Colorimetric Method of Loop-Mediated Isothermal Amplification with the Pre-Addition of Calcein for Detecting Flavobacterium columnare and its Assessment in Tilapia Farms

Flavobacterium columnare, the causative agent of columnaris disease in fish, affects many economically important freshwater fish species. A colorimetric method of loop-mediated isothermal amplification with the pre-addition of calcein (LAMP–calcein) was developed and used to detect the presence of F. columnare in farmed tilapia (Nile Tilapia Oreochromis niloticus and red tilapia [Nile Tilapia × Mozambique Tilapia O. mossambicus]) and rearing water. The detection method, based on a change in color from orange to green, could be performed within 45 min at 63°C. The method was highly specific, as it had no cross-detections with 14 other bacterial species, including other fish pathogens and two Flavobacterium species. The method has a minimum detection limit of 2.2 × 10² F. columnare CFU; thus, it is about 10 times more sensitive than conventional PCR. With this method, F. columnare was detected in gonad, gill, and blood samples from apparently healthy tilapia broodstock as well as in samples of fertilized eggs, newly hatched fry, and rearing water. The bacteria isolated from the blood were further characterized biochemically and found to be phenotypically identical to F. columnare. The amplified products from the LAMP–calcein method had 97% homology with the DNA sequence of F. columnare.

Received May 21, 2014; accepted August 10, 2014     

Effect of Nanophyetus salmincola and Bacterial Co-Infection on Mortality of Juvenile Chinook Salmon

The freshwater trematode Nanophyetus salmincola has been demonstrated to impair salmonid immune function and resistance to the marine pathogen Vibrio anguillarum, potentially resulting in ocean mortality. We examined whether infection by the parasite N. salmincola similarly increases mortality of juvenile Chinook Salmon Oncorhynchus tshawytscha when they are exposed to the freshwater pathogens Flavobacterium columnare or Aeromonas salmonicida, two bacteria that juvenile salmonids might encounter during their migration to the marine environment. We used a two-part experimental design where juvenile Chinook Salmon were first infected with N. salmincola through cohabitation with infected freshwater snails, Juga spp., and then challenged with either F. columnare or A. salmonicida. Cumulative percent mortality from F. columnare infection was higher in N. salmincola-parasitized fish than in nonparasitized fish. In contrast, cumulative percent mortality from A. salmonicida infection did not differ between N. salmincola-parasitized and nonparasitized groups. No mortalities were observed in the N. salmincola-parasitized-only and control groups from either challenge. Our study demonstrates that a relatively high mean intensity (>200 metacercariae per posterior kidney) of encysted N. salmincola metacercariae can alter the outcomes of bacterial infection in juvenile Chinook Salmon, which might have implications for disease in wild fish populations.

Received February 24, 2015; accepted September 7, 2015     

Gill Infection Model for Columnaris Disease in Common Carp and Rainbow Trout

Challenge models generating gill lesions typical for columnaris disease were developed for the fry of both Common Carp Cyprinus carpio and Rainbow Trout Oncorhynchus mykiss by means of an immersion challenge and Flavobacterium columnare field isolates were characterized regarding virulence. Carp inoculated with highly virulent isolates revealed diffuse, whitish discoloration of the gills affecting all arches, while in trout mostly unilateral focal lesions, which were restricted to the first two gill arches, occurred. Light microscopic examination of the gills of carp exposed to highly virulent isolates revealed a diffuse loss of branchial structures and desquamation and necrosis of gill epithelium with fusion of filaments and lamellae. In severe cases, large parts of the filaments were replaced with necrotic debris entangled with massive clusters of F. columnare bacterial cells enwrapped in an eosinophilic matrix. In trout, histopathologic lesions were similar but less extensive and much more focal, and well delineated from apparently healthy tissue. Scanning and transmission electron microscopic observations of the affected gills showed long, slender bacterial cells contained in an extracellular matrix and in close contact with the destructed gill tissue.

