Abstract This study compares the effects of virus-cell interactions among SARS-CoV-2 variants of concern (VOCs) isolated in Brazil in 2021, hypothesizing a correlation between cellular alterations and mortality and between viral load and transmissibility. For this purpose, reference isolates of Alpha, Gamma, Zeta, and Delta variants were inoculated into monolayers of Vero-E6 cells. Viral RNA was quantified in cell supernatants by RT‒PCR, and infected cells were analyzed by Transmission Electron Microscopy (TEM) for qualitative and quantitative evaluation of cellular changes 24, 48, and 72 hours postinfection (hpi). Ultrastructural analyses showed that all variants of SARS-CoV-2 altered the structure and function of mitochondria, nucleus, and rough endoplasmic reticulum of cells. Monolayers infected with the Delta variant showed the highest number of modified cells and the greatest statistically significant differences compared to those of other variants. Viral particles were observed in the cytosol and the cell membrane in 100 % of the cells at 48 hpi. Alpha showed the highest mean particle diameter (79 nm), and Gamma and Delta were the smallest (75 nm). Alpha and Gamma had the highest particle frequency per field at 48 hpi, while the same was observed for Zeta and Delta at 72 hpi and 24 hpi, respectively. The cycle threshold of viral RNA varied among the target protein, VOC, and time of infection. The findings presented here demonstrate that all four VOCs evaluated caused ultrastructural changes in Vero-E6 cells, which were more prominent when infection occured with the Delta variant. viruscell virus SARSCoV2 SARSCoV SARS CoV 2 SARS-CoV- (VOCs 2021 transmissibility purpose VeroE6 VeroE Vero E6 E Vero-E RTPCR RT PCR RT‒PCR TEM (TEM 7 . (hpi) mitochondria nucleus 10 4 79 (7 nm, nm , nm) 75 nm. respectively protein VOC SARS-CoV 202 (hpi 1 ( 20

Abstract In 2015-2016, the Zika virus (ZIKV) caused a major epidemic in the Americas, increasing cases of microcephaly and Guillain-Barré syndrome. During this period, the discovery of ZIKV sexual transmission intensified studies on the impact of this virus on the reproductive organs. For this study, 2-month-old male BALB/c mice were infected with 1.26 x 106 PFU/mL of ZIKV in solution via the intravenous route. After three, seven, and fourteen days post-infection (DPI), blood and testicle samples were obtained to detect ZIKV RNA. The authors observed that the infected animals had slower weight gain than the control group. Viremia occurred only at 3DPI, and the ZIKV RNA was detected in one testis sample at 7DPI. The histopathological analysis of this organ revealed intense disorganization of the seminiferous tubules' structure, inflammatory infiltrate, necrosis, hemorrhage, fluid accumulation, congestion of blood vessels, and reduced sperm count. Ultrastructural analysis showed nuclear changes in tubule cells, activation of interstitial cells, and morphological changes in spermatozoa, in addition to fragmentation and decreased electron density of the genetic material of these cells. Thus, despite causing predominantly asymptomatic infections, ZIKV can cause significant subclinical and transient damage, including to male reproductive organs. 20152016, 20152016 2015 2016, 2016 2015-2016 (ZIKV Americas GuillainBarré Guillain Barré syndrome period organs study 2monthold monthold 2 month old BALBc BALB c 126 1 26 1.2 10 PFUmL PFU mL route three seven postinfection post infection DPI, DPI , (DPI) group 3DPI 7DPI tubules structure infiltrate necrosis hemorrhage accumulation vessels count cells spermatozoa Thus infections damage 2015201 201 2015-201 12 1. (DPI 201520 20 2015-20 20152 2015-2 2015-

