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Salmonellosis of pet birds

Salmonellosis of pet birds

D.E. Salmon (1885) first isolated Salmonella from the infected pigs. It was considered as a cause of 'hog cholera' until the discovery of the etiological virus.

 The nomenclature of the bacteria (‘Salmonella’) was done in memory of Salmon. In 1889, Klein (United Kingdom) first isolated Salmonella Gallinarum from chickens with ‘fowl typhoid’. Loeffler first described Salmonella Typhimurium from a natural outbreak of typhoid like infection in mice. Among the pet birds, Salmonellosis was first described in ducks (Vandervort 1954; Keymer 1958), and parakeets (Kaye et al. 1961; Madewell and McChesney 1975).

Etiology:

The genus Salmonella is classified under the family Enterobacteriaceae that belongs to the order Enterobacteriales. There are two major species under the genus Salmonella i.e. S. enterica (>2440 serovars) and S. bongori (20 serovars). A third proposed species is S. subterranea, yet to be recognized as a true species. Salmonella enterica is considered as type species of the genus at present. S. enterica has 6 subspecies (ssp.): salamae, arizonae, diarizonae, houtenae, indica, enterica.
 Most of the pathogenic Salmonella are designated as ‘serovar’ under the S. enterica ssp. enterica. Virulent serovars are: Typhi, Typhimurium, Dublin, Choleraesuis, Pullorum, Gallinarum, Abortusovis. Salmonella Typhimurium (var. copenhagen) is the most frequently isolated serovar from different outbreaks in psittacine birds. In passerine birds (perching/song birds), S. Typhimurium phage types DT40, DT41, DT56, DT160 are adapted. It acts as either a primary pathogen or it causes sub-clinical infection in young and immunocompromised birds. If the density of flock is high and the quantity of available feed is low, most of the birds become debilitated and are susceptible to S. Typhimurium infection.
 Other sub-species and serovars of Salmonella isolated from pet birds include S. houtenae, S. arizonae, S. Rissen, S. Enteritidis, S. Pullorum, S. Gallinarum, S. Newport, S. Panama, S. Rublislaw, S. Aberdeen, S. Thompson and S. Wasenaar. Among them, S. Gallinarum can infect canaries, ring dove, pheasants, peacocks and peafowl. A novel Salmonella serovar (S. Pajala) was isolated from Peregrine falcon (Falco peregrinus) nestlings.

Host Susceptibility:

Clinical outbreaks of salmonellosis are frequently detected in passerine and psittacine birds. Among passerine birds, finches (Fringillidae) and sparrows (Passeridae) seem to be particularly susceptible to Salmonella spp. infection. Salmonellosis is reported from canary (Serinus canaria), eurasian siskin (Carduelis spinus), zebra finch (Taeniopygia guttata), bengalese finch (Lonchura striata domestica) and picoplat (Sporophila intermedia). Fatal outbreaks with high mortality were reported in psittacine birds such as lories and lorikeets (Trichoglossus, Lorius, Eos spp.), budgerigars (Melopsittacus undulatus), parakeets (Psephotus spp., Psittacula spp.), and sulphur crested cockatoo (Cacatua galerita galerita). The birds with caecum [e.g. macaw (Ara sp.), amazon parrot (Amazona sp.)] can asymptomatically carry Salmonella spp. like poultry. Both free-ranging and captive blue-fronted amazon parrots (Amazona aestiva) are detected to carry Salmonella spp. Although, lilac-crowned amazon parrots (Amazona finschi Schlater) was found died due to S. Enteritidis infection. Moreover, S. Typhimurium is detected as a primary pathogen causing death of blue and gold macaws (Ara ararauna).
 Occasionally, raptors or hunter birds (e.g. falcon, red kite), game birds [e.g. red-legged partridge (Alectoris rufa)], free-ranging sparrow (Passer domesticus), gull (Laridae), wild birds such as temminck’s seedeater (Sporophila falcirostris), chestnut-capped blackbird (Chrysomus ruficapillus), brown-headed cowbirds (Molothrus ater), white-throated sparrows (Zonotrichia albicollis) can also harbour Salmonella spp.

Transmission:

Infected pet birds, rodents, reptiles, wild birds, contaminated water, feed and eggs act as source of Salmonella spp. When the pet birds are gathered in an exhibition the healthy birds come in direct contact to the infected birds. The rodents, reptiles and wild birds having access to the open-air aviary can contaminate the place. The bacteria can survive for extensive periods on wood and dust and can live for 28 months in avian faeces. The contaminated places become a constant source of infection. In a pet shop, iguana (Iguana iguana) was identified as a source of Salmonella spp. infection in a cockatoo. Ingestion of contaminated feed and drinking water is the major route of transmission in pet birds. Sometimes, the infection is also transmitted by owners or attendants through their contaminated hands, feet and clothes. Trans-ovarian transmission is common in poultry, although, is not frequently observed in pet birds.