This is the first study to reveal gill lesions typical for columnaris disease at macroscopic, light microscopic, and ultrastructural levels in both Common Carp and Rainbow Trout following a challenge with F. columnare. The results provide a basis for research opportunities to examine pathogen–gill interactions.

Received January 10, 2014; accepted July 27, 2014     

Raised environmental temperature and food rationing as means of restricting growth of the replacement pullet

1. Four rearing temperature regimes (15, 20, 25, 30 °C) and three feeding schedules (ad libitum, restricting to the ad libitum intake of the 12th week and feeding 70% of this rate) were carried out with layer replacement pullets to 170 d of age. From this age, during lay, birds were kept at either 21 °C or on a 24‐h cycle of 21 for 18 h and 28 °C for the 6 h before lights out. Both a white and a brown egg‐laying strain were used.

2. Body weight at 169 d of age varied from, on average, 1409 g (15 °C, ad libitum) to 943 g (30 °C, 70% schedule) for the white strain and 1947 to 1250 g for the same treatments respectively for the brown strain. Sexual maturity was considerably delayed by the 70% feeding schedule, only slightly by rearing at 30 °C.

3. Rearing at 30 °C tended to depress subsequent egg output. The 70% feeding schedule at least maintained egg output compared with birds fed ad libitum in rearing.

4. There was a highly significant effect of temperature treatment during lay on food intake. The reduction in food intake due to the 21–28 °C cycle, however, appeared small.     

Improved Broth Microdilution Method for Antimicrobial Susceptibility Testing of Francisella Noatunensis Orientalis

In this project we optimized a minimal inhibitory concentration testing protocol for Francisella noatunensis orientalis. Thirty-three F. noatunensis orientalis isolates recovered from different fish species and locations were tested, and Escherichia coli ATCC 25922 was used as a quality control reference strain. A modified cation-adjusted Mueller Hinton broth supplemented with 2% IsoVitalex and 0.1% glucose (MMH) was tested at a pH of 6.4 ± 0.1, 7.1 ± 0.1, and 7.3 ± 0.1. Growth curves generated for F. noatunensis orientalis indicated that MMH at a pH of 6.4 ± 0.1 provided optimal growth. There were no significant differences in the growth curves obtained from isolates recovered from different fish species or from fresh or marine water. The pH of 6.4 ± 0.1 in the MMH media interfered with the inhibitory properties of the potentiated sulfonamides (ormetoprim-sulfadimethoxine and trimethoprim-sulfamethoxazole) when using the E. coli ATCC reference strain. Minimal inhibitory concentrations of eight antimicrobials (gentamicin, enrofloxacin, ampicillin, oxytetracycline, erythromycin, florfenicol, flumequine, and oxolinic acid) were similar for all F. noatunensis orientalis isolates. The in vitro susceptibility data provided here can provide a baseline for monitoring the development of antimicrobial resistance among F. noatunensis orientalis isolates, as well as provide valuable data in the development of potential therapeutics.

Received October 27, 2015; accepted April 13, 2016 Published online August 2, 2016     

Flavobacterium psychrophilum Infections in Salmonid Broodstock and Hatchery-Propagated Stocks of the Great Lakes Basin

