Abstract Thirteen Haemophylus influenzae invasive strains isolated from patients at Clinical Hospital of State University of Campinas, from May 2013 through August 2019, was submitted to Illumina genome sequencing HiSeq platform. Further in silico analysis of serogroup and Multi Locus Sequence Typing (MLST) from whole DNA sequencing had demonstrated the actual clonal distribution in the Campinas Metropolitan region. Thus, results showed the existence of a new ST Haemophilus influenzae found in the Brazilian territory and an increase of strains belonging to serogroup a (three strains also belonging to ST23). In conclusion, we observed an increase of non-typable H. influenzae (NTHi) and a strain involved in invasive diseases in the Campinas – São Paulo region after frequent detection of those serotypes and genotypes in other Brazilian regions.

ABSTRACT Acrostichum danaeifolium, a Neotropical fern, occurs preferentially in marshy areas or at the margins of lakes and mangroves. Microlepidoptera larvae burrow through the petioles of the fern, preferentially on the non-expanded leaves. The galleries in the petiole create a new microhabitat, harboring a rich fauna of arthropods. The aim of the present study was to assess the richness of ants associated with its petiole. The study was conducted in a population of A. danaefolium from the Atlantic Forest in Rio de Janeiro state, Southeastern Brazil. Six collections were carried out every two months (2009-2010), three in the dry and three in the rainy season. The leaves were divided into three development stages: non-expanded leaves (NEL), expanded leaves (EL) and senescent leaves (SL). Seven leaves from each phase were randomly collected from seven individuals. A total of fifteen ant species were recorded. The species with the highest frequency and density in fern petioles were Camponotus crassus and Crematogaster curvispinosa. The highest ant richness and abundance was found in senescent leaves. The high number of ants found in the petioles of Acrostichum danaefolium qualifies it as a potential key species in the marshes and flooded areas where it occurs.

Abstract: Pteridium is a cosmopolitan genus of ferns that possess nectaries on its fronds (fern leaves), thereby attracting ants. Foliar (or extrafloral) nectaries are nectar-producing glands that are not related to pollination, but rather attract ants and other arthropods. Foliar nectaries are found in 101 fern species, belonging to 11 genera and six families. The aim of the study is to characterize the community of ants that visit the fronds of Pteridium esculentum subsp. arachnoideum, as well as daily and seasonal ant abundance in different frond development stages. The study was conducted in the Atlantic Forest of Rio de Janeiro state, Brazil. Bimonthly collections were established, where 30 expanding fronds and 30 fully expanded fronds were randomly marked. In each 1-hour shift starting at 8:30 am and ending at 5:30 pm, the fronds were observed for the presence of ants. Thirty three ant species were recorded on the Pteridium esculentum subsp. arachnoideum fronds, distributed into six subfamilies and 13 genera. The most abundant species were Solenopsis sp.1 and Ectatomma tuberculatum. Eight ant species were observed foraging the nectaries of tropical bracken fern fronds. Ectatomma tuberculatum has been observed feeding on the nectaries and patrolling the fronds. Ant activity peak was on mid-day during the rainy season. The tropical bracken fern Pteridium esculentum subsp. arachnoideum has a rich (the highest recorded until now on Pteridium species) and diverse ant community on its fronds, mainly on the expanding fronds. The presence of generalist predatory ants (Ectatomma tuberculatum and Solenopsis sp.1) during the entire study period suggests a positive interaction between ants and Pteridium esculentum subsp. arachnoideum.

