S. K. Kritas

Occurrence, virulence genes and antibiotic resistance of Escherichia coli O157 isolated from raw bovine, caprine and ovine milk in Greece

The examination of 2005 raw bovine (n = 950), caprine (n = 460) and ovine (n = 595) bulk milk samples collected throughout several regions in Greece for the presence of Escherichia coli serogroup O157 resulted in the isolation of 29 strains (1.4%) of which 21 were isolated from bovine (2.2%), 3 from caprine (0.7%) and 5 from ovine (0.8%) milk. Out of the 29 E. coli O157 isolates, only 12 (41.4%) could be classified as Shiga-toxigenic based on immunoassay and PCR results. All 12 Shiga-toxigenic E. coli serogroup O157 isolates belonged to the E. coli O157:H7 serotype. All except one of the 12 Shiga-toxin positive isolates were stx(2)-positive, five of which were also stx(1)-positive. The remaining isolate was positive only for the stx(1) gene. All stx-positive isolates (whether positive for stx(1), stx(2) or stx(1) and stx(2)) were also PCR-positive for the eae and ehxA genes. The remaining 17 E. coli O157 isolates (58.6%) were negative for the presence of the H7 flagellar gene by PCR, tested negative for Shiga-toxin production both by immunoassay and PCR, and among these, only four and three strains were PCR-positive for the eae and ehxA genes, respectively. All 29 E. coli O157 isolates displayed resistance to a wide range of antimicrobials, with the stx-positive isolates being, on average, resistant to a higher number of antibiotics than those which were stx-negative.

Relationships between tail biting in pigs and disease lesions and condemnations at slaughter

Two matched case-control studies were performed at an abattoir with a capacity of 780 pigs per hour, each study using the approximately 7000 pigs slaughtered on one day. In the first study, the severity of tail biting and pneumonia were recorded in pigs with bitten or intact tails. In the second study, the tail score, sex, and the presence of pleuritis, externally visible abscesses and trimming were recorded in pigs with bitten or intact tails. In study 1, there was no significant association between the tail score and the percentage of lung tissue affected by lesions typical of enzootic pneumonia, but there was a significant association between the severity of tail biting and the prevalence of lungs with abscesses and/or pleuritic lesions (P<0·0001). In study 2, there were significant associations between the severity of tail biting, and the prevalence of external carcase abscesses and carcase trimming; the carcases of castrated males had evidence of tail biting more frequently than the carcases of females (P<0·05).

Caprine PRNP polymorphisms at codons 171, 211, 222 and 240 in a Greek herd and their association with classical scrapie

The association between PRNP variation and scrapie incidence was investigated in a highly affected Greek goat herd. Four mutations were identified at codons 171Q/R, 211R/Q, 222Q/K and 240P/S. Lysine at codon 222 was found to be associated with the protection from natural scrapie (P=0.0111). Glutamine at codon 211 was observed in eight animals, all of them being scrapie-negative, indicating a possible protective role of this polymorphism although statistical analysis failed to support it (P=0.1074). A positive association (P=0.0457) between scrapie-affected goats and the wild-type Q(171)R(211)Q(222)S(240) allele is presented for the first time. In addition, a novel R(171)RQS allele, which is identical to the A(136)R(154)R(171) allele that has been associated with resistance to classical scrapie in sheep, was observed in low frequency. Resistant alleles that include K(222) and Q(211) are absent or rare in sheep and can provide the basis for the development of a feasible breeding programme for scrapie eradication in goats.

Efficacy of moxidectin against sarcoptic mange and effects on milk yield of ewes and growth of lambs.

Forty-eight dairy ewes and 45 lambs naturally infested with sarcoptic mange, were divided into three equal groups and treated with moxidectin 1% inj. sol. at a dose rate of 0.2 mg moxidectin kg(-1) bodyweight once or twice ten days apart. During the study, the skin lesions were scored and the number of mites recovered in facial skin scrapings from the animals were recorded. Moxidectin was found to be effective against ovine sarcoptic mange, when given twice; the percentage reduction of mites was 100% (p < 0.0001) and the skin lesions disappeared completely (p = 0.0001). In sheep treated once only, the percentage reduction of mites varied from 75% to 92%. Treated ewes produced up to 22.4% more milk than the infested ones (p = 0.0001). Treated lambs had up to 15% greater final weight and up to 23.7% more growth than infested controls (p = 0.0001). It is concluded that two injections of moxidectin 1% inj. sol. at a dose rate of 0.2 mg kg(-1) bodyweight, given ten days apart, were effective against ovine sarcoptic mange. Furthermore, it is considered that sarcoptic mange adversely affects the production of the infested animals.

