A simple heat-based alternative method for deparaffinization of histological sections significantly improves acid-fast staining results for Mycobacteria in tissue.

Histopathology is the study of how disease alters human and animal tissue and is based on the microscopic examination of stained tissue sections. To maintain tissue integrity, preserving it from degradation, it is initially fixed, primarily with formalin, before being treated with alcohol and organic solvents, allowing the infiltration of paraffin wax. The tissue can then be embedded in a mold and sectioned, usually at a thickness between 3 and 5 µm, before staining with dyes or antibodies to demonstrate specific components. As the paraffin wax is insoluble in water, it is necessary to remove it from the tissue section before applying any aqueous or water-based dye solution, to allow the tissue to successfully interact with the stain. This deparaffinization/hydration step is normally carried out using xylene, an organic solvent, followed by hydration using graded alcohols. However, this use of xylene has been shown to have detrimental effects on acid-fast stains (AFS), such as those employed to demonstrate Mycobacterium, including the causative agent of tuberculosis (TB), as the integrity of the lipid-rich wall present in these bacteria may be compromised using xylene. A simple, novel method, Projected Hot Air Deparaffinization (PHAD) removes the solid paraffin from the tissue section without the use of any solvents, which produces significantly improved staining results using AFS. PHAD relies on the projection of hot air onto the histological section to melt and remove paraffin from the tissue, which can be achieved using a common hairdryer. •PHAD relies on the projection of hot air onto the histological section which can be achieved using a common hairdryer.•The blowing force is such that melted paraffin is removed from the tissue in 20 min.•Subsequent hydration allows for using aqueous histological stains with success, such as the fluorescent auramine O acid-fast-stain.

A novel and simple heat-based method eliminates the highly detrimental effect of xylene deparaffinization on acid-fast stains.

OBJECTIVES: Histopathology is an important method for diagnosing extrapulmonary tuberculosis, yet tissue sections are often negative for mycobacteria after use of acid-fast stain (AFS). This study investigated the mechanism of AFS use and the detrimental effect of histologic processing-in particular, xylene deparaffinization-on AFS and mycobacterial detection. METHODS: The target of the fluorescent Auramine O (AuO) AFS was investigated using triple staining with DNA- and RNA-specific dyes. The effect of xylene deparaffinization on the acid fastness of mycobacteria in cultures or tissue sections was studied using AuO fluorescence as a quantitative marker. The xylene method was compared with a novel, solvent-free projected-hot-air deparaffinization (PHAD). RESULTS: Co-localization of AuO with DNA/RNA stains suggests that intracellular nucleic acids are the true target of AFS, producing highly specific patterns. Xylene reduces mycobacterial fluorescence significantly (P < .0001; moderate effect size, r = 0.33). The PHAD process yielded significantly higher fluorescence than xylene deparaffinization in tissues (P < .0001; large effect size, r = 0.85). CONCLUSIONS: Auramine O can be applied for nucleic acid staining of mycobacteria in tissues producing typical beaded patterns. Acid-fast staining depends heavily on the integrity of the mycobacterial cell wall, which xylene appears to damage. A solvent-free tissue deparaffinization method has the potential to increase mycobacterial detection significantly.

Human Gammaherpesvirus 8 Oncogenes Associated with Kaposi's Sarcoma.

Kaposi's sarcoma-associated herpesvirus (KSHV), also known as human gammaherpesvirus 8 (HHV-8), contains oncogenes and proteins that modulate various cellular functions, including proliferation, differentiation, survival, and apoptosis, and is integral to KSHV infection and oncogenicity. In this review, we describe the most important KSHV genes [ORF 73 (LANA), ORF 72 (vCyclin), ORF 71 or ORFK13 (vFLIP), ORF 74 (vGPCR), ORF 16 (vBcl-2), ORF K2 (vIL-6), ORF K9 (vIRF 1)/ORF K10.5, ORF K10.6 (vIRF 3), ORF K1 (K1), ORF K15 (K15), and ORF 36 (vPK)] that have the potential to induce malignant phenotypic characteristics of Kaposi's sarcoma. These oncogenes can be explored in prospective studies as future therapeutic targets of Kaposi's sarcoma.

A competitive family to the Beta and Kumaraswamy generators: Properties, Regressions and Applications.

We define two new flexible families of continuous distributions to fit real data by compoun-ding the Marshall-Olkin class and the power series distribution. These families are very competitive to the popular beta and Kumaraswamy generators. Their densities have linear representations of exponentiated densities. In fact, as the main properties of thirty five exponentiated distributions are well-known, we can easily obtain several properties of about three hundred fifty distributions using the references of this article and five special cases of the power series distribution. We provide a package implemented in R software that shows numerically the precision of one of the linear representations. This package is useful to calculate numerical values for some statistical measurements of the generated distributions. We estimate the parameters by maximum likelihood. We define a regression based on one of the two families. The usefulness of a generated distribution and the associated regression is proved empirically.

