Bacterial identity and characteristics in healthy and unhealthy respiratory tracts of sheep and calves.

The aim of this study was to compare different bacteriological aspects of the respiratory systems of healthy (H) versus unhealthy (UH) animals with respiratory signs. The prevalence of different bacterial species was determined in the upper and lower respiratory tract of H and UH Najdi sheep, Somali sheep and Holstein calves. The characteristics of Pasteurella spp. isolates, and the biotype of Pasteurella haemolytica were identified in H and UH animals, Eighteen out of 28 (64.3%) of the identified bacterial species in the upper respiratory tract were more prevalent in the nasal cavities of UH Najdi and Somali sheep and Holstein calves with respiratory signs than in apparently healthy animals; four of the most prevalent bacteria in the upper respiratory system of UH sheep were Moraxella spp., Pseudomonas pseudomallei, Erysipelothrix spp., Pasteurella multocida, while three of the most prevalent bacteria in UH calves were Pasteurella haemolytica, Actinomyces spp., and Pseudomonas aeruginosa. The prevalence of six different bacterial species was greater in the lungs of UH animals, namely Actinomyces pyogenes, Erysipelothrix spp., P. haemolytica, Pasteurella ureae, Staphylococcus aureus, and Staphylococcus epidermidis, which could be risk factors in the complexity of the prevalent respiratory diseases of the animals surveyed. Of the biochemical, cytological and colonial characteristics studied in the identified P. haemolytica and P. multocida, two characters were significantly different (p < 0.05) in organisms isolated from UH as compared to those from H animals. These were the higher loss of haemolytic power by the strains of P. haemolytica and the decreased fermentation of trehalose by all the strains of P. multocida recovered from healthy animals. The only biotype of P. haemolytica isolated from H animals was biotype A, while both biotypes A (88.0% of the isolates) and T (12.0% of the isolates) were recovered from UH animals.

Subtyping of Bacillus cereus by total cell protein patterns and arbitrary primer polymerase chain reaction.

Bacillus cereus is a ubiquitous sporeforming Gram-positive rod that is associated with foodborne outbreaks as well as several opportunistic infections. Inspite of the prevalence of B. cereus associated foodborne outbreaks, subtyping of the species using molecular typing assays was not attempted. In this study we have recovered 58 B. cereus isolates from natural and clinical sources and initially characterized them, along with a B. cereus strain (ATCC 14579) and B. thuringiensis natural isolate, by biotyping, antibiotic susceptibility testing, and SDS-PAGE of total cell proteins. Our data have shown the existence of 1 biotype, 3 anti-biograms and 22 (38%) total cell protein patterns among the 58 B. cereus isolates. B. thuringiensis had a different protein pattern. SDS-PAGE of total cell proteins data denote clonal heterogeneity within B. cereus. Protein pattern 4 (pp4) was the most predominant with 13 isolates of B. cereus showing this pattern. Eight out of the 13 isolates with pp4 and one B. cereus strain (ATCC 14579) were further subtyped by using the arbitrary primer polymerase chain reaction (AP-PCR) assay. Eight (88.8%) different PCR patterns out of the 9 B. cereus isolates were obtained. Patterns obtained by SDS-PAGE of total cell proteins and AP-PCR were reproducible. These results indicate that SDS-PAGE of total cell proteins allows the differentiation among species within Bacillus and of strains within B. cereus. The typability of the method was 100% and the simpson's discrimination index of diversity was 98%. The utility of SDS-PAGE of total cell proteins in a pilot epidemiologic study was assessed and results obtained demonstrate its typing potential. AP-PCR allows further subtyping of the species. Both methods if used in conjunction may be useful for further clinical and epidemiologic studies of the spectrum of diseases caused by B. cereus.

Evaluation of bacterins containing three predominant phage types of Salmonella enteritidis for prevention of infection in egg-laying chickens.

Six Salmonella enteritidis bacterin formulations differing in adjuvant content and whole-cell inactivation procedures were evaluated in egg-laying chickens. Chickens given S enteritidis bacterins containing modified Freund's incomplete adjuvant had greater humoral immune responses to S enteritidis than did birds given other bacterin formulations (P < 0.05). Better protection against infection by S enteritidis phage types 8, 13a, and 23 was obtained in birds vaccinated with bacterin 5. Bacterin 5 contained S enteritidis cells inactivated by 20% acetone and modified Freund's incomplete adjuvant.

Evaluation of cell culture propagated and in vivo propagated hemorrhagic enteritis vaccines in turkeys.

