Gut-organ axis: a microbial outreach and networking.

Human gut microbiota (GM) includes a complex and dynamic population of microorganisms that are crucial for well-being and survival of the organism. It has been reported as diverse and relatively stable with shared core microbiota, including Bacteroidetes and Firmicutes as the major dominants. They are the key regulators of body homeostasis, involving both intestinal and extra-intestinal effects by influencing many physiological functions such as metabolism, maintenance of barrier homeostasis, inflammation and hematopoiesis. Any alteration in GM community structures not only trigger gut disorders but also influence other organs and cause associated diseases. In recent past, the GM has been defined as a 'vital organ' with its involvement with other organs; thus, establishing a link or a bi- or multidirectional communication axis between the organs via neural, endocrine, immune, humoral and metabolic pathways. Alterations in GM have been linked to several diseases known to humans; although the exact interaction mechanism between the gut and the organs is yet to be defined. In this review, the bidirectional relationship between the gut and the vital human organs was envisaged and discussed under several headings. Furthermore, several disease symptoms were also revisited to redefine the communication network between the gut microbes and the associated organs.

A practical criterion for focusing of unstained cell samples using a digital holographic microscope.

Digital holographic microscopy (DHM) is an important technique that may be used for quantitative phase imaging of unstained biological cell samples. Since the DHM technology is not commonly used in clinics or bioscience research labs, at present there is no well-accepted focusing criterion for unstained samples that users can follow while recording image plane digital holograms of cells. The usual sharpness metrics that are useful for auto-focusing of stained cells do not work well for unstained cells as there is no amplitude contrast. In this work, we report a practical method for estimating the best focus plane for unstained cells in the digital hologram domain. The method is based on an interesting observation that for the best focus plane the fringe pattern associated with individual unstained cells predominantly shows phase modulation effect in the form of bending of fringes and minimal amplitude modulation. This criterion when applied to unstained red blood cells shows that the central dip in the doughnut-like phase profile of cells is maximal in this plane. The proposed methodology is helpful for standardizing the usage of DHM technology across different users and application development efforts. LAY DESCRIPTION: Digital holographic microscopy (DHM) is slowly but steadily becoming an important microscopy modality and gaining acceptability for basic bio-science research as well as clinical usage. One of the important features of DHM is that it allows users to perform quantitative imaging of unstained transparent cells. Instead of using dyes or fluorescent labelling, DHM systems use quantitative phase as a contrast mechanism which depends on the natural refractive index variation within the cell samples. Since minimal wet lab processing is required in order to image cell samples with a DHM, cells can be imaged in their natural state. While DHM is gaining popularity among users, the imaging protocols across the labs or users need to be standardized in order to make sure that the same quantitative phase parameters are used for tasks such as quantitative phased based cell classification. One of the important operational tasks for any microscopy work is to focus the sample under study. While focusing comes naturally to users of brightfield microscopes based on image contrast, the focusing is not straightforward when samples are unstained so that they do not offer any amplitude contrast. When performing quantitative phase imaging, defocus can actually change the phase profile of the cell due to near-zone (Fresnel) diffraction effects. So unless a standardized focusing methodology is used, it will be difficult for multiple DHM users (potentially at different sites) to agree on quantitative results out of their phase images. DHM literature has prior works which perform numerical focusing of recovered complex wave-field in the hologram plane to find the best focus plane. However such methods are not user friendly and do not allow user the same focusing experience as in a brightfield microscope. The numerical focusing is therefore a reasonably good method for an optics researcher but not necessarily so for a microscopy technician looking at cell samples with a DHM system in a clinical setting. The present work provides a simple focusing criterion for unstained samples that works directly in the hologram domain. The technique is based on an interesting observation that the when an unstained cell sample is in the best-focus plane, its corresponding hologram (or fringe pattern) predominantly shows phase modulation manifested by bending of fringes at the location of the cell. This criterion can be converted into a simple numerical method as we have used to find the best-focus plane using a stack of through focus holograms. We believe that the technique can be used manually by visually observing the holograms or can be converted to an auto-focus algorithm for a motorized DHM system.

Multiparametric imaging-based differentiation of lymphoma and glioblastoma: using T1-perfusion, diffusion, and susceptibility-weighted MRI.

