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Booming innovation area at NIZO food research: Discovering the role of microbiota in human and animal health

Published: 04-02-2014, | Member: NIZO

Scientists believe these microbial ecosystems play a crucial role in human health. Growing evidence suggests that the microbiota are intimately connected  to the immune system and help in calibrating fundamental metabolic functions. It has also been reported that the microbiota may influence the enteric (ENS)  and central nervous system (CNS), having an impact on behavior and mood. Hence,  therapeutic targeting of the microbiome might be an effective treatment strategy  for specific disorders of the gut, skin or oral niche, and may also, for example, modulate metabolic disorders like obesity and diabetes. However, we are not  yet able to establish clear causal relationships or define the ideal microbiota profile to prevent or delay the onset of specific diseases.

Characterization of microbiota
As methods for microbiota analyses rapidly evolve and improve, our knowledge of species and functional composition of the various microbiomes swiftly increases. NIZO food research is among the pioneers in this challenging and intriguing area. “First, we have to characterize these complex microbial ecosystems,” says Dr. Harro Timmerman, Group Leader Microbiota at NIZO. He continues: “This characterization should not only evaluate the composition of the microbiome, but—and this is very important—should also address its functionality because it is NIZO’s goal to help clients improve their products. Furthermore, before we can unambiguously identify microbiological biomarkers related to health and disease, we need to understand which normal temporal variations occur in the microbiome within one individual, and how the microbiota can be influenced by external factors like medication, dietary or lifestyle habits, and how these external influences interact with the impact of the genotype of the host.” Hence, researchers are studying interpersonal and intrapersonal variations in the microbiome, as well as the origin of these variations.

Microbiota profiling can be done in several ways. NIZO has a state-of-the-art microbiota profiling and quantification platform at its disposal. Well-established techniques include fluorescent in situ hybridization (FISH), quantitative polymerase chain reaction (qPCR), (high-resolution) Amplified Fragment Length Polymorphism (AFLP), as well as the use of intestinal tract chips. “One of the newest technologies in microbiota analysis is barcoded pyrosequencing,” Timmerman explains enthusiastically. “With this technique, randomly amplified and highly conserved genes, like the 16s ribosomal RNA gene of bacteria, are analysed by massive parallel sequencing of the generated amplicons. This profiling can be performed in a high-throughput manner, with low costs per sample.” This approach allows researchers to compare hundreds of samples, generates compositional profiles of the microbiota down to the genus level (even of unknown bacteria) and quantifies the relative proportions of organisms present.

However, the massive data output of the technology requires advanced interpretation strategies, to extract information from the data in a comprehensive manner. This is easier said than done. NIZO bio-informaticians have developed methods to comprehensively visualize the composition of the microbiota and to statistically compare the differences between groups of people or animals. “This enables us to identify the bacterial profile related to a specific health problem,” explains Timmerman. “With these tools, the multidisciplinary NIZO team is able to translate the sequencing data into biological perspectives.”

Profiling skin, oral and gastrointestinal microbiota: setting the scene for nutritional intervention studies
NIZO has extensive experience in studying intestinal microbiota, revealing—step by step—the dynamics and possible mechanisms and relationships between the intestinal microbiota and the host, and thereby the role of the microbes in physiological, metabolic or pathological conditions. “We determine who’s there, what they are capable of, what they are doing and what they have done,” quips Timmerman. In addition to the intestinal microbiota, NIZO is actively involved in analyses of the human skin microbiome, as well as the microbial inhabitants of the oral cavity.

In a collaborative project funded by national and regional governments in the Netherlands, NIZO is actively partnering with both the Radboud University Nijmegen Medical Centre and Nutreco, an international animal nutrition and fish feed company. One project line targets resistance to infections in piglets, chickens and fish. “Barcoded pyrosequencing revealed an association between the presence of specific groups of bacteria in the small intestines of weaning piglets and animal growth” says Timmermans. “Based on this research, Nutreco is able to develop new products to support the gut health of these animals, ideally leading to a reduction in the use of antibiotics.”

In a second project line, researchers are creating inventories of the skin bacteria of various patients with skin diseases and of control groups. At least 250 different genera live on the human skin, and this flora composition differs with the anatomical location. Research themes include the relationship between streptococcal bacteria and psoriasis, as well as the relationship between Staphylococcus aureus and eczema. Last but not least, researchers are considering the link between the gut microbiota and skin diseases.

Links between skin flora and conditions like acne, dandruff and malodor also seem plausible. Analogously, NIZO is studying the influence of the oral microbial ecosystem on breath quality (halitosis) in collaboration with the University Medical Centre Groningen and the Clinic of Parodontology in Amsterdam. Volatile sulfur compounds from oral bacteria influence breath quality. NIZO researchers have demonstrated that morning breath is a suitable target for intervention studies (Snel et al., 2011a). Furthermore, women tend to have higher concentrations of these volatile components when they wake up. “Ongoing studies involving the dorsal tongue microbiota composition demonstrate clear differences between healthy volunteers and halitosis patients,” reports Timmerman.

