Similar cognitive deficits in mice and humans in the chronic phase post-stroke identified using the touchscreen-based paired-associate learning task.

For many chronic stroke survivors, persisting cognitive dysfunction leads to significantly reduced quality of life. Translation of promising therapeutic strategies aimed at improving cognitive function is hampered by existing, disparate cognitive assessments in animals and humans. In this study, we assessed post-stroke cognitive function using a comparable touchscreen-based paired-associate learning task in a cross-sectional population of chronic stroke survivors (≥ 5 months post-stroke, n = 70), age-matched controls (n = 70), and in mice generated from a C57BL/6 mouse photothrombotic stroke model (at six months post-stroke). Cognitive performance of stroke survivors was analysed using linear regression adjusting for age, gender, diabetes, systolic blood pressure and waist circumference. Stroke survivors made significantly fewer correct choices across all tasks compared with controls. Similar cognitive impairment was observed in the mice post-stroke with fewer correct choices compared to shams. These results highlight the feasibility and potential value of analogous modelling of clinically meaningful cognitive impairments in chronic stroke survivors and in mice in chronic phase after stroke. Implementation of validated, parallel cross-species test platforms for cognitive assessment offer the potential of delivering a more useful framework for evaluating therapies aimed at improving long-term cognitive function post-stroke.

Opposing Associations of Stress and Resilience With Functional Outcomes in Stroke Survivors in the Chronic Phase of Stroke: A Cross-Sectional Study.

Stroke survivors report significant levels of psychological distress post stroke. To date, most studies conducted have focused on the relationship between psychological stress and functional outcomes in the acute phase of stroke. However, no studies had considered the role of stress over the chronic phase, where stress may continue to exert negative effects on cognitive and psychological processes. Further, the role of potentially modulatory variables, such as psychological resilience, on stroke outcomes has been understudied. The purpose of this study was to consider the relationships between stress and resilience with functional outcomes in long-term survivors of stroke. People (N = 70) who had experienced a stroke between 5 months and 28 years ago were included in the cross-sectional study, along with age-matched controls (N = 70). We measured stress using both the Perceived Stress Scale and biological markers, and resilience using both the Brief Resilience Scale and the Connor-Davidson Resilience Scale. Stroke outcomes were assessed using the Stroke Impact Scale. We found that, compared with age-matched controls, stroke survivors reported greater levels of perceived stress, and lower levels of resilience. In stroke survivors, both perceived stress and resilience were independently associated with stroke outcomes in linear regression models. In particular, these relationships were observed for cognitive outcomes including mood, memory, and communication. The association between stress and stroke outcome did not differ across time post stroke. Given that resilience is a modifiable psychological construct, future research may consider whether strategies directed at enhancing resilience may improve recovery from stroke. Australia and New Zealand Clinical Trials Registry: ACTRN12617000736347.

Exploring the relationship between fatigue and circulating levels of the pro-inflammatory biomarkers interleukin-6 and C-reactive protein in the chronic stage of stroke recovery: A cross-sectional study.

BACKGROUND: The precise mechanisms underlying the aetiology of post-stroke fatigue remain poorly understood. Inflammation has been associated with clinically significant fatigue across a number of neurological disorders; however, at present there is a lack of evidence regarding the association of fatigue and inflammation in the chronic phase of stroke recovery. AIMS: The aim of this study was to examine fatigue in a cohort of stroke survivors in the chronic phase of stroke, compared with matched controls, and to explore associations between the pro-inflammatory cytokine interleukin-6, high-sensitivity C-reactive Protein and fatigue. METHODS: We performed an exploratory cross-sectional study of 70 people in the chronic phase of stroke recovery, and 70 age matched controls. Fatigue was assessed using the Fatigue Assessment Scale. Interleukin-6 was measured in serum using a commercially available enzyme immunoassay kit. Both outcome measures were assessed contemporaneously. RESULTS: Clinically significant fatigue, defined as a score ≥24 on the Fatigue Assessment Scale, was reported by 60% of stroke survivors, and 15.7% of controls. The odds of experiencing clinically significant fatigue was 8.04 times higher among stroke survivors compared to control participants (odds ratio 8.045; 95% CI: 3.608, 17.939; P â€‹< â€‹0.001). The fatigue score was significantly correlated with the level of both interleukin-6 and high-sensitivity c-reactive protein, however once entered into a linear regression model with cardiovascular covariables, this relationship was no longer statistically significant. CONCLUSIONS: This study shows that fatigue may be associated with systemic inflammation in the chronic phase of stroke. The pathological mechanisms underlying post-stroke fatigue and its clinical implications require further study.

