Analysis of transmission-related third-generation cephalosporin-resistant Enterobacterales by electronic data mining and core genome multi-locus sequence typing.

BACKGROUND: To contain intra-hospital transmission of third-generation cephalosporin-resistant Enterobacterales (3GCR-E), contact isolation precautions are recommended. AIM: To quantify transmissions of 3GCR Escherichia coli and 3GCR Klebsiella pneumoniae within a hospital. METHODS: An automated outbreak detection system (AODS) was used to identify clusters (N≥2) of 3GCR Enterobacterales for the years 2016, 2018 and 2020. Clusters were defined by phenotypic agreement of microbiological results and spatial and temporal relationship. Core genome multi-locus sequence typing (cgMLST) was used to confirm whether the cluster isolates were transmitted between patients. FINDINGS: A total of 4343 3GCR E. coli and 1377 K. pneumoniae isolates were analysed. Among the 3GCR E. coli isolates, the AODS identified 304 isolates as cluster isolates, the median cluster size was two (range: 2-5). The cgMLST analysis revealed that a total of 23 (7.5%) 3GCR E. coli cluster isolates were transmission-associated, of which 20 isolates (87%) were detected in intensive care patients. Among the 3GCR K. pneumoniae isolates, the AODS identified 73 isolates as cluster isolates, the median cluster size was two (range: 2-4). CgMLST revealed that 35 (48%) 3GCR K. pneumoniae cluster isolates were transmission associated, of which 27 isolates (77%) were detected in intensive care patients. CONCLUSION: For 3GCR K. pneumoniae, cgMLST confirmed the AODS results more frequently than for 3GCR E. coli. Therefore, contact isolation precautions for 3GCR K. pneumoniae may be appropriate on intensive care units, but only in certain circumstances, such as outbreaks, for Enterobacterales with lower transmissibility, such as E. coli.

Blood and liver telomere length, mitochondrial DNA copy number, and hepatic gene expression of mitochondrial dynamics in mid-lactation cows supplemented with l-carnitine under systemic inflammation.

The current study was conducted to examine the effect of l-carnitine (LC) supplementation on telomere length and mitochondrial DNA copy number (mtDNAcn) per cell in mid-lactation cows challenged by lipopolysaccharide (LPS) in blood and liver. The mRNA abundance of 31 genes related to inflammation, oxidative stress, and the corresponding stress response mechanisms, the mitochondrial quality control and the protein import system, as well as the phosphatidylinositol 3-kinase/protein kinase B pathway, were assessed using microfluidics integrated fluidic circuit chips (96.96 dynamic arrays). In addition to comparing the responses in cows with or without LC, our objectives were to characterize the oxidative and inflammatory status by assessing the circulating concentration of lactoferrin (Lf), haptoglobin (Hp), fibrinogen, derivates of reactive oxygen metabolites (dROM), and arylesterase activity (AEA), and to extend the measurement of Lf and Hp to milk. Pluriparous Holstein cows were assigned to either a control group (CON, n = 26) or an LC-supplemented group (CAR; 25 g LC/cow per day; d 42 ante partum to d 126 postpartum (PP), n = 27). On d 111 PP, each cow was injected intravenously with LPS (Escherichia coli O111:B4, 0.5 µg/kg). The mRNA abundance was examined in liver biopsies of d -11 and +1 relative to LPS administration. Plasma and milk samples were frequently collected before and after the challenge. After LPS administration, circulating plasma fibrinogen and serum dROM concentrations increased, whereas AEA decreased. Moreover, serum P4 initially increased by 3 h after LPS administration and declined thereafter irrespective of grouping. The Lf concentrations increased in both groups after LPS administration, with the CAR group showing greater concentrations in serum and milk than the CON group. After LPS administration, telomere length in blood increased, whereas mtDNAcn per cell decreased; however, both remained unaffected in liver. For mitochondrial protein import genes, the hepatic mRNA abundance of the translocase of the mitochondrial inner membrane (TIM)-17B was increased in CAR cows. Moreover, TIM23 increased in both groups after LPS administration. Regarding the mRNA abundance of genes related to stress response mechanisms, 7 out of 14 genes showed group × time interactions, indicating a (local) protective effect due to the dietary LC supplementation against oxidative stress in mid-lactating dairy cows. For mtDNAcn and telomere length, the effects of the LPS-induced inflammation were more pronounced than the dietary supplementation of LC. Dietary LC supplementation affected the response to LPS primarily by altering mitochondrial dynamics. Regarding mRNA abundance of genes related to the mitochondrial protein import system, the inner mitochondrial membrane translocase (TIM complex) seemed to be more sensitive to dietary LC than the outer mitochondrial membrane translocase (TOM complex).

