Worker laying in leafcutter ant Acromyrmex subterraneus brunneus （Formicidae, Attini）

We studied the process of offspring production in queenless colonies of Acromyrmex subterraneus brunneus, and particularly evaluated the ovary development of workers as a function of their age. For this, subcolonies were set up and evaluated at different periods of isolation from the queen （2, 4 and 6 months）, besides individually labeled age groups. The subcolonies were assessed according to offspring production and ovaries containing oocytes or not. The evaluations showed worker oviposition and development of males originating from worker-laid eggs. At 2 months＇ absence of the queen, eggs and larvae were found, with eggs in a higher proportion than larvae. After 4 months, the proportion of eggs had reduced while larvae had increased, and a pupa was found in one subcolony. At 6 months, besides a higher share of larvae, one pupa and one adult male were found. Dissection of workers revealed ovaries containing oocytes during the periods of evaluation. Only a group of medium-sized and large workers, 23.3%, 20.9% and 37.5% of the population from each period assessed in queenless subcolonies respectively, presented developed oocytes in the ovary. The same was observed in colonies with a queen, with 17.6%, 19.6% and 7.8% of the group of dissected workers from each time period, respectively. With respect to worker age, we observed by dissection of the ovary, that the greatest percentage of individuals with ovarioles containing oocytes occurred at 45 days （6 weeks） up to 90 days （12 weeks）. These results probably are associated with the workers reproduction and the laying of trophic and reproductive eggs in colonies with and without a queen; these eggs have distinct functions in each situation.     

长木蜂蜂粮中微生物丰度及抑菌活性菌株的筛选

长木蜂属于野生蜂类,后代的生存和繁衍完全依靠雌蜂所提供的蜂粮,蜂粮保鲜时间的长短对蜂群的繁衍具有关键性作用。本文采用营养琼脂培养基、改良"LB"培养基、MRS(4.5)、MRS(6.5)培养基分别对芍药的手采花粉、长木蜂蜂花粉和不同酿制时间的蜂粮样品中的微生物进行菌落计数、分析和分离鉴定。酿制初期的蜂粮样品在营养琼脂培养基生长的细菌菌落数占优势,随酿制时间的延长而迅速下降;7d、8d蜂粮样品在改良"LB"培养基上生长的细菌菌落数占优势;10d之后的蜂粮样品在MRS(6.5)培养基上微生物菌落数占优势;分离的68株菌株,其中细菌43株,酵母17株,放线菌8株。从蜂粮中分离的酵母菌和放线菌菌株均来自长木蜂雌蜂的体表。分离的68株菌株中有12株有较强抑菌活性,尤其是放线菌NF-34菌株对真菌和细菌均有抑菌活性。     

Characterization of contracting cardiomyocyte colonies in the primary culture of neonatal rat myocardial cells: A model of in vitro cardiomyogenesis

The unmet clinical need for myocardial repair after irreversible ischemic injury requires a better understanding of the biological properties of cardiac stem cells (CSCs). Using a primary culture of neonatal rat myocardial cells, we describe the formation and maturation of contracting cardiomyocyte colonies stemming from c-kit⁺, Sca⁺, or Isl1⁺ CSCs, which occurs in parallel to the hypertrophy of the major cardiac myocyte population. The contracting cardiomyocyte colonies (~1–2 colonies per 1 × 10⁵ of myocardial cells) were identified starting from eighth day of culturing. At first, spontaneous weak, asynchronous, and arrhythmic contractions of the colonies at a rate of 2–3 beats/min were registered. Over time, the contractions of the colonies became more synchronous and frequent, with a contraction rate of 58–60 beats/min by the 30th day of culturing. The colonies were characterized by the CSCs subtype-specific pattern of growth and structure. The cells of the colonies were capable of spontaneous cardiomyogenic differentiation, demonstrating expression of both sarcomeric α-actinin and α-sarcomeric actin as well as the maturation of contractile machinery and typical Ca²⁺ responses to caffeine (5 mМ) and K⁺ (120 mМ). Electromechanical coupling, characterized by cardiac muscle-specific Ca²⁺-induced Ca²⁺ release, was evident at 3 weeks of culturing. Thus, the co-cultivation of CSCs with mature cardiac cells resulted in the formation of contracting cardiomyocyte colonies, resembling the characteristics of in vivo cardiomyogenesis. The proposed model can be used for the investigation of fundamental mechanisms underlying cardiomyogenic differentiation of CSCs as well as for drug testing and/or other applications.     

