Development of a capture ELISA for the detection of antibodies to enteropathogenic Escherichia coli (EPEC) in rabbit flocks using intimin-specific monoclonal antibodies

A capture enzyme-linked immunosorbent assay (cELISA) was developed using intimin-specific monoclonal antibodies to detect specific antibody in rabbits that have been in contact with enteropathogenic Escherichia coli (EPEC). Sera from 121 EPEC-negative, minimum-disease-level (MDL) rabbits were used for negative controls, and sera from 25 MDL rabbits, experimentally infected with EPEC of bio-/serotype 3-/O15, for positive controls. These were used to determine a cut-off value for a positive cELISA result. The value selected gave the test a sensitivity of 80.0% and a specificity of 98.4% on an individual level. At this value, a flock level sensitivity and specificity of 79.2 and 85.2%, respectively were calculated for a flock with a prevalence of seven per cent, if 40 animals were tested, and a minimum of two reactors were obtained. The test characteristics improve with increasing prevalence. To evaluate the diagnostic potential of the cELISA, sera from 40 to 50 slaughter rabbits per flock from 25 rabbit flocks with bacteriologically determined EPEC status were tested. The results demonstrated that this test can be a useful tool to determine the EPEC status of a rabbitry, provided that it is used at regular intervals.     

The diversity of Mycoplasma hyopneumoniae within and between herds using pulsed-field gel electrophoresis

Over the years, pulsed-field gel electrophoresis (PFGE) has been proven a robust technique to type isolates with a high resolution and a good reproducibility. In this study, a PFGE protocol is described for the typing of Mycoplasma hyopneumoniae isolates. The potential of this technique was demonstrated by comparing M. hyopneumoniae isolates obtained from the same as well as from different herds. The use of two different restriction enzymes, SalI and ApaI, was evaluated. For each enzyme, the resulting restriction profiles were clustered using the unweighted pair group method with arithmetic means (UPGMA). For both obtained dendrograms, the included isolates of the related M. flocculare species clustered separately from all M. hyopneumoniae isolates, forming the root of the dendrograms. The PFGE patterns of the M. hyopneumoniae isolates of different herds were highly diverse and clustered differently in both dendrograms, illustrated by a Pearson's correlation coefficient of only 0.33. A much higher similarity was observed with isolates originating from different pigs of a same herd. The PFGE patterns of these isolates always clustered according to their herd and this for both dendrograms. In conclusion, the results indicate a closer relationship of M. hyopneumoniae isolates within a herd compared to isolates from different herds and this for both restriction enzymes used. Since the described PFGE technique was shown to be highly discriminative and reproducible, it will be a helpful tool to further elucidate the epidemiology of M. hyopneumoniae.     

Effect of supplemental tryptophan, vitamin E, and a herbal product on responses by pigs to vibration

Economic losses related to increased stress during the transport of pigs are well documented. The effects of supplementing of tryptophan (Trp), vitamin E, or a herbal product via feed or drinking water were investigated in terms of effects on stress response in pigs during transport simulation. The study consisted of three analogous experiments. For the testing in each experiment, the pigs (23.5+/-3.2 kg) were allocated to one of two treatments, with and without supplementation of a product. The applied doses were Trp (5 g/L drinking water for 3 d), vitamin E (additional amount of 300 mg/kg feed for 21 d, as-fed basis), and Sedafit (2.5 g/L drinking water for 2 d). Sedafit is a commercial herbal product containing Valeriana officinalis L. and Passiflora incarnata L. as active components. In each experiment of the study, at least 47 pigs were involved, which were treated in groups of 3. The day before transport simulation, a Holter device was attached to the pigs to produce an electrocardiogram during the night (rest values), as well as during vibration in the transport simulator (1.2 Hz, 1 m/s2), where the behavior of the pigs (standing-sitting-lying) was also observed. Samples of saliva (taken before, during, and after [3x] vibration) and blood (taken before and after vibration) were analyzed for cortisol and intermediate metabolites (glucose, lactate, creatine kinase, and nonesterified fatty acids), respectively. Pigs supplemented with Trp tended to spend more time lying down during the second hour of vibration (P < 0.05). Vitamin E decreased the peak heart rate (P < 0.05), ventricular ectopic beats (P < 0.01), and ST elevation (P < 0.10). The supplementation of Sedafit resulted in smaller increases of the investigated heart variables (minimum heart rate, P < 0.05; ventricular ectopic beats, P < 0.05; ST elevation, P < 0.01) during and after stress evocation compared with the control group. None of the tested products influenced the intermediate metabolites; one possible explanation for this finding may be that peak values were reached before the time of bleeding. In conclusion, Trp had a positive behavioral effect in this experiment, and vitamin E and Sedafit mediated an increase in some heart variables, suggesting sedative and antianxiety effects.     

Lingual abscesses in three dogs

Lingual abscessation is a rare condition in dogs. Very little information is available on the diagnosis and treatment of lingual abscesses in the major surgical textbooks and current veterinary literature. The common clinical signs of lingual abscesses are macroglossia, hypersalivation and a reluctance to open the mouth, but these can vary depending on the time course of the disease and the location of the abscess. This article presents three cases of tongue abscess in the dog outlining treatment and outcomes. A thorough diagnostic work up, consisting of anamnesis, clinical and haematological examinations, oral inspection under sedation and the use of diagnostic imaging techniques should be mandatory before surgical exploration of the abscess. Surgery is followed by drainage and systemic antibiotics, complemented by systemic fluid support and pain management. Conservative management of lingual abscesses can be fatal. Sharp trauma from an unknown object is suspected to be the underlying cause for the abscesses in the present cases.     