Antimicrobial Activity of Ceftolozane-Tazobactam, Ceftazidime-Avibactam, and Cefiderocol against Multidrug-Resistant Pseudomonas aeruginosa Recovered at a German University Hospital.

Multidrug-resistant (MDR) Pseudomonas aeruginosa increasingly causes health care-associated infections. In this study, we determined the activity of ceftolozane-tazobactam, ceftazidime-avibactam, and cefiderocol against 223 MDR P. aeruginosa clinical isolates recovered from 2013 to 2017 at the University Hospital Frankfurt by using MIC test strips. Furthermore, we evaluated the presence of genes encoding major ß-lactamases, such as VIM, IMP, NDM, GIM, SPM, and KPC; the extended spectrum ß-lactamase (ESBL)-carbapenemase GES; and the virulence-associated traits ExoS and ExoU, as in particular ExoU is thought to be associated with poor clinical outcome. For MDR P. aeruginosa isolates, the MIC50/MIC90 values of ceftolozane-tazobactam, ceftazidime-avibactam, and cefiderocol were 8/>256 mg/L, 16/>256 mg/L, and 0.25/1 mg/L, respectively. Cefiderocol showed the highest susceptibility rate (97.3%) followed by ceftazidime-avibactam (48.4%) and ceftolozane-tazobactam (46.6%). In 81 (36.3%) isolates, carbapenemase gene blaVIM was detected, and in 5 (2.2%) isolates, blaGES was detected (with a positive association of exoU and blaVIM). More than half of the isolates belong to the so-called international P. aeruginosa "high-risk" clones, with sequence type 235 (ST235) (24.7%) being the most prevalent. This study underlines that ceftolozane-tazobactam, ceftazidime-avibactam, and cefiderocol are important options for the treatment of infections due to MDR P. aeruginosa, with cefiderocol currently being the most active available antipseudomonal ß-lactam agent. According to our clinical experience, the outcome of cefiderocol therapy (8 patients) was favorable especially in cases of MDR P. aeruginosa-associated complicated urinary tract infections. IMPORTANCE After testing ceftolozane-tazobactam, ceftazidime-avibactam, and cefiderocol against a collection of 233 multidrug-resistant (MDR) Pseudomonas aeruginosa, we showed that cefiderocol is the most active antipseudomonal ß-lactam agent (susceptibility rates were 46.6%, 48.4%, and 97.4%, respectively). The most prevalent one was sequence type 235 (ST235) (24.7%), followed by ST244, ST175, and ST233, with all belonging to the top 10 P. aeruginosa high-risk clones with worldwide distribution. Our data indicate that during surveillance studies special attention should be paid to the MDR and highly virulent VIM- and ExoU-producing variant of ST235. Furthermore, in the case of infections caused by carbapenemase-producing MDR P. aeruginosa, cefiderocol is the preferred treatment option, while outcomes of complicated urinary tract infections and hospital-acquired pneumonia with cefiderocol were favorable.

Prolonged Orbital Relaxation by Locally Modified Phonon Density of States for the SiV

Coherent quantum systems are a key resource for emerging quantum technology. Solid-state spin systems are of particular importance for compact and scalable devices. However, interaction with the solid-state host degrades the coherence properties. The negatively charged silicon vacancy center in diamond is such an example. While spectral properties are outstanding, with optical coherence protected by the defects symmetry, the spin coherence is susceptible to rapid orbital relaxation limiting the spin dephasing time. A prolongation of the orbital relaxation time is therefore of utmost urgency and has been tackled by operating at very low temperatures or by introducing large strain. However, both methods have significant drawbacks: the former requires use of dilution refrigerators and the latter affects intrinsic symmetries. Here, a novel method is presented to prolong the orbital relaxation with a locally modified phonon density of states in the relevant frequency range, by restricting the diamond host to below 100 nm. Subsequently measured coherent population trapping shows an extended spin dephasing time compared to the phonon-limited time in a pure bulk diamond. The method works at liquid helium temperatures of few Kelvin and in the low-strain regime.

