Prevention of fetal infection in heifers challenged with bovine viral diarrhoea virus type 1a by vaccination with a type 1c or type 1a vaccine.

AIMS: To evaluate a vaccine containing type 1c bovine viral diarrhoea (BVD) virus for prevention of fetal infection in pregnant heifers when challenged with New Zealand BVD virus type 1a 6 months after vaccination, compared to unvaccinated heifers and heifers vaccinated with a vaccine containing type 1a BVD virus. METHODS: Fifty five crossbred Friesian heifers, free from BVD virus and antibody, were randomly allocated to three groups. Twenty five heifers were vaccinated twice with a vaccine containing type 1c BVD virus (T1c group), and 10 heifers with a vaccine containing type 1a BVD virus (T1a group), and 20 heifers were unvaccinated (NC group). After oestrus synchronisation the heifers were bred by artificial insemination followed by natural bull mating. Six months after booster vaccination 15 heifers from the T1c group, eight from the T1a group, and 15 from the NC group, were exposed to four calves that were persistently infected with type 1a BVD virus, for 4 weeks. At the beginning of the challenge phase 36/38 heifers were 72-74 days pregnant and 2/38 heifers were approximately 53 days pregnant. Approximately 52 days after the start of the challenge the heifers were subjected to euthanasia and fetal tissues were collected for the detection of BVD virus by ELISA in fetal heart blood and PCR in fetal tissues. RESULTS: Based on PCR results, BVD virus was detected in 15/15 fetuses in the NC group, compared to 4/14 fetuses in the T1c group and 3/8 fetuses in the T1a group. The proportion of BVD virus-positive fetuses was lower in both vaccinated groups compared to the NC group (p<0.002), but there was no difference in proportions between the vaccinated groups (p=1.00). Fetal protection, expressed as the prevented fraction, was 71.4 (95% CI=41.9-91.6)% and 62.5 (95% CI=24.5-91.5)% for the T1c and T1a groups, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: The vaccines containing killed type 1c and type 1a BVD viruses significantly reduced fetal infection following challenge with a New Zealand type 1a BVD virus. Prevention of fetal infection by vaccination may not be 100%, and the risk of persistently infected calves being born to some vaccinated cattle should be acknowledged and managed as part of a BVD control programme.

Improved procedure for implanting radiotransmitters in the coelomic cavity of snakes.

OBJECTIVE: To investigate the expulsion of radiotransmitters in snakes and modify the surgical technique for coelomic implantation to prevent its occurrence. DESIGN: To enable monitoring of snakes for an ecological study, radiotransmitters were implanted in 23 south-west carpet pythons (Morelia spilota imbricata) using the standard surgical technique. In a further 23 pythons we used a refinement of the technique, which anchored the tracking device, using non-dissolvable sutures, to the snake's rib-cage. We also investigated the potential mechanisms for expelling the radiotransmitters in one snake that underwent an exploratory coeliotomy. RESULTS: Of the initial group of snakes, 12 (52%) expelled the radiotransmitter between 4 days and 3 years post implantation. In the later group, which underwent the refined technique of implantation, none of the radiotransmitters was expelled and no adverse responses were observed. CONCLUSION: An appropriately sized radiotransmitter anchored to the rib-cage of the snake will prevent expulsion of the device and appears to be well tolerated. Non-attachment of the tracking device enables it to migrate along the length of the body, particularly during feeding and reproduction. Caudal positioning of the transmitter's antenna provides a possible pathogenesis for expulsion into the cloaca.

Ablation of Teflon granulomas in the canine larynx with the free-electron laser.

This study develops a canine model for the treatment of laryngeal Teflon granulomas and demonstrates endoscopic ablation using the free-electron laser (FEL) set at a wavelength of 8.5 microm. Laryngeal Teflon granulomas may cause dysphonia and airway obstruction, and they are difficult to remove. The infrared absorption spectrum of Teflon reveals a strong absorption peak centered at 8.5 microm. In this study, 12 dogs had the right vocal cord injected with Teflon paste. Two months later, Teflon granuloma formation was confirmed histologically. Laser incisions into the granulomas were performed at 3 different wavelengths: 7.4 microm (FEL), 8.5 microm (FEL), and 10.6 microm (carbon dioxide laser). Histopathologic analysis was performed at 1 week and 6 weeks after the laser incisions. The FEL at the 8.5-microm wavelength was found to optimally ablate the Teflon granulomas, but the granulomas persisted in the specimens treated with 7.4 microm (FEL) and 10.6 microm (carbon dioxide laser).

Structural characterizations of nonpeptidic thiadiazole inhibitors of matrix metalloproteinases reveal the basis for stromelysin selectivity.

