Hemagglutinin destabilization in H3N2 vaccine reference viruses skews antigenicity and prevents airborne transmission in ferrets.

During influenza virus entry, the hemagglutinin (HA) protein binds receptors and causes membrane fusion after endosomal acid activation. To improve vaccine efficiency and pandemic risk assessment for currently-dominant H3N2 influenza viruses, we investigated HA stability of 6 vaccine reference viruses and 42 circulating viruses. Recent vaccine reference viruses had destabilized HA proteins due to egg-adaptive mutation HA1-L194P. Virus growth in cell culture was independent of HA stability. In ferrets, the vaccine reference viruses and circulating viruses required a relatively stable HA (activation and inactivation pH < 5.5) for airborne transmissibility. The recent vaccine reference viruses with destabilized HA proteins had reduced infectivity, had no airborne transmissibility unless reversion to HA1-P194L occurred, and had skewed antigenicity away from the studied viruses and circulating H3N2 viruses. Other vaccine reference viruses with stabilized HAs retained infectivity, transmissibility, and antigenicity. Therefore, HA stabilization should be prioritized over destabilization in vaccine reference virus selection to reduce mismatches between vaccine and circulating viruses.

Swine H1N1 Influenza Virus Variants with Enhanced Polymerase Activity and HA Stability Promote Airborne Transmission in Ferrets.

Understanding how animal influenza A viruses (IAVs) acquire airborne transmissibility in humans and ferrets is needed to prepare for and respond to pandemics. Here, we investigated in ferrets the replication and transmission of swine H1N1 isolates P4 and G15, whose majority population had decreased polymerase activity and poor hemagglutinin (HA) stability, respectively. For both isolates, a minor variant was selected and transmitted in ferrets. Polymerase-enhancing variant PA-S321 airborne-transmitted and propagated in one ferret. HA-stabilizing variant HA1-S210 was selected in all G15-inoculated ferrets and was transmitted by contact and airborne routes. With an efficient polymerase and a stable HA, the purified minor variant G15-HA1-S210 had earlier and higher peak titers in inoculated ferrets and was recovered at a higher frequency after airborne transmission than P4 and G15. Overall, HA stabilization played a more prominent role than polymerase enhancement in the replication and transmission of these viruses in ferrets. The results suggest pandemic risk-assessment studies may benefit from deep sequencing to identify minor variants with human-adapted traits. IMPORTANCE Diverse IAVs circulate in animals, yet few acquire the viral traits needed to start a human pandemic. A stabilized HA and mammalian-adapted polymerase have been shown to promote the adaptation of IAVs to humans and ferrets (the gold-standard model for IAV replication, pathogenicity, and transmissibility). Here, we used swine IAV isolates of the gamma lineage as a model to investigate the importance of HA stability and polymerase activity in promoting replication and transmission in ferrets. These are emerging viruses that bind to both α-2,6- and α-2,3-linked receptors. Using isolates containing mixed populations, a stabilized HA was selected within days in inoculated ferrets. An enhanced polymerase was also selected and propagated after airborne transmission to a ferret. Thus, HA stabilization was a stricter requirement, yet both traits promoted transmissibility. Knowing the viral traits needed for pandemic potential, and the relative importance of each, will help identify emerging viruses of greatest concern.

The Impact of CRISPR-Cas9 on Age-related Disorders: From Pathology to Therapy.

With advances in medical technology, the number of people over the age of 60 is on the rise, and thus, increasing the prevalence of age-related pathologies within the aging population. Neurodegenerative disorders, cancers, metabolic and inflammatory diseases are some of the most prevalent age-related pathologies affecting the growing population. It is imperative that a new treatment to combat these pathologies be developed. Although, still in its infancy, the CRISPR-Cas9 system has become a potent gene-editing tool capable of correcting gene-mediated age-related pathology, and therefore ameliorating or eliminating disease symptoms. Deleting target genes using the CRISPR-Cas9 system or correcting for gene mutations may ameliorate many different neurodegenerative disorders detected in the aging population. Cancer cells targeted by the CRISPR-Cas9 system may result in an increased sensitivity to chemotherapeutics, lower proliferation, and higher cancer cell death. Finally, reducing gene targeting inflammatory molecules production through microRNA knockout holds promise as a therapeutic strategy for both arthritis and inflammation. Here we present a review based on how the expanding world of genome editing can be applied to disorders and diseases affecting the aging population.

