Lack of correlation between growth, stress, and virulence phenotypes in strains of Salmonella enterica serovar Enteritidis, S. Typhimurium DT104, S. 4,12, b:- and S. Liverpool.

Strains of Salmonella Enteritidis (SEnt, n = 10) and S. Typhimurium (STm, n = 11), representing clones with high impact on human health, and strains of S. 4,12: b:- (S412B n = 11) and S. Liverpool (SLiv, n = 4), representing clones with minor impact on human health were characterized for 16 growth, stress, and virulence phenotypes to investigate whether systematic differences exist in their performance in these phenotypes and whether there was correlation between performance in different phenotypes. The term serotype was not found to be predictive of a certain type of performance in any phenotype, and surprisingly, on average, strains of SEnt and STm were not significantly better in adhering to and invading cultured intestinal cells than the less pathogenic types. Forest analysis identified desiccation tolerance and the ability to grow at 42°C with high salt as the characters that separated serovars with low human health impact (S412B/SLiv) from serovars with high human health impact (SEnt/STm). The study showed that variation in phenotypes was high even within serovars and correlation between phenotypes was low, i.e. the way that a strain performed phenotypically in one of the tested conditions had a low predictive value for the performance of the strain in other conditions.

Development and survival of Ascaris suum eggs in deep litter of pigs.

SUMMARY Indoor transmission of Ascaris suum partly depends on the physico-chemical conditions in bedding material. Temperature, pH, aqueous ammonia, moisture, occurrence and development of A. suum eggs were therefore compared in different areas (resting, intermediate and latrine) of two deep litter pens on an organic farm in four seasons. There was some variation, but mean ammonia levels were generally very low (1·0-2·6 mm) and pH levels were moderate (8·04-8·88) in all three areas. Relatively, resting areas were characterized by overall moderate moisture (36%) and moderately high temperature (35·7 °C) levels. The area contained few eggs (50 eggs g-1 DM) of which 17% were viable, and though only 4% were larvated and 0·7% appeared infective, it was more than in the other areas. Intermediate areas had moderate moisture (43%) and high temperature (43·6 °C) levels. There were many eggs (523 eggs g-1 DM), but overall viability was very low (5%) and few eggs were larvated (0·004%) or even infective (0·002%). Latrines typically had high moisture (79%) and moderate temperature (30 °C) levels. The concentration of eggs was very high (1444 egg g-1 DM) and though 32% were viable, none had developed larval stages. The large majority of A. suum eggs appear to die and only few become infective while in the deep litter. However, a large fraction of eggs may remain viable for some time and could thus contaminate agricultural land and develop to infectivity, if the manure is not composted appropriately.

Pathogenicity of wild-type and small-colony variants of Streptococcus equi subsp. zooepidemicus in layer chickens.

Wild-type (WT) and small-colony-variant (SCV) strains of Streptococcus equi subsp. zooepidemicus have recently been isolated from a layer flock in Denmark experiencing high mortality. To investigate the disease-causing potential of SCV compared with WT, a 2-week long infection study was performed in 45-week-old brown layer chickens. Four groups of 11 chickens each were inoculated with a WT or SCV strain by the intravenous or intra-tracheal route: WT-IV, SCV-IV or WT-IT, SCV-IT, respectively. Clinical signs were observed in most chickens in the WT-IV group (9/11). Mortality was observed in the SCV-IV (4/11) and WT-IV (2/11) groups. Ten chickens in the WT-IV and WT-IT groups, respectively, developed gross lesions including oophoritis/peritonitis, hepatitis and airsacculitis cervicalis. Bronchopneumonia was common in the SCV-IT group (6/11), and valvular endocarditis in the SCV-IV group (4/11). Histological lesions in liver tissue were frequently observed in the chickens of the SCV-IV group (9/11), followed by the WT-IT (7/11), WT-IV (6/11), and SCV-IT (2/11) groups. The lesions in the SCV-IV group were dominated by deposition of eosinophilic material with infiltration of inflammatory cells (6/9). Bacteriological re-isolation of either strain type was achieved from all chickens of the WT-IV and WT-IT groups, and from nine and seven out of 11 chickens for each of the SCV-IV and SCV-IT groups, respectively. In summary, we were able to reproduce clinical signs and lesions as observed during the natural outbreak, which included an overall initial onset in WT-infected chickens as opposed to a late onset and possible recurring infection seen in the SCV-infected chickens.

Population dynamics of Ascaridia galli following single infection in young chickens.

