Protection of chicks against salmonella infection induced by spray application of intestinal microflora in the hatchery

The efficacy of spray application of intestinal microflora of the adult bird in protection of broilers against salmonella infections was studied in 3 experiments under laboratory and field conditions. In chicks treated soon after hatching (in the hatchers at approximately 30 per cent hatch or in the chick delivery boxes in the hatchery) with crop-caecum homogenate or a mixture of aerobically and strict anaerobically cultured intestinal flora, very good results were achieved. A very obvious protective effect against a high infection dose (3 X 10(6) c.f.u.S. infantis bacteria per chick) and complete protection against natural infection with different salmonella types was induced. A significant improvement of growth rate was observed in broilers treated by spray in the hatchery and reared for 7 weeks under field conditions in an environment heavily contaminated with different salmonella types.     

Protection of chicks against Salmonella infantis infection induced by strict anaerobically cultured intestinal microflora

Summary

In two experiments newly hatched broilers were orally inoculated either with intestinal microfloras cultured under different conditions or with a suspension of intestinal homogenate and challenged with high doses (3 x 10⁵ c.f.u. per chick) of Salmonella infantis organisms.

Inocula were prepared from intestinal material of mature SPF WL hens under aerobic or strictly anaerobic conditions (< 5 ppm oxygen), and protected against atmospheric oxygen during storage. A very significant reduction of the number q/Salmonella infantis bacteria in caecal content was achieved one and two weeks after challenge in those groups of chicks which were inoculated with anaerobically cultured flora or with intestinal homogenate. In order to induce protection, obligate anaerobes had to be accompanied by facultative anaerobes. Amprolium or Monensin dit not interfere with protection. Protection induced by intestinal microflora depends on anaerobiosis during preparation, culturing, storage, and application of the protective bacteria. Starvation on the first day of life appeared to have a negative effect on protection.     

Reduction of salmonella infection of broilers by spray application of intestinal microflora: a longitudinal study

The Nurmi concept for diminishing salmonella infection of poultry by application of gastrointestinal microflora from adult birds into newly hatched chicks was tested in a field study in which 8 million broilers were involved. Half of the 284 flocks was treated in the hatchery by spray application of an undefined microflora propagated in SPF chicks. A significant reduction in salmonella-contaminated flocks, as well as in the number of infected broilers in these flocks was demonstrated. No undesirable side effects were detected. It was concluded that hygienic improvements in transportation and slaughtering of broilers are needed in order to maintain the reduced salmonella contamination rate.     

A questionnaire-based,retrospective field study of persistence of Salmonella Enteritidis and Salmonella Typhimurium in Danish broiler houses

A questionnaire-based, retrospective field study was conducted in 78 Danish broiler houses (analytical units) on 42 farms. In spring 1997, all these broiler houses had been infected with Salmonella Enteritidis, phage type 8, and/or Salmonella Typhimurium, definitive-type 66, by day-old chicks delivered from the same hatchery. Because these two salmonella types had not been detected in the Danish broiler sector before and the broiler houses were infected in a period of low salmonella prevalence, the study could focus on farm-related factors, without being distorted by other salmonella infections.

The same person visited all 78 broiler houses, and farm-related factors were listed both from personal observations and by interrogating the person in charge of attending to the broilers. Additional factors (noted at the visits) were recorded in a later telephone interview. Altogether, 187 variables were analyzed.

Broiler houses were divided into single-infection houses (which had one or both of the salmonella types in only one crop), and multiple-infection houses (where infection occurred in multiple crops). This was done both generally and separately for each of the salmonella types.

Factors associated with house status—but that were beyond the control of the farmer—were the salmonella type and the percentage of salmonella positive samples in the first crop. Factors which could be controlled by the farmer and which were associated with reduced prevalence of multiple-infection houses included antiseptic soap and water for washing hands in the anteroom, hygiene barriers when removing dead broilers, gravel alongside the broiler house, systematic checks of indoor rodent-bait depots, and combined surface and pulse-fogging disinfection.     

Influence of temperature, social, and dietary stress on development and stability of protective microflora in chickens against S. typhimurium

In the day-old chick, physiological stress in the form of high or low temperatures or feed and water deprivation either interfered with colonization or altered the protection provided by normal intestinal microflora against subsequent challenge with Salmonella typhimurium. The response was more obvious after individual exposure of chicks to salmonella than after exposure to infected seeders. Physiological and dietary stress at 2 weeks of age did not appreciably alter the S. typhimurium excretion pattern in treated chicks. The protective microflora appears to be quite stable under these stress conditions.     

