Gene Signatures of Symptomatic and Asymptomatic Clinical-Immunological Profiles of Human Infection by Leishmania (L.) chagasi in Amazonian Brazil.

Individuals infected with Leishmania (L.) chagasi may present different asymptomatic and symptomatic stages of infection, which vary in the clinical-immunological profiles that can be classified as asymptomatic infection (AI), subclinical resistant infection (SRI), indeterminate initial infection (III), subclinical oligosymptomatic infection (SOI), and symptomatic infection (SI) (=American visceral leishmaniasis, AVL). However, little is known about the molecular differences between individuals having each profile. Here, we performed whole-blood transcriptomic analyses of 56 infected individuals from Pará State (Brazilian Amazon), covering all five profiles. We then identified the gene signatures of each profile by comparing their transcriptome with those of 11 healthy individuals from the same area. Symptomatic individuals with SI (=AVL) and SOI profiles showed higher transcriptome perturbation when compared to those asymptomatic III, AI and SRI profiles, suggesting that disease severity may be associated with greater transcriptomic changes. Although the expression of many genes was altered on each profile, very few genes were shared among the profiles. This indicated that each profile has a unique gene signature. The innate immune system pathway was strongly activated only in asymptomatic AI and SRI profiles, suggesting the control of infection. In turn, pathways such as MHC Class II antigen presentation and NF-kB activation in B cells seemed to be specifically induced in symptomatic SI (=AVL) and SOI profiles. Moreover, cellular response to starvation was down-regulated in those symptomatic profiles. Overall, this study revealed five distinct transcriptional patterns associated to the clinical-immunological (symptomatic and asymptomatic) profiles of human L. (L.) chagasi-infection in the Brazilian Amazon.

Molecular tools confirm natural Leishmania (Viannia) guyanensis/L. (V.) shawi hybrids causing cutaneous leishmaniasis in the Amazon region of Brazil.

Seven isolates from patients with American cutaneous leishmaniasis in the Amazon region of Brazil were phenotypically suggestive of Leishmania (Viannia) guyanensis/L. (V.) shawi hybrids. In this work, two molecular targets were employed to check the hybrid identity of the putative hybrids. Heat shock protein 70 (hsp70) gene sequences were analyzed by three different polymerase chain reaction (PCR) approaches, and two different patterns of inherited hsp70 alleles were found. Three isolates presented heterozygous L. (V.) guyanensis/L. (V.) shawi patterns, and four presented homozygous hsp70 patterns involving only L. (V.) shawi alleles. The amplicon sequences confirmed the RFLP patterns. The high-resolution melting method detected variant heterozygous and homozygous profiles. Single-nucleotide polymorphism genotyping/cleaved amplified polymorphic site analysis suggested a higher contribution from L. (V.) guyanensis in hsp70 heterozygous hybrids. Additionally, PCR-RFLP analysis targeting the enzyme mannose phosphate isomerase (mpi) gene indicated heterozygous and homozygous cleavage patterns for L. (V.) shawi and L. (V.) guyanensis, corroborating the hsp70 findings. In this communication, we present molecular findings based on partial informative regions of the coding sequences of hsp70 and mpi as markers confirming that some of the parasite strains from the Brazilian Amazon region are indeed hybrids between L. (V.) guyanensis and L. (V.) shawi.

Value of the oral swab for the molecular diagnosis of dogs in different stages of infection with Leishmania infantum.