BACKGROUND Dengue is a disease caused by dengue virus (DENV-1 through -4). Among the four serotypes, DENV-4 remains the least studied. Acute kidney injury is a potential complication of dengue generally associated with severe dengue infection. OBJECTIVES The goal of this study was to investigate the alterations caused by experimental dengue infection in the kidney of adult BALB/c mice. METHODS In this study, BALB/c mice were infected through the intravenous route with a DENV-4 strain, isolated from a human patient. The kidneys of the mice were procured and subject to histopathological and ultrastructural analysis. FINDINGS The presence of the viral antigen was confirmed through immunohistochemistry. Analysis of tissue sections revealed the presence of inflammatory cell infiltrate throughout the parenchyma. Glomerular enlargement was a common find. Necrosis of tubular cells and haemorrhage were also observed. Analysis of the kidney on a transmission electron microscope allowed a closer look into the necrotic tubular cells, which presented nuclei with condensed chromatin, and loss of cytoplasm. MAIN CONCLUSIONS Even though the kidney is probably not a primary target of dengue infection in mice, the inoculation of the virus in the blood appears to damage the renal tissue through local inflammation. DENV1 DENV 1 (DENV- 4. 4 . -4) serotypes DENV4 DENV- studied BALBc BALB c strain patient analysis immunohistochemistry parenchyma find observed chromatin cytoplasm inflammation (DENV -4 -

BACKGROUND According to the last 2023 Monkeypox (Mpox) Outbreak Global Map from the Centres for Disease Control and Prevention (CDC), more than 100 countries with no Mpox infection report cases. Brazil stands out in this group and is the second country with the highest number of cases in the last outbreak. OBJECTIVE To contribute to knowledge of the virus infection effects in a cellular model, which is important for diagnosis infections not yet included in a provider´s differential diagnosis and for developing viral inhibition strategies. METHODS We describe a virus isolation protocol for a human clinical sample from a patient from Brazil, the viral growth in a cell model through plaque forming units (PFU) assay, reverse transcriptase polymerase chain reaction (RT-PCR) and transmission electron microscopy (TEM). FINDINGS We follow the viral isolation in Vero cell culture from a Mpox positive clinically diagnosed sample and show the infection effects on cellular structures using a TEM. MAIN CONCLUSIONS Understanding the impact of viral growth on cellular structures and its replication kinetics may offer better strategies for the development of new drugs with antiviral properties. 202 (Mpox CDC, CDC , (CDC) 10 outbreak providers provider s PFU (PFU assay RTPCR RT PCR (RT-PCR TEM . (TEM) properties 20 (CDC 1 (TEM 2

BACKGROUND The coronaviruses (CoVs) called the attention of the world for causing outbreaks of severe acute respiratory syndrome (SARS-CoV), in Asia in 2002-03, and respiratory disease in the Middle East (MERS-CoV), in 2012. In December 2019, yet again a new coronavirus (SARS-CoV-2) first identified in Wuhan, China, was associated with a severe respiratory infection, known today as COVID-19. This new virus quickly spread throughout China and 30 additional countries. As result, the World Health Organization (WHO) elevated the status of the COVID-19 outbreak from emergency of international concern to pandemic on March 11, 2020. The impact of COVID-19 on public health and economy fueled a worldwide race to approve therapeutic and prophylactic agents, but so far, there are no specific antiviral drugs or vaccines available. In current scenario, the development of in vitro systems for viral mass production and for testing antiviral and vaccine candidates proves to be an urgent matter. OBJECTIVE The objective of this paper is study the biology of SARS-CoV-2 in Vero-E6 cells at the ultrastructural level. METHODS In this study, we documented, by transmission electron microscopy and real-time reverse transcription polymerase chain reaction (RT-PCR), the infection of Vero-E6 cells with SARS-CoV-2 samples isolated from Brazilian patients. FINDINGS The infected cells presented cytopathic effects and SARS-CoV-2 particles were observed attached to the cell surface and inside cytoplasmic vesicles. The entry of the virus into cells occurred through the endocytic pathway or by fusion of the viral envelope with the cell membrane. Assembled nucleocapsids were verified inside rough endoplasmic reticulum cisterns (RER). Viral maturation seemed to occur by budding of viral particles from the RER into smooth membrane vesicles. MAIN CONCLUSIONS Therefore, the susceptibility of Vero-E6 cells to SARS-CoV-2 infection and the viral pathway inside the cells were demonstrated by ultrastructural analysis.

