In vitro replicative fitness of early Transmitted founder HIV-1 variants and sensitivity to Interferon alpha.

Type I interferons, particularly interferon-alpha (IFN-α), play a vital role in the host's anti-viral defenses by interfering with viral replication. However, the virus rapidly evolves to exploit the IFN-α response for its replication, spread, and pathogenic function. In this study, we attempted to determine IFN-α susceptibility and productivity of infectious transmitted/founder (TF) (n = 8) and non-transmitted (NT) viruses (n = 8) derived from HIV-1 infected infants. Independent experiments were carried out to determine IFN-α resistance, replication fitness, and viral productivity in CD4+ T cells over a short period. In vitro studies showed that TF viruses were resistant to IFN-α during the very near moment of transmission, but in the subsequent time points, they became susceptible to IFN-α. We did not observe much difference in replicative fitness of the TF viruses in cultures treated with and without IFN-α, but the difference was significant in the case of NT viruses obtained from the same individual. Despite increased susceptibility to IFN-α, NT viruses produced more viral particles than TF viruses. Similar results were also obtained in cultures treated with maraviroc (MVC). The study identified unique characteristics of TF viruses thus prompting further investigation into virus-host interaction occurring during the early stages of HIV infection.

Current trends in tuberculosis vaccine.

Despite the global efforts made to control tuberculosis (TB) and the large number of available new anti-TB drugs, TB still affects one-third of the world population. The conventional vaccine bacille Calmette-Guérin (BCG) shows varying efficacy in different populations, and there are safety issues in immunocompromised patients. Hence, there is an urgent requirement for a new and better TB vaccine candidate than BCG. There are several alternate vaccines available for TB such as DNA, subunit, adjuvant, and live-attenuated vaccines. Use of auxotrophic vaccine is an emerging technology. Newer vaccine technologies include vaccine delivery methods such as adenovirus- and cytomegalovirus (CMV)-based vector delivery, chimeric monoclonal antibody, single-chain fragment variable, RNA-lipoplexes, and nanoparticle-based technology. Based on its application, TB vaccines are classified as conventional, prophylactic, booster, therapeutic, and reinfection preventive vaccines. Currently, there are 12 vaccine candidates in clinical trials. In this review, we have briefly discussed about each of these vaccines in different phases of clinical trials. These vaccines should be analyzed further for developing a safe and more efficacious vaccine for TB.

Synthesis and biological evaluation of 1-benzyl-N-(2-(phenylamino)pyridin-3-yl)-1H-1,2,3-triazole-4-carboxamides as antimitotic agents.

A library of 1-benzyl-N-(2-(phenylamino)pyridin-3-yl)-1H-1,2,3-triazole-4-carboxamides (7a-al) have been designed, synthesized and screened for their anti-proliferative activity against some selected human cancer cell lines namely DU-145, A-549, MCF-7 and HeLa. Most of them have shown promising cytotoxicity against lung cancer cell line (A549), amongst them 7f was found to be the most potent anti-proliferative congener. Furthermore, 7f exhibited comparable tubulin polymerization inhibition (IC50 value 2.04 µM) to the standard E7010 (IC50 value 2.15 µM). Moreover, flow cytometric analysis revealed that this compound induced apoptosis via cell cycle arrest at G2/M phase in A549 cells. Induction of apoptosis was further observed by examining the mitochondrial membrane potential and was also confirmed by Hoechst staining as well as Annexin V-FITC assays. Furthermore, molecular docking studies indicated that compound 7f binds to the colchicine binding site of the ß-tubulin. Thus, 7f exhibits anti-proliferative properties by inhibiting the tubulin polymerization through the binding at the colchicine active site and by induction of apoptosis.

2-Anilino-3-Aroylquinolines as Potent Tubulin Polymerization Inhibitors.