Low oxygen post conditioning prevents thalamic secondary neuronal loss caused by excitotoxicity after cortical stroke.

In the current study, we were interested in investigating whether Low oxygen post-conditioning (LOPC) was capable of limiting the severity of stroke-induced secondary neurodegeneration (SND). To investigate the effect of LOPC we exposed adult male C57/BL6 mice to photothrombotic occlusion (PTO) of the motor and somatosensory cortex. This is known to induce progressive neurodegeneration in the thalamus within two weeks of infarction. Two days after PTO induction mice were randomly assigned to one of four groups: (i) LOPC-15 day exposure group; (ii) a LOPC 15 day exposure followed by a 15 day exposure to normal atmosphere; (iii) normal atmosphere for 15 days and (iv) normal atmosphere for 30 days (n = 20/group). We observed that LOPC reduced the extent of neuronal loss, as indicated by assessment of both area of loss and NeuN+ cell counts, within the thalamus. Additionally, we identified that LOPC reduced microglial activity and decreased activity within the excitotoxic signalling pathway of the NMDAR axis. Together, these findings suggest that LOPC limits neuronal death caused by excitotoxicity in sites of secondary damage and promotes neuronal survival. In conclusion, this work supports the potential of utilising LOPC to intervene in the sub-acute phase post-stroke to restrict the severity of SND.

The role of human Metapneumovirus genetic diversity and nasopharyngeal viral load on symptom severity in adults.

BACKGROUND: Human metapneumovirus (HMPV) is established as one of the causative agents of respiratory tract infections. To date, there are limited reports that describe the effect of HMPV genotypes and/or viral load on disease pathogenesis in adults. This study aims to determine the role of HMPV genetic diversity and nasopharyngeal viral load on symptom severity in outpatient adults with acute respiratory tract infections. METHODS: Severity of common cold symptoms of patients from a teaching hospital was assessed by a four-category scale and summed to obtain the total symptom severity score (TSSS). Association between the fusion and glycoprotein genes diversity, viral load (quantified using an improved RT-qPCR assay), and symptom severity were analyzed using bivariate and linear regression analyses. RESULTS: Among 81/3706 HMPV-positive patients, there were no significant differences in terms of demographics, number of days elapsed between symptom onset and clinic visit, respiratory symptoms manifestation and severity between different HMPV genotypes/sub-lineages. Surprisingly, elderly patients (≥65 years old) had lower severity of symptoms (indicated by TSSS) than young and middle age adults (p = 0.008). Nasopharyngeal viral load did not correlate with nor predict symptom severity of HMPV infection. Interestingly, at 3-5 days after symptom onset, genotype A-infected patients had higher viral load compared to genotype B (4.4 vs. 3.3 log10 RNA copies/µl) (p = 0.003). CONCLUSIONS: Overall, HMPV genetic diversity and viral load did not impact symptom severity in adults with acute respiratory tract infections. Differences in viral load dynamics over time between genotypes may have important implications on viral transmission.

Growth Hormone Improves Cognitive Function After Experimental Stroke.

BACKGROUND AND PURPOSE: Cognitive impairment is a common outcome for stroke survivors. Growth hormone (GH) could represent a potential therapeutic option as this peptide hormone has been shown to improve cognition in various clinical conditions. In this study, we evaluated the effects of peripheral administration of GH at 48 hours poststroke for 28 days on cognitive function and the underlying mechanisms. METHODS: Experimental stroke was induced by photothrombotic occlusion in young adult mice. We assessed the associative memory cognitive domain using mouse touchscreen platform for paired-associate learning task. We also evaluated neural tissue loss, neurotrophic factors, and markers of neuroplasticity and cerebrovascular remodeling using biochemical and histology analyses. RESULTS: Our results show that GH-treated stroked mice made a significant improvement on the paired-associate learning task relative to non-GH-treated mice at the end of the study. Furthermore, we observed reduction of neural tissue loss in GH-treated stroked mice. We identified that GH treatment resulted in significantly higher levels of neurotrophic factors (IGF-1 [insulin-like growth factor-1] and VEGF [vascular endothelial growth factor]) in both the circulatory and peri-infarct regions. GH treatment in stroked mice not only promoted protein levels and density of presynaptic marker (SYN-1 [synapsin-1]) and marker of myelination (MBP [myelin basic protein]) but also increased the density and area coverage of 2 major vasculature markers (CD31 and collagen-IV), within the peri-infarct region. CONCLUSIONS: These findings provide compelling preclinical evidence for the usage of GH as a potential therapeutic tool in the recovery phase of patients after stroke.

