Introduction and History

(cited by Calnek et al., 1997) Arizonosis is an acute septicemic disease, primarily of young turkey poults, caused by the bacterium Salmonella arizonae. S. arizonae is one of the most frequently identified salmonella serotypes in turkeys in the united states and is related to significant morbidity and mortality. The disease is clinically indistinguishable from Salmonellosis and is referred to as arizona infection or avian arizonosis (AA). Avian arizonosis is of considerable economic significance to the turkey ibdustry of North America and certain parts of the world.through reduced egg production and hatchability.

S. arizonae represents an antigenically diverse group of bacteria (over 300 serotypes have been identified) which can be distinguished biochemically from other species in the genus Salmonella.
Historically, Caldwall and Ryerson were the first to isolate the Salmonella-like organisms from diseased reptilesfrom the semiarid regions surrounding Tucson, Arizona. It is probable, however, that the bacteria were isolated earlier from poultry. The organisms now classified as S. arizonae were included in the genus Arizona, and have commonly been referred to as the Arizona group, Arizonas and paracolons. In 1982, the international subcommittee on taxonomy of Enterobacteriaceae decided that the Arizona group should be classed as two subspecies of the genus Salmonella based on the relatedness of their DNA with that of Salmonella sp.

Epidemiology

Incidence and distribution

In Turtles

Spices and herbs

A total of 160 samples of 55 different spices and herbs originating from six different suppliers and retailed at outlets in Vienna, Austria, were screened for their microbiological quality, based on plate count and selective identification techniques . Although max. values of total aerobic mesophilic bacteria of >107/g could be observed with some spices such as black pepper, chillies and China spice, more than 50% of the sample material contained between 104 and 106 c.f.u./g. Bacilli were found in nearly all samples, showing counts >105 c.f.u./g in almost 40% of the products. Enterobacteria, pseudomonades, and aeromonades as well as lactobacilli and enterococci were present mainly at levels of <102 c.f.u./g. Anaerobic sporeformers could be detected only three products.

One black pepper sample contained S. arizonae. Although several samples gave colonies indicating the presence of coagulase-positive staphylococci, representative isolates were not identified as Staphylococcus aureus. Some samplesespecially ginger and curry, had Bacillus cereus counts as high as 105 c.f.u./g; many others contained relatively low numbers of this species. Clostridium perfringens was detected in only one caraway sample (Kneifel-W and Berger-E, 1994).

In Sheep and Cattle

The laboratory infection and colonisation of S. arizonae in sheep with the development of mild proliferative rhinitis is reported. Specific pathogen-free Columbia yearling rams were divided into 2 groups and inoculated either with a culture of S. arizonae containing 1.6 X 108 CFU/ml isolated from a turbinate of a sheep with a proliferative rhinitis (Group 1: 5 lambs) or with a homogenate prepared from cultured frozen resected turbinates of this sheep (Group 2: 5 lambs).

Each lamb received 2 ml of the respective culture into each nostril; the mucosa of the right nostril of each lamb was brushed after inoculation. S. arizonae was isolated from the nares of all 5 lambs in Group 1 throughout the 6 month study period. In Group 2 lambs, S. arizonae was isolated consistently from only the right nostril of one lamb. S. arizonae was not isolated from faeces of any lamb in either group. One lamb from each group was killed on days 8, 28, 64, 118 and 184. Lambs with lower numbers of S. arizonae in nasal passages were killed first. No lesions were seen in the turbinates of any lambs until day 184.

Histological examination of any area of roughened mucosa on the cranial portion of the right turbinate (conchae) of one lamb in Group 1 detected a mild proliferative response with intracellular bacilli that were identified as S. arizonae (Brogden-KA et al., 1994).
Of 55 specimens submitted from aborting ewes during two lambing periods 25% were found to be due to infective agents and S. arizonae was isolated from 3 cases(Schweighardt-H et al., 1991).
In the spring of 1989 investigation of an abortion of triplet lambs which appeared to be a complication of pregnancy toxemia yielded a heavy pure culture of S. arizonae from the fetal tissues. No gross or histological lesions were found in these lambs. Investigation of the flock showed that 11 of the 40 ewes and one of the rams were persistent faecal shedders of S. arizonae (Pritchard-J, 1990).
Samples of faeces, jejunal lymph nodes, bile and carcass swabs from 264 slaughtered sheep were examined bacteriologically. Salmonella spp. were present in faeces from 14 sheep, lymph nodes from 10, and carcass swabs from one sheep. 25 of 26 isolates were S. arizonae and the remaining isolate was S. typhimurium. Precautions to prevent Salmonella contamination of sheep carcasses were listed (Jethon-H, 1990).