Blood and adipose tissue steroid metabolomics and mRNA expression of steroidogenic enzymes in periparturient dairy cows differing in body condition.

In high-yielding dairy cows, the rapidly increasing milk production after parturition can result in a negative nutrient balance, since feed intake is insufficient to cover the needs for lactation. Mobilizing body reserves, mainly adipose tissue (AT), might affect steroid metabolism. We hypothesized, that cows differing in the extent of periparturient lipomobilization, will have divergent steroid profiles measured in serum and subcutaneous (sc)AT by a targeted metabolomics approach and steroidogenic enzyme profiles in scAT and liver. Fifteen weeks antepartum, 38 multiparous Holstein cows were allocated to a high (HBCS) or normal body condition (NBCS) group fed differently until week 7 antepartum to either increase (HBCS BCS: 3.8 ± 0.1 and BFT: 2.0 ± 0.1 cm; mean ± SEM) or maintain BCS (NBCS BCS: 3.0 ± 0.1 and BFT: 0.9 ± 0.1 cm). Blood samples, liver, and scAT biopsies were collected at week -7, 1, 3, and 12 relative to parturition. Greater serum concentrations of progesterone, androsterone, and aldosterone in HBCS compared to NBCS cows after parturition, might be attributed to the increased mobilization of AT. Greater glucocorticoid concentrations in scAT after parturition in NBCS cows might either influence local lipogenesis by differentiation of preadipocytes into mature adipocytes and/or inflammatory response.

Improvement in tinnitus distress, health-related quality of life and psychological comorbidities by cochlear implantation in single-sided deaf patients.

BACKGROUND: Patients with single-sided deafness (SSD) lack the ability localize sound sources and have difficulty with speech-in-noise hearing. In addition, there is a high prevalence of tinnitus distress. These problems may result in reduced health-related quality of life (HRQoL) and psychological comorbidities. This study aimed to investigate the influence of treatment with a cochlear implant (CI) on HRQoL, tinnitus distress, psychological comorbidities, and audiological parameters in SSD patients. METHODS: This retrospective study included 20 patients with postlingually acquired SSD (13 women, 7 men, mean age 57.0 years). Data on HRQoL were collected with the Nijmegen Cochlear Implant Questionnaire (NCIQ) and the Medical Outcome Study Short Form 36 Survey (SF-36). Tinnitus distress was assessed with the Tinnitus Questionnaire (TQ), psychological comorbidities were evaluated with validated questionnaires, and speech perception and hearing ability were measured with validated speech tests. RESULTS: Postoperatively, the mean total NCIQ score (p < 0.0001) and four subscores improved significantly, as did tinnitus distress (p < 0.05). Anxiety symptoms (Generalized Anxiety Disorder questionnaire, GAD-7) decreased significantly after CI. Speech perception improved significantly, particularly "hearing with background noise" (p < 0.05, Oldenburg Inventory, OI) and "localization" (p < 0.001, OI), as well as the Oldenburg Sentence Test (OlSa) scores for with vs. without CI when speech was presented from the SSD side and noise was presented from the normal hearing side (SSSDNNH; p < 0.005). CONCLUSION: CI in SSD patients is a powerful procedure to improve HRQoL, reduce tinnitus distress, and improve psychological comorbidities. Additionally, it is beneficial for hearing improvement, particularly in noise and for directional hearing.

[Improvement in tinnitus distress, health-related quality of life and psychological comorbidities by cochlear implantation in single-sided deaf patients. German version]. / Verbesserung von Tinnitusdistress, Lebensqualität und psychologischen Komorbiditäten durch Cochleaimplantation einseitig ertaubter Patienten.