Genome Sequence of a Novel HIV-1 Circulating Recombinant Form (CRF77_cpx) Identified among Blood Donors in Malaysia.

We report here the first HIV-1 circulating recombinant form (CRF) complex identified among the blood donors in Malaysia. The CRF77_cpx mosaic genome consists of parental subtypes B', C, and CRF01_AE and is structurally related to CRF07_BC. The identification of CRF77_cpx underlines the genetic complexity and mobility of HIV-1 among the blood donors.

Extensive Genetic Diversity of HIV-1 in Incident and Prevalent Infections among Malaysian Blood Donors: Multiple Introductions of HIV-1 Genotypes from Highly Prevalent Countries.

Transfusion-transmissible infections including HIV-1 continue to pose major risks for unsafe blood transfusions due to both window phase infections and divergent viruses that may not be detected by donor screening assays. Given the recent emergence of several HIV-1 circulating recombinant forms (CRFs) in high-risk populations in the Southeast Asia region, we investigated the genetic diversity of HIV-1 among the blood donors in Kuala Lumpur, Malaysia. A total of 211 HIV-positive plasma samples detected among 730,188 donations to the National Blood Centre between 2013 and 2014 were provided (90.5% male, median age: 27.0 years old). Recent or long-term infection status at the time of donation was determined using a limiting antigen avidity enzyme immunoassay (LAg-Avidity EIA). HIV-1 gag-pol genes were amplified and sequenced from residual plasma for 149 cases followed by genotype determination using phylogenetic and recombination analyses. Transmitted antiretroviral resistance mutations were not observed among the blood donors, among which 22.7% were classified as recent or incident infections. Major circulating HIV-1 genotypes determined by neighbour-joining phylogenetic inference included CRF01_AE at 40.9% (61/149), CRF33_01B at 21.5% (32/149), and subtype B at 10.1% (15/149). Newly-described CRFs including CRF54_01B circulated at 4.0%, CRF74_01B at 2.0%, and CRF53_01B and CRF48_01B at 0.7% each. Interestingly, unique HIV-1 genotypes including African subtype G (8.7%), CRF45_cpx (1.3%), CRF02_AG (0.7%) and CRF07_BC (0.7%) from China were detected for the first time in the country. A cluster of subtype G sequences formed a distinct founder sub-lineage within the African strains. In addition, 8.7% (13/149) of HIV-infected donors had unique recombinant forms (URFs) including CRF01_AE/B' (4.7%), B'/C (2.7%) and B'/G (1.3%) recombinants. Detailed analysis identified similar recombinant structures with shared parental strains among the B'/C and B'/G URFs, some of which were sequenced from recently infected individuals, indicating the possible emergence and on-going spread of foreign clades of CRF candidates among the local population. The findings demonstrate extensive molecular complexity of HIV-1 among the infected blood donors in Malaysia, driven in part by the increased spread of recently described CRFs and multiple introductions of previously unreported genotypes from highly prevalent countries.

Genetic diversity, seasonality and transmission network of human metapneumovirus: identification of a unique sub-lineage of the fusion and attachment genes.