Risk Factors Associated with Salmonella Status of Broiler Flocks Delivered to Grow‐Out Farms

In a prospective field observational study in the southeastern USA, we sampled gastrointestinal (GI) tracts from chicks of 65 broiler flocks delivered to conventional grow‐out farms for rearing. The flocks were hatched at seven broiler hatcheries. The mean within‐flock prevalence of Salmonella‐positive samples was 6.5% and ranged from 0% to 86.7%. Of the 65 flocks studied, 25 (38.5%) had at least one Salmonella‐positive sample. Accounting for confounding variability among the hatcheries and broiler companies, we tested whether the probability of detecting Salmonella in GI tracts of the chicks delivered was associated with certain characteristics of parent breeder flocks; hatchery production volume; hatchery ventilation system; hatchery egg‐room conditions; egg incubation, candling, hatching, eggshell and bird separation, and bird‐processing procedures; management of hatchery‐to‐farm transportation; day of week of hatch; weather conditions during transportation; or season of the hatch. Two risk factor models were adopted. The first model indicated that a greater number of parent flocks, manual separation of eggshell and bird, and a greater amount of fluff and feces on tray liners used during hatchery‐to‐farm transportation at delivery were associated with increased probability of detecting Salmonella in chick GI tracts, whereas a greater number of birds in the delivery vehicle was associated with decreased probability. The second model indicated that broiler flocks hatched on Tuesdays versus either Mondays or Thursdays (with no hatches on Wednesdays, Fridays or week‐ends), increased average hatchability of the eggs from the parent flocks, and greater amounts of fluff and feces on the transport tray liners at delivery were all associated with increased probability of detecting Salmonella in chick GI tracts. The results of this study suggest potential management decisions to lessen Salmonella contamination of broilers supplied by commercial hatcheries and areas for further research.     

Effect of Vaccination Against Coccidiosis in Combination with an Antibacterial Oregano (Origanum vulgare) Compound in Organic Broiler Production

Effects on production and health in broilers vaccinated against coccidiosis (Paracox®, Schering-Plough Animal Health) in combination with an antibacterial organic compound (Orego-Stim®, Meriden Animal Health Ltd.) were investigated. To obtain an environment and infection level similar to practical conditions, experimental floor pens were set up in a commercial broiler chicken house. Orego-Stim® supplement increased live weight and feed intake up to 48 days of age, whilst feed conversion ratio was not significantly affected. Vaccinated birds had a significantly higher live weight between 34 and 48 days, compared to non-vaccinated, but vaccination did not significantly affect feed intake or feed conversion ratio. Coccidial numbers in the litter were low, and only minor intestinal lesions were found. There were no significant differences in lesion scores between the different treatments. Orego-Stim® supplement decreased caecal numbers of Clostridium perfringens at 31 days, but not at 52 days, and numbers were not affected at all by anticoccidial vaccination. Bird health was overall good. The results indicate that vaccination against coccidiosis in combination with an in-feed antibacterial compound may be an alternative control method for intestinal health in organically produced chickens.     

Lysolecithin,but not lecithin,improves nutrient digestibility and growth rates in young broilers

ABSTRACT

1. The potential of lecithin and lysolecithin to improve lipid digestion and growth performance was investigated in three experiments: 1. an in vitro model that mimics the intestinal conditions of the chick, 2. a digestibility trial with chicks (5–7 days of age), and 3. a performance trial until 21 days of age.

2. In experiment 1, palm oil (PO), palm oil with lecithin (PO+L), and palm oil with lysolecithin (PO+LY) were subjected to in vitro hydrolysis and applied to Caco-2 monolayers to assess lipid absorption.

3. The in vitro hydrolysis rate of triglycerides was higher in PO+LY (k = 11.76 × 10³/min) than in either PO (k = 9.73 × 10³/min) or PO+L (k = 8.41 × 10³/min), and the absorption of monoglycerides and free fatty acids was highest (P < 0.01) for PO+LY. In experiment 2, 90 broilers were assigned to three dietary treatments: a basal diet with 4% palm oil, and the basal diet supplemented with either 250 ppm lecithin or lysolecithin.

