Diagnosis and treatment of conjunctivitis in house finches associated with mycoplasmosis in Minnesota.

An ongoing outbreak of Mycoplasma gallisepticum-associated conjunctivitis in house finches (Carpodacus mexicanus) that began in 1994 in the eastern United States has been spreading westward. House finches presenting with the clinical signs of M. gallisepticum-associated conjunctivitis were first seen at the Wildlife Rehabilitation Center of Minnesota (Minnesota, USA) in July of 1996, and 42 cases were admitted from 26 December 1996 to 10 August 1997. A nested PCR was designed for sensitive and specific detection of the presence of the organism. Twelve birds were treated with oral enrofloxacin (15 mg/kg, twice daily for 21 days) and ophthalmic gentamicin (twice daily for 21 days). All treated birds showed resolution of clinical signs. Following treatment, finches were held for up to 6 mo and tested for the presence of M. gallisepticum by culture and nested polymerase chain reaction (PCR). Eight of twelve finches (67%) were positive for M. gallisepticum by nested-PCR and four (33%) were positive by culture. The results suggest that oral enrofloxacin and opthalmic gentamicin are not an effective treatment for the eradication of M. gallisepticum in house finches. Further, the results show that nested PCR is an effective method for detection of M. gallisepticum in house finches and was more sensitive than culture.

Sulfated polyester interactions with the CD4 molecule and with the third variable loop domain (v3) of gp120 are chemically distinct.

Sulfated polyesters (SP) that inhibit HIV infection interact with both the gp120 binding region of CD4 molecules and with the v3 domain loop of gp120 molecules (gp120/v3) but the contributions of these interactions to the inhibition of HIV env-mediated fusion are presently unclear. In order to characterize the molecular mechanisms by which SP inhibit HIV env-mediated fusion, we studied the effect of SP treatment on env-mediated fusion of CD4+ cells driven by recombinant vaccinia virus (vPE-16) and on the binding of anti-HIV MAbs to cellular gp120 or purified, rgp120. SP were more effective than neutralizing anti-gp120/v3 MAbs in inhibiting env-mediated fusion. In addition, SP interacted with the v3 loop of gp120 to inhibit the binding of the neutralizing MAb 9284 but not the binding of 9305, a neutralizing anti-gp120/v3 MAb that binds to an adjacent epitope. Because SP are polyanions, we compared the chemical properties of the SP-gp120/v3 and SP-CD4 interactions. Whereas the ability of SP to inhibit the binding of MAb 9284 and rgp120 was relatively independent of NaCl concentrations, the ability of SP to interfere with rCD4-rgp120 binding depended on the NaCl concentration and was maximal at low NaCl concentrations. In addition, the SP-gp120 interaction was found to be reversible, in contrast to the SP-rCD4 interaction which was previously shown to be relatively irreversible at low salt. These data are consistent with the notions that the interaction of SP with CD4 is primarily electrostatic, but that the interaction of SP with gp120 has complex characteristics that implicate a role for protein conformation.

The G protein of vesicular stomatitis virus has free access into and egress from the smooth endoplasmic reticulum of UT-1 cells.

We have investigated the role of the smooth endoplasmic reticulum (SER) of UT-1 cells in the biogenesis of the glycoprotein (G) of vesicular stomatitis virus (VSV). Using immunofluorescence microscopy, we observed the wild type G protein in the SER of infected cells. When these cells were infected with the mutant VSV strain ts045, the G protein was unable to reach the Golgi apparatus at 40 degrees C, but was able to exit the rough endoplasmic reticulum (RER) and accumulate in the SER. Ribophorin II, a RER marker, remained excluded from the SER during the viral infection, ruling out the possibility that the infection had destroyed the separate identities of these two organelles. Thus, the mechanism that results in the retention of this mutant glycoprotein in the ER at 39.9 degrees C does not limit its lateral mobility within the ER system. We have also localized GRP78/BiP to the SER of UT-1 cells indicating that other mutant proteins may also have access to this organelle. Upon incubation at 32 degrees C, the mutant G protein was able to leave the SER and move to the Golgi apparatus. To measure how rapidly this transfer occurs, we assayed the conversion of the G protein's N-linked oligosaccharides from endoglycosidase H-sensitive to endoglycosidase H-resistant forms. After a 5-min lag, transport of the G protein followed first order kinetics (t1/2 = 15 min). In contrast, no lag was seen in the transport of G protein that had accumulated in the RER of control UT-1 cells lacking extensive SER. In these cells, the transport of G protein also exhibited first order kinetics (t1/2 = 17 min). Possible implications of this lag are discussed.

The M protein of vesicular stomatitis virus associates specifically with the basolateral membranes of polarized epithelial cells independently of the G protein.

Using monoclonal antibodies and indirect immunofluorescence microscopy, we investigated the distribution of the M protein in situ in vesicular stomatitis virus-(VSV) infected MDCK cells. M protein was observed free in the cytoplasm and associated with the plasma membrane. Using the ts045 mutant of VSV to uncouple the synthesis and transport of the VSV G protein we demonstrated that this distribution was not related to the presence of G protein on the cell surface. Sections of epon-embedded infected cells labeled with antibody to the M protein and processed for indirect horseradish peroxidase immunocytochemistry revealed that the M protein was associated specifically with the basolateral plasma membrane. The G and M proteins of VSV have therefore evolved features which bring them independently to the basolateral membrane of polarized epithelial cells and allow virus to bud specifically from that membrane.