Evaluating a Measles and Rubella Multiplex Bead Assay for Countries in the WHO Global Measles and Rubella Laboratory Network

INTRODUCTION: Measles and rubella are highly contagious viral diseases. Measles remains one of the leading causes of vaccine-preventable deaths, and rubella during pregnancy can cause congenital rubella syndrome. The enzyme immunoassay (EIA) is most frequently used to determine antibody responses to measles and rubella, and the plaque reduction neutralization assay (PRN) is considered the gold standard for determining measles immunity. A measles and rubella multiplex bead assay (MBA) is currently being evaluated at the Centers for Disease Control and Prevention for its utility in serosurveillance within the World Health Organization Global Measles and Rubella Laboratory Network, and the MBA has several benefits over these assays. The MBA requires a smaller sample volume, measures several diseases simultaneously, is faster, and has less technician-dependency than the PRN. AIM: Study objectives are to compare accuracy of measles serological status from both the MBA and EIA when compared with the PRN, and to evaluate whether country or participant age are associated with test accuracy. METHODS: Samples for participants of varying ages from the United States, Tajikistan, and Bangladesh (n=300) that had been tested by each assay were used. Results were dichotomized as positive or negative according to respective assay cutoff values. Logistic regression models were applied to estimate point estimates and confidence intervals for sensitivity and specificity of the MBA and EIA with relation to the PRN gold standard. RESULTS: Across the three age groups and countries, EIA has higher median values (mIU/ml) compared with MBA and PRN (582.55, 399.28, and 378.95, respectively, for Tajikistan participants aged 13 and older). McNemar’s Test of Agreement comparing MBA and PRN suggests disagreement (p DISCUSSION: When compared with the PRN, the MBA has similar accuracy to the EIA with regards to sensitivity and specificity, and in several groupings of country and age, these percentages are higher for the MBA. Many of the benefits of the MBA over conventional assays have led to its increased application in research, and these results suggest the MBA can be used in settings where the EIA is currently used

A SUSTAINABLE APPROACH TO THE CONTROL OF PATHOGENS: THE FATE OF STREPTOCOCCI IN EQUINE COMPOST.

Streptococcus equi subspecies equi (S. equi), causes the potentially fatal respiratory disease called “strangles” in horses, while the closely related Streptococcus equi subspecies zooepidemicus (S. zooepidemicus) causes potentially fatal infections in humans. A study was undertaken to determine the survival of these two organisms in equine compost. Compost piles of equine bedding and feed waste were inoculated with 10 x 1010 c.f.u. of S. zooepidemicus and samples taken at 48, 96, 168 and 336 hours relative to samples placed in the pile at 0 hours. No Streptococci were isolated at 48 hours or subsequent time-points. Next, S. equi was similarly inoculated into equine compost, with samples taken at 2, 4, 8, 12, 24, 48, 168 and 336 hours later. No Streptococci were isolated at any time-point. To rule out killing of S. equi by microflora in equine waste, samples of soiled bedding, both autoclaved and un-autoclaved (with water added to match autoclaved moisture) were inoculated with 10 x 1010 c.f.u. of S. zooepidemicus and sampled at 0, 6, 12, 24, 48, 72, 120, 168 and 264 hours. In autoclaved bedding, S. zooepidemicus was isolated from 0 – 120 hours, but replaced by other flora at 264 hours. In un-autoclaved samples, Streptococci were not present after 48 hours. A repeated trial with S. equi yielded similar results. This data suggest that microbial activity of equine waste bedding may eliminate streptococci within 24 - 48 hours, indicating that normal microflora may provide sustainable methods for the control of human and animal pathogens

A SUSTAINABLE APPROACH TO THE CONTROL OF PATHOGENS: THE FATE OF STREPTOCOCCI IN EQUINE COMPOST.

Streptococcus equi subspecies equi (S. equi), causes the potentially fatal respiratory disease called “strangles” in horses, while the closely related Streptococcus equi subspecies zooepidemicus (S. zooepidemicus) causes potentially fatal infections in humans. A study was undertaken to determine the survival of these two organisms in equine compost. Compost piles of equine bedding and feed waste were inoculated with 10 x 1010 c.f.u. of S. zooepidemicus and samples taken at 48, 96, 168 and 336 hours relative to samples placed in the pile at 0 hours. No Streptococci were isolated at 48 hours or subsequent time-points. Next, S. equi was similarly inoculated into equine compost, with samples taken at 2, 4, 8, 12, 24, 48, 168 and 336 hours later. No Streptococci were isolated at any time-point. To rule out killing of S. equi by microflora in equine waste, samples of soiled bedding, both autoclaved and un-autoclaved (with water added to match autoclaved moisture) were inoculated with 10 x 1010 c.f.u. of S. zooepidemicus and sampled at 0, 6, 12, 24, 48, 72, 120, 168 and 264 hours. In autoclaved bedding, S. zooepidemicus was isolated from 0 – 120 hours, but replaced by other flora at 264 hours. In un-autoclaved samples, Streptococci were not present after 48 hours. A repeated trial with S. equi yielded similar results. This data suggest that microbial activity of equine waste bedding may eliminate streptococci within 24 - 48 hours, indicating that normal microflora may provide sustainable methods for the control of human and animal pathogens

Thromboembolism After Intramedullary Nailing for Metastatic Bone Lesions.