The binding of two 5-substituted-1,3,4-thiadiazole-2-thione inhibitors to the matrix metalloproteinase stromelysin (MMP-3) have been characterized by protein crystallography. Both inhibitors coordinate to the catalytic zinc cation via an exocyclic sulfur and lay in an unusual position across the unprimed (P1-P3) side of the proteinase active site. Nitrogen atoms in the thiadiazole moiety make specific hydrogen bond interactions with enzyme structural elements that are conserved across all enzymes in the matrix metalloproteinase class. Strong hydrophobic interactions between the inhibitors and the side chain of tyrosine-155 appear to be responsible for the very high selectivity of these inhibitors for stromelysin. In these enzyme/inhibitor complexes, the S1' enzyme subsite is unoccupied. A conformational rearrangement of the catalytic domain occurs that reveals an inherent flexibility of the substrate binding region leading to speculation about a possible mechanism for modulation of stromelysin activity and selectivity.

Posterior glottic stenosis: a canine model.

Posterior glottic stenosis is a disabling disease in which the vocal folds are fixed near the midline. This allows adequate vocal fold adduction for voicing, but does not permit useful abduction for ventilation. The most common cause is prolonged endotracheal intubation for mechanical ventilation, and the incidence is estimated at 4% for intubations between 5 and 10 days. Currently, our understanding and treatment modalities are based on retrospective reviews of small, nonrandomized clinical experiences. The purpose of this project was to develop an animal model that would improve our understanding of histologic changes and allow future prospective randomized trials for therapeutic intervention. Twelve dogs, 15 to 25 kg, were randomly divided into three groups. Animals in group I had a superficial injury produced in the tissue over the right cricoarytenoid joint; animals in group 2 had a deep soft tissue injury produced; and animals in group 3 underwent joint opening. The animals were allowed to recover for 2 months. Morphometric analysis of the harvested larynges demonstrated clinically significant limitation in motion in the animals with deep soft tissue injury and in animals with joint disruption. Histologic analysis revealed various degrees of injury, from loss of subepithelial soft tissue to cartilaginous resorption and fusion of the arytenoid to the cricoid. These findings were directly related to the depth of the initial injury. It is possible to produce posterior glottic stenosis in the canine species. This will serve as a reliable animal model for future study.

Cation binding to the integrin CD11b I domain and activation model assessment.

BACKGROUND: The integrin family of cell-surface receptors mediate cell adhesion through interactions with the extracellular matrix or other cell-surface receptors. The alpha chain of some integrin heterodimers includes an inserted 'I domain' of about 200 amino acids which binds divalent metal ions and is essential for integrin function. Lee et al. proposed that the I domain of the integrin CD11b adopts a unique 'active' conformation when bound to its counter receptor. In addition, they proposed that the lack of adhesion in the presence of Ca2+ ion reflected the stabilization of an 'inactive' I-domain conformation. We set out to independently determine the structure of the CD11 b I domain and to evaluate the structural effects of divalent ion binding to this protein. RESULTS: We have determined the X-ray structure of a new crystal form of the CD11 b I domain in the absence of added metal ions by multiple isomorphous replacement (MIR). Metal ions were easily introduced into this crystal form allowing the straight-forward assessment of the structural effects of divalent cation binding at the metal ion dependent adhesion site (MIDAS). The equilibrium binding constants for these ions were determined by titration calorimetry. The overall protein conformation and metal-ion coordination of the I domain is the same as that observed for all previously reported CD11 a I-domain structures and a CD11 b I-domain complex with Mn2+. These structures define a majority conformation. CONCLUSIONS: Addition of the cations Mg2+, Mn2+ and Cd2+ to the metal-free I domain does not induce conformational changes in the crystalline environment. Moreover, we find that Ca2+ binds poorly to the I domain which serves to explain its failure to support adhesion. We show that the active conformation proposed by Lee et al, is likely to be a construct artifact and we propose that the currently available data do not support a dramatic structural transition for the I domain during counter-receptor binding.

Delayed regional metastasis from midfacial squamous carcinomas.