BACKGROUND: Patients with single-sided deafness (SSD) lack the ability localize sound sources and have difficulty with speech-in-noise hearing. In addition, there is a high prevalence of tinnitus distress. These problems may result in reduced health-related quality of life (HRQoL) and psychological comorbidities. This study aimed to investigate the influence of treatment with a cochlear implant (CI) on HRQoL, tinnitus distress, psychological comorbidities, and audiological parameters in SSD patients. METHODS: This retrospective study included 20 patients with postlingually acquired SSD (13 women, 7 men, mean age 57.0 years). Data on HRQoL were collected with the Nijmegen Cochlear Implant Questionnaire (NCIQ) and the Medical Outcome Study Short Form 36 Survey (SF-36). Tinnitus distress was assessed with the Tinnitus Questionnaire (TQ), psychological comorbidities were evaluated with validated questionnaires, and speech perception and hearing ability were measured with validated speech tests. RESULTS: Postoperatively, the mean total NCIQ score (p < 0.0001) and four subscores improved significantly, as did tinnitus distress (p < 0.05). Anxiety symptoms (Generalized Anxiety Disorder questionnaire, GAD-7) decreased significantly after CI. Speech perception improved significantly, particularly "hearing with background noise" (p < 0.05, Oldenburg Inventory, OI) and "localization" (p < 0.001, OI), as well as the Oldenburg Sentence Test (OlSa) scores for with vs. without CI when speech was presented from the SSD side and noise was presented from the normal hearing side (SSSDNNH; p < 0.005). CONCLUSION: CI in SSD patients is a powerful procedure to improve HRQoL, reduce tinnitus distress, and improve psychological comorbidities. Additionally, it is beneficial for hearing improvement, particularly in noise and for directional hearing.

Comparison of performance and metabolism from late pregnancy to early lactation in dairy cows with elevated v. normal body condition at dry-off.

Excessive mobilization of body reserves during the transition from pregnancy to lactation imposes a risk for metabolic diseases on dairy cows. We aimed to establish an experimental model for high v. normal mobilization and herein characterized performance, metabolic and endocrine changes from 7 weeks antepartum (a.p.) to 12 weeks postpartum (p.p.). Fifteen weeks a.p., 38 pregnant multiparous Holstein cows were allocated to two groups that were fed differently to reach either high or normal body condition scores (HBCS: 7.2 NEL MJ/kg dry matter (DM); NBCS: 6.8 NEL MJ/kg DM) at dry-off. Allocation was also based on differences in body condition score (BCS) in the previous and the ongoing lactation that was further promoted by feeding to reach the targeted BCS and back fat thickness (BFT) at dry-off (HBCS: >3.75 and >1.4 cm; NBCS: <3.5 and <1.2 cm). Thereafter, both groups were fed identical diets. Blood samples were drawn weekly from 7 weeks a.p. to 12 weeks p.p. to assess the serum concentrations of metabolites and hormones. The HBCS cows had greater BCS, BFT and BW than the NBCS cows throughout the study and lost more than twice as much BFT during the first 7 weeks p.p. compared with NCBS. Milk yield and composition were not different between groups, except that lactose concentrations were greater in NBSC than in HBCS. Feed intake was also greater in NBCS, and NBCS also reached a positive energy balance earlier than HBCS. The greater reduction in body mass in HBCS was accompanied by greater concentrations of non-esterified fatty acids, and ß-hydroxybutyrate in serum after calving than in NBCS, indicating increased lipomobilization and ketogenesis. The mean concentrations of insulin across all time-points were greater in HBCS than in NBCS. In both groups, insulin and IGF-1 concentrations were lower p.p than in a.p. Greater free thyroxine (fT4) concentrations and a lower free 3-3'-5-triiodothyronine (fT3)/fT4 ratio were observed in HBCS than in NBCS a.p., whereas p.p. fT3/fT4 ratio followed a reverse pattern. The variables indicative for oxidative status had characteristic time courses; group differences were limited to greater plasma ferric reducing ability values in NBSC. The results demonstrate that the combination of pre-selection according to BCS and differential feeding before dry-off to promote the difference was successful in obtaining cows that differ in the intensity of mobilizing body reserves. The HBCS cows were metabolically challenged due to intense mobilization of body fat, associated with reduced early lactation dry matter intake and compromised antioxidative capacity.

Effects of sodium salicylate on glucose kinetics and insulin signaling in postpartum dairy cows.