Several 2-anilino-3-aroylquinolines were designed, synthesized, and screened for their cytotoxic activity against five human cancer cell lines: HeLa, DU-145, A549, MDA-MB-231, and MCF-7. Their IC50 values ranged from 0.77 to 23.6 µm. Among the series, compounds 7 f [(4-fluorophenyl)(2-((4-fluorophenyl)amino)quinolin-3-yl)methanone] and 7 g [(4-chlorophenyl)(2-((4-fluorophenyl)amino)quinolin-3-yl)methanone] showed remarkable antiproliferative activity against human lung cancer and prostate cancer cell lines. The IC50 values for inhibiting tubulin polymerization were 2.24 and 2.10 µm for compounds 7 f and 7 g, respectively, and were much lower than that of the reference compound E7010 [N-(2-(4-hydroxyphenylamino)pyridin-3-yl)-4-methoxybenzenesulfonamide]. Furthermore, flow cytometric analysis revealed that these compounds arrest the cell cycle at the G2 /M phase, leading to apoptosis. Apoptosis was also confirmed by mitochondrial membrane potential, Annexin V-FITC assay, and intracellular ROS generation. Immunohistochemistry, western blot, and tubulin polymerization assays showed that these compounds disrupt tubulin polymerization. Molecular docking studies revealed that these compounds bind efficiently to ß-tubulin at the colchicine binding site.

Combretastatin linked 1,3,4-oxadiazole conjugates as a Potent Tubulin Polymerization inhibitors.

A new class of combretastatin linked 1,3,4-oxadiazoles were designed, synthesized and screened for their cytotoxic activity against five human cancer cell lines such as HeLa, DU-145, A549, MDA-MB-231 and B16. These compounds showed significant cytotoxicity with IC50 values in the range 0.118-54.32µM. Conjugate 5m displayed potent antiproliferative activity against DU-145 cell line. Flow cytometric analysis revealed that these compounds arrested the cell cycle in G2/M phase. Moreover, the tubulin polymerization assay and immunofluorescence analysis indicate that 5m exhibits potent inhibitory effect on the tubulin assembly. Further, DNA fragmentation and Hoecst staining assays confirm that 5m induces apoptosis. Molecular docking studies and competitive binding assay indicated that 5m effectively bind at the colchicine binding site of the tubulin.

Associations of total and free 25OHD and 1,25(OH)2D with serum markers of inflammation in older men.

UNLABELLED: Vitamin D is hypothesized to suppress inflammation. We tested total and free vitamin D metabolites and their association with inflammatory markers. Interleukin-6 levels were lower with higher 25-hydroxyvitamin D. 1,25-dihydroxyvitamin D and free 25OHD associations mirrored those of 25OHD. However, associations for the two metabolites diverged for tumor necrosis factor alpha (TNF-α) soluble receptors. INTRODUCTION: Vitamin D is hypothesized to suppress inflammation, and circulating 25-hydroxyvitamin D (25OHD) and inflammatory markers are inversely correlated. However, total serum 25OHD may not be the best indicator of biologically active vitamin D. METHODS: We tested serum total 25OHD, total 1,25(OH)2D, vitamin D binding protein (DBP), and estimated free 25OHD and free 1,25(OH)2D associations with inflammatory markers serum interleukin-6 (IL-6), TNF-α and their soluble receptors, interleukin-10 (IL-10), and C-reactive protein (CRP) as continuous outcomes and the presence of ≥2 inflammatory markers in the highest quartile as a dichotomous outcome, in a random subcohort of 679 men in the Osteoporotic Fractures in Men (MrOS) study. RESULTS: IL-6 was lower in men with higher 25OHD (-0.23 µg/mL per standard deviation (SD) increase in 25OHD, 95 % confidence intervals (CI) -0.07 to -0.38 µg/mL) and with higher 1,25(OH)2D (-0.20 µg/mL, 95 % CI -0.0004 to -0.39 µg/mL); free D associations were slightly stronger. 25OHD and DBP, but not 1,25(OH)2D, were independently associated with IL-6. TNF-α soluble receptors were inversely associated with 1,25(OH)2D but positively associated with 25OHD, and each had independent effects. The strongest association with ≥2 inflammatory markers in the highest quartile was for free 1,25(OH)2D (odds ratios (OR) 0.70, 95 % CI 0.54 to 0.89 per SD increase in free 1,25(OH)2D). CONCLUSIONS: Associations of 1,25(OH)2D and free 25OHD with IL-6 mirrored those of 25OHD, suggesting that 1,25(OH)2D and free D do not improve upon 25OHD in population-based IL-6 studies. However, associations for the two metabolites diverged for TNF-α soluble receptor, warranting examination of both metabolites in studies of TNF-α and its antagonists.

