Optimization of cottontail rabbit papilloma virus challenge technique.

Disease induced by Cottontail Rabbit Papilloma Virus (CRPV) scarification in domestic rabbits shares many attributes with disease induced by human papilloma virus (HPV). CRPV induces squamous papillomas in domestic rabbits, of which approximately 70% transform into invasive carcinomas. In advanced tumors, virus is often undetectable, and occasionally, some rabbits undergo spontaneous regression of papillomas. Techniques utilized to scarify rabbit skin are diverse, often labor intensive and time consuming with the possibility for significant variability. Using four unique infection techniques, resultant papilloma incidence, time to onset, and total papilloma volumes were compared to determine an optimal challenge method. Five rabbits were each infected with CRPV via a tattoo gun with and without ink, an intradermal injection, manual use of a tattoo needle, or a sterile blade followed by manual use of a tattoo needle. Papilloma formation was monitored weekly after inoculation for 6 weeks. CRPV papillomas began as pinpoint foci at 3 weeks post challenge and grew exponentially throughout the course of measurement. Individual foci coalesced rapidly to form larger papilloma aggregates. Although intradermal injection was well tolerated and easily performed, it was the worst method of papilloma production (2.2 mm(3) at 6 weeks). The best method, a sterile blade followed by manual use of a tattoo needle, produced significantly larger papillomas over all time periods (>1100 mm(3) at 6 weeks, P<0.01). Inoculation of CRPV using this method produces highly repeatable papillomas beginning 3 weeks post-infection.

A novel surfactant nanoemulsion with a unique non-irritant topical antimicrobial activity against bacteria, enveloped viruses and fungi.

A novel non-ionic surfactant nanoemulsion designated 8N8 has been tested for its biocidal activity. One percent 8N8 produced effective bactericidal activity against Bacillus cereus, Bacillus subtilis, Haemophilus influenzae, Neisseria gonorrhoeae, Streptococcus pneumoniae, and Vibrio cholerae in 15 minutes. In contrast, most enteric gram-negative bacteria were resistant to 8N8. One percent 8N8 was also virucidal within 15 minutes for all tested enveloped viruses, including Herpes simplex type 1, influenza A and vaccinia viruses. One percent 8N8 also demonstrated fungistatic activity on Candida albicans. The rapid and non-specific inactivation of vegetative bacteria and enveloped viruses, in addition to its fungistatic activity and low toxicity in experimental animals, makes 8N8 a potential candidate for use as a topical biocidal agent.

Anisocoria and middle cerebral artery saccular (berry) aneurysm in a rhesus macaque (Macaca mulatta).

A 27-year-old female rhesus macaque (Macaca mulatta) developed anisocoria. The left pupil was dilated and unresponsive to light. The macaque was euthanized because of unrelated reasons and the body was submitted for necropsy. On gross examination, a berry aneurysm of the right middle cerebral artery causing marked compression of the right optic tract was found. Arteriosclerotic changes were observed microscopically in the right middle cerebral and in the internal carotid arteries. The left iris was markedly degenerated, with atrophy of the constrictor muscle. Compression of the right optic tract may cause homonimus hemianopsia. A dilated and unresponsive left pupil indicated a lesion in the ipsilateral parasympathetic efferent pathway. In the absence of appreciable lesions of the left oculomotor nerve, the most likely cause of mydriasis was the iridic lesion. Intracranial aneurysms are common in humans (2 to 5%), but not in other species. Only about 10% of unruptured aneurysms are associated with neurologic deficits related to mechanical compression, such as visual deficits or anisocoria. Meticulous investigation of the ocular vascular and neural pathways led us to conclude that the anisocoria was unrelated to the aneurysm. To our knowledge, this report represents the first documented case of a naturally occurring intracranial aneurysm in nonhuman primates.

Prevention of murine influenza A virus pneumonitis by surfactant nano-emulsions.