BACKGROUND: The risk of venous thromboembolism (VTE) in patients undergoing intramedullary nailing for skeletal metastatic disease is currently undefined. The purpose of our study was to determine the risk of thromboembolic events, to define the risk factors for VTE, and to define the rate of wound complications in this population. METHODS: A retrospective review of surgical databases at three National Cancer Institute (NCI)-designated cancer centers identified 287 patients with a total of 336 impending or pathologic long-bone fractures that were stabilized with intramedullary nailing between February 2001 and April 2013. Statistical analysis was performed utilizing multivariable logistic regression and Fisher exact tests. RESULTS: The overall rate of VTE was twenty-four (7.1%) of the 336; thirteen (3.9%) were pulmonary embolism (PE), and eleven (3.3%), deep venous thrombosis (DVT). In two patients, adequate anticoagulation data were not available. We found no significant relationship between the type of anticoagulant used and VTE. There was a significant positive correlation found between lung-cancer histology and the development of VTE (p \u3c 0.001) or PE (p \u3c 0.001). The absence of radiation therapy approached significance (p = 0.06) with respect to decreased overall VTE risk. Wound complications were documented for 11 (3.3%) of the operations. CONCLUSIONS: There is a high rate of VTE among those with skeletal metastatic disease who undergo intramedullary nailing, even while receiving postoperative thromboembolic prophylaxis. Current anticoagulation protocols may be inadequate. Wound-complication risk with anticoagulant use in this population is low and should not be a deterrent to adequate anticoagulant use for this population

Progressive tauopathy disrupts breathing stability and chemoreflexes during presumptive sleep in mice

Rationale: Although sleep apnea occurs in over 50% of individuals with Alzheimer’s Disease (AD) or related tauopathies, little is known concerning the potential role of tauopathy in the pathogenesis of sleep apnea. Here, we tested the hypotheses that, during presumptive sleep, a murine model of tauopathy (rTg4510) exhibits: 1) increased breathing instability; 2) impaired chemoreflex function; and 3) exacerbation of these effects with tauopathy progression.Methods: rTg4510 mice initially develop robust tauopathy in the hippocampus and cortex, and eventually progresses to the brainstem. Type I and II post-sigh apnea, Type III (spontaneous) apnea, sigh, and hypopnea incidence were measured in young adult (5–6 months; n = 10–14/group) and aged (13–15 months; n = 22–24/group) non-transgenic (nTg), monogenic control tetracycline transactivator, and bigenic rTg4510 mice using whole-body plethysmography during presumptive sleep (i.e., eyes closed, curled/laying posture, stable breathing for >200 breaths) while breathing room air (21% O2). Peripheral and central chemoreceptor sensitivity were assessed with transient exposures (5 min) to hyperoxia (100% O2) or hypercapnia (3% and 5% CO2 in 21% O2), respectively.Results: We report significant increases in Type I, II, and III apneas (all p < 0.001), sighs (p = 0.002) and hypopneas (p < 0.001) in aged rTg4510 mice, but only Type III apneas in young adult rTg4510 mice (p < 0.001) versus age-matched nTg controls. Aged rTg4510 mice exhibited profound chemoreflex impairment versus age matched nTg and tTA mice. In rTg4510 mice, breathing frequency, tidal volume and minute ventilation were not affected by hyperoxic or hypercapnic challenges, in striking contrast to controls. Histological examination revealed hyperphosphorylated tau in brainstem regions involved in the control of breathing (e.g., pons, medullary respiratory column, retrotrapezoid nucleus) in aged rTg4510 mice. Neither breathing instability nor hyperphosphorylated tau in brainstem tissues were observed in young adult rTg4510 mice.Conclusion: Older rTg4510 mice exhibit profound impairment in the neural control of breathing, with greater breathing instability and near absence of oxygen and carbon-dioxide chemoreflexes. Breathing impairments paralleled tauopathy progression into brainstem regions that control breathing. These findings are consistent with the idea that tauopathy per se undermines chemoreflexes and promotes breathing instability during sleep

APOE4, Age, and Sex Regulate Respiratory Plasticity Elicited by Acute Intermittent Hypercapnic-Hypoxia