Outcome of patients on second line antiretroviral therapy under programmatic condition in India.

BACKGROUND: The National AIDS Control Organization of India has been providing free second line antiretroviral therapy (ART) since 2008. This observational study reports the survival and virologic suppression of patients on second-line ART under programmatic condition and type of mutations acquired by those failing therapy. METHODS: 170 patients initiated on second-line therapy between 2008 and 2012 were followed up till 2013. Viral Load (VL) was repeated at 6 months for all patients and at 12 months for those with VL >400 copies/ml at 6 months. Adequate virological response was defined as plasma HIV-1 VL <400 copies/ml and virological failure was defined as VL >1000 copies/ml. Genotyping was done in 16 patients with virological failure. RESULTS: Out of 170 patients, 110 (64.7 %) were alive and on therapy and 35 (20.5 %) expired. In the first year the occurrence of death was 13.7 /100 person years while between 1 and 5 year it was 3.88 /100 person years. In the first year, duration of immunological failure >12 months, weight <45 kg, WHO clinical stage 3 and 4 and WHO criteria CD4 count less than pretherapy baseline [hazard ratio HR 4.2. 15.8, 11.9 & 4.1 respectively] and beyond first year poor first and second line adherence and first line CD4 count < 200/µL [HR 5.2,15.8, 3.3 respectively] had high risk of death. 119/152 (78.2 %) had adequate virological response and 27/152 (17.7 %) had virological failure. High viral load at baseline and poor second line adherence (Odds Ratio 3.4 & 2.8 respectively) had increased risk of virological failure. Among those genotyped, 50 % had major Protease Inhibitor mutation (M46I commonest) however 87.5 % were still susceptible to darunavir. CONCLUSIONS: Second line therapy has shown high early mortality but good virological suppression under programmatic conditions.

An unusual complication of a common gynaecological procedure.

A case of bilateral, but more of massive right sided transudative pleural effusion associated with bilateral ureteric trauma following laparoscopy for endometriosis is reported. The diagnosis of urinothorax was confirmed by demonstrating a pleural fluid to serum creatinine ratio of greater than one. Management of ureteric injury by insertion of Double J (DJ) stents on both sides resulted in resolution of the pleural effusion.

Initial virologic response and HIV drug resistance among HIV-infected individuals initiating first-line antiretroviral therapy at 2 clinics in Chennai and Mumbai, India.

Human immunodeficiency virus drug resistance (HIVDR) in cohorts of patients initiating antiretroviral therapy (ART) at clinics in Chennai and Mumbai, India, was assessed following World Health Organization (WHO) guidelines. Twelve months after ART initiation, 75% and 64.6% of participants at the Chennai and Mumbai clinics, respectively, achieved viral load suppression of <1000 copies/mL (HIVDR prevention). HIVDR at initiation of ART (P <.05) and 12-month CD4 cell counts <200 cells/µL (P <.05) were associated with HIVDR at 12 months. HIVDR prevention exceeded WHO guidelines (≥ 70%) at the Chennai clinic but was below the target in Mumbai due to high rates of loss to follow-up. Findings highlight the need for defaulter tracing and scale-up of routine viral load testing to identify patients failing first-line ART.