Nanocolonies: Detection, cloning, and analysis of individual molecules

Nanocolonies (other names molecular colonies or polonies) are formed upon template nanomolecule (DNA or RNA) amplification in immobilized medium with efficient pore size in the nanometer range. This work deals with the principle, invention, development, and diverse nanocolony applications based on their unique abilities to compartmentalize amplification and expression of individual DNA and RNA molecules, including studying reactions between single molecules, digital molecular diagnostics, in vitro gene cloning and expression, as well as identification of the molecular cis-elements including DNA sequencing, analysis of single-nucleotide polymorphism, and alternative splicing investigation.     

Nonlinearity in the Growth of Bacterial Colonies: Conditions and Causes

The universally recognized kinetic model of colony growth, introduced by Pirt, predicts a linear increase of colony size. The linearity follows from the assumption that the colony expands through the growth of only such cells that are located immediately behind the moving colony front, in the so-called peripheral zone of constant width and density. In this work, Pirt's model was tested on two bacteria—Alcaligenes sp. and Pseudomonas fluorescens—having markedly distinct cultural properties and grown on an agarized medium with pyruvate. The colony size dynamics was followed for different densities of the inoculum, ranging from a single cell to a microdroplet of bacterial suspension (10⁵–10⁶ cells), and for different depths of the agar layer, determining the amount of available substrate. A linear growth mode was observed only with P. fluorescens and only in the case of growth from a microdroplet. When originating from a single cell, colonies of both organisms displayed nonlinear growth with a distinct peak of K _r (the rate of colony radius increase) occurring after 2–3 days of growth. The growth of P. fluorescens colonies showed virtually no dependence on the depth of the agarized medium, whereas the rate of colony size increase of Alcaligenes sp. turned out to be directly related to the medium layer thickness. The departure from linearity is consistently explained by a new kinetic scheme stipulating a possible contribution to the colony growth not only of peripheral cells but also (much more distinct in Alcaligenes) of cells at the colony center. The colony growth dynamics is determined not only by the concentration of the limiting substrate but also by the amount of autoinhibitor, the synthesis of which is governed by the age of cells. The distinctions of growth from a single cell and microdroplet could also originate as a result of dissociation into the R- and S-forms and competition between the corresponding subpopulations for oxygen and the common substrate.     

Continuous Growth and Differentiation of Trypanosoma (Megatrypanum) freitasi Rego, Magalhaes & Siqueira, 1957, In Vitro

Trypanosoma (Megatrypanum) freitasi, a parasite of didelphid opossum, was known to be very difficult to cultivate in conventional media. Co-cultivation with L929 cell line in Baltz's medium at 27.5° C resulted in luxuriant growth of the trypanosome with the production of epimastigote colonies that adhered to the surface of culture flasks or tubes, and transformation into metacylics. Further transformation was stimulated by raising the incubation temperature. At 37° C the population was of the bloodstream type and resistant to lysis by complement.     

Longevity of U cells of differentiated yeast colonies grown on respiratory medium depends on active glycolysis

Colonies of Saccharomyces cerevisiae laboratory strains pass through specific developmental phases when growing on solid respiratory medium. During entry into the so-called alkali phase, in which ammonia signaling is initiated, 2 prominent cell types are formed within the colonies: U cells in upper colony regions, which have a longevity phenotype and activate the expression of a large number of metabolic genes, and L cells in lower regions, which die more quickly and exhibit a starvation phenotype. Here, we performed a detailed analysis of the activities of enzymes of central carbon metabolism in lysates of both cell types and determined several fermentation end products, showing that previously reported expression differences are reflected in the different enzymatic capabilities of each cell type. Hence, U cells, despite being grown on respiratory medium, behave as fermenting cells, whereas L cells rely on respiratory metabolism and possess active gluconeogenesis. Using a spectrum of different inhibitors, we showed that glycolysis is essential for the formation, and particularly, the survival of U cells. We also showed that β-1,3-glucans that are released from the cell walls of L cells are the most likely source of carbohydrates for U cells.     

单菌落PCR法直接快速鉴定重组克隆

利用单菌落PCR法直接筛选含有GFP、LTB-ST外源基因的重组克隆,阳性克隆可以扩增出目的条带,和质粒PCR扩增的结果一致。同时,单菌落PCR法也可应用于重组质粒转化后的农杆菌的筛选,单菌落PCR法的扩增结果和农杆菌液扩增的结果一致。结果表明,单菌落PCR法是一个有效简便的鉴定重组阳性克隆的方法。