Bacterial coldwater disease (BCWD), caused by Flavobacterium psychrophilum, threatens wild and propagated salmonids worldwide and leads to substantial economic losses. In addition to being horizontally transmitted, F. psychrophilum can be passed from infected parents to their progeny, furthering the negative impacts of this pathogen. In Michigan, both feral and captive salmonid broodstocks are the gamete sources used in fishery propagation efforts. A 5-year study was initiated to follow the prevalence of systemic F. psychrophilum infections in feral broodstocks of four species (steelhead Oncorhynchus mykiss [potadromous Rainbow Trout]; Coho Salmon O. kisutch; Chinook Salmon O. tshawytscha; and Atlantic Salmon Salmo salar) residing in three Great Lakes watersheds. Additionally, captive broodstocks of four species (Rainbow Trout, Brown Trout Salmo trutta, Lake Trout Salvelinus namaycush, and Brook Trout Salvelinus fontinalis) maintained at two facilities were assessed for the presence of F. psychrophilum. The resultant offspring from each broodstock population were sampled for F. psychrophilum infections multiple times throughout hatchery residency. Using selective flavobacterial culture and PCR confirmation, F. psychrophilum was detected in all broodstocks except the captive Lake Trout and Brook Trout. Logistic regression analysis demonstrated that among the infected feral broodstocks, Chinook Salmon from the Lake Michigan watershed had the highest prevalence of systemic F. psychrophilum infection (mean = 63.2%). Among the captive broodstocks, the Gilchrist Creek strain of Brown Trout had the highest infection prevalence (mean = 5%). Collectively, the captive broodstocks were found to have significantly lower infection prevalence than the feral broodstocks. Despite the high prevalence of systemic F. psychrophilum infections in many broodstock populations, the bacterium was rarely detected in their progeny during hatchery rearing. However, heavy losses associated with clinical BCWD outbreaks did occur. Collectively, our results reinforce that BCWD continues to threaten Great Lakes basin salmonids.

Received April 6, 2015; accepted August 25, 2015     

Use of Penicillin and Streptomycin to Reduce Spread of Bacterial Coldwater Disease II: Efficacy of Using Antibiotics in Diluents and During Water Hardening

Bacterial coldwater disease, caused by Flavobacterium psychrophilum, has lead to the loss of significant numbers of hatchery-reared salmonids. The bacteria can be spread from parent to progeny within contaminated sperm and ovarian fluid and can enter the egg during fertilization. The addition of antibiotics to diluents and water-hardening solutions could prevent the spread of the disease. In separate trials, a mixture of 0.197 mg/mL penicillin plus 0.313 mg/mL streptomycin was added to both a 0.5% sodium chloride fertilization diluent and hatchery well water during hardening. Tests showed that the addition of the antibiotics to the diluent and during up to 60 min of water hardening had no effect on the eye-up, hatch and deformity rates of Rainbow Trout Oncorhynchus mykiss eggs compared with the nonantibiotic-treated controls. Also, significant reductions in the prevalence of F. psychrophilum on the surface and inside eggs were observed when compared with controls. These results indicate that the addition of penicillin and streptomycin to diluents and during water hardening can prevent the vertical transmission of bacterial coldwater disease.

Received May 1, 2014; accepted July 10, 2014     

Spermatozoa DNA and plasma membrane integrity after pellet optimized processing for cryopreservation in meat type chicken breeders

1. Aim of this study was the development of an optimised cryopreservation pellet procedure for chicken semen and the assessment of DNA and membrane integrity in frozen/thawed spermatozoa in a Hubbard F15 meat type selected strain.

2. The following semen processing conditions were studied: spermatozoa working concentration (SWC), 1.5 vs 2 × 10⁹ cells/ml in pre-freezing extender; equilibration of diluted semen at 5°C, 20 vs 40 min; dimethylacetamide concentration, 6% vs 9%; dimethylacetamide equilibration time at 5°C, 1 vs 30 min; thawing at 60°C for 10 vs 50°C for 30 sec. Spermatozoa viability (EtBr exclusion procedure – stress test), mobility (Accudenz® swim-down test) and subjective motility were assessed in fresh and frozen-thawed semen.

3. The lower SWC (1.5 × 10⁹ cells/ml) and the higher dimethylacetamide concentration (9%) had positive significant effects on the recovery rate of motile (22% vs 16%) and viable spermatozoa (39 vs 34%), respectively.

4.Membrane (SYBR14-PI staining) and DNA integrity (comet assay) were assessed before and after freezing/thawing according to the optimised protocol.