4. ATTD of crude fat was higher in broilers supplemented with lysolecithin, but was lower in broilers supplemented with lecithin. DM digestibility and AMEn in birds supplemented with lysolecithin were significantly higher (3.03% and 0.47 MJ/kg, respectively).

5. In experiment 3, 480 broilers were randomly allocated to four dietary treatments: basal diet with soybean oil (2%), basal diet with lecithin (2%), soybean oil diet with 250 ppm lysolecithin, or lecithin oil diet with 250 ppm lysolecithin.

6. Lecithin diets significantly reduced weight at day 10 and 21 compared with soybean oil. However, the addition of lysolecithin to lecithin-containing diets significantly improved bird performance.

7. The results of these studies showed that, in contrast to lecithin, lysolecithin was able to significantly improve the digestibility and energy values of feed in young broilers.     

Effects of dietary supplementation with a herbal extract on the performance of broilers infected with a mixture of Eimeria species

1. A herbal extract containing a blend of three essential oils, derived from oregano, laurel leaf and lavender, was investigated as a feed additive alternative to the conventional anticoccidial sodium monensin.

2. Broilers were infected with a mixture of Eimeria species or left uninfected.

Both infected and uninfected broilers were provided with diets containing either herbal extract (HEX), monensin (MON) or without these supplements (CON). The HEX group had 50?mg herbal extract/kg diet and the MON group 100?mg monensin/kg diet.

3. All of the uninfected broilers exhibited higher body weight gain and better feed conversion when compared with their infected counterparts at d 28 and 42 of age. Both HEX and MON supplements caused significant improvements in performance in the infected broilers, but failed to have any effect on uninfected broilers.

4. Faecal oocyst output measured daily by sampling excreta, and expressed on a per bird basis, was lower in the HEX and MON groups than in the CON group. However, the herbal extract was not as effective as monensin in reducing oocyst excretion. Coccidial infection caused a significant increase in total intestinal length and caecal weight, but the dietary treatments did not influence these measurements.

5. These results indicate that providing a herbal extract in the diet was not as effective as monensin in protecting broilers exposed to a coccidial challenge.     

Comparison of the effect of live Newcastle disease vaccine Clone 30 in broilers administered at day 1 or at day 7 and the effect of H120 vaccination at 17 days of age: a field experiment

To analyse the results of a vaccination on the first day of age against Newcastle disease (ND) and on the 17th day of age against Infectious Bronchitis (IB) resp. with spray vaccines with Clone 30 and H120 vaccine. These vaccinations are compared in field circumstances with other vaccination methods. A serological examination and challenge test were used to be informed about the response and protection. From the present study the following conclusions can be drawn: Clear indications are obtained that following a spray vaccination against ND with Clone 30 vaccine of one-day-old broilers which possessed maternal antibodies, birds received a moderately good protection against ND, in spite of very low levels of HI antibodies. A spray vaccination against IB with H120 vaccine of broilers at 17 days of age gave some protection from two weeks after vaccination, however making a good conclusion about the protection is impossible and further investigation is required.     

Hatchery hygiene evaluation as measured by microbiological examination of samples of fluff

The microbial content of 1,180 samples of fluff from hatchers at 18 chick and 6 turkey hatcheries has been determined. The samples were taken (a) 12–15 hr before removal of birds from the hatchers, (b) just before or after theremoval of chicks orpoults, (c) after fumigation and (d) inreplicate from the same hatcher.

Fifty‐eight per cent of the samples of chick fluff contained one million or more bacteria but only 24 per cent contained more than 100,000 coli‐aerogenes bacteria. Moulds were not found in 68 per cent of the samples. Some samples of fluff were examined for coagulase positive staphylococci and fluorescent pseudomonads but these organisms were generally not detected. Bacillus cereus was recovered in very large numbers from a few samples. Bacterial and coli‐aerogenes counts from poult fluff were generally higher than those from chick fluff but the numbers of pseudomonads and coagulase positive staphylococci recovered were generally negligible.

In an intensive study of samples of fluff from one turkey hatchery, it was found that the level of contamination of fluff increased during hatching and the laying season and was accompanied by a decrease in poult quality. Sampling from any location in the hatcher appears to provide a representative indication of overall contamination at that time. Fumigation of fluff and debris generally caused a reduction in the level of contamination in the fluff.