Pathogenesis:

Salmonella Typhimurium
Following oral route of transmission, Salmonella is deposited in the intestine, where they invade enterocytes. The bacteria can invade the epithelial cells throughout the intestine, although, caeca and ileocaecal junction are the preferred site. In the intestine, low pH, peristalsis, intestinal mucus, lysozyme in secreations and moreover normal microbial flora try to destabilize the bacterial colonization. Normal microflora in adults prevents the Salmonella colonization by occupying their receptors, known as ‘competitive exclusion’. Young are more susceptible as their intestinal microfloral range is not fully developed. Bacterial fimbriae, lipopolysaccharide (LPS), pathogenicity island (SP-1-T3SS associated proteins) help in adherence with the enterocytes. The interaction between the T3SS proteins (SipA, SipC) with the actin cytoskeleton of the enterocytes causes cytoskeletal rearrangements to generate an uneven surface (membrane ruffle).
 The organisms are trapped within the ruffled membrane and are internalized by the enterocytes. Within the enterocytes, the bacteria reside in a membrane bound vacuole i.e. Salmonella containing vacuoles (SCV). As the SCV matures, it migrates from the luminal border of the enterocyte to the basal membrane. In the basal membrane, the SCV enter the macrophages associated with peyer’s patches in the sub mucosal space. The SCV never fuses with lysosome within macrophages and thus avoid phago-lysosome fusion which is necessary to kill the bacteria. Additional factors such as SP-2-T3SS (SipC protein), SP-3 associated proteins also help in intracellular survival. The formation of Salmonella induced fibrils (SIF) help in bacterial replication in an unknown way. It is evident that major portion of the infection is cleared by the macrophages, only certain part can survive leading to chronic infection or carrier state with persistent faecal shedding. Sometimes, invasion of Salmonella takes place beyond the intestine which causes bacterimia, survival and replication of the bacteria in reticulo-endothelial cells of liver and spleen. In passerine birds (canary, finch and starling), esophagus and crop are the preferred site for bacterial colonization after bacterimia.

Clinical Symptoms:

Acute and chronic form of salmonellosis is detected in pet birds. In acute form, huge mortality without any prior cardinal signs was observed in a flock of canaries. A S. Typhimurium infected macaw (Ara ararauna) showed depression, anorexia, delay in the emptying of crops (ingluvies), laboured breathing and diarrhoea for 3–4 days before death. Greenish-yellow diarrhoea was observed in adult budgerigers (Melopsittacus undulatus) infected with S. Gallinarum.
 The chronic form shows numerous general symptoms such as anorexia, diarrhoea, dyspnoea, lethargy, cachexia, ruffled feathers, subcutaneous granuloma, crop stasis, conjunctivitis, arthritis and panophthalmitis. Adult budgerigars with chronic salmonellosis showed loss of condition, unwillingness to fly, inability to perch, and gathering in the bottom of the cage (Fig. 2.3). Drop in egg production and increased embryonic mortality rate was observed. In pigeons, in addition to the general syndrome, polyuria, arthritis of the joint, nervous symptoms and dermatitis followed by death was detected.
 Stress associated with environment, season, change of diet and housing, transport, breeding, concurrent infection, and introduction of new birds without quarantine are the major predisposing factors for salmonellosis in pet birds.

Lesion:

In acute form of salmonellosis in pet birds, no specific gross lesion is observed. In chronic infection, congestion of lung and intestine, spleenomegaly, hepatomegaly, liver with necrotic foci is observed (Fig. 2.4). The passerines birds infected with S. Typhimurium shows the granulomatous lesions like pseudotuberculosis. Esophagitis and ingluvitis with necrotic plaques in esophagus and crop, respectively, are consistently detected in wild or free-range passerine birds. In European goldfinch (Carduelis carduelis), instead of esophagus lesions are sometimes developed in the subcutaneous tissues. In canaries, congestion of vital organs and necrotic foci on the liver with a nodular and miliary appearance is detected.
 Intestinal content become dark in colour due to haemorrhagic diasthesis. In lories and lorikeets, petechial hemorrhages on the serosal surface of the proventriculus, ventriculus, and cardiac muscle along with atrial dilation are observed. Sometimes, bacterial emboli occur in liver, spleen, lung, kidney, and proventriculus. In S. Typhimurium infected macaws, pectoral muscle atrophy, fibrinous exudate on intestinal mucosa, white-greyish nodules on intestinal serosa, myocardium, lungs and ingluvies mucosa are commonly observed. In young budgerigars, fibrin deposit is observed as a white, thick layer on the pericardium. Petechial haemorrhages are detected on the surfaces of pericardium, myocardium, gizzard, duodenum and ileo-caecum. In garden birds infected with S. Typhimurium, esophageal ulcers, granulomata in soft tissues, hepatomegaly and spleenomegaly are observed.
 Microscopic inspection of histopathological sections shows necrosis of parenchymatous organs specially the liver with granulocyte infiltration and fibrin deposition. In young and adult budgerigars, the blood vessel walls become hyalinized in appearance with various-sized microthrombi. The chronic cases are characterized by formation of a granuloma. A typical granuloma consists a necrotic centre which is surrounded by granulocytes and macrophages containing Salmonella spp. Multinucleated giant cells are found in chronic infection. In passerine birds, epithelial surface of the esophagus is ulcerated and it forms a thick layer of necrotic cellular debris composed of degenerated and intact leukocytes with gram-negative bacteria. Infiltration of heterophils, lymphocytes and plasma cells occur into the underlying sub-mucosa.