Low-grade inflammation has been implicated as a contributor to metabolic disease during the transition to lactation. In previous work, administration of sodium salicylate (SS) for 7 d led to hypoglycemia in mature dairy cows in early lactation. The purpose of this study was to identify the mode of action underlying this response to SS. Twenty mature (parity 3) cows were assigned alternately at time of calving to either control or SS treatments; the control received a molasses placebo in drinking water, whereas SS received 2.3 g/L of SS with the molasses carrier in drinking water for 7 d after parturition. Blood samples were collected daily. A glucose turnover assay was performed on d 7, followed by liver, muscle, and adipose tissue biopsies. There were no treatment effects on intake of dry matter or water. Tumor necrosis factor α mRNA abundance tended to be decreased by SS in adipose tissue but not in muscle or liver, and plasma haptoglobin and adiponectin concentrations were not altered by treatment. Treatment did not significantly alter plasma glucose or insulin concentrations, but plasma glucagon concentration tended to be increased by SS and the insulin:glucagon molar ratio was significantly decreased. Cows on SS had a tendency for a 25% decrease in glucose turnover rate compared with control cows. However, there were no differences in transcript abundance of pyruvate carboxylase (PC) or glucose-6-phosphatase (G6PC) in liver or of glucose transporter 4 (GLUT4) in any of the tissues. Finally, SS did not alter insulin receptor substrate-1 phosphorylation in muscle or adipose, but tended to increase phosphorylation of AMP-activated protein kinase and decrease protein kinase B phosphorylation in adipose tissue. These findings may be explained by enhanced hepatic insulin sensitivity leading to posttranscriptional suppression of gluconeogenesis and adaptive responses to decreased glucose supply in the pancreas and adipose tissue.

Tufted angiomas of the head and neck.

BACKGROUND: Tufted angiomas (TAs) are uncommon benign vascular tumors mostly located cutaneously or subcutaneously in the skin of the neck and upper trunk, which appear during childhood and grow slowly. TAs are a variant of lobular capillary hemangiomas. This is the first study to provide an overview of the current literature in combination with the authors' own clinical experience on this rare entity in the head and neck area with non-dermatological localization. METHODS: A selective Medline and Google Scholar database search was performed. Additionally, we conducted an ICD-10-based database search on hemangiomas (D18.0) in the Charité SAP patient records. RESULTS: We identified 13 reports in the literature consisting of 16 cases of TA of the head and neck. Males were predominantly affected by TAs (70.6%). The mean age of the patients at the time of surgery was 31.5 years; the mean maximum diameter of the tumors was 16.3 mm. Additionally, we report on one case of TA in our Department of Head and Neck Surgery. A male patient presented with swelling and redness of the left upper eyelid. Radiological examination with computed tomography showed a suprabulbar tumor with a diameter of 13 mm. The tumor was surgically removed via a transcutaneous blepharoplasty approach with the patient under general anesthesia. DISCUSSION: There is only one other case report on the surgical excision of an intra-orbital TA described to date. Here, we present an epidemiological overview, therapeutic considerations, and differential diagnoses.

[Tufted angiomas of the head and neck. German version]. / Büschelartige Hämangiome ("tufted angiomas") im Kopf-Hals-Bereich.

BACKGROUND: Tufted angiomas (TAs) are uncommon benign vascular tumors mostly located cutaneously or subcutaneously in the skin of the neck and upper trunk, which appear during childhood and grow slowly. TAs are a variant of lobular capillary hemangiomas. This is the first study to provide an overview of the current literature in combination with the authors' own clinical experience on this rare entity in the head and neck area with non-dermatological localization. METHODS: A selective Medline and Google Scholar database search was performed. Additionally, we conducted an ICD-10-based database search on hemangiomas (D18.0) in the Charité SAP patient records. RESULTS: We identified 13 reports in the literature consisting of 16 cases of TA of the head and neck. Males were predominantly affected by TAs (70.6%). The mean age of the patients at the time of surgery was 31.5 years; the mean maximum diameter of the tumors was 16.3 mm. Additionally, we report on one case of TA in our Department of Head and Neck Surgery. A male patient presented with swelling and redness of the left upper eyelid. Radiological examination with computed tomography showed a suprabulbar tumor with a diameter of 13 mm. The tumor was surgically removed via a transcutaneous blepharoplasty approach with the patient under general anesthesia. DISCUSSION: There is only one other case report on the surgical excision of an intra- orbital TA described to date. Here, we present an epidemiological overview, therapeutic considerations, and differential diagnoses.

Effect of increasing body condition on oxidative stress and mitochondrial biogenesis in subcutaneous adipose tissue depot of nonlactating dairy cows.