Resumo: O conteúdo polínico de amostras de mel coletadas nos anos de 2017 e 2019 de apiários experimentais de Melipona seminigra pernigraMoure & Kerr 1950, instalado dentro de uma vegetação de campo rupestre em um afloramento de canga na Serra dos Carajás, sudeste da Amazônia, foi analisado para entender a variabilidade local dos recursos florais em áreas naturais e perturbadas. Aproximadamente 100% dos tipos polínicos foram identificados e pertencem principalmente às famílias Fabaceae, Myrtaceae e Euphorbiaceae (31, 6 e 5 espécies, respectivamente). Áreas de mineração apresentaram a maior riqueza de pólen, quase o dobro daquelas identificadas em áreas perturbadas. 80% dos tipos de pólen são raros com concentrações ≤ 2.000 grãos de pólen/10g, enquanto que os restantes foram os mais abundantes, frequentes e fontes primárias para as abelhas. Este últimos correspondem principalmente a plantas nativas como Tapirira guianensis Aubl., Protium spp., Aparisthmium cordatum (A.Juss.) Baill., Mimosa acutistipula var. ferrea Barneby, Periandra mediterrânea (Vell.) Taub., Miconia spp., Pleroma carajasense K.Rocha, Myrcia splendens (Sw.) DC., Serjania spp. e Solanum crinitum Lam. Todos os tipos polínicos foram identificados durante ambas as estações, mas altas concentrações estão relacionadas ao período seco (junho-setembro). A análise estatística indicou que não houve diferença significativa nos dados de pólen de mel entre áreas naturais e áreas anteriormente degradadas, uma vez que as fontes primárias das abelhas deste estudo são amplamente utilizadas na revegetação de áreas mineradas.

Abstract: Honey pollen samples of Melipona seminigra pernigraMoure & Kerr 1950 sampled between 2017 and 2019 from experimental apiaries installed in campo rupestre on canga (CRC) vegetation of the Serra dos Carajás aimed to evaluated seasonal floral availability of undisturbed and mining-influenced areas. Around one hundred pollen types were identified mainly belonging to Fabaceae, Myrtaceae and Euphorbiaceae (31, 6 and 5 species, respectively). Mining area presented the highest pollen richness, almost twice those identified in the undisturbed areas. 80% of the pollen types are rare with concentrations ≤ 2,000 pollen grains/10 g, while the remaining were the most abundant, frequent and the primary bee sources. These latter correspond mostly to native plants species such as Tapirira guianensis Aubl., Protium spp., Aparisthmium cordatum (A.Juss.) Baill., Mimosa acutistipula var. ferrea Barneby, Periandra mediterranea (Vell.) Taub., Miconia spp., Pleroma carajasense K.Rocha, Myrcia splendens (Sw.) DC., Serjania spp. and Solanum crinitum Lam. All pollen types were identified during both seasons, but higher concentration values are related to the dry period (June-September). The statistical analysis of the pollen data indicated that there was no significant difference between undisturbed and mining-influenced areas, since primary bee sources of this study are widespread used in revegetation of mined areas.

BACKGROUND The impact of arbovirus cocirculation in Brazil is unknown. Dengue virus (DENV) reinfection may result in more intense viraemia or immunopathology, leading to more severe disease. The Zika virus (ZIKV) epidemic in the Americas provided pathogenicity evidence that had not been previously observed in flavivirus infections. In contrast to other flaviviruses, electron microscopy studies have shown that ZIKV may replicate in viroplasm-like structures. Flaviviruses produce an ensemble of structurally different virions, collectively contributing to tissue tropism and virus dissemination. OBJECTIVES AND METHODS In this work, the Aedes albopictus mosquito cell lineage (C6/36 cells) and kidney epithelial cells from African green monkeys (Vero cells) were infected with samples of the main circulating arboviruses in Brazil [DENV-1, DENV-2, DENV-3, DENV-4, ZIKV, Yellow Fever virus (YFV) and Chikungunya virus (CHIKV)], and ultrastructural studies by transmission electron microscopy were performed. FINDINGS We observed that ZIKV, the DENV serotypes, YFV and CHIKV particles are spherical. ZIKV, DENV-1, -2, -3 and -4 presented diameters of 40-50 nm, and CHIKV presented approximate diameters of 50-60 nm. Viroplasm-like structures was observed in ZIKV replication cycle. MAIN CONCLUSIONS The morphogenesis of these arboviruses is similar to what has been presented in previous studies. However, we understand that further studies are needed to investigate the relationship between viroplasm-like structures and ZIKV replication dynamics.