Reduced-Beclin1-Expressing Mice Infected with Zika-R103451 and Viral-Associated Pathology during Pregnancy.

Here, we used a mouse model with defective autophagy to further decipher the role of Beclin1 in the infection and disease of Zika virus (ZIKV)-R103451. Hemizygous (Becn1+/-) and wild-type (Becn1+/+) pregnant mice were transiently immunocompromised using the anti-interferon alpha/beta receptor subunit 1 monoclonal antibody MAR1-5A3. Despite a low mortality rate among the infected dams, 25% of Becn1+/- offspring were smaller in size and had smaller, underdeveloped brains. This phenotype became apparent after 2-to 3-weeks post-birth. Furthermore, the smaller-sized pups showed a decrease in the mRNA expression levels of insulin-like growth factor (IGF)-1 and the expression levels of several microcephaly associated genes, when compared to their typical-sized siblings. Neuronal loss was also noticeable in brain tissues that were removed postmortem. Further analysis with murine mixed glia, derived from ZIKV-infected Becn1+/- and Becn1+/+ pups, showed greater infectivity in glia derived from the Becn1+/- genotype, along with a significant increase in pro-inflammatory molecules. In the present study, we identified a link by which defective autophagy is causally related to increased inflammatory molecules, reduced growth factor, decreased expression of microcephaly-associated genes, and increased neuronal loss. Specifically, we showed that a reduced expression of Beclin1 aggravated the consequences of ZIKV infection on brain development and qualifies Becn1 as a susceptibility gene of ZIKV congenital syndrome.

Morphine counteracts the antiviral effect of antiretroviral drugs and causes upregulation of p62/SQSTM1 and histone-modifying enzymes in HIV-infected astrocytes.

Accelerated neurological disorders are increasingly prominent among the HIV-infected population and are likely driven by the toxicity from long-term use of antiretroviral drugs. We explored potential side effects of antiretroviral drugs in HIV-infected primary human astrocytes and whether opioid co-exposure exacerbates the response. HIV-infected human astrocytes were exposed to the reverse transcriptase inhibitor, emtricitabine, alone or in combination with two protease inhibitors ritonavir and atazanavir (ERA) with and without morphine co-exposure. The effect of the protease inhibitor, lopinavir, alone or in combination with the protease inhibitor, abacavir, and the integrase inhibitor, raltegravir (LAR), with and without morphine co-exposure was also explored. Exposure with emtricitabine alone or ERA in HIV-infected astrocytes caused a significant decrease in viral replication and attenuated HIV-induced inflammatory molecules, while co-exposure with morphine negated the inhibitory effects of ERA, leading to increased viral replication and inflammatory molecules. Exposure with emtricitabine alone or in combination with morphine caused a significant disruption of mitochondrial membrane integrity. Genetic analysis revealed a significant increase in the expression of p62/SQSTM1 which correlated with an increase in the histone-modifying enzyme, ESCO2, after exposure with ERA alone or in combination with morphine. Furthermore, several histone-modifying enzymes such as CIITA, PRMT8, and HDAC10 were also increased with LAR exposure alone or in combination with morphine. Accumulation of p62/SQSTM1 is indicative of dysfunctional lysosomal fusion. Together with the loss of mitochondrial integrity and epigenetic changes, these effects may lead to enhanced viral titer and inflammatory molecules contributing to the neuropathology associated with HIV.

Toll-like receptor 3 regulates Zika virus infection and associated host inflammatory response in primary human astrocytes.

