Integrating central nervous system metagenomics and host response for diagnosis of tuberculosis meningitis and its mimics.

The epidemiology of infectious causes of meningitis in sub-Saharan Africa is not well understood, and a common cause of meningitis in this region, Mycobacterium tuberculosis (TB), is notoriously hard to diagnose. Here we show that integrating cerebrospinal fluid (CSF) metagenomic next-generation sequencing (mNGS) with a host gene expression-based machine learning classifier (MLC) enhances diagnostic accuracy for TB meningitis (TBM) and its mimics. 368 HIV-infected Ugandan adults with subacute meningitis were prospectively enrolled. Total RNA and DNA CSF mNGS libraries were sequenced to identify meningitis pathogens. In parallel, a CSF host transcriptomic MLC to distinguish between TBM and other infections was trained and then evaluated in a blinded fashion on an independent dataset. mNGS identifies an array of infectious TBM mimics (and co-infections), including emerging, treatable, and vaccine-preventable pathogens including Wesselsbron virus, Toxoplasma gondii, Streptococcus pneumoniae, Nocardia brasiliensis, measles virus and cytomegalovirus. By leveraging the specificity of mNGS and the sensitivity of an MLC created from CSF host transcriptomes, the combined assay has high sensitivity (88.9%) and specificity (86.7%) for the detection of TBM and its many mimics. Furthermore, we achieve comparable combined assay performance at sequencing depths more amenable to performing diagnostic mNGS in low resource settings.

Prognostic implications of baseline anaemia and changes in haemoglobin concentrations with amphotericin B therapy for cryptococcal meningitis.

OBJECTIVES: Anaemia represents a common toxicity with amphotericin B-based induction therapy in HIV-infected persons with cryptococcal meningitis. We sought to examine the impact of amphotericin-related anaemia on survival. METHODS: We used data from Ugandan and South African trial participants to characterize the variation of haemoglobin concentrations from diagnosis to 12 weeks post-diagnosis. Anaemia severity was classified based on the haemoglobin concentration at cryptococcal meningitis diagnosis, and nadir haemoglobin values during amphotericin induction. Cox proportional hazard models were used to estimate 2- and 10-week mortality risk. We also estimated 10-week mortality risk among participants with nadir haemoglobin < 8.5 g/dL during amphotericin induction and who survived ≥ 2 weeks post-enrolment. RESULTS: The median haemoglobin concentration at meningitis diagnosis was 11.5 g/dL [interquartile range (IQR) 9.7-13 g/dL; n = 311] with a mean decline of 4.2 g/dL [95% confidence interval (CI) -4.6 to -3.8; P < 0.001; n = 148] from diagnosis to nadir value among participants with baseline haemoglobin ≥ 8.5 g/dL. The median haemoglobin concentration was 8.1 g/dL (IQR 6.5-9.5 g/dL) at 2 weeks, increasing to 9.4 g/dL (IQR 8.2-10.9 g/dL) by 4 weeks and continuing to increase to 12 weeks. Among participants with haemoglobin < 8.5 g/dL at diagnosis, mortality risk was elevated at 2 weeks [hazard ratio (HR) 2.7; 95% CI 1.5-4.9; P < 0.01] and 10 weeks (HR 1.8; 95% CI 1.1-2.2; P = 0.03), relative to those with haemoglobin ≥ 8.5 g/dL. New-onset anaemia occurring with amphotericin therapy did not have a statistically significant association with 10-week mortality (HR 2.0; 95% CI 0.5-9.1; P = 0.4). CONCLUSIONS: Amphotericin induced significant haemoglobin declines, which were mostly transient and did not impact 10-week mortality. Individuals with moderate to life-threatening anaemia at baseline had a higher mortality risk at 2 and 10 weeks post-enrolment.

Improved diagnostic sensitivity for tuberculous meningitis with Xpert(®) MTB/RIF of centrifuged CSF.

BACKGROUND: TB meningitis (TBM) diagnosis is difficult and novel diagnostic methods are needed. The World Health Organization recommends Xpert(®) MTB/RIF as the initial TBM diagnostic test based on two studies reporting suboptimal sensitivity (~50-60%). OBJECTIVE: To study the effect of cerebrospinal fluid (CSF) centrifugation on Xpert performance for TBM detection. DESIGN: A total of 107 predominantly human immunodeficiency virus (HIV) infected adults with presumed meningitis were screened prospectively in Kampala, Uganda. CSF was tested using 1) microscopy for acid-fast bacilli; 2) MGIT™ culture; 3) Xpert of 2 ml of unprocessed CSF; and 4) Xpert of centrifuged CSF. Diagnostic performance was measured against an a priori composite reference standard of any positive CSF tuberculosis test. RESULTS: Of 107 participants, 18 (17%) had definite TBM. When CSF was centrifuged, Xpert had better sensitivity (13/18, 72%) than when using 2 ml of unprocessed CSF (5/18, 28%; P = 0.008). The median centrifuged CSF volume was 6 ml (IQR 4-10). Mycobacterial culture yielded 71% (12/17) sensitivity at a median delay of 27 days. Only 39% were positive by both culture and centrifuged Xpert, with additional cases detected by Xpert and culture. CONCLUSIONS: CSF centrifugation optimizes the diagnostic performance of Xpert in the detection of TBM. A combination of culture and Xpert detected the largest number of cases.