5. Recovery rates of spermatozoa with undamaged plasma membrane and DNA were 41% and 76%, respectively. The distribution of spermatozoa in classes of DNA damage was also analysed and discussed.

6. It was concluded that pellet cryopreservation was a damaging process mainly for plasma membrane rather than nuclear DNA in chicken spermatozoa.     

Thermoregulation at high ambient temperature in genetically fat and lean broiler hens fed ad libitum or on a controlled‐feeding regime

1. Hens of lines divergently selected for fatness and leanness, fed either ad libitum or on a controlled regimen, were compared for susceptibility to heat stress.

2. The rate of increase in deep‐body (rectal) temperature during exposure to 32°C was used as the index of thermoregulatory ability. Comb and foot surface temperatures were measured as indicators of peripheral vasomotor tone.

3. Because body temperature control depends on the balance between heat production and heat loss, heat production was measured to determine whether fat line hens had a higher heat production, which they would then have to dissipate.

4. During the first hour of heat exposure, rectal temperature in the ad libitum‐fed birds increased twice as rapidly as in the corresponding lean line sample and 6 times more rapidly than in the control‐fed fat‐line group.

5. Surface temperatures of comb and foot increased from 27°C to 37°C within the first hour at 32°C, with no effect of either genotype or feeding regimen on rate of increase.

6. Heat production was unrelated to genotype but was reduced by 23% by controlled feeding, largely because of the reduction in body weight.

7. The results demonstrated that ad libitum‐fed fat‐line birds are susceptible to heat stress and that this is related not to increased heat production, but to a decreased ability to lose heat. Elevation of blood viscosity by plasma triglycerides is suggested as a causal factor.     

The thermoneutral temperature zone and seasonal acclimatisation in the hen

1. Oxygen consumption, body temperature, respiratory frequency and respiratory water loss of White Leghorn x Rhode Island hens were measured for short periods at six air temperatures between 2 and 32 °C. The hens were kept between tests in an open shed. The experiments were carried out over 3 years.

2. The upper critical temperature (Tcu) was estimated by the air temperature at which: 1, respiratory frequency increased above 60 respirations/min and 2, body temperature increased by 0.3 °C above that at the lower critical temperature. These responses to the test temperatures were examined as a function of the. acclimatisation temperature (Ta) represented by the mean daily temperature during experimental periods.

3. A seasonal change in Tcu was observed, which correlated with Ta(r = 0.836). The seasonal 10 °C change in the Ta brought about a 3 °C change in Tcu, compared with an 8.5 °C change in the lower critical temperature.

4. Thermoneutral temperature zone decreased with increasing Ta; the two critical temperatures tended to merge at a Ta of 32 °C. The latter probably represents an upper limit for acclimatisation to heat.     

Acceleration of post‐mortem changes in Tsaiya duck (Anas platyrhynchos) breast muscle by lactic acid marination

1. The effect of lactic acid marination at 5°C on post mortem changes in breast muscle pectoralis major of spent layer Tsaiya duck was studied.

2. Myofibrils were prepared from 0.1 M and 0.2 M lactic acid marinated muscle and control (non‐marinated samples) sampled at 0, 1, 3, 7 and 14 d post mortem.

3. Changes in myofibril fragmentation index (MFI), myofibrillar proteins and Z‐line structure were examined.

4. Marination of duck breast muscle in lactic acid at 5°C enhanced fragmentation of myofibrils and degradation of myofibrillar proteins and Z‐line structure as compared to control samples.

5. In summary, lactic acid marination at 5°C can accelerate the post mortem degradation of myofibrils in Tsaiya duck breast muscle.     

Optimum incubation conditions of the domestic guinea fowl egg

1. During artificial incubation of 8000 guinea fowl eggs, the effects of temperature, relative humidity, rate of inflowing air and age of the laying flocks were determined.

