Evaluation of Phytate-Degrading Lactobacillus Culture Administration to Broiler Chickens

Abstract

Probiotics have been demonstrated to promote growth, stimulate immune responses, and improve food safety of poultry. While widely used, their effectiveness is mixed, and the mechanisms through which they contribute to poultry production are not well understood. Microbial phytases are increasingly supplemented in feed to improve digestibility and reduce antinutritive effects of phytate. The microbial origin of these exogenous enzymes suggests a potentially important mechanism of probiotic functionality. We investigated phytate degradation as a novel probiotic mechanism using recombinant Lactobacillus cultures expressing Bacillus subtilis phytase. B. subtilis phyA was codon optimized for expression in Lactobacillus and cloned into the expression vector pTRK882. The resulting plasmid, pTD003, was transformed into Lactobacillus acidophilus, Lactobacillus gallinarum, and Lactobacillus gasseri. SDS-PAGE revealed a protein in the culture supernatants of Lactobacillus pTD003 transformants with a molecular weight similar to that of the B. subtilis phytase. Expression of B. subtilis phytase increased phytate degradation of L. acidophilus, L. gasseri, and L. gallinarum approximately 4-, 10-, and 18-fold over the background activity of empty-vector transformants, respectively. Phytase-expressing L. gallinarum and L. gasseri were administered to broiler chicks fed a phosphorusdeficient diet. Phytase-expressing L. gasseri improved weight gain of broiler chickens to a level comparable to that for chickens fed a control diet adequate in phosphorus, demonstrating proof of principle that administration of phytate-degrading probiotic cultures can improve performance of livestock animals. This will inform future studies investigating whether probiotic cultures are able to provide both the performance benefits of feed enzymes and the animal health and food safety benefits traditionally associated with probiotics. Lactobacillus species are important inhabitants of the gastrointestinal tracts of humans and animals and are increasingly being used as probiotic microorganisms due to their health-promoting properties (1, 2). Probiotics, sometimes called direct-fed microbials (DFM) when used in animals (3), are live microorganisms administered to confer a health benefit upon the host (4). Administration of probiotic Lactobacillus to poultry has been demonstrated to promote growth at levels similar to antibiotics (5, 6) and to reduce gastrointestinal colonization of human foodborne pathogens, including Campylobacter (7, 8), Clostridium (9), and Salmonella (10, 11). Because of concern over antibiotic-resistant pathogens and pressure from both consumers and regulatory agencies, probiotics have received increased interest as potential alternatives to antibiotic growth promoters (12). While probiotics are used widely in livestock production (13), their effectiveness is varied, and the mechanisms responsible for their benefits are not well understood.Phosphorus is an essential nutrient in poultry production (14), with dietary deficiencies leadin...

Synthetic gene design—The rationale for codon optimization and implications for molecular pharming in plants

“…marginale (Pierlé et al 2013), R. pomeroyi (Green et al 2013), L. acidophilus (Askelson et al 2014), C. reinhardtir (Erpel et al 2016), Y. lipolytica (Matthaus et al 2014), S. elongates (van der Woude et al 2016) & baculovirus (Maghodia et al 2016). While it is encouraging that codon optimization software programs are used for a variety of species and purposes, most of the papers…”

“…Many of the codon optimization software lack citations. Of these software packages, citations have been found for JCat and OPTIMIZER for expression systems such as E. coli (Fahimi et al, 2016;Guo et al, 2016;Karkhah and Amani, 2016;Zhao et al, 2016), S. cerevisiae (Ask et al, 2013;Guadalupe-Medina et al, 2013;Li et al, 2015;Milne et al, 2015;Solis-Escalante et al, 2013), N. benthamiana (Binder et al, 2014), HEK293 cells (Shah et al, 2015), B. subtilis (Reilman et al, 2014), Caulobacter crescentus (Ko et al, 2013), Pseudomonas putida (Dammeyer et al, 2011(Dammeyer et al, , 2013, Salmonella typhimurium (Manuel et al, 2011), S. lividans (Dubeau et al, 2009), wheat (Mih alik et al, 2015, transplastomic tobacco plants (chloroplast translation system) (Occhialini et al, 2015), Plasmodium berghei (Singer et al, 2015), Spodoptera frugiperda (Geisler et al, 2015), S. pneumoniae (Overkamp et al, 2013), A. marginale (Pierl e et al, 2013), R. pomeroyi (Green et al, 2013), L. acidophilus (Askelson et al, 2014), C. reinhardtir (Erpel et al, 2016), Y. lipolytica (Matthaus et al, 2014), S. elongates (van der Woude et al, 2016), and baculovirus (Maghodia et al, 2016). While it is encouraging that codon optimization software programs are used for a variety of species and purposes, most of the papers do not compare yields of native and codon optimized sequences, so yield comparisons cannot be made.…”

Assessment of the potential of the multi‐enzyme producer Bacillus amyloliquefaciens US573 as alternative feed additive

“…When administered in adequate amounts, probiotics confer a health benefit on the host by balancing its intestinal microbes . The most common microorganisms used as probiotics are lactic acid bacteria and bifidobacteria, thanks to their recognition as members of the indigenous microflora of animals, their safety and the evidence supporting their positive role . However, the use of these microorganisms in poultry industry imposes a supplementary cost for survival during long storage periods and feed pelleting processes .…”

“…[3][4][5] The most common microorganisms used as probiotics are lactic acid bacteria and bifidobacteria, thanks to their recognition as members of the indigenous microflora of animals, their safety and the evidence supporting their positive role. [6][7][8] However, the use of these microorganisms in poultry industry imposes a supplementary cost for survival during long storage periods and feed pelleting processes. 9 Among the microorganisms used as DFM, members of the spore-forming Bacillus genus have been increasingly supplemented as feed additives in poultry rations [10][11][12][13] owing to their outstanding capacity to resist harsh environmental conditions (high temperatures applied during feed pelleting, extremes of pH, dehydration, high pressures and caustic chemicals) and long storage life.…”

Phytase-Producing Potential and Other Functional Attributes of Lactic Acid Bacteria Isolates for Prospective Probiotic Applications

“…Also anti-nutritional properties of phytate could be minimized or omitted with the application of phytase/phytase-producing probiotics [6]. Improving availability and digestibility of nutrients in feeds is very important considering increased demand of food/feed world over, and recently, the WHO approved the phytase Aspergillus niger for use in foods [17].…”