The connection between Zika virus (ZIKV) and neurodevelopmental defects is widely recognized, although the mechanisms underlying the infectivity and pathology in primary human glial cells are poorly understood. Here we show that three isolated strains of ZIKV, an African strain MR766 (Uganda) and two closely related Asian strains R103451 (Honduras) and PRVABC59 (Puerto Rico) productively infect primary human astrocytes, although Asian strains showed a higher infectivity rate and increased cell death when compared to the African strain. Inhibition of AXL receptor significantly attenuated viral entry of MR766 and PRVABC59 and to a lesser extend R103451, suggesting an important role of TAM receptors in ZIKV cell entry, irrespective of lineage. Infection by PRVABC59 elicited the highest release of inflammatory molecules, with a 8-fold increase in the release of RANTES, 10-fold increase in secretion of IP-10 secretion and a 12-fold increase in IFN-ß secretion when compared to un-infected human astrocytes. Minor changes in the release of several growth factors, endoplasmic reticulum (ER)-stress response factors and the transcription factor, NF-κB were detected with the Asian strains, while significant increases in FOXO6, MAPK10 and JNK were detected with the African strain. Activation of the autophagy pathway was evident with increased expression of the autophagy related proteins Beclin1, LC3B and p62/SQSTM1 with all three strains of ZIKV. Pharmacological inhibition of the autophagy pathway and genetic inhibition of the Beclin1 showed minimal effects on ZIKV replication. The expression of toll-like receptor 3 (TLR3) was significantly increased with all three strains of ZIKV; pharmacological and genetic inhibition of TLR3 caused a decrease in viral titers and in viral-induced inflammatory response in infected astrocytes. We conclude that TLR3 plays a vital role in both ZIKV replication and viral-induced inflammatory responses, irrespective of the strains, while the autophagy protein Beclin1 influences host inflammatory responses.

Complementary Mechanisms Potentially Involved in the Pathology of Zika Virus.

Zika virus (ZIKV) has emerged as a global health threat due to its neuro-teratogenic effect and wide range of transmission routes. Most recently, ZIKV infection has been linked with both autoimmune disorders in adults and neurodevelopmental disorders in newborns. Researchers are exploring potential cellular and molecular mechanisms underlying the neuro-teratogenicity and related consequences by using various in vitro cell culture methods and in vivo animal models. Though some of the putative viral entry receptors have been identified for ZIKV entry into the target cells, the exact mechanism of ZIKV entry or induced pathology are still not clear. Some of the important host cellular pathways including the toll-like receptor (TLR), autophagy, apoptosis and unfolded protein response (UPR) pathways are considered potential mechanism(s) for ZIKV induced neuroinflammation and for neurodevelopmental disorders. Since there is still a dire need for efficient treatment and vaccine to prevent ZIKV mediated disorders, a better understanding of the interaction between virus and host cellular pathways could pave the way for development of targeted therapeutic intervention. In this review, we are focusing on the recent advances and current knowledge regarding the interaction of ZIKV with abovementioned pathways so as to provide basic understanding to execute further research that could aid in the development of novel therapeutic strategy.

Signaling pathways and therapeutic perspectives related to environmental factors associated with multiple sclerosis.

Multiple sclerosis (MS) is an immune-mediated demyelinating disorder of unknown etiology. Both genetic-susceptibility and environment exposures, including vitamin D deficiency, Epstein-Barr viral and Herpesvirus (HHV-6) infections are strongly implicated in the activation of T cells and MS-pathogenesis. Despite precise knowledge of how these factors could be operating alone or in combination to facilitate and aggravate the disease progression, it is clear that prolonged induction of inflammatory molecules and recruitment of other immune cells by the activated T cells results in demyelination and axonal damage. It is imperative to understand the risk factors associated with MS progression and how these factors contribute to disease pathology. Understanding of the underlying mechanisms of what factors triggers activation of T cells to attack myelin antigen are important to strategize therapeutics and therapies against MS. Current review provides a detailed literature to understand the role of both pathogenic and non-pathogenic factors on the impact of MS.

Critical Role of Beclin1 in HIV Tat and Morphine-Induced Inflammation and Calcium Release in Glial Cells from Autophagy Deficient Mouse.