2. Total duration of incubation was divided into setter (0 to 24 days) and hatcher (25 to 28 days) periods. Eggs transferred at the end of the setter period, (ET, % of fertile eggs) and hatching rate (HR, % of transferred eggs) were calculated, as well as the total hatching rate expressed as percentage of fertile eggs.

3. In the range 36 to 39°C, temperature affected significantly (P< 0.001) ET and HR, with optima at 37.2 and 37.0°C ± 0.1°C respectively.

4. During incubating period, relative humidity in the range 40 to 64% (water vapour partial pressure, Ph₂o ⁼ 17 to 34 Torr at 36 to 39°C), significantly (P< 0.001) affected the total diffusive water loss and hatchling mass, both expressed as percentage of fresh egg mass. ET was significantly affected by water loss, the highest ET being for water loss of 13.3 ± 0.5%. Optimal relative humidity was calculated to be 48 to 52% (Ph₂o = 23 to 25 Torr at 37.2°G).

5. The rate of inflowing air significantly (P< 0.001) affected HR, with an optimum at 3.1 Lstpd/(h . egg).

6. The age of the laying flock significantly (P< 0.001) increased water loss; this was explained by a parallel increase of the mass specific shell conductance to water vapour.

7. Finally optimum incubation conditions were deduced, giving total hatching rates of 78 to 81% of fertile eggs, improving by 5 to 8% the best results obtained routinely in commercial practice.     

In ovo injection of ascorbic acid and higher incubation temperature modulate blood parameters in response to heat exposure in broilers

1. This study analysed whether in ovo injection of ascorbic acid before incubation and at high incubation temperature influenced blood characteristics and performance in broilers reared in different temperature conditions.

2. A total of 3,000 fertile eggs from broiler breeders (Cobb®) were randomly divided into three incubation treatments: no ascorbic acid injection and egg incubation at 37.5°C (control); no ascorbic acid injection and egg incubation at 39°C; in ovo ascorbic acid injection prior to incubation (6 µg AA/100 µl water) and egg incubation at 39°C.

3. Male chicks hatched from the three incubation treatments were submitted to three distinct rearing temperatures (control, cold and hot) from the third week of age onwards (540 chicks were divided into 6 treatments with 5 replicates per treatment).

4. Measurements at 42 d showed that, after egg incubation at 39°C, the haematocrit, haemoglobin values, ionised calcium and glucose concentrations were increased and base excess values were reduced. However, in ovo injection of ascorbic acid normalised all these parameters.

5. Partial CO₂ and O₂ pressure were higher with increased rearing temperature. Blood pH was lower when eggs were incubated at 39°C and injected with ascorbic acid. In ovo injection of ascorbic acid induced leucocytosis due to lymphocytosis and heterophilia, restored basophils rate and led to monocytopoenia. Leucocytosis was triggered by hot rearing temperature due to lymphocytosis, eosinophilia and heterophilia.

6. The results obtained in this study showed that in ovo injection of ascorbic acid before incubation may serve as a long-term stimulator and modulator of the broiler immune system, and that high incubation temperatures induce adaptations in the electrolytic balance, minimising or avoiding the occurrence of respiratory alkalosis under hot rearing temperature.     

Use of Penicillin and Streptomycin to Reduce Spread of Bacterial Coldwater Disease I: Antibiotics in Sperm Extenders

