Spatio-temporal study of environmental nontuberculous mycobacteria isolated from Wardha district in Central India

During the last two decades, nontuberculous mycobacteria (NTM) have gained in importance but there is still a paucity of data, particularly for environmental isolates. We studied, over a period of two years, the spatio-temporal features of NTM isolates obtained from different environmental sources in Wardha district, India. A total of 1398 samples (699 each of soil and water) were tested and 170 (12.2%) yielded NTM isolates, including 123 from soil and 47 from water samples. Out of 170 NTM isolates, 107 (63%) belonged to potentially pathogenic mycobacteria (PPM) and 63 (37%) to the less pathogenic mycobacterial (LPM) group. Overall, maximum isolation was obtained in rainy season (20.3%) followed by winter (13.5%), post rainy (8.7%) and summer seasons (5.8%). Mycobacterium fortuitum, Mycobacterium gordonae and Mycobacterium avium complex (MAC) were common isolates followed by Mycobacterium flavescens, Mycobacterium scrofulaceum, Mycobacterium simiae and Mycobacterium marinum. From soil, isolation of NTM was highest from grounds used for community gatherings (42.8%) followed by soil from residential premises (27.7%) and near the wells (26.0%). From drinking water sources, highest NTM isolation was obtained from wells (15.4%) followed by treated water tanks (6.9%), household receptacles (6.3%), hand pumps (5.6%) and tap water supply (3.5%). Isolation from natural canal water was 6.6%, while from drainage and waste water ponds isolation was 8.3%. The results of the study revealed that in Wardha district, NTM are present both in the soil and drinking water. As NTM can be pathogenic, particularly in immune-compromised individuals, these can be of potential risk to the human population.     

Comparison of Some Plant DNA Extraction Methods

DNA isolation is a routine procedure when performed in laboratory environment, yet in the field it may still remain problematic. This is especially true of some crop species bred for useful metabolites that may also hinder DNA extraction. Here we compare the efficiency of DNA extraction protocols and commercial DNA isolation kits when used on samples from Helianthus and Allium. Since extraction of DNA is known to be compromised by co-extraction of PCR-inhibiting metabolites, the isolation of DNA was followed by PCR as a testing procedure for the isolation step. The MagnoPrime Fact and MagnoPrime Uni DNA isolation kits were better suited for field work due to faster processing times and smaller required amount of starting material (20 mg fresh/0.5 mg dry). In all cases the subsequent PCR managed to amplify the DNA fragments of interest well enough to be useful in further research.     

Invasion of the Fungal Pathogen <Emphasis Type="Italic">Batrachochytrium dendrobatidis</Emphasis> on California Islands

Batrachochytrium dendrobatidis (Bd), an amphibian fungal pathogen, has infected >500 species and caused extinctions or declines in >200 species worldwide. Despite over a decade of research, little is known about its invasion biology. To better understand this, we conducted a museum specimen survey (1910–1997) of Bd in amphibians on 11 California islands and found a pattern consistent with the emergence of Bd epizootics on the mainland, suggesting that geographic isolation did not prevent Bd invasion. We propose that suitable habitat, host diversity, and human visitation overcome isolation from the mainland and play a role in Bd invasion.     

Molecular biological properties of new isolates of plum pox virus strain Winona

Plum pox virus (PPV, genus Potyvirus, family Potyviridae) is the most important virus pathogen of stone fruit cultures of the genus Prunus from an economical standpoint. Winona strain (PPV-W) is the most variable of nine known virus strains and one of the most widespread in the European part of Russia. Six new PPV-W isolates were first discovered in green plantations of Moscow (Kp2U, Avang, Pulk, Pulk-1), in Taldomsky district of Moscow region (Karm), and in Kovrovsky district of Vladimir region (Vlad-4) on wild trees of plum Prunus domestica. 3’-Terminal segment of genome of new isolates was notable for high variability level. The study on the relationship with other isolates of this strain by means of phylogenetic analysis of gene sequence of the coat protein showed the lack of clusterization of Russian PPV-W isolates according to geographical principle. Inoculation of Nicotiana benthamiana plants by hop plant louse Phorodon humili from plum trees infected with Avang and Pulk isolates and by thistle aphid Brachycaudus cardui from the tree infected with Kp2U isolate led to the systemic viral infection in indicator plants, suggesting the possibility of PPV-W spread by both aphid species in nature. Transmission of PPV-W through seeds was not observed.     

