To determine how the bed bug, Cimex lectularius, survives in a dry environment for many months without feeding, water-balance characteristics were compared for all stages from first-instar nymphs to adults. This species is characterized by a low net transpiration rate averaging < 0.2%/h, high tolerance for dehydration (30-40% loss in body water), and an impermeable cuticle as indicated by a high critical transition temperature (CTT) in the 35-40 degrees C range, implying that this insect is adapted for desiccation-hardiness. The capacity of adults to survive for 2 weeks at 0.00a(v) (a(v) = % RH/100) with no access to food or water exemplifies this trait. In contrast to more mature stages, first-instar nymphs contain more water, lose water at a faster rate, experience abrupt water loss at a lower temperature, and survive less time in dry air, suggesting that this stage is the most sensitive to water stress. This insect relies on blood to replenish water stores; none of the stages examined have the capacity to absorb water vapor (critical equilibrium activity, CEA > or = 0.99a(v)), and they drank only sparingly when offered free water. As the bed bugs progress through their development, they gradually reduce their water requirements while increasing their desiccation resistance. Surviving water stress is considerably enhanced behaviorally by quiescence, characterized by prolonged periods of inactivity, and by the formation of clusters that generate a water-conserving group effect.

Human papillomavirus 16 (HPV16) is a worldwide health threat and an etiologic agent of cervical cancer. To understand the antigenic properties of HPV16, we pursued a structural study to elucidate HPV capsids and antibody interactions. The cryo-electron microscopy (cryo-EM) structures of a mature HPV16 particle and an altered capsid particle were solved individually and as complexes with fragment of antibody (Fab) from the neutralizing antibody H16.V5. Fitted crystal structures provided a pseudoatomic model of the virus-Fab complex, which identified a precise footprint of H16.V5, including previously unrecognized residues. The altered-capsid-Fab complex map showed that binding of the Fab induced significant conformational changes that were not seen in the altered-capsid structure alone. These changes included more ordered surface loops, consolidated so-called "invading-arm" structures, and tighter intercapsomeric connections at the capsid floor. The H16.V5 Fab preferentially bound hexavalent capsomers likely with a stabilizing effect that directly correlated with the number of bound Fabs. Additional cryo-EM reconstructions of the virus-Fab complex for different incubation times and structural analysis provide a model for a hyperstabilization of the capsomer by H16.V5 Fab and showed that the Fab distinguishes subtle differences between antigenic sites. IMPORTANCE Our analysis of the cryo-EM reconstructions of the HPV16 capsids and virus-Fab complexes has identified the entire HPV.V5conformational epitope and demonstrated a detailed neutralization mechanism of this clinically important monoclonal antibody against HPV16. The Fab bound and ordered the apical loops of HPV16. This conformational change was transmitted to the lower region of the capsomer, resulting in enhanced intercapsomeric interactions evidenced by the more ordered capsid floor and "invading-arm" structures. This study advances the understanding of the neutralization mechanism used by H16.V5. Human papillomavirus (HPV) is a nonenveloped doublestranded DNA virus that can induce several epithelial cancers, especially cervical cancer (1-3). HPV16 is the most prevalent high-risk type of HPV (4, 5) and has been a primary target for the development of prophylactic vaccines (6, 7). HPV is epitheliotropic, and its replication is tightly associated with terminal differentiation of keratinocytes. This restricted tropism makes the production of high-titer preparations of authentic virion challenging. Alternative production methods have been developed to produce high-titer stocks of virus-like particles (VLP) (8), pseudovirions (PsV) (9), and quasivirions (QV) (10) while preserving the main attributes of the native capsid structure. These particles have been used successfully for vaccine development and for studies of antigenicity, receptor usage, entry mechanisms, and capsid structure.The infectious HPV has a Tϭ7 icosahedral capsid (55 to 60 nm in diameter), composed of 72 L1 capsid protein pentamers and up to 72 copies of L2 capsid protein located bene...

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The primary oligomerization domain of poliovirus polymerase, 3Dpol, is stabilized by the interaction of the back of the thumb subdomain of one molecule with the back of the palm subdomain of a second molecule, thus permitting the head-to-tail assembly of 3Dpol monomers into long fibers. The interaction of Arg-455 and Arg-456 of the thumb with Asp-339, Ser-341, and Asp-349 of the palm is key to the stability of this interface. We show that mutations predicted to completely disrupt this interface do not produce equivalent growth phenotypes. Virus encoding a polymerase with changes of both residues of the thumb to alanine is not viable; however, virus encoding a polymerase with changes of all three residues of the palm to alanine is viable. Biochemical analysis of 3Dpol derivatives containing the thumb or palm substitutions revealed that these derivatives are both incapable of forming long fibers, suggesting that polymerase fibers are not essential for virus viability. The RNA binding activity, polymerase activity, and thermal stability of these derivatives were equivalent to that of the wild-type enzyme. The two significant differences observed for the thumb mutant were a modest reduction in the ability of the altered 3CD proteinase to process the VP0/VP3 capsid precursor and a substantial reduction in the ability of the altered 3Dpol to catalyze oriI-templated uridylylation of VPg. The defect to uridylylation was a result of the inability of 3CD to stimulate this reaction. Because 3C alone can substitute for 3CD in this reaction, we conclude that the lethal replication phenotype associated with the thumb mutant is caused, in part, by the disruption of an interaction between the back of the thumb of 3Dpol and some undefined domain of 3C. We speculate that this interaction may also be critical for assembly of other complexes required for poliovirus genome replication.The RNA-dependent RNA polymerase (RdRP) 1 is the key component of the replication machinery of RNA viruses. The RdRP from poliovirus (3Dpol) serves as a paradigm for this class of nucleic acid polymerases. The crystal structure for 3Dpol (1) revealed that this polymerase has the typical topology observed for other nucleic acid polymerases and can be compared with a cupped, right hand with fingers, palm, and thumb subdomains (Fig. 1A). A unique feature of this polymerase, however, is the presence of two extensive regions of polymerase-polymerase interactions, referred to as interface I and interface II (1). Polymerase molecules interact in a "headto-tail" fashion to form long, extended fibers via interface I, and these fibers interact with each other via interface II. Interface I is formed by an interaction between the back of the thumb of one polymerase molecule and the back of the palm of a second polymerase molecule (Fig. 1B). A few of the critical interactions required for integrity of interface I are shown in Fig. 1C. Specifically, Arg-455 and Arg-456 of the thumb subdomain of one polymerase molecule interact with Asp-339, Ser-341, and Asp-349 of...

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