Rapid detection of Staphylococcus aureus bacteremia and methicillin resistance by real-time PCR in whole blood samples.

We prospectively evaluated a real-time polymerase chain reaction (PCR) approach for the rapid diagnosis of Staphylococcus aureus bacteremia and presence of the mecA gene in 902 blood samples from 468 infectious episodes of 384 patients. Eight of 12 blood culture (BC)-confirmed samples were positive by the S. aureus-specific PCR. In addition, the mecA gene PCR correctly detected all cases of BC-confirmed methicillin-resistant Staphylococcus aureus (MRSA) infection. A positive PCR result was also obtained in ten of 462 BC-negative infectious episodes, including three patients with culture-confirmed S. aureus infection at other body sites.

Influence of different dietary vitamin C levels on vitamin E and C content and oxidative stability in various tissues and stored m. longissimus dorsi of growing pigs.

Both vitamin E and C have antioxidative properties and may act synergistically. To examine a possible interaction between vitamin E, C and oxidative stability in various tissues, 40 barrows (25-105kg body weight) were allocated to four cereal-based diets (13.4MJ digestible energy/kg, 168g crude protein/kg, 140mg dietary dl-α-tocopheryl acetate) which were supplemented with 0 (B), 150 (C100), 300 (C200) or 600 (C400)mg/kg crystalline ascorbic acid. The influence of storage time on these factors in m. longissimus dorsi samples (LD) was investigated. Samples of liver (LI), heart (HT), spleen (SP), backfat outer layer (BF) and LD were obtained. Dry matter, vitamin E, vitamin C and thiobarbituric acid reactive substances (TBARS) were analyzed in all investigated tissues and in addition color (L(∗), a(∗), b(∗) values) and drip loss measurements in LD at day 0, 1, 2, 3, 4 and 8 were collected. The treatments did not influence growth performance and slaughter data of the pigs. Neither the vitamin E nor the vitamin C content of investigated tissues showed differences due to feeding treatments. There was also a lack of response to the amount in TBARS. Parameters in LD samples were only affected by storage time and not by diets. The results show that dietary use of both vitamins simultaneously did not further improve qualitative characteristics of the investigated pork tissues.

Effect of dietary energy supply and fat source on the fatty acid pattern of adipose and lean tissues and lipogenesis in the pig.

Forty Large White barrows were used to determine whether the effects of dietary fat source (tallow or soy oil at 5% of the diet) on lipogenesis and fatty acid profile of porcine adipose and lean tissue were dependent on dietary digestible energy density (8.8 vs 14.0 MJ DE/kg). Barrows were allocated to one of four groups and offered a fixed amount of feed (170 g x BW0.569/d) from 27 to 105 kg BW. The fatty acid composition of the backfat layers (BF), omental fat (OF), and i.m. adipose tissue of longissimus muscle as well as the activity of lipogenic enzymes of the adipose tissues were determined. Growth performance and carcass characteristics were affected by the dietary energy level (P < 0.01) but not by fat source. In accordance with the lower carcass fat deposition, the activity of lipogenic enzymes were decreased in the low-energy groups (P < 0.01). Within dietary energy level, inclusion of soy oil resulted in increased proportion of PUFA that was compensated by decreased saturated (SFA) and monounsaturated fatty acid (MUFA) proportions (P < 0.01). The SFA changes accounted for 23 (BF) and 24% (OF) of the PUFA changes in the high-energy and 31 (BF) and 39% (OF) in the low-energy diets. The differences in the fatty acid proportions between the soy oil and tallow group were more pronounced in the low-energy groups (fat source x energy density interactions: P < 0.01). Pigs fed the soy oil, low-energy diet had decreased SFA (BF: 28%; OF: 30%) and MUFA (BF: 13%; OF: 19%) concentration, whereas PUFA concentration was increased (BF: 59%; OF: 88%) compared with pigs fed the soy oil, high-energy diet. However, in the tallow groups, pigs fed the low-energy diets had slightly decreased SFA (BF: 14%; OF: 12%) and relatively constant MUFA (BF: 3%; OF: 1%), whereas PUFA concentration increased (BF: 39%; OF: 62%) relative to pigs fed the tallow high-energy diet. Lipid content of the i.m. adipose tissue was decreased in the low-energy groups (P < 0.05). Contrary to what was observed in the adipose tissues, increased PUFA concentration in the neutral and polar lipid fractions of the longissimus muscle was predominantly compensated by reduced MUFA deposition. In the polar lipid fraction, the proportions of both SFA and MUFA were decreased by the low-energy diet. Thus, the extent to which tissue concentration of fatty acids are altered from dietary fats differing in the degree of unsaturation depends on the dietary energy level.