In Guineapigs

Meat Hygiene

Etiology

Classification

Since 1939, many attempts have been made to find a generally acceptable taxonomic position for this group of bacteria; Several classification systems have been used and a wide variety of names and designations has been applied to the organism.
Edward and associates established the biochemical and antigenic similarity of the arizonae and salmonellae. Enough differences were found between the groups, however to justify classification of the arizonae in a separae genus. Kaufmanna and Edwards first employed the name Arizona arizonae, which was also used by Ewing for members of the genus Arizona (Genus II of the tribe Salmonellae). A new type specicies name Arizona hinshawii had been proposed by Ewing to pay honour to the pioneering work of W.R Hinshaw on AA in turkeys, reptiles, and other animals.

Kauffmann subsequently included the arizonae in his subgenus III of the genus Salmonella, designating them S. arizonae and listing their antigenic formulas only in the simplified Kauffmann-White scheme. The arizonae have been classified in the Salmonella genus in 8th edition of Bergy's Manual and all organisms in the group are designated Salmonella arizonae.
Ewing and His Colleagues have further clarified the definition by which the biochemical and antigenic characteristics of members of the genus Arizona may be readily differentiated from other enterobacteriaceae. S. arizonae is the terminology used by Centers for Disease Control and Prevention.

Morphology and staining

Growth requirements

Biochemical properties. Cultures possessing the following biochemical characteristics are almost invariably classifiable serologically as members of S. arizonae:

• Dextrose Fermented with gas
• Lactose Fermented, as a rule slowly and promptly
• Sucrose Not fermented, as a rule
• Mannitol Fermented with gas
• Maltose Fermented with gas
• Dulcitol Not fermented
• Inositol Not fermrnted
• Indole Not produced, as aule
• Methyl red positive
• Voges-Proskauer Negative
• Jordan's tartarate No reaction
• Hydrogen sulphide Positive
• Urea Not hydrolysed
• Gelatin Liqified slowly
• Potassium cyanide Negative, as a rule
• Nitrates Reduced
• Motility Positive
• Betagalactosidase Positive
• Decarboylases
• Lysine Positive
• Arginine Positive, usually delayed
• Ornithine Positive
• Malonate Positive
• Phenylalanine Negative
• Deaminase

Most isolates from poultry, unlike salmonellae, ferment lactose usually within 7-10 days incubation. Failure of cultures to ferment dulcitol and inositol or to use D-tatarate, their slow liquifaction of gelatin, and their positive reaction in sodium malonate and betagalactosidase are most useful in distinguishing them from other members of the Salmonella group.
Citrobacter. For purposes of classification and identification, the S. arizonae must be differentiated not only from other Salmonellae, but also from the antigenically related genus Citrobacter of the tribe Salmonellae. Members of this genus are not known to be pathogenic for poultry, but from a diagnostic standpoint they may be confused with Salmonella cultures on initial isolation from fecal specimens. The former Bethesda-Ballerup "paracolons" (P.intermedium) are included in the genus Citrobacter along with cultures previously classified as Escherichia freundi.

Antigenic Structure

S. arizonae strains are related serologically to the salmonellae and other Enterobacteriaceae, and procedures for study and identification of their antigenic structure are identical to those for paratyphoid organisms. Thirty-four somatic (O) and 43 flagellar (H) antigens have been demonstrated.The serotype nomenclature system used in designating members of the genus salmonella has been applied to S. arizonae .

In writing antigenic formulas, commas are used to separate O antigen factors, a colon to distinguish the O and H antigens, commas to separate H antigenic factors within a single phase , and a hyphen or dash to separate the first phase from the second and the second from the third, etc. Thus the monophasic type species would be designated S arizonae 18:Z4,Z32. Evolution of the nomenclature for salmonellae has resulted in some confusion over the identification of strains. Two serotypes that were previously designated as 7:1,7,8 and 7:1,2,6 are now recognized as 18:Z4,Z32 and 18:Z4,Z23, respectively. Confusion also exist because, even though there are only 34 O and 43 H antigens, sometimes an isolate is designated as 65:Z52,Z53. The later is the designation that conforms to the system recognized by the Centers for Disease Control and Prevention , and the World Health Organization.

Drug resistance

Natural and experimental infection

Detection And Isolation

Detection by lectin

Detection by PCR

A polymerase chain reaction (PCR) assay with 2 nested pairs of primers selected from conserved sequences within a 2.3-kb randomly cloned DNA fragment from the S. typhimurium chromosome was developed. The nested PCR assay correctly identified 12 of 129 Salmonella strains belonging to subspecies I, II, IIIb and IV. One strain of S. arizona (ssp. IIIa) tested negative. No PCR products were obtained from any of the 31 non-Salmonella strains examined. The sensitivity of the assay was 2 c.f.u., as determined by analysis of proteinase K-treated boiled lysates of S. typhimurium. The performance of the assay was evaluated for environmental water, sewage and food samples spiked with S. typhimurium. Water and sewage samples were filtered and filters were enriched overnight in a non-selective medium. Prior to PCR, the broth cultures were subjected to a rapid and simple preparation procedure consisting of centrifugation, proteinase K treatment and boiling.