Resumo: Pteridium é um gênero cosmopolita de samambaias que possui nectários em suas frondes (folhas de samambaias), atraindo formigas. Nectários foliares (ou extraflorais) são glândulas produtoras de néctar que não estão relacionadas com a polinização, mas podem atrair formigas e outros artrópodes. Nectários foliares já foram registrados em 101 espécies de samambaias, pertencentes a 11 gêneros e seis famílias. O objetivo do estudo é caracterizar a comunidade de formigas que visitam as frondes de Pteridium esculentum subsp. arachnoideum, bem como a abundância diária e sazonal das formigas em diferentes estágios foliares. O estudo foi realizado na Mata Atlântica do estado do Rio de Janeiro, Brasil. Foram estabelecidas coletas bimestrais, onde foram marcadas aleatoriamente 30 frondes em expansão e 30 frondes totalmente expandidas. Em cada turno de 1 hora com início às 8h30 e término às 17h30, as frondes marcadas foram observadas quanto à presença de formigas. Trinta e três espécies de formigas foram registradas nas frondes do Pteridium esculentum subsp. arachnoideum, distribuídas em seis subfamílias e 13 gêneros. As espécies mais abundantes foram Solenopsis sp.1 e Ectatomma tuberculatum. Foram registradas oito espécies de formigas forrageando os nectários foliares da samambaia. Ectatomma tuberculatum foi observada se alimentando nos nectários e patrulhando as frondes. As formigas tiveram o pico de atividade ao meio-dia e na estação chuvosa. A samambaia tropical Pteridium esculentum subsp. arachnoideum tem uma elevada riqueza (a maior já registrada até o momento para espécies de Pteridium) e diversidade de formigas em suas frondes, com maior frequência nas frondes não totalmente expandidas. A presença de formigas predadoras generalistas (Ectatomma tuberculatum e Solenopsis sp.1) durante todo o período de estudo sugere uma interação positiva entre as formigas e Pteridium esculentum subsp. arachnoideum.

Abstract Brazilian Purpuric Fever (BPF) is a hemorrhagic pediatric illness caused by Haemophilus influenzae biogroup aegyptius (Hae), a bacterium that was formerly associated with self-limited purulent conjunctivitis. BPF is assumed to be eradicated. However, the virulence mechanisms inherent to Hae strains associated with BPF is still a mystery and deficient in studies. Here, we aim to analyze the role of the autotransporter genes related to adherence and colonization las, tabA1, and hadA genes through RT-qPCR expression profiling and knockout mutants. Relative quantification by real-time PCR after infection in human cells and infant rat model suggests that las was initially downregulated probably duo to immune evasion, tabA1, and hadA were overexpressed in general, suggesting an active role of TabA1 and HadA1 adhesins in Hae in vitro and in vivo. Transformation attempts were unsuccessful despite the use of multiple technical approaches and in silico analysis revealed that Hae lacks genes related to competence in Haemophilus, which could be part of the elucidation of the difficulty of genetically manipulating Hae strains.

Abstract S. pneumoniae, commonly known as pneumococcus, is a naturally competent Gram-positive bacterium and is the major cause of pneumonia in elderly and children in developing countries. This pathogen is associated with respiratory diseases affected by pollution. The objective of this work was determining the effect of ash and environmental dust from the burning of sugarcane on pneumococci bacterial transformation. The transformation capacity of the Pn360 pneumococci strain was performed using the assays of DNA donor of mutant for luxS gene. Thus, the transformation tests were performed in contact with dust collected in the southwestern region of Brazil (important region where burning of sugar cane is present in the agriculture). The use of degradative practices in the sugar cane agriculture in Brazil was involved in the transformation capacity of the S. pneumoniae. This phenomenon includes important consequences for public health concerning to resistance acquisition and new virulence factors of this important infection. In conclusion, we obtained important results concerning the action of environmental pollution in Streptococcus pneumoniae transformation, increasing the DNA acquisition for this pathogen.

Glicosídeos cianogênicos são substâncias de defesa que são capazes de produzir ácido cianídrico quando sofrem hidrólise em consequência da herbivoria, processo chamado de cianogênese. Galhas são estruturas neoformadas de tecidos vegetais geradas por interações espécie-específica entre um organismo indutor e uma planta hospedeira. Estudos realizados demonstraram que há um polimorfismo cianogênico em espécies de Microgramma. Microgramma squamulosa é uma samambaia epífita que pode conter galhas caulinares induzidas por Tortrimosaica polypodivora (Lepidoptera: Tortricidae). Desse modo, o objetivo do presente trabalho foi realizar uma prospecção da cianogênese nos tecidos do caule, folhas e galhas de Microgramma squamulosa. O estudo foi realizado em populações de M. squamulosa localizadas no estado do Rio de Janeiro, Brasil. A cianogênese foi avaliada pelo teste de papel Feigl-Anger. Foram analisadas 260 amostras de tecidos galhados e não galhados, sendo 45 amostras de galhas, 67 de folhas estéreis, 43 de folhas férteis, 103 de caules e 2 de báculos. A cianogênese foi detectada em apenas três amostras de folhas estéreis (4,5%). Em nenhuma das amostras o caule ou as galhas foram cianogênicas. A frequência da cianogênese não diferiu significativamente entre as folhas estéreis, folhas férteis, caules, galhas e báculos.