Evaluation of probiotics as a substitute for antibiotics in a large pig nursery

ANTIMICROBIALS, when used as growth promoters, are claimed to improve daily weight gain by 3 to 9 per cent and feed utilisation by 2 to 7 per cent, with fewer scour problems (Visk 1978, Hedde 1984, Doyle 2001). For this reason they are customarily used in pigs even on high-health status farms. They appear to act by reducing the pathogenic bacteria and modifying the microflora in the gut, providing more nutrient availability for the animal itself and less substrate for the bacterial organisms to use for their own growth (Visk 1978, Hedde 1984). Dritz and others (2002) showed that antibiotics are justified for use only in nursery pigs but not in growers and finishers. Despite their beneficial effects, recent concerns regarding the presence of drug residues in edible animal products and the potential transfer of antibiotic resistance to human pathogens has directed research towards alternative solutions such as probiotics. Legislation to phase out the routine feeding to food animals of eight classes of antibiotics as feed additives (penicillin, tetracycline, bacitracin, macrolides, lincomycin, streptogramin, aminoglycoside and sulphonamides) judged to be directly or indirectly of human medical importance has been recently introduced in the USA and has already been applied in the EU. Thus, alternative solutions should always be sought. Probiotics are live cultures of harmless bacteria or yeast species (for example, Lactobacillus, Streptococcus and Saccharomyces species) that equilibrate intestinal microflora, to the benefit of the animal (Fuller 1989, Ferencik and others 2000). They may promote growth by competing with harmful gut flora, and by stimulating the immune system of the animal and therefore increasing its resistance to infectious agents (Tannock 1980, Fuller 1989, Khajarern and Khajarern 1994). Several studies in various animal species have indicated a beneficial role of probiotics against diseases; for example, they may control some Escherichia coli-induced diseases (Khajarern and Khajarern 1994, Zani and others 1998, Kyriakis and others 1999, Genovese and others 2000). Other studies including untreated, antibiotic-free control pigs have shown that the average daily weight gain (ADG) and feed conversion ratio (FCR) are significantly improved after probiotic treatment (Kyriakis and others 1999, 2003, Kritas and others 2000). This short communication describes a study to examine the ability of probiotics to act as a substitute for growthpromoting antibiotics on the health and productivity of weaned pigs on one farm with high-health status. The study was carried out from October 2002 to October 2003, on a commercial, three-site production farm of 1600 sows in Minnesota, USA. All breeding animals were of the same breed and they were vaccinated against Aujeszky’s disease, porcine reproductive and respiratory syndrome, parvovirus infection, erysipelas, leptospirosis and E coli. The piglets on the farm were weaned in weekly batches of (mean [sd]) 520 (50) animals at the age of 20 (2) days and, after separation by sex and bodyweight, were transferred to two adjacent off-site nurseries, where they remained until the age of 64 (3) days. Each nursery contained four small rooms of 16 pens and two large rooms of 32 pens, with approximately 22 pigs per pen. The nursery rooms were filled sequentially one after another, and each small room contained pigs of the same week of age, or of two sequential weeks of age in the large rooms. A common outer corridor connected the rooms in each nursery, and footbaths in the corridor outside each room were used before personnel entered. A separate feed bin supplied each room. The pigs were fed ad libitum rations containing low doses of antibiotics: 400 ppm neomycin (Neomix; Phibro Animal Health) for the first five to seven days postweaning; 100 ppm neomycin plus 100 ppm oxytetracycline (Neo-Terramycin; Phibro Animal Health) for the next seven days, and thereafter 20 ppm tylosin (Tylan 10; Elanco Animal Health) up to the age of 70 days. The programme had been started five years previously to prevent E coli postweaning diarrhoea. Postweaning mortality in the nurseries was historically low (<2 per cent). To test the effects of probiotics compared with subtherapeutic doses of antibiotics, the product BioPlus 2B (Chr Hansen) containing 1012 colony-forming units total Bacillus content (Bacillus licheniformis and Bacillus subtilis) per 0·45 kg of product was used instead of the antibiotics. The US Food and Drug Administration has approved this product for use in feeds, and the Bacillus species strains included do not produce any antibiotic (Anon 2005). The product was included in the usual rations of the nursery period at the standard dose of 0·45 kg/tonne of complete feed. Forty-two nursery rooms were used chronologically but in an alternate mode; for example, the pigs of the first room received feed with probiotics, those of the second room feed with antibiotics, those of the third room feed with probiotics, and so on. At the end of the trial, the two treatment groups, that is, pigs receiving the usual feed that contained antibiotics (21 rooms) and pigs receiving the same type of feed but containing probiotics instead of antibiotics for the same period (21 rooms), were compared. As shown in Table 1, the average age and bodyweight of the pigs between the groups did not differ significantly at the beginning of the trial. Mortality and reasons of death were recorded by the farmer for each group of pigs up to the end of the nursery period. At that time, the pigs were weighed again. The feed consumption per group was also recorded during this period, and the ADG, the average daily feed intake (ADFI) and the FCR were calculated. At least 20 treated and 20 control rooms were needed to detect a 1·50 per cent difference in mortality (α=0·05, power 80 per cent, sd 1·5 per cent). Each parameter was recorded and analysed with ‘room’ as the experimental unit by one-way analysis of variance using the SPSS for Windows statistical package (release 11.0.1; SPSS). The results shown in Table 1 were derived from a total of 10,955 pigs in 21 probiotic rooms and 10,800 pigs in 21 antibiotic rooms. In both treatment groups, the nursery periods had the same average length. The bodyweight of pigs at the end of the nursery period, the ADG, ADFI and FCR were similar for both groups. The cost of feed per pig and per kg of bodyweight was also similar. There was no difference in morVeterinary Record (2005) 156, 447-448