We previously showed that autophagy is an important component in human immunodeficiency virus (HIV) replication and in the combined morphine-induced neuroinflammation in human astrocytes and microglia. Here we further studied the consequences of autophagy using glial cells of mice partially lacking the essential autophagy gene Atg6 (Beclin1) exposed to HIV Tat and morphine. Tat is known to cause an inflammatory response, increase calcium release, and possibly interact with autophagy pathway proteins. Following Tat exposure, autophagy-deficient (Becn1+/-) glial cells had significantly and consistently reduced levels in the pro-inflammatory cytokine IL-6 and the chemokines RANTES and MCP-1 when compared to Tat-treated cells from control (C57BL/6J) mice, suggesting an association between the inflammatory effects of Tat and Beclin1. Further, differences in RANTES and MCP-1 secretion between C57BL/6J and Becn1+/- glia treated with Tat and morphine also suggest a role of Beclin1 in the morphine-induced enhancement. Analysis of autophagy maturation by immunoblot suggests that Beclin1 may be necessary for Tat, and to a lesser extent morphine-induced arrest of the pathway as demonstrated by accumulation of the adaptor protein p62/SQSTM1 in C57BL/6J glia. Calcium release induced by Tat alone or in combination with morphine in C57BL/6J glia was significantly reduced in Becn1+/- glia while minimal interactive effect of Tat with morphine in the production of reactive oxygen or nitrogen species was detected in glia derived from Becn1+/- or C57BL/6J. Overall, the data establish a role of Beclin1 in Tat and morphine-mediated inflammatory responses and calcium release in glial cells and support the notion that autophagy mediates Tat alone and combined morphine-induced neuropathology.

Author Correction: Intranasal drug delivery of small interfering RNA targeting Beclin1 encapsulated with polyethylenimine (PEI) in mouse brain to achieve HIV attenuation.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Importance of Autophagy in Mediating Human Immunodeficiency Virus (HIV) and Morphine-Induced Metabolic Dysfunction and Inflammation in Human Astrocytes.

Under physiological conditions, the function of astrocytes in providing brain metabolic support is compromised under pathophysiological conditions caused by human immunodeficiency virus (HIV) and opioids. Herein, we examined the role of autophagy, a lysosomal degradation pathway important for cellular homeostasis and survival, as a potential regulatory mechanism during pathophysiological conditions in primary human astrocytes. Blocking autophagy with small interfering RNA (siRNA) targeting BECN1, but not the Autophagy-related 5 (ATG5) gene, caused a significant decrease in HIV and morphine-induced intracellular calcium release. On the contrary, inducing autophagy pharmacologically with rapamycin further enhanced calcium release and significantly reverted HIV and morphine-decreased glutamate uptake. Furthermore, siBeclin1 caused an increase in HIV-induced nitric oxide (NO) release, while viral-induced NO in astrocytes exposed to rapamycin was decreased. HIV replication was significantly attenuated in astrocytes transfected with siRNA while significantly induced in astrocytes exposed to rapamycin. Silencing with siBeclin1, but not siATG5, caused a significant decrease in HIV and morphine-induced interleukin (IL)-8 and tumor necrosis factor alpha (TNF-α) release, while secretion of IL-8 was significantly induced with rapamycin. Mechanistically, the effects of siBeclin1 in decreasing HIV-induced calcium release, viral replication, and viral-induced cytokine secretion were associated with a decrease in activation of the nuclear factor kappa B (NF-κB) pathway.

Impact of mass drug administration for elimination of lymphatic filariasis in Nepal.

