Epidemiology and impact of methicillin-sensitive Staphylococcus aureus with ß-lactam antibiotic inoculum effects in adults with cystic fibrosis.

Staphylococcus aureus is the most prevalent cystic fibrosis (CF) pathogen. Several phenotypes are associated with worsened CF clinical outcomes including methicillin-resistance and small-colony-variants. The inoculum effect (IE) is characterized by reduced ß-lactam susceptibility when assessed at high inoculum. The IE associates with worse outcomes in bacteremia and other high-density infections, and may therefore be relevant to CF. The prevalence of IE amongst a CF cohort (age ≥18 years), followed from 2013 to 2016, was investigated. Yearly methicillin-sensitive S. aureus (MSSA) isolates were screened at standard (5 × 105 CFU/mL) and high (5 × 107 CFU/mL) inoculum against narrow-spectrum anti-Staphylococcal ß-lactams and those with anti-pseudomonal activity common to CF. A ≥ 4-fold increase in minimum inhibitory concentration between standard and high inoculum defined IE. Isolates underwent blaZ sequencing and genotyping and were compared against published genomes. Fifty-six percent (99/177) of individuals had MSSA infection. MSSA was observed at ≥105 CFU/mL in 44.8% of entry sputum samples. The prevalence of the IE was 25.0%-cefazolin; 13.5%-cloxacillin; 0%-meropenem; 1.0%-cefepime; 5.2%-ceftazidime; and 34.4%-piperacillin-tazobactam amongst baseline MSSA isolates assessed. blaZ A associated with cefazolin IE (P = 0.0011), whereas blaZ C associated with piperacillin-tazobactam IE (P < 0.0001). Baseline demographics did not reveal specific risk factors for IE-associated infections, nor were long-term outcomes different. Herein, we observed the IE in CF-derived MSSA disproportionally for cefazolin and piperacillin-tazobactam and this phenotype strongly associated with underlying blaZ genotype. The confirmation of CF being a high density infection, and the identification of high prevalence of MSSA with IE in CF supports the need for prospective pulmonary exacerbation treatment studies to understand the impact of this phenotype.

The Caenorhabditis elegans Shugoshin regulates TAC-1 in cilia.

The conserved Shugoshin (SGO) protein family is essential for mediating proper chromosome segregation from yeast to humans but has also been implicated in diverse roles outside of the nucleus. SGO's roles include inhibiting incorrect spindle attachment in the kinetochore, regulating the spindle assembly checkpoint (SAC), and ensuring centriole cohesion in the centrosome, all functions that involve different microtubule scaffolding structures in the cell. In Caenorhabditis elegans, a species with holocentric chromosomes, SGO-1 is not required for cohesin protection or spindle attachment but appears important for licensing meiotic recombination. Here we provide the first functional evidence that in C. elegans, Shugoshin functions in another extranuclear, microtubule-based structure, the primary cilium. We identify the centrosomal and microtubule-regulating transforming acidic coiled-coil protein, TACC/TAC-1, which also localizes to the basal body, as an SGO-1 binding protein. Genetic analyses indicate that TAC-1 activity must be maintained below a threshold at the ciliary base for correct cilia function, and that SGO-1 likely participates in constraining TAC-1 to the basal body by influencing the function of the transition zone 'ciliary gate'. This research expands our understanding of cellular functions of Shugoshin proteins and contributes to the growing examples of overlap between kinetochore, centrosome and cilia proteomes.

Clinical Outcomes Associated With Escherichia coli Infections in Adults With Cystic Fibrosis: A Cohort Study.