Application of geometric morphometrics for intraspecific variability analysis in mesopelagic fishes of Sternoptychidae and Myctophidae families

We present the results of application of geometric morphometrics for studies of variability in populations of the members of two families of South Atlantic mesopelagic fishes, Sternoptyx diaphana (Sternoptychidae) and Ceratoscopelus warmingii (Myctophidae), in which an almost complete genetic isolation was previously discovered within the borders of hydrographic regions. A system of landmark points is developed, describing both anatomical features and configuration of luminescent organs, photophores, which allows to consider not only the influence of external physical and ecological conditions but also the existence of population-specific signaling system as a potential cause of genetic isolation. Morphological similarities are demonstrated for S. diaphana throughout the whole studied portion of the distribution area, while certain differences affecting the head shape and position of individual photophores on the body are discovered between C. warmingii populations.     

Pulsatile arterial wall-blood flow interaction with wall pre-stress computed using an inverse algorithm

Background

The computation of arterial wall deformation and stresses under physiologic conditions requires a coupled compliant arterial wall-blood flow interaction model. The in-vivo arterial wall motion is constrained by tethering from the surrounding tissues. This tethering, together with the average in-vivo pressure, results in wall pre-stress. For an accurate simulation of the physiologic conditions, it is important to incorporate the wall pre-stress in the computational model. The computation of wall pre-stress is complex, as the un-loaded and un-tethered arterial shape with residual stress is unknown. In this study, the arterial wall deformation and stresses in a canine femoral artery under pulsatile pressure was computed after incorporating the wall pre-stresses. A nonlinear least square optimization based inverse algorithm was developed to compute the in-vivo wall pre-stress.

Methods

First, the proposed inverse algorithm was used to obtain the un-loaded and un-tethered arterial geometry from the unstressed in-vivo geometry. Then, the un-loaded, and un-tethered arterial geometry was pre-stressed by applying a mean in-vivo pressure of 104.5 mmHg and an axial stretch of 48% from the un-tethered length. Finally, the physiologic pressure pulse was applied at the inlet and the outlet of the pre-stressed configuration to calculate the in-vivo deformation and stresses. The wall material properties were modeled with an incompressible, Mooney-Rivlin model derived from previously published experimental stress-strain data (Attinger et al., 1968).

Results

The un-loaded and un-tethered artery geometry computed by the inverse algorithm had a length, inner diameter and thickness of 35.14 mm, 3.10 mm and 0.435 mm, respectively. The pre-stressed arterial wall geometry was obtained by applying the in-vivo axial-stretch and average in-vivo pressure to the un-loaded and un-tethered geometry. The length of the pre-stressed artery, 51.99 mm, was within 0.01 mm (0.019%) of the in-vivo length of 52.0 mm; the inner diameter of 3.603 mm was within 0.003 mm (0.08%) of the corresponding in-vivo diameter of 3.6 mm, and the thickness of 0.269 mm was within 0.0015 mm (0.55%) of the in-vivo thickness of 0.27 mm. Under physiologic pulsatile pressure applied to the pre-stressed artery, the time averaged longitudinal stress was found to be 42.5% higher than the circumferential stresses. The results of this study are similar to the results reported by Zhang et al., (2005) for the left anterior descending coronary artery.

Conclusions

An inverse method was adopted to compute physiologic pre-stress in the arterial wall before conducting pulsatile hemodynamic calculations. The wall stresses were higher in magnitude in the longitudinal direction, under physiologic pressure after incorporating the effect of in-vivo axial stretch and pressure loading.

