Leo van Leengoed

Clostridium difficile PCR ribotype 078 toxinotype V found in diarrhoeal pigs identical to isolates from affected humans

In diseased piglets from two Dutch pig-breeding farms with neonatal diarrhoea for more than a year, culture and PCR analyses identified the involved microorganism as Clostridium difficile PCR ribotype 078 harbouring toxin A (tcdA) and B (tcdB), and binary toxin genes. Isolated strains showed a 39 bp deletion in the tcdC gene and they were ermB gene-negative. A number of 11 porcine and 21 human isolated C. difficile PCR ribotype 078 toxinotype V strains were found genetically related by multiple-locus variable-number tandem-repeat analysis (MLVA). Moreover, a clonal complex was identified, containing both porcine and human isolates. The porcine isolates showed an antimicrobial susceptibility profile overlapping that of isolates from Dutch human patients. On the basis of these pheno- and genotypical analyses results, it was concluded that the strains from affected piglets were indistinguishable from increasingly encountered C. difficile PCR ribotype 078 strains of human C. difficile infections in the Dutch population and that a common origin of animal and humans strains should be considered.

Clostridium difficile PCR Ribotype 078: an Emerging Strain in Humans and in Pigs?

In a recent paper, Keel and colleagues concluded that Clostridium difficile PCR ribotype 078 was the most common PCR ribotype among isolates from swine (83% of 119 isolates) and isolates from calves (94% of 33 isolates) in The United States ([1][1]). In contrast, only 1 of 23 human isolates

In Vivo Buccal Delivery of the Peptide Drug Buserelin with Glycodeoxycholate as an Absorption Enhancer in Pigs

AbstractPurpose. To study the potential of buccal delivery of the peptide drug in pigs. Methods. Intravenous administration and buccal delivery without and with 10 mM sodium glycodeoxycholate (GDC) as absorption enhancer were investigated as a randomised cross-over study in six pigs. The buccal delivery device consisted of an application chamber with a solution of buserelin and was attached to the buccal mucosa for 4 hours using an adhesive patch. Results. Buccal administration of buserelin resulted in rapidly reached steady state plasma levels. The absolute bioavailability of the peptide after buccal delivery for 4 hours could be increased from 1.0 ± 0.3 to 5.3 ± 1.1% (mean ± S.D.) by co-administration of 10 mM GDC (0.45% w/v)). Conclusions. The results of this study demonstrate that buccal administration with the use of absorption enhancers is a useful approach for the delivery of peptide drugs such as buserelin.

Induction of protective immunity by aerosol or oral application of candidate vaccines in a dose-controlled pig aerosol infection model.

In order to outline basic concepts for the design of a bacterial aerosol infection model, the development of a pig model with Actinobacillus pleuropneumoniae is described. First, reproducibility of aerosol parameters should be maintained by optimizing generating and sampling conditions. Survival rates of the chosen strain must be predictable. Secondly, inhalation conditions for the recipients have to be standardized to enable the determination of deposition sites and the dose administered. Subsequently, dose-response relationship should be evaluated to find a suitable challenge dose. Furthermore, it seems necessary to establish methods to obtain local specimens for determination of the local immune responses. The present study demonstrates that after aerosol challenge pigs were completely protected after inhalation and partially protected after oral application of A. pleuropneumoniae vaccines and describes techniques to administer bacteria in a dose-dependent, viable way. Using the infection model several stages of the disease from acute pleuropneumonia to chronic infection can be induced for research purposes.

Atypical Porcine Pestivirus: A Possible Cause of Congenital Tremor Type A‐II in Newborn Piglets

Congenital tremor type A-II in piglets has been regarded as a transmissible disease since the 1970s, possibly caused by a very recently-described virus: atypical porcine pestivirus (APPV). Here, we describe several strains of APPV in piglets with clinical signs of congenital tremor (10 of 10 farms tested). Piglets on a farm with no history of congenital tremor were PCR-negative for the virus. To demonstrate a causal relationship between APPV and disease, three gilts were inoculated via intramuscular injection at day 32 of pregnancy. In two of the three litters, vertical transmission of the virus occurred. Clinical signs of congenital tremor were observed in APPV-infected newborns, yet also two asymptomatic carriers were among the offspring. Piglets of one litter were PCR-negative for the virus, and these piglets were all without congenital tremors. Long-term follow up of farm piglets born with congenital tremors showed that the initially high viremia in serum declines at five months of age, but shedding of the virus in feces continues, which explains why the virus remains present at affected farms and causes new outbreaks. We conclude that trans-placental transmission of APPV and subsequent infection of the fetuses is a very likely cause of congenital tremor type A-II in piglets.

Pleuritis in slaughter pigs: relations between lung lesions and bacteriology in 10 herds with high pleuritis.