Lactobacillus brevis strain 1E1 administered to piglets through milk supplementation prior to weaning maintains intestinal integrity after the weaning event.

Early colonisation in the gastrointestinal tract by commensal microbes influences the progressive development and maturity of digestive and immune system functionality in the neonate. Application of strategically selected direct-fed microbials to neonatal pigs may provide an opportunity to dictate a portion of the intestinal microbial community and exert a beneficial influence on these developmental processes. Experiments were conducted to determine the effects of early administration of Lactobacillus brevis strain 1E1 to neonatal piglets (n=224) via a milk supplement system on gastrointestinal microbial counts, villous architecture, and immune cell phenotypes during the lactation phase and after weaning. Pigs administered the direct-fed microbial had lower Escherichia coli counts in the jejunum and ileum (P<0.05), and lower coliform counts in the jejunum compared to unsupplemented pigs (P<0.05). The villous height:crypt depth ratio was greater in the ileum at 9 days of age when pigs were provided L. brevis 1E1 compared to unsupplemented pigs (P<0.05), as well as in the duodenum of pigs supplemented with L. brevis 1E1 at 22 days of age (P<0.05). The number of leukocytes expressing CD2 (P<0.05), CD4 (P=0.07) and MHC-II (P=0.07) was lower in the jejunum of pigs administered L. brevis 1E1 compared to unsupplemented pigs, however direct-fed microbial treatment had no effect on the number of leukocytes expressing CD8, CD25 or SWC3. These data demonstrate that early colonisation of the porcine gastrointestinal tract with L. brevis strain 1E1 during the lactation phase influences the progression of intestinal structure, immune system development, and pathogen establishment, indicating a relationship between early microbial colonisation and development of intestinal maturity and integrity.

Effect of source and quantity of dietary vitamin D in maternal and creep diets on bone metabolism and growth in piglets.

Piglets are born with reduced plasma concentrations of 25-hydroxycholecalciferol (25-OH-D(3)) and are thus highly predisposed to vitamin D deficiency. Furthermore, sow milk contains little vitamin D, and the slow intestinal vitamin D absorption of sows limits the efficacy of dietary vitamin D supplementation. Hence, the neonate depends, to a large extent, on the vitamin D stores built up in fetal tissues from maternal sources. The current study was undertaken to evaluate whether the source and quantity of dietary vitamin D provided to the gestating and lactating sow, and also directly in the form of creep feed to the piglet, would influence the vitamin D status, growth performance, and skeletal development of piglets. A total of 39 primiparous and multiparous sows were randomly assigned to 1 of 3 dietary treatments (13 in each treatment), supplemented with either 5 or 50 µg of the commonly used cholecalciferol (vitamin D(3)) or 50 µg of 25-OH-D(3) per kilogram of feed. By wk 3 of lactation, piglets were offered a creep diet with vitamin D supplementation according to the treatment of the dam, and they were offered the same creep diets after weaning at d 35 of age until they reached a BW of approximately 20 kg. When dietary 25-OH-D(3) was provided, circulating concentrations of 25-OH-D(3) in piglet serum increased (P < 0.05) as early as d 21 and later at d 33 and 77, indicating greater body stores in those animals. Bone-breaking strength and cortical bone mineral content and density at the tibial midshaft of piglets were reduced (P < 0.05) when vitamin D(3) was supplemented at 5 µg/kg compared with the bone traits of other groups, but no differences (P > 0.05) were observed between the 2 other groups. After weaning, ADFI was greater (P < 0.05) and growth performance tended (P = 0.08) to improve when doses of 50 µg/kg were administered, regardless of the vitamin D source. In conclusion, supplementation of the diet with 50 µg/kg of either source of vitamin D was proved to be adequate in meeting the needs of gestating sows and in permitting the accumulation of vitamin D in fetal tissues, as well as for normal skeletal mineralization and growth in the offspring. Furthermore, the markedly improved vitamin D status of piglets whose mothers received 25-OH-D(3) possibly resulted from greater tissue reserves present at birth and a greater availability of vitamin D when released from those stores.