     

Overexpression of <Emphasis Type="Italic">PSP1</Emphasis> enhances growth of transgenic Arabidopsis plants under ambient air conditions

Cellulose biosynthesis is mediated by cellulose synthases (CesAs), which constitute into rosette-like cellulose synthase complexe (CSC) on the plasma membrane. Two types of CSCs in Arabidopsis are believed to be involved in cellulose synthesis in the primary cell wall and secondary cell walls, respectively. In this work, we found that the two type CSCs participated cellulose biosynthesis in differentiating xylem cells undergoing secondary cell wall thickening in Populus. During the cell wall thickening process, expression of one type CSC genes increased while expression of the other type CSC genes decreased. Suppression of different type CSC genes both affected the wall-thickening and disrupted the multilaminar structure of the secondary cell walls. When CesA7A was suppressed, crystalline cellulose content was reduced, which, however, showed an increase when CesA3D was suppressed. The CesA suppression also affected cellulose digestibility of the wood cell walls. The results suggest that two type CSCs are involved in coordinating the cellulose biosynthesis in formation of the multilaminar structure in Populus wood secondary cell walls.     

Bacterial diversity obtained by culturable approaches in the gut of <Emphasis Type="Italic">Glossina pallidipes</Emphasis> population from a non sleeping sickness focus in Tanzania: preliminary results

Background

Glossina pallidipes is a haematophagous insect that serves as a cyclic transmitter of trypanosomes causing African Trypanosomiasis (AT). To fully assess the role of G. pallidipes in the epidemiology of AT, especially the human form of the disease (HAT), it is essential to know the microbial diversity inhabiting the gut of natural fly populations. This study aimed to examine the diversity of G. pallidipes fly gut bacteria by culture-dependent approaches.

Results

113 bacterial isolates were obtained from aerobic and anaerobic microorganisms originating from the gut of G. pallidipes. 16S rDNA of each isolate was PCR amplified and sequenced. The overall majority of identified bacteria belonged in descending order to the Firmicutes (86.6%), Actinobacteria (7.6%), Proteobacteria (5.5%)and Bacteroidetes (0.3%). Diversity of Firmicutes was found higher when enrichments and isolation were performed under anaerobic conditions than aerobic ones. Experiments conducted in the absence of oxygen (anaerobiosis) led to the isolation of bacteria pertaining to four phyla (83% Firmicutes, 15% Actinobacteria, 1% Proteobacteria and 0.5% Bacteroidetes, whereas those conducted in the presence of oxygen (aerobiosis) led to the isolation of bacteria affiliated to two phyla only (90% Firmicutes and 10% Proteobacteria). Phylogenetic analyses placed these isolates into 11 genera namely Bacillus, Acinetobacter, Mesorhizobium, Paracoccus, Microbacterium, Micrococcus, Arthrobacter, Corynobacterium, Curtobacterium, Vagococcus and Dietzia spp.which are known to be either facultative anaerobes, aerobes, or even microaerobes.

Conclusion

This study shows that G. pallidipes fly gut is an environmental reservoir for a vast number of bacterial species, which are likely to be important for ecological microbial well being of the fly and possibly on differing vectorial competence and refractoriness against AT epidemiology.

     

A new species ofScopulariopsis from soil

A new species ofScopularoipsis described from soil is characterized by the conidiophores, branches, and sporogenous cells being distinctly swollen. The conidia have a well-marked germ slit running more or less longitudinally down the spore. TheScopulariopsis state ofMicroascus doguetii is re-examined and evidence is presented which indicates that the striations on the walls of the spores are germ slits.     

Two new bioluminescent <Emphasis Type="Italic">Henlea</Emphasis> from Siberia and lack of molecular support for <Emphasis Type="Italic">Hepatogaster</Emphasis> (Annelida,Clitellata, Enchytraeidae)