BACKGROUND: Lymphatic filariasis (LF) is a neglected tropical disease transmitted by mosquitoes. Nepal has implemented a national effort to eliminate LF by 2020 through mass drug administration (MDA) using diethylcarbamazine (DEC) and albendazole (ALB). We assessed the impact of MDAs on LF in selected districts of Nepal after the recommended six MDA rounds had been completed. METHODOLOGY AND PRINCIPAL FINDINGS: Baseline surveys were conducted in seven districts and mapping data were used as baseline in the other three districts before starting MDA in 2009. LF antigen (Ag) prevalence ranged from 1.06% to 20% among districts included in the baseline and mapping study. The number of people who received DEC and ALB were recorded during each MDA round and population-based cluster surveys were conducted at least once in each district during the life of the program. The reported MDA coverage in five districts was consistently at least 65%. Two districts achieved the targeted coverage in four out of five rounds and the rest three districts achieved the target only in the first round. A pre-transmission assessment survey (pre-TAS) was conducted in one sentinel site and at least one spot check site in each of the districts after five MDA rounds. In pre-TAS, all the sites of five districts (Pyuthan, Arghakhanchi, Kaski, Bhaktapur, and Kathmandu) and all but one spot check site of Lalitpur district had LF Ag < 2% (ranging from 0.0% to 1.99%). Transmission assessment survey (TAS) was conducted in six evaluation units (EUs) consisting of six districts qualified on pre-TAS. Though MDA coverage of 65% was not achieved in three districts (Kathmandu, Lalitpur and Bhaktapur), Nepal government in consultation with World Health Organization (WHO) decided to conduct TAS. All six EUs achieved the LF Ag threshold required to stop MDA in TAS, despite the low reported MDA coverage in those three districts. CONCLUSIONS: Although Nepal has achieved significant progress towards LF elimination, five rounds of MDA were not sufficient to disrupt the transmission cycle in all districts, probably because of high baseline prevalence.

Interplay between Autophagy, Exosomes and HIV-1 Associated Neurological Disorders: New Insights for Diagnosis and Therapeutic Applications.

The autophagy-lysosomal pathway mediates a degradative process critical in the maintenance of cellular homeostasis as well as the preservation of proper organelle function by selective removal of damaged proteins and organelles. In some situations, cells remove unwanted or damaged proteins and RNAs through the release to the extracellular environment of exosomes. Since exosomes can be transferred from one cell to another, secretion of unwanted material to the extracellular environment in exosomes may have an impact, which can be beneficial or detrimental, in neighboring cells. Exosome secretion is under the influence of the autophagic system, and stimulation of autophagy can inhibit exosomal release and vice versa. Neurons are particularly vulnerable to degeneration, especially as the brain ages, and studies indicate that imbalances in genes regulating autophagy are a common feature of many neurodegenerative diseases. Cognitive and motor disease associated with severe dementia and neuronal damage is well-documented in the brains of HIV-infected individuals. Neurodegeneration seen in the brain in HIV-1 infection is associated with dysregulation of neuronal autophagy. In this paradigm, we herein provide an overview on the role of autophagy in HIV-associated neurodegenerative disease, focusing particularly on the effect of autophagy modulation on exosomal release of HIV particles and how this interplay impacts HIV infection in the brain. Specific autophagy-regulating agents are being considered for therapeutic treatment and prevention of a broad range of human diseases. Various therapeutic strategies for modulating specific stages of autophagy and the current state of drug development for this purpose are also evaluated.

Intranasal drug delivery of small interfering RNA targeting Beclin1 encapsulated with polyethylenimine (PEI) in mouse brain to achieve HIV attenuation.

We previously reported that activation of the host autophagic protein, Beclin1, by HIV-1 infection represents an essential mechanism in controlling HIV replication and viral-induced inflammatory responses in microglial cells. Existing antiretroviral therapeutic approaches have been limited in their ability to cross the blood-brain barrier effectively and recognize and selectively eliminate persistent HIV-infected brain reservoirs. In the present study and for the first time, the bio-distribution and efficacy of noninvasive intranasal delivery of small interfering RNA (siRNA) against the Beclin1 gene using the cationic linear polyethylenimines (PEI) as a gene carrier was investigated in adult mouse brain. Fluorescein isothiocyanate (FITC)-labeled control siRNA delivered intranasally was found in the cytoplasm of neurons and glial cells of the prefrontal cortex at 4 and 24 hours post-delivery, with no major adverse immune reaction encountered. Intranasal delivery of the siRNA targeting Beclin1 significantly depleted the target protein expression levels in brain tissues with no evidence of toxicity. Binding of siRNA to PEI-polymer was characterized and confirmed by Raman spectroscopy. These results indicate that the intranasal drug delivery allows for the direct delivery of the PEI-siRNA nano-complex to the central nervous system, which could potentially offer an efficient means of gene silencing-mediated therapy in the HIV-infected brain.

