Loredana Baffoni

Beneficial microorganisms for honey bees: problems and progresses

Nowadays, honey bees are stressed by a number of biotic and abiotic factors which may compromise to some extent the pollination service and the hive productivity. The EU ban of antibiotics as therapeutic agents against bee pathogens has stimulated the search for natural alternatives. The increasing knowledge on the composition and functions of the bee gut microbiota and the link between a balanced gut microbiota and health status have encouraged the research on the use of gut microorganisms to improve bee health. Somehow, we are assisting to the transfer of the “probiotic concept” into the bee science. In this review, we examine the role of the honey bee gut microbiota in bee health and critically describe the available applications of beneficial microorganisms as pest control agents and health support. Most of the strains, mainly belonging to the genera Lactobacillus, Bifidobacterium and Bacillus, are isolated from honey bee crop or gut, but some applications involve environmental strains or formulation for animal and human consumption. Overall, the obtained results show the favourable effect of applied microbial strains on bee health and productivity, in particular if strains of bee origin are used. However, it is actually not yet possible to conclude whether this strategy will ever work. In particular, many aspects regarding the overall setup of the experiments, the dose, the timing and the duration of the treatment need to be optimized, also considering the microbiological safety of the hive products (i.e. pollen and honey). In addition, a deep investigation about the effect on host immunity and physiology is envisaged. Lastly, the final users of the formulations, i.e. beekeepers, should be taken into account for the achievement of high-quality, cost-effective and easy-to-use products.

Bioavailability and biological effect of engineered silver nanoparticles in a forest soil

The extensive use of silver nanoparticles (SNPs) as antimicrobial in food, clothing and medicine, leads inevitably to a loss of such nanomaterial in soil and water. Little is known about the effects of soil contamination, in particular, on microbial cells, which play a fundamental ecological role. In this work, the impact of SNPs on forest soil has been studied, investigating eco-physiological indicators of microbial biomass and microbial diversity with culture-dependent and independent techniques. Moreover, SNPs bioavailability and uptake were assessed. Soil samples were spiked with SNPs at two different concentrations (10 and 100 μg g(-1)dw) and incubated with the relative controls for 30, 60 and 90 days. The overall parameters showed a significant influence of the SNPs on the soil microbial community, revealing a marked shift after 60 days of incubation.

Effect of dietary supplementation of Bifidobacterium and Lactobacillus strains in Apis mellifera L. against Nosema ceranae.

Nosema ceranae is a widespread microsporidium of European honeybee Apis mellifera L. affecting bee health. The ban of Fumagillin-B (dicyclohexylammonium salt) in the European Union has driven the search for sustainable strategies to prevent and control the infection. The gut microbial symbionts, associated to the intestinal system of vertebrates and invertebrates and its impact on host health, are receiving increasing attention. In particular, bifidobacteria and lactobacilli, which are normal inhabitants of the digestive system of bees, are known to protect their hosts via antimicrobial metabolites, immunomodulation and competition. In this work, the dietary supplementation of gut bacteria was evaluated under laboratory conditions in bees artificially infected with the parasite and bees not artificially infected but evidencing a low natural infection. Supplemented bacteria were selected among bifidobacteria, previously isolated, and lactobacilli, isolated in this work from healthy honeybee gut. Four treatments were compared: bees fed with sugar syrup (CTR); bees fed with sugar syrup containing bifidobacteria and lactobacilli (PRO); bees infected with N. ceranae spores and fed with sugar syrup (NOS); bees infected with N. ceranae and fed with sugar syrup containing bifidobacteria and lactobacilli (NP). The sugar syrup, with or without microorganisms, was administered to bees from the first day of life for 13 days. N. ceranae infection was carried out individually on anesthetised 5-day-old bees. Eight days after infection, a significant (P<0.05) lower level of N. ceranae was detected by real-time PCR in both NP and PRO group, showing a positive effect of supplemented microorganisms in controlling the infection. These results represent a first attempt of application of bifidobacteria and lactobacilli against N. ceranae in honeybees.