RESUMO O trabalho teve como objetivo identificar e descrever a morfologia de microfósseis não polínicos ocorrentes nos sedimentos de um testemunho retirado próximo à barra da Lagoa Comprida, Estado do Rio de Janeiro, Brasil. Foram identificadas oito formas de fungos, dois tipos de algas Chlorococcales em estágio colonial, um tipo de zygósporo de alga Zygnematales, quatro de diatomáceas, um de microforaminífero, dois de cistos de dinoflagelados, um de microesclera de Porifera, um de esporo de briófita e nove de esporos de samambaias e licófitas. Informações ecológicas sobre os organismos de origem foram fornecidas para prover material de referência para a reconstrução paleoambiental ao longo do Holoceno em áreas de restinga.

ABSTRACT The aim of this work was to identify and describe the morphology of non-pollen microfossils present in the sediments of a core collected near the Lagoa Comprida barrier, Rio de Janeiro State, Brazil. Eight forms of fungi, two types of Chlorococcales algae at the colonial stage, one zygospore of Zygnematales, four of diatoms, one of microforaminifer, two cysts of dinoflagellates, one spicule of Porifera, one spore of bryophyte, and nine spores of ferns and lycophytes were identified. Ecological information on the organisms of origin was given to provide reference material for paleoenvironmental reconstruction along the Holocene in restinga areas.

The lack of an experimental animal model for the study of dengue pathogenesis is a limiting factor for the development of vaccines and drugs. In previous studies, our group demonstrated the susceptibility of BALB/c mice to infection by dengue virus (DENV) 1 and 2, and the virus was successfully isolated in several organs. In this study, BALB/c mice were experimentally infected intravenously with DENV-4, and samples of their saliva were collected. Viral RNA extracted from the saliva samples was subjected to qRT-PCR, with a detection limit of 0.002 PFU/mL. The presence of DENV-4 viral RNA was detected in the saliva of two mice, presenting viral titers of 109 RNA/mL. The detection of DENV RNA via saliva sampling is not a common practice in dengue diagnosis, due to the lower detection rates in human patients. However, the results observed in this study seem to indicate that, as in humans, detection rates of DENV RNA in mouse saliva are also low, correlating the infection in both cases. This study reports the first DENV detection in the saliva of BALB/c immunocompetent mice experimentally infected with non-neuroadapted DENV-4.

Zika virus (ZIKV) has infected thousands of Brazilian people and spread to other American countries since 2015. The introduction of ZIKV brought a strong impact to public health in Brazil. It is of utmost importance to identify a susceptible cell line that will enable the isolation and identification of the virus from patient samples, viral mass production, and testing of drug and vaccine candidates. Besides real-time reverse transcriptase polymerase chain reaction diagnosis for detecting the viral genome, virus isolation in cell lines was useful in order to study the structure of the viral particle and its behaviour inside cells. Analysis of ZIKV infected cell lines was achieved using transmission electron microscopy (TEM). Blood was obtained from a Brazilian patient during the first days after presenting with signs of the disease, and ZIKV from the patient’s blood was isolated in the C6/36 mosquito cell line. Afterwards, Vero cells were inoculated with the viral suspension, fixed six days after inoculation, embedded in polymers, and ultra-thin cut. Like dengue viruses, this flavivirus showed numerous virus particles present inside cellular vesicles thereby confirming the susceptibility of the Vero cell line to ZIKV replication. TEM is a unique technique available to make the virus visible.

In cell culture, cell structures suffer strong impact due to centrifugation during processing for electron microscope observation. In order to minimise this effect, a new protocol was successfully developed. Using conventional reagents and equipments, it took over one week, but cell compression was reduced to none or the lowest deformation possible.