Host condition and individual risk of cowpox virus infection in natural animal populations: cause or effect?

Recent studies have provided evidence that endemic pathogens may affect dynamics in animals. However, such studies have not typically considered that infected individuals might have a preceding underlying poor condition. We examined whether individuals in poor condition are more likely to become infected by an endemic pathogen, using as a system the dynamics of cowpox virus in field voles. With data from monthly sampled vole populations, a nested case-control study evaluated whether susceptible individuals with poorer condition had higher probabilities of contracting cowpox. The influence of condition was found to be considerable, especially for males. At times when a susceptible male with good body condition had a relatively low probability of becoming infected, a susceptible male with poor body condition was twice as likely to contract cowpox; if this male was also anaemic, the chances were almost quadrupled. We discuss the care needed when interpreting the findings of wildlife disease studies.

Energy and protein needs of cats for maintenance, gestation and lactation.

In the present investigation, data on the energy intakes and energy needs, as well as protein and fat accretion, of queens during pregnancy, during lactation and after lactation are given. Eleven adult cats were used as experimental animals. Data were collected during the fourth and seventh week of pregnancy, the second and sixth week of lactation and the second and sixth week after lactation. The cats were fed dry kitten food. During gestation and after lactation, all measurements were performed with respiration chambers. During lactation, balance trials without respiration chambers were performed. Body weight was measured and nitrogen, carbon and energy balances were calculated. From these, protein and fat accretion, as well as the metabolisable energy intake, was calculated. The weight gain during gestation was linearly independent of the number of kittens. During lactation, all cats lost weight; nevertheless, all cats except one were heavier 2 weeks after lactation than at mating. The energy intake of the cats during gestation was 1.8 times the maintenance requirement in the fourth week and two times maintenance requirement in the seventh week, and these energy intakes differed greatly among individuals. The energy intake of the cats during lactation was clearly higher than that recommended by National Research Council (NRC)(1), whereas the recommended protein intake in the second week of lactation was met. As the calculated protein balance was negative, the NRC recommendation for protein intake seems to be too low. In comparison to previous data, the cats showed a higher energy intake during lactation (median 502kJ/kgBW/d, second week lactation), and the weight loss was much lower. Further investigations on pregnant and lactating cats are necessary to complete the database.

Additional data on energy requirements of young adult cats measured by indirect calorimetry.

Measurements of nitrogen, carbon and energy balances were done on eight female adult cats aged approximately 1.5 years with the help of respiration chambers. The cats were fed with a marked dry food for kittens (Biomill kitten). Faeces and urine samples were collected with the help of a special cat toilet that allows the single collection of both materials. The calculated energy requirement of these eight female cats was 239.6 kJ ME/kg BW/day (R(2) = 0.98). Including the data, on 12 young male adult cats, of Läuger, (2001), an energy requirement of 238 kJ ME/kg BW/day (R(2) = 0.95) could be calculated. From these data, it can be followed that the energy requirements of young, active cats are higher than that of the older cats. The method of indirect calorimetry does not necessarily lead to lower energy requirements.

Radiation hybrid mapping of 18 positional and physiological candidate genes for arthrogryposis multiplex congenita on porcine chromosome 5.

We report the chromosomal assignment of 18 porcine genes to human homologues using the INRA-Minnesota swine radiation hybrid panel (IMpRH). These genes (CACNA1C, COL2A1, CPNE8, C3F, C12ORF4, DDX11, GDF11, HOXC8, KCNA1, MDS028, TMEM106C, NR4A1, PHB2, PRICKLE1, Q6ZUQ4, SCN8A, TUBA8 and USP18) are located on porcine chromosome 5 (SSC5) and represent positional and functional candidates for arthrogryposis multiplex congenita (AMC), which maps to SSC5. CPNE8, PRICKLE1, Q6ZUQ4 and TUBA8 were mapped to the interval for pig AMC between microsatellites SW152 and SW904. Three SNPs in TUBA8 co-segregated with the AMC phenotype in 230 pigs of our research population without recombination and could be used as a genetic marker test for AMC. In addition, we provide evidence that a small chromosomal region of HSA22q11.2 evolutionarily corresponds to SSC5q12-q22 (and contains the human homologues of porcine SW152, Q6ZUQ4, TUBA8 and USP18), while the regions flanking HSA22q11.2 on SSC5 correspond to HSA12p13 and HSA12q12. We identified seven distinct chromosomal blocks, further supporting extensive rearrangements between genes on HSA12 and HSA22 in the AMC region on SSC5.