Role of Mast Cells in Atherosclerosis: A Classical Inflammatory Disease

Atherosclerosis is an inflammatory disease and hyperlipidaemia is one of the main risk factors for aging, hypertension and diabetes. Variance in plasma LDL cholesterol concentration may be associated with differences in cardiovascular disease risk and high levels of lipids are associated with increased risk of developing atherosclerosis. Macrophages, which generate pro-inflammatory cytokines, mainly interleukin-1 (IL-1) and tumor necrosis factor-α (TNF-alpha), are deeply involved in atherosclerosis, as well as mast cells which generate several cytokines, including IL-6 and IFN-gamma, and chemokines such as eotaxin, MCP-1 and RANTES involved in monocyte recruitment and differentiation in the arterial wall. In addition, mast cells participate in lipid retention and vascular cell remodeling, and are mediators of innate and adaptive immunity during atherosclerosis. Mast cells which accumulate in the human arterial intima and adventitia during atherosclerotic plaque progression, release vasoactive and angiogenic compounds, and pro-inflammatory mediators, such as arachidonic acid metabolites, histamine, cytokines/chemokines, platelet activating factor (PAF) and proteolytic enzymes. Mast cells can be activated by pro-inflammatory stimuli, including cytokines, hypercholesterolemia, and hyperglycemia, and trigger the endothelial expression of adhesion molecules such as P-selectin, vascular cell adhesion molecule-1 (VCAM-1) and chemokines which mediate the recruitment and adhesion of leukocytes. The participation of mast cells in atherosclerosis is still an enigma and it may be of therapeutic interest to clarify this process.

Effect of salinomycin in the control of Clostridium perfringens type C infection in suckling pigs

The ability of salinomycin to control Clostridium perfringens type C infection in sows and their offspring was examined under field conditions. Two groups of sows and their offspring were offered feed either medicated with 60 ppm salinomycin or free of antibiotics, and their performance was compared. The number of piglets with diarrhoea, the duration of the diarrhoea, and the mortality of the piglets during the lactation period were markedly lower in the group given salinomycin. In addition, laboratory examinations showed that the numbers of carrier piglets and sows were reduced after treatment with salinomycin. Finally, the sows treated with salinomycin lost less weight during the lactation period and weaned more and heavier piglets than the untreated sows. It was concluded that salinomycin incorporated in the diet can be used for controlling C perfringens type C infection in sows and their offspring.