This study was based on the need to employ a sensitive and specific method with samples that could be easily collected for diagnosing dogs infected with Leishmania infantum. To this end, we used real time-PCR (qPCR) to assess the value of the oral swab (OS) in detecting infected sick dogs (SD; n=62), including, for the first time, the analysis of apparently healthy infected dogs (AD; n=30), both from endemic areas for visceral leishmaniasis (VL). For comparison, we also evaluated the performance of the conjunctival swab (CS), blood (BL), lymph node (LN) and serology. We detected the presence of Leishmania DNA in the oral cavity in 62 out of the 92 dogs studied. The OS positivity (67.4%) was equivalent to the CS (68.5%) (p>0.05), higher than BL (52.2%) (p≤0.05), and lower than LN (84.8%) (p≤0.05). OS and CS performed well in SD dogs (82.3% and 83.9%, respectively) but not in AD dogs (36.7% for both samples). BL showed the lowest positivity (52.2%) and provided equivalent results between AD (60.0%) and SD (48.4%) dogs (p>0.05). LN yielded the highest positivity (84.8%), and it was also higher in the SD population (93.5%) compared to the AD population (66.7%) (p≤0.05). Parasite load was high in LN, moderate in OS and CS, and low in BL, showing the relationship between the levels of parasitism and the positivity rates found in these samples. Serology was positive in 82.2% of the SD group and in 70% of the AD dogs (p>0.05). Among the 20 seronegative dogs, seven (35%) were positive in either OS or CS, and 12 (60%) were positive when both noninvasive samples were jointly considered. The OS/CS combination resulted in a significant increase of positivity (p≤0.05) for the AD dogs (from 36.7% to 63.4%), as well as OS/serology (80%) and OS/CS/serology (83.4%). For the SD population, positivity reached up to 95.2% with the same combinations, showing that combination of samples and/or tests is required for the identification of dogs infected with L. infantum and that the OS and CS combination based on qPCR notably improves the detection of both AD and SD dogs. In conclusion, OS proved to be a suitable sample for the molecular diagnosis of infected dogs with clinical signs of VL, but not for dogs with inapparent infection. For these, we recommend the combination of OS results with CS and/or serology in order to reach relevant positivity for L. infantum. Finally, another advantage of using OS or both noninvasive samples is the increased likelihood of diagnosing seronegative dogs.

Salivary gland homogenates from wild-caught sand flies Lutzomyia flaviscutellata and Lutzomyia (Psychodopygus) complexus showed inhibitory effects on Leishmania (Leishmania) amazonensis and Leishmania (Viannia) braziliensis infection in BALB/c mice.

During the natural transmission of Leishmania parasites, the infected sand fly female regurgitates promastigotes into the host's skin together with its saliva. It has been reported that vector saliva contains immunomodulatory molecules that facilitate the establishment of infection. Thus, the main objective of this study was to evaluate the specificity of Lutzomyia (Lu.) flaviscutellata and Lu. (Psychodopygus) complexus salivas on the infectivity of Leishmania (L.) (Leishmania) amazonensis and L. (Viannia) braziliensis, respectively. BALB/c mice were inoculated into the skin of hind footpad with L. (L.) amazonensis and L. (V.) braziliensis promastigotes in the absence or presence of Lu. flaviscutellata and Lu. (P.) complexus salivary gland homogenates (SGHs). The evolution of the infection was evaluated by lesion size, histopathological analysis and determination of the parasite load in the skin biopsies collected from the site of infection at 4 and 8 weeks PI. The lesion size and the parasite load of both groups of mice infected in the presence of SGHs were smaller than the control groups. The histopathological features showed that the inflammatory reaction was less prominent in the groups of mice infected in the presence of both SGHs when compared to the control group. The results showed that the presence of SGHs of Lu. flaviscutellata and Lu. (P.) complexus led to induction of processes that were disadvantageous to parasite establishment during infection by L. (L.) amazonensis and L. (V.) braziliensis. An inhibitory effect on Leishmania infection could be observed in both groups inoculated with SGHs, especially when the SGH from Lu. (P.) complexus was used.

Evaluation of anti-inflammatory activity of derivatives from aerial parts of Baccharis uncinella.

