Early migration pattern of Avibacterium paragallinarum in the nasal passage of experimentally infected chicken and Japanese quail by immunohistochemistry.

Infectious coryza (IC) is often a curse for poultry farmers when it occurs concurrently with several pathogens causing swollen head syndrome. The disease is caused by Avibacterium paragallinarum, which inflicts initial damage to the nasal and respiratory epithelium. This facilitates the progression of disease pathology across the nasal cavity, thereby providing a platform for multiplication of opportunistic microbes. In this study, we attempted to investigate the early entrance and migration pattern of A. paragallinarum in chicken and Japanese quail following experimental infection, by employing an in-house developed polyclonal antiserum against this pathogen. Antigenic-specificity of the raised antiserum was subsequently evaluated through immune-dot blot techniques and counter-current immunoelectrophoresis (CIE). The resultant antiserum characterized the antigen localization within formalin-fixed and partially decalcified nasal tissue sections though immunohistochemistry (IHC). Japanese quail showed prominent localization of the bacterial antigen at 12 h post-infection in anterior turbinates. However, the chicken exhibited a higher level of the bacterial pathogen with intense immuno-reactivity at 24 and 48 h post-inoculation. The decline in immunostaining intensity in the nasal tissue of chicken as well as Japanese quail by 72 h post-infection signifies either an attempt to resolve the infection by the resident immune cells across the nasal passage of the host, or its dissipation by certain inherent innate immune factors present across the nasal passage that are still unknown to us. In the present study, we used a moderately virulent pathogen (A. paragallinarum) that inflicted a mild to moderate degree of damage to histo-architecture of the nasal passage and provided a discernible migratory pattern with fewer alterations, along with provision toward unravelling basics of the immuno-pathogenetic mechanism. This knowledge will support efforts towards the development of a future mucosal nasal vaccine in birds affected with IC.

Hypervitaminosis D3 in broiler chicks: histopathological, immunomodulatory and immunohistochemical approach.

The present study was undertaken to investigate the toxic effects of higher doses (1,00,000 IU i.e. 2.5 mg/kg body weight (BW)) of vitamin D3, concomitantly with bacterial endotoxins (lipopolysaccharides: LPS) to study the immunomodulatory potential of vitamin D3 in IBL-80 broiler chicks. The chicks were divided into four groups [group I (NSS), group II (LPS), group III (Vit. D3 + NSS), and group IV (Vit. D3 + LPS)] containing eight chicks in each group, treated accordingly for 21 days. Birds were kept under close observation for apparent clinical signs and symptoms. Clinically, vitamin D3 treated chicks were dull, off feed, showed polydipsia, polyuria, watery faeces, rigidity of limbs, severe dehydration, weakness and significant progressive emaciation. Grossly, the bones were soft whereas most organs revealed congestion and hemorrhages in visceral organs. Histopathologically, renal tubular epithelium showed coagulative necrosis and metastatic calcification. The lung parenchyma and bronchi showed hemorrhages, congestion with diffuse heterophilic cell infiltration in inter-alveolar septa and infiltration of heterophils in alveoli along with proteinacious fluid in LPS treated chicks of treatment group, suggesting immunomodulatory action of vitamin D3. Vitamin D3 treated chicks showed strong expression of Calbindin D28k in intestine and kidney but weak expression in lung, which can be linked to nephrocalcinosis seen in kidney and from its prospective role in cellular calcium homeostasis.

Histopathological and immunohistochemical diagnosis of infectious bursal disease in poultry birds.

AIM: The aim of the present study was to diagnose infectious bursal disease (IBD) using gross, histopathological, and immunopathological approaches and to compare efficacy of immunohistochemical techniques with conventional diagnostic techniques. MATERIALS AND METHODS: A total of 33 samples were collected from the six different poultry farms from Ludhiana and the nearby districts. Upon gross analysis of the necropsied birds, the relevant tissue samples such as bursa, kidney, junction of proventriculus and gizzard, heart, and muscles were then processed for histopathological and immunohistochemical studies. RESULTS: Varied macroscopic changes were noted in bursa, characterized as swollen, hemorrhages to atrophy in size. Nonetheless, hemorrhages over thigh muscles were rarely seen. Histologically, the bursa showed prominent fibrotic and atrophic changes. Rarefaction of bursal follicles with intermittent infiltration of lympho-mononuclear cells with chronic cystic changes was additional changes, considered to be paramount for IBD. Expression and localization of IBD specific viral antigens were noticed mainly intracellular to the rarefied areas of bursal follicle section(s), in conjunction to inner lining of the cystic cavities of affected follicles. In addition, the junction of proventriculus and gizzard, the heart muscle, respiratory ciliated epithelium, and proventriculus also revealed positive expression to IBD virus (IBDV) antigen. Advanced immunopathological techniques, i.e., immunofluorescence further testified the evidence of antigen as positive green signal within affected follicles. Further consideration to the reliability of various techniques employed, positive correlation (r=0.64623) was emerged out with conventional pathological scoring. CONCLUSION: It is concluded that the bursa acts as an organ of choice for demonstrating IBDV antigen for specific diagnosis of disease using immunohistochemistry (IHC), and IHC staining is a precise, specific, rapid, and reliable method to demonstrate the IBDV antigen in the altered tissues due to IBDV infection.