Molecular characterisation of an endophyte showing a strong antagonistic activity against Pseudomonas syringae pv. actinidiae

Background and aimsEndophytic bacteria have been often studied as biological control agents of plant pathogens and many of their secondary metabolites involved in antagonism are Non Ribosomal Peptides (NRPs). In this study, the molecular basis of the biocontrol properties of the endophyte Pseudomonas synxantha, isolated from Actinidia chinensis, against Pseudomonas syringae pv. actinidiae (Psa), the causal agent of bacterial kiwi canker, has been investigated.MethodsAntagonism-deficient mutants of P. synxantha strain DLS65 were generated by insertion of minitransposon mTn5-GNm in its genome. Southern blot analysis allowed the selection of single transposon insertion-mutants. Amplification of the transposon flanking regions by means of arbitrary and single primer PCR in selected mutants was perfomed to obtain amplicons for sequencing purposes.ResultsSequencing results of the amplicons obtained from three antagonism-deficient mutants led to the localization of the transposonin three genes, which implies their involvement in the antagonism of P. synxantha: an acyl-homoserine lactone acylase gene (pvdQ), a glucose-6-phosphate dehydrogenase gene (zwf) and an mbtH-like gene were identified. It is known from the literature that these three genes are involved directly or indirectly in NRPs synthesis.ConclusionsWe suggest that a molecule with antibiotic properties, produced by NRP synthetases, contributes to the antagonistic activity of P. synxantha.

Identification of Bifidobacterium spp. using hsp60 PCR-RFLP analysis: An update

A PCR-RFLP technique has been applied on 13 species of Bifidobacterium in order to update a previous study carried out by Baffoni et al. This method is based on the restriction endonuclease activity of HaeIII on the PCR-amplified hsp60 partial gene sequence, and allows a rapid and efficient identification of Bifidobacterium spp. strains at species and subspecies level.

Evidence of Campylobacter jejuni reduction in broilers with early synbiotic administration

C. jejuni is considered a food safety concern to both public health authorities and consumers since it is the leading bacterial cause of food-borne gastroenteritis in humans. A high incidence of C. jejuni in broiler flocks is often correlated to pathogen recovery in retail poultry meat, which is the main source of human infection. In this work broiler chickens were fed with a synbiotic product mixed with conventional feed using two different administration strategies. The synbiotic was formulated with the microencapsulated probiotic Bifidobacterium longum PCB133 and a xylo-oligosaccharide (XOS). 1-day old chicks were infected with C. jejuni strain M1 (105 cells) and the synbiotic mixture was then administered starting from the first and the 14th day of chicken life (for animal groups GrpC and GrpB respectively). The goal of this study was to monitor C. jejuni load at caecum level at different sampling time by real-time PCR, identifying the best administration strategy. The microbiological analysis of the caecal content also considered the quantification of Campylobacter spp., Bifidobacterium spp. and B. longum. The supplemented synbiotic was more successful in reducing C. jejuni and Campylobacter spp. when administered lifelong, compared to the shorter supplementation (GrpB). Bifidobacterium spp. quantification did not show significant differences among treatments and B. longum PCB133 was detected in both supplemented groups evidencing the successful colonization of the strain. Moreover, the samples of the control group (GrpA) and GrpC were analysed with PCR-denaturing gradient gel electrophoresis (PCR-DGGE) to compare the caecal microbial community profiles at the beginning and at the end of the trial. Pattern analysis evidenced the strong influence of the early synbiotic supplementation, although a physiological change in the microbial community, occurring during growth, could be observed. Experimental results demonstrate that the synbiotic approach at farm level can be an effective strategy, combined with biosecurity measures, to improve the safety of poultry meat.

Impact of beneficial bacteria supplementation on the gut microbiota, colony development and productivity of Apis mellifera L.