Non-ionic surfactant nano-emulsions have extensive anti-microbial activity and are biocompatible with skin and mucous membranes at effective concentrations. Two nano-emulsion formulations (8N8 and 20N10) made from soybean oil, tributyl phosphate and Triton X-100, were tested for their ability to prevent murine influenza virus pneumonia in vivo. In the initial study, CD-1 mice were administered various dilutions of the nano-emulsions intranasally, and safe dosages and concentrations were determined. Non-toxic concentrations of the nano-emulsions were then mixed with influenza virus and applied to the nares of mice. Animals receiving mixtures of two different emulsions (8N8 or 20N10) and a LD50 of virus survived the challenge without evidence of viral infection. To determine if the nano-emulsions could prevent influenza virus infection in vivo when used as a prophylactic treatment, the nano-emulsions (8N8 at 1.0% and 20N10 at 1.0% or 0.2%) were applied to mouse nares 90 min before exposure to 5x10(5) p.f.u./ml virus by nebulized aerosol. Animals pretreated with the nano-emulsions had significantly decreased clinical signs of infection. Only 26.0% (8N8 at 1.0%), 31.25% (20N10 at 1.0%) and 37.0% (20N10 at 0.2%) of animals pretreated with nano-emulsion died from pneumonitis, whereas >80.0% of mock pretreated animals succumbed to infection (P<0.005). These findings suggest that non-ionic surfactant nano-emulsions have therapeutic potential for the prevention of influenza virus infection in vivo.

Inhibition of viral adhesion and infection by sialic-acid-conjugated dendritic polymers.

Multiple sialic acid (SA) residues conjugated to a linear polyacrylamide backbone are more effective than monomeric SA at inhibiting influenza-induced agglutination of red blood cells. However, "polymeric inhibitors" based on polyacrylamide backbones are cytotoxic. Dendritic polymers offer a nontoxic alternative to polyacrylamide and may provide a variety of potential synthetic inhibitors of influenza virus adhesion due to the wide range of available polymer structures. We evaluated several dendritic polymeric inhibitors, including spheroidal, linear, linear-dendron copolymers, comb-branched, and dendrigraft polymers, for the ability to inhibit virus hemagglutination (HA) and to block infection of mammalian cells in vitro. Four viruses were tested: influenza A H2N2 (selectively propagated two ways), X-31 influenza A H3N2, and sendai. The most potent of the linear and spheroidal inhibitors were 32-256-fold more effective than monomeric SA at inhibiting HA by the H2N2 influenza virus. Linear-dendron copolymers were 1025-8200-fold more effective against H2N2 influenza, X-31 influenza, and sendai viruses. The most effective were the comb-branched and dendrigraft inhibitors, which showed up to 50000-fold increased activity against these viruses. We were able to demonstrate significant (p < 0.001) dose-dependent reduction of influenza infection in mammalian cells by polymeric inhibitors, the first such demonstration for multivalent SA inhibitors. Effective dendrimer polymers were not cytotoxic to mammalian cells at therapeutic levels. Of additional interest, variation in the inhibitory effect was observed with different viruses, suggesting possible differences due to specific growth conditions of virus. SA-conjugated dendritic polymers may provide a new therapeutic modality for viruses that employ SA as their target receptor.

Review of exertional rhabdomyolysis and a case in a rhesus monkey (Macaca mulatta).

This is the first confirmed report of exertional rhabdomyolysis in a non-human primate. The monkey was singly housed and presented with anorexia and reluctance to move. There was no external evidence of trauma. Clinicopathologic findings included mild azotemia, marked elevation in serum creatine phosphokinase (CPK), alanine aminotransferase, aspartate aminotransferase, and myoglobinuria. Two days post-incident, the peripheral skeletal muscle had marked multifocal myonecrosis and fibrillar disruption without an inflammatory reaction. Treatment included diuresis and pain relief, and urinary output was monitored. The monkey recovered over the next two weeks. The major significance of skeletal muscle damage is the potential of released myoglobin to cause acute renal failure in the presence of other co-factors such as hypovolemia, acidosis, or ischemia. CPK levels can be highly variable and are inconsistent with the degree of muscle damage; however, CPK is thought to be the most sensitive enzyme marker for muscle necrosis. Because of the potential life-threatening sequelae, exertional rhabdomyolysis should be included as a differential diagnosis when similar clinical and pathological signs are observed.