Bacterial coldwater disease caused by Flavobacterium psychrophilum has led to the loss of significant numbers of hatchery-reared salmonids. The bacteria can be spread from parent to progeny within contaminated sperm and ovarian fluid. Methods for disinfecting ovarian fluid and unfertilized eggs are available, but methods for disinfecting sperm have not been described. In this study we determined whether sperm extenders containing a mixture of penicillin and streptomycin can be used to eliminate F. psychrophilum. In vitro trials demonstrated that when Rainbow Trout Oncorhynchus mykiss sperm is mixed with an extender, a 15-min exposure to 0.197 mg penicillin plus 0.313 mg/mL streptomycin is effective at killing the bacteria and has no effect on sperm motility. Small-scale trials showed that egg fertilization rates were not reduced when sperm held in an extender solution containing the same antibiotic mixture for 15 min was used to fertilize eggs. Production-scale trials, however, showed a roughly 18% decrease in egg fertilization rate when sperm stored in an antibiotic containing extender was used. To determine why a reduction in fertilization capacity was observed, a small-scale experiment testing the fertilization of eggs with larger quantities of sperm was performed and showed that increasing the volume of sperm used did not increase fertilization rates. Our results demonstrate that extenders containing penicillin and streptomycin can be used to disinfect sperm, especially when small quantities of eggs are fertilized, but factors negatively affecting egg fertilization and survival on a production scale still need further investigation.

Received May 1, 2014; accepted August 10, 2014     

Thermal panting and respiratory alkalosis in the laying hen

1. Changes in respiratory rate (f), rectal temperature (T_r ) and blood acid‐base values were measured in laying hens exposed to ambient temperatures (T_a) of 32, 35, 38 or 41 °G.

2. At T_a 32 °G there was no panting. At T_a 35 °G panting occurred without any increase in T_r but there was a slight alkalosis (pH 7.55).

3. At T_a 38 °G T_r increased and panting was accompanied by moderate alkalosis (pH 7.58).

4. At T_a 41 °G T_r increased considerably and severe alkalosis developed (pH 7.65).

5. From the relation between T_r , f and pH it is concluded that some degree of alkalosis is a normal response to panting in the laying hen.     

Plumping fluid added to storage medium increases twofold the functional life span of fowl spermatozoa in vitro at 4°C

1. The objective of this study was to examine whether addition of plumping fluid (PF) to Lake's solution (LS) for storage of fowl spermatozoa in vitro at 4°C can prolong survival and improve the quality of spermatozoa.

2. In experiment 1, aliquots of spermatozoa were stored in vitro in LS alone and LS containing 10%, 25%, 50% and 75% (v:v) PF for 0.5, 24, 48, 72, 96 and 120 h at 4°C. After the end of each storage period, spermatozoa were evaluated for their viability, mobility and penetrability. Viability was determined using SYBR-14 and propidium iodide (PI) staining. Mobility was assessed using an Accudenz assay. Penetrability was assessed using spermatozoa-inner perivitelline layer (IPL) interaction assay.

3. In experiment 2, aliquots of spermatozoa were stored in vitro in LS alone and LS containing 25% and 50% (v:v) PF for 0.5, 24, 48 and 72 h at 4°C, and then fertility of the spermatozoa was evaluated using intravaginal artificial insemination (AI) in hens.

4. Storage of spermatozoa in LS alone resulted in loss of viability, mobility, penetrability and fertility within 48 h. In contrast, no loss of viability and penetrability was observed for the spermatozoa stored for 48, 96, 72 and 48 h in LS containing 10%, 25%, 50% and 75% (v:v) PF, respectively. In particular, fertilising capacity was not lost for the spermatozoa stored in the presence of 25% or 50% PF in LS for 48 and 24 h, respectively.

5. In conclusion, these findings demonstrated that in vitro exposure of fowl spermatozoa to PF during hypothermic storage in LS prolonged spermatozoa survival. A 25% (v:v) level of inclusion of PF in LS may be effective for the improvement of viability, penetrability and fertilising ability of the stored spermatozoa.     

Cooling of broiler hatching eggs during incubation

1. Broiler hatching eggs were cooled to 22°C for periods of 8 or 24 h on day 16 of incubation. Cooling had no significant effects on hatchability or chick quality compared with controls.

2. Cooling to 22°C for 24 h on days 14, 15, 16, 17 or 18 yielded similar results but cooling on day 13 increased late embryonic mortality.

