A.P. Berisvil

In vitro and in vivo screening of native lactic acid bacteria toward their selection as a probiotic in broiler chickens.

Among 360 isolates from the gastrointestinal tract (GIT) of broilers, eleven isolates which showed in vitro probiotic properties were identified and selected for further tests. After the in vitro screening, three strains were chosen for the in vivo study of persistence of fresh cultures and then one strain was selected for the in vivo study of persistence of lyophilized culture. Lyophilized Lactobacillus salivarius DSPV 001P was capable of persisting in broilers during a complete rearing, even 28 days following cessation of administration. L. salivarius DSPV 001P administered to broilers and recovered from GIT was compared by pulsed-field gel electrophoresis (PFGE) to ensure that the same genotype was persistently identified. A combination of in vitro and in vivo screening of native lactic acid bacteria (LAB) described in this study may offer a method for selecting the most suitable strain for potential application as a broiler probiotic supplement.

Probióticos en pollos parrilleros: una estrategia para los modelos productivos intensivos ☆

The broiler industry has become an important economic activity in Argentina. Global production of broiler meat has been growing in Argentina faster than for any other meats, possibly due to declining poultry prices and increasing incomes. Modern rearing systems can produce broilers ready to slaughter in 50 days, with the required 2.7kg of weight and a feed conversion of about 1.6kg feed/kg of meat. Nevertheless, broilers raised under these intensive conditions are exposed to various stressors every day. For many years, feed supplementation with antibiotics was widely used to stabilize the gut flora, improve general parameters and prevent avian diseases. However, the utility of antibiotics has been questioned because of the emergence and spread of antibiotic-resistant bacteria in meat. Therefore, there is a renewed interest in finding viable alternatives to antibiotics. One potential method is the supplementation of broiler diets with probiotics. This review provides an updated summary of the use of probiotics to improve sanitary conditions and enhance performance in broilers, demonstrating the role of probiotics as a reliable option to replace antimicrobial growth promoters.

Quantification of FITC-labelled probiotic Lactobacillus salivarius DSPV 001P during gastrointestinal transit in broilers

The knowledge related to the fate of probiotics in the complex environment of the intestinal microbiota in broilers is just beginning to be elucidated; however, it is not yet well understood. A good method to investigate the mechanisms by which probiotics mediate their effects is to mark probiotic bacteria and trace them. The aim of this research was to develop a new method to estimate in vivo fluorescein isothiocyanate (FITC)-labelled Lactobacillus salivarius DSPV 001P counts during passage through the gastrointestinal tract (GIT) of broilers. Forty-five, 1 d old Cobb broilers were used in this trial. Programmed necropsies were performed 30 min, 6 h, and 12 h after the administration of the probiotic bacterium, and samples of liver, crop, duodenum, caecum, and bursa of fabricius were collected. To determine the spatial and temporal transit of L. salivarius DSPV 001P in broilers, the number of bacteria as well as its respective fluorescent signal produced by FITC were measured. In order to observe the relationship between the variables, a logistic regression analysis was applied. The amount of fluorescence could be used as an indicator of fluorescent probiotic bacteria in the crop and duodenum 30 min after probiotic bacterium supplementation. In addition, the fluorescent signal could be used to estimate bacterial counts in caecum 6 and 12 h after L. salivarius DSPV 001P administration. To the best of our knowledge, this research is the first in vivo trial to employ the bacterial FITC-labelling technique in order to enumerate probiotic bacteria during gastrointestinal transit in broilers.

Genetic diversity of thermotolerant Campylobacter spp. isolates from different stages of the poultry meat supply chain in Argentina

The objective of this study was to investigate a clonal relationship among thermotolerant Campylobacter spp. isolates from different stages of the poultry meat supply chain in Argentina. A total of 128 thermotolerant Campylobacter spp. (89 C. jejuni and 39 C. coli) isolates from six poultry meat chains were examined. These isolates were from: a) hens from breeder flocks, b) chickens on the farm (at ages 1 wk and 5 wk), c) chicken carcasses in the slaughterhouse, and d) chicken carcasses in the retail market. Chickens sampled along each food chain were from the same batch. Campylobacter spp. isolates were analyzed using pulsed-field gel electrophoresis to compare different profiles according to the source. Clustering of C. jejuni isolates resulted in 17 profiles, with four predominant genotypes and many small profiles with just a few isolates or unique patterns, showing a very high degree of heterogeneity among the C. jejuni isolates. Some clusters included isolates from different stages within the same chain, which would indicate a spread of strains along the same poultry meat chain. Moreover, twenty-two strains of C. coli clustered in seven groups and the remaining 17 isolates exhibited unique profiles. Evidence for transmission of thermotolerant Campylobacter spp. through the food chain and cross contamination in the slaughterhouses were obtained. This collective evidence should be considered as the scientific basis to implement risk management measures to protect the public health.