AIM: To compare the diagnostic performance of T1 perfusion magnetic resonance imaging (MRI), diffusion-weighted imaging (DWI), and susceptibility-weighted imaging (SWI) for differentiating primary central nervous system lymphoma (PCNSL) and glioblastoma (GBM). MATERIALS AND METHODS: This retrospective study comprised a cohort of 70 patients with glioblastoma and 30 patients with PCNSL. T1 perfusion MRI-derived rCBV_corr (leakage corrected relative cerebral blood volume), apparent diffusion coefficient (ADC) derived from DWI, and intratumoural susceptibility signals intensity (ITSS) measured on SWI were evaluated in these 100 patients. The Mann-Whitney U-test was used for pairwise comparison between groups. The diagnostic performance for differentiating PCNSL from glioblastoma was evaluated by using univariate and multivariable logistic regression analyses and receiver operating characteristic (ROC) analysis. RESULTS: Minimum ADC, maximum rCBVs_corr, kep (back flux exchange rate), and ITSS scores were significantly lower in patients with PCNSL than in those with glioblastoma (p<0.05). On ROC analysis, ITSS showed the best discrimination ability for differentiation of GBM and PCNSL with an area under the ROC curve (AUC) of 0.80. rCBV_corr and ADC showed AUCs of 0.68 and 0.63, respectively. Multiparametric assessment using ADC, rCBV_corr, kep, and ITSS scores significantly increased the diagnostic ability for differentiating PCNSL from GBM as compared to mean ADC, mean rCBV_corr, and ITSS alone or a combination of these parameters. The multiparametric model could correctly discriminate 84% of tumours with a sensitivity and specificity of 90% and 70% with an AUC of 0.92. CONCLUSION: Multiparametric MRI evaluation using DWI, T1 perfusion MRI, and SWI enabled reliable differentiation of PCNSL and GBM in the majority patients, and these results support an integration of advanced MRI techniques for the diagnostic work-up of patients with these tumours.

Differential promoter methylation of DAZL gene in bulls with varying seminal parameters.

In India, cross-breeding of indigenous cattle with exotic cattle such as Holstein Friesian and Jersey has been going on since last four decades to improve milk production. Although it has led to increased milk yield, the subfertility in male cross-bred progeny has remained a significant problem. Epigenetic modifications (DNA methylation, histone modifications and chromatin remodelling) are regarded as key players influencing gene expression. DAZL gene plays an important role in germline development and gametogenesis. The methylation and mRNA expression level of this gene have been significantly negatively correlated in the testes of cattle-yak hybrids and their parents. This study analysed the methylation profile of DAZL gene promoter in bull spermatozoa in an attempt to speculate its role in cross-bred cattle subfertility. Semen samples from Sahiwal, Holstein Friesian and Frieswal bulls (Sahiwal X Holstein Friesian) with varying semen motility parameters were collected, and DNA was isolated. Methylation-specific primers were used to amplify part of promoter and exon 1 of DAZL gene using bisulphite-converted DNA. The amplified products were sequenced after cloning in pTZ57R/T vector. Sequence analysis revealed significantly higher DNA methylation of DAZL gene in Frieswal bulls with poor motility (28.26%) as compared to medium (15.21%) and high motility phenotype (6.52%). In pure-bred counterparts, Sahiwal and Holstein Friesian, epigenetic marks were more in the former (15.21%) than the latter (4.34%), but in both cases, the values were lower as compared to the poor motility Frieswal bulls. This suggests that differential hypermethylation of the CpG islands could possibly influence reproductive parameters in bovines.

Analysis of genetic relatedness among Indian cattle (Bos indicus) using genotyping-by-sequencing markers.

Genetic relatedness of 24 animals belonging to seven Indian cattle breeds was studied using high throughput genotyping-by-sequencing (GBS) markers. GBS produced 93.6 million reads with an average of about 3.9 million reads per animal. A total of 107 488 SNPs were identified in these individuals. When only one SNP per read was considered, a total of 60 261 SNPs representing independent reads were identified with an average SNP-to-SNP distance of 45 kb across the bovine reference genome. About 24% of the GBS-SNP markers were more than 100 kb apart. Of these, 58 322 SNPs mapped to autosomes, 1645 to the X chromosome and 28 to the Y chromosome. The average SNP-to-SNP distance on the X chromosome was 91.3 kb, whereas on the Y chromosome it was 1546.4 kb. The minor allele frequency within the Indian cattle varied from 0.103 (Ongole) to 0.177 (Siri), whereas Holstein cattle had the lowest value of 0.089. This is the first application of GBS in cattle of South Asia. The baseline information generated in this study might prompt implementation of GBS in breeding of cattle belonging to this region.

