Selective breeding for high natural antibody level increases resistance to avian pathogenic Escherichia coli (APEC) in chickens.

Keyhole limpet hemocyanin (KLH)-binding natural antibody (NAb) titers in chickens are heritable, and higher levels have previously been associated with a higher survival. This suggests that selective breeding for higher NAb levels might increase survival by means of improved general disease resistance. Chickens were divergently selected and bred for total NAb levels binding KLH at 16 weeks of age for six generations, resulting in a High NAb selection line and a Low NAb selection line. To for test differences in disease resistance, chickens were challenged with avian pathogenic Escherichia coli (APEC) in two separate experiments. Chickens at 8 days of age received one of four intratracheal inoculations of 0.2 mL phosphate buffered saline (PBS): 1) mock inoculate, 2) with 0.2 mL PBS containing 108.20 colony-forming units (CFU)/mL APEC, 3) with 0.2 mL PBS containing 106.64 CFU/mL APEC, and 4) with 0.2 mL PBS containing 107.55 CFU/mL APEC. Mortality was recorded during 7 days post inoculation. Overall, 50-60% reduced mortality was observed in the High line compared to the Low line for all APEC doses. In addition, morbidity was determined of the surviving chickens at 15 days of age. The High line had lower morbidity scores compared to the Low line. We conclude that selective breeding for high KLH-binding NAb levels at 16 weeks of age increase APEC resistance in early life. This study and previous studies support the hypothesis that KLH-binding NAb might be used as an indicator trait for to selective breed for general disease resistance in an antigen non-specific fashion.

Field and laboratory findings following the large-scale use of intermediate type infectious bursal disease vaccines in Denmark.

Following a period of clinical outbreaks of very virulent infectious bursal disease virus (vvIBDV) in Denmark, the histological bursal lesion score (HBLS) was used on a national scale to screen broiler flocks vaccinated with intermediate IBD vaccines for lesions indicative of IBDV challenge. High lesion scores were detected in a high percentage of healthy and well performing flocks despite the lack of other indications of the presence of vvIBDV. RT-PCR and subsequent sequencing showed the frequent presence of H253Q and H253N IBDV strains that were genetically close to the sequence of the intermediate vaccines with a relative risk ratio of 13.0 (P < 0.0001) in intermediate vaccine A or B vaccinated flocks compared to unvaccinated flocks. The relevance of these H253Q and H253N strains was tested under experimental conditions using a protocol derived from the European Pharmacopoeia for safety of live IBD vaccines. The results confirmed the higher pathogenicity for the bursa of these strains compared to intermediate vaccines as well as the negative effect on antibody response to a Newcastle disease (ND) vaccination performed at the peak of the bursa damage. The efficacy of the ND vaccination was still 100% showing that the H253N and H253Q IBDV strains would be considered as safe vaccine viruses. In conclusion, the use of the HBLS to screen commercial broiler flocks vaccinated with intermediate IBD vaccines for the presence of vvIBDV does not seem to be a reliable method due to the frequent occurrence of H253N and H253Q strains in those flocks. For screening of IBD vaccinated flocks for the presence of vvIBDV or other field strains, the RT-PCR with subsequent sequencing seems to be most suitable.

Signs Indicating Imminent Death in Escherichia coli-Infected Broilers.

Broilers were observed during 9 days for clinical signs after intratracheal inoculation at 8 days of age with 107 E. coli 506. It was determined if these signs were predictive for imminent death. Hourly observations were made daily from a distance of 1-2 m and nightly by camera observation, with respect to the following parameters: level of attention, locomotory activity, posture and appearance, interaction, and impairment of respiration. For deviations of the normal state for these five parameters (i.e., typical clinical signs of disease), scores were defined in up to four classes. The periods of time elapsing from attaining a score for the first time to death were registered per bird for each score for each parameter. Of 114 birds, 85 did not present typical signs of illness as described, and 29 presented the following clinical history: 25 died after presenting signs of illness, 2 died without previous signs, 1 fell ill but survived, and 1 fell ill and recovered. Extended clinical examination was performed in birds presenting clinical signs; temperature, heart rate, respiratory rate, and subcutaneous capillary refill time were measured. The level of attention, and posture and appearance were affected most often in ill birds; 25% of these birds died within 5 and 4 hr, respectively; 50% died within 12 hr; and 75% died within 20 and 19 hr, respectively. Any of these typical signs of illness visible from 1-2 m indicated imminent death, with 75% of the birds dying within 20 hr. Measurements resulting from extended clinical examination proved of lesser predictive value. From these observations, a protocol for intervention to prevent animal suffering may be designed.