BACKGROUND: Analysis of "emerging" pathogens in cystic fibrosis (CF) lung disease has focused on unique pathogens that are rare in other human diseases or are drug resistant. Escherichia coli is recovered in the sputum of up to 25% of patients with CF, yet little is known about the epidemiology or clinical impact of infection. METHODS: We studied patients attending a Canadian adult CF clinic who had positive sputum cultures for E coli from 1978 to 2016. Infection was categorized as transient or persistent (≥3 positive sputum cultures, spanning >6 months). Those with persistent infection were matched 2:1 with age, sex, and time-period controls without history of E coli infection, and mixed-effects models were used to assess pulmonary exacerbation (PEx) frequency, lung function decline, hospitalization, and intravenous antibiotic days. RESULTS: Forty-five patients (12.3%) had E coli recovered from sputum samples between 1978 and 2016, and 18 patients (40%) developed persistent infection. Nine patients (24%) had PEx at incident infection, and increased bioburden was predictive of exacerbation (P = .03). Risk factors for persistent infection included lower nutritional status (P < .001) and lower lung function (P = .009), but chronic infection with Pseudomonas aeruginosa was protective. There was no difference in annual lung function decline, need for hospitalization or intravenous antibiotics, or risk of PEx in patients with persistent infection. CONCLUSIONS: Persistent E coli infection was frequent and was more common in CF patients with low nutritional status and lung function. However, this does not predict clinical decline. Multicenter studies would allow better characterization of the epidemiology and clinical impact of E coli infection.

Decay of Thermal Tolerance in Queensland Fruit Fly Eggs (Bactrocera tryoni, Diptera: Tephritidae) Following Non-Lethal Heat Hardening.

Quarantine disinfestation treatments for Queensland fruit fly (Bactrocera tryoni (Froggatt)) have been developed which use high temperatures to kill preimaginal life stages within fruit prior to export. However, thermal tolerance of individuals can be increased if they are exposed to elevated temperatures before disinfestation treatment. The rate that this thermal conditioning decays after exposure, and the effect of temperature on this decay process, were investigated. Eggs of B. tryoni were exposed to a nonlethal hot water treatment at 38°C for 15 min, 1 or 3 h, then held in air at 25°C for times ranging from 15 min to 12 h, before being exposed to hot water disinfestation at 46°C for various times. From each of these cohorts, the lethal time for 99% mortality (LT99) was calculated. The LT99 of B. tryoni eggs increased with longer conditioning times at 38°C. For each conditioning time, the LT99 decreased with longer delay periods at 25°C prior to disinfestation. The rate of decrease was greatest during the first hour of delay, after which the rate of decrease slowed and tended toward zero. This induction and decay was modeled using a double-exponential equation. These experiments show that thermal conditions prior to disinfestation, and the time delay before the procedure commences, both influence the response of the insect to the disinfestation treatment. These results have implications for the specification of postharvest quarantine treatments, which are usually expressed only in terms of a fruit-center target temperature.

Prevalence and Outcomes of Achromobacter Species Infections in Adults with Cystic Fibrosis: a North American Cohort Study.

Achromobacter species are increasingly being detected in cystic fibrosis (CF) patients, with an unclear epidemiology and impact. We studied a cohort of patients attending a Canadian adult CF clinic who had positive sputum cultures for Achromobacter species in the period from 1984 to 2013. Infection was categorized as transient or persistent (≥50% positive cultures for 1 year). Those with persistent infection were matched 2:1 with age-, sex-, and time-matched controls without a history of Achromobacter infection, and mixed-effects models were used to assess pulmonary exacerbation (PEx) frequency and lung function decline. Isolates from a biobank were retrospectively assessed, identified to the species level by nrdA sequencing, and genotyped using pulsed-field gel electrophoresis (PFGE). Thirty-four patients (11% of those in our clinic), with a median age of 24 years (interquartile range [IQR], 20.3 to 29.8 years), developed Achromobacter infection. Ten patients (29%) developed persistent infection. Persistence did not denote permanence, as most patients ultimately cleared infection, often after years. Patients were more likely to experience PEx at incident isolation than at prior or subsequent visits (odds ratio [OR], 2.7 [95% confidence interval {CI}, 1.2 to 6.7]; P = 0.03). Following persistent infection, there was no difference in annual lung function decline (-1.08% [95% CI, -2.73 to 0.57%] versus -2.74% [95% CI, -4.02 to 1.46%]; P = 0.12) or the odds of PEx (OR, 1.21 [95% CI, 0.45 to 3.28]; P = 0.70). Differential virulence among Achromobacter species was not observed, and no cases of transmission occurred. We demonstrated that incident Achromobacter infection was associated with a greater risk of PEx; however, neither transient nor chronic infection was associated with a worsened long-term prognosis. Large, multicenter studies are needed to clarify the clinical impact, natural history, and transmissibility of Achromobacter.