A suggested assessment of hatchery hygiene based on the microbial examination of fluff is given and the advantages and disadvantages of this method are discussed.     

Validation of Fermentation and Processing Procedures for the Commercial-Scale Production of a Live,Attenuated Edwardsiella ictaluri Vaccine for Use in Channel Catfish Aquaculture

Abstract

Mortality associated with Edwardsiella ictaluri infection is a serious impediment to the commercial production of fingerling Channel Catfish Ictalurus punctatus. A patented, live, attenuated, orally delivered vaccine has been developed that offers exceptional protection against E. ictaluri infection in both laboratory and small-scale pond trials. Further vaccine development is contingent on the successful completion of large-scale field trials that accurately reflect industry conditions. This current work focuses on the validation of fermentation protocols and the optimization of downstream processing procedures to produce sufficient quantities of vaccine to conduct commercial-scale field trials. Eight vaccine serials were produced from a master seed stock (S97-773-340X2) in a 50-L floor model fermenter over two consecutive years. Following fermentation, cells were harvested, concentrated 10-fold, and cryogenically stored (?74°C). To assess processing protocols and determine shelf life of cryogenically stored vaccine, serials were tested for cell viability and vaccine potency at various intervals over 24 months. There were no significant differences in cell viability between the fresh vaccine and the stored frozen product. All serials provided a high level of protection (77–100% relative percent survival) against E. ictaluri infection in juvenile Channel Catfish and exhibited excellent poststorage viability. This data demonstrates that the live, attenuated, orally delivered vaccine can be stored at ?74°C for at least 2 years with no reduction in cell viability or vaccine potency.

Received May 17, 2016; accepted January 19, 2017 Published online April 4, 2017     

Inclusion of Bacillus amyloliquefaciens strain TOA5001 in the diet of broilers suppresses the symptoms of coccidiosis by modulating intestinal microbiota

Coccidiosis is an intestinal parasitic infection and one of the most prevalent and economically damaging diseases of chickens. Furthermore, coccidia‐induced mucogenesis promotes secondary colonization by Clostridium perfringens, a major pathogen of chickens that causes necrotic enteritis. Our previous work found that supernatant of a culture of Bacillus amyloliquefaciens strain TOA5001 (BA) inhibited the growth of C. perfringens on Gifu anaerobic broth medium. Accordingly, we evaluated the effectiveness of dietary BA administration in inhibiting C. perfringens colonization of the intestine in broilers that were experimentally infected with coccidia. Ten healthy broilers from a BA‐supplemented (2 × 10⁵ colony‐forming units/g of feed) broiler group and 10 from a non‐treated group were challenged with Eimeria tenella and E. maxima (5000 oocysts of each species/chick) at 28 days old. At 36 days old, five chicks from each group were slaughtered, whereas the remaining five in each group were killed at 49 days old. Dietary BA administration into Eimeria‐challenged birds reduced coccidial symptoms such as intestinal lesions. It also modified the cecal microbiota through suppressing C. perfringens and E. coli colonization, and inducing domination of Faecalibacterium prausnitzii, the Lactobacillus group and unknown Lachnospiraceae genera by bacterial DNA‐based metagenome analyses. B. amyloliquefaciens TOA5001 supplementation suppressed the symptoms of coccidiosis by modulating cecal microbiota in Eimeria‐challenged broilers.     

In vivo screening of four phytochemicals/extracts and a fungal immunomodulatory protein against an Eimeria acervulina infection in broilers

Background: Besides the anticoccidial drug resistance problem, increasing consumer concerns about food safety and residues have propelled the quest for alternative prevention and control strategies amongst which phytotherapy has gained appeal due to a renewed interest in natural medicine.Objective: The objective was in vivo screening of four phytochemicals/extracts and a fungal immunomodulatory protein (FIP) against an Eimeria acervulina infection in broilers.Animals and methods: Four phytochemicals/extracts (extract from Echinacea purpurea, betaine (Betain?), curcumin, carvacrol (two different doses)), and a recombinant FIP from Ganoderma lucidum cloned and expressed in Escherichia coli were investigated for their anticoccidial potential. The experiment was conducted in a battery cage trial with 54 cages of eight birds each. Broilers infected with E. acervulina (a low and high infection dose of 10⁴ and 10⁵ sporulated oocysts, respectively) and treated with the phytochemicals/extracts or the FIP were compared with broilers treated with the anticoccidial salinomycin sodium (Sacox®) and with an untreated uninfected and an untreated infected control group. Coccidiosis lesion scores, body weight gains and oocyst shedding were used as parameters.Results: The results showed a coccidiosis infection dose effect on the mean coccidiosis lesion scores. The phytochemicals/extracts and the FIP failed to reduce coccidiosis lesion scores and oocyst shedding, while salinomycin efficiently controlled the E. acervulina infection and enabled significantly higher body weight gains.Conclusion: In conclusion, the selected phytochemicals/extracts and the FIP did not reduce the lesions of an experimentally induced E. acervulina infection.     