Overview on the Current Status of Zika Virus Pathogenesis and Animal Related Research.

There is growing evidence that Zika virus (ZIKV) infection is linked with activation of Guillan-Barré syndrome (GBS) in adults infected with the virus and microcephaly in infants following maternal infection. With the recent outpour in publications by numerous research labs, the association between microcephaly in newborns and ZIKV has become very apparent in which large numbers of viral particles were found in the central nervous tissue of an electively aborted microcephalic ZIKV-infected fetus. However, the underlying related mechanisms remain poorly understood. Thus, development of ZIKV-infected animal models are urgently required. The need to develop drugs and vaccines of high efficacy along with efficient diagnostic tools for ZIKV treatment and management raised the demand for a very selective animal model for exploring ZIKV pathogenesis and related mechanisms. In this review, we describe recent advances in animal models developed for studying ZIKV pathogenesis and evaluating potential interventions against human infection, including during pregnancy. The current research directions and the scientific challenges ahead in developing effective vaccines and therapeutics are also discussed.

Mammalian microRNA: an important modulator of host-pathogen interactions in human viral infections.

MicroRNAs (miRNAs), which are small non-coding RNAs expressed by almost all metazoans, have key roles in the regulation of cell differentiation, organism development and gene expression. Thousands of miRNAs regulating approximately 60 % of the total human genome have been identified. They regulate genetic expression either by direct cleavage or by translational repression of the target mRNAs recognized through partial complementary base pairing. The active and functional unit of miRNA is its complex with Argonaute proteins known as the microRNA-induced silencing complex (miRISC). De-regulated miRNA expression in the human cell may contribute to a diverse group of disorders including cancer, cardiovascular dysfunctions, liver damage, immunological dysfunction, metabolic syndromes and pathogenic infections. Current day studies have revealed that miRNAs are indeed a pivotal component of host-pathogen interactions and host immune responses toward microorganisms. miRNA is emerging as a tool for genetic study, therapeutic development and diagnosis for human pathogenic infections caused by viruses, bacteria, parasites and fungi. Many pathogens can exploit the host miRNA system for their own benefit such as surviving inside the host cell, replication, pathogenesis and bypassing some host immune barriers, while some express pathogen-encoded miRNA inside the host contributing to their replication, survival and/or latency. In this review, we discuss the role and significance of miRNA in relation to some pathogenic viruses.

Virological and Immunological Status of the People Living with HIV/AIDS Undergoing ART Treatment in Nepal.

Antiretroviral therapy (ART) has increased the life span of the people living with HIV (PLHIV), but their virological and immunological outcomes are not well documented in Nepal. The study was conducted at a tertiary care center including 826 HIV-1 seropositive individuals undergoing ART for at least six months. Plasma viral load (HIV-1 RNA) was detected by Real Time PCR and CD4(+) T-lymphocyte (CD4(+)) counts were estimated by flow cytometry. The mean CD4(+) count of patients was 501 (95% CI = 325-579) cells/cumm, but about 35% of patients had CD4(+) T cell counts below 350 cells/cumm. With increasing age, average CD4(+) count was found to be decreasing (p = 0.005). Of the total cases, 82 (9.92%) were found to have virological failure (viral load: >1000 copies/ml). Tenofovir/Lamivudine/Efavirenz (TDF/3TC/EFV), the frequently used ART regimen in Nepal, showed virological failure in 11.34% and immunological failure in 37.17% of patients. Virological failure rate was higher among children < 15 years (14.5%) (p = 0.03); however, no association was observed between ART outcomes and gender or route of transmission. The study suggests there are still some chances of virological and immunological failures despite the success of highly active ART (HAART).

Evaluation of SD Bioline HIV/syphilis Duo rapid test kits in Nepal.

