Towards a bioengineered uterus: bioactive sheep uterus scaffolds are effectively recellularized by enzymatic preconditioning.

Uterine factor infertility was considered incurable until recently when we reported the first successful live birth after uterus transplantation. However, risky donor surgery and immunosuppressive therapy are factors that may be avoided with bioengineering. For example, transplanted recellularized constructs derived from decellularized tissue restored fertility in rodent models and mandate translational studies. In this study, we decellularized whole sheep uterus with three different protocols using 0.5% sodium dodecyl sulfate, 2% sodium deoxycholate (SDC) or 2% SDC, and 1% Triton X-100. Scaffolds were then assessed for bioactivity using the dorsal root ganglion and chorioallantoic membrane assays, and we found that all the uterus scaffolds exhibited growth factor activity that promoted neurogenesis and angiogenesis. Extensive recellularization optimization was conducted using multipotent sheep fetal stem cells and we report results from the following three in vitro conditions; (a) standard cell culturing conditions, (b) constructs cultured in transwells, and (c) scaffolds preconditioned with matrix metalloproteinase 2 and 9. The recellularization efficiency was improved short-term when transwells were used compared with standard culturing conditions. However, the recellularization efficiency in scaffolds preconditioned with matrix metalloproteinases was 200-300% better than the other strategies evaluated herein, independent of decellularization protocol. Hence, a major recellularization hurdle has been overcome with the improved recellularization strategies and in vitro platforms described herein. These results are an important milestone and should facilitate the production of large bioengineered grafts suitable for future in vivo applications in the sheep, which is an essential step before considering these principles in a clinical setting.

Caries incidence and risk assessment during a five-year period in adolescents living in south-eastern Sweden.

OBJECTIVES: The aim was to examine the caries incidence in adolescents using the Public Dental Service (PDS) during a 5-year period in relation to their caries experience at baseline and risk classification. METHODS: A 5-year retrospective cohort study based of the dental records from 17 PDS clinics in south-eastern Sweden was conducted; 159 individuals born in 1997 were included, and their caries risk was classified at 12 and 17 years of age. Caries prevalence and documented risk groups were assessed at baseline and after 5 years. RESULTS: The increment of caries (both initial and manifest caries) was higher, to a statistically significant degree, after 5 years in adolescents who were recorded as caries-free at baseline compared to individuals with caries at baseline (P < .001). In individuals with caries at baseline, the greatest increment of caries was found at approximal sites (P < .001). At baseline, individuals were classified as low (94%), medium (6%) and high risk (0%). After 5 years, the figures were 74%, 20% and 6%, respectively. Although classified in a low-caries risk group, 9% had ≥6 decayed or filled surfaces at baseline, and 23% did after 5 years. Approximately 62% of individuals were registered as caries-free at baseline, and 45% were after 5 years. CONCLUSIONS: There was an increase in caries over 5 years, especially among adolescents without caries experience at baseline. The majority of adolescents had the same risk classification after 5 years. Further research with a larger sample size is needed to evaluate risk assessment for caries.

Experimental inoculation of Treponema pedis T A4 failed to induce ear necrosis in pigs.

Ear necrosis is a syndrome affecting pigs shortly after weaning and is regarded as an animal welfare issue. The etiology is unknown but Treponema spp., predominantly Treponema pedis, are commonly detected in the lesions. Oral treponemes have been suggested as source of infection, transferred by biting and licking behavior. In this study, five pigs were intradermally inoculated with Treponema pedis strain T A4 with the aim of investigating if this strain would induce ear lesions. Three pigs served as controls. The inoculation was repeated after 29 days, and the study continued for 56 days. Serum samples were collected throughout the study and analyzed by ELISA for IgG antibodies towards T. pedis T A4 lysate. Skin biopsies were taken from the inoculation area at the end of the study. Gingival samples were collected and cultivated for treponemes, for comparison to the inoculation strain and to follow colonisation. The challenged pigs did not develop any clinical signs of infection and no spirochetes were detected in sections from skin biopsies. The number of Treponema-positive gingival samples increased during the study. In the challenge group, IgG towards the bacterial lysate peaked 7 days after each inoculation and decreased rapidly hereafter. In the control group a weak IgG response was observed after the second inoculation, possibly caused by the oral treponemes.

Identification of Treponema pedis as the predominant Treponema species in porcine skin ulcers by fluorescence in situ hybridization and high-throughput sequencing.

Skin lesions often seen in pig production are of great animal welfare concern. To study the potential role of Treponema bacteria in porcine skin ulcers, we investigated the presence and distribution of these organisms in decubital shoulder ulcers (n=51) and ear necroses (n=54) by fluorescence in situ hybridization (FISH) and high-throughput sequencing. In addition, two cases of facial ulcers and five cases of other skin ulcers were included in the study. Samples from all 112 skin lesions and intact skin from pigs without skin ulcers (n=14) were screened by FISH. Three different oligonucleotide probes targeting 16S rRNA were used, specific for domain bacterium, Treponema spp. and species T. pedis. Screening showed that two cases each of facial and other ulcers, 35 (69%) of shoulder ulcers and 32 (59%) of ear necroses were positive for Treponema spp. T. pedis was the unequivocally, predominant species typically constituting more than 90% of the treponemes in a lesion, assessed visually by microscopy. Altogether, T. pedis was demonstrated in 69 of the 71 Treponema spp. positive lesions. We conclude that Treponema spp. are frequently present and abundant in various skin ulcers of pigs. The results from this study point toward an important role of T. pedis as a secondary bacterial infection in porcine skin ulcers, especially in severe and chronic lesions.