Diagnosis:

Clinical Specimens
Clinical samples include faeces or cloacal swabs, blood/serum of live birds and affected tissues, such as liver, spleen, heart, intestine/caeca, lung, esophagus/crop, brain and kidney in 10% buffered formalin. Before collection of cloacal swabs, pericloacal asepsis with iodized alcohol is performed. Blood samples are collected from jugular, wing or ulnar vein. The environmental samples, such as pooled faeces, litter and dust from the cages, feed and drinking water should be examined to know about an outbreak, if any. Specimens should be collected before antibiotic treatment of the birds. After death, the collection should be done immediately from fresh carcasses. For ‘pre-enrichment’, swabs should be collected in buffered peptone water. Pre-enrichment in buffered peptone water helps in survival of Salmonella from freezing, heating and desiccation. The cold chain (4–5 °C) should be
maintained during transportation of the samples to the laboratory.

Diagnostic Techniques:

(a) Clinical signs and lesions after necropsy, history of direct contact with infected birds give a tentative diagnosis.

(b) Direct Examination:
An impression smear prepared from clinical samples such as cloacal swab/faeces/tissues, is stained by Gram’s Method. Salmonella appears as gram negative small rods with no distinct characteristics. The tissue samples of heart, lung, liver, spleen, kidney, and intestine are fixed in 10% buffered formalin, embedded in paraffin, sectioned at 3 mm, and stained with hematoxylin-eosin and periodic acid-Schiff for direct examination of the bacteria.

(c) Isolation of bacteria from clinical samples:
Clinical samples require pre-enrichment and enrichment for growth. For pre-enrichment, the samples collected in buffered peptone water are kept at 40 °C for 24 h. The pre-enriched clinical samples are transferred into enrichment medium such as selenite or tetrathionate broth and are incubated at 40 °C for another 24 h.
 From the supernatant, the samples are plated in brilliant green agar (BGA) or xylose-lysine-deoxycolate agar (XLD) and are re-incubated for another 24 h at 40 °C. Convex, pale red, translucent colonies in BGA and red coloured colonies with black centres in XLD agar are presumably diagnosed as Salmonella spp. S. Pullorum produces small, paler colonies than other salmonellae in BGA. The suspected colonies can be confirmed by different biochemical tests such as catalase, oxidase, IMViC, TSI, carbohydrate fermentation profile and lysine decarboxylase test.

(d) Serological tests:
1- Rapid whole blood/serum agglutination test: It can be used for rapid detection of Salmonella spp. with crystal violet stained or unstained Salmonella polyvalent ‘O’-antigen. Equal amount of suspected whole blood or serum is gently mixed with the antigen on a white tile. Tile is agitated gently for 2 min and is observed for reading. In a positive case clumping of antigen is visible within 2 min. The antigen is commercially available or it can be obtained from Veterinary institutes in different countries.
2- Tube agglutination test
3- Immunodiffusion
4- Immunofluorescence
5- ELISA

(e) Molecular Biology:
For rapid and reliable detection of Salmonella, conventional PCR based diagnostic techniques targeting invA or other genes are used. Genus and serovar (S. Enteritidis and S. Typhimurium) specific real-time PCR system have also developed. For phylogenetic analysis of the Salmonella isolates, pulsed field gel electrophoresis (PFGE) and randomly amplified polymorphic DNA (RAPD) can be used.

Zoonosis:

Zoonotic transmission of Salmonella spp. to human from parakeet kept as a pet was documented. A salmonellosis outbreak associated with dissection of an owl was reported among the elementary school children. The possibility of children infection (below 5 years age) is more in those families who rear a pet bird (odd ratio: 2.7) or a lizard (odd ratio: 3). In open air aviaries and children’s zoos, the transmission of Salmonella spp. was reported between wild birds, pet birds and human. Special care for designing such aviaries should be adopted.

Treatment and Control Strategy:

Antibiotics against Salmonella spp. in infected pet birds can be administered after doing the sensitivity of the bacterial isolates. Antimicrobial resistance of Salmonella spp. is a global concern at present. Successful treatment of infected canaries with 10% (w/v) enrofloxacin solution provided as 200 mg/l in drinking water for 5–7 days was observed. Treatment with kanamycin, gentamicin, trimethoprim/sulfamethoxazole suspension along with anti-diarrhoeals such as daolin and pectin combination is recommended.
 General control and prevention strategies such as isolation of diseased birds from the rest of the flock, cleaning and disinfection of cages, water and feed utensils with 10% (v/v) solution of sodium hypochlorite or commercial disinfectants are recommended. If the feedstuffs are suspected it should be replaced with new batch immediately.

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