Two bioluminescent enchytraeids, Henlea petushkovi sp. n. and Henlea rodionovae sp. n., are described from the Krasnoyarsk and Irkutsk regions in Eastern Siberia. These large potworms exhibit the typical light-production pattern reported repeatedly in the genus and recently elucidated by Russian researchers in its main biophysical and biochemical aspects. Morphology and DNA indicate that the two species are very closely related, but clearly divergent in the strength of the body wall (thick and opaque in H. petushkovi), structure of the prostomium (in H. rodionovae unprecedentedly wrinkled and mobile), brain shape (almost equilateral in H. petushkovi), size of coelomocytes (60–85 μm in H. petushkovi) and structure of intestinal diverticula (tulip-shaped in H. petushkovi, apple-shaped in H. rodionovae). Limited hybridization seems to occur between them, supported by a single case of conflict between COI and morphology, and a few intermediate morphotypes were noted in greenhouse populations. The new species are phylogenetically distant from all known congeners so far DNA-barcoded, even those that, like them, respond to the diagnosis of the putative subgenus Hepatogaster ?ejka, 1910 (multitubular gut diverticula in VIII, indented brain, dorsal blood vessel from IX, prominent spermathecal glands, and nephridia from 5/6). In fact, our phylogenetic analyses dismiss Hepatogaster as an artificial (polyphyletic) taxon. Issues related to the definition of H. nasuta (Eisen, 1878), H. ochracea (Eisen, 1878) and H. irkutensis Burov, 1929, three species originally described from Siberia, indicate that Henlea taxonomy is still in a state of flux, as regards not only species interrelationships but also species definitions.     

Extracellular hydrolases of strain <Emphasis Type="Italic">Bacillus</Emphasis> sp. 739 and their involvement in the lysis of micromycete cell walls

The mycolytic bacterial strain Bacillus sp. 739 produces extracellular enzymes which degrade in vitro the cell walls of a number of phytopathogenic and saprophytic fungi. When Bacillus sp. 739 was cultivated with Bipolaris sorokiniana, a cereal root-rot pathogen, the fungus degradation process correlated with the levels of the β-1,3-glucanase and protease activity. The comparative characteristic of Bacillus sp. 739 enzymatic preparations showed that efficient hydrolysis of the fungus cell walls was the result of the action of the complex of enzymes produced by the strain when grown on chitin-containing media. Among the enzymes of this complex, chitinases and β-1,3-glucanases hydrolyzed most actively the disintegrated cell walls of B. sorokiniana. However, only β-1,3-glucanases were able to degrade the cell walls of native fungal mycelium in the absence of other hydrolases, which is indicative of their key role in the mycolytic activity of Bacillus sp. 739.     

Reproductive Isolation Among <Emphasis Type="Italic">Drosophila arizonae</Emphasis> from Geographically Isolated Regions of North America

A long-standing goal of speciation research is to describe how reproductive isolating barriers develop, when they arise along the ‘speciation continuum’, and to measure the strength with which they restrict gene flow. Drosophila arizonae and D. mojavensis are a recently diverged sister species pair distributed from the southwestern United States through southern Mexico. While incipient speciation in D. mojavensis has been studied for decades, relatively little attention has been directed toward D. arizonae, despite the fact that previous studies have revealed evidence for significant genetic differentiation among populations separated by geographic barriers. Here, we examine the potential for both pre- and post-mating reproductive isolation in D. arizonae from geographically isolated parts of North America. We find evidence for strong premating isolation between flies from northern mainland Mexico and southern mainland Mexico, but no evidence for postmating isolation in any cross. This study highlights the utility of the D. arizonae system for further investigation into the early evolution of premating isolation, and reinforces the potential of the D. arizonae/D. mojavensis system as a whole for studying the evolution of reproductive isolation at a range of divergence times.     