BACKGROUND: Metastases from mucosal and cutaneous carcinomas can present in a delayed fashion, and this late presentation may confer a different prognosis after conventional treatment. METHODS: We present a series of patients in which there was a significant time delay between the treatment of a squamous carcinoma of the skin or mucosa of the midface and the detection of regional metastases in 12 of the 13 cases. Primary tumors were located on the lower lip and commissure (n = 3), nasal tip (n = 2), nasal ala (n = 1), columella (n = 1), nasofacial crease (n = 2), maxillary alveolus (n = 3), and mandibular alveolus (n = 1). Metastatic spread manifested by palpable perifacial or submandibular lymph nodes was not evident until greater than 11 months after the treatment of the primary site in 12 of 13 patients (range, 3-45 months). Nine of the patients were clinically staged as N1, whereas there was one each in the N2a, N2b, N2c, and N3 categories. Eleven of the 13 patients were initially seen with palpable disease involving the perifacial nodes within or around the submandibular gland. All patients were treated with neck dissection except one, who refused surgical treatment and underwent a second course of radiotherapy to the cervical region. The nine patients initially seen with clinical stage N1 disease underwent neck dissection with preservation of the sternocleidomastoid, internal jugular vein, and accessory nerve. RESULTS: Of 10 patients with perifacial node metastases who underwent neck dissection, 8 required sacrifice of the marginal mandibular nerve and overlying platysma to gain adequate margin. Extracapsular spread was present in 11 patients, (8 of 9 who were clinically N1). Postoperative radiotherapy was recommended to all patients with extracapsular spread, although only 7 of the 11 received radiotherapy. There were no regional recurrences after a minimum follow-up of 1 year (range, 12-65 months; mean, 31.4 months). Histologic grade appeared to have no influence on prognosis. CONCLUSIONS: This cohort demonstrates the ability of midfacial squamous cell carcinoma to manifest regional metastatic disease over a delayed time. This delayed presentation appears to confer a more favorable response to treatment. For midfacial cancers, the perifacial nodes are at greatest risk for metastatic spread. For tumors in this region, primary treatment of the neck is probably not warranted, but careful extended follow-up for the potential of delayed cervical metastasis is prudent.

Ablation of teflon granuloma with the free-electron laser emitting in the eight- to nine-micron range.

In this study, we developed a rat model for Teflon granuloma and used this model to evaluate the removal of the granuloma at laser wavelengths at which Teflon has a maximal absorption. Twenty-four Teflon granulomas were created in 12 rats, and the gross and histologic effects from laser incision at four different wavelengths (8.25, 8.5, 8.75, and 10.6 microm) were evaluated acutely and at 7 and 14 days postoperatively. Polytetrafluoroethylene, or Teflon, is a relatively inert substance that has been used over the past 4 decades for endoscopic injection into the thyroarytenoid muscle of the larynx for the purposes of laryngeal rehabilitation in cases of unilateral vocal fold paralysis or incomplete glottic closure. In certain cases in which formation of granulomatous reaction to the Teflon occurs, patients may have significant dysphonia or airway compromise. Once Teflon has infiltrated the surrounding tissue planes, it is exceedingly difficult to remove endoscopically. Endoscopic removal of this granuloma is usually attempted with the carbon dioxide (CO2) laser and has had limited success. Examination of the infrared absorption spectrum of polytetrafluoroethylene reveals strong absorption in the mid-infrared region in the 8- to 9-microm range, with minimal absorption at 10.6 microm. Therefore, this absorption spectrum predicts a more efficient vaporization of Teflon at wavelengths near 8.5 microm. Using the free-electron laser to generate 8.25-, 8.5-, and 8.75-microm laser light, we found Teflon granuloma ablation was far superior to CO2 laser ablation at 10.6 microm. The 8.25-, 8.5-, and 8.75-microm wavelengths selectively ablated Teflon granuloma with minimal to no collateral thermal injury to tissue. The differences in thermal effects observed while actually using the lasers were confirmed histologically.

Histologic study of oral mucosa wound healing: a comparison of a 6.0- to 6.8-micrometer pulsed laser and a carbon dioxide laser.

Incisional wound healing in the canine oral mucosa was histologically monitored at 3, 7, and 14 days after incision. Healing was compared from a scalpel, a carbon dioxide (CO2) laser at 10.6 microm, and the Vanderbilt free-electron laser tuned to 6.0, 6.45, and 6.8 microm. A significant delay in wound healing was observed when incisions were made with the CO2 laser, probably attributable to the excess thermal damage caused by the continuous-wave laser beam. When using the short pulsed, free-electron laser, a much smaller delay comparable to the scalpel wound healing was observed. This smaller delay tended to decrease with increasing tissue absorption. The results emphasize the greater importance of laser pulse duration rather than wavelength in relation to the subsequent wound healing.

Characterization of the protective borreliacidal antibody response in humans and hamsters after vaccination with a Borrelia burgdorferi outer surface protein A vaccine.