Influence of plasma pretreatment on shear bond strength of self-adhesive resin cements to polyetheretherketone.

OBJECTIVES: The aim of this study is to evaluate the adhesion between PEEK and two self-adhesive resin cements after plasma treatment. METHODS: Eight hundred sixty-four polyetheretherketone (PEEK) disks were cut and polished to silicon carbide (SIC) P4000. One half of the specimens were randomly selected and pretreated with plasma, whereas the remaining 432 specimens remained untreated. Subsequently, specimens were randomly allocated to four groups (n = 108/group): Visio.link (Bredent), Signum PEEK Bond (Heraeus Kulzer), Ambarino P60 (Creamed), and a control group without additional treatment. Half of the specimens of each group (n = 54) were then cemented with either RelyX Unicem Automix 2 (3 M ESPE) or with Clearfil SA (Kuraray). All specimens were stored in water for 24 h (37 °C). Afterwards, specimens were divided into three groups (n = 18) for different aging levels: (1) no aging (baseline measurement), (2) thermal aging for 5,000 cycles (5/55 °C), and (3) thermal aging for 10,000 cycles (5/55 °C). Thereafter, shear bond strengths (SBS) were measured, and failure types (adhesive, mixed, and cohesive) were assessed. Data were analyzed using descriptive statistics, four- and one-way ANOVA followed by a post hoc Scheffé test (p < 0.05). RESULTS: No adhesion could be established without adhesive pretreatment, irrespectively, whether plasma was applied or not. Also, no bond strength was measured when Ambarino P60 was applied. In contrast, adhesive pretreatment resulted in SBS ranging between 8 and 15 MPa. No significant differences were found between the resin cements used. In general, no cohesive failures were observed. Groups without plasma treatment combined with Visio.link or Signum PEEK Bond showed predominantly mixed failure types. Control groups, plasma treated, or treated using Ambarino P60 groups fractured predominantly adhesively. CONCLUSION: The use of methyl methacrylate (MMA)-based adhesives allows bonding between PEEK and self-adhesive resin cements. Plasma treatment has no impact on bond to resin cements. CLINICAL SIGNIFICANCE: PEEK reconstructions can be cemented using self-adhesive resin cements combined with pretreatment with MMA-based adhesives.

Amphetamine withdrawal does not produce a depressive-like state in rats as measured by three behavioral tests.

Administration of amphetamine (AMPH) can induce symptoms of psychosis in humans and locomotor sensitization in rats; in contrast, withdrawal from a period of AMPH intake is most often associated with symptoms of human endogenous depression. The aim of this study was to determine whether AMPH withdrawal produces a depressive-like state in rats. The present study examined the effects of withdrawal from an escalating-dose AMPH schedule (ESC; three daily injections over 6 days, 1-5 mg/kg, i.p.) and an intermittent-dose AMPH schedule (INT; one daily injection over 6 days, 1.5 mg/kg, i.p.) on animals' performance in three behavioral paradigms related to depression: the Porsolt swim test, the learned helplessness assay and operant responding for sucrose on a progressive ratio schedule. ESC and INT AMPH withdrawal had no effect on any of these tests or on stress responsiveness as measured by increased plasma levels of corticosterone (CORT) and adrenocorticotropin following the swim test, although basal CORT levels were higher in AMPH-withdrawn animals compared to controls. Finally, we confirmed the presence of locomotor sensitization for both AMPH schedules after 30 days of withdrawal. Our results suggest that the ability of AMPH withdrawal to produce symptoms of depression may not be evident in all behavioral screens for depressive symptoms in the rat.

Peripartum sex steroid profiles and endocrine correlates of postpartum maternal behavior in captive gorillas (Gorilla gorilla gorilla).

An association between pregnancy levels of estrogen and progesterone and maternal behavior has been demonstrated in several taxonomic orders of nonprimate and primate mammals, but has not so far been investigated in the gorilla. In this study we investigated whether prepartum titers of urinary estrone conjugates (E1C) or pregnanediol-3alpha-glucuronide (PdG) were related to postpartum maternal behavior in eight multiparous Western lowland gorilla females (Gorilla gorilla gorilla) housed in four zoological gardens. Urine samples were collected from each study animal for 14 days prepartum and 14 days postpartum, and measures of maternal responsiveness were scored during the first 15 days postpartum. Urine samples were assayed with radioimmunoassay for E1C and PdG. Results for the peripartum profiles of urinary E1C, as well as postpartum profiles of PdG, agree with previous findings for the gorilla, while results for late-pregnancy profiles of urinary PdG were inconclusive in confirming a prepartum increase or decrease. Neither prepartum levels of E1C or PdG, nor the E1C/PdG ratio were found to be related to measures of postpartum maternal behavior. This lack of association between late-pregnancy E1C titers per se and postpartum maternal behavior is contrary to findings in nonprimate and other primate species.