Human metapneumovirus (HMPV) is an important viral respiratory pathogen worldwide. Current knowledge regarding the genetic diversity, seasonality and transmission dynamics of HMPV among adults and children living in tropical climate remains limited. HMPV prevailed at 2.2% (n = 86/3,935) among individuals presented with acute respiratory tract infections in Kuala Lumpur, Malaysia between 2012 and 2014. Seasonal peaks were observed during the northeast monsoon season (November-April) and correlated with higher relative humidity and number of rainy days (P < 0.05). Phylogenetic analysis of the fusion and attachment genes identified the co-circulation of three known HMPV sub-lineages, A2b and B1 (30.2% each, 26/86) and B2 (20.9%, 18/86), with genotype shift from sub-lineage B1 to A2b observed in 2013. Interestingly, a previously unrecognized sub-lineage of A2 was identified in 18.6% (16/86) of the population. Using a custom script for network construction based on the TN93 pairwise genetic distance, we identified up to nine HMPV transmission clusters circulating as multiple sub-epidemics. Although no apparent major outbreak was observed, the increased frequency of transmission clusters (dyads) during seasonal peaks suggests the potential roles of transmission clusters in driving the spread of HMPV. Our findings provide essential information for therapeutic research, prevention strategies, and disease outbreak monitoring of HMPV.

Genetic Characterization of a Panel of Diverse HIV-1 Isolates at Seven International Sites.

HIV-1 subtypes and drug resistance are routinely tested by many international surveillance groups. However, results from different sites often vary. A systematic comparison of results from multiple sites is needed to determine whether a standardized protocol is required for consistent and accurate data analysis. A panel of well-characterized HIV-1 isolates (N = 50) from the External Quality Assurance Program Oversight Laboratory (EQAPOL) was assembled for evaluation at seven international sites. This virus panel included seven subtypes, six circulating recombinant forms (CRFs), nine unique recombinant forms (URFs) and three group O viruses. Seven viruses contained 10 major drug resistance mutations (DRMs). HIV-1 isolates were prepared at a concentration of 107 copies/ml and compiled into blinded panels. Subtypes and DRMs were determined with partial or full pol gene sequences by conventional Sanger sequencing and/or Next Generation Sequencing (NGS). Subtype and DRM results were reported and decoded for comparison with full-length genome sequences generated by EQAPOL. The partial pol gene was amplified by RT-PCR and sequenced for 89.4%-100% of group M viruses at six sites. Subtyping results of majority of the viruses (83%-97.9%) were correctly determined for the partial pol sequences. All 10 major DRMs in seven isolates were detected at these six sites. The complete pol gene sequence was also obtained by NGS at one site. However, this method missed six group M viruses and sequences contained host chromosome fragments. Three group O viruses were only characterized with additional group O-specific RT-PCR primers employed by one site. These results indicate that PCR protocols and subtyping tools should be standardized to efficiently amplify diverse viruses and more consistently assign virus genotypes, which is critical for accurate global subtype and drug resistance surveillance. Targeted NGS analysis of partial pol sequences can serve as an alternative approach, especially for detection of low-abundance DRMs.

Co-infections and transmission networks of HCV, HIV-1 and HPgV among people who inject drugs.

Co-infections with human immunodeficiency virus type 1 (HIV-1) and human pegivirus (HPgV) are common in hepatitis C virus (HCV)-infected individuals. However, analysis on the evolutionary dynamics and transmission network profiles of these viruses among individuals with multiple infections remains limited. A total of 228 injecting drug users (IDUs), either HCV- and/or HIV-1-infected, were recruited in Kuala Lumpur, Malaysia. HCV, HIV-1 and HPgV genes were sequenced, with epidemic growth rates assessed by the Bayesian coalescent method. Based on the sequence data, mono-, dual- and triple-infection were detected in 38.8%, 40.6% and 20.6% of the subjects, respectively. Fifteen transmission networks involving HCV (subtype 1a, 1b, 3a and 3b), HIV-1 (CRF33_01B) and HPgV (genotype 2) were identified and characterized. Genealogical estimates indicated that the predominant HCV, HIV-1 and HPgV genotypes were introduced into the IDUs population through multiple sub-epidemics that emerged as early as 1950s (HCV), 1980s (HIV-1) and 1990s (HPgV). By determining the difference in divergence times between viral lineages (ΔtMRCA), we also showed that the frequency of viral co-transmission is low among these IDUs. Despite increased access to therapy and other harm reduction interventions, the continuous emergence and coexistence of new transmission networks suggest persistent multiple viral transmissions among IDUs.