A new quality-of-life measure for men experiencing involuntary childlessness.

BACKGROUND: Infertility may considerably reduce quality-of-life. Many of the existing generic quality-of-life measures, which often focus on physical impairments, do not represent the specific complaints of infertile patients. In this article, we report on the development and validation of the TLMK (Tübinger Lebensqualitätsfragebogen für Männer mit Kinderwunsch), an instrument for measuring quality-of-life in male patients with involuntary childlessness. METHODS: The first version of the questionnaire, which consisted of 91 items, was administered to 275 men who attended andrology and gynaecology clinics for fertility evaluations. After the questionnaires were scored, item analysis and reduction, principal component analysis and internal consistency analyses were conducted. RESULTS: The final version of the TLMK consists of 35 items in four scales and provides an internally consistent quality-of-life profile for men experiencing involuntary childlessness. Convergent and discriminant validity was supported through the correlation of the TLMK scales with established questionnaires on life satisfaction (FLZ) and partnership (PFB). CONCLUSION: The TLMK provides information about the quality-of-life in men experiencing involuntary childlessness and was found to be easy to administer and acceptable to patients. It may be used to assess patients' baseline and ongoing quality-of-life during fertility treatment and as an outcome variable in the evaluation of integrated psychological counselling.

Comparative pharmacokinetics and clinical experience with a new cephalosporin-derivative: cefazolin.

In a cross-over study in 12 normal individuals, the pharmacokinetic parameters of cefalotin, cefradine and cefazolin were determined after intravenous injection of 1,000 mg of each substance. The microbiological activities in urine and serum were determined using the agar diffusion test; the pharmacokinetic data were calculated by a computer system on the basis of a Fortran programme. Cefazolin has significantly higher serum concentrations than the other two cephalosporins, distinctly longer serum half-lives, higher protein binding, and smaller apparent volumina of distribution. In 36 inpatients with mainly chronical and acute infections of the urinary tract, we tested the antibacterial effectivity, the compatibility, and the application modalities of cefazolin. In 29 patients we had a satisfactory clinical result, in 25 cases we achieved the elimination of bacteria by the end of the therapy. The compatibility of cefazolin was good; apart from a minor, reversible, liver-specific increase in enzymes in 6 patients, no side effects could be detected.

Vitamin E content of different animal products: influence of animal nutrition.

The alpha-tocopherol content of different meat cuts was examined. Chicken thigh had the highest vitamin E content, followed by chicken breast and pork shoulder (p < 0.05). The lowest concentrations were found in longissimus dorsi muscle from pork, beef, veal and in beef shoulder. Considering the average daily lean meat consumption (105 g) in Switzerland, recommendation for daily vitamin E intake was met to 3%. Supplementation of 200 mg alpha-tocopherol acetate/kg feed to pigs and laying hens significantly increased the alpha-tocopherol content in all examined products. The alpha-tocopherol accumulation differed according to the following ranking: egg yolk > liver > adipose tissue > musculus longissimus dorsi. The alpha-tocopherol:energy ratios were 28.8, 7.3, 0.9 and 1.2 mg/MJ for egg yolk, liver, adipose tissue and longissimus dorsi muscle of the vitamin E supplemented groups, respectively. The results showed that meat, with the exception of chicken thigh, is not an important supplier of vitamin E, not even from animals fed a vitamin E enriched diet. Egg yolk became a good source of vitamin E for human nutrition by dietary modification.

Functional interaction between the human cytomegalovirus 86-kilodalton IE2 protein and the cellular transcription factor CREB.

