Much evidence suggests that the major immediate-early (IE) transactivator of human cytomegalovirus (HCMV), IE-2, is likely to be critical for efficient viral replication; however, the lack of an IE-2 mutant HCMV has precluded an experimental test of this hypothesis. As an initial step toward characterizing an IE-2 mutant, we first cloned the HCMV Towne genome as a bacterial artificial chromosome (BAC) and analyzed the ability of transfected Towne-BAC DNA (T-BACwt) to produce plaques following introduction into permissive human fibroblasts. Like Towne viral DNA, transfected T-BACwt DNA was infectious in permissive cells, and the resulting virus stocks were indistinguishable from Towne virus. We then used homologous recombination in Escherichia coli to delete the majority of UL122, the open reading frame encoding the unique portion of IE-2, from T-BACwt. From this deleted BAC, a third BAC clone in which the deletion was repaired with wild-type UL122 was created. In numerous transfections of permissive human foreskin fibroblast cells with these three BAC DNA clones, the rescued BAC and T-BACwt consistently yielded plaques, while the UL122 mutant BAC never generated plaques, even after 4 weeks. Protein and mRNA of other IE genes were readily detected from transfected UL122 mutant BAC DNA; however, reverse transcription-PCR failed to detect mRNA expression from any of five early genes examined. The generalized failure of this mutant to express early genes is consistent with expectations from in vitro assays which have demonstrated that IE-2 transactivates most HCMV promoters. These experiments provide the first direct demonstration that IE-2 is required for successful HCMV infection and indicate that virus lacking IE-2 arrests early in the replication cycle.

The aim of this work was to develop green and bioactive films with sodium alginate incorporating guava leaf extracts. Seven formulations were performed with a different sodium alginate: Guava leaf water extract (WE)/ethanolic extract (EE) proportions (100:0, 90:10, 85:15, 80:20), and glycerol were used as a plasticizer. The HPLC-PDA analysis showed the main phenolic compounds in WE were gallic acid, ellagic acid, quercetin-3-O-β-D-xylopyranoside, avicularin and quercetin. The main polyphenols in EE were rutin, isoquercitrin, quercetin-3-O-β-D-xylopyranoside, avicularin, quercitrin, quercetin and kaempferol. Guava leaf extracts could greatly enhance the antioxidant activity, antibacterial activity, tensile strength and water solubility of the sodium alginate film as well as the water barrier property, while inducing a decrease in the moisture content and elongation at the break. The FTIR and SEM analyses indicated that intermolecular hydrogen bonding between the guava leaf extract and sodium alginate resulted in a more compact structure in the composite films. These results indicated that sodium alginate-guava leaf extract films might be developed into antiradical and antimicrobial food packaging materials.

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Owning to its highly viscous and adhesive properties, viscous crude oil tends to easily adhere onto separation materials, resulting in serious fouling. Herein, we report an ampholytic nanofibrous membrane composed of a pair of oppositely charged biopolymerspositive chitosan (CS) and negative sodium alginate (ALG). CS and ALG were homogeneously complexed with each other to form a well-defined, interconnected, and entangled nanofibrous structure in a cosolvent of the LiOH/KOH/urea aqueous system after sequential treatment with chemical and physical cross-linking reactions. Benefitting from the coexistence of cationic and anionic groups on the surfaces of the CS/ALG composite nanofibers, the resulting CS/ALG composite nanofibrous membrane (CAM) has a strong hydrate layer to repel various oils including viscous crude oil. Moreover, The CAM could retain its wettability in harsh environments (e.g., acid, alkali, and salt). Furthermore, it could effectively separate the most troublesome crude oil-in-water emulsions with a superior separation efficiency (99%) and behaved well in terms of reusability. Meanwhile, CAM could realize multifunctional water treatment, where water-soluble dyes and heavy metal ions could be effectively wiped off by CAM from oily wastewater. The superiorities in low-cost, biodegradability, antioil fouling performance, oil/water separation properties, as well as multifunctional water treatment endow CAM with great potential in dealing with practical oily wastewater treatment.