Fundamentals of host immune response against Brucella abortus: what the mouse model has revealed about control of infection

The studies reviewed here evaluated the role cellular immune system components play in control of brucellosis by conducting comparative studies with brucella-resistant C57BL/10 or C57BL/6 mice and susceptible BALB/c mice. We have shown by both in vitro and in vivo studies that activation of macrophages with interferon-gamma (IFN-γ) is an important factor for control of infection with B. abortus in the mouse model and that the mechanism of anti-brucella activity largely involved reactive oxygen intermediates. Differences in control of the organism by resistant and susceptible mice was not related to inherent differences in the ability of their macrophages to control infection either with or without IFN-γ activation nor was it attributable to NK cells since we found no role for them in control of brucellosis in either mouse strain. However, relative resistance to brucellosis did correlate with increased production of IFN-γ by CD4 T cells during the first weeks after infection while IL-10 contributed to susceptibility in BALB/c mice. Moreover, by 3 weeks post-infection splenocytes from the susceptible BALB/c mice failed to produce IFN-γ and relied on TNF- as well as CD8 T cells to control infection until the end of the plateau phase around 6 weeks post-infection when IFN-γ production resumed and clearance began. In contrast, IFN-γ was crucial for control throughout the infection in the more resistant C57BL/6 mice and the mice died in its absence by 6 weeks post-infection compared to 12 weeks for the more susceptible mice that relied on additional mechanisms of control. In contrast to the IFN-γ knock-out mice, both β2 microglobulin knock-out C57BL/6 mice, which do not express conventional MHC class I molecules and thus cannot present antigen to CD8 T cells, or perforin knock-out C57BL/6 mice, which have no T cell cytotoxic activity, controlled and cleared the infection as well as normal C57BL/6 mice. The hiatus of IFN-γ production in BALB/c mice correlated with very high levels of total IL-12 and it was postulated that the lack of IFN-γ was a consequence of p40 homodimer blocking activity. However, reduction of p40 IL-12 in vivo through administration of indomethacin reduced the infection without a concomitant measurable increase in IFN-γ. Current studies are aimed at elucidating the mechanism of the IFN-γ hiatus.     

Persistent Frenulum of the Epiglottis in Four Foals

OBJECTIVE: To report the clinical signs and management of 4 foals with persistent frenulum of the epiglottis. STUDY DESIGN: Case report. ANIMALS: Four newborn foals. METHODS: Foals were admitted with a complaint of oronasal reflux after nursing. Variable systemic signs of aspiration pneumonia were evident. Nasal endoscopy confirmed persistent dorsal displacement of the soft palate. Persistent frenulum of the epiglottis, confirmed by oral endoscopic examination, was transected. RESULTS: After surgery, all foals had the epiglottis positioned normally, dorsal to the soft palate. Clinical signs of oronasal reflux resolved by the second nursing attempt in 3 foals, whereas this was difficult to assess in one foal that was nursed intermittently because of the mares behavior. This foal died 2 days later. The other 3 foals have had normal epiglottic function for 2-4 years. CONCLUSIONS: Persistent frenulum of the epiglottis should be considered in foals with oronasal reflux from birth. With appropriate medical and surgical management the prognosis for resolution should be good.     

Molecular typing of Mycoplasma agalactiae: tracing European-wide genetic diversity and an endemic clonal population

Mycoplasma agalactiae causes chronic infections in small ruminants and remains endemic in many regions of the world, despite intensive and costly eradication programs. In this study, the innate genomic plasticity of M. agalactiae was exploited to design and assess a combination of molecular epidemiological tools to trace the pathogen in different geographic locations and to understand its emergence or re-emergence after eradication campaigns. For this purpose, two collections of M. agalactiae isolates, representing European outbreaks or localized endemic disease in a single region of France, were subjected to RFLP (Restriction Fragment Length Polymorphism) analyses using two sets of DNA probes (distributed across the genome and specific for the vpma gene locus), and a previously described VNTR (Variable Number Tandem Repeats) analysis. A combination of four genome-specific DNA probes and two VNTRs gave the highest discriminative power. Molecular typing revealed that, while isolates from diverse geographical origins fell into clearly different groups, the endemic disease repeatedly observed in the Western Pyrenees region over the past 30 years has been caused by a unique subtype of M. agalactiae. This indicates that the re-emergence of the pathogen after seemingly successful eradication programs is not due to the importation of exotic strains, but to the persistence of local reservoirs of infection.     