BACKGROUND: Accurate and prompt diagnosis of HIV and syphilis simultaneously has reinforcing effect on their control program because of their prevalent co-infection. Availability of a simple user-friendly two-pronged and affordable detection tools brings down the cost of health care. They are important in the antenatal clinics, with added opportunity for intervention and prevention of mother to child transmission. In cooperation with rapid test kit manufacturers, SD Bioline, NPHL and NCASC, an evaluation of commercially available HIV/syphilis Duo rapid test kit (SD Bioline) to assess its performance and operational characteristics was done in the present study. METHOD: A prospective laboratory-based cross sectional study was conducted at a large Women's Hospital. Ten thousand pregnant women, visiting the Hospital for antenatal care or for delivery, were enrolled in study. Tests were performed by the SD Bioline HIV/Syphilis Duo kit as well as national algorithm for HIV and syphilis diagnosis which were considered gold standard. Sensitivity, Specificity, positive predictive value and negative predictive value along with kappa coefficient were calculated for the kit under evaluation. RESULT: The sensitivity, specificity, Negative predictive value and Positive predictive value of the kit for HIV diagnosis were 100 % (95 % CI 83.18-100 %, 99.96-100 %, 83.18-100 %, and 99.96-100 %, respectively). Kappa value was found to be 1.0. Out of total cases, results of 9985 (99.85 %) cases were concordant with National algorithm for syphilis diagnosis. Thirteen (0.13 %) cases were found false positive while two were false negative. The sensitivity of the kit for syphilis diagnosis was found to be 95.45 % (95 % CI 84.86-98.74 %) and specificity was 99.87 % (95 % CI; 99.78-99.92 %). Positive predictive value was 76.36 % (95 % CI; 63.65-85.63 %) and Negative predictive value was 99.89 % (95 % CI; 99.39-99.99 %). Kappa value was found to be 0.85. CONCLUSION: The performance characteristics of SD Bioline HIV/Syphilis duo kit were found almost concordant with the kits being used for HIV and Syphilis diagnosis separately. Its implementation in antenatal clinics/VCTs could be an added opportunity for simultaneous diagnosis of HIV and syphilis.

Lower respiratory tract infections among HIV positive and control group in Nepal.

Lower respiratory tract infections (LRTIs) are the most frequent respiratory diseases among HIV infected patients and are frequently the first clinical manifestations of the HIV infections. LRTIs are common not only among the HIV seropositive cases but also the commonest domiciliary and nosocomial infections among the general population. The present study was carried out to determine the comparative prevalence of common bacterial and fungal organism among the HIV positive and control population. This cross sectional study was conducted among 220 people attending National Public Health Laboratory, Kathmandu, Nepal. Out of them 120 were HIV sero-positive and rest were HIV sero-negative. Sputum samples were collected and processed soon after its collection. Macroscopic examination was done to determine the sample integrity. Gram stain, AFB stain and KOH preparation was performed for preliminary identification of the pathogens. Culture was carried out for bacterial and fungal pathogens. Antibiotic susceptibility test (Kirby-Bauer disc diffusion method) was performed from the isolated organisms. The 85 out of 120 HIV sero-positive patients were found to be infected with one or more microbial pathogens. The overall infection rate was found to be significantly lower in HIV seronegative people (27 %). Among HIV seropositive cases prevalence of LRTIs was strongly associated with lower CD4 counts (<200/mm(3)). The prevalence of mycobacterium tuberculosis was found to be 10 % among HIV/AIDS patients which was significantly higher than among the non-HIV cases (3 %). The bacterial pathogens was observed among 46.6 % of HIV positive and 22.0 % of HIV negative people. Among the positive cases, K. pneumoniae was the predominant bacterial pathogens, followed by E. coli and S. pneumoniae. C. albicans was found to be predominant fungal pathogen followed by Aspergillus spp. germ tube negative Candida spp. and Penicillium spp. Similar types of organisms were found to be associated with LRTIs among HIV positive and negative people. The prevalence of both fungal and bacterial infections was significantly higher among HIV seropositive people than HIV seronegative people. All in all, lower respiratory tract illness is significantly higher in HIV/AIDS cases than in HIV seronegative cases.