Enterococcus hirae-associated endocarditis outbreaks in broiler flocks: clinical and pathological characteristics and molecular epidemiology.

BACKGROUND: Enterococcus hirae-associated endocarditis, characterized by a peak in mortality during the second week of the grow-out, and occasionally lameness, was diagnosed at Dutch broiler farms. OBJECTIVES: Field cases were studied to increase knowledge on clinical and pathological characteristics, pathogenesis and epidemiology of these infections. ANIMALS AND METHODS: In total, 1266 birds of 25 flocks from 12 farms were examined. Post-mortem examinations, bacteriology, histopathology, PCR and DNA fingerprinting was carried out. Six flocks were followed longitudinally (n = 1017 birds). RESULTS: Average mortality was 4.1% for the entire grow-out, of which 36% was attributed to endocarditis. Fibrinous thromboendocarditis of the right atrioventricular (AV) valve was found in 24% of hearts, compared to 7% and 4% with lesions of left and both AV valves, respectively. Thrombotic lesions were found in 24% (n = 432) of lungs, but only in larger branches of the Arteria pulmonalis. Occasionally, thrombi were found in the Arteria ischiadica externa and in liver and brain vessels. Enterococcus was cultured from 54% (n = 176) of heart and in 75% (n = 28), 62% (n = 106) and 31% (n = 16) of liver, bone marrow and lung samples, respectively. Further identification, using the Rapid ID Strep 32 API system and a PCR targeting mur-2 and mur-2(ed) genes was carried out on a subset of Enterococcus positive isolates (n = 65): both techniques identified the isolates as Enterococcus hirae. Pulsed-field gel electrophoresis did not indicate evidence of clonality between farms and flocks. CONCLUSIONS: The relevance of these findings for pathogenesis and epidemiology of E. hirae infections is discussed. CLINICAL IMPORTANCE. This study may facilitate diagnosis of field cases and may contribute to the design of further research and development of control measures.

Cloacal reflexes and uptake of fluorescein-labeled polystyrene beads in broiler chickens.

Experiments were performed (1) to quantify reflex movements and volume uptake of physiological salt solution by the cloaca and (2) to evaluate the conditioned cloacal uptake of fluorescein-labeled polystyrene. In experiment 1, measurements were done on birds (n = 12) once a day at 3, 4, and 5 d of age and during 5 consecutive days at wk 3, 5, 7, and 9 of age. The reflexes and volume uptake after applying saline droplets were studied simultaneously during 30 s. The median number and range of reflexes per 30 s during the first week of age were 45 (28 to 54), at 3 wk 35 (18 to 52), at 5 wk 44 (27 to 60), at 7 wk 47 (32 to 61), and at 9 wk 44 (23 to 56). The median volume uptake and range in wk 1, 3, 5, 7, and 9 were 0.10 (0.05 to 0.30), 0.25 (0.05 to 0.60), 0.58 (0.25 to 1.15), 1.05 (0.50 to 2.25), and 1.15 (0.30 to 3.05) mL per 30 s, increasing significantly with time. In experiment 2, a solution containing 10(7) polystyrene beads/mL was applied to the cloaca of broilers (3 aged 2 wk and 3 aged 9 wk) during 30 s. Most beads were found in the bursa of Fabricius. In the bursal lumen, a median of 10(6.43) beads/mL was found; the median number found in the follicular tissue was 5 (range 3 to 38) beads per tissue section. In the lumen content of ileum, cecum, and rectum of all birds together, it was 10(5.87), 0, and 10(6.32) beads/ ml, respectively. Polystyrene beads were never found intramuraly.