Impact of HIV-1 Subtype on the Time to CD4+ T-Cell Recovery in Combination Antiretroviral Therapy (cART)-Experienced Patients.

Human immunodeficiency virus type 1 (HIV-1) subtypes have been shown to differ in the rate of clinical progression. We studied the association between HIV-1 subtypes and the rate of CD4+ T-cell recovery in a longitudinal cohort of patients on combination antiretroviral therapy (cART). We studied 103 patients infected with CRF01_AE (69%) and subtype B (31%) who initiated cART between 2006 and 2013. Demographic data, CD4+ T-cell counts and HIV-1 viral load were abstracted from patient medical charts. Kaplan-Meier was used to estimate the time to CD4+ T-cell count increase to ≥350 between subtypes and effects of covariates were analysed using Cox proportional hazards. An 87% of the study population were male adults (mean age of 38.7 years old). Baseline CD4+ T-cell counts and viral loads, age at cART initiation, sex, ethnicity and co-infection did not differ significantly between subtypes. A shorter median time for CD4+ T-cell count increase to ≥350 cells/µL was observed for CRF01_AE (546 days; 95% confidence interval [CI], 186-906 days; P = .502) compared to subtype B (987 days; 95% CI, 894-1079 days). In multivariate analysis, female sex was significantly associated with a 2.7 times higher chance of achieving CD4+ T-cell recovery (adjusted hazard ratio [HR], 2.75; 95% CI, 1.21-6.22; P = .025) and both baseline CD4+ T-cell count (P = .001) and viral load (P = .001) were important predictors for CD4+ T-cell recovery. Immunological recovery correlated significantly with female sex, baseline CD4+ T-cell counts and viral load but not subtype.

Genetic Characterization of a Novel HIV-1 Circulating Recombinant Form (CRF74_01B) Identified among Intravenous Drug Users in Malaysia: Recombination History and Phylogenetic Linkage with Previously Defined Recombinant Lineages.

In many parts of Southeast Asia, the HIV-1 epidemic has been driven by the sharing of needles and equipment among intravenous drug users (IDUs). Over the last few decades, many studies have proven time and again that the diversity of HIV-1 epidemics can often be linked to the route of infection transmission. That said, the diversity and complexity of HIV-1 molecular epidemics in the region have been increasing at an alarming rate, due in part to the high tendency of the viral RNA to recombine. This scenario was exemplified by the discovery of numerous circulating recombinant forms (CRFs), especially in Thailand and Malaysia. In this study, we characterized a novel CRF designated CRF74_01B, which was identified in six epidemiologically unlinked IDUs in Kuala Lumpur, Malaysia. The near-full length genomes were composed of CRF01_AE and subtype B', with eight breakpoints dispersed in the gag-pol and nef regions. Remarkably, this CRF shared four and two recombination hotspots with the previously described CRF33_01B and the less prevalent CRF53_01B, respectively. Genealogy-based Bayesian phylogenetic analysis of CRF74_01B genomic regions showed that it is closely related to both CRF33_01B and CRF53_01B. This observation suggests that CRF74_01B was probably a direct descendent from specific lineages of CRF33_01B, CRF53_01B and subtype B' that could have emerged in the mid-1990s. Additionally, it illustrated the active recombination processes between prevalent HIV-1 subtypes and recombinants in Malaysia. In summary, we report a novel HIV-1 genotype designated CRF74_01B among IDUs in Kuala Lumpur, Malaysia. The characterization of the novel CRF74_01B is of considerable significance towards the understanding of the genetic diversity and population dynamics of HIV-1 circulating in the region.

Genome Sequence of a Complex HIV-1 Unique Recombinant Form Involving CRF01_AE, Subtype B, and Subtype B' Identified in Malaysia.

A complex HIV-1 unique recombinant form involving subtypes CRF01_AE, B, and B' was recently identified from an injecting drug user in Malaysia. A total of 13 recombination breakpoints were mapped across the near-full-length genome of isolate 10MYPR226, indicating the increasingly diverse molecular epidemiology and frequent linkage among various high-risk groups.

A newly emerging HIV-1 recombinant lineage (CRF58_01B) disseminating among people who inject drugs in Malaysia.