Molecular characterization of Chikungunya virus during an outbreak in South India.

INTRODUCTION: Re-emergence of Chikungunya is a major public health problem in the southern states of India. OBJECTIVES: This study was undertaken to investigate an outbreak of Chikungunya, in June-August 2008 using PCR and determine the prevalent genotypes of Chikungunya virus (CHIKV) associated with the outbreak. MATERIALS AND METHODS: Samples of blood were collected (in heparinized vacutainer tubes) from suspected patients of CHIKV infection from both Government Taluk Hospital in Kerala and a tertiary care hospital in Chennai, Tamil Nadu. A one-step RT-PCR was carried out on a block thermo-cycler targeting the E2 gene that codes for the viral envelope protein. The amplicons were verified for 305 bp size by standard agarose gel electrophoresis. The PCR products were purified, sequenced, and compared with other CHIKV strains reported from different geographical regions. A phylogenetic tree was constructed using MEGA 4. RESULTS: Altogether 118 samples were collected from patients who presented with sudden onset of fever and/or joint pain, myalgia, and headache. CHIKV infection was confirmed by RT-PCR in 14 patients and all these cases were from Kerala. The positivity correlated with the early stage of the disease as all these patients had fever of less than seven days duration. The study isolates have been allotted the GenBank accession nos. GQ272368-GQ272381. Phylogenetic analysis of recent CHIKV isolates by partial sequencing of E2 region shows that isolates are closely related to strains from neighboring states and the African type. CONCLUSION: RT-PCR is a useful technique for the early detection of CHIKV infection during outbreaks. Molecular characterization of the strains indicates that majority of the strains have originated from the Central/East African strains of CHIKV.

An association of Alternaria alternata and Scopulariopsis brevicaulis in cutaneous phaeohyphomycosis.

Rare molds are increasingly emerging as a cause of deep and invasive fungal infections. We report here a rare case of cutaneous phaeohyphomycosis of the lower limbs due to Alternaria alternata associated with extra-ungual localization of Scopulariopsis brevicaulis. Diagnosis was made based on repeated, direct, microscopic mycological and histological examinations. The study revealed hyphae and fungal cells in a granulomatous dermal infiltrate. Identification of the molds was based on macroscopic appearance on culture of samples from the lesions on Sabouraud's dextrose agar and microscopic appearance on Lactophenol cotton blue following slide culture.

Dysregulation of proinflammatory and regulatory cytokines in HIV infected persons with active tuberculosis.

Proinflammatory and regulatory cytokines have been implicated to play important role in immunopathology of HIV and tuberculosis (TB) infection. Capacity of unstimulated and mitogen-stimulated peripheral blood mononuclear cells (PBMCs) to secrete cytokines (interleukin (IL)-2, interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), IL-4, IL-10 and IL-6) was estimated for 15 HIV-TB coinfected patients, 22 HIV seropositives without TB, 32 HIV negative TB patients, and 36 healthy subjects. Dually infected patients had suppression of both Th1 and Th2 cytokine secretion as evidenced by significantly lower production of IL-2, IFN-gamma and TNF-alpha as well as IL-4 and IL-10. Production of IL-2 and TNF-alpha was significantly decreased only in case of HIV infection. Significantly higher IL-6 secretion was found in unstimulated cultures in dually infected patients. The mitogen induced cytokine secretion was generally lower in HIV-TB coinfected patients indicating profound perturbation of both Th1 and Th2 responses.

Superantigens - concepts, clinical disease and therapy.