Clinicopathological Studies on Vitamin D(3) Toxicity and Therapeutic Evaluation of Aloe vera in Rats.

A study was conducted to examine the clinical signs, hematological, biochemical and histopathological changes in vitamin D(3) toxicity at a dose rate 2 mg/kg b.wt. of vitamin D(3) and to assess the protective effect of Aloe vera in vitamin D(3) toxicity. The clinical signs observed were anorexia, progressive weight loss, difficulty in movement and respiration, diarrhea, epistaxis, subnormal body temperature and nervous signs before death. Mortality was observed in treated rats between day 10 and day 19 of treatment. The gross postmortem changes observed were severe emaciation, white chalky deposits on epicardial surface of heart, pin point white deposits on cortical surface of kidneys with pale yellow discoloration and diffused white deposits on serosal surface of stomach and intestine with bloody ingesta in lumen. The hematological changes included non-significant increase in hemoglobin and total leukocyte count and significant increase in relative neutrophil count. The biochemical changes observed were significant increase in plasma concentration of calcium, phosphorus and blood urea nitrogen, whereas a significant decrease in the concentration of albumin and total plasma protein was observed. The histopathological lesions included calcification of various organs, viz., tongue, stomach, intestines, kidney, heart, aorta, larynx, trachea, lungs, spleen, choroid plexus arteries of brain and vas deferens. The Aloe vera juice (2.5% in drinking water) has no protective effect on vitamin D(3) toxicity (2 mg/kg b.wt.).

Immunolocalization of pulmonary intravascular macrophages, TLR4, TLR9 and IL-8 in normal and Pasteurella multocida-infected lungs of water buffalo (Bubalus bubalis).

Water buffalo are of considerable economic significance in South East Asia, but these animals suffer from many bacterial respiratory diseases including haemorrhagic septicaemia caused by Pasteurella multocida. Bacterial respiratory diseases of animals cause lung inflammation that is characterized by the activation of Toll-like receptors (TLRs) expressed on macrophages, expression of chemokines and recruitment of neutrophils and monocytes. Pulmonary intravascular macrophages (PIMs) present in the alveolar septa play a critical role in lung inflammation, but there are no data on the immunolocalization of PIMs or the expression of TLRs and chemokines such as interleukin (IL)-8, in the lungs of water buffalo. The present study compares the occurrence of PIMs, TLR4, TLR9 and IL-8 in the lungs of normal water buffalo and those infected with P. multocida. Labelling of PIMs with the anti-human macrophage antibody (MCA874G) demonstrated an increase in this population in inflamed lungs. TLR4 and IL-8 were detected in the alveolar septa, airway epithelium and endothelium of large blood vessels of normal lungs. TLR9 expression was similar to that of TLR4, but TLR9 was not expressed by the endothelium of arteries and veins. While the expression of TLR9 and IL-8 was increased in the inflamed lungs compared with normal lungs, TLR4 labelling intensity remained unchanged or was reduced. The expression of these molecules potentially plays a role in the regulation of lung inflammation.

Chlorpyrifos chronic toxicity in broilers and effect of vitamin C.

An experiment was conducted to study chlorpyrifos chronic toxicity in broilers and the protective effect of vitamin C. Oral administration of 0.8 mg/kg body weight (bw) (1/50 LD50) chlorpyrifos (Radar(®)), produced mild diarrhea and gross lesions comprised of paleness, flaccid consistency and slightly enlargement of liver. Histopathologically, chlorpyrifos produced degenerative changes in various organs. Oral administration of 100 mg/kg bw vitamin C partially ameliorated the degenerative changes in kidney and heart. There was insignificant alteration in biochemical and haematological profiles. It is concluded that supplementation of vitamin C reduced the severity of lesions induced by chronic chlorpyrifos toxicity in broilers.

Neuropathological studies of chickens following exposure to chlorpyrifos.