Quarnet Inference Rules for Level-1 Networks

An important problem in phylogenetics is the construction of phylogenetic trees. One way to approach this problem, known as the supertree method, involves inferring a phylogenetic tree with leaves consisting of a set X of species from a collection of trees, each having leaf-set some subset of X. In the 1980s, Colonius and Schulze gave certain inference rules for deciding when a collection of 4-leaved trees, one for each 4-element subset of X, can be simultaneously displayed by a single supertree with leaf-set X. Recently, it has become of interest to extend this and related results to phylogenetic networks. These are a generalization of phylogenetic trees which can be used to represent reticulate evolution (where species can come together to form a new species). It has recently been shown that a certain type of phylogenetic network, called a (unrooted) level-1 network, can essentially be constructed from 4-leaved trees. However, the problem of providing appropriate inference rules for such networks remains unresolved. Here, we show that by considering 4-leaved networks, called quarnets, as opposed to 4-leaved trees, it is possible to provide such rules. In particular, we show that these rules can be used to characterize when a collection of quarnets, one for each 4-element subset of X, can all be simultaneously displayed by a level-1 network with leaf-set X. The rules are an intriguing mixture of tree inference rules, and an inference rule for building up a cyclic ordering of X from orderings on subsets of X of size 4. This opens up several new directions of research for inferring phylogenetic networks from smaller ones, which could yield new algorithms for solving the supernetwork problem in phylogenetics.     

Disjunct Occurrence of <Emphasis Type="Italic">Trichadenotecnum s.str</Emphasis>. in Southeastern Brazil (Psocodea: “Psocoptera”: Psocidae), with Description of a New Species

A new barklouse species belonging to the Trichadenotecnum s.str. clade (Psocodea: “Psocoptera”: Psocidae) was described from southeastern Brazil (Minas Gerais state). This locality is highly isolated (about 3400 km) from the closest known distributional range of this clade. It also represents the southernmost distributional record of Trichadenotecnum s.str. Phylogenetic placement of the species and biogeography of Neotropical Trichadenotecnum were briefly discussed. Trichadenotecnum punctipenne New, 1972 described from Brazil was synonymized with Trichadenotecnum pardus Badonnel, 1955.     

New species of unicellular obligate parasite, <Emphasis Type="Italic">Olpidiopsis pyropiae</Emphasis> sp. nov., that plagues <Emphasis Type="Italic">Pyropia</Emphasis> sea farms in Korea

Pyropia (Porphyra) sea farms are plagued with many diseases, similar to a land crop field. Olpidiopsis disease has been one of the most serious diseases causing multimillion dollars of economic loss every year. From 3 years of epidemiological studies in Pyropia sea farms, we found that the pathogen of Olpidiopsis disease in Korea is different from the oomycete, Olpidiopsis porphyrae, which is known to infect the commercially cultivated Pyropia yezoensis in Japan. The Korean species showed a clearly different small subunit (SSU) 18S rRNA sequence (90.4 % identity) and lacked four intron-like insertions, which are present in O. porphyrae. We therefore established a new species named Olpidiopsis pyropiae. The infection process and asexual life history of O. pyropiae were similar to O. porphyrae. Infection started when zoospores attached to the surface of Pyropia blade, lost flagella, and produced thin germ tubes that penetrated the cell walls of the host. Spherical multinucleate thalli developed within the cytoplasm of its algal host, which grew into mature sporangia within the next 2 days. The shape and size of sporangium was similar to that of the Japanese species, except for longer discharge tubes in O. pyropiae. Transmission electron microscopy revealed the presence of K-body-like organelles with tubular inclusions located close to the nucleus, which is one of the key characters of the genus. However, phylogenetic analysis based on SSU ribosomal RNA (rRNA) gene data showed a loose affinity of the Korean species with the other marine Olpidiopsis spp.     

Existence of two strains of <Emphasis Type="Italic">Habrobracon hebetor</Emphasis> (Hymenoptera: Braconidae): a complex in Thailand and Japan

Habrobracon hebetor Say (Hymenoptera: Braconidae) is a cosmopolitan gregarious ectoparasitoid that attacks larvae of several species of Lepidoptera. Although there are two genetically different strains within H. hebetor, distribution of the strains has been poorly understood. In 2010, in Thailand, where H. hebetor has been known as a parasitoid of stored grain pests, it was found that H. hebetor attacked Opisina arenosella Walker (Lepidoptera: Oecophoridae), which is an invasive pest of coconut palm. For correct identification of this H. hebetor, we conducted DNA analysis and cross tests using populations collected from O. arenosella and stored grain pests in Thailand and populations in Japan known as H. hebetor. We obtained 413 bp of mitochondrial cytochrome oxidase I (COI) sequences and 414 bp of 16S rRNA gene sequences, and both indicated that there are two distinct clades within H. hebetor: one contains insects from Thailand, Spain, India, and Barbados; the other contains insects from Japan and the USA. There were no genetic differences or sexual isolation between Thai populations from different hosts. Our results also showed that populations in Thailand were sexually isolated from a H. hebetor population in Japan.     