With the onset of lactation, dairy cows with a body condition score >3.5 are sensitive to oxidative stress and metabolic disorders. Adipose tissue (AT) can adapt to varying metabolic demands and energy requirements by the plasticity of its size during lactation. In AT, angiogenesis is necessary to guarantee sufficient oxygen and nutrient supply for adipocytes. Cellular energy metabolism is reflected mainly by mitochondria, which can be quantified by the mitochondrial DNA copy number per cell. In the present study, we aimed to investigate the effect of overconditioning on angiogenesis and mitochondrial biogenesis in AT of nonlactating cows, irrespective of the physiological influences of lactation and pregnancy. Eight nonpregnant, nonlactating cows received a ration of increasing energy density for 15 wk, during which body weight and body condition increased substantially. Subcutaneous AT was biopsied every 8 wk, and blood was sampled monthly. The blood concentrations of indicators of oxidative stress increased continuously throughout the experimental period, possibly damaging mitochondrial DNA. Concomitantly, HIF-1α, a major marker for hypoxia, increased until wk 8, indicating insufficient angiogenesis in the rapidly expanding AT. Based on the observation that the number of apoptotic cells decreased with increasing hypoxia, the increasing mitochondrial DNA copy numbers might compensate for the hypoxia, reinforcing the production of oxidative stressors. Key transcription factors of mitochondrial biogenesis were largely unaffected. Thus, increased oxidative stress does not impair mitochondrial DNA.

Short Communication: Immunohistochemical localization of the immune cell marker CD68 in bovine adipose tissue: impact of tissue alterations and excessive fat accumulation in dairy cows.

With the onset of lactation energy from feed intake is mostly insufficient to meet the requirements of dairy cows. Lipid mobilization from adipose tissue (AT) could lead to a compromised inflammatory response enhancing the incidence for diseases. In addition, tissue alterations can occur, displaying areas of necrosis and inflammation. Furthermore, over-conditioned cows mobilizing more lipids from AT than thin cows are prone to develop metabolic disorders. This might lead to an increased infiltration of phagocytic immune cells into AT. In the present study, CD68 positive cells were localized in AT from 10 early lactating Holstein cows displaying different grades of AT alterations. Biopsies were sampled from visceral and subcutaneous AT and the number of CD68 positive cells was immunohistochemically determined. In addition, AT biopsies from over-conditioned, non-pregnant, non-lactating cows (n=8) were immunohistochemically analyzed for CD68 positive cells. The percentage of CD68 positive cells was less than 2% in AT biopsies with tissue alterations and in AT from over-conditioned cows. Therefore, immune cell infiltration demonstrated via the localization of CD68 positive cells seems to play only a minor role in AT from over-conditioned cows as well as in different bovine AT depots with tissue alterations.

Short communication: Telomere lengths in different tissues of dairy cows during early and late lactation.

Telomeres create a protective cap on the ends of chromosomes that shorten with cell division and are influenced by stressful conditions. With the onset of lactation, high-yielding dairy cows are exposed to metabolic stress. In the present study, we aimed to analyze telomere length (TL) in key metabolic organs, such as liver, subcutaneous (sc) adipose tissue (AT), and mammary gland, as well as in peripheral blood cells during early and late lactation in German Holstein cows (n=21). Animals were fed according to their requirement, and biopsies from scAT, liver, and mammary gland as well as blood cells were collected in early and late lactation. The relative quantity of telomere products (qT), which is proportional to the average TL, was determined in genomic DNA by multiplex quantitative PCR. In this study, relative qT varied widely in the investigated tissues and blood. In late lactation, slowly proliferating tissues, such as liver and scAT, had the highest qT, whereas peripheral blood cells and in the mammary gland had the lowest qT. Comparing early with late lactation, relative qT attrition was limited to blood and mammary gland. Relationships between relative qT in blood, mammary gland, scAT, and liver suggest that blood qT might serve as a surrogate marker for tissue-specific qT. Cows with high initial qT in tissues and blood in early lactation had greater qT attrition during the course of lactation than cows with lower qT. The determination of qT could be included when phenotyping dairy cattle to test for associations with performance and fitness traits.

Mitochondrial DNA copy number and biogenesis in different tissues of early- and late-lactating dairy cows.