Reconstruction of non-contained acetabular defects with impaction grafting, a reinforcement mesh and a cemented polyethylene acetabular component.

This review summarises the technique of impaction grafting with mesh augmentation for the treatment of uncontained acetabular defects in revision hip arthroplasty. The ideal acetabular revision should restore bone stock, use a small socket in the near-anatomic position, and provide durable fixation. Impaction bone grafting, which has been in use for over 40 years, offers the ability to achieve these goals in uncontained defects. The precepts of modern, revision impaction grafting are that the segmental or cavitary defects must be supported with a mesh; the contained cavity is filled with vigorously impacted morselised fresh-frozen allograft; and finally, acrylic cement is used to stabilise the graft and provide rigid, long-lasting fixation of the revised acetabular component. Favourable results have been published with this technique. While having its limitations, it is a viable option to address large acetabular defects in revision arthroplasty. Cite this article: Bone Joint J 2017;99-B(1 Supple A):25-30.

A review of fundamental principles for animal models of DOHaD research: an Australian perspective.

Epidemiology formed the basis of 'the Barker hypothesis', the concept of 'developmental programming' and today's discipline of the Developmental Origins of Health and Disease (DOHaD). Animal experimentation provided proof of the underlying concepts, and continues to generate knowledge of underlying mechanisms. Interventions in humans, based on DOHaD principles, will be informed by experiments in animals. As knowledge in this discipline has accumulated, from studies of humans and other animals, the complexity of interactions between genome, environment and epigenetics, has been revealed. The vast nature of programming stimuli and breadth of effects is becoming known. As a result of our accumulating knowledge we now appreciate the impact of many variables that contribute to programmed outcomes. To guide further animal research in this field, the Australia and New Zealand DOHaD society (ANZ DOHaD) Animals Models of DOHaD Research Working Group convened at the 2nd Annual ANZ DOHaD Congress in Melbourne, Australia in April 2015. This review summarizes the contributions of animal research to the understanding of DOHaD, and makes recommendations for the design and conduct of animal experiments to maximize relevance, reproducibility and translation of knowledge into improving health and well-being.

Development and Validation of a PCR Assay To Detect the Prairie Epidemic Strain of Pseudomonas aeruginosa from Patients with Cystic Fibrosis.

The monitoring of epidemic Pseudomonas aeruginosa is important for cystic fibrosis (CF) infection control. The prairie epidemic strain (PES) is common in western Canadian CF clinics. Using whole-genome sequencing, we identified a novel genomic island and developed a PCR assay for PES. Against a collection of 186 P. aeruginosa isolates, the assay had 98% sensitivity and 100% specificity.

Infection control knowledge, beliefs and behaviours amongst cystic fibrosis patients with epidemic Pseudomonas aeruginosa.

BACKGROUND: Epidemic P. aeruginosa (ePA) infections are common in cystic fibrosis (CF) and have been associated with accelerated clinical decline. Factors associated with ePA are unclear, and evidence based infection control interventions are lacking. METHODS: We prospectively collect all bacterial pathogens from adult CF patients. We performed PA strain typing on retrospectively collected enrollment samples and recent isolates to identify patients infected with ePA. All patients attending our clinic were approached to complete a survey on infection control knowledge, beliefs and exposures. We analyzed responses of those with ePA relative to the entire cohort without ePA as well as those infected with unique strains of P. aeruginosa to assess for risk factors for ePA and differences in infection control knowledge, beliefs or behaviours. RESULTS: Of 144 participants, 30 patients had ePA (two Liverpool epidemic strain, 28 Prairie epidemic strain), 83 % of which had established infection prior to transition to the adult clinic. Risk of concomitant infecting pathogens was no different between groups although, Staphylococcus aureus and non-tuberculous mycobacteria were less common in those with ePA. Patients with ePA were more likely to have attended CF-camp and have a history of CF fundraising. Patients with ePA did not differ with respect to beliefs regarding pathogens or transmission risk, except they believed indirect contact posed little risk. Furthermore, patients with ePA were more likely to continue to associate with others with CF despite extensive counselling. Use of peer-peer online networking was minimal in both groups. CONCLUSION: Infections with ePA are closely linked to past exposures, now routinely discouraged. As socialization is the greatest risk factor for ePA, infection control strategies for ePA must focus on discouraging face-to-face interactions amongst CF patients. As peer support remains a desire amongst patients, investment in technologies and strategies that enable indirect communication and support are required.