Comparison of the effects of infant handling, isolation, and nonhandling on acoustic startle, prepulse inhibition, locomotion, and HPA activity in the adult rat.

This study examined whether early isolation (EI), early handling (EH), or early nonhandling (NH) in infant rats alters (a) prepulse inhibition (PPI) of the acoustic startle response (ASR) or its disruption by apomorphine, (b) motor activity or its stimulation by amphetamine, or (c) corticosterone activity (because of its modulation of dopamine activity), in adulthood and in comparison with a normal-husbandry postnatal control environment. EI did not affect PPI, reduced PPI disruption by apomorphine in males, and increased amphetamine-stimulated activity in males. NH increased the ASR, reduced activity in the open field, and increased corticosterone reactivity in males. In all paradigms, the effects of EH were similar to those of the control environment. This study provides an important contribution to the evidence on the relationship between postnatal experience and long-term neurobehavioral development in the rat and the relevance of this approach to animal models of neuropsychiatric disorder.

Comparative aspects of the metabolism and excretion of cortisol in three individual nonhuman primates.

A radiometabolism study is described to provide the first comparative data on the time course, route, and characteristics of excreted [3H]cortisol metabolites in three nonhuman primates: the common marmoset (Callithrix jacchus), the long-tailed macaque (Macacafascicularis), and the chimpanzee (Pan troglodytes). A low dose (40-100 microCi) of 3H-labeled cortisol was administered intravenously to one adult male of each species and the excreta collected over a 5-day period postinjection. The major proportion of radioactivity was excreted in the urine (>80%). Peak radioactivity in urine was recovered within 5.5 h following injection in all three species, while in the feces peak levels of radioactivity were recovered within 26 h postinjection. In all three species, urinary metabolites were primarily excreted as conjugates (61-87%), whereas the percentage of conjugated metabolites in feces was 50% or less. The number and relative abundance of urinary and fecal [3H]cortisol metabolites were determined by reverse-phase high-performance liquid chromatography (HPLC) and immunoreactivity of the radioactivity peaks was assessed by screening HPLC fractions with established cortisol, corticosterone, and 11-oxoetiocholanolone enzyme immunoassays (EIA), the latter being a group-specific assay for measuring 11,17-dioxoandrostanes. HPLC separation of urinary and fecal extracts revealed multiple peaks of radioactivity, several of which were common to all three species. The relative proportion of these peaks, however, differed considerably among species and between urine and feces. HPLC indicated that native cortisol was a major urinary excretory product in the marmoset, while comparatively small amounts were present in the urine of the macaque and chimpanzee. In contrast, in feces, cortisol was only detected in low amounts in the marmoset and was virtually absent in the macaque and chimpanzee. In all three species, one of the major radioactivity peaks showed a retention time comparable to 11-oxoetiocholanolone and high immunoreactivity in the 11-oxoetiocholanolone EIA. The measurement of urinary- and/or fecal-immunoreactive 11,17-dioxoandrostanes is therefore implicated for noninvasive assessment of adrenal function in Old World monkeys, New World monkeys, and great apes.

Evidence from urinary cortisol that maternal behavior is related to stress in gorillas.

By studying western lowland gorillas (Gorilla gorilla gorilla, n = 8) in zoological gardens via ethological and non-invasive physiological techniques, we have demonstrated that their postpartum maternal behavior is related negatively to their postpartum urinary titers of cortisol. On the basis of this finding, it is proposed that postpartum stress contributes to disrupted maternal behavior in the gorilla in captivity. Morning urine samples were collected with a mean sampling interval of 1.6 days from Day 14 prepartum to Day 14 postpartum (n = 11 pregnancies). Creatinine-indexed (Cr) urinary cortisol titers declined significantly between Day 9 to 1 prepartum (0.634 +/- 0.014 microg/mg of Cr, mean +/- SEM) and Day 1 to 6 postpartum (0.396 +/- 0.030 microg/mg of Cr, mean +/- SEM; p < 0.01-0.001). For each pregnancy, the relative postpartum decline in urinary cortisol was calculated as (microg of cortisol/mg of Cr Day 1 to 4)/(microg of cortisol/mg of Cr Day -4 to -1). Values ranged from 0.35 to 1.12, were independent of absolute prepartum cortisol titers, and were interpreted as evidence of inter-female differences in postpartum hypothalamo-pituitary-adrenal axis activity and, therefore, postpartum stress. This postpartum stress index was negatively correlated with the amount of time (0-100%) that females carried and supported their 0-14 day-old infants in a ventral position during locomotion (r(s) = -0.68, p < 0.05) and tended to be negatively correlated with the total amount of time (0-100%) they spent in ventro-ventral contact with their infants (r(s) = -0.58; p < 0.10). This study provides the first physiological evidence that postpartum stress is an important etiologic factor in gorilla maternal failure in captive environments.