CONTEXT: Species of Baccharis exhibit antibiotic, antiseptic, and wound-healing properties, and have been used in the traditional medicine of South America for the treatment of inflammation, headaches, diabetes, and hepatobiliary disorders. OBJECTIVE: To investigate the anti-inflammatory activity of organic phases from EtOH extract of the aerial parts of Baccharis uncinella DC (Asteraceae). MATERIALS AND METHODS: The crude EtOH extract from the aerial parts of B. uncinella was subjected to partition procedures and the corresponding CH(2)Cl(2) and EtOAc phases were subjected to several chromatographic separation procedures. Thus, these phases and their purified compounds were assayed for evaluation of anti-inflammatory activity. RESULTS: The CH(2)Cl(2) phase from EtOH extract from B. uncinella contained two triterpenoids (oleanolic and ursolic acids) and one flavonoid (pectolinaringenin), whereas the respective EtOAc phase showed to be composed mainly by two phenylpropanoid derivatives (caffeic and ferulic acids). The CH(2)Cl(2) and EtOAc phases as well as their isolated compounds exhibited anti-inflammatory effects against inflammatory reactions induced by phospholipase A2 (from Crotalus durissus terrificus venom) and by carrageenan. DISCUSSION AND CONCLUSION: The results suggested that the components obtained from partition phases of EtOH extract of B. uncinella could represent lead molecules for the development of anti-inflammatory agents. Additionally, the results confirmed the use of Baccharis genus in the traditional medicine of South America for the treatment of inflammation and other heath disorders. To date, the present work describes for the first time the anti-inflammatory effects of compounds isolated from B. uncinella.

Anti-leishmanial effects of purified compounds from aerial parts of Baccharis uncinella C. DC. (Asteraceae).

Species of Baccharis exhibit antibiotic, antiseptic, wound-healing, and anti-protozoal properties, and have been used in the traditional medicine of South America for the treatment of several diseases. In the present work, the fractionation of EtOH extract from aerial parts of Baccharis uncinella indicated that the isolated compounds caffeic acid and pectolinaringenin showed inhibitory activity against Leishmania (L.) amazonensis and Leishmania (V.) braziliensis promastigotes, respectively. Moreover, amastigote forms of both species were highly sensible to the fraction composed by oleanolic + ursolic acids and pectolinaringenin. Caffeic acid also inhibited amastigote forms of L. (L.) amazonensis, but this effect was weak in L. (V.) braziliensis amastigotes. The treatment of infected macrophages with these compounds did not alter the levels of nitrates, indicating a direct effect of the compounds on amastigote stages. The results presented herein suggest that the active components from B. uncinella can be important to the design of new drugs against American tegumentar leishmaniases.

The effect of phospholipase A2 from Crotalus durissus collilineatus on Leishmania (Leishmania) amazonensis infection.

In this study, the effect of phospholipase A2 (PLA2) derived from Crotalus durissus collilineatus was evaluated in vitro and in vivo on experimental cutaneous leishmaniasis. The promastigote and amastigote forms treated with PLA2 presented increased growth rate. In vivo studies showed that PLA2-treated Leishmania (Leishmania) amazonensis promastigotes increased the size of lesions in BALB/c mice, and histopathological analysis showed numerous necrotic regions presenting a higher density of polymorphonuclear, mononuclear, and amastigote cells. Additionally, infected macrophages treated with PLA2 were able to generate prostaglandin E2 (PGE2). Cytokine quantification showed that the supernatant from infected macrophages presented moderate and high amounts of IL-2 and IL-10, respectively. However, in PLA2-treated infected macrophages, suppression of IL-2 levels occurred, but not of IL-10 levels. Observation also revealed that both the supernatant and lysate of L. (L.) amazonensis promastigotes exhibited PLA2 activity, which, in the presence of dexamethasone, showed no reduction in their activities; while glucocorticoid maintained the ability of promastigote forms to infect macrophages, which presented values similar to controls. In conclusion, the results indicate that PLA2 may be a progression factor for cutaneous leishmaniasis, since the PLA2 effect suppressed IL-2 levels and generated PGE2, an inflammatory lipid mediator.

