Torunn Forberg

GIARDIA DUODENALIS CYSTS ISOLATED FROM WILD MOOSE AND REINDEER IN NORWAY : GENETIC CHARACTERIZATION BY PCR-RFLP AND SEQUENCE ANALYSIS AT TWO GENES

There are few genotyping studies of Giardia duodenalis isolates from cervid hosts, although a previous study suggested that cervids may be a source of infection for humans and cattle. Giardia duodenalis isolates collected from wild moose (Alces alces) and reindeer (Rangifer tarandus) in Norway during 2002 and 2003 were characterized by polymerase chain reaction–restriction fraction length polymorphism (PCR-RFLP) at the β-giardin gene, and sequence analysis at both the β-giardin and glutamate dehydrogenase (gdh) genes. All results suggested that these isolates (n=25) belonged to assemblage A. Three different restriction patterns were obtained with PCR-RFLP, one of which has previously been associated with assemblage A. At the β-giardin gene, sequences from six reindeer isolates and one moose isolate were identical to a previously published assemblage A sequence from G. duodenalis cysts isolated from dairy calves. The other 10 moose isolates could be divided into five groups, with between two and 14 single nucleotide polymorphisms (SNPs) from the published genotype A2. At the gdh gene, three different sequences were obtained, differing from each other by between one and 15 SNPs and which have all been previously published as genotype A1, but with different specific hosts. Grouping of the isolates based on the sequences from both genes gave complex results; whereas all the G. duodenalis isolates from reindeer grouped together, two moose isolates, which had identical sequences at the β-giardin gene, had sequences that differed from each other by 15 SNPs at the gdh gene. The results of these studies, together with the large Norwegian populations of these cervids and the amount of fecal matter they produce, indicate that moose and reindeer may be significant reservoirs of G. duodenalis infection in Norway, which may be of importance to veterinary and public health.

Molecular and Genetic Characterization of a Novel Bacteriocin Locus in Enterococcus avium Isolates from Infants

ABSTRACT Enterococci are among the most common human intestinal lactic acid bacteria, and they are known to produce bacteriocins. In this study, fecal enterococci were isolated from infants and screened for bacteriocin production. Bacteriocin-producing Enterococcus avium isolates were obtained, and a new pediocin-like bacteriocin was purified and characterized. This bacteriocin, termed avicin A, was found to be produced by isolates from two healthy infants. It was purified to homogeneity from culture supernatant by ion-exchange and reversed-phase chromatography, and part of its amino acid sequence was obtained. The sequence of a 7-kb DNA fragment of a bacteriocin locus was determined by PCR and DNA sequencing. The bacteriocin locus was organized into four operon-like structures consisting of (i) the structural genes encoding avicin A and its immunity protein, (ii) a divergicin-like bacteriocin (avicin B) gene, (iii) an ABC bacteriocin transporter gene and two regulatory genes (histamine protein kinase- and response regulator-encoding genes), and (iv) induction peptide pheromone- and transport accessory protein-encoding genes. It was shown that the production of avicin A was regulated by the peptide pheromone-inducible regulatory system. Avicin A shows very high levels of similarity to mundticin KS and enterocin CRL35. This bacteriocin showed strong antimicrobial activity against many species of Gram-positive bacteria, including the food-borne pathogen Listeria monocytogenes. The avicin A locus is the first bacteriocin locus identified in E. avium to be characterized at the molecular level.

Cryptosporidium parvum Infections in Bergen, Norway, during an Extensive Outbreak of Waterborne Giardiasis in Autumn and Winter 2004

ABSTRACT During a large waterborne giardiasis outbreak in Norway, many diarrheic patients were found to have Cryptosporidium infections. Gene sequencing identified these infections as Cryptosporidium parvum infections, although they were not identical. Whether these infections were due to a simultaneous outbreak of waterborne cryptosporidiosis or reflected background levels not normally detected is discussed.

A small outbreak of human cryptosporidiosis associated with calves at a dairy farm in Norway

Cryptosporidiosis is rarely diagnosed in Norway. This is the first report of a human outbreak in Norway and involved 3 calves and 5 people. Sequencing studies were performed. Although 4 of the people acquired their infections from the calves, the other probably became infected from other calves or contaminated pens.

Correlation between microbiota and growth in Mangrove Killifish ( Kryptolebias marmoratus ) and Atlantic cod ( Gadus morhua )

The vertebrate gut is host to large communities of bacteria, and one of the beneficial contributions of this commensal gut microbiota is the increased nutritional gain from feed components that the host cannot degrade on its own. Fish larvae of similar age and under the same rearing conditions often diverge with regards to growth. The underlying reasons for this could be differences in genetic background, feeding behavior or digestive capacity. Both feeding behavior and digestion can be influenced by differences in the microbiota. To investigate possible correlations between the size of fish larvae and their gut microbiota, we analyzed the microbiota small and large genetically homogenous killifish and genetically heterogeneous cod larvae by Bray-Curtis Similarity measures of 16S DNA DGGE patterns. A significant difference in richness (p = 0.037) was observed in the gut microbiota of small and large killifish, but the overall gut microbiota was not found to be significantly different (p = 0.13), indicating strong genetic host selection on microbiota composition at the time of sampling. The microbiota of small and large cod larvae was significantly different with regards to evenness and diversity (p = 0.0001), and a strong correlation between microbiota and growth was observed.