Characterization of Treponema spp. isolates from pigs with ear necrosis and shoulder ulcers.

Ear necrosis and shoulder ulcers in pigs are animal welfare problems and ethical issues that can cause economic losses for producers. Spirochetes have been observed microscopically in scrapings from pig ulcers since the early 1900s, but have until recently not been cultured and therefore not characterized. In this study, 12 Treponema spp. isolates were acquired from porcine ear necrosis, shoulder ulcers and gingiva. DNA analysis of the 16S rRNA-tRNA(Ile) intergenic spacer region (ISR2) or the 16S rRNA gene revealed relatedness to oral treponemes found in dogs and humans. All isolates except one aligned into two clusters, Treponema pedis and Treponema sp. OMZ 840-like. The 16S rRNA gene of the remaining isolate shared 99% nucleotide identity with Treponema parvum. Genetic fingerprinting of the isolates was performed through random amplification of polymorphic DNA (RAPD). In addition, the isolates were characterized by biochemical tests, including api(®)ZYM, tryptophanase and hippuricase activity, and by testing the antimicrobial susceptibility to tiamulin, valnemulin, tylosin, tylvalosin, lincomycin and doxycycline using broth dilution. All isolates except two showed unique RAPD fingerprints, whereas metabolic activity tests could not differentiate between the isolates. The MICs of all antimicrobial agents tested were low.

Occurrence of Treponema spp. in porcine skin ulcers and gingiva.

Porcine shoulder ulcers and ear necrosis are a significant animal welfare concern and impair efficient livestock production. Although spirochetes have been detected in both types of lesions the potential role of these bacteria in lesion propagation has received little attention. The objective of this study was to investigate the occurrence of spirochetes of the genus Treponema in shoulder ulcers or ear necrosis in pigs and compare these with treponemes from porcine gingiva. Samples were collected from gingiva and necrotic ulcers in 169 pigs. Presence of spirochetes was observed in silver stained histological sections and by phase contrast microscopy in scrapings from the necrotic lesions. Additionally, PCR of the 16SrRNA-tRNA(Ile) intergenic spacer region (ISR2) was used to detect Treponema spp. in all samples. Combined analysis showed that 73% of the shoulder ulcers and 53% of the ear necroses were positive for spirochetes. Treponema spp. were detected in 9.7% of the gingival samples. Comparative DNA sequence analysis of the ISR2 sequences revealed the presence of three distinct genetic phylotypes of Treponema spp. corresponding to Treponema pedis, and as yet two unnamed phylotypes represented by GenBank sequences C1UD1 (Acc. No. AY342041) and C1BT2-8 (Acc. No. AY342046). Detection of identical ISR2 sequences from gingiva and ulcer samples indicates that oral Treponema spp. are spread from mouth to ulcer. We conclude that Treponema spp. frequently occur in shoulder ulcers and ear necrosis in pigs, and suggest a possible infection route through biting and licking.

Gender differences when using sedative music during colonoscopy.

Colonoscopy is a procedure often experienced as uncomfortable and worrying. Music has been reported to reduce discomfort during colonoscopy; however, no study in a Swedish setting has been found. The purpose of this randomized controlled trial was to analyze the effects of sedative music on patients' experience of anxiety, pain, relaxation, and well-being during colonoscopy. Prior to colonoscopy, adult patients (n = 120), aged 18-80 years, were randomly assigned to either an intervention group (n = 60) who listened to sedative instrumental music with 60-80 beats per minute during the colonoscopy or a control group. After the colonoscopy, both groups completed a questionnaire on anxiety, the State Trait Anxiety Inventory, and an anxiety Visual Analogue Scale. Pain, relaxation, and well-being were also measured with Visual Analogue Scales. Women in the intervention group had a lower level of anxiety during the colonoscopy than those in the control group (p = .007) and well-being was significantly higher in the intervention group, especially among men, than in the controls (p = .006 and p = .025, respectively). Men in the intervention group were more relaxed during the colonoscopy than those in the control group (p = .065). Listening to sedative music decreased anxiety among women and increased well-being among men during colonoscopy.

Low occurrence of Clostridium difficile in retail ground meat in Sweden.

This pilot study was conducted to evaluate the occurrence of Clostridium difficile in samples of ground meat in Sweden. From April to September 2008, 82 meat samples were collected from randomly selected retail shops in Uppsala County (central Sweden). C. difficile was isolated from 2 (2.4%; both ground beef) of the 82 meat samples. No C. difficile was detected in pork, hamburger, sheep, poultry, or other type of meat samples. The two C. difficile isolates produced both toxin A and toxin B. These findings indicate that C. difficile might be present in ground meat samples in Sweden. However, further studies are necessary to confirm these preliminary data and to elucidate the public health significance of meat contamination by C. difficile in Sweden.