Cyanogenic glycosides are defense substances that can produce hydrocyanic acid when they undergo hydrolysis as a result of herbivory, a process called cyanogenesis. Galls are neoformed structures of plant tissues induced by species-specific interactions between an inducer organism and a host plant. Earlier studies in Microgramma species have demonstrated cyanogenic polymorphism. Microgramma squamulosa is an epiphytic fern that may contain stem galls induced by Tortrimosaica polypodivora (Lepidoptera: Tortricidae). Thus, the objective of present study was to conduct a cyanogenesis prospection in the tissues of the stem, leaves and galls of Microgramma squamulosa. The study was conducted in populations located in the Rio de Janeiro state, Brazil. Cyanogenesis was assessed using the Feigl-Anger paper test. A total of 260 galled and non-galled tissues were analyzed, 45 gall samples, 67 sterile leaves, 43 fertile leaves, 103 stems and 2 croziers. Cyanogenesis was detected in only three sterile leaf samples (4.5%). In none of the samples were the stems or galls cyanogenic. Cyanogenesis frequency did not differ significantly between sterile leaves, fertile leaves, stems, galls and croziers.

Abstract Galls are neoformed plant structures created by cell hyperplasia and hypertrophy induced by a number of organisms, especially insects. After adult insects hatch, senescent galls may remain on the host plant and be occupied by a succession of fauna, the most important and dominant being ants. This study aimed at characterizing the ant fauna successor of stem galls induced by microlepidoptera in Microgramma squamulosa (Kaulf.) de la Sota (Polypodiaceae). Four collections were carried out in the municipality of Nova Friburgo, Rio de Janeiro state, Brazil. The galls were packed in plastic bags and taken to the laboratory. Ants were euthanized and conserved in 70° GL alcohol and later identified. A total of 49 stem galls were collected and analyzed, 15 containing microlepidoptera galler larvae, one a parasitoid wasp and 33 without the microlepidoptera or parasitoid (67%). Twelve of these galls (39%) contained ants. Six ant species were recorded (Camponotus crassus, Crematogaster curvispinosa, Crematogaster sericea, Procryptocerus sampaioi, Tapinoma atriceps, and Wasmannia auropunctata), all native to Brazil. Ant occupation in M. squamulosa seems to be associated with senescent galls due to hatching of the galler insect, which leaves a hole that allows ants to colonize it, in other words, an opportunistic domatia. Senescent galls resulting from the death of galler insects do not seem to facilitate ant occupation.

Nanotechnology is currently one of the fastest growing scientific fields. The products of this science have become part of our everyday lives. However, to date, regulatory agencies have not yet established a single definition for nanomaterials and nanotechnology. Therefore, each country has its own definitions and legislation to control products containing nanomaterials. Being relatively new materials, there are no long-term studies showing their impact on human health and the environment. Consequently, countries control the amount of nanomaterials present in cosmetics, allowing the end consumer to choose which cosmetic to use, by choosing products with or without nanomaterials. Therefore, the primary objective of this study was to identify the most used nanomaterials in cosmetics and verify whether these formulations are in accordance with the laws in force in the United States, the European Union and Brazil, thereby determining if the cosmetics on the market are in line with the existing laws in these three economic powers. This study is unique and will contribute to furthering the discussion on existing laws pertinent to the use of nanotechnology in cosmetics.

The Brazilian Purpuric Fever (BPF) is a systemic disease with many clinical features of meningococcal sepsis and is usually preceded by purulent conjunctivitis. The illness is caused by Haemophilus influenza biogroup aegyptius, which was associated exclusively with conjunctivitis. In this work construction of the las gene, hypothetically responsible for this virulence, were fusioned with ermAM cassette in Neisseria meningitidis virulent strains and had its DNA transfer to non BPF H. influenzae strains. The effect of the las transfer was capable to increase the cytokines TNFα and IL10 expression in Hec-1B cells line infected with these transformed mutants (in eight log scale of folding change RNA expression). This is the first molecular study involving the las transfer to search an elucidation of the pathogenic factors by horizontal intergeneric transfer from meningococci to H. influenzae.