Giardia cysts in sewage influent in Bergen, Norway 15–23 months after an extensive waterborne outbreak of giardiasis

Aims:  In autumn/winter 2004, a large outbreak of waterborne giardiasis occurred in Bergen, Norway. Over 1 year later, the concentrations and genotypes of Giardia cysts occurring in sewage influent were studied to investigate the impact of the outbreak event on Giardia infections in the community.

Bacterial composition and activity determines host gene-expression responses in gnotobiotic Atlantic cod (Gadus morhua) larvae

In this study the effects of different live versus dead bacteria on the survival and gene-expression patterns in Atlantic cod (Gadus morhua) larvae were examined. Seventeen different gnotobiotic conditions were tested in a 5-day experiment, and the effects of these conditions on the expression of selected genes involved in immune response, xenobiotic breakdown and nutrition (C3, Gpx, Cyp1a1 and Fdps) were investigated. For some of the conditions where dead bacteria were added, we observed significantly higher survival compared to the axenic control. Microbial specificity of host-responses was observed for all four genes, for both live and dead bacteria, although expression of Gpx and Fdps was more influenced by active bacteria. Live bacteria generally resulted in higher transcript levels of all the genes investigated, indicating that these host-responses are partly dependent on microbial activity, and not just due to interactions with bacteria as particles. The observed microbial specificity toward dead bacteria highlights the need to take into account the role of bacterial components when investigating host-microbe relationships.

Demographics of Giardia infections in Bergen, Norway, subsequent to a waterborne outbreak

Young women have previously been reported as the predominant group infected during a waterborne giardiasis outbreak, due to elevated water consumption. Here, the demographics of those subsequently infected are described, and young women again predominate. As secondary cases were not waterborne, this cannot be attributed to drinking habits.

Investigating Fish Larvae-Microbe Interactions in the 21st Century: Old Questions Studied with New Tools

All animals need a mutualistic interaction with their microbiota for proper development and functioning. Also for the fish-microbiota interaction considerable research has been done, and especially for reared fish larvae this interaction is crucial for their viability. However, during the 1980s and 1990s a number of findings revealed at that time current methods were not suitable for studying the total microbial community and that data on composition of microbiota was biased. Several recent methodological revolutions have boosted the possibilities for addressing questions related to fish larvae-microbiota interactions that previously lacked suitable tools for proper evaluation. These methodological achievements include the development of experimental rearing systems including gnotobiotic systems for fish, new visualization tools, and molecular “omics” tools for characterizing the response of the host on a variety of levels and for characterizing both composition and activity of fish microbiota. We present and review these tools and give examples on how they have been used to improve our understanding of fish larvae-microbiota interactions. With respect to understanding, this includes in particular how the microbiota is established and maintained, what the functionality of the microbiota is and how it affects fish health, and finally how we can apply this knowledge for management of a healthy and beneficial microbiota in aquaculture settings.

Managing the microbial community of marine fish larvae: a holistic perspective for larviculture

The availability of high-quality juveniles is a bottleneck in the farming of many marine fish species. Detrimental larvae-microbe interactions are a main reason for poor viability and quality in larval rearing. In this review, we explore the microbial community of fish larvae from an ecological and eco-physiological perspective, with the aim to develop the knowledge basis for microbial management. The larvae are exposed to a huge number of microbes from external and internal sources in intensive aquaculture, but their relative importance depend on the rearing technology used (especially flow-through vs. recirculating systems) and the retention time of the water in the fish tanks. Generally, focus has been on microbes entering the system, but microbes from growth within the system is normally a substantial part of the microbes encountered by larvae. Culture independent methods have revealed an unexpected high richness of bacterial species associated with larvae, with 100–250 operational taxonomic units associated with one individual. The microbiota of larvae changes rapidly until metamorphosis, most likely due to changes in the selection pressure in the digestive tract caused by changes in host-microbe and microbe-microbe interactions. Even though the microbiota of larvae is distinctly different from the microbiota of the water and the live food, the microbiota of the water strongly affects the microbiota of the larvae. We are in the early phase of understanding larvae-microbe interactions in vivo, but some studies with other animals than fish emphasize that we so far have underestimated the complexity of these interactions. We present examples demonstrating the diversity of these interactions. A large variety of microbial management methods exist, focusing on non-selective reduction of microbes, selective enhancement of microbes, and on improvement of the resistance of larvae against microbes. However, relatively few methods have been studied extensively. We believe that there is a lot to gain by increasing the diversity of approaches for microbial management. As many microbial management methods are perturbations of the microbial community, we argue that ecological theory is needed to foresee and test for longer term consequences in microbe-microbe and microbe-larvae interactions. We finally make some recommendations for future research and development.