Effects of omega-3 and omega-6 fatty acids on human placental cytokine production.

INTRODUCTION: Dietary supplementation with omega-3 long chain polyunsaturated fatty acids (n-3 PUFAs) may exert benefits in pregnancy through inhibition of placental inflammation. However, studies on the anti-inflammatory effects of n-3 PUFAs in the placenta are lacking. We compared the cytokine responses of human placental explants in vitro after 4 days pre-incubation with either: a) individual n-3 or n-6 PUFAs (20 µM), or b) physiologically relevant combinations of low, medium or high n-3 or n-6 PUFA concentrations. METHODS: Placental cytokine (IL-6 and TNF-α) mRNA expression and protein production were assessed at 4 h and 12 h, respectively. Cytokine and fatty acid concentrations were also measured in placentas delivered at term by women who ingested either low (n = 12) or high (n = 10) amounts of fish/fish oil in the month prior to delivery. RESULTS: Pre-exposure to n-3 PUFAs as individual fatty acids results in reduced placental IL-6 production (P < 0.05), whereas exposure to complex fatty acid mixtures enriched in n-3 PUFAs (high n-3:n-6 ratios) results in a significant stimulation of IL-6 production (P < 0.05). There were no differences in placental n-3 or n-6 PUFA levels between women with either high or low dietary fish oil intake and no differences in cytokine expression. DISCUSSION: In summary, data from our complex lipid explant model and an observational cohort study do not support a role for n3 PUFAs in the suppression of pro-inflammatory cytokine expression in the human placenta. Results from studies of placental tissues exposed to single n-3 and n-6 PUFAs should be interpreted with considerable caution.

Neurological improvement following reinstitution of a low phenylalanine diet after 20 years in established phenylketonuria.

A 41-year-old woman had meaningful functional improvement following reinstitution of a low phenylalanine diet. She was diagnosed at birth with phenylketonuria and followed strict dietary adherence till the age of 16. Thereafter the diet was discontinued. She subsequently presented with subacute profound visual loss, cognitive dysfunction and paraparesis such that she was bed bound requiring full nursing care. Following dietary intervention there was meaningful improvement such that she was no longer demented and while her vision remains poor she is now independent for activities of daily living. This case report suggests that consideration of reimplementation of dietary intervention is warranted even after a prolonged period of time.

Kiss1 and Kiss1r mRNA expression in the rat placenta: changes with gestational age and regulation by glucocorticoids.

INTRODUCTION: Kisspeptin, the neuropeptide product of the KISS1 gene, is synthesized by neurons within the hypothalamus and is critical for fertility. Human placenta also expresses KISS1 and kisspeptin receptor (KISS1R) mRNA within the trophoblast compartment, where it is thought to act as a physiological invasion inhibitor. METHODS: We determined the expression of Kiss1 mRNA in rat placenta and examined the effect of gestational age and feto-placental growth restriction, achieved through excess maternal glucocorticoid exposure. RESULTS: Dexamethasone induced fetal growth restriction at both day 16 and day 22 of gestation, but placental growth restriction only at day 22. Real-time quantitative RT-PCR revealed an increase in Kiss1 and Kiss1r mRNA from day 16-22 in the labyrinth and junctional zones of the rat placenta. Immunolocalization confirmed kisspeptin expression in the placenta and was prominent in trophoblast tissue. Dexamethasone exposure elevated the expression of Kiss1 mRNA in the labyrinth and junctional zones of day 16 placentas. In contrast, Kiss1 mRNA in the labyrinth zone was reduced following dexamethasone-treatment at day 22. Kiss1r expression was increased in both placental zones at day 16 and 22 in response to dexamethasone-treatment. CONCLUSIONS: We confirm the presence of Kiss1 and Kiss1r mRNA in the rat placenta with expression increasing over the final third of pregnancy, suggestive of a role in restricting placental growth. Furthermore, the effects of dexamethasone on placental Kiss1/Kiss1r suggest glucocorticoid-induced placental growth retardation could be mediated, in part, via early stimulation of Kiss1 and the subsequent inhibition of trophoblast proliferation and invasion.