Avian leukosis virus subtype J in ovo -infected specific pathogen free broilers harbour the virus in their feathers and show feather abnormalities.

In the present paper, abnormal feather development associated with an in ovo avian leukosis virus (ALV) subgroup J infection in specific pathogen free broilers is reported. A description of gross lesions is given, together with the results of light microscopic, immunohistochemical and ultrastructural studies to further elucidate the characteristics of the feather pathology and to assess the presence of ALV subtype J in broiler feathers. The feather lesions found, which were mainly restricted to primary and secondary remiges, were thinness and increased transparency of the calamus and sparseness of the vane. In a few cases, feather growth retardation was present, while the vane remained ensheathed. Light microscopy did not reveal abnormalities, although immunohistochemically prominent anti-p27 staining was found in the central area of the feather pulp between cells forming the barbular epithelium. Transmission electron microscopy showed the presence of retroviral particles in various structures of the feather (epidermal collar and intermediate layer of feather epidermis), most virus being present in the same area that stained most prominently by immunohistochemistry and where apparently intra-epithelial gaps were seen. ALV was isolated from the upper two-thirds of primary remiges of almost all inoculated birds and 40% of contact birds. Polymerase chain reaction of cell cultures showed that the virus isolates belonged to subgroup J. This study shows that subtype J virus can be massively present in feather tissue of infected birds, which could have consequences for its transmission and diagnosis.

Polymyxin E-1 (colistin sulphate) (neuro-)intoxication in young ostriches ( Struthio camelus spp.).

Colistin (polymyxin E) is a cyclic polypeptide with a potent bactericidal action against most gramnegative bacilli. When used parenterally, polymyxins should be given with great care as they have a very small safety range, and easily induce neurotoxicity and nephrotoxicity. A dose of 39.5mg/kg body weight colistin sulphate injected subcutaneously induced rapid (within 1 to 3 h) mortality in young ostriches. Clinical signs of apathy, lethargy and hypotonia indicative of neurotoxicity of the compound were observed. At postmortem, vascular congestion of brain vessels was seen while, on histology, severe acute oedema was present in the epicardium and the intestinal serosa. Congestion of villi, swelling and vacuolization of the plexus ofAuerbach, as well as intermuscular and perivascular oedema of the heart, were also observed. In view of our observations in ostriches and in other species studied, a dose of >5mg/kg body weight polymyxin E is not considered safe for parenteral administration in ostriches.

Mycotic spondylitis in broilers caused by Aspergillus fumigatus resulting in partial anterior and posterior paralysis.

This report describes outbreaks of mycotic spondylitis in two broiler flocks of 17 and 19 days old. Postmortem examination showed kyphosis and compression of the spinal cord, due to osteomyelitis of either the cervical or the first thoracic vertebrae. These lesions resulted in partial anterior paralysis affecting wing mobility and posterior paralysis. Aspergillus fumigatus was cultured from affected vertebrae.

Observations on the influence of intestinal spirochaetosis in broiler breeders on the performance of their progeny and on egg production.

Observations were made on the effect of avian intestinal spirochaetosis (AIS) in eight broiler breeder flocks on the performance of their commercial broiler progeny flocks, and also on the production of eggs and the selection of hatching eggs. The effect of treatment on egg production was analysed. The broiler breeder flocks were examined for the presence or absence of antibodies, spirochaetes and clinical signs of AIS, and correlated with the performance data from 136 commercial broiler flocks produced by them. Broiler flocks from affected breeders with clinical signs of AIS had a poorer feed conversion of approximately 90 g per kg growth, an increased number of weak chicks, slower growth and poorer feed digestion than the offspring of unaffected flocks including those previously infected with AIS but without clinical signs. Breeder flocks with clinical signs produced 7.5% less eggs than unaffected flocks. A larger number of eggs (+ 3%) was considered to have insufficient weight for successful hatching. Treatment of hens before the onset of lay prevented the negative effects on egg production but later treatment was less effective. These findings indicate that AIS in broiler breeder flocks is associated with significant production losses through decreased broiler flock performance and decreased egg production and quality.