The HIV epidemic is primarily characterised by the circulation of HIV-1 group M (main) comprising of 11 subtypes and sub-subtypes (A1, A2, B-D, F1, F2, G, H, J, and K) and to date 55 circulating recombinant forms (CRFs). In Southeast Asia, active inter-subtype recombination involving three main circulating genotypes--subtype B (including subtype B', the Thai variant of subtype B), CRF01_AE, and CRF33_01B--have contributed to the emergence of novel unique recombinant forms. In the present study, we conducted the molecular epidemiological surveillance of HIV-1 gag-RT genes among 258 people who inject drugs (PWIDs) in Kuala Lumpur, Malaysia, between 2009 and 2011 whereby a novel CRF candidate was recently identified. The near full-length genome sequences obtained from six epidemiologically unlinked individuals showed identical mosaic structures consisting of subtype B' and CRF01_AE, with six unique recombination breakpoints in the gag-RT, pol, and env regions. Among the high-risk population of PWIDs in Malaysia, which was predominantly infected by CRF33_01B (>70%), CRF58_01B circulated at a low but significant prevalence (2.3%, 6/258). Interestingly, the CRF58_01B shared two unique recombination breakpoints with other established CRFs in the region: CRF33_01B, CRF48_01B, and CRF53_01B in the gag gene, and CRF15_01B (from Thailand) in the env gene. Extended Bayesian Markov chain Monte Carlo sampling analysis showed that CRF58_01B and other recently discovered CRFs were most likely to have originated in Malaysia, and that the recent spread of recombinant lineages in the country had little influence from neighbouring countries. The isolation, genetic characterization, and evolutionary features of CRF58_01B among PWIDs in Malaysia signify the increasingly complex HIV-1 diversity in Southeast Asia that may hold an implication on disease treatment, control, and prevention.

Molecular diversity of HIV-1 among people who inject drugs in Kuala Lumpur, Malaysia: massive expansion of circulating recombinant form (CRF) 33_01B and emergence of multiple unique recombinant clusters.

Since the discovery of HIV-1 circulating recombinant form (CRF) 33_01B in Malaysia in the early 2000 s, continuous genetic diversification and active recombination involving CRF33_01B and other circulating genotypes in the region including CRF01_AE and subtype B' of Thai origin, have led to the emergence of novel CRFs and unique recombinant forms. The history and magnitude of CRF33_01B transmission among various risk groups including people who inject drugs (PWID) however have not been investigated despite the high epidemiological impact of CRF33_01B in the region. We update the most recent molecular epidemiology of HIV-1 among PWIDs recruited in Malaysia between 2010 and 2011 by population sequencing and phylogenetic analysis of 128 gag-pol sequences. HIV-1 CRF33_01B was circulating among 71% of PWIDs whilst a lower prevalence of other previously dominant HIV-1 genotypes [subtype B' (11%) and CRF01_AE (5%)] and CRF01_AE/B' unique recombinants (13%) were detected, indicating a significant shift in genotype replacement in this population. Three clusters of CRF01_AE/B' recombinants displaying divergent yet phylogenetically-related mosaic genomes to CRF33_01B were identified and characterized, suggestive of an abrupt emergence of multiple novel CRF clades. Using rigorous maximum likelihood approach and the Bayesian Markov chain Monte Carlo (MCMC) sampling of CRF33_01Bpol sequences to elucidate the past population dynamics, we found that the founder lineages of CRF33_01B were likely to have first emerged among PWIDs in the early 1990 s before spreading exponentially to various high and low-risk populations (including children who acquired infections from their mothers) and later on became endemic around the early 2000 s. Taken together, our findings provide notable genetic evidence indicating the widespread expansion of CRF33_01B among PWIDs and into the general population. The emergence of numerous previously unknown recombinant clades highlights the escalating genetic complexity of HIV-1 in the Southeast Asian region.

Genome sequences of a novel HIV-1 CRF53_01B identified in Malaysia.

A novel HIV-1 genotype designated CRF53_01B was recently characterized from three epidemiologically unrelated persons in Malaysia. Here we announced three recently isolated full-length genomes of CRF53_01B, which is likely to be phylogenetically linked to CRF33_01B, circulating widely in Southeast Asia. The genome sequences may contribute to HIV-1 molecular surveillance and future vaccine development in the region.