The COVID-19 Pandemic in Brazil: Some Aspects and Tools.

The article presents some aspects related to the COVID-19 pandemic in Brazil including public health, challenges facing healthcare workers and adverse impacts on the country's economy. Its main contribution is the availability of two web applications for online monitoring of the evolution of the pandemic in Brazil and South America. The applications provide the possibility to download data in different formats, view interactive maps and graphs of the cumulative confirmed cases, deaths and lethality rates, in addition to presenting plots of moving averages for states and municipalities. The predictions about new cases and new deaths caused by COVID-19, in states and regions of Brazil, are also reported using GAMLSS models. The forecasts can be easily used by public managers for effective decision-making.

Covid-19 in Brazil: A sad scenario.

The main contribution of this article is to report general statistics about COVID-19 in Brazil, based on analysis of accumulated series of confirmed cases, deaths and lethality rates, in addition to presenting graphs of moving averages for states and municipalities. The data show that the pandemic in Brazil has grown rapidly since February 25th (date of the first reported case). Furthermore, the lethality rate of COVID-19 in Brazil is greater than in many other Latin American countries (Chile, Argentina, Uruguay and Paraguay). However, the number of new confirmed cases in Brazil has little statistical relevance because only a small part of the population has been tested. In relation to Brazilian municipalities, we highlight the 10 states with the highest lethality rates, ranked from highest to lowest. Also, predictions about the increaseor decrease innew cases and deaths for states and capital cities are presented. These results can help managers and researchers to better guide their decisions regarding COVID-19.

The advertisement call of Boana freicanecae (Carnaval amp; Peixoto, 2004) (Anura: Hylidae), an endemic and poorly known treefrog from northeastern Brazil.

Boana freicanecae is a medium-sized hylid (males SVL= 37.3-42.2 mm) belonging to the B. pulchella group (Carnaval Peixoto 2004). This species is associated with streams in Atlantic montane forests (450-750 m elevation) located from the Pernambuco Biogeographic Sub-Region (Silva Casteleti 2003), in the states of Pernambuco and Alagoas, northeastern Brazil (Carnaval Peixoto 2004; Cardoso et al. 2006). Currently, B. freicanecae is classified as 'Data Deficient' according to the Brazilian Red List of Threatened species (ICMBio 2018) and many aspects of its natural history (e.g., advertisement call) are poorly known. Here we describe the advertisement call of this treefrog based on calls obtained from two populations.

Conspecific Presence Improves Episodic-Like Memory in Rats.

A number of studies have provided evidence that animals, including rats, remember past episodes. However, few experiments have addressed episodic-like memory from a social perspective. In the present study, we evaluated Wistar rats in the WWWhen/ELM task as single setups and in dyads, applying a long retention interval. We also investigated behaviors that could subserve the emergence of this type of memory. We found that only rats tested in the social setting were able to recollect an integrated episodic-like memory that lasted 24 h. Additionally, rats in dyads presented higher levels of exploration during the task. When exposed to the testing environment, the dyads exhibited affiliative behavior toward each other and presented fewer anxiety-like responses. Our findings indicate that the presence of a conspecific could act as a facilitating factor in memory evaluations based on spontaneous exploration of objects and provide empirical support for applying more naturalistic settings in investigations of episodic-like memory in rats.

A Study of the Disruptive Effect of the Acetate Fraction of Punica granatum Extract on Cryptococcus Biofilms.

Cryptococcosis, caused by yeasts of the genus Cryptococcus, is an infectious disease with a worldwide distribution. Cryptococcus neoformans and Cryptococcus gattii are the species that commonly cause this disease in humans; however, infections caused by Cryptococcus laurentii, especially in immunocompromised patients, are increasingly being reported. Owing to the increase in the resistance of fungi to antifungals, and a lack of treatment options, it is important to seek new therapeutic alternatives such as natural products. Among these are plant species such as Punica granatum, which is used in folk medicine to treat various diseases. This study aimed to evaluate the activity of the acetate fraction of P. granatum leaf extract against environmental and clinical isolates of Cryptococcus. Three environmental isolates of C. laurentii, PMN, PMA, and PJL II, isolated from soils of different municipalities in the state of Maranhão, a clinical isolate, C. gattii, from a patient with neurocryptococcosis, and a standard strain of C. gattii (ATCC 32068) were used. The minimum and fractional inhibitory concentrations (MIC and FIC, respectively) and time-kill curve of the extract and fluconazole were determined to assess the susceptibility profile of the fungal isolates. Larvae of Tenebrio molitor were infected with Cryptococcus strains, and the effects of acetate fraction of P. granatum extract and fluconazole on the survival and fungal burden were determined. The extract activity was tested against pre-formed biofilms. The acetate fraction of P. granatum extract showed promising antifungal activity against all the species of Cryptococcus evaluated in this study, with an MIC value lower than that of fluconazole. The indices obtained in the FIC test indicated that the antimicrobial effect of the combination of the extract and antifungal was indifferent for 80% of the isolates. The P. granatum acetate fraction reduced the pre-formed biofilm of some isolates, showing better activity than fluconazole, which is consistent with results from fluorescence microscopy. This is the first study on the use of P. granatum and its ability to inhibit Cryptococcus biofilms; therefore, further studies and tests are needed to investigate the components and mechanism of action of P. granatum against cryptococcosis agents.