This assay enabled detection of 10 c.f.u./100 ml water with background levels of up to 8700 heterotrophic organisms per ml and 10 000 c.f.u. of coliform organisms per 100 ml water. Spiked food samples were analyzed with and without overnight enrichment in a non-selective medium using the same assay as above. Nested PCR performed on enriched broth enabled detection of <10 c.f.u./g food. Variable results were obtained for food samples examined without prior enrichment and most results were negative. It is concluded that this rapid and simple assay provides a sensitive and specific means of screening drinking water or environmental water samples, as well as food samples, for the presence of Salmonella spp (Waage-AS et al., 1999).

Prevention & control

Egg treatment

Immunostimulation

In a series of experiments, male broilers (experiments 1 and 2, n=576) were grown for 21 days on diets containing beta-hydroxy-beta-methylbutyrate (HMB) 0, 0.01. 0.05 and 0.10%. In experiment 3 (n=240), chickens were fed diets containing 0, 0.05, 0.075 and 0.10% HMB. HMB dietary supplementation did not significantly affect live weight gain in any experiment. However, a trend toward increased mean weight gain per bird was observed in experiments 1 and 3 when HMB was consumed at 0.10% of the diet. Mean feed to gain ratio was not affected by the inclusion of HMB. In experiment 3, HMB-supplemented diets did not affect bursa of Fabricius, thymus and spleen weights at 21 days of age. Cutaneous basophilic hypersensitivity response against pokeweed mitogen was higher (P_0.05) at 48 and 72 h post-injection in chickens fed 0.05% dietary HMB (experiment 1).

In experiment 2, this increase occurred 24 h post-injection in chickens fed HMB at 0.01% of the diet. On the contrary, the T-cell mediated response against PHA-P mitogen was comparable between all dietary treatments in multiple experiments. Macrophage function profiles were determined at 21 days of age. All chickens in experiments 1 and 2 on HMB-supplemented diets showed an increase in the recruitment of Sephadex-G50 R-elicited abdominal exudate cells (AEC). A 2-fold increase in AEC numbers occurred at 0.10% HMB (experiment 1, P_0.05). Although HMB supplementation did not significantly affect the phagocytic potential of the abdominal macrophages, nitrite levels in the macrophage culture supernatants were higher in 0.01 and 0.05% treatment groups compared with controls (experiment 2, P_0.04; experiment 3, P_0.05). HMB supplementation did not alter the ability to clear E. coli or S. arizona from the blood stream.

From 7 days' post-hatch, chickens were injected intravenously with a suspension of 7% sheep red blood cells. Serum samples were collected to determine primary and secondary antibody response. Chickens receiving 0.1% HMB diet in experiments 1 and 2 exhibited increased IgG and total anti-sheep red blood cell (SRBC) antibody levels during the primary response. During the secondary response, chickens consuming the 0.10% HMB diet had elevated IgM levels as well as increased total anti-SRBC levels over the controls in experiments 1 and 3. The study showed that HMB supplementation improves several immunological functions in young broilers, leading to decreased mortality (Peterson-AL et al., 1999).

Competitive Exclusion

Cultured Lactobacillus reuteri CCRC 14625 (9.25 X 108 c.f.u/ml) was repeatedly centrifuged and washed with H2O. The cell pellets were collected and incubated anaerobically in a 250 mM glycerol solution at 30°C for 2 h. GC-MS analysis indicated that glycerol was converted to reuterin by this process. The molecular weight of reuterin was estimated to be 74 (monomer) and 148 (cyclic dimer). The minimum inhibitory concentration (MIC), an indicator of the antimicrobial activity, was determined as the minimum concentration of the reuterin needed to inhibit the growth of various bacterial test strains.

The MIC (v/v) were: 0.5% for E. coli CCRC 12567, 0.8% for Bacillus subtilis CCRC 11408, 4% for Enterococcus faecalis 31, 4% for E. faecium 43, 1% for Enterobacter sakazakii CCRC 13988, 4% for Lactobacillus reuteri CCRC 14625, 4% for L. acidophilus Cl, 4% for L. rhamnosus GK-1, 0.8% for Salmonella choleraesuis subsp. arizonae CCRC 10742, and 0.6% for Shigella sonnei CCRC 15966, respectively. The IC50 (v/v) (the concentration of the reuterin required for 50% growth inhibition) for the reuterin preparation was 0.18% for colon adenocarcinoma, 0.17% for hepatocellular carcinoma, 0.18% for breast adenocarcinoma, and 0.08% for acute lymphoblastic leukaemia. A (10% v/v) reuterin was also bactericidal against E. coli CCRC 12567 (Chen-ChinChu et al., 1999).