Adolescência, vida sexual e planejamento reprodutivo de escolares de Serra Pelada, Pará / Adolescence, sexual life and reproductive planning of adolescents of Serra Pelada, Pará

As condições de vida em que um adolescente se desenvolve determinam suas escolhas e influenciam sua vida sexual. O presente estudo visou caracterizar o cotidiano e vida sexual de 108 adolescentes entre 11 e 15 anos de idade, residentes em Serra Pelada, município de Curionópolis, PA, através de análise quantitativa e qualitativa de dados, diferenciando-os quanto ao gênero. Os resultados mostraram a forte presença das relações de gênero em alguns casos. Foi encontrada alguma relação entre religião e vida sexual somente no sexo feminino, em que se observa que 58,5 porcento delas eram adeptas a alguma religião e 66,7 porcento não haviam iniciado a vida sexual aos 12 anos ou menos. Quando questionados se mantinham, ou não, relações sexuais enquanto namoravam, 93,6 porcento das meninas afirmaram que não, enquanto 59,6 porcento dos meninos disseram o mesmo, mostrando que as meninas evitam mais a relação sexual. A grande maioria das meninas afirmou ter feito uso de método para evitar filho na primeira relação, enquanto mais de 50 porcento dos meninos afirmaram o contrário. Essa diferença reflete que a responsabilidade da contracepção é julgada como pertencente ao sexo feminino. Cerca de 70 porcento das meninas afirmam ter a intenção de se casar com a pessoa com quem tem relação sexual atualmente, enquanto 60 porcento dos meninos afirmam o contrário, o que mostra que as meninas se mostram mais receosas quanto a ter filho cedo, em relação aos garotos, e mostram-se mais abertas ao compromisso.

The life conditions in which an adolescent develops determine his choices and influence his sexual life. The present study aimed to characterize the daily activities and sexual life of 108 adolescents aged between 11 and 15 years, living in Serra Pelada, a municipality of Curionópolis, state of Pará, through a quantitative and qualitative data analysis, distinguishing them according to gender. Results showed the strong presence of gender relations in some cases. Some relation was found between religion and sexual life only in the female sex: 58.5% of them followed some religion and 66.7% had not initiated sexual life at the age of 12 or below. When the adolescents were asked whether they had sexual intercourse with their boyfriends or girlfriends, 93.6% of the girls answered they did not, while 59.6% of the boys answered the same, showing that the girls avoid more the sexual intercourse. The great majority of the girls stated they used a contraceptive method in the first intercourse, while more than 50% of the boys said the opposite. This difference reveals that the responsibility for contraception is deemed as belonging to the female sex. Approximately 70% of the girls state they intend to marry the person with whom they are currently having sexual intercourse, while 60% of the boys say the opposite, which shows that the girls are more afraid to have children early, and more open to commitments.

What to expect in the excluded stomach mucosa after vertical banded Roux-en-Y gastric bypass for morbid obesity.

Mucosal alterations after vertical banded Roux-en-Y gastric bypass have not been clearly evaluated. The aim of this paper was to analyze the histological findings and the presence of Helicobacter pylori in the excluded stomach. Forty consecutive patients who underwent Roux-en-Y gastric bypass longer than 36 months were selected for double-balloon enteroscopy. The excluded stomach was reached in 35/40 patients (88%). Morphological alterations were analyzed through hematoxilin and eosin and the presence of H. pylori was confirmed with Giemsa staining. Thirty patients (86%) were female, and the mean age was 43 years old. The mean postoperative time was 78 months (36-110 months). Histologically, all patients had chronic gastritis in the bypassed stomach, with pangastritis in 33/35 (94%). Five cases (5/35, 14%) presented atrophy and four of them also had intestinal metaplasia. Helicobacter pylori was detected in 7/35 (20%) of the excluded stomach and in 12/35 (34%) of the functional pouch. All patients positive for H. pylori in the excluded stomach were also positive in the functional pouch, p = 0.0005. Helicobacter pylori is still present in the excluded stomach after Roux-en-Y gastric bypass and might be considered for treatment. Histological findings indicated high prevalence of atrophy and intestinal metaplasia in this selected population.