The 86-kDa IE2 protein (IE86) of human cytomegalovirus (HCMV) has been described as a promiscuous transactivator of viral, as well as cellular, gene expression. Investigation of the mechanism used by IE86 to activate gene expression from the early UL112/113 promoter of HCMV revealed the existence of three binding sites for IE86 located between nucleotides -290 and -120 relative to the transcriptional start site (H. Arlt, D. Lang, S. Gebert, and T. Stamminger, J. Virol. 68:4117-4125, 1994). As shown previously, deletion of these target sites resulted in a reduction of IE86-mediated transactivation by approximately 70%. The remaining promoter, however, could still be stimulated about 40-fold, indicating the presence of an additional responsive element within these sequences. Here, we provide evidence that a binding site for the cellular transcription factor CREB can also act as a target for IE86 transactivation. By DNase I protection analysis, a binding sequence for CREB could be detected between nucleotides -78 and -56 within the respective promoter region. After in vitro mutagenesis of this CREB-binding site within the context of the entire UL112/113 promoter, a marked reduction in transactivation levels was evident. Moreover, when individual CREB-binding sites were positioned upstream of a minimal, TATA box-containing UL112/113 promoter, they were able to confer strong IE86 responsiveness, whereas a mutated sequence did not exert any effect. In far Western blot and pull-down experiments, a direct interaction of IE86 with the cellular transcription factor CREB could be observed. The in vivo relevance of this in vitro interaction was confirmed by using various GAL4 fusion proteins in the presence or absence of IE86 which revealed a strong activation only in the presence of both a GAL4-CREB fusion and IE86. This shows that at least one specific member of the ATF/CREB family of transcription factors is involved in mediating transactivation by the HCMV IE86 protein.

[Chromium supplements in the feed for growing pigs and meat quality]. / Chromzulagen zum Schweinemastfutter: Einfluss auf Wachstum und Fleischqualität.

An experiment with 40 female growing pigs from 27.4 to 106.5 kg body weight (BW) in individual pens was conducted to evaluate the effect of different chromium supplements (Cr-chloride, Cr-yeast and Cr-picolinate) according to 0.5 ppm Cr in the diet compared with a control diet without any additional chromium. The influence on growth performance and carcass as well as meat composition was studied. In comparison with the control diet body weight gain and feed conversion ratio in the finishing period (60 to 106.5 kg BW) were significant increased and lowered respectively in the treatment with Cr-chloride and with the other Cr supplements tendentially. The results of the carcass composition as well as the fatty acid profile of neutral and complex lipids in the muscle (M. longissimus dorsi) at the 10th rib did not indicate a statistically significant effect of the Cr supplements. Furthermore energy utilization on the base of digestibility was not affected. Concerning the Longissimus muscle area and the intramuscular fat content there were positive tendencies of the investigated Cr supplements.

Identification of binding sites for the 86-kilodalton IE2 protein of human cytomegalovirus within an IE2-responsive viral early promoter.

The 86-kDa IE2 protein (IE86) of human cytomegalovirus (HCMV) can act as both an activator and a repressor of gene expression. The mechanisms for both of these functions are not well defined. It has recently been demonstrated that this protein has sequence-specific DNA binding properties: it interacts directly with a target sequence that is located between the TATA box and the cap site of its own promoter. This sequence, termed the CRS (cis repression signal) element, is required for negative autoregulation of the IE1/IE2 enhancer/promoter by IE2. We demonstrate now that binding of this protein to DNA is not confined to this site but occurs also within an early promoter of HCMV that has previously been shown to be strongly IE2 responsive. By DNase I protection analysis using a purified, procaryotically expressed IE2 protein, we could identify three binding sites within the region of -290 to -120 of the UL112 promoter of HCMV. Competition in DNase I protection experiments as well as gel retardation experiments showed that the identified binding sites are specific and have high affinity. Deletion of IE2 binding sites from this promoter reduced the level of transactivation; however, the remaining promoter could still be stimulated about 40-fold. Constructs in which IE2 binding sites were fused directly to the TATA box of the UL112 promoter did not reveal a significant contribution of these sequences to transactivation. However, if an IE2 binding site was reinserted upstream of nucleotide -117 of the UL112 promoter, an increase in transactivation by IE2 was obvious, whereas a mutated sequence could not mediate this effect. This finding suggests that DNA-bound IE2 can contribute to transactivation but seems to require the presence of additional transcription factors. Moreover, a comparison of the detected IE2 binding sites could not detect a strong homology, suggesting that this protein may be able to interact with a broad spectrum of different target sequences.