A cataract outbreak in broiler breeders.

Varying degrees of lens cataract and iritis were found in birds of a flock of grandparent broiler breeders. Major histopathological findings included stromal lens alterations with liquefaction (phacolysis) and globule formation, and a lymphocytic-plasmocytic anterior uveitis. Transmission experiments were performed to assess a possible infectious aetiology. However, a cause for the eye alterations could not be established.

Keratoglobus lesions in the eyes of rearing broiler breeders.

This report describes the occurrence of keratoglobus lesions in a rearing flock of broiler breeder grandparent stock (female line), and its ophthalmological and ocular pathomorphological features. Keratoglobus is known as a recessive sex-linked (Z chromosome) trait in some lines of chickens. The first cases were encountered at 5 weeks of age. At the end of the rearing period a total of 1.5% of the flock was affected. In the flock and affected birds, no signs of any other disease were detected. Compared to eyes of healthy flock mates, the most prominent symptom was ocular protrusion due to bulging and thinning of the cornea, and increase in anterior chamber depth. In addition, there were mononuclear, granulocytic and plasma cell infiltrates in the iris and corpus ciliare, and mild to minimal exudate in the anterior chamber in all cases examined. Although the corneal epithelium and collagen bundles appeared normal on electron microscopy, the corneal endothelium showed some loss of cells and the keratocytes some degenerative changes. The cornea had not lost its transparency and did not show any signs of inflammation, whereas the conjunctiva contained mononuclear infiltrations. Other eye structures did not show any clinical or pathological changes. The pathogenesis of keratoglobus lesions in chickens is unknown. Although a hereditary origin seems probable, it is not known whether the inflammatory changes found caused the presented pathology or were secondary to the extreme corneal bulging.

A syndrome associated with growth depression and amyloid arthropathy in layers: a preliminary report.

In heavy breed layers a new syndrome was encountered, associated with impaired growth. The disease developed from 5 to 6 weeks onwards. Morbidity in most flocks was 1 to 4%, but in some cases up to 20%. The affected birds were smaller in size, made a peeping sound like that of younger chickens, and showed a characteristic stiff gait. The stifle and hock joints were swollen. At necropsy 12 out of 66 affected birds (18%) had bronze-coloured livers and 23 out of 66 (35%) had orange-coloured joint deposits. Histology indicated that these birds appeared to have amyloidosis. The amyloid was found to react positively with anti-duck and anti-bovine amyloid protein A antisera on PAP-staining, indicating it to be of the reactive type. The cause of the syndrome is unknown. A possible association with infectious agents (Streptococcus faecalis and reovirus) is discussed.

Infection of broiler parent hens with avian intestinal spirochaetes: effects on egg production and chick quality.

Broiler parent hens were inoculated with avian intestinal spirochaetes several weeks before the onset of egg production. The infection persisted, wet droppings developed, and egg production, mean egg weight and carotenoid contents of the eggs were decreased. Hatching eggs were collected and incubated. In broilers which hatched from these eggs, reduced gain in body weight at 2 and 3 weeks of age, wet droppings, low plasma carotenoid concentration and elevated alkaline phosphatase activity in the blood plasma were observed. Spirochaetes were not detected in these broilers. These findings demonstrated the deleterious effects on chick quality of parental infection with avian intestinal spirochaetes. Avian intestinal spirochaetosis was diagnosed in about 2.5% of all submissions from reproductive flocks in 1991.

Infection of broiler chicks (Gallus domesticus) with human intestinal spirochaetes.