One hundred and fifty-one methicillin-resistant z (MRSA) strains have been isolated from patients admitted in tertiary care hospitals in two metropolitan areas (Campinas City and Ribeirão Preto City) in the southeast region of Brazil and analyzed through PCR-based techniques [(PCR amplification of spa, coa, and housekeeping genes (arcC, aroE, gmk, pta, tpi, yqiL)] and further restriction fragment typing of coa and of housekeeping genes. The heterogeneity of spa gene was determined directly by agarose gel electrophoresis migration. The results obtained indicate the existence of three (A, B, C) main clusters. Since the strain distribution in these three clusters is much characteristic, it denotes the existence of three main clones. All strains isolated in Campinas were grouped in clusters A and B, while most of the strains isolated in Ribeirão Preto were grouped in cluster C. This distribution denotes the existence of different founder strains that undergo independent genetic variability. The strains considered representative of the Brazilian Epidemic Clone (BEC) were categorized as cluster A. These results indicate a possible higher variability among Brazilian MRSA strains than currently described and indicate that the techniques herein used can be used as an alternative to Pulsed Field Gel Electrophoresis (PFGE).

Several pathogenic or opportunistic bacteria have the ability to either induce or inhibit host cell apoptosis. The capacity to modulate cell pathways that result in the induction or delay of host cell apoptosis is considered to be an important bacterial virulence mechanism. These processes could be mediated by different host cell signaling pathways that are subverted by the bacteria. Pathogens are able to activate apoptotic proteins, such as caspases, or inactivate anti-apoptotic proteins, such as NFkB and the MAPKKs, or even up-regulate the endogenous receptor/ligand system that induces apoptosis, generally when the bacteria are bound to the host cell surface. The bacteria-induced apoptotic or anti-apoptotic processes are often related with the fact that the bacteria acquire the ability to reach the host tissues. However, apoptosis is also considered to be a host defense mechanism against infectious agents. Thus, the apoptosis phenomenon plays a central role in host-pathogen interactions.

Forty-five Haemophilus influenzae strains isolated from patients were characterized based on biochemical characteristics. Their capsular types were determined by polymerase chain reaction (PCR); they were compared, using two molecular methods [ribotyping with a specific DNA probe amplified from the 16S rDNA region from H. influenzae and through restriction fragment length polymorphism (RLFP) of an amplified 16S DNA region]. The strains were better discriminated by the ribotyping technique that used the 16S probe and by the combination of both techniques. Biotypes I and IV were the most common, followed by biotypes VI, VIII and III. Biotypes II and VII were not found. Most of the capsular samples were nontypable (89%), with capsular types a and b found in 2 and 9% of the samples, respectively. We concluded that there is a very close genetic identity among pathogenic and non-pathogenic strains.

A técnica de REP (Repetitive extragenic palindrome)-PCR foi utilizada para avaliar a variabilidade genética de 49 amostras de Escherichia coli patogênicas para aves (APEC), isoladas de aves de corte (frangos) em diferentes surtos de septicemia (n=24), síndrome da cabeça inchada (n=14) e onfalite (n=11). Trinta amostras comensais, isoladas de frangos sem sinais de doença, foram utilizadas como controle. A análise do perfil eletroforético obtido por reação de REP-PCR utilizando DNA purificado das amostras evidenciou a amplificação de 0 a 15 bandas de DNA com pesos moleculares variando entre 100 pb e 6.1 Kb. A análise deste padrão permitiu a construção de um dendrograma demonstrando o agrupamento das 79 amostras em 49 perfis distintos. Embora a técnica de REP-PCR tenha apresentado grande poder discriminatório, as amostras patogênicas e não patogênicas não foram discriminadas entre si assim como não foi observado o agrupamento de amostras causadoras do mesmo tipo de doença. Por outro lado, demonstramos recentemente que outras técnicas tais como ERIC-PCR e a análise de isoenzimas foram eficientes quando utilizadas para esta mesma finalidade. Concluindo, REP-PCR parece não ser uma técnica eficiente e universal para discriminar entre amostras APEC. Porém, a estrutura clonal populacional obtida com o uso de REP-PCR não deve ser desprezada, particularmente se considerarmos que os mecanismos de patogenicidade de APEC ainda não são completamente conhecidos.