A pathogenic myxoma virus in vaccinated and non-vaccinated commercial rabbits

A case of a myxoma virus strain in vaccinated and non-vaccinated rabbits is described, and genetic identification of that strain was performed in this study. In two commercial farms being 150km apart, myxomatosis has been occurred after the import of animals from a common supplier. The disease was manifested firstly in the existing non-immune population of does and fatteners, and later in all vaccinated animals, being 2-3 months immune at the time of first symptoms. Morbidity was almost 100% with nasal discharge, listlessness, fever, eyelid swelling, eye and nasal purulent discharge, papules in the ears, facial oedema, and swelling of the anagenital region, with result always the death of the animals. Examination by PCR had shown the presence of a 492-bp specific product in all the symptomatic animals tested from both farms, having 100% nucleotide sequence identity with the homologous region of the myxoma virus Lausanne strain. The simultaneous occurrence of myxomatosis in the vaccinated and non-vaccinated rabbits of both farms, suggests that the supplier was possibly the source of a viral isolate with increased virulence.

Influence of oestrus on the heat stability and other characteristics of milk from dairy goats.

We examined the heat stability, somatic cell count (SCC), pH, fat, protein and lactose content of milk from goats during the oestrous period, in order to investigate evidence of possible oestrus effects on milk physical and chemical properties. Goats free from mammary infections were ranked on average SCC from three tests so that they could be stratified randomly in pairs to synchronized oestrus or left as unsynchronized non-oestrus controls. The synchronisation consisted of insertion of an intravaginal progesterone-releasing device for 17 d, and introduction of the bucks the day of the device removal (D0). The repeated measurements analysis of variance model included the fixed effects of the experimental group (oestrus or control) and day and the corresponding interaction and also the random effect of doe. Reduced milk-heat stability, increased SCC, increased protein content and reduced pH were found in the milk samples of the oestrus group on D1, 2 and 3. The fat and lactose content of the milk was not affected by oestrus. These data indicate that the milk of goats during the mating period has reduced heat stability and, therefore, that dilution into bulk tanks should be recommended to avoid clotting when milk is intended for high thermal treatment.

The role of lactate dehydrogenase, alkaline phosphatase and aspartate aminotransferase in the diagnosis of subclinical intramammary infections in dairy sheep and goats

The objective was to investigate the changes occurring in the activities of the enzymes lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and aspartate aminotransferase (AST) in sheep and goat milk as a result of subclinical intramammary infections (IMI) and to evaluate the use of these enzymes for the diagnosis of subclinical IMI in dairy sheep and goats. A total of 206 samples of sheep milk and 162 samples of goat milk, obtained from equal udder halves, were used in the study. For each species they were divided into two groups: a no-infection group and a subclinical infection group. Activities of LDH, ALP and AST were significantly higher in the subclinical infection group than in the no-infection group (P<0.05) in both sheep (LDH: 350.42+/-11.25 v. 120.91+/-4.41; ALP: 2773.43+/-105.18 v. 2189+/-94.24; AST: 29.57+/-0.74 v. 17.32+/-0.46) and goats (LDH: 354.07+/-13.33 v. 103.79+/-3.75; ALP: 311.13+/-25.74 v. 137.24+/-19.62; AST: 27.59+/-6.42 v. 15.87+/-0.45). The activity of LDH was identified as indicator for subclinical IMI in both sheep and goats. The optimum cut-off values for LDH activity, offering the highest diagnostic sensitivity (DSn) and diagnostic specificity (DSp), determined by receiver operating characteristic (ROC) analysis, were at 197 U/l, 185 U/l and 197 U/l for sheep, goats and both species, respectively. DSn for sheep, goats and both species at these cut-off values was 92.8%, 98.2% and 94.0%, whereas DSp was 95.4%, 96.3% and 96.3%, respectively. It was concluded that the determination of LDH activity in milk serum is a sensitive and reliable method for the detection of subclinical IMI in dairy sheep and goats.