Mycotoxigenic infestation in samples of cereal straw from Bihar, India

A survey of different types of cereal straw samples viz. paddy, maize and wheat, from Bihar State, India, was conducted in order to examine the mould flora and mycotoxin contamination. Out of 170 samples examined for mould flora,Aspergillus flavus group of fungi had highest level of incidence followed byA niger. Isolates ofA flavus, A ochraceus, Fusarium verticillioides andPenicillium citrinum were screened for their mycotoxins producing abilities. Out of 75, 63 and 68 isolates ofA flavus group obtained from stored straw of paddy, maize and wheat samples, respectively, 27 (36%), 14 (22%) and 24 (35%) were found to be toxigenic which produced different combinations of aflatoxins in different concentrations. The percentage toxigenicity was comparatively lower in the isolates of other mycotoxigenic fungi from all types of samples. Out of 222 samples of straw analysed for natural occurrence of different mycotoxins, besides the aflatoxins present, zearalenone, ochratoxin A and citrinin were also recorded alone or as co-contaminants. A conducive climate together with the socioeconomic conditions of this region are important determinants for the high incidence of mycotoxins in cereal straw samples.     

Simultaneous Detection of Major Pome Fruit Viruses and a Viroid

A rapid and sensitive two-step RT-PCR protocol for simultaneous detection of major apple viruses, namely Apple mosaic virus (ApMV), Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV), Apple chlorotic leaf spot virus (ACLSV) and Apple scar skin viroid (ASSVd), was developed. Five specific primer pairs were tested and confirmed for these viruses and viroid together in a single tube, giving amplicons of ~198, ~330, ~370, ~547 and ~645 bp corresponding to ASGV, ASSVd, ASPV, ApMV and ACLSV, respectively. Using a guanidinium-based extraction buffer along with a commercial kit resulted in better quality RNA as compared to kit, suited for multiplex RT-PCR. A rapid CTAB method for RNA isolation from apple tissue was developed, which produce good yield and saves time. To the best of our knowledge, this is the first report on the simultaneous detection of five pathogens (four viruses and a viroid) from apple with NADH dehydrogenase subunit 5 (nad5) as an internal control.     

Acute Neuronal Injury and Blood Genomic Profiles in a Nonhuman Primate Model for Ischemic Stroke

The goal of this study was to characterize acute neuronal injury in a novel nonhuman primate (NHP) ischemic stroke model by using multiple outcome measures. Silk sutures were inserted into the M1 segment of the middle cerebral artery of rhesus macaques to achieve permanent occlusion of the vessel. The sutures were introduced via the femoral artery by using endovascular microcatheterization techniques. Within hours after middle cerebral artery occlusion (MCAO), infarction was detectable by using diffusion-weighted MRI imaging. The infarcts expanded by 24 h after MCAO and then were detectable on T2-weighted images. The infarcts seen by MRI were consistent with neuronal injury demonstrated histologically. Neurobehavioral function after MCAO was determined by using 2 neurologic testing scales. Neurologic assessments indicated that impairment after ischemia was limited to motor function in the contralateral arm; other neurologic and behavioral parameters were largely unaffected. We also used microarrays to examine gene expression profiles in peripheral blood mononuclear cells after MCAO-induced ischemia. Several genes were altered in a time-dependent manner after MCAO, suggesting that this ischemia model may be suitable for identifying blood biomarkers associated with the presence and severity of ischemia. This NHP stroke model likely will facilitate the elucidation of mechanisms associated with acute neuronal injury after ischemia. In addition, the ability to identify candidate blood biomarkers in NHP after ischemia may prompt the development of new strategies for the diagnosis and treatment of ischemic stroke in humans.Abbreviations: MCAO, middle cerebral artery occlusion; NHP, nonhuman primate; PBMC, peripheral blood mononuclear cellsStroke is a debilitating neurologic condition, and little progress has been made in the development of neuroprotective treatments for acute stroke. The Stroke Therapy Academic Industry Roundtable (STAIR) report suggested that preclinical candidates for stroke therapy should be validated by testing in large animals with similarities to humans, such as nonhuman primates (NHP).²⁶ NHP stroke models have been developed in several species, including rhesus monkeys, marmosets, and baboons, by using a variety of techniques for middle cerebral artery occlusion (MCAO).^{4,10,12,13,14,25,32} The rhesus macaque is ideal for stroke studies because of its structural similarities to human brain. The rhesus brain is gyrencephalic, which makes it preferable to those of lissencephalic primates (for example, marmosets) and is functionally similar to human brain.⁶ In addition, the immunologic profile of rhesus macaques is similar to that of humans; therefore these animals are the preferred model for the study of immune responses to infectious diseases such as HIV/SIV, Dengue virus, and others.^17,23,30In addition to their use for neuroprotection assessment, NHP stroke models can facilitate efforts to develop diagnostic tools for identifying and treating stroke symptoms. The use of genomics in peripheral blood cells has been shown to be an excellent method to identify candidate biomarkers and cellular mechanisms associated with stroke.^28,29 Blood biomarkers can be used to rapidly determine the occurrence, timing, subtype, and severity of stroke.^11,15 One possible reason for the lack of viable stroke biomarkers may be the research models used to search for these markers. Although rodent stroke models have yielded a wealth of information on the mechanisms associated with brain ischemia, the findings have not translated well to human clinical trials.²⁶ Recent studies in human patients showed promising results when genomic tools have been used to screen for novel stroke biomarkers.^3,16,27 However, validation of human studies is limited by the need for large data sets in light of heterogeneity in stroke onset, subtype, comorbidities, and other factors. In addition, it is also impossible to know the exact time of stroke onset in most patients.Here we characterized acute neuronal injury in a novel, minimally invasive permanent ischemic stroke model involving rhesus macaques. Using endovascular catheterization techniques, we introduced silk sutures into the M1 segment of the middle cerebral artery and permanently occluded it. This procedure reliably produced infarcts that could be measured by MRI of the macaque brains during the acute phase period. The procedure resulted in discrete and limited neurobehavioral deficits, indicating the potential of this stroke model for chronic neuroprotection studies in the future. In addition, we used microarrays to identify blood genomic profiles that were altered in a time-dependent manner after ischemia. These studies characterize a preclinical model that is suitable for elucidating the mechanisms associated with cerebral ischemia and that may aid in identifying strategies for the diagnosis and treatment of stroke in humans.     