Systematic and phylogenetic implications of the wood anatomy of six Neotropical genera of Primulaceae

Our main goals were to identify diagnostic characters at the species, genus, and subfamily levels, find anatomical features with potential for future morphological and molecular (combined) phylogenetic analyses, and to reconstruct the evolution of wood anatomical characters in two subfamilies of Primulaceae in a molecular phylogenetic framework. We investigated twenty-seven species from the woody Myrsinoideae (4 genera) and Theophrastoideae (2 genera) using scanning electron, light, and epifluorescence microscopy. Samples were prepared using standard protocols. Based on the wood anatomical characters, we were able to identify synapomorphies and to detect evolutionary trends of interest for the genera and subfamilies. Both subfamilies share the presence of diffuse porosity, simple perforation plates, septate fibres, and scanty paratracheal axial parenchyma. Theophrastoideae species have rays?>?10 cells wide and short (<?350 µm) vessel elements, and Myrsinoideae have breakdown areas in rays and longer vessel elements. Ardisia and Stylogyne have scalariform intervessel pits, Myrsine exhibit breakdown areas in rays, and two Cybianthus species from subgenus Weilgetia have distinguishing features (e.g., scalariform perforation plate in C. nemoralis and the absence of rays in C. densiflorus). Overall, when combining characters, we were able to segregate the Neotropical Primulaceae subfamilies and genera from each other and from the subfamily Maesoideae based on wood anatomy.     

An improved method of DNA purification from secondary metabolites rich medicinal plants using certain chaotropic agents

The presented work describes good quality DNA isolation method from mature leaves of some medicinally important plant species, viz. Asparagus racemosus, Withania somnifera, Abrus precatorius, Commiphora wightii and Carissa carandas. These plants hold immense medicinal values due to presence of certain secondary metabolites like polyphenols, terpenes, flavonoids, alkaloids, gums, resins, etc. Although these metabolites are accountable for important medicinal properties and authorize these plants to precedence over others, the same compounds disappoint the researcher while isolating high quality DNA. To overcome this problem, we propose a simple method in which DNA is adroitly bounded to diatomaceous earth in a solution of different chaotropic agent and alienated from intrusive compounds. Presented method affirms that secondary products, along with polysaccharides and proteins, can be perceptibly reduced by using silica matrix along with chaotropic agents. The described method is fast, simple and highly reliable for the isolation of DNA from obstinate plant species.     

Bacterial and fungal biodeterioration of discolored building paints in Lagos,Nigeria

Microbial induced discolorations are an unsightly feature occurring on painted walls in Lagos, the commercial hub of Nigeria. Very few studies have been carried out conventionally about the microbial community structure of discolored painted walls in Nigeria therefore, knowledge of the true microbial diversity is elusive. To further our understanding of the phylogenetic diversity of representative microbial community on 40 discolored and three clean-looking buildings, a comparative DNA sequence analysis of 16S rDNA genes was undertaken. Following DNA extraction, portions of the rDNA genes were amplified by PCR and sequenced. Resulting sequences were compared with GenBank data base sequences. Fifteen unique fungal sequences and one bacterial sequence were obtained. Majority (37.50%) of rDNA sequences analyzed, represent the genus Meyerozyma of which the novel fungus Meyerozyma guilliermondii, which to our knowledge, has not yet been implicated in painted walls was detected. Clones from the discolored painted wall isolates also produced a data set in which 31.25% of sequences were related to Fusarium proliferatum and 6.25% were Pseudomonas aeruginosa. The remaining sequences clustered with members of the genera Candida (6.25%), Aspergillus (12.5%) and Cerrena (6.25%). The study provides reliable data on microbial communities on painted walls and information for paint biocide formulation.