MDCT of hand and wrist infections: emphasis on compartmental anatomy.

Hand and wrist infections can present with a spectrum of manifestations ranging from cellulitis to deep-space collections. The various infectious processes can be categorised as superficial or deep infections based on their respective locations relative to the tendons. Superficial hand infections are located superficial to the tendons and are comprised of cellulitis, lymphangitis, paronychia, pulp-space infections, herpetic whitlow, and include volar as well as dorsal subcutaneous abscesses. Deep hand infections are located deep to the tendon sheaths and include synovial space infections, such as infectious tenosynovitis, deep fascial space infections, septic arthritis, necrotising fasciitis, and osteomyelitis. Knowledge of hand and wrist compartmental anatomy is essential for the accurate diagnosis and management of hand infections. Although early and superficial infections of the hand may respond to non-surgical management, most hand infections are surgical emergencies. Multidetector computed tomography (MDCT), with its muliplanar reformation (MPR) and three-dimensional (3D) capabilities, is a powerful tool in the emergency setting for the evaluation of acute hand and wrist pathology. The clinical and imaging features of hand and wrist infections as evident on MDCT will be reviewed with emphasis on contiguous and closed synovial and deep fascial spaces. Knowledge of hand compartmental anatomy enables accurate characterisation of the infectious process and localise the extent of disease in the acute setting.

Evidence of positive selection and concerted evolution in the rapidly evolving PRDM9 zinc finger domain in goats and sheep.

Meiotic recombination contributes to augmentation of genetic diversity, exclusion of deleterious alleles and proper segregation of chromatids. PRDM9 has been identified as the gene responsible for specifying the location of recombination hotspots during meiosis and is also the only known vertebrate gene associated with reproductive isolation between species. PRDM9 encodes a protein with a highly variable zinc finger (ZF) domain that varies between as well as within species. In the present study, the ZF domain of PRDM9 on chromosome 1 was characterized for the first time in 15 goat breeds and 25 sheep breeds of India. A remarkable variation in the number and sequence of ZF domains was observed. The number of ZF repeats in the ZF array varied from eight to 12 yielding five homozygous and 10 heterozygous genotypes. The number of different ZF domains was 84 and 52 producing 36 and 26 unique alleles in goats and sheep respectively. The posterior mean of dN/dS or omega values were calculated using the codeml tool of pamlx to identify amino acids that are evolving positively in goats and sheep, as positions -1, +3 and +6 in the ZF domain have been reported to experience strong positive selection across different lineages. Our study identified sites -5, -1, +3, +4 and +6 to be experiencing positive selection. Small ruminant zinc fingers were also found to be evolving under concerted evolution. Our results demonstrate the existence of a vast diversity of PRDM9 in goats and sheep, which is in concert with reports in many metazoans.

Exon skipping creates novel splice variants of DMC1 gene in ruminants.

Disrupted meiotic cDNA1 (DMC1) recombinase plays a pivotal role in homology search and strand exchange reactions during meiotic homologous recombination. In the present study, full length coding sequence of DMC1 gene was sequence characterized for the first time from four ruminant species (cattle, buffalo, sheep and goat) and phylogenetic relationship of ruminant DMC1 with other eukaryotes was analyzed. DMC1 gene encodes a putative protein of 340 amino acids in cattle, sheep and buffalo and 341 amino acids in goat. A high degree of evolutionary conservation at both nucleotide and amino acid level was observed for the four ruminant orthologs. In cattle and sheep, novel alternatively spliced mRNAs with skipping of exons 7 and 8 (Transcript variant 1, TV1) were isolated in addition to the full length (FL) transcript. Novel transcript variants with partial skipping of exon 7 and complete skipping of exon 8 (Transcript variant 2, TV2) were found in sheep and goat. The presence of these variants was validated by amplifying cDNA isolated from testis tissue of ruminants using two oligonucleotides flanking the deleted region. To accurately estimate their relative proportions, real-time PCR was performed using primers specific for each variant. Expression level of DMC1-FL was significantly higher than that of TV1 in cattle and TV2 in goat (P < 0.05). Relative ratio for expression of DMC1-FL: TV1: TV2 in sheep was 6.78: 1.43: 1. In-silico analysis revealed presence of splice variants of DMC1 gene across other mammalian species underpinning the role of alternative splicing in functional innovation.