Identification of Hammondia heydorni oocysts by a heminested-PCR (hnPCR-AP10) based on the H. heydorni RAPD fragment AP10

Toxoplasma gondii, Hammondia hammondi, Neospora caninum, Neospora hughesi and Hammondia heydorni are members of the Toxoplasmatinae sub-family. They are closely related coccidians with similarly sized oocysts. Molecular diagnostic techniques, especially those based on polymerase chain reaction (PCR), can be successfully applied for the differentiation of Hammondia-like oocysts. In this paper, we describe a rapid and simple method for the identification of H. heydorni oocysts among other members of the Toxoplasmatinae sub-family, using a heminested-PCR (hnPCR-AP10) based on a H. heydorni RAPD fragment available in molecular database. DNA of oocysts of H. heydorni yielded a specific fragment of 289-290 bp in the heminested-PCR assay. No product was yielded when the primers were used for the amplification of DNA extracted from T. gondii, N. caninum, N. hughesi and H. hammondi, thus allowing the differentiation of H. heydorni among other members of the Toxoplasmatinae sub-family. The hnPCR-AP10 was capable of detecting H. heydorni genetic sequences from suspensions with at least 10 oocysts. In conclusion, the hnPCR-AP10 proved to be a reliable method to be used in the identification of H. heydorni oocysts from feces of dogs.     

Use of the caudolateral curvilinear osteophyte as an early marker for future development of osteoarthritis associated with hip dysplasia in dogs

OBJECTIVE: To determine the relationship between the caudolateral curvilinear osteophyte (CCO) and osteoarthritis associated with hip dysplasia in dogs. DESIGN: Longitudinal cohort study. ANIMALS: 48 Labrador Retrievers from 7 litters. PROCEDURE: In each of 24 sex- and size-matched pairs fed the same diet, a restricted-fed dog was fed 25% less than a control dog for life. The dogs' hips were evaluated in the standard ventrodorsal hip-extended radiographic projection at 16, 30, and 52 weeks of age and then yearly for life. Histologic examination of hip joint tissues was performed on 45 dogs. RESULTS: Median age at death was 11.2 years. Adjusting for feeding group, dogs with a CCO were 3.7 times as likely to develop radiographic signs of osteoarthritis than those without a CCO. Stratified by diet, 100% of the control dogs with a CCO developed radiographic signs of osteoarthritis and 55% of restricted-fed dogs with a CCO developed radiographic signs of osteoarthritis. The CCO was the first radiographic change seen in 22 of 29 (76%) dogs with osteoarthritis. Overall, 35 of 37 (95%) dogs with a CCO had histopathologic lesions of osteoarthritis. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate a relationship between a CCO on the femoral neck and subsequent development of radiographic signs of osteoarthritis in Labrador Retrievers evaluated over their life span. A CCO is an important early radiographic indication of osteoarthritis associated with canine hip dysplasia.     

Comparison of different intracellular cryoprotectants on the solid surface vitrification of red-rumped agouti (Dasyprocta Leporina Lichtenstein, 1823) ovarian tissue

In contributing to the conservation of wild rodents, the aim of this study was to evaluate the use of distinct cryoprotectants, separately or in combination, for solid surface vitrification (SSV) of red-rumped agouti ovarian tissue. Ovarian cortex from nine females was recovered and fragmented. Fresh fragments (control) were used to analyse the pre-antral follicle (PF) morphology using a histologic procedure, viability using the Trypan blue test, cell proliferation by counting the argyrophilic nucleolar organizing regions (Ag-NORs technique) and DNA integrity using the TUNEL assay. The remaining fragments were vitrified using SSV method with 3 M or 6 M ethylene glycol (EG) or dimethyl sulfoxide (DMSO), or in combination (3 M EG/3 M DMSO), and further evaluated as reported for the fresh samples. All cryoprotectants were effective at preserving PFs morphology compared to the control group (80.7 ± 5.21%), except 6 M EG and 3 M DMSO that provoked a significant (p < .05) decrease on the values of morphologically normal primary (60.0 ± 19.0%) and primordial (44 ± 4.5%) follicles, respectively. Regarding viability, all cryoprotectants provided values similar to that verified for the control group (79.0%), but a significant decrease (p < .05) was observed with EG/DMSO combination (59%). Using Ag-NORs technique, the highest (p < .05) cell proliferative capacity was detected when using EG at each tested concentration. The TUNEL proved the preservation of DNA integrity regardless of the cryoprotectant. In summary, we suggest the use of 3 M EG for the solid surface vitrification of red-rumped agouti ovarian tissue.     