Honey bees are important pollinators of several crops and ecosystems, having a great ecological and economic value. In Europe, the restricted use of chemicals and therapeutic agents in the beekeeping sector has stimulated the search for natural alternatives with a special focus on gut symbionts. The modulation of the gut microbiota has been recognised as a practical and successful approach in the entomological field for the management of insect-related problems. To date, only a few studies have investigated the effect of bacterial supplementation on the health status of colonies, colony productivity and gut symbionts. To this purpose, a preparation of sugar syrup containing bifidobacteria and lactobacilli isolated from bee gut was sprayed on the frames of an apiary located in open field once a week for four weeks. Treated and control hives were monitored for two months for brood extension, honey and pollen harvest. The presence of beneficial gut microorganisms within bee gut was investigated with denaturing gradient gel electrophoresis and next generation sequencing. The administered bacteria led to a significant increase of brood population (46.2%), pollen (53.4%) and harvestable honey in honey supers (59.21%). Analysis of the gut microbiota on the new generation of bees in treated hives showed an increase in relative abundance of Acetobacteraceae and Bifidobacterium spp., which are known to be involved in bee nutrition and protection.

Probiotics for Honeybees’ Health

Honeybee is certainly one of the most familiar flying insects of terrestrial habitats. Honeybees are critically important in the environment, sustaining biodiversity and providing essential pollination for a wide range of crops and wild plants. Extensive losses of honeybee colonies in recent years are becoming a major cause of concern. These social insects continuously face threats (diseases, climate change, and management practices) that weaken their health. In this chapter, we will focus on the western honeybee A. mellifera, focusing on gut symbionts, their functions and role on honeybee health

Probiotics and Prebiotics for the Health of Companion Animals

Companion animals, also referred as pets, are animals kept primarily as companions at home or having close daily relationship with humans. Dogs and cats are the major human companions and their health and wellness are essential for pet owners. Ensuring a wholesome and balanced diet is a fundamental duty of a responsible pet owner. In the last decades, the growing knowledge in canine and feline nutrition has greatly contributed to improve longevity and well-being of companion animals and interesting outcomes have been obtained with the introduction of dietary fibres and beneficial microorganisms in pets’ diet.

Three-Month Feeding Integration With Bifidobacterium Strains Prevents Gastrointestinal Symptoms in Healthy Newborns

Infantile functional gastrointestinal disorders are common in the first months of life. Their pathogenesis remains unknown although evidences suggest multiple independent causes, including gut microbiota modifications. Feeding type, influencing the composition of intestinal microbiota, could play a significant role in the pathogenesis. Previous studies supported probiotic supplementation success against colics, however mainly Lactobacillus spp. were tested. The aim of this study was to evaluate the effectiveness against functional gastrointestinal disorders of a Bifidobacterium breve based probiotic formulation including in the study both breast-fed and bottle-fed subjects. Two hundred and sixty-eight newborns were enrolled within 15 days from birth. One hundred and fifty-five of them effectively entered the study and were randomized in probiotic and placebo group, receiving the formulation for 90 days. The probiotic formulation consists of a 1:1 mixture of 2 strains of B. breve prepared in an oily suspension and administered in a daily dosage of 5 drops containing 108 CFU of each strain. Absolute quantification of selected microbial groups in the faeces was performed using qPCR. Anthropometric data, daily diary minutes of crying, number of regurgitations, vomits and evacuations, and colour and consistency of stools were evaluated before and after treatment. The study confirmed the positive role of breast milk in influencing the counts of target microbial groups, in particular the bifidobacteria community. No adverse events upon probiotic administration were reported, suggesting the safety of the product in this regimen. B. breve counts increased significantly in all administered newborns (p < 0.02). The study demonstrates that a 3 months treatment with B. breve strains in healthy breast-fed newborns helps to prevent functional gastrointestinal disorders, in particular reducing 56% of daily vomit frequency (p < 0.03), decreasing 46.5% of daily evacuation over time (p < 0.03), and improving the stool consistency (type 6 at the Bristol Stool chart instead of type 5) in those at term (p < 0.0001). Moreover, a significant reduction (8.65 vs. 7.98 LogCFU/g of feces, p < 0.03) of B. fragilis in the bottle-fed group receiving the probiotic formulation was observed.