Isolation and biological characteristics of multipotent mesenchymal stromal cells derived from chick embryo intestine

ABSTRACT

1. Over the past decade, rapid advancement in isolation methods for identifying markers of the once elusive intestinal stem cell (ISC) populations has laid the foundation for unravelling their complex interrelationships during homeostasis. Study on ISC in avian intestinal tissue might play a pivotal foundation for further studies on the epithelial-to-mesenchymal transition (EMT) in gastrointestinal disease and cell-based therapy as well as intestinal tissue engineering.

2. The following experiment isolated a population of fibroblast-like, plastic adhering cells derived from chick embryo intestine, showing a strong self-renewing and proliferative ability, which was maintained in vitro up to passage 25. The findings included growth characteristics, detected expression of cell surface markers and characterised the capability of these cells to differentiate towards the osteogenic, adipogenic, and chondrogenic cell lineages.

3. RT-PCR analysis showed that these cells from chick embryos expressed mesenchymal stromal cell markers CD44, CD90 and VIMENTIN as well as ISC-specific genes LGR5, MI1, SMOC2, BMI1, and HOPX. Immunofluorescence and flow cytometry confirmed this biology characterisation further.

4. In conclusion, cells were isolated from the intestine of 18-day-old chicken embryos that exhibited the biological characteristics of mesenchymal stromal cells as well as markers of intestinal stem cells. Our findings may provide a novel insight for in vitro cell culture and characteristics of ISCs in avian species, which may also indicate a benefit for obtaining cell source for intestinal tissue engineering as well as cell-based investigation for gastrointestinal disease and treatment.     

Loma sp. in Salmonids from the Eastern United States: Associated Lesions in Rainbow Trout

Abstract

A microsporidian of the genus Loma was noted in the gills of rainbow trout Oncorhynchus mykiss from a state hatchery (Buford Trout Hatchery) in Georgia. Mortalities of varying severity occur at this hatchery every fall, and the microsporidian was noted during an experiment from August 1991 to January 1992 to determine the effects of water source on disease. Infections first appeared to be systemic in the October sample; xenomas were observed in heart, spleen, and peripheral vessel walls. The presence of unidentified intracellular material preceded the appearance of xenomas in all tissues, but whether this material was associated with inflammation or represented immature stages of the parasite has yet to be determined. These structures were also noted in the intestine and liver, although xenomas were not noted in these organs. Mature xenomas did not elicit an inflammatory response but appeared to be short-lived. When the xenoma wall ruptured and released spores, an inflammatory response was again observed. The prevalence and severity of the infection were determined in fish maintained in troughs with well water, Chattahoochee River water, or hatchery (treated river) water. The infection tended to be more prevalent and more severe in fish maintained in the hatchery or river water than in those maintained in the well water. Stress induced by poor water quality may increase mortality from this parasite. This report extends the range of Loma sp. into the eastern United States.     

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

Summary

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:

(1) Hatchery waste eggs are highly contaminated (69.1%) and enterobacteriaceae predominate (26.6%).

(2) Chloramphenicol is the most effective antimicrobial tested.

(3) 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.

(4) Hatchability of either group of dipped eggs is reduced in comparison with fumigated eggs.

(5) 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.     

Comparison of two topical treatments on wound healing 7 days after disbudding of calves using thermocautery

ABSTRACT

Aims To compare wound healing 7 days after cautery disbudding of dairy calves treated immediately after disbudding with a traditional antimicrobial spray (OXY) or with a topical anaesthetic gel containing cetrimide, adrenaline and two local anaesthetics, lignocaine and bupivacaine (TA).