The inflammatory state of the rat placenta increases in late gestation and is further enhanced by glucocorticoids in the labyrinth zone.

INTRODUCTION: Inflammation plays central roles in key aspects of successful reproduction: ovulation, implantation and parturition. In this study we characterised the inflammatory profile of the rat placenta in late gestation with and without maternal glucocorticoid (dexamethasone) treatment. METHODS: Placentas (n = 6/group) were collected from untreated (Con) rats at days 16 and 22 (term = day 23) and from dexamethasone-treated (Dex) rats at day 22. mRNA and protein expression was determined for enzymes of prostaglandin synthesis and metabolism (Ptgs-1, Ptgs-2, 15-Pgdh), pro-inflammatory cytokines (Tnf-α, Il-1ß, Il-6), and the macrophage marker Emr-1 in the junctional (JZ) and labyrinth (LZ) zones of the placenta. RESULTS: Tnf-α, Il-1ß and Il-6 mRNAs all increased (2- to 4-fold) in both placental zones between days 16 and 22 (P < 0.01). Ptgs-2 mRNA (30-fold; P < 0.01) and PTGS-2 protein (2.4-fold; P < 0.05) similarly increased in LZ. In contrast, 15-Pdgh expression increased in JZ but decreased in LZ; these changes were accompanied by decreased levels of PGE2 in the JZ and a trend towards increased LZ levels. Dex treatment inhibited fetal and placental growth, but had minimal effects on expression of Ptgs-1, Ptgs-2 or 15-Pdgh. Nevertheless, Dex treatment increased LZ PGE2 levels (5-fold, P < 0.01) at the end of gestation. Dex treatment increased Tnf-α mRNA expression in LZ (40%; P < 0.05), but modestly suppressed cytokine protein expression in JZ. CONCLUSIONS: These data demonstrate that the inflammatory state of the LZ increases near term coincident with the known increase in local glucocorticoid levels. This suggests the classic anti-inflammatory actions of glucocorticoids do not occur in the placental LZ.

Are we discussing SUDEP?--A retrospective case note analysis.

PURPOSE: Sudden unexplained death in epilepsy (SUDEP) is uncommon. Discussing the risk of SUDEP can be difficult, particularly in those where the risk is considered low, and previous studies have suggested that clinical practice varies widely. The Scottish Intercollegiate Guidelines Network (SIGN) suggest information on SUDEP is "essential" and National Institute of Clinical Excellence (NICE) recommend that "tailored information on the person's relative risk of SUDEP should be part of the counselling process…". The study aimed to evaluate if discussion of SUDEP risk is being documented in clinical records and to determine if there is an association between documented discussion and risk factors for SUDEP. METHODS: A retrospective case note review was undertaken in those with an established diagnosis of epilepsy attending clinic between 1st January 2009 and 30th June 2009. RESULTS: Overall, a documented SUDEP discussion was noted in 14/345 (4%) cases. Patients were statistically more likely to have a documented SUDEP discussion if they had ongoing generalised tonic-clonic seizures, with a trend also towards informing those non-compliant with medication. CONCLUSION: Patients were more likely to be informed of SUDEP if they had potentially modifiable risk factors identified. There was, however, no documented evidence to suggest that SUDEP is being discussed in the majority of cases.

A rhythmic placenta? Circadian variation, clock genes and placental function.