The immune response of turkeys to a liquid, was compared with a previously frozen, cell culture propagated hemorrhagic enteritis (HE) vaccine. The liquid cell culture propagated HE vaccine was able to induce 100% seroconversion in turkeys 4 weeks after being vaccinated at 3.5 weeks of age; however, the previously frozen cell culture propagated HE vaccine induced 80% seroconversion 4 weeks post vaccination (P < 0.05). The average seroconversion in turkey flocks administered the liquid cell culture propagated HE was 97% in comparison with 98.5% in flocks given the splenic vaccine (P > 0.05). The complete absence of HE antigens in spleens of birds 5 days after being challenged with the virulent HE virus (40,000 TCID50 per bird) at an age of 9.5 weeks, was used as a model for successful protection against HE disease. The HE antigens were absent from spleens of all challenged birds that were previously vaccinated by the liquid cell culture propagated HE vaccine or splenic vaccine.

O-serotypes of Pseudomonas aeruginosa from animal and inanimate sources in Saudi Arabia.

A total of 1012 samples were examined for Pseudomonas aeruginosa and 257 (25.4%) were positive. The incidence of Ps. aeruginosa in samples collected from animal sources (N = 730) was significantly higher (28.7%) than that in 282 samples of inanimate sources (16.7%). The percentage of samples infected with these organisms was lowest in poultry feed (2.8%) and highest in sewage effluent (57.1%). Nine serotypes were defined from all sources. P5 was the common predominant individual O type in infected chicken navels and in the nasal cavities of Najdi sheep (a Saudi Arabian sheep breed), while P3 and P6 were predominate in the nasal cavities of Somali sheep (a breed imported from Somalia). No Ps. aeruginosa serotype was predominant in sheep faeces. In inanimate sources, P4 was predominant in water and sewage effluent. The isolate from the animal feed was untypeable. In using the slide agglutination technique for serotyping, most of the unusual agglutination reaction types of Ps. aeruginosa (70%) were of strains isolated from Somali sheep.

Mastitis in Camelus dromedarius in Saudi Arabia.

The correlation between camels' milk samples collected from abnormal inflamed udders and samples positive in the California Mastitis Test (CMT) was +0.803 (P less than 0.01). The bacterial count ranges of milk samples differed significantly (P less than 0.05) for those with a negative CMT and those with a positive CMT. Infection with many but not all bacterial species was associated with positive CMT results. The highest percentage of camel milk samples was included in the bacterial count range of 3.0 x 10(2) to 3.0 x 10(3) cfu/ml rather than in the greater than 3.0 x 10(3) cfu/ml range for most of the bacterial species. The most predominant bacterial isolates were Micrococcus spp., Staphylococcus aureus, Streptococcus spp. and Corynebacterium spp. followed by eight other flora. Chloramphenicol was the most effective antimicrobial agent of six tested against 118 bacterial isolates. Preliminary observations are made on chemotherapy of mastitis cases in camels.

Use of epidemiologic markers to identify the source of Escherichia coli infections in poultry.

During an epidemiologic study of poultry colisepticemia on 2 Saudi Arabian poultry broiler farms, Escherichia coli was isolated from 101 (40.4%) of the 250 specimens examined. The antigenic structure and the drug resistance pattern of 65.4% of the E coli isolates from different sources were used as epidemiologic markers to trace the source of the infection. The predominant E coli serotypes involved in infections of 2 poultry broiler progeny farms were 033:H4 (51.8%) and 078:H- (19.6%) that had the following respective drug resistance patterns: furazolidone-streptomycin-sulfathiazole and streptomycin-sulfathiazole-tetracycline. Escherichia coli strains with typical epidemiologic markers were isolated from various sources on a broiler breeding farm, but not from well waters of the infected progeny farm. Three other E coli serotypes (045:H10[14.3%], 0119:H27[1.8%], and 0145:H25[1.8%]) were involved in poultry infection, but to a lesser extent. These 3 serotypes were multiply resistant against 5 to 6 of the antimicrobials evaluated.

Production of H2S by Escherichia coli isolated from poultry: an unusual character useful for epidemiology of colisepticemia.

Eleven isolates of H2S-producing Escherichia coli were recovered from necropsy materials of chickens with symptoms and lesions of colisepticemia on Saudi Arabian broiler farms. Results of 19 out of 20 biochemical reactions studied were typical for E. coli. Hydrogen sulfide production by the E. coli isolates was used as an epidemiological marker to pinpoint a breeding farm as the probable source of these strains, which were then transferred to progeny farms, where colisepticemia occurred. This finding was confirmed by the presence of the same antigenic structure (O78:H-) and by the same drug-resistance pattern (a multiple resistance to streptomycin, sulfathiazole, and tetracycline) in the isolates.

Reduction of bacterial infections in newly hatched chicks by the use of antimicrobial dips: preliminary approaches.