Energy balance in dairy cows changes during the course of lactation due to alterations in voluntary feed intake and energy required for milk synthesis. To adapt to the demands of lactation, energy metabolism needs to be regulated and coordinated in key organs such as adipose tissue (AT), liver, and mammary gland. Mitochondria are the main sites of energy production in mammalian cells and their number varies depending on age, organ, and physiological condition. The copy number of the mitochondrial genome, the mitochondrial DNA (mtDNA), reflects the abundance of mitochondria within a cell and is regulated by transcriptional and translational factors. Environmental, physiological, and energetic conditions change during lactation and we thus hypothesized that these changes may influence the mtDNA copy number and the abundance of genes regulating mitochondrial biogenesis. Therefore, we aimed to provide an overview of mitochondrial biogenesis in liver, subcutaneous (sc)AT, mammary gland, and peripheral blood cells during early and late lactation in dairy cows. German Holstein cows (n=21) were fed according to their requirements, and biopsies from scAT, liver, mammary gland, and blood were collected in early and late lactation and assayed for relative mtDNA copy numbers and the mRNA abundance of genes regulating mitochondrial biogenesis, such as nuclear-respiratory factor 1 and 2 (NRF-1, NRF-2), mitochondrial transcription factor A (TFAM), and peroxisome proliferator-activated receptor-gamma coactivator 1-α (PGC-1α). The number of mtDNA copies increased from early to late lactation in all tissues, whereas that in peripheral blood cells was greater in early compared with late lactation. Moreover, mitochondrial activity enzymes (i.e., citrate synthase and cytochrome c oxidase) increased from early to late lactation in scAT. Comparing the number of mtDNA copies between tissues and blood in dairy cows, the highest mtDNA content was observed in liver. The mRNA abundance of genes related to mitochondrial biogenesis changed in a tissue-specific manner when comparing early versus late lactation. The mtDNA copy number was associated with transcriptional factors only in AT, suggesting nontranscriptional regulation of mtDNA in the other tissues. We detected strong correlations between peripheral blood mtDNA and tissue mtDNA content in early lactation. Peripheral blood forms an appropriate medium to display the cellular content of mtDNA copy numbers and consequently the cellular energy status of tissues during early lactation.

Short communication: localization and expression of monocyte chemoattractant protein-1 in different subcutaneous and visceral adipose tissues of early-lactating dairy cows.

The present study aimed to examine the mRNA abundance of the monocyte chemoattractant protein-1 (MCP-1) and to localize the MCP-1 protein in different subcutaneous (s.c.) and visceral (v.c.) fat depots in high-yielding dairy cows. Early-lactating German Holstein cows (n=25) were divided into a control (CON) and a conjugated linoleic acids (CLA)-supplemented group to investigate potential effects of dietary CLA treatment on MCP-1. The MCP-1 was localized in different s.c. and v.c. adipose tissue (AT) by immunohistochemistry, whereas the mRNA abundance was investigated using quantitative PCR. Albeit the infiltration of immune cells into bovine AT has been demonstrated to be only marginal, both MCP-1 protein and mRNA could be detected in bovine AT depots. The MCP-1 protein was localized both in the cytoplasm of adipocytes and in the cytoplasm of cells from the stromal vascular fraction; however, the number of MCP-1-positive cells was low. The mRNA abundances of MCP-1 were higher in v.c. compared with s.c. AT. Moreover, neither mRNA abundance nor protein expression of MCP-1 was seriously influenced by CLA supplementation of early-lactating dairy cows.

Detection of 11 beta-hydroxysteroid dehydrogenase type 1, the glucocorticoid and mineralocorticoid receptor in various adipose tissue depots of dairy cows supplemented with conjugated linoleic acids.