Whole blood and tissue fungal DNA quantification in the diagnosis of canine sino-nasal aspergillosis

Various combinations of tests are used to confirm the diagnosis of canine sino-nasal aspergillosis (SNA) because false-positive and false-negative results can occur with each test. Therefore, the aim of this study was to evaluate whether detection of fungal DNA in blood and nasal tissue samples was of value in the clinical diagnosis of this disease. Four groups were included in the study (dogs with SNA, lymphoplasmacytic rhinitis or nasal neoplasia, and control animals). Real-time PCR assays detecting DNA from all Penicillium and Aspergillus species (PenAsp assay) or species-specific DNA from A. fumigatus, A. terreus, A. flavus and A. niger were applied to whole blood and nasal tissue samples. Results obtained by PCR were compared between the groups. Sensitivity, specificity, positive and negative predictive values (PPV and NPV) for fungal DNA detection were compared with those for alternative diagnostic procedures including histopathology, serology and fungal culture. Significantly more fungal DNA was detected by the PenAsp assay in tissue biopsies from dogs with SNA than in the three other groups. Sensitivity, specificity, PPV and NPV for this method were 1.00, 0.06, 0.32 and 1.00. A. fumigatus DNA was detected in seven tissue biopsies from dogs with SNA and in one biopsy from a dog with a nasal tumour. Sensitivity, specificity, PPV and NPV for this diagnostic test were 0.50, 0.97, 0.87 and 0.82. No significant difference was found between the groups with respect to the amount of DNA detected in blood by the PenAsp assay. Sensitivity, specificity, PPV and NPV for this method were 0.71, 0.24, 0.31 and 0.64. A. fumigatus DNA was detected in the blood of three dogs with SNA and sixteen dogs without SNA. Sensitivity, specificity, PPV and NPV for this diagnostic tool were 0.21, 0.45, 0.15 and 0.54. Detection of A. fumigatus DNA in nasal tissue had the highest specificity, PPV and NPV but sensitivity of this method was low. Detection of fungal DNA in whole blood was of no value in the diagnosis of SNA.     

Understanding Ebola virus and other zoonotic transmission risks through human–bat contacts: Exploratory study on knowledge,attitudes and practices in Southern Cameroon

The ecology of Ebola virus (EBV) remains largely unknown, but the previous detection of viral RNA and anti‐EBV antibodies in African bats suggests that they might play a role in the EBV reservoir. Moreover, African bats also carry other potentially zoonotic agents such as Henipah‐like viruses, coronaviruses and lyssaviruses. Today only little information is available on interactions between humans and bats. The objective of our exploratory study was to describe the extent and modes of contacts between humans and bats in southern Cameroon, considered as an area at risk for future EBV outbreaks. The survey was conducted in 11 villages of four distinct rural areas in southern Cameroon. A total of 135 respondents were interviewed using semi‐structured questionnaires, between February and May 2017. The study showed that direct contacts between bats and humans are relatively common. Bat bushmeat appeared to be an occasional meat resource; 40% of respondents consume bats with a median annual consumption of three, and 28% of respondents hunt them. About 22% of the respondents reported children catching bats. Indirect contact also appeared to be common; 55% of hunters use caves as shelters and 67% of interviewees eat fruits previously chewed by bats. Bat consumption varied significantly between regions (from 0% to 87%) and between pygmies and bantus in the extreme south‐east of Cameroon. The study revealed considerable diversity in practices among interviewees, most of them being subsistence cultivators and relying on self‐hunted bushmeat. Geographical diversity of contacts and perceptions regarding bats in Cameroon emphasizes the need to adjust zoonotic pathogen surveillance and education campaigns to the specificities of the communities and their context of interaction with wildlife.     

Evaluation of the cold chain of fresh-cut endive from farmer to plate

In this study, the cold chain and its impact on food stuff microbiology was evaluated for fresh-cut endive. A survey was carried out to analyze endive temperature throughout the supply chain from producer, via processor and distributor to the restaurant. Data loggers accompanying the endive on its route provided a temperature profile of the endive. The effect of the outdoor temperature on initial cooling was evaluated and critical points regarding cold chain management in the supply chain were identified. Our experiments indicate that the cold chain is generally properly maintained. In parallel with temperature monitoring, indicator microorganisms were assessed at different points in the supply chain to examine the effects of endive temperature, temperature fluctuations, and the outdoor temperature on microbial food safety. Small temperature fluctuations in the supply chain had a small effect on the total level of aerobic mesophilic bacteria. However, at the best before-date, total coliforms and Enterobacteriaceae levels were significantly higher in endive samples subject to temperature fluctuations in the supply chain, compared to endive stored in optimal conditions. To our knowledge, this is the first study that combines microbiological analysis with the temperature profile of fresh-cut produce in a realistic food supply chain.     

A multiplex PCR to identify porcine mycoplasmas present in broth cultures

Mycoplasma hyopneumoniae, Mycoplasma hyorhinis and Mycoplasma flocculare can be present in the lungs of pigs at the same time. These three mycoplasma species all require similar growth conditions and can be recovered from clinical samples using the same media. We have developed a multiplex PCR as a helpful tool for rapid differentiation of these three species in the course of isolation. Based on the 16S ribosomal DNA sequences, three different forward primers and a single reverse primer were selected. Each forward primer was compared to available mycoplasma sequences, showing the primers to be specific. The three amplification products observed of 1129 bp (M. hyorhinis), 1000 bp (M. hyopneumoniae) and 754 bp (M. flocculare) were clearly distinguishable on a 1% agarose gel. In addition, no cross-reaction with Mycoplasma hyosynoviae, another porcine mycoplasma, was noted. This multiplex PCR using the proposed set of primers is the first reported assay that allows the simultaneous identification of the different Mycoplasma species isolated from the lungs of pigs.