The population dynamics of Ascaridia galli was studied in 70 ISA Brown layer pullets, 42 of them were each experimentally infected with 500 embryonated A. galli eggs and 28 chickens were kept as uninfected controls. Six chickens from the infected group and 4 from the control group were necropsied at 3, 7, 10, 14, 21, 28 and 42 days post-infection (d.p.i.). The mean worm recovery varied from 11-20% of the infection dose with the highest recovery at 3 d.p.i. and the lowest at 21 and 42 d.p.i. (P < 0·05). More larvae were recovered from the intestinal wall than from the content (P < 0·0001) and intestinal content larvae were longer than those from the wall (mean length 1·6 and 1 mm, respectively, P < 0·0001). Although larvae were growing over time, a population of small-sized larvae (length < 1 mm) was recovered at all d.p.i. During the first week of infection most of the larvae were located in the anterior half of the jejunoileum but they moved posteriorly with the age of infection. Thus, a subpopulation of larvae mainly in the lumen grew with time while another subpopulation remained small and associated with the mucosa. During the infection both subpopulations moved to a more posterior localization in the gastrointestinal (GI) tract.

Environmental and public health related risk of veterinary zinc in pig production - Using Denmark as an example.

At great economic cost, important steps have been taken over the last many decades to reduce and control emissions of heavy metals in order to protect the environment and public health. Monitoring has confirmed the success of these policies with progressive declines of heavy metals in for example air, sewage sludge and environmental samples. For zinc, such improvements may nevertheless be counter-acted by its widely usage as a feed additive and veterinary medicinal product to piglets in the post-weaning period resulting in reduced occurrence of diarrhea and improvement of daily weight gain. This review therefore focuses on two major concerns associated with veterinary use of zinc, namely the quantifiable risks to the environment and promotion of (multi) resistant bacteria like LA-MRSA in pig farms. Denmark is used as an informed and realistic worst-case scenario, representing the largest pig production per capita in Europe. It is furthermore, one of the countries where most recent information can be found regarding soil monitoring data and zinc consumption within the pig production. An average increase in soil concentration by >45% was recently reported within the period 1998-2014. In order to predict future risk, this review presents new and simplified model predictions using current soil concentrations, annual load rates and predicted accumulation rates. In conclusion, it is estimated that within 25 years, continued agricultural practice of current zinc loads may result in a situation where almost all soils receiving manure from intensive piglet production may be at risk, but also other pig production types may result in scenarios with predicted risk to soil dwelling species, especially in sandy soils. Besides the quantifiable risks to soil ecosystems, high levels of zinc furthermore co-select for the persistence of LA-MRSA CC398 and other resistant bacteria on pig farms.

Do antimicrobial mass medications work? A systematic review and meta-analysis of randomised clinical trials investigating antimicrobial prophylaxis or metaphylaxis against naturally occurring bovine respiratory disease.

A distinct difference between veterinary and human medicine is the routine use of antimicrobial mass medications (prophylaxis, metaphylaxis) to healthy individuals. The need for antimicrobial mass medications is based on beliefs that group/s of animals will contract a bacterial disease (i.e. morbidity) and/or die (i.e. mortality). Bovine respiratory disease (BRD) represents the major indication for cattle antimicrobials worldwide. The objectives were to perform a systematic review and meta-analysis of randomised controlled clinical trials (RCTs) for naturally occurring BRD investigating antimicrobial prophylaxis/metaphylaxis to prevent morbidity/mortality. In total, 58 publications met the inclusion criteria summarizing 169 individual RCTs, spanning 50 years (1966-2016). Antimicrobial prophylaxis and metaphylaxis demonstrated moderate, yet highly variable relative risk reductions in BRD morbidity. These were dependent on the antimicrobial classes used, dependent on metaphylaxis definition, BRD attack rates and duration of the RCTs. Best relative risk reductions were from broad-spectrum critically important antimicrobials, or combinations. BRD prophylaxis/metaphylaxis represents major antimicrobial consumption for highly variable short-term gains in absolute risk reduction of morbidity/mortality. Despite widespread use of prevention products, the need for antimicrobial mass medications should be re-evaluated since the underlying problem is more likely the segmented infrastructure of the feedlot and veal calf industries compared to the disease itself.

The use of genetically marked infection cohorts to study changes in establishment rates during the time course of a repeated Ascaridia galli infection in chickens.