Probiotic effect on calves infected with Salmonella Dublin: haematological parameters and serum biochemical profile

The aim of this study was to evaluate the effect of a probiotic/lactose inoculum on haematological and immunological parameters and renal and hepatic biochemical profiles before and during a Salmonella Dublin DSPV 595T challenge in young calves. Twenty eight calves, divided into a control and probiotic group were used. The probiotic group was supplemented with 100 g lactose/calf/d and 1010 cfu/calf/d of each strain of a probiotic inoculum composed of Lactobacillus casei DSPV318T, Lactobacillus salivarius DSPV315T and Pediococcus acidilactici DSPV006T throughout the experiment. The pathogen was administered on day 11 of the experiment, at an oral dose of 109 cfu/animal (LD50). Aspartate aminotransferase (AST), gamma glutamyl transpeptidase (GGT), urea, red blood cells, haemoglobin, haematocrit, mean cell haemoglobin (MCH), mean corpuscular volume, mean corpuscular haemoglobin concentration (MCHC), white blood cells, lymphocytes, neutrophils, band neutrophils, monocytes, eosinophils, basophils and the neutrophils/lymphocytes ratio were measured on days 1, 10, 20 and 27 of the experiment. In addition, animals were necropsied to evaluate immunoglobulin A (IgA) production in the jejunal mucosa. The most significant differences caused by the administration of the inoculum/lactose were found during the acute phase of Salmonella challenge (9 days after challenge), when a difference between groups in neutrophils/lymphocytes ratio were detected. These results suggest that the probiotic/lactose inoculum administration increases the calfs ability to respond to the disease increasing the systemic immune response specific. No differences were found in haemoglobin, haematocrit, MCH, MCHC, AST, urea, GGT, band neutrophils, eosinophils, monocytes and IgA in the jejunum between the two groups of calves under the experimental conditions of this study. Further studies must be conducted to evaluate different probiotic/pathogens doses and different sampling times, to achieve a greater understanding of the effects of this inoculum on intestinal infections in young calves and of its mechanism of action.

Development and storage studies of high density macrocapsules containing Lactobacillus spp. strains as nutritional supplement in young calves

The aim of this study was to evaluate different production methodologies of probiotic macrocapsules with high bacterial densities destined to lactating calves. Three types of capsules containing Lactobacillus casei DSPV318T and Lactobacillus plantarum DSPV354T were prepared from an overnight culture in whey medium: (1) mixing the culture with calcium alginate and then, reincubating the capsules in whey (RC); (2) concentrating the biomass by centrifugation and mixing the pellet with calcium alginate (CC) at different concentrations with respect to the initial culture (5X and 12.5X); (3) CC with cryoprotectants: whey permeate (Per) and glycerol (Gly). Chitosan coating was evaluated. Capsules were freeze-dried and viability was assessed before freezing, after freeze-drying and every two weeks for 84 days of storage at room temperature, 4°C and -20°C. CC showed higher cell densities than RC. Storage temperature affected viability: greater viability at lower temperature. Moreover, the effect of temperature was influenced by other factors, such as capsule coating, culture neutralization and cryoprotectants. Coating improved viability at room temperature; however no effect was observed at 4°C. Culture neutralization allowed greater survival during storage. Cryoprotectants improved viability during freezing, but they also generated a positive or negative effect depending on storage temperature. The best results were: at refrigeration Gly12.5X exhibited counts above 109CFU/capsule until day 70 and Per12.5X until day 56 of storage and at -20°C Gly12.5X showed counts above 109CFU/capsule until the end of the study (84 days). A 109CFU capsule is the daily dose per calf which would facilitate the administration of this probiotic inoculum to field animals.