Method Eighty-one female dairy calves between 6–8 weeks of age were disbudded using a standard cautery disbudding protocol (sedation, cornual block, and analgesia), with complete removal of the horn bud. After disbudding, the wound on the right and left horn buds within each animal were randomly allocated to receive either OXY or TA. One week after disbudding, wounds were visually assessed for the presence of exudate, necrotic tissue, crust, or granulation tissue which were each assigned a grade from 1–3, where 1?=?no evidence, 2?=?moderate presence; and 3?=?marked presence.

Results At 7 days after disbudding, the prevalence of wounds with exudate or necrotic tissue was very low, and independent of treatment. The odds of a disbudding wound treated with TA being scored as having granulation tissue and was 5.2 (95% CI?=?1.72–15.7) times that of a wound treated with OXY. Conversely, the odds of wounds treated with TA being scored as having crusts was 0.18 (95% CI?=?0.06–0.57) that of a wound treated with OXY. No sign of infection was seen in any calves.

Conclusion and clinical relevance The greater prevalence of granulation tissue and reduction in crusting in disbudding wounds treated with topical anaesthetic gel compared to those treated with the antimicrobial spray suggests that use of the topical anaesthetic gel may speed wound healing compared to using antimicrobial spray. This should be further tested in a larger study undertaken over a longer period.     

Vegetative Bacillus amyloliquefaciens cells do not confer protection against necrotic enteritis in broilers despite high antibacterial activity of its supernatant against Clostridium perfringens in vitro

1. In this study, the effect of Bacillus amyloliquefaciens on Clostridium perfringens was tested in vitro and in vivo.

2. Using an agar well diffusion assay, the inhibitory activity of B. amyloliquefaciens supernatant was analysed against a large collection of netB-positive and netB-negative C. perfringens strains. Although strong growth inhibiting activity was detected against all C. perfringens isolates, it was significantly higher against virulent netB-positive C. perfringens strains compared with avirulent netB-negative isolates.

3. Subsequently, the efficacy of in-feed administration of lyophilised vegetative cells of B. amyloliquefaciens to prevent necrotic enteritis was tested in vivo using an established experimental infection model in broilers. Ross 308 broilers received either B. amyloliquefaciens supplemented or unsupplemented feed throughout the experiment. No significant differences could be detected between the untreated positive control group and the B. amyloliquefaciens treated group in body weight, the number of chickens that developed necrotic lesions and in pathological lesion scores.

4. These results demonstrate that despite its substantial inhibitory activity in vitro, lyophilised vegetative B. amyloliquefaciens cells had no beneficial effect against necrotic enteritis in the in vivo model used here.

     

Salmonellosis in Turkeys: Evaluation of Bacteriological and Serological Evidence of Infection

Bacteriological and serological studies were made of Salmonella infections in turkeys under three conditions; in commercial hatchery supply flocks and their embryo and poult progeny, in survivors of a natural infection over a ten-month period, and in experimentally infected adult turkeys.

The investigations revealed the following: 1. Carrier birds yielded Salmonella from cloacal swabs only intermittently which emphasized that failure to so isolate the organism did not necessarily indicate absence of infection in flocks. On the other hand, recovery of Salmonella from swabs or from dead-in-shell embryos foretold the likelihood of clinical disease in the progeny.

2. Bacteriological examination of ten per cent of the embryos from infected individual hatches did not regularly yield Salmonella but culture of a series of hatches revealed the disease status of the progeny.

3. The percentage of infected embryos in one hatchery which frequently disseminated S. typhimurium was estimated to range from two to twenty per cent in different hatches.

4. Environmental sanitation in the flock and hatchery appeared to be a major influence in the frequency of clinical outbreaks in the poults.

5. Two hatchery supply flocks were found to be infected with at least two serotypes; a third serotype appeared in the progeny of one in addition to the two in the parent flock.

6. Hatchery supply flocks with no history of clinical infection but having a latent infection transmitted the disease as readily as those which had survived clinical outbreaks.

7. The overall performance of the bacterial agglutination and hemagglutination tests demonstrated an equal but unsatisfactory degree of efficacy insofar as detection of infected birds was concerned. Both tests failed to identify half of the infected birds and also one infected flock.

8. A high incidence of non-specific reactions was observed in the hemagglutination tests of sera from non-infected birds. The number was greatly reduced by selecting strains whose extracts were more suitable for modifying erythrocytes. The possibility of the practical application of this test as a screening procedure would appear to depend on overcoming the problem arising from frequent non-specific reactions.