Early lactating cows mobilize adipose tissue (AT) to provide energy for milk yield and maintenance and are susceptible to metabolic disorders and impaired immune response. Conjugated linoleic acids (CLA), mainly the trans-10, cis-12 isomer, reduce milk fat synthesis and may attenuate negative energy balance. Circulating glucocorticoids (GC) are increased during parturition in dairy cows and mediate differentiating and anti-inflammatory effects via glucocorticoid (GR) and mineralocorticoid receptors (MR) in the presence of the enzyme 11ß-hydroxysteroid dehydrogenase type 1 (11ßHSD1). Activated GC are the main ligands for both receptors in AT; therefore, we hypothesized that tissue-specific GC metabolism is effected by varying amounts of GR, MR and 11ßHSD1 and/or their localization within AT depots. Furthermore, the lipolytic and antilipogenic effects of CLA might influence the GC/GR/MR system in AT. Therefore, we aimed to localize GR and MR as well as the expression pattern and activity of 11ßHSD1 in different AT depots during early lactation in dairy cows and to identify potential effects of CLA. Primiparous German Holstein cows were divided into a control (CON) and a CLA group. From day 1 post-partum (p.p.) until sample collection, the CLA group was fed with 100 g/d CLA (contains 10 g each of the cis-9, trans-11 and the trans-10, cis-12-CLA isomers). CON cows (n = 5 each) were slaughtered on day 1, 42 and 105 p.p., while CLA cows (n = 5 each) were slaughtered on day 42 and 105 p.p. Subcutaneous fat from tailhead, withers and sternum, and visceral fat from omental, mesenteric and retroperitoneal depots were sampled. The localization of GR and 11ßHSD1 in mature adipocytes - being already differentiated - indicates that GC promote other effects via GR than differentiation. Moreover, MR were observed in the stromal vascular cell fraction and positively related to the pre-adipocyte marker Pref-1. However, only marginal CLA effects were observed in this study.

Effect of increasing body condition on key regulators of fat metabolism in subcutaneous adipose tissue depot and circulation of nonlactating dairy cows.

In response to negative energy balance, overconditioned cows mobilize more body fat than thin cows and subsequently are prone to develop metabolic disorders. Changes in adipose tissue (AT) metabolism are barely investigated in overconditioned cows. Therefore, the objective was to investigate the effect of increasing body condition on key regulator proteins of fat metabolism in subcutaneous AT and circulation of dairy cows. Nonlactating, nonpregnant dairy cows (n=8) investigated in the current study served as a model to elucidate the changes in the course of overcondition independent from physiological changes related to gestation, parturition, and lactation. Cows were fed diets with increasing portions of concentrate during the first 6wk of the experiment until 60% were reached, which was maintained for 9wk. Biopsy samples from AT of the subcutaneous tailhead region were collected every 8wk, whereas blood was sampled monthly. Within the experimental period cows had an average BW gain of 243±33.3 kg. Leptin and insulin concentrations were increased until wk 12. Based on serum concentrations of glucose, insulin, and nonesterified fatty acids, the surrogate indices for insulin sensitivity were calculated. High-concentrate feeding led to decreased quantitative insulin sensitivity check index and homeostasis model assessment due to high insulin and glucose concentrations indicating decreased insulin sensitivity. Adiponectin, an adipokine-promoting insulin sensitivity, decreased in subcutaneous AT, but remained unchanged in the circulation. The high-concentrate diet affected key enzymes reflecting AT metabolism such as AMP-activated protein kinase and hormone-sensitive lipase, both represented as the proportion of the phosphorylated protein to total protein, as well as fatty acid synthase. The extent of phosphorylation of AMP-activated protein kinase and the protein expression of fatty acid synthase were inversely regulated throughout the experimental period, whereas the extent of phosphorylation of hormone-sensitive lipase was consistently decreasing by the high-concentrate diet. Overcondition in nonpregnant, nonlactating dairy cows changed the expression of key regulator proteins of AT metabolism and circulation accompanied by impaired insulin sensitivity, which might increase the risk for metabolic disorders.

Murine solid tumours as a novel model to study bacterial biofilm formation in vivo.

Bacteria of many species are able to invade and colonize solid tumours in mice. We have focused on Salmonella enterica serovar Typhimurium. Detailed analysis revealed that such tumour-invading Salmonella form biofilms, thus providing a versatile in vivo test system for studying bacterial phenotypes and host-pathogen interactions. It appears that biofilm formation by S. typhimurium is induced as a defence against the immune system of the host, and in particular against neutrophils. Further, we extended our work to the clinically more relevant biofilm infection by Pseudomonas aeruginosa. The induction of P. aeruginosa biofilms in neoplastic tissue appears to be elicited as a reaction against the immune system. Reconstitution experiments reveal that T cells are responsible for biofilm induction. Isogenic mutants that are no longer able to form biofilms can be used for comparison studies to determine antimicrobial resistance, especially therapeutic efficacy against P. aeruginosa located in biofilms.