The crystal structure of the Hazara virus nucleocapsid protein

Background: Hazara virus (HAZV) is a member of the Bunyaviridae family of segmented negative stranded RNA viruses, and shares the same serogroup as Crimean-Congo haemorrhagic fever virus (CCHFV). CCHFV is responsible for fatal human disease with a mortality rate approaching 30 %, which has an increased recent incidence within southern Europe. There are no preventative or therapeutic treatments for CCHFV-mediated disease, and thus CCHFV is classified as a hazard group 4 pathogen. In contrast HAZV is not associated with serious human disease, although infection of interferon receptor knockout mice with either CCHFV or HAZV results in similar disease progression. To characterise further similarities between HAZV and CCHFV, and support the use of HAZV as a model for CCHFV infection, we investigated the structure of the HAZV nucleocapsid protein (N) and compared it to CCHFV N. N performs an essential role in the viral life cycle by encapsidating the viral RNA genome, and thus, N represents a potential therapeutic target. Results: We present the purification, crystallisation and crystal structure of HAZV N at 2.7 Å resolution. HAZV N was expressed as an N-terminal glutathione S-transferase (GST) fusion protein then purified using glutathione affinity chromatography followed by ion-exchange chromatography. HAZV N crystallised in the P212121 space group with unit cell parameters a = 64.99, b = 76.10, and c = 449.28 Å. HAZV N consists of a globular domain formed mostly of alpha helices derived from both the N- and C-termini, and an arm domain comprising two long alpha helices. HAZV N has a similar overall structure to CCHFV N, with their globular domains superposing with an RMSD = 0.70 Å, over 368 alpha carbons that share 59 % sequence identity. Four HAZV N monomers crystallised in the asymmetric unit, and their head-to-tail assembly reveals a potential interaction site between monomers. Conclusions: The crystal structure of HAZV N reveals a close similarity to CCHFV N, supporting the use of HAZV as a model for CCHFV. Structural similarity between the N proteins should facilitate study of the CCHFV and HAZV replication cycles without the necessity of working under containment level 4 (CL-4) conditions

Occurrence of blaNDM-1 & absence of blaKPC genes encoding carbapenem resistance in uropathogens from a tertiary care centre from north India

Background & objectives: Carbapenem resistance mediated by carbapenemases is increasingly being reported worldwide. This study was conducted to know the occurrence of important carbapenem resistance encoding genes in Gram-negative bacilli (GNB) causing complicated urinary tract infection (CUTI), and to look at the genetic diversity of these isolates. Methods: The study was carried out on 166 consecutive carbapenem resistant uropathogens (CRU) isolated from cases with CUTI during 2008 and 2012. Carbapenemase production was characterized phenotypically and polymerase chain reaction was used to detect bla VIM , bla IMP , bla KPC , and bla NDM-1 . BOX- PCR was done on 80 randomly selected isolates for molecular typing. Results: The bla VIM gene was present in 34 (43.6%), bla IMP in five (6.4%) and none of the isolates from 2008 had bla NDM-1 or bla KPC genes. Among the isolates from 2012, bla NDM-1 gene was present in 47 (53.4%), bla VIM in 19 (24.4%), bla IMP in one (1.1%) and none had bla KPC . There were nine isolates during the two years which had multiple genes encoding carbapenemases; while 66 did not have any of the genes tested. Of the 80 isolates subjected to BOX-PCR, 58 could be used for analysis and showed, presence of multiple clusters of carbapenem resistant isolates and absence of a single dominant clone. Interpretation & conclusions: The bla NDM-1 gene was absent in our isolates obtained during 2008 but was present amongst Enterobacteriaceae isolated in 2012. The bla KPC gene was also not found. Nine isolates obtained during the two years had multiple genes encoding carbapenemases confirming the previous reports of emergence of GNB containing genes encoding multiple carbapenemases. Typing using BOX-PCR indicated that this emergence was not because of clonal expansion of a single strain, and multiple strains were circulating at a single point of time

Viral markers in patients with hemophagocytosis: A prospective study in a tertiary care hospital

Background : Hemophagocytic syndrome (HPS) is a rare clinicopathological condition characterized by the activation of macrophages with prominent hemophagocytosis in bone marrow and other reticulo-endothelial systems. HPS can be familial or secondary to infections including viruses. Aim : To study the viral markers in patients with HPS. Materials and Methods : Serum samples of patients with HPS and control group were screened for anti EBV VCA IgM, and IgG, anti-Parvo B19 IgM, and anti-CMV IgM antibodies using commercially available ELISA kits and CMV and ParvoB19 DNA by polymerase chain reaction (PCR). Results and Discussion : The present prospective study reports the profile of viral markers in HPS cases from north India. Among the 14 HPS cases 43% (6/14) were positive for at least one viral marker tested, of which EBV was found to be the most prevalent (3/6: 50%) followed by parvovirus B19(2/6: 33%) and cytomegalovirus (1/6: 17%). Mortality was noted in 33% of virus associated HPS patients. Our study highlights the higher association of Epstein-Barr virus (EBV) with HPS as compared to other viruses along with higher rate of mortality in both parvovirus B 19 and EBV associated HPS

Hospital acquired urinary tract infection by multidrug-resistant Brevundimonas vesicularis

Infections caused by Brevundimonas vesicularis, a nonfermenting Gram-negative bacterium, are very rare. Here, we report the first case of multidrug-resistant hospital acquired urinary tract infection by B. vesicularis. Patient was successfully treated with antimicrobial therapy with piperacillin-tazobactam and amikacin