The objective of this study was to determine the putative neuropathological effects in young chickens after administration of a single dose of 55 mg/kg bw chlorpyrifos. The gross lesions of the nervous system comprised of congestions in the brain. Microscopic examination of brain showed mild congestion of cerebral blood vessels and mild perivascular cuffing of lymphomononuclear cells in the cerebral cortex and necrosis of the neurons. The interesting findings were the presence of cytoplasmic vacuolations of cerebral neurons and swelling of the endothelial cell of the cerebral capillaries. Cerebellum showed congestion and hemorrhages in the granular layer and necrosis of Purkinje cell. Sciatic nerve exhibited mild edema, swelling and degeneration of axons, and swelling of Schwann cells. There was a significant inhibition of plasma cholinesterase enzyme activity in chickens administered with chlorpyrifos compared to chickens of control group. The study revealed that administration of chlorpyrifos produces neuropathological lesions in chickens shortly after exposure.

Toxic impacts of cypermethrin on behavior and histology of certain tissues of albino rats.

In the present investigation, the behavioral, morphological, and histopathological effects of cypermethrin, a widely used synthetic pyrethroid insecticide, was ascertained in male and female albino rats (Rattus norvegicus). Cypermethrin administered at repeated oral doses of 5 and 20 mg/kg/day for 30 days produced varying degree of mild to moderate toxic symptoms and behavioral changes in both male and female rats. The lower dose produced very mild toxicosis characterized by intermittent diarrhea, decreased feed intake, and thick eye discharge, whereas higher dose displayed mild to moderate toxicosis with diarrhea, decreased feed intake, loss of body weight, dyspnoea, ataxia, eye discharge, and salivation. Two female and one male albino rats died between 23 to 28 days after displaying signs of incoordination and tremors. Repeated oral doses of cypermethrin for 30 days enhanced the relative weight of liver and heart, but significantly decreased that of brain, kidneys, and testes. Microscopically, cypermethrin produced neuronal degeneration and increase in glial cells in brain, and disorganization of hepatic laminae, increase in sinusoid, and necrosis of hepatocytes in liver. Section of kidney displayed hemorrhage and sloughing off renal epithelial cell in the convoluted tubules, shrinkage of glomeruli, and necrosis of renal tubules. Repeated administration of cypermethrin also produced hemorrhages within myocardium, disruption of branching structure, and loss of striation of cardiac tissue; thickening of alveolar septa in lungs, partial to extensive loss of various stages of spermatogenesis in testes, and loss of follicular cells and oocytes in ovaries. The study suggested that repeated oral exposure of cypermethrin has considerable harmful effects on body organs in R. norvegicus.

A Case of Intranasal Hemangioma and Concurrent Tetracycline-induced Ulcerative Gastritis in Dogs.

Incidence of drug-induced gastritis and ulceration in human medicine is well established. Besides, unilateral hemangioma, a unique concurrent case of tetracycline induced gastric toxicity in a dog, characterized by gastritis and ulceration is being reported here. Grossly, the appearance of gastric ulcers mimicked the appearance of Italian pizza. Histological examination further supported drug-induced etiology in this case. This is probably the one of the few cases in the annals of veterinary medicine to be documented as drug-induced gastric toxicity in dog.

A protective effect of vitamin E and selenium in ameliorating the immunotoxicity of malathion in chicks.

A study was conducted to determine whether a combination of vitamin E and selenium was able to counteract the immunosuppressive effects of chronic toxicity of malathion in chicks. The chicks were divided into three groups. The first group received malathion 10 mg/kg body weight; the second group received the same dose of malathion but was supplemented with vitamin E and selenium for 60 days; and the third group served as the control. The concentration of total immunoglobulins and circulatory immune complexes decreased following the administration of malathion, whereas the supplementation of the diet with vitamin E and selenium partially counteracted this effect. The histopathological changes in the bursa of Fabricius due to malathion were consistent with these findings. Supplementation with vitamin E and selenium partially ameliorated the degenerative changes in the bursa of Fabricius.

Studies on some blood parameters of crossbred calves with experimental Theileria annulata infections.

Subcutaneous inoculation of 1 ml of ground Theileria annulata tick tissue stabilate (0.75 tick equivalent) into crossbred calves (n = 6, average age 53 days) resulted in the development of acute theileriosis. The percentage parasitaemia was 71.7% +/- 3.3% on day 20 after inoculation. Macroschizonts were observed in lymphocytes and monocytes. Phagocytosed schizonts were observed in neutrophils, along with cytoplasmic vacuolation in monocytes and neutrophils. There was progressive decrease (p < 0.05) in the haemoglobin and packed cell volume, along with a marked reticulocytosis. Serum analysis revealed a decrease (p <0.05) in the concentrations of calcium, cholesterol and triglycerides, while there was an increase (p < 0.05) in the concentrations of blood urea nitrogen as compared to day 0 values. The total serum proteins, albumin and serum immunoglobulin concentrations and the albumin-to-immunoglobulin ratio showed marked decreases (p<0.05). Coagulopathies included thrombocytopenia and an increased prothrombin time, along with a non-significant increase in the bleeding time and activated partial thromboplastin time during the terminal stages of the disease. There was an increase in the osmotic fragility of erythrocytes during the disease. Morphological alterations in the erythrocytes were observed with the developing parasitaemia.

