Screening for Bacillus Isolates in the Broiler Gastrointestinal Tract

Abstract

Spores from a number of different Bacillus species are currently being used as human and animal probiotics, although their mechanisms of action remain poorly understood. Here we describe the isolation of 237 presumptive gut-associated Bacillus spp. isolates that were obtained by heat and ethanol treatment of fecal material from organically reared broilers followed by aerobic plating. Thirty-one representative isolates were characterized according to their morphological, physiological, and biochemical properties as well as partial 16S rRNA gene sequences and screening for the presence of plasmid DNA. The Bacillus species identified included B. subtilis, B. pumilus, B. licheniformis, B. clausii, B. megaterium, B. firmus, and species of the B. cereus group, whereas a number of our isolates could not be classified. Intrinsic properties of potential importance for survival in the gut that could be advantageous for spore-forming probiotics were further investigated for seven isolates belonging to five different species. All isolates sporulated efficiently in the laboratory, and the resulting spores were tolerant to simulated gastrointestinal tract conditions. They also exhibited antimicrobial activity against a broad spectrum of bacteria, including food spoilage and pathogenic organisms such as Bacillus spp., Clostridium perfringens, Staphylococcus aureus, and Listeria monocytogenes. Importantly, the isolates were susceptible to most of the antibiotics tested, arguing that they would not act as donors for resistance determinants if introduced in the form of probiotic preparations. Together, our results suggest that some of the sporeformers isolated in this study have the potential to persist in or transiently associate with the complex gut ecosystem.The widespread use of antibiotics as therapeutic and prophylactic agents and as growth promoters in animal husbandry has led to a worldwide increase in antibiotic resistance and the emergence of untreatable multidrug-resistant strains of bacteria (4, 33). This has prompted the European Union to phase out the use of these compounds as animal feed additives by 2006 (47). The use of competitive exclusion agents and probiotic feed additives in the livestock industry is therefore attracting increased attention as a cost-effective alternative to controlling animal disease and improving breeding performance (54).The genus Bacillus comprises a diverse collection of aerobic endospore-forming bacteria whose spores consist of several protective layers surrounding the nucleoid in the spore core (20, 51). This structural organization makes the spores extremely resistant to external physical and chemical insults and in part determines their exceptional longevity in the environment (20, 38). The resilient and ubiquitous nature of these bacteria results in considerable daily intake of these organisms by humans and animals. This ingestion, which is often in the form of spores, occurs primarily through contaminated food and water.Bacillus spores are being used as human and animal probiotics...

Identification and characterization of Bacillus subtilis from grass carp (Ctenopharynodon idellus) for use as probiotic additives in aquatic feed

“…The results elucidated that the spores were not sensitive to 80 C and 90 C, and survived even up to 100 C, although the survival rate decreased with the increase of the heating time. Furthermore, the spores showed strong resistances to low pH and high bile concentration, which were also reported by many previous studies [4,29,51]. Thus, the isolates got the accession to feed additives.…”

“…Firstly, the spores are produced in nature and more heat-stable than other non-spore forming bacteria such as Lactobacillus spp, namely, the products can be stored at room temperature in desiccated form with little deleterious effect on viability [6]. Secondly, the spores are capable of surviving at high percentage of bile and low pH of the gastric barrier [4,7], which is not the case for all species of Lactobacillus [8]. Furthermore, the spores have the abilities to survive and germinate in the gut, to form biofilm, and to secrete antimicrobials [4,9,10].…”

“…Secondly, the spores are capable of surviving at high percentage of bile and low pH of the gastric barrier [4,7], which is not the case for all species of Lactobacillus [8]. Furthermore, the spores have the abilities to survive and germinate in the gut, to form biofilm, and to secrete antimicrobials [4,9,10]. Finally, extensive investigations confirmed that B. subtilis is an attractive vehicle for delivery of heterologous antigens to gastrointestinal tract, suggesting that B. subtilis can be used as oral vaccine delivery system to enhance immune responses and prevent diseases [11].…”

Selection of Bacillus species for targeted in situ release of prebiotic galacto-rhamnogalacturonan from potato pulp in piglets

“…The strains were isolated from mainly feces of humans, chickens and pigs or from soil (Supplementary Table S1). Strain 10894 that is identical to isolate 37 in (Barbosa et al 2005 (Schaeffer et al 1965). For evaluation of biofilm formation, the strains were grown in biofilm medium (LB medium supplemented with 0.1 mM MnCl 2 , and 3 % v/v glycerol) (Trejo et al 2013).…”

Influência do jejum alimentar, probióticos e antibiótico na população de enterobactérias, bactérias ácido lácticas, Bacillus e Salmonella sp. em cecos e papos de frangos de corte

“…Apesar de a maioria dos estudos utilizar BAL como probióticos, atenção vem sendo dada a microrganismos formadores de esporos, principalmente Bacillus sp.. As vantagens deste tipo de bactérias sobre as formas vegetativas são fundamentalmente comerciais, pela facilidade de preparação, resistência e menor custo para processos de produção, além de vida útil maior em amplos limites de temperaturas (BARBOSA et al, 2005).…”