Abstract Pollen is used in the human diet as a food supplement because of its high nutritional value; however, this product is prone to fungal contamination that could potentially generate toxins that are harmful to human health. This study aimed to verify the floral diversity of commercial brands of bee pollen and their mycotoxicological safety for human consumption. A total of 27 bee pollen samples were analyzed; these samples represented commercial brands, either showing an inspection seal or not, marketed in the State of Rio de Janeiro. The analyzed parameters included floral diversity through palynological analysis, water activity, fungal counts, identification and toxigenic profiles. The palynological analysis identified nine plant families, of which the Asteraceae was predominant. Analysis of hygienic quality based on fungal load showed that 92% of samples were reproved according to the commercial, sanitary, and food safety quality indicators. Aspergillus, Cladosporium and Penicillium were the most common genera. Toxigenic evaluation showed that 25% of the A. flavus strains produced aflatoxins. The high rate of contamination of products bearing an inspection seal emphasizes the need to monitor the entire procedure of bee pollen production, as well as to revise the current legislation to ensure safe commercialization of this product.

A própolis é uma substância resinosa coletada de diversas partes das plantas por abelhas e misturada à cera, pólen e secreções salivares. Sua composição é complexa e está relacionada, principalmente, às características da vegetação de cada região. Trinta e três amostras de própolis foram coletadas em quatro regiões brasileiras (Nordeste, Sudeste, Sul e Centro-Oeste) e analisadas quanto à sua capacidade antioxidante, pela oxidação acoplada do sistema β-caroteno/ácido linoleico. Variação significativa na capacidade antioxidante (51,33-92,70%) foi observada de acordo com a região onde a amostra foi coletada. A capacidade antioxidante das amostras de própolis da região Nordeste foi menor que a das amostras das regiões Centro-Oeste, Sul e Sudeste, respectivamente.

Propolis is a resinous substance collected by bees from many parts of plants and mixed with wax, pollen and salivary secretions. Its composition is complex and closely related to the vegetation features of each region. Thirty-three propolis samples were collected from four Brazilian regions (Northeast, Southeast, South and Central-West) and had their antioxidant activity analyzed by the connected oxidation of b-carotene/linoleic acid. A significant variation was observed for the antioxidant activity (51.33-92.70%), according to the region where the sample was collected. The antioxidant activity of propolis samples collected in the Northeast region was lower than those collected in the Central-West, South and Southeast regions, respectively.

Fourier transform infrared attenuated total reflectance (FT-IR ATR) spectroscopy was used to determine 14 different measurands in northeast Brazilian honey samples. Nine different honey samples (six monofloral and three polyfloral) from 2009 obtained from the company CEARAPI underwent FT-IR ATR, palynological, color, and sensorial analysis to obtain preliminary results for these types of honey. The results showed that there are five monofloral, three bifloral, and one extrafloral honey, and also that mid-infrared spectrometry can be used as a screening method for the routine analysis of Brazilian honey, with the advantages of being rapid, nondestructive, and accurate.

O uso de microrganismos patogênicos em atos de bioterrorismo é já há algum tempo objeto de grande preocupação em vários países. O presente trabalho apresenta a possível aplicação de vírus e bactérias para fins bélicos e terroristas, bem como o diagnóstico laboratorial para a identificação desses agentes. Foram salientados, entre outros, como agentes de infecções humanas visando o bioterrorismo, os vírus da varíola (ortopoxvírus), os de febres hemorrágicas e os pertencentes aos filovírus. Entre as bactérias foram destacadas as do antrax ( Bacillus anthracis ), da peste ( Yersinia pestis ), do botulismo ( Clostridium botulinum ) e da tularemia ( Francisella tularensis ), incluindo ainda a ricina ( Ricinus communis ) como componente do grupo B de agentes.

In recent years the use of pathogenic microorganisms in acts of bioterrorism has been the subject of major concern in many countries. This paper presents a possible application of viruses and bacteria for warfare and terrorist purposes, as well as a laboratory diagnosis to identify those agents. The viruses of smallpox (orthopoxvirus), of hemorrhagic fever and those belonging to filovirus have been highlighted, inter alia, as agents of human infection with bioterrorist intent. Among the bacteria, the emphasis has been on anthrax (Bacillus anthracis), the plague (Yersinia pestis), botulism (Clostridium botulinum) and tularemia (Francisella tularensis), not to mention ricin (Ricinus communis), as one of the Group B agents.