Description of new species of Pterygorhabditis Timm, and Aspidonema (Sachs, ) Andrássy, (Nematoda: Bunonematoidea) in aquatic habitats from India.

The paper contains descriptions of two new species of the genera Pterygorhabditis Timm, 1957 and Aspidonema (Sachs, 1949) Andrássy, 1958 belonging to the families Pterygorhabditidae Goodey 1963 and Bunonematidae Micoletzky 1922, respectively. Species were procured from fixed samples, collected earlier from aquatic habitats. Pterygorhabditis punctata n. sp. is characterized by a cuticle with flattened hexagonal blocks arranged in eight longitudinal rows in both sexes; each metastegostomal plate with a minute denticle, and males with long, slender, fused spicules and nine pairs of post-cloacal, prominently setose genital papillae. Aspidonema formosa n. sp. is the first report of the genus from India. The species is characterized by the right side provided with 35-48 pairs of warts flanking a row of smooth membranous shields and surrounded by a well-developed network and an anisomorphic metastegostom without discernible armature. Species are compared with other congeners, together with the diagnoses of amended genera, and keys to the identification of species are provided.

Association of mucoid degeneration of anterior cruciate ligament with knee meniscal and cartilage damage.

OBJECTIVE: To assess the prevalence of anterior cruciate ligament (ACL) mucoid degeneration in patients referred for routine knee magnetic resonance (MR) imaging, and its association with age and structural joint damage. METHOD: Four independent radiologists assessed 413 consecutive knee MR examinations for the presence of a normal or ruptured ACL, or ACL mucoid degeneration. Knees with ACL mucoid degeneration were frequency matched by age, sex, and MR field strength with consecutive control knees with a normal ACL (1:2 ratio). Differences in meniscal and cartilage damage of the tibiofemoral compartments, as determined by the Whole-Organ MR Imaging Score (WORMS) system, were compared by Mann-Whitney U tests. Multivariable logistic regression analysis identified the association of ACL mucoid degeneration with severe MTFC cartilage damage (WORMS≥5). RESULTS: Patients with ACL mucoid degeneration (n = 36; 36% males; median age 55.5 years, range: 26-81) were older than patients with a normal (P < 0.001) or ruptured ACL (P < 0.001), without sex predilection (P = 0.76), and were more frequently diagnosed at 3 T (12%) compared to 1.5 T (2%). Knees with ACL mucoid degeneration had statistically significantly more medial meniscal (P < 0.001) and central and posterior medial tibiofemoral compartment (MTFC) cartilage (P < 0.001) damage compared with control knees (n = 72), but there were no differences in patients ≤50 years (P = 0.09 and 0.32, respectively). In multivariable logistic regression, severe MTFC cartilage damage (WORMS≥5) was significantly associated with ACL mucoid degeneration (odds ratio 4.09, 95% confidence interval 1.29-12.94, P = 0.016). CONCLUSION: There is a strong association between ACL mucoid degeneration and cartilage damage in the central and posterior MTFC, especially in patients >50 years.

Novel trends in development of dietary fiber rich meat products-a critical review.

Meat and meat products are generally recognized as good sources of high biological value proteins, fat-soluble vitamins, minerals, trace elements and bioactive compounds. Changes in socioeconomic factors in recent years have increased the consumer's preference for ready to eat foods including meat products. The processing of meat and meat products leads to generation of many functional compounds beneficial to human health but most of those foods are rich in fat, added salts but deficient in complex carbohydrates like dietary fiber and pose a health hazard that somehow is proved to be a predisposing factor for cardiovascular diseases, colon cancer, obesity including diabetes mellitus. With increasing consciousness among consumers about their nutrition and well being, there is a growing concern over nutritional diseases of affluence. Therefore an increase in dietary fiber inclusion in daily diet has been recommended. For adults, the recommended acceptable intakes of dietary fiber are 28-36 g/day, 70-80 % of which must be insoluble fiber. The insoluble fraction of dietary fiber has been related to intestinal regulation whereas soluble fiber is associated with decrease in cholesterol level and absorption of intestinal glucose. So incorporation of dietary fibers from different sources in meat products would help to enhance their desirability. Dietary fiber sources are generally agricultural byproducts that are comparatively cheap and incorporation in meat products reduces its overall cost. Whole grains and cereal brans are the rich source of insoluble fiber and pectins, gums, starch and other storage polysaccharides have high content of the soluble fraction. With this background, the effect of various dietary fibers on the quality attributes of meat and meat products with its physiological role has been reviewed here.

