists; and describes issues surrounding risk management recommendations that have been made by regulatory organizations on the use of antibiotics in food animal production. A more harmonized approach to defining criticality in its various contexts (e.g., for human versus animal health, enteric diseases versus other systemic infections, and direct versus indirect selection of resistance) is needed in order to identify shared contextual features, aid in their translation into risk management, and identify the best ways to maintain the health of food animals, all while keeping in mind the wider risks of antimicrobial resistance, environmental impacts, and animal welfare considerations.

From farm management to bacteriophage therapy: strategies to reduce antibiotic use in animal agriculture.

To reduce the use of antibiotics in animal agriculture, a number of effective or commercially viable alternatives have been implemented by food animal producers or are under development. Perhaps the most well-established strategies are flock and herd management practices to mitigate disease introduction and spread, and, subsequently, reduce the need for antibiotic use. While vaccines in food animal production have been used to prevent both bacterial and viral diseases, but historically, most vaccines have targeted viral diseases. Though vaccines against viral diseases can help reduce the need for antibiotic use by controlling the spread of secondary bacterial infections, more recent vaccines under development specifically target bacteria. New developments in selecting and potentially tailoring bacteriophages provide a promising avenue for controlling pathogenic bacteria without the need for traditional small-molecule antibiotics. In this article we discuss these established and emerging strategies, which are anticipated to reduce the reliance on antibiotics in food animal production and should reduce the prevalence and transmission to humans of antimicrobial resistant bacteria from these systems.

Antimicrobial-resistant bacterial infections from foods of animal origin: understanding and effectively communicating to consumers.

Consumers are increasingly interested in the attributes of the food they consume. This includes what is in the food and how it was raised; and at least some consumers are willing to pay a premium for products with specific attributes. However, the current plethora of labels on the market does not adequately address this issue; rather than providing actionable information, most labels add to the consumer confusion. In addition, there is a tendency toward "absence labels" that can contribute to a negative consumer perception of conventional products that may or may not include the attribute in question. Communication with consumers about the complex and highly technical issue of antimicrobial resistance (AMR) is challenging, and experiences from communication efforts about food safety-related issues demonstrate exactly how challenging this is to communicate clearly. General lessons learned from the science of risk communication can help guide efforts to communicate about the challenging issue of AMR. There are efforts underway to chart out a new approach. A new labeled animal production certification program is under development to provide choice for consumers, while reducing consumer confusion, which mandates antibiotic stewardship practices.

Patients Armed With Their Own Genetic Data Raise Tough Questions.

With the market for direct-to-consumer genetic testing expanding rapidly, clinicians are playing catch-up.

As Zika Reaches US Shores, States And Cities Struggle To Respond.

Local health officials have been forced to make do with public health resources stretched thin.

PATJ, a tight junction-associated PDZ protein, is a novel degradation target of high-risk human papillomavirus E6 and the alternatively spliced isoform 18 E6.

The E6 protein from high-risk human papillomavirus types interacts with and degrades several PDZ domain-containing proteins that localize to adherens junctions or tight junctions in polarized epithelial cells. We have identified the tight junction-associated multi-PDZ protein PATJ (PALS1-associated TJ protein) as a novel binding partner and degradation target of high-risk types 16 and 18 E6. PATJ functions in the assembly of the evolutionarily conserved CRB-PALS1-PATJ and Par6-aPKC-Par3 complexes and is critical for the formation of tight junctions in polarized cells. The ability of type 18 E6 full-length to bind to, and the subsequent degradation of, PATJ is dependent on its C-terminal PDZ binding motif. We demonstrate that the spliced 18 E6* protein, which lacks a C-terminal PDZ binding motif, associates with and degrades PATJ independently of full-length 18 E6. Thus, PATJ is the first binding partner that is degraded in response to both isoforms of 18 E6. The ability of E6 to utilize a non-E6AP ubiquitin ligase for the degradation of several PDZ binding partners has been suggested. We also demonstrate that 18 E6-mediated degradation of PATJ is not inhibited in cells where E6AP is silenced by shRNA. This suggests that the E6-E6AP complex is not required for the degradation of this protein target.

The positive transcription elongation factor activity of the vaccinia virus J3 protein is independent from its (nucleoside-2'-O-) methyltransferase and poly(A) polymerase stimulatory functions.

Previous genetic and biochemical experiments have shown that the vaccinia virus J3 protein has three different roles in mRNA synthesis and modification. First, J3 is a (nucleoside-2'-O-)methyltransferase which methylates the 2' position of the first transcribed nucleotide, thus converting a cap-0 to a cap-1 structure at the 5' ends of mRNAs. Second, J3 is a processivity factor for the virus coded poly(A) polymerase. Third, J3 has recently been shown to have intermediate and late gene positive transcription elongation factor activity in vivo. Previous experiments have shown that the poly(A) polymerase stimulatory activity and the (nucleoside-2'-O-)methyltransferase activity are two independent functions of the protein that can be genetically separated through site-directed mutagenesis. In this article, the relationship between the J3-mediated transcription elongation activity and the two other functions of the protein was investigated by constructing several site-directed mutant viruses that contain specific defects in either methyltransferase or poly(A) polymerase processivity functions. The results demonstrate that the J3 positive transcription elongation factor activity is a third independent function of the protein that is genetically separable from its two other functions in mRNA modification. The results also show that neither the poly(A) polymerase stimulatory nor the methyltransferase activities of the J3 protein is essential for virus growth in cell culture.