3. When eggs were cooled to 22°C on day 16 of incubation there were no major differences in hatchability up to 48 h of cooling but chick quality deteriorated rapidly after 30 h and the limit for embryonic survival at this temperature was about 72 h.

4. There were no significant differences when eggs were cooled on day 16 and held for 24 h at 21.1, 23.9 or 26.7°C but hatchability and late deaths were significantly affected by cooling to 18.3°C.     

Effect of post‐mortem temperature on chicken M. pectoralis major: Isometric tension and pH profiles

1. The relationship between isometric tension development and pH, as a function of storage temperature between 0° and 40°C, was examined in chicken M. pectoralis major (PM) muscle during the critical 24 h post‐mortem period.

2. The muscle strips incubated at 0°C developed a peak isometric tension of 53.3 g/cm². This occurred after only 17 min incubation when the pH was 7.02, demonstrating the potential of chicken PM muscle to cold shorten. Peak isometric tension at 5°C was considerably lower than that generated at 0°C. However, as this occurred when the muscle pH was still high (6.70), this also indicated some potential to cold shorten at 5°C.

3. At 10° to 30°C, the muscle strips developed mean peak isometric tensions of 18 g/cm² after 6 h incubation by which time the muscle pH had fallen to 6.00, demonstrating a limited potential to rigor shorten. In contrast, those incubated at 40°C developed a peak tension of 54.5 g cm² after 75 min when the muscle pH was also around 6.00, thus indicating the potential for intensive rigor shortening at this temperature. Incubation temperature and the resultant muscle pH therefore determine the potential of chicken PM muscle to either cold shorten or rigor shorten.

4. Despite the differences found in isometric tension profiles, cooked meat texture after isometric tension measurement was not significantly different at any of the temperatures studied primarily because the muscle strips were essentially prevented from shortening.     

Genotypic and phenotypic diversity differences of presumptive commensal and avian pathogenic E. coli

1. The objective of the experiment was to characterise the genotypic and phenotypic differences between presumptive commensal E. coli and avian pathogenic E. coli (APEC) of poultry.

2. DNA was extracted from 65 confirmed APEC E. coli from chicken, 100 presumptive commensal E. coli from healthy turkey and 35 from healthy chicken. Enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and virulence factors genotyping was performed to characterise genetic features.

3. Carbon source utilisation and antimicrobial susceptibility tests were performed to characterise phenotypic features of isolates.

4. The genetic divergence between E. coli strains tested by ERIC-PCR profiles and virulence-associated genes showed a clear genetic separation between E. coli APEC and turkey E. coli strains.

5. The carbon utilisation profile of turkey isolates was different from chicken and APEC strains; whereas antimicrobial susceptibility was highest for turkey isolates (53%), and lowest for APEC strains (33.8%).

6. The study showed a significant negative correlation between utilisation of arabitol and adonitol with different virulence determinants tested, which suggests that the ability to utilise some uncommon carbon sources may be used to discriminate between presumptive commensal E. coli and APEC.     

Microbiological and shelf life assessment of chilled eviscerated whole chicken broilers in Saudi Arabia

1. The microbiological quality and shelf life of chicken carcasses marketed in Riyadh, Saudi Arabia were assessed.

2. The mean initial microbial counts (log₁₀ count/cm²) were 4·67, 4·14, 2·21, 2·78 and 2·96 for total aerobes, psychrotrophs, coliforms, Staphylococcus aureus and yeasts and moulds, respectively; these counts suggest a moderate level of contamination during processing.

3. Yeasts and moulds were present in relatively large numbers and constituted a considerable portion of the spoilage flora.

4. The mean shelf life of chicken broilers was 9·6, 6 and 4·4 d at 4, 7 and 10°C, respectively. Storage at 4°C resulted in better keeping than storage at 7°C or 10°C, while there was no significant difference between 7° and 10°C.

5. The initial totals of aerobes, psychrotrophs and yeasts and moulds were found to negatively correlate with shelf life.