This study investigated the changes in establishment rates during the time course of a 6 week trickle infection of chickens with Ascaridia galli at two different dose levels, using a molecular marker. To differentiate early and late infection, two different egg cohorts (haplotype a and haplotype b, genetically identified using PCR-linked restriction fragment length polymorphism on the cox1 gene of the mitochondrial DNA) were used. Cohort-specific egg batches were produced by harvesting eggs from the uteri of female worms of the specific cohort. Fifty-six 8 week old Lohmann Brown Lite chickens were divided into seven groups and the infectivity of the egg batches was compared between two groups of chickens (P=0.6). The remaining chickens were allocated to four infection regimes and one control group. Group ab100 was trickle infected for 3 weeks with 100 eggs of haplotype a (twice weekly) followed by the same dose of eggs of haplotype b for another 3 weeks. Group ba100 was treated similarly but in the opposite order (haplotype b preceding a). A similar infection regime was applied for groups ab25 and ba25 but with a lower inoculation dose (25 eggs). All of the birds in these five groups (four infected and one control) were euthanased 2 weeks after the last inoculation. It was found that in the low-dose groups both the early and late infections established equally well, whereas in the high-dose groups the early infection was recovered in a significantly (P<0.001) higher proportion of chickens than the late infection, irrespective of genetic cohorts. Moreover, relatively higher proportions of the larvae from both the early and late infections were found in the posterior section of the small intestine. This result indicates the presence of dose-dependent resistance against reinfection and this resistance seems to act by reducing the establishment of late infection and by relocating the larvae from early infection.

Survival of Ascaris suum and Ascaridia galli eggs in liquid manure at different ammonia concentrations and temperatures.

Eggs of Ascaris suum from pigs are highly resistant and commonly used as a conservative indicator of pathogen inactivation during slurry storage. Eggs of Ascaridia galli, the poultry ascarid, are also known to be highly resistant but the suitability as an indicator of pathogen inactivation has never been tested. Pig slurry has to be stored for several months to inactivate pathogens but chemical treatment of slurry may reduce this time. The suitability of A. galli as an indicator of slurry sanitation was tested by comparing the survival of eggs of A. suum and A. galli in pig slurry. In addition, the effect of urea treatment on inactivation of ascarid eggs in relation to storage time was also tested. Nylon bags with 10,000 eggs of either species were placed in 200 ml plastic bottles containing either urea-treated (2%) or untreated pig slurry for up to 120 days at 20°C, 6 days at 30°C, 36h at 40°C or 2h at 50°C. At all the temperatures in both slurry types, A. galli eggs were inactivated at a significantly faster rate (P<0.05) compared to A. suum eggs. For each 10°C raise in temperature from 20°C, T50 (time needed to inactivate 50% of eggs) for both types of eggs was reduced markedly. At all temperatures, viability of eggs of both species was significantly higher (P<0.05) in untreated slurry compared to urea-treated slurry except A. galli eggs at 20°C where no significant difference was detected. In untreated slurry, the levels of pH (6.33-9.08) and ammonia (0.01-1.74 mM) were lower (P<0.0001) compared to that of urea-treated slurry (pH: 8.33-9.28 and ammonia 1-13 mM). The study demonstrated that A. galli eggs are more sensitive to unfavourable conditions compared to A. suum eggs. The use of A. galli eggs as hygiene indicator may thus be suitable to assess inactivation of pathogens that are more sensitive than A. galli eggs. Addition of urea may markedly reduce the storage time of slurry needed to inactivate A. suum and A. galli eggs.

Localization of Ascaridia galli larvae in the jejunum of chickens 3 days post infection.

The normal habitat of the parasitic stages of Ascaridia galli is in the small intestine of poultry but the exact localization is poorly understood. Therefore, a histological study was conducted in order to localize the larvae during the early phase of infection. Six layer pullets seven-week old were infected orally with 20,000 embryonated A. galli eggs each, whereas four chickens were left as un-infected controls. At necropsy 3 days after infection the first half of jejunum/ileum was divided into two equally sized sections (J1 and J2). After taking samples for histology from the middle of J1 and J2 and the junction between these determined JX, the two sections were subjected to parasitological examination. A higher number of A. galli larvae were recovered from section J2 than J1 and the majority of larvae were recovered from the most profound layers. Based on histology 144 larvae were identified and their location was noted. The highest number of larvae was observed in the JX sample as compared to J1 and J2 (P<0.001). Most of them were located in the profound crypt zone of the mucosa (51%) as compared to the other zones (P<0.05). The number of larvae was higher in the lumen (63%) compared to the epithelium (32%) and lamina propria (5%) (P<0.001). A significantly higher number of eosinophils were found in lamina propria of the infected group compared to the control group (P<0.001). This experiment clearly showed that only few larvae had penetrated the epithelium and were positioned in the lamina propria at 3 days post infection. It was far more common that the larvae were localized within the epithelium or in the lumen of the crypts. It is therefore suggested that at least in this early phase "mucosal phase" is a more appropriate term to be used for the A. galli larval localization as compared to the term "histotrophic phase" currently used in many textbooks.