COMPARISON OF THORACIC ULTRASONOGRAPHY AND RADIOGRAPHY FOR THE DETECTION OF INDUCED SMALL VOLUME PNEUMOTHORAX IN THE HORSE

Small volume pneumothorax can be challenging to diagnose in horses. The current standard method for diagnosis is standing thoracic radiography. We hypothesized that thoracic ultrasonography would be more sensitive. Objectives of this prospective, experimental study were to describe a thoracic ultrasound method for detection of small volume pneumothorax in horses and to compare results of radiography and ultrasound in a sample of horses with induced small volume pneumothorax. Six mature healthy horses were recruited for this study. For each horse, five 50 ml air boluses were sequentially introduced via a teat cannula into the pleural space. Lateral thoracic radiographs and standardized ultrasound (2D and M‐mode) examinations of both hemithoraces were performed following administration of each 50 ml air bolus. Radiographs and ultrasound images/videos were analyzed for detection of pneumothorax by four independent investigators who were unaware of treatment status. Sensitivity, specificity, positive predictive values, negative predictive values, and agreement among investigators (Kappa test, κ) were calculated for radiography, 2D and M‐mode ultrasound. Comparisons were made using a chi‐squared exact test with significance set at P < 0.05. Two‐dimensional (84%) and M‐mode (80%) ultrasound were more sensitive than radiography (48%) for pneumothorax detection (P = 0.02 and P = 0.04, respectively). Specificity and positive predictive values were similar for all three imaging modalities (P = 1). Agreement between investigators for pneumothorax detection was excellent for 2D ultrasound (κ = 1), very good for M‐mode ultrasound (κ = 0.87), and good for radiography (κ = 0.79). Findings from this experimental study supported the use of thoracic ultrasonography as a diagnostic method for detecting pneumothorax in horses.     

Influence of three anesthetic protocols on glomerular filtration rate in dogs

OBJECTIVE: To investigate renal function in clinically normal dogs when awake and during anesthesia with medetomidine; xylazine, ketamine, and halothane (XKH) combination; or propofol. ANIMALS: 10 adult female Beagles. PROCEDURES: At intervals of 15 days, dogs were administered medetomidine (0.05 mg/kg, IV); XKH combination (xylazine [1 mg/kg, IV], ketamine [5 mg/kg, IV], and halothane [1% end-tidal concentration]); or propofol (6 mg/kg, IV) to induce anesthesia or no treatment. Glomerular filtration rate was assessed on the basis of renal uptake (RU; determined via renal scintigraphy) and plasma clearance (CL) of technetium 99m-labeled diethylenetriamine pentaacetic acid ((99m)Tc-DTPA). RESULTS: In awake dogs, mean +/- SEM RU was 9.7 +/- 0.4% and CL was 3.86 +/- 0.23 mL/min/ kg. Renal uptake and CL of (99m)Tc-DTPA were not significantly modified by administration of XKH (RU, 11.4 +/- 0.9%; CL, 4.6 +/- 0.32 mL/min/kg) or propofol (RU, 9.7 +/- 0.3%; CL, 3.78 +/- 0.37 mL/min/kg). Half-life elimination time of plasma (99m)Tc-DTPA decreased significantly in XKH-anesthetized dogs, compared with the value in awake dogs (14.4 minutes and 28.9 minutes, respectively). However, glomerular filtration rate was significantly decreased by administration of medetomidine (RU, 3.9 +/- 0.1%), and the time to maximum kidney activity was significantly increased (867 +/- 56 seconds vs 181 +/- 11 seconds without anesthesia). CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that anesthesia with propofol or an XKH combination did not alter renal function in healthy Beagles, but anesthesia with medetomidine decreased early RU of (99m)Tc-DTPA.