Genetic structure of four socio-culturally diversified caste populations of southwest India and their affinity with related Indian and global groups

Background  

A large number of microsatellites have been extensively used to comprehend the genetic diversity of different global groups. This paper entails polymorphism at 15 STR in four predominant and endogamous populations representing Karnataka, located on the southwest coast of India. The populations residing in this region are believed to have received gene flow from south Indian populations and world migrants, hence, we carried out a detailed study on populations inhabiting this region to understand their genetic structure, diversity related to geography and linguistic affiliation and relatedness to other Indian and global migrant populations.     

Phylogenetic analysis of cubilin (CUBN) gene

Cubilin, (CUBN; also known as intrinsic factor-cobalamin receptor [Homo sapiens Entrez Pubmed ref {"type":"entrez-nucleotide","attrs":{"text":"NM_001081.3","term_id":"126091151","term_text":"NM_001081.3"}}NM_001081.3; {"type":"entrez-nucleotide","attrs":{"text":"NG_008967.1","term_id":"212549718","term_text":"NG_008967.1"}}NG_008967.1; GI: 119606627]), located in the epithelium of intestine and kidney acts as a receptor for intrinsic factor – vitamin B12 complexes. Mutations in CUBN may play a role in autosomal recessive megaloblastic anemia. The current study investigated the possible role of CUBN in evolution using phylogenetic testing. A total of 588 BLAST hits were found for the cubilin query sequence and these hits showed putative conserved domain, CUB superfamily (as on 27^th Nov 2012). A first-pass phylogenetic tree was constructed to identify the taxa which most often contained the CUBN sequences. Following this, we narrowed down the search by manually deleting sequences which were not CUBN. A repeat phylogenetic analysis of 25 taxa was performed using PhyML, RAxML and TreeDyn softwares to confirm that CUBN is a conserved protein emphasizing its importance as an extracellular domain and being present in proteins mostly known to be involved in development in many chordate taxa but not found in prokaryotes, plants and yeast.. No horizontal gene transfers have been found between different taxa.     

Detection of Prunus necrotic ring spot virus in plum,cherry and almond by serological and molecular techniques from India

Stone fruits are cultivated in the temperate and sub-temperate regions of India. During surveys in stone fruit growing areas, viral symptoms were observed in almond, cherry and plum. These samples were brought to the laboratory for further detection at serological and molecular levels to check the presence of virus. In the present study, incidence of PNRSV is reported on plum (Prunus domestica), almond (Prunus dulcis) and cherry (Prunus avium) using serological and molecular techniques. Coat protein gene of PNRSV was amplified from almond, cherry and plum. This is the first molecular evidence of PNRSV on these stone fruits reported from India.