Novel polymorphism of AA-NAT gene in Indian goat breeds differing in reproductive traits.

This is the first description of the polymorphisms of arylalkylamine-N-acetyltransferase (AA-NAT) gene in Indian goats with different reproductive traits (twinning percentage and age of sexual maturity). Based on the important role of AA-NAT in reproduction, it is considered as a possible candidate gene for this trait. Two novel synonymous SNPs, C825T (exon2) and C1249T (exon3) were identified. All three possible genotypes (CC, CT and TT) were identified for C825T mutation whereas two genotypes were observed (CC and CT) for C1249T mutation. SNPs C825T and C1249T changed recognition site of restriction enzyme BtsCI (GGA T G) and AciI (C C GC) and thus can be genotyped by the relatively simple and cost effective technique of PCR-RFLP for establishing further association with reproductive traits. Present results add to the meager existing knowledge and extend the spectrum of genetic variation of caprine candidate genes of reproductive traits, which is another step towards improvement of goat genetic resources and breeding.

Gradient-echo in-phase and opposed-phase chemical shift imaging: role in evaluating bone marrow.

Chemical shift imaging (CSI) provides valuable information for assessing the bone marrow, while adding little to total examination time. In this article, we review the uses of CSI for evaluating bone marrow abnormalities. CSI can be used for differentiating marrow-replacing lesions from a range of non-marrow-replacing processes, although the sequence is associated with technical limitations and pitfalls. Particularly at 3 T, susceptibility artefacts are prevalent, and optimal technical parameters must be implemented with appropriate choices for echo times.

Selection of appropriate isolation method based on morphology of blastocyst for efficient derivation of buffalo embryonic stem cells.

The efficiency of embryonic stem cell (ESC) derivation from all species except for rodents and primates is very low. There are however, multiple interests in obtaining pluripotent cells from these animals with main expectations in the fields of transgenesis, cloning, regenerative medicine and tissue engineering. Researches are being carried out in laboratories throughout the world to increase the efficiency of ESC isolation for their downstream applications. Thus, the present study was undertaken to study the effect of different isolation methods based on the morphology of blastocyst for efficient derivation of buffalo ESCs. Embryos were produced in vitro through the procedures of maturation, fertilization and culture. Hatched blastocysts or isolated inner cell masses (ICMs) were seeded on mitomycin-C inactivated buffalo fetal fibroblast monolayer for the development of ESC colonies. The ESCs were analyzed for alkaline phosphatase activity, expression of pluripotency markers and karyotypic stability. Primary ESC colonies were obtained after 2-5 days of seeding hatched blastocysts or isolated ICMs on mitomycin-C inactivated feeder layer. Mechanically isolated ICMs attached and formed primary cell colonies more efficiently than ICMs isolated enzymatically. For derivation of ESCs from poorly defined ICMs intact hatched blastocyst culture was the most successful method. Results of this study implied that although ESCs can be obtained using all three methods used in this study, efficiency varies depending upon the morphology of blastocyst and isolation method used. So, appropriate isolation method must be selected depending on the quality of blastocyst for efficient derivation of ESCs.

Microsatellite analysis of genetic population structure of zebu cattle (Bos indicus) breeds from north-western region of India.