Haematological and biochemical studies in broiler chicks fed ochratoxin and inoculated with inclusion body hepatitis virus, singly and in concurrence.

Day-old broiler chicks, which had been shown to be negative for maternal antibodies against inclusion body hepatitis (IBH) virus and for viral antigen in cloacal swabs, were divided into four groups of 20 chicks each. One group was fed ochratoxin-A at 0.5 ppm from 3 to 38 days of age, another group was inoculated with 1 ml of IBH virus containing 10(6.5) EID50 per 0.2 ml. A third group was given both ochratoxin-A and infected with IBH virus. The fourth group served as the control. Anaemia was observed in all three treated groups but the changes were more pronounced in the combined group. The biochemical changes also suggested a cumulative damaging effect by ochratoxin-A and IBH virus.

Haematological and biochemical studies on experimental Theileria annulata infection in crossbred calves.

Administration of ground-up tick tissue stabilate (0.75 tick equivalent) by the subcutaneous route to crossbred calves aged 1 week to 1 month led to the development of acute theileriosis. Haematological studies revealed significant progressive decreases in haemoglobin concentration, packed cell volume and red blood cell count, whereas the total leukocyte count showed an initial non-significant leukocytosis followed by a significant leukopenia. Analysis of serum revealed significant increases in levels of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, creatinine kinase and gamma-glutamyltransferase, and in the concentrations of uric acid, blood urea nitrogen and bilirubin. The concentrations of total protein, albumin, glucose, cholesterol and calcium showed non-significant decreases, while phosphorus decreased significantly during the terminal stages of the disease.

An outbreak of bacillary haemoglobinuria in sheep in India.

An outbreak of bacillary haemoglobinuria was recorded in 60 out of 110 sheep in Ludhiana, Punjab, India. The condition was clinically characterised by fever, haemoglobinuria, constipation, weakness of hind quarters followed by recumbency, respiratory distress and death in 16 sheep. Haematological studies revealed moderate to severe degrees of anaemia associated with leucocytosis. Plasma gamma-glutamyl transferase, alkaline phosphatase and creatinine phosphokinase activities were significantly higher in haemoglobinuric sheep. Babesiosis and copper poisoning were ruled out on stained blood film examination and from blood mineral profiles, respectively. Post-mortem examination of affected sheep revealed no gross changes. Pure cultures of Clostridium haemolyticum isolated from heart blood, liver, kidney and spleen of freshly killed sheep confirmed the disease. Parenteral administration of procaine penicillin was effective in the treatment of affected sheep.

Effect of alkyl sulfides on diazomethane-induced methylation of DNA in vitro.

Organosulfur compounds found in Allium species such as garlic and onions inhibit carcinogenesis induced by alkylating agents. One potential mechanism for this activity involves scavenging of ultimate carcinogenic species by the nucleophilic sulfur atom. Carcinogenic methylating agents such as methyl-nitrosourea produce, among others, O6-methylguanine and N7-methylguanine in DNA both in vivo and in vitro. In the present study we have determined the effect of several alkyl sulfides on the formation of O6-methylguanine and N7-methylguanine in DNA reacted with the methylating agent diazomethane in vitro. Dipropyl sulfide and diallyl sulfide affect guanine methylation by increasing the O6/N7 ratio without drastic alterations in the total amount of adduct formed. Three similar compounds--diallyl disulfide, allyl methyl sulfide and diallyl ether--had no appreciable effect on the amount of alkylation at either position. These data suggest that scavenging of diazomethane-like ultimate carcinogens does not play a major role in the inhibition of carcinogenesis by organic sulfides.

Outbreak of mycotic tracheitis in domestic fowl.

An outbreak of mycotic tracheitis was observed in 8000 2-month-old, female White Leghorn birds. The birds were showing difficult respiration and there was mortality of 7-8 birds daily in the flock. On post-mortem examination of the affected birds, the trachea was found to be occluded with a white caseous mass. Microscopically hyphae were found invading the tracheal epithelium, cartilage and serosal layer along with infiltration of macrophages and lymphocytes. Aspergillus fumigatus was isolated in pure culture from the trachea. The birds responded to oral copper sulphate treatment. The ubiquitous occurrence of the organism and the conditions of the harvesting season have been found to be responsible for the outbreak of the disease.

Three congenital anomalies of domestic fowl.

Of 35,000 necropsies performed on domestic poultry, three cases of anomalies were recorded. The anomalies observed were duplicate polydactyly, accessory hepatic lobe, and reduplication of caeca.