The term superantigen was introduced in microbiology and immunology only a decade ago. The unique feature of superantigen is that it bypasses the antigen processing mechanism and specifically binds to TCR v segment and forms a trimolecular complex along with major histocompatibility complex class II. Since its discovery, several studies have been carried out to unravel the properties of superantigens. They are implicated in the pathogenesis of diseases of bacterial, viral and fungal origin. They are also known to play a role in autoimmune diseases. Therapy for these superantigens is being worked upon. Studies have shown that intravenous gamma globulin containing specific antibodies to these superantigens inhibits the activation of T-Cells and also the cytokine production by them resulting in dramatic recovery from superantigen mediated diseases. The use of superantigen in the therapy of cancer is being explored.

Low-dose valaciclovir prophylaxis against cytomegalovirus disease in renal transplant recipients.

High-dose valaciclovir at up to 8 g/day has been shown to be effective in prophylaxis against cytomegalovirus (CMV) disease in renal transplant recipients. We report our experience with low-dose valaciclovir prophylaxis of up to 3 g/day, adjusted to creatinine clearance. A group of patients at high risk of developing CMV disease who received prophylaxis were selected as the study group. This included all CMV-positive patients who received antilymphocyte therapy (R+, n=20) and all CMV-negative recipients of CMV-positive organs (D+R-, n=15). D+R- patients receiving antilymphocyte therapy were excluded, as most of the patients in the control group had received ganciclovir prophylaxis. A historical control group was used, which consisted of patients who did not receive prophylaxis. Low-dose valaciclovir prophylaxis resulted in a statistically significant decrease (8.5 vs 37%, P=0.004) in CMV disease in the study group at 6 months. On subgroup analysis the decrease was statistically significant only in the R+ group (5 vs 45%, P=0.003), not in the D+R- group (13.3 vs 26.6%, P=0.651). Low-dose valaciclovir prophylaxis seems to be adequate for R+ patients receiving antilymphocyte therapy. The role of low-dose valaciclovir prophylaxis needs to be assessed further in a prospective trial.

Consistent subtype-specific anti-HIV type 1 T lymphocyte responses in Indian subjects recently infected with HIV type 1.

Anti-HIV-1-specific T cell responses in early HIV-1 infection have been found to be important in deciding the course of disease progression. But there are few data concerning nonsubtype B HIV infection. HIV-1 subtype C is the most prevalent subtype in India. HIV-1 Gag-specific T cell responses in 12 Indian subjects with recent HIV-1 infection were characterized by an ELISpot assay at two consecutive visits and their correlation with plasma viral load and CD4(+) T lymphocyte counts was studied. Ten of the 12 subjects demonstrated T cell responses to either one or both Gag B and C peptides, on at least one visit. Five of 10 responders showed a consistent response (response at both visits): 4 exhibited a Gag C-specific consistent response and 1 showed a consistent response to Gag B. The remaining five patients, showing response at only one of the two visits, were considered inconsistent responders. None of the individuals showed a consistent response to both B and C Gag peptides. Marginally significant correlation was observed between consistency of the response and lower plasma viral load (p = 0.062). The subtype-specific Gag C response was also found to be correlated with lower viral load as compared with the response to Gag B (r = -0.336, p = 0.054 for subtype C and r = -0.234, p = 0.13 for subtype B). The data suggest that the patients exhibiting consistent subtype-specific responses to HIV-1 Gag might have better control of viral replication in early HIV infection.

Role of surface swab, core swab and fine needle aspiration in isolating the core bacteria in inflammed tonsils.

The microbial flora of 75 patients with chronic tonsillitis were sudied to identify the commonest organism causing chronic tonsillitis and to compare surface swab, fine needle aspiration (FNA) and core swab in isolating the pathogens. Staphylococcus aureus was the predominant isolate from surface core swab FNA and surface swab. In our study the correlation between surface swab with core swab was 47.9% and between surface swab with FNA 57.3%. The correlation between FNA with core swab of 69.3% showed that FNA for culture was superior to surface swab in predicting the core culture. Hence FNA of the tonsil has more advantage over the routinely performed surface swab in representing the core pathogen. Thus the rationale of treating chronic tonsillitis medically should be based on the knowledge of the common core pathogen.