Bacteriological examination of hatchery waste eggs, identification of the isolated bacteria, and susceptibility testing against seven antimicrobial agents were used in an attempt to establish a rational basis for reducing bacterial infections in newly hatched chicks. Chloramphenicol at 1000 ppm was selected as the antibiotic for preliminary dipping trials and 0.45% iodophore (Wescodyne) was added for later trials. The control treatment consisted of formaldehyde fumigation. The following conclusions can be drawn: Hatchery waste eggs are highly contaminated (69.1%) and enterobacteriaceae predominate (26.6%). Chloramphenicol is the most effective antimicrobial tested. Dip treatments with either chloramphenicol alone or chloramphenicol plus Wescodyne result in a reduced percentage of abnormal navels (8.4% and 10.4%), as compared with 21.9% for the control treatment. Hatchability of either group of dipped eggs is reduced in comparison with fumigated eggs. Dip treatment with chloramphenicol plus Wescodyne significantly reduces the anal carrier rates for Escherichia coli, Pseudomonas aeruginosa, Salmonella spp., and Staphylococcus aureus. This treatment reduces the incidence of bacterial infection in abnormal navels to zero.

Effect of management changes on hatchability and bacterial infections on a Saudi Arabian poultry breeding farm.

A new management programme for improvement of hatchability and reduction of transmission of bacterial potential pathogens from breeders to hatchery products was applied on a Saudi Arabian poultry farm. The technical changes included better nest engineering, improved cleaning of hatching eggs and better criteria for selecting eggs for incubation. The new nests led to a change in laying behaviour with 86.6% of eggs layed in nests which was significantly higher than was achieved previously. Bacterial penetration in dirty floor eggs was significantly higher than in clean nest eggs. The average hatchability was significantly higher for the flock under the new management than for the same age flock under the old management. The new management reduced transmission of paratyphoid organisms and Escherichia coli from infected breeders to their hatchery products. The old and new managements differed in the percentage of products infected with Salmonella.

A non-motile Salmonella mutant (6, 7:-:-) and two serological variants encountered in Saudi Arabia.

A non-motile mutant of Salmonella group C1 and two serological variants were isolated during the performance of a national Salmonella surveillance program in Saudi Arabia. Most of the biochemical reactions of the strains conformed to those of the genus Salmonella except for some atypical reactions. The three strains exhibited different susceptibility patterns to 21 antimicrobial agents. The serological analysis revealed the following antigenic structures: 1, 4, 12, 27:b: - Salmonella sofia var. monophasic; 1, 42:z4, z23: - Salmonella gera var. monophasic and a Salmonella strain reported as a non-motile strain of group C1 by the World Health Organization International Collaborating Center for Reference and Research on Salmonella.

Distribution of paratyphoids on Saudi Arabian poultry farms and pathogenicity studies of predominant serotypes.

A total of 412 feed samples and 632 litter samples from 15 poultry farms (2 breeding farms and 13 rearing farms) were examined for salmonella. Twelve of these farms had salmonella in litter, five farms had salmonella in the feed and four had salmonella in both feed and litter. Seventeen feed samples (4.13%) and 121 litter samples (19.15%) were contaminated with salmonella. Sixteen salmonella serotypes were encountered, of which six were found in both feed and litter. Salmonella concord and S. livingstone were present in the litter of one breeding farm and its progeny farms. The five most frequently isolated salmonella serotypes in feed and litter were S. concord (17.39%), S. coeln (15.94%), S. livingstone (15.22%), S. manhattan (11.59%), and S. paratyphi B var. java (8.69%). The pathogenicities of those serotypes were determined by calculating their median lethal doses (LD50) 24 and 48 hr postinjection of 1,050 one-day-old broiler chicks via the navel into the yolk sac. The composite 48-hr LD50s (viable cells) were: S. concord, less than 8.8 X 10(3); S. livingstone, 1.1 X 10(5); S. manhattan, 3.5 X 10(5); S. coeln, 1.25 X 10(7); and S. paratyphi B var. java, 1.73 X 10(7).

Occurrence of salmonellae in animal feed ingredients in Saudi Arabia.

Of 346 samples of animal feed ingredients (144 complete feed, 88 feed concentrate, 42 calcium carbonate, 30 meat meal, 29 fish meal, and 13 mixed meat and bone meal), 42 (12.13%) were contaminated with salmonellae. The rate of Salmonella contamination varied according to the nature of the feed sample. Seven (53.84%) of the mixed meat and bone meal, 14 (48.27%) of the fish meal, 12 (13.63%) of the feed concentrate, 7 (4.86%) of the complete feed, and 2 (4.76%) of the calcium carbonate samples contained salmonellae. The 49 Salmonella isolates represented 18 serotypes; the 5 most frequent serotypes in decreasing order were S lille, S new-haw, S livingstone, S kentucky, and S typhimurium. The 4 most common somatic serogroups encountered were C1, E2, B, and C3.

Isolation of salmonella and some other potential pathogens from two chicken breeding farms in Saudi Arabia.