One-day-old broiler chicks were infected orally with spirochaetes isolated from cases of intestinal disorder in humans. Three different isolates were studied in an experiment of 22 days duration. No signs of clinical disease were observed; the infection did not cause changes in the concentration of carotenoids or in the activity of alkaline phosphatase in the serum. Spirochaetes were shed in the caecal faeces. At autopsy, no lesions were seen in the intestinal tracts of the birds. Large numbers of spirochaetes were present in the caecum of 30 to 100% of the infected birds; in a few birds small numbers of spirochaetes were observed in scrapings from the mucosa of the small intestine. On histological examination massive colonisation of the caecal mucosa was observed. Spirochaetes of two isolates had invaded the caecal mucosa and for one of these, they were found between enterocytes, but not below the basement membrane. The other isolate had produced gap-like lesions, subepithelial accumulations of spirochaetes and focal erosion. No signs of an inflammatory reaction were observed.

Influence of infection with avian intestinal spirochaetes on the faeces of laying hens.

Laying hens of 20 weeks of age were infected with avian intestinal spirochaetes (isolate 1380). After 20 weeks it was found that most infected birds shed 10(7) or more spirochaetes/ml caecal faeces. Faecal dry matter content was not significantly influenced by the infection, but the amount of crude fat in the faecal dry matter increased by more than 25%.

Spirochaetosis in broilers.

One-day-old broilers were inoculated with spirochaetes (isolate 1380); the inocula consisting either of spirochaetes derived from plate culture or from intestinal homogenates of previously infected birds. After 13 to 15, and 21 days after inoculation growth was depressed, serum concentrations of protein lipid, carotenoids and bilirubin were decreased and the fat content of the faeces was increased. The activity of alkaline phosphatase in the serum was increased but the activities of tau-glutamyl transferase, sorbitol dehydrogenase, aspartate aminotransferase and alanine aminotransferase of the infected groups were not affected. These findings indicate impaired resorption in the smaller intestine which cannot be explained from a direct effect of the spirochaetes on the smaller intestinal wall nor from pathological changes in the liver.

Observations on avian intestinal spirochaetosis.

Three SPF-laying hens were inoculated into the crop with avian intestinal spirochaetes which previously had been passaged in broiler chicks by oral inoculation (isolate 1380). Mild persisting gastrointestinal disorder developed; at nine months post inoculation spirochaetes were readily demonstrated in caecal faeces. Histologic examination of the caecal mucosa revealed many spirochaetes covering the mucosal surface and filling up the crypts lumina. Spirochaetes were found in intra- and subepithelial locations and in gaps running through the epithelium. These gaps often opened into subepithelial cavities crowded with spirochaetes ('gaplike lesions'). These lesions were seen mostly on the tips of the villi and in the deeper parts of the crypts. Massive erosion or desquamation of epithelium heavily infested by spirochaetes occurred. These findings indicate colonisation of the mucosal surface and of the crypts, penetration of the mucosa and colonisation of subepithelial compartments with spirochasetes in poultry suffering from intestinal spirochaetosis.

Incidence of spirochaetal infections in cases of intestinal disorder in chickens.

The incidence of intestinal spirochaetes was determined using direct fluorescent antibody microscopy on faecal and mucosal samples from chickens. Of 134 flocks with intestinal disorders 27.6% were found to be positive, but only 4.4% (of 45 flocks) were positive where signs of enteritis were absent. Flocks housed in cages or with access to litter were equally affected. No evidence was found for pigs as an aetiological factor.

Infectious typhlitis in chickens caused by spirochetes.

A weakly haemolytic spirochete was detected with an unabsorbed fluorescent antiserum to Treponema hyodysenteriae in smears and cultures of scrapings of caecal mucosa of laying hens with diarrhoea. Two groups of experimental chickens were fed a pure culture of this spirochete or homogenated intestinal contents of affected birds. Both groups showed clinical signs of disease such as increased water content of faecal material and slight retardation of growth. A non-specific typhlitis which histologically resembled milder forms of swine dysentery was seen in the birds from which spirochetes were isolated. The isolate obtained differed in cultural, biochemical, anti-genic and morphological characteristics from T. hyodysenteriae. The pathological significance of intestinal spirochetes and their possible epidemiological relation to swine dysentery are discussed.