AdequacyModel: An R package for probability distributions and general purpose optimization.

Several lifetime distributions have played an important role to fit survival data. However, for some of these models, the computation of maximum likelihood estimators is quite difficult due to presence of flat regions in the search space, among other factors. Several well-known derivative-based optimization tools are unsuitable for obtaining such estimates. To circumvent this problem, we introduce the AdequacyModel computational library version 2.0.0 for the R statistical environment with two major contributions: a general optimization technique based on the Particle Swarm Optimization (PSO) method (with a minor modification of the original algorithm) and a set of statistical measures for assessment of the adequacy of the fitted model. This library is very useful for researchers in probability and statistics and has been cited in various papers in these areas. It serves as the basis for the Newdistns library (version 2.1) published in an impact journal in the area of computational statistics, see https://CRAN.R-project.org/package=Newdistns. It is also the basis of the Wrapped library (version 2.0), see https://CRAN.R-project.org/package=Wrapped. A third package making use of the AdequacyModel library can be found in https://CRAN.R-project.org/package=sglg. In addition, the proposed library has proved to be very useful for maximizing log-likelihood functions with complex search regions. The library provides a greater control of the optimization process by introducing a stop criterion based on a minimum number of iterations and the variance of a given proportion of optimal values. We emphasize that the new library can be used not only in statistics but in physics and mathematics as proved in several examples throughout the paper.

A new class of lifetime models and the evaluation of the confidence intervals by double percentile bootstrap.

In this paper, we introduce a new three-parameter distribution by compounding the Nadarajah-Haghighi and geometric distributions, which can be interpreted as a truncated Marshall-Olkin extended Weibull. The compounding procedure is based on the work by Marshall and Olkin 1997. We prove that the new distribution can be obtained as a compound model with mixing exponential distribution. It can have decreasing, increasing, upside-down bathtub, bathtub-shaped, constant and decreasing-increasing-decreasing failure rate functions depending on the values of the parameters. Some mathematical properties of the new distribution are studied including moments and quantile function. The maximum likelihood estimation procedure is discussed and a particle swarm optimization algorithm is provided for estimating the model parameters. The flexibility of the new model is illustrated with an application to a real data set.

Advertisement calls, intraspecific variation and species diagnosis of six Brazilian species of Elachistocleis (Anura: Microhylidae: Gastrophryninae).

We describe for the first time the advertisement calls of three Elachistocleis species (E. surumu, E. magna, E. carvalhoi) and provide additional data on the calls of three other species (E. matogrosso, E. helianneae, E. muiraquitan). Additionally, we evaluate intraspecific variation in morphology and color patterns, and comment on species diagnosis in the genus. The advertisement calls of the six species of Elachistocleis have a stereotyped temporal and spectral structure, which consists of a long sustained multipulsed note with an upsweep in frequency at its onset, followed by a constant frequency along the note. Newly collected Elachistocleis from northern and western Brazil showed intraspecific variation in morphology and coloration that had not been previously reported. Coloration was listed as a diagnostic feature in the original descriptions of the Elachistocleis species evaluated in this study, but color patterns were variable in some species and sometimes did not allow for species differentiation. Our results indicated that the three species with greater degrees of morphological similarity (E. helianneae, E. matogrosso, E. muiraquitan) differed in microtemporal and spectral traits of their calls. Moreover, pulse duration in E. helianneae was very distinct, lasting approximately twice the average pulse duration in calls of the other species studied. Therefore, pulse duration might be an informative trait when assigning specimens to E. helianneae, which is widely distributed across Amazonian lowland forests and in a region of Amazonian savanna. Our results bring novelties on species diagnosis and intraspecific variation in Elachistocleis based on acoustic evidence and morphological/color patterns, and accentuate the importance of taking into consideration multiple lines of evidence as a means for accurate species-level identification in this genus of Neotropical frogs.