In the present study the repetitive extragenic palindromic (REP) polymerase chain reaction (PCR) technique was used to establish the clonal variability of 49 avian Escherichia coli (APEC) strains isolated from different outbreak cases of septicemia (n=24), swollen head syndrome (n=14) and omphalitis (n=11). Thirty commensal strains isolated from poultry with no signs of these illnesses were used as control strains. The purified DNA of these strains produced electrophoretic profiles ranging from 0 to 15 bands with molecular sizes varying from 100 bp to 6.1 kb, allowing the grouping of the 79 strains into a dendrogram containing 49 REP-types. Although REP-PCR showed good discriminating power it was not able to group the strains either into specific pathogenic classes or to differentiate between pathogenic and non-pathogenic strains. On the contrary, we recently demonstrated that other techniques such as ERIC-PCR and isoenzyme profiles are appropriate to discriminate between commensal and APEC strains and also to group these strains into specific pathogenic classes. In conclusion, REP-PCR seems to be a technique neither efficient nor universal for APEC strains discrimination. However, the population clonal structure obtained with the use of REP-PCR must not be ignored particularly if one takes into account that the APEC pathogenic mechanisms are not completely understood yet.

O objetivo deste trabalho foi estudar características biológicas tais como a expressão de fímbrias e adesinas, capacidade de absorção do corante Vermelho Congo, produção de hemolisina e aerobactina, adesão e invasão a células HeLa e adesão a células do epitélio urinário em amostras de Eschericia coli isoladas de pacientes com sinais clínicos de infecção do trato urinário (UTI). A presença dos genes responsáveis pela expressão de fímbrias (apa, afa e spa) e das Citotoxinas Necrotizantes CNF1, CNF2 foi avaliada por PCR. Esses dados foram comparados com a estrutura clonal das amostras obtidas por análises de isoenzimas (MLEE), Ribotipagem (RFLP) e ERIC-PCR. Com uma única exceção, os isolados apresentaram combinação de ao menos duas das características estudadas, fato que sugere a existência de Ilhas de Patogenicidade (PAIs). A maioria das amostras apresentaram um padrão difuso de aderência a células HeLa. Os resultados indicam que a capacidade de adesão a células epiteliais do sistema urinário poderia ser um teste mais específico e correlacionado à patogenicidade. Embora os estudos com microscopia óptica indicassem que quatro linhagens pudessem ser invasivas, dados de microscopia eletrônica não confirmaram tais achados. As técnicas de MLEE, Ribotipagem e ERIC-PCR separaram os isolados em diferentes grupos principais mas estes não foram correlacionados à patogenicidade.

The aim of the present study was to determine biological characteristics such as expression of fimbriae, Congo red binding, production of hemolysin and aerobactin, adhesion to HeLa and uroepithelial cells and invasion of HeLa cells by Escherichia coli isolates obtained from patients showing clinical signs of urinary tract infection (UTI). Also, the presence of genes (apa, afa, spa) for fimbria expression and cytotoxic necrotizing factors (CNF1, CNF2) was assayed using specific primers in PCR. The data obtained were compared with the clonal relationships obtained by analysis of multilocus enzyme electrophoresis (MLEE), restriction fragment length polymorphism (RFLP) of the rDNA (ribotyping) and enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR). All isolates but one presented a combination of at least two of the characteristics studied, a fact suggesting the presence of pathogenicity islands (PAIs). Diffuse adherence type to HeLa cells was observed to occur in most of the strains, but adhesion to uroepithelial cells seems to be a more reliable test to verify pathogenicity. Although four strains seemed to be able to invade HeLa cells when assayed by light microscopy, electron microscopy studies demonstrated that these strains were not invasive. MLEE, RFLP and ERIC-PCR were able to group the isolates differently into main clusters that were not correlated with the presence of pathogenic traits.