Rational Acute intermittent hypoxia (AIH) shows promise for enhancing motor recovery in chronic spinal cord injuries and neurodegenerative diseases. However, human trials of AIH have reported significant variability in individual responses. Objectives Identify individual factors (eg, genetics, age, and sex) that determine response magnitude of healthy adults to an optimized AIH protocol, acute intermittent hypercapnic-hypoxia (AIHH). Methods In 17 healthy individuals (age = 27 ± 5 yr), associations between individual factors and changes in the magnitude of AIHH (15, 1-min O2 = 9.5%, CO2 = 5% episodes) induced changes in diaphragm motor-evoked potential (MEP) amplitude and inspiratory mouth occlusion pressures (P0.1) were evaluated. Single nucleotide polymorphisms (SNPs) in genes linked with mechanisms of AIH induced phrenic motor plasticity (BDNF, HTR2A, TPH2, MAOA, NTRK2) and neuronal plasticity (apolipoprotein E, APOE) were tested. Variations in AIHH induced plasticity with age and sex were also analyzed. Additional experiments in humanized (h)ApoE knock-in rats were performed to test causality. Results AIHH-induced changes in diaphragm MEP amplitudes were lower in individuals heterozygous for APOE4 (i.e., APOE3/4) compared to individuals with other APOE genotypes (P = 0.048) and the other tested SNPs. Males exhibited a greater diaphragm MEP enhancement versus females, regardless of age (P = 0.004). Additionally, age was inversely related with change in P0.1 (P = 0.007). In hApoE4 knock-in rats, AIHH-induced phrenic motor plasticity was significantly lower than hApoE3 controls (P \u3c 0.05). Conclusions APOE4 genotype, sex, and age are important biological determinants of AIHH-induced respiratory motor plasticity in healthy adults. Addition To Knowledge Base AIH is a novel rehabilitation strategy to induce functional recovery of respiratory and non-respiratory motor systems in people with chronic spinal cord injury and/or neurodegenerative disease. Figure 5 Since most AIH trials report considerable inter-individual variability in AIH outcomes, we investigated factors that potentially undermine the response to an optimized AIH protocol, AIHH, in healthy humans. We demonstrate that genetics (particularly the lipid transporter, APOE), age and sex are important biological determinants of AIHH-induced respiratory motor plasticit

Vancomycin-induced gut microbial dysbiosis alters enteric neuron-macrophage interactions during a critical period of postnatal development

Vancomycin is a broad-spectrum antibiotic widely used in cases of suspected sepsis in premature neonates. While appropriate and potentially lifesaving in this setting, early-life antibiotic exposure alters the developing microbiome and is associated with an increased risk of deadly complications, including late-onset sepsis (LOS) and necrotizing enterocolitis (NEC). Recent studies show that neonatal vancomycin treatment disrupts postnatal enteric nervous system (ENS) development in mouse pups, which is in part dependent upon neuroimmune interactions. This suggests that early-life antibiotic exposure could disrupt these interactions in the neonatal gut. Notably, a subset of tissue-resident intestinal macrophages, muscularis macrophages, has been identified as important contributors to the development of postnatal ENS. We hypothesized that vancomycin-induced neonatal dysbiosis impacts postnatal ENS development through its effects on macrophages. Using a mouse model, we found that exposure to vancomycin in the first 10 days of life, but not in adult mice, resulted in an expansion of pro-inflammatory colonic macrophages by increasing the recruitment of bone-marrow-derived macrophages. Single-cell RNA sequencing of neonatal colonic macrophages revealed that early-life vancomycin exposure was associated with an increase in immature and inflammatory macrophages, consistent with an influx of circulating monocytes differentiating into macrophages. Lineage tracing confirmed that vancomycin significantly increased the non-yolk-sac-derived macrophage population. Consistent with these results, early-life vancomycin exposure did not expand the colonic macrophage population nor decrease enteric neuron density in CCR2-deficient mice. Collectively, these findings demonstrate that early-life vancomycin exposure alters macrophage number and phenotypes in distinct ways compared with vancomycin exposure in adult mice and results in altered ENS development

Omics-driven identification and elimination of valerolactam catabolism in Pseudomonas putida KT2440 for increased product titer.

Pseudomonas putida is a promising bacterial chassis for metabolic engineering given its ability to metabolize a wide array of carbon sources, especially aromatic compounds derived from lignin. However, this omnivorous metabolism can also be a hindrance when it can naturally metabolize products produced from engineered pathways. Herein we show that P. putida is able to use valerolactam as a sole carbon source, as well as degrade caprolactam. Lactams represent important nylon precursors, and are produced in quantities exceeding one million tons per year (Zhang et al., 2017). To better understand this metabolism we use a combination of Random Barcode Transposon Sequencing (RB-TnSeq) and shotgun proteomics to identify the oplBA locus as the likely responsible amide hydrolase that initiates valerolactam catabolism. Deletion of the oplBA genes prevented P. putida from growing on valerolactam, prevented the degradation of valerolactam in rich media, and dramatically reduced caprolactam degradation under the same conditions. Deletion of oplBA, as well as pathways that compete for precursors L-lysine or 5-aminovalerate, increased the titer of valerolactam from undetectable after 48 h of production to ~90 mg/L. This work may serve as a template to rapidly eliminate undesirable metabolism in non-model hosts in future metabolic engineering efforts