In a project with the multinational dairy company FrieslandCampina, NIZO determined the persistence of beneficial bacteria in the oral cavity. Beneficial microorganisms such as lactic acid bacteria might offer opportunities as oral probiotics by outcompeting the unwanted bacteria or by reducing their activity, for instance, with respect to fighting bad breath and tooth decay.

NIZO scientists developed an effective way to determine the persistence of bacteria in the mouth – persistence being one of the requirements for developing strategies to decrease the activity or abundance of the detrimental bacteria. With this new selection method, 69 food-grade lactic acid bacteria strains from the LactobacillusLactococcus and Streptococcus genera were evaluated for their persistence in vivo in the human oral cavity. It was found that candidate probiotic strains differed greatly in their persistence in the oral cavity. In particular, they recovered strains of Lactobacillus fermentum and Lactobacillus salivarius, and some of these strains presented in the mouth of healthy volunteers for more than two weeks after consumption (Snel et al., 2011b). This study illustrates a first step in selecting candidate probiotic strains aimed at promoting oral health.

Potential effect of probiotic interventions
Numerous health benefits have been attributed to probiotic microorganisms, although not all effects have been evaluated in in vivo (human) conditions yet. For example, probiotics may have a direct interaction with the intestinal microbiota, prevent pathogenic bacterial overgrowth, influence intestinal barrier function at the level of the epithelium or mucus layer, and affect the mucosal and even the systemic immune system. It is important to note that each probiotic strain has its own specific properties. Even closely related probiotic strains of the same species turn out to have different physiological effects. In collaboration with FrieslandCampina, Wageningen University, Rijnstate Hospital and Radboud University Nijmegen Medical Centre, NIZO has demonstrated strain-specific immunomodulatory effects of Lactobacillus plantarum in birch-pollen-allergic persons (Snel et al., 2011c).

In a unique randomized controlled trial, carried out in the framework of TIFN and co-sponsored by FrieslandCampina and Unilever, scientists from NIZO and Wageningen University, together with the Southeast Asian Ministers of Education Organization Regional Centre for Food and Nutrition (SEAMEO RECFON), demonstrated that the probiotic Lactobacillus reuteri prevents acute diarrhea in children from socioeconomically depressed communities in Indonesia (Agustina et al., 2012).

In a six-month, double blind, placebo-controlled trial, 494 apparently healthy children ages 1 to 6 were randomly assigned to receive low-lactose milk with or without Lactobacillus casei 431 or Lactobacillus reuteri DSM17938. Incidence of all diarrhea episodes (≥ 2 loose/liquid stools in 24 hours) was reduced by 32% in the reuteri group as compared to the placebo group. Furthermore, incidence of WHO-defined episodes (≥ 3 loose/liquid stools in 24 hours) tended to be lower in the reuteri group among children with lower nutritional status. “Introduction of barcoded pyrosequencing and other high-throughput molecular techniques in this type of dietary intervention studies generates novel insights into the mechanism of action of probiotic interventions, which can prove  to be crucial information for health claim approval by EFSA,” says Timmerman.

“In studying the microbiota composition in health and disease, we also identify co- and anti-occurrence of pathogens with other members of the microbial community,” Timmerman adds. In itself, these commensal, respectively potentially beneficial microorganisms might be an alternative target for specific nutritional intervention. “Understanding the metabolic interrelationship between these pathogenic organisms and the commensal inhabitants will contribute to our capacities to develop innovative food products that enable the selective outcompeting of specific pathogens,” Timmerman concludes.”In studying the microbiota composition in health and disease, we also identify co- and anti-occurrence of pathogens with other members of the microbial community,” Timmerman adds. In itself, these commensal, respectively potentially beneficial microorganisms might be an alternative target for specific nutritional intervention. “Understanding the metabolic interrelationship between these pathogenic organisms and the commensal inhabitants will contribute to our capacities to develop innovative food products that enable the selective outcompeting of specific pathogens,” Timmerman concludes.


Agustina et al. 2012 Pediatrics. 129:e1155–64. Randomized trial of probiotics and calcium on diarrhea and respiratory tract infections in Indonesian children.

Snel et al. Arch Oral Biol. 2011a, 56:29–34. Volatile sulphur compounds in morning breath of human volunteers.

Snel et al. Appl Environ Microbiol. 2011b, 77:8445–50. Competitive selection of lactic acid bacteria that persist in the human oral cavity.

Snel et al. Clin Exp Allergy. 2011c, 41:232–42. Strain-specific immunomodulatory effects of Lactobacillus plantarum strains on birch-pollen-allergic subjects out of season.

NIZO food research