Presencia de serotipos de Campylobacter jejuni O:19 en la cadena avícola Argentina

Thermotolerant species of Campylobacter have been focus of attention in the last years because they are the major agent causing zoonotic foodborne diseases. In addition, Campylobacter jejuni O:19 serotype was associated with Guillain Barré syndrome. The aim of this study was to determine the proportion of C. jejuni O:19 serotype isolated at different stages of three poultry meat supply chain in Santa Fe, Argentina. The analysis showed that 18% of isolated C. jejuni belong to serotype O:19. It was also determined that the presence of these strains is given in almost all production stages. These results reflect a significant risk to public health of consumers. Epidemiological studies of Campylobacter should be considered to establish a risk manager policy.

Impact of lyophilized Lactobacillus salivarius DSPV 001P administration on growth performance, microbial translocation, and gastrointestinal microbiota of broilers reared under low ambient temperature

This study was undertaken with the aim of investigating the effects of dietary supplementation of probiotic strain Lactobacillus salivarius DSPV 001P on growth performance, microbial translocation, and gastrointestinal microbiota of broilers reared under low ambient temperature. Two hundred and forty, one-day-old male Cobb broilers were randomly distributed into two treatment groups, a probiotic group and a control group, with four replicates per treatment and 30 broilers per replicate. The temperature of the broiler house was maintained at 18-22°C during the first three weeks, after which the temperature was at range of 8°C to 12°C. The results showed that probiotic treatment significantly improved body weight of broilers when compared with the control group. After 42days, the weight means were 2905±365.4g and 2724±427.0g, respectively. Although there were no significant differences, dietary inclusion of L. salivarius tended to increase feed intake and to reduce feed conversion ratio during the six-week experimental period. Similarly, supplementation tended to reduce the rate of mortality, with 12 deaths occurring in the probiotic group, and 20 in the control group. However, no differences were observed in intestinal bacterial concentrations of Enterobacteriaceae, E.coli, and lactic acid bacteria in both crop and caecum among treatments. Through our study, it appears that L. salivarius DSPV 001P was non-pathogenic, safe and beneficial to broilers, which implies that it could be a promising feed additive, thus enhancing the growth performance of broilers and improving their health.

Development of cecal-predominant microbiota in broilers during a complete rearing using denaturing gradient gel electrophoresis

Understanding of the intestinal microbiota is crucial to enhance intestinal health and performance parameters in animals. A more exhaustive research of the intestinal microbiota of broilers could be of interest to implement appropriate intervention measures. The aim of the present study was to investigate the development of the predominant cecal microbiota in broilers that were fed a Lactobacillus salivarius DSPV 001P strain during a complete rearing using denaturing gradient gel electrophoresis (DGGE). Bacterial DNA from cecal samples of 24 broilers at different ages were amplified by PCR and analysed by DGGE. A total of 35 DGGE products were excised and sequenced. Distinctive differences in bacterial communities were observed in the caecum as broilers age. At early stages, identified bacteria within the caecum of broilers were predominantly Clostridium-related species. Also, some sequences had the closest match to the genus Escherichia/Shigella. Furthermore, the caecum was a reservoir rich in uncultured bacteria. The major difference observed in our study was an increase of potentially beneficial Lactobacillus at Day 45. These results may be attributed to modulation of the microbiota by the probiotic supplementation. The obtained data could be relevant for future studies related to the influence of the microbiota resulting from probiotic supplementation on the performance and the immunological parameters of broilers.

Poultry blood from slaughterhouses: development of a biopreservation system to improve microbiological quality prior to transforming blood into by-products

Abstract The aim of this study was to investigate the use of indigenous lactic acid bacteria (LAB) with specific additives as a Biopreservation System (BS) for poultry blood during its storage in slaughterhouses. The BS consisted of two LAB (Enterococcus faecalis DSPV 008SA or Lactobacillus salivarius DSPV 032SA) with 4 additives (lactose 2 g/l, yeast extract 0.4 g/l, ammonium citrate 0.4 g/l and NaCl 1 g/l). After 24 h storage at 30ºC, lower counts of enterobacteria, coliforms, Pseudomonas spp. and Staphylococcus aureus were evident in blood treated with the BS than in untreated blood. The ability of LAB to prevent haemolysis was evident. A decrease in pH was associated with control of spoilage microorganisms but it needed to be regulated to prevent coagulation of proteins. On the basis of these results it is recommended to supplement blood with a BS to avoid undesirable changes during blood storage before processing.