The present study aims to understand the existing genetic diversity and structure of six native cattle breeds (Rathi, Tharparkar, Nagori, Mewati, Gir, and Kankrej) adapted to the north-western arid and semi-arid region of India based on microsatellite loci. Various diversity estimates, mean number of alleles (12.84); effective number of alleles (5.02); gene diversity (0.769), and observed heterozygosity (0.667) reflected the existence of substantial within-breed diversity in all the investigated cattle breeds. Mean estimates of F-statistics: F(IT) = 0.144 ± 0.023, F(IS) = 0.071 ± 0.021, and F(ST) = 0.078 ± 0.014 were significantly different from zero (P < 0.05). The interbreed relationships indicated moderate level of breed differentiation between the six cattle breeds with least differentiation between Kankrej-Mewati pair. The phylogeny structuring further supported close grouping of Kankrej and Mewati breeds. Correspondence analysis plotted Rathi, Tharparkar, and Gir individuals into three separate areas of multivariate space; whereas, Kankrej, Mewati, and Nagori cattle showed low breed specific clustering. This reflected the existence of discrete genetic structure for Tharparkar, Rathi, and Gir, the prominent dairy breeds of the region; whereas, admixture was observed for Kankrej, Mewati, and Nagori individuals.

Genetic diversity and structure of two prominent zebu cattle breeds adapted to the arid region of India inferred from microsatellite polymorphism.

This study aims to assess the genetic diversity and population structure of two major zebu dairy breeds (Tharparkar and Rathi) adapted to the arid region of Rajasthan state of India. Various variability estimates indicate the existence of sufficient within-breed genetic diversity. Mean estimates of F-statistics are significantly different from zero: F (IS) = 0.112 +/- 0.029, F (IT) = 0.169 +/- 0.033, F (ST) = 0.065 +/- 0.017. The overall positive value of F (IS) (0.112) and an F (IT) value (0.169) that is more than the F (ST) (0.065) indicate departure from random mating. The drift-based estimates reflect a moderate yet significant level of breed differentiation between the Tharparkar and Rathi breeds. The evaluation of an exact test, showing that allele frequencies across all the loci differed significantly, supports the population differentiation. This is paralleled by the outcome of neighbor-joining clustering based on allele-sharing distance measures. The allocation of a high percentage of individuals (95.7%) to their population of origin and correspondence analysis further substantiates the existence of a cohesive genetic structure in both the breeds.

Successful use of biliary accessories in antegrade dilation of complex upper esophageal stricture due to chemoradiation and surgery.

Endoscopic management of complete or near complete upper esophageal strictures is challenging. Current methods such as retrograde esophageal access are high risk and may require additional abdominal surgery. A biliary cannulation technique with a 0.035 inch guidewire was utilized to obtain antegrade esophageal access in a patient with near complete high esophageal stricture due to chemo radiation and surgery for head and neck cancer. Biliary accessories including bougie and balloon dilators were used for the initial dilation of the esophageal stricture, followed by the traditional approach of stricture dilation using over-the-wire dilators. The procedure was successfully performed in a patient with near complete upper esophageal stricture due to chemo radiation and surgery for recurrent laryngeal cancer. The dysphagia of this patient was resolved following serial esophageal dilations and his esophageal stricture was wide open on the last upper endoscopy. Biliary accessories can be safely used for obtaining antegrade esophageal access and dilation of near complete upper esophageal strictures. This approach should be considered in patients with complex esophageal strictures, especially after chemo radiation or surgery for head and neck cancer and prior to seeking other more complex alternatives involving retrograde esophageal access.

Genetic diversity and bottleneck analysis of Indian bellary sheep by microsatellite markers.

Bellary sheep population variability and structure was investigated genetically utilizing FAO recommended microsatellite markers. Genetic variation at 20 microsatellite loci, population structure, and genetic bottleneck hypothesis were examined. Estimates of genetic variability such as effective number of alleles and gene diversities revealed substantial genetic variation frequently displayed by microsatellite markers. A total of 133 alleles were detected. Average polymorphism across the studied loci and expected gene diversity in the population were 1.419 +/- 0.405 and 0.684 +/- 0.140, respectively. No significant genotypic linkage disequilibrium was detected across population, suggesting no evidence of linkage between loci. The population was observed to be significantly differentiated into different groups, showed fairly high level of inbreeding (f = 0.253 +/- 0.050) and global heterozygote deficit. Population structure analysis indicated the intermixing/introduction of unique/rare alleles in these migrating flocks. A normal L-shaped distribution of mode-shift test, non-significant heterozygosity excess on the basis of different models, as revealed from Sign, Standardized differences and Wilcoxon sign rank tests suggested that there was no recent bottleneck. The study revealed that even breed with increasing population trend needs genetic management for the conservation and improvement.