Significant borreliacidal antibody was induced in volunteers and hamsters 60 days after primary and secondary vaccination with high concentrations of recombinant outer surface protein A (rOspA). However, the borreliacidal antibody response waned rapidly. Only 1 person had detectable cidal activity 180 days after vaccination. Similarly, the borreliacidal antibody response waned rapidly in hamsters by week 10 of vaccination. By contrast, the total anti-rOspA antibody response remained elevated in volunteers and hamsters. When isolates of Borrelia burgdorferi sensu lato were incubated in sera from vaccinated humans or hamsters, only the vaccine-specific isolate was killed. These results were confirmed by challenging rOspA-vaccinated hamsters with different isolates of B. burgdorferi sensu lato. The results showed that monitoring total rOspA antibody is inappropriate for evaluating the efficacy of an rOspA vaccine. The rOspA vaccine must be improved to yield comprehensive protection and maintain sustained levels of protective borreliacidal antibodies.

An investigation of acute facial paralysis in animals induced by exposure of the tympanic membrane to cold air.

The goal of this investigation was to test the hypothesis that tympanic membrane exposure to cold air is a cause of acute facial palsy. A series of acute invasive experiments and a series of chronic noninvasive experiments were conducted in both cats and dogs. In the acute studies, stimulation was applied intracranially to the facial nerve root through a stereotaxically placed microelectrode and recordings of compound action potentials obtained extracranially from the facial nerve. Nerve conduction was monitored continuously during the application of cold air to the tympanic membrane. Nerve conduction disturbances were observed in all animals tested (8), and reduction in compound action potential amplitude ranged from 33% to 96%. Histologic analysis of the intratemporal portion of the facial nerve was performed in the animal exhibiting the greatest block in conduction, representative of a near-total paralysis. Axon swelling, demyelinization, and degeneration (Bungner's bands) without inflammation were apparent along the entire tympanic membrane segment. Interstitial swelling of nerve endoneurium was also present at the second genu and vertical segment. In the chronic studies, animals were exposed to cold air and monitored daily for facial paralysis after recovery from anesthesia. None of the animals demonstrated any detectable behavioral facial paralysis.

Detection of borreliacidal antibodies by flow cytometry. An accurate, highly specific serodiagnostic test for Lyme disease.

BACKGROUND: Borreliacidal antibodies can be detected in serum samples from patients with early or late Lyme disease symptoms. When these serum samples are incubated with Borrelia burgdorferi and complement, spirochetes are rapidly killed. Detection of these antibodies can be used as a serodiagnostic test. METHODS: Individual serum samples containing IgM or IgG borreliacidal antibodies were Used to develop a method for detection using flow cytometry. An additional 10 case-defined Lyme disease serum samples and 10 normal serum samples were used to confirm appropriate flow cytometric parameters. To determine specificity, 157 normal serum samples and 104 potential cross-reactive serum samples were tested for borreliacidal activity and antibodies to B burgdorferi using indirect fluorescent antibody or enzyme immunoassay. RESULTS: Flow cytometry can be used to detect borreliacidal activity within 16 to 24 hours after incubation of B burgdorferi organisms. Lyme disease serum, and complement. Significant borreliacidal activity was detected in all Lyme disease serum samples. The percentages of positive normal serum samples were comparable (6% to 10%) using all three assays. In addition, the indirect fluorescent antibody and enzyme immunoassay identified 41 (39%) and 47 (45%) potential cross-reactive serum samples as positive, respectively. In contrast, significant borreliacidal activity was not detected in any potential cross-reactive serum samples. CONCLUSION: Detection of borreliacidal antibody, unlike indirect fluorescent antibody and enzyme immunoassay, is an accurate, highly specific serodiagnostic test for detection of Lyme disease.

Lyme disease: clinical features, classification, and epidemiology in the upper midwest.

Lyme disease can be classified using the terminology of syphilis. In this series of 95 cases from the upper midwest, early cases, defined as an illness of less than 2 months, were more likely to have lived in or recently visited a highly endemic area. Unlike late cases, early cases presented entirely in the nonwinter months (p less than .001). Early disease was further subdivided into primary and secondary disease. Ninety percent of primary and 43% of secondary cases had erythema migrans, while no late cases had active erythema migrans (p less than .001). Clinical manifestations of nonspecific inflammation, except for arthralgia, were more common in early than late disease (p less than .01). In secondary cases, monoarticular arthritis was slightly more common than polyarticular arthritis, with the reverse occurring in late disease (p less than .05). Indirect fluorescent antibody testing revealed a ratio of IgM to IgG antibodies to be helpful in distinguishing early from late disease. Antibacterial therapy in early, primary cases caused Jarisch-Herxheimer reaction 7% of the time. Despite longer and more frequent parenteral therapy, late Lyme disease frequently required retreatment, owing to poor clinical response (p less than .05).