Different salmonella serotypes were recovered from various sources in meat (A) and layer (B) poultry-breeder farms of Saudi Arabia. On farm A, salmonella were recovered from intestines of poultry breeders (7.41%), inshelled chicken embryos (2.87%), day-old chicks (19.23%), and mice (21.43%). Salmonellae in shell contents and on shell surfaces of hatching eggs contaminated the same percentage of eggs (1.24%), and litter and feed contaminations were 4.0% and 1.67%, respectively. On farm B, salmonella were recovered from intestines of poultry breeders (27.59%), mice intestines (15.0%), shells of hatching eggs (2.48%), hatching egg contents (0.35%), litter (33.33%), and feed (5.0%). No salmonella were recovered from intestines of inshelled chick embryos or day-old chicks on farm B. The total number of salmonella serotypes isolated from both farms was 14, and the serotypes common to both farms were Salmonella concord and S. livingstone. Salmonella contamination of shells of hatching eggs, pooled from both farms, was higher for dirty floor eggs than for clean eggs collected from floors and nests. The percentages of hatching eggs penetrated by bacteria other than salmonella on farms A and B, respectively, were: dirty floor eggs (25.53%, 22.83%), clean floor eggs (2.67%, 2.06%) and clean nest eggs (0.00%, 1.08%). The bacterial flora isolated from egg contents of unincubated hatching eggs (in decreasing order of frequency) were Staphylococcus, coliforms, Streptococcus, Bacillus, Salmonella, and Proteus.

Salmonella species and serotypes isolated from farm animals, animal feed, sewage, and sludge in Saudi Arabia.

A total of 264 salmonellae representing 65 different species and serotypes were isolated for the first time in Saudi Arabia, from various animal species, animal feed, sewage, and sludge. The six most frequently isolated Salmonella species or serotypes were: livingstone, concord, "S. schottmuelleri" (invalid), lille, S. typhimurium, and cerro.

In vitro susceptibility of salmonellae to eight antimicrobial agents.

The Bauer-Kirby disk diffusion method was used to determine the susceptibility of 333 strains of Salmonella isolated in Saudi Arabia to eight antimicrobial agents. The following susceptibilities were observed: Cephalothin 99.40%, furadoine 99.10%, chloramphenicol 98.80%, ampicillin 98.20%, polymyxin 96.70%, kanamycin 95.80%, streptomycin 90.99%. Only 50.45% of the strains were susceptible to tetracycline, whereas 44.14% and 5.41% were respectively intermediate and resistant to this drug. The antibiograms displayed many resistant patterns. Some of the salmonellae were singly resistant while others were resistant to 2 or more drugs in various combinations. In view of the potential for salmonellae to have inherent or acquired resistance to certain commonly used antimicrobial agents, susceptibility of all isolates of salmonellae, particularly of all clinically significant strains should be determined periodically.

Salmonella serotypes encountered in animal feed additives in Lebanon.

Animal feed-additive samples (n = 300) were examined for the presence of salmonellae, using the selenite-F broth-enrichment method followed by subculturing on Salmonella-Shigella and brilliant green agar with sulfadiazine selective agar plates. Samples consisted of a variety of feed additives: 119 bone meal samples, 77 meat meal samples, 40 fish meal samples, and 64 miscellaneous meal samples. Results of examination found 49 (41.2%) of the bone meal samples, 6 (7.8%) of the meat meal samples and 2 (5%) of the fish meal samples contained salmonellae. Of 57 isolates representing 24 serotypes, 4 most frequently isolated serotypes were Salmonella meleagridis (35.1%), Salmonella tennessee (7%), Salmonella chester (5.2%), and Salmonella senftenberg (5.2%). This study shows a high Salmonella-contamination rate of bone meal compared with meat meal and fish meal samples. Of 12 known positive bone meal samples that were examined, 100% of 25-g samples, compared with 70% to 100% of 2.5-g samples and 30% to 90% of 0.25-g samples and 30% to 90% of 0.25-g samples, were positive for salmonellae.

Isolation, purification and characterization of a pyocin.

Seventeen strains of P. aeruginosa were tested for pyocin production using the broth method. Mitomycin C was used for the induction of pyocin in TSB shaked cultures incubated at 32 degrees. The highest pyocin activity was produced by strain 14. TSB shaked cultures in fluting flasks were found to be superior to Erlenmeyer flasks for pyocin production. Pyocin 14 was purified by chemical and chromatographic methods. It was found to be protein in nature and free from carbohydrates. It is stable between pH 5 and 9 and is partially inactivated when kept for one month at 4 degrees. Freezing and thawing, lyophilization and heating at 65 degrees for 5 min completely destroyed its activity. The addition of 4 mg/ml bovine albumin or 2 mg/ml gelatin caused about 60% increase in activity. However, treatment with 10 mM EDTA caused a 98% drop in its activity.