Physiological rhythms entrained by the circadian clock are present in virtually all organs including those of the reproductive system. In mammals, circadian timing is driven by a 'master clock' in the suprachiasmatic nucleus that influences peripheral tissue clocks via endocrine, autonomic and behavioral cues. The molecular clock machinery comprises a network of 'clock' genes, namely Clock, Bmal1, Per1, Per2, Per3, Cry1 and Cry2. These clock genes generate endogenous oscillations that drive rhythmic expression of downstream genes and thus physiological and behavioral processes. Importantly, disturbances in clock gene expression are implicated in a range of pathologies including cancer and obesity. The recent recognition that clock genes are expressed in the placenta, together with observations linking circadian disruption with compromised placental function, suggests that circadian variation may be an important component of the normal placental phenotype. In this review we consider this possibility in the context of maternal circadian physiology in pregnancy. While there is good evidence for rhythmic expression of several genes in the rodent placenta, the conventional transcriptional-translational feedback loops of the clock machinery appear less robust and coordinated. Further study is needed to elucidate the function of the placental clock genes across gestation and among different species, particularly those in which greater circadian development occurs in utero. Such studies will likely provide important insights into placental physiology and pathology.

Oxysterols inhibit differentiation and fusion of term primary trophoblasts by activating liver X receptors.

Oxygenated cholesterol metabolites known as oxysterols display potent biological activities ranging from regulation of lipid homeostasis to cytotoxicity. Oxysterols have previously been shown to inhibit the invasion of first trimester trophoblasts, an effect which involves activation of the nuclear liver X receptors (LXRs). In the present study, we investigated the effects of several oxysterols on syncytialisation (differentiation and fusion) in term placental trophoblasts. Treatment of cultured term primary trophoblast cells with oxysterols [25-hydroxycholesterol, 7-ketocholesterol, 22(R)-hydroxycholesterol] and the synthetic LXR agonist T0901317 at non-toxic doses decreased expression of GCM-1 and HERV-W mRNA and reduced hCG secretion and placental alkaline phosphatase activity, indicative of diminished trophoblast differentiation. Furthermore, treatment with these compounds also decreased cell fusion measured by E-cadherin immunostaining and quantification of syncytialised nuclei. Treatment with an LXR antagonist (geranylgeranyl diphosphate) abrogated the inhibitory effects of oxysterols and T0901317 on trophoblast syncytialisation indicating that these effects are mediated by LXR. These findings suggest that oxysterols impair differentiation and fusion of term trophoblast cells via an LXR-dependent mechanism.

IFPA Meeting 2009 workshops report.

Workshops are an important part of the annual meeting of the International Federation of Placenta Associations (IFPA). At IFPA Meeting 2009 diverse topics were discussed in twelve themed workshops. Topics covered included: immune response to pregnancy; signaling between fetus and placenta; bioactive lipids in placenta; placenta in agricultural species; epigenetics and placentation; trophoblast deportation; glucocorticoids and placental function; endothelium; placental transport; genes and placenta; uteroplacental blood flow and placental stem cells. This report is a full summary of the various topics covered.

Placental and fetal growth retardation following partial progesterone withdrawal in rat pregnancy.

This study investigated placental expression of the two main isoforms of the progesterone receptor and the regulation of placental and fetal growth by progesterone over the final third of rat pregnancy, the period of maximal fetal growth. Expression patterns of mRNAs encoding the two major progesterone receptor isoforms (PR-A and PR-B) were measured by real-time RT-PCR in the two morphologically- and functionally-distinct regions of the placenta, the basal and labyrinth zones, at days 16 and 22 of pregnancy (term=day 23). PR-A and PR-B mRNA expression was extremely low in labyrinth zone on days 16 and 22, close to the limits of detection. In contrast, the basal zone exhibited much higher levels of mRNA expression for both PR-A (>10-fold higher than in labyrinth zone) and PR-B (3-fold higher at day 16). To assess the role of progesterone in placental growth, maternal progesterone was reduced from day 16 by ovariectomy with full estradiol replacement and partial progesterone replacement until day 22. Progesterone reduction lowered fetal (10%), whole placental (24%), basal zone (37%) and labyrinth zone (14%) weights at day 22 compared with sham-controls, whereas fetal and placental weights (both zones) were maintained in ovariectomised rats given full estradiol/progesterone replacement. The effects of progesterone withdrawal were not associated with changes in placental expression of either IGF-II or IGFBP-2, both important players in growth factor regulation of placental growth. Importantly, however, IGF-II expression remained elevated in the labyrinth zone but fell markedly in basal zone ( approximately 7-fold) between days 16 and 22 of normal pregnancy, consistent with the growth patterns of these two placental regions over this period.