Studies on modifying the tackiness and drug release of Kollicoat EMM 30 D coatings.

In the search for antitack additives for Kollicoat EMM 30 D (ethyl acrylate-methyl methacrylate 30% dispersion, Ph. Eur.) film coatings, various possibilities were investigated. The best results were obtained using a combination of simethicone and talc. This mixture was tested on propranolol, theophylline, and verapamil HCl blank pellets in a previously developed Kollicoat EMM 30 D basic formulation. Almost any desired drug release rate can be obtained with all three pellet formulations by varying the two pore formers hypromellose 3mPas and microcrystalline cellulose type 105. A thin application of colloidal silica onto the coated pellets additionally prevents them from sticking together during storage.

Influence of dietary vitamin E and C supplementation on vitamin E and C content and thiobarbituric acid reactive substances (TBARS) in different tissues of growing pigs.

To investigate the influence and possible interactions of dietary vitamin E and C supplementation on vitamin content of both vitamins and oxidative stability of different pork tissues 40 Large White barrows from 25 kg to 106 kg were allocated to four different cereal based diets: Basal diet (B), dl-alpha-tocopherylacetate + 200 mg/kg (E), crystalline ascorbic acid + 300 mg/kg (C) or both vitamins (EC). At slaughtering samples of liver, spleen, heart, kidney, backfat outer layer, ham and M. tongissimus dorsi were obtained. Growth performance of the pigs and carcass characteristics were not influenced by feeding treatments. Dietary vitamin E supplementation had a significant effect on the vitamin E and alpha-tocopherol concentration in all investigated tissues. Backfat outer layer, liver, spleen, kidney and heart had higher vitamin E concentrations than ham and M. longissimus dorsi. Dietary vitamin C supplementation tended towards enhanced vitamin E levels except for ham samples. Therefore, some synergistic actions without dietary vitamin E supplementation between the two vitamins could be shown. The vitamin C concentration and TBARS were increased or at least equal in all tissues due to vitamin C supplementation. Dietary alpha-tocopherol supplementation resulted in lower TBARS in backfat outer layer (malondialdehyde 0.35 mg/kg in B vs. 0.28 mg/kg in E), but increased in heart and ham. When both vitamins were supplemented (EC) TBARS were lower in M. longissimus dorsi and backfat outer layer, equal in heart and higher in liver and ham compared to a single vitamin C supplementation. Rancimat induction time of backfat outer layer was 0.3 h higher in C compared to B and 0.17 h higher in EC than in E. Correlations between levels of both vitamins were positive for kidney (r = 0.169), M. longissimus dorsi (r = 0.499) and ham (r = 0.361) and negative for heart (r = -0.350). In liver and spleen no interaction could be found. In backfat outer layer vitamin E was positively correlated with rancimat induction time (r = 0.550) and negatively with TBARS (r = -0.202), but provided no evidence that dietary vitamin E supply led to better oxidative stability.

The UL84 protein of human cytomegalovirus acts as a transdominant inhibitor of immediate-early-mediated transactivation that is able to prevent viral replication.

The 86-kilodalton immediate-early (IE) 2 protein (IE2-p86) of human cytomegalovirus (HCMV) is a multifunctional regulator of HCMV gene expression which appears to be essential for triggering the lytic replicative cycle. IE2-p86 functions as a promiscuous transactivator of both viral and cellular gene expression and can repress transcription from its own promoter. In this study we demonstrate that a viral early protein, termed pUL84, which is able to interact with IE2-p86 both in vivo and in vitro, modulates IE2-p86 in a specific manner. First, pUL84 acts as a transdominant inhibitor of IE2-p86-mediated transactivation of both homologous and heterologous promoters. Second, negative autoregulation by IE2-p86 is augmented in the presence of pUL84. Using two in vivo assays, we obtained evidence that expression of pUL84 during the IE phase of the viral replicative cycle leads to an inhibition of viral early gene expression which prevents replication of HCMV and results in a persistent infection of UL84-positive cell lines. Transdominant inhibition of a viral IE function by a protein expressed during the later phases of replication appears to be a novel principle used by herpesviruses which could account for the slow replication of HCMV and may be useful in the development of new antiviral strategies.