Pleuritis in slaughter pigs has increased in recent years in the Netherlands. The aim of the present study was to determine what respiratory pathogens were involved in pleuritis. In total, lungs of 968 slaughter pigs from 10 herds with high prevalence of pleuritis were morphologically examined for size, location, and type of lesions. Moreover, histology and bacteriology were performed. Examination of gross lung lesions showed 45% pleuritis, 14% pleuropneumonia and 38% catarrhal pneumonia. Peribronchiolar cuffing was found in 61 of 142 samples. Actinobacillus pleuropneumoniae was cultured from 22 lung samples from four herds. Pasteurella multocida was cultured from 55 lung samples in eight herds. No specific pattern with respect to the causal pathogens was found. In conclusion, no single infectious cause of pleuritis was found. A variety of infectious agents combined with environmental factors should be considered as a cause of pleuritis.

Differential effects of pentoxifylline on the hepatic inflammatory response in porcine liver cell cultures. Increase in inducible nitric oxide synthase expression

Pentoxifylline (PTX) has been shown to exert hepatoprotective effects in various liver injury models. However, little information is available about the effect of PTX on the hepatic acute phase response. In the present study, the effect of PTX on a lipopolysaccharide (LPS)-induced acute phase response in primary porcine liver cell cultures was examined. During 72 hr of incubation with or without LPS, the ability of PTX to influence the secretion of tumour necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), acute phase proteins, and nitric oxide (NO) was assessed. PTX completely inhibited LPS-induced TNF-alpha production and attenuated IL-6 only after 48 hr of incubation. In contrast, PTX potentiated NO production and the expression of inducible nitric oxide synthase (iNOS) in hepatocytes after stimulation with LPS. The increased expression of iNOS and concurrent production of NO was also observed when liver cell cultures were incubated with dibutyryl cyclic adenosine monophosphate. No effect of PTX on acute phase protein secretion was observed during 72 hr of incubation. The present results show that PTX differentially affects the endotoxin-induced inflammatory response in primary porcine liver cell cultures by suppressing TNF-alpha and IL-6 while potentiating NO production.

Effect of Spatial Separation of Pigs on Spread of Streptococcus suis Serotype 9

The spread of an infectious agent in a population can be reduced by interfering in the infectiousness or susceptibility of individuals, and/or in their contact structure. The aim of this study was to quantify the effect of prevention of direct contact between infectious and susceptible pigs on the transmission of Streptococcus suis (S. suis). In three replicate experiments, S. suis-free pigs were housed in boxes either in pairs (25 pairs) or alone (15 pigs). The distance between the boxes was ±1 m. At 7 weeks of age, one pig of each pair was inoculated intranasally with S. suis serotype 9; the other pigs were exposed to S. suis by either direct (pairs) or indirect contact (individually housed pigs). Tonsillar brush and saliva swab samples from all pigs were collected regularly for 4 weeks post inoculation to monitor colonization with S. suis. All inoculated pigs became infected, and their pen mates became colonized within 2 days. Thirteen indirectly exposed pigs became positive within 7–25 days after exposure. The rate of direct transmission βdir was estimated to be 3.58 per pig per day (95% CI: 2.29–5.60). The rate of indirect transmission increased in time, depending on the cumulative number of days pigs tested positive for the presence of S. suis. The estimate β’ind was 0.001 (95% CI: 0.0006–0.0017) new infections per pig per day for each day that an infected pig was tested positive for S. suis. We conclude that prevention of direct contact reduces the rate at which susceptible pigs become colonized. Simulation studies using these parameters showed, however, that such intervention measure would not limit S. suis serotype 9 spread in a commercial pig farm to a relevant extent, implying that spatial separation of groups op pigs within a compartment would not be effective on a farm.

Effect of Simultaneous Exposure of Pigs to Streptococcus suis Serotypes 2 and 9 on Their Colonization and Transmission, and on Mortality

The distribution of Streptococcus suis serotypes isolated from clinically infected pigs differs between geographical areas, and varies over time. In several European countries, predomination of serotype 2 has changed to serotype 9. We hypothesize a relation, with one serotype affecting the other in colonization and invasion. The aim of this study was to evaluate whether simultaneous exposure of pigs to serotypes 2 and 9 affects colonization and transmission of each type, and mortality. Thirty-six caesarean-derived/colostrum-deprived piglets were randomly assigned to three groups, and there housed pair-wise. At six weeks old, one pig per pair was inoculated with either one (serotype 2 or 9; mono-group) or two serotypes simultaneously (dual-group); the other pig was contact-exposed. Tonsillar and nasal samples were collected within three weeks post inoculation. Bacterial loads in samples were quantified using multiplex real-time polymerase chain reaction (PCR). Transmission rates of the serotypes among pigs were estimated using a mathematical Susceptible-Infectious (SI) model. Bacterial loads and transmission rates did not differ significantly between serotypes. Compared to the mono-group, in the dual-group the average serotype 2 load in tonsillar samples from contact pigs was reduced on days 1 to 4 and on day 6. Simultaneous exposure to the serotypes reduced the mortality hazard 6.3 times (95% C.I.: 2.0–19.8) compared to exposure to serotype 2 only, and increased it 6.6 times (95% C.I.: 1.4–30.9) compared to exposure to serotype 9 only. This study indicates that serotype 2 load and mortality were affected in pigs exposed to these two serotypes.