Frequency of Infection of Lutzomyia Phlebotomines with Leishmania braziliensis in a Brazilian Endemic Area as Assessed by Pinpoint Capture and Polymerase Chain Reaction

Leishmania infected of Lutzomyia spp. are rare in endemic areas. We tested the hypothesis that there is clustering of infected vectors by combining pinpoint capture with sensitive L. braziliensis kDNA minicircle specific PCR/dot blot in an endemic area in the State of Bahia. Thirty out of 335 samples (10 to 20 sand flies/sample; total of 4,027 female sand flies) were positive by PCR analysis and dot blot leading to a underestimated overall rate of 0.4% positive phlebotomines. However, 83.3% of the positive samples were contributed by a single sector out of four sectors of the whole studied area. This resulted in a rate of 1.5% Leishmania positive phlebotomines for this sector, far above rates of other sectors. Incidence of American cutaneous leishmaniasis cases for this sector was about twice that for other sectors. Our results show that there is a non-homogeneous distribution of Leishmania-infected vectors. Such a clustering may have implications in control strategies against leishmaniasis, and reinforces the necessity of understanding the ecological and geographical factors involved in leishmanial transmission.

Larval dispersal and predation in experimental populations of Chrysomya albiceps and Cochliomyia macellaria (Diptera: Calliphoridae)

In this study we investigated the larval dispersal associated with larval predation in experimental populations of Chrysomya albiceps and Cochliomyia macellaria. Frequency distribution of sampling units (G test) in the substrate was used to evaluate variation in larval dispersal. An experimental acrylic channel (1 x 0.1 x 0.2 m) covered with wood shavings was used to observe larval dispersal prior to pupation. The acrylic channel was graduated at 0.05 m intervals, each representing a sampling unit; hence, 20 sampling units were set up. A Petri dish containing third instar larvae of single and double species was deposited at one edge of the acrylic channel allowing larvae to disperse. The number of buried pupae (0, 1, 2, …n) present in each sampling unit was recorded. For double species, the number of recovered larvae of C. albiceps was similar to the number initially released on the dish Petri. On the other hand, the number of recovered larvae of C. macellaria was significantly smaller than the initially released number. The results show that C. albiceps attacks C. macellaria larvae during the larval dispersal process. The larval distribution of C. albiceps did not differ significantly from C. macellaria in double species, but it differed significantly in single species. The larval aggregation level of C. macellaria decreased when C. albiceps was present and the larval aggregation level of C. albiceps increased when C. macellaria was present. The implications of such findings for the population dynamics of these species are discussed.

Parasitological characteristics of Schistosoma mansoni infection in swiss mice with underlying malnutrition

The effects of a protein-restricted diet (8% protein, 81% carbohydrate and 11% lipids) on Schistosoma mansoni infectivity, fecal egg excretion and intestinal egg distribution in Swiss (SW) mice were studied. Pregnant mice received a deficient diet from the middle of gestation until delivery. Seven-days-old mice were exposed to 50 cercariae (BH strain, Brazil). Offspring mice had a free access to the deficient diet since lactation until adulthood. The controls were fed with a commercial mice diet. A parasitological examination was performed between six and eight weeks post-infection while both groups were necropsied one week later. Mice on the experimental diet showed a significant loss in body weight. There was no significant difference (p > 0.05) in pre-patent period, kinetics of egg excretion and worm recovery from mice on either diet. Significant differences (p < 0.05) were found concerning to the percentage of deposited eggs in the distal segment of the small intestine from hosts on the experimental diet.Our data suggest that experimental malnutrition induced for a long term has no detrimental effect on the acute schistosomiais infection in SW mice.

Trypanosoma cruzi: genetic group with peculiar biochemical and biological behavior

Thirty-two Trypanosoma cruzi strains, isolated from chronic chagasic patients in the northwest of the state of Paraná (Brazil), were analyzed using molecular, biochemical and biological characteristics. Genotypic analysis using randomly amplified polymorphic DNA and simple sequence repeat-anchored polymerase chain reaction amplified profiles showed a large, genetically well-correlated group that contained the majority of the strains and a divergent group that included the PR-150 strain. For glycoconjugate composition, the PR-150 strain was different from the other strains considering the absence or presence of specific bands in aqueous or detergent phases. This strain was also totally different from the others in one out of the six parameters related to in vitro and in vivo biological behavior. We highlight the fact that the PR-150 was totally resistant to benznidazole. For the other biological parameters this strain was not totally distinct from the others, but it showed a peculiar behavior.

Helicobacter pylori detection in gastric biopsies, saliva and dental plaque of Brazilian dyspeptic patients

Helicobacter pylori is an important human pathogen that causes chronic gastritis and is associated with the development of peptic ulcer disease and gastric malignancies. The oral cavity has been implicated as a potential H. pylori reservoir and may therefore be involved in the reinfection of the stomach, which can sometimes occur following treatment of an H. pylori infection. The objectives of this paper were (i) to determine the presence of H. pylori in the oral cavity and (ii) to examine the relationship between oral H. pylori and subsequent gastritis. Gastric biopsies, saliva samples and dental plaques were obtained from 78 dyspeptic adults. DNA was extracted and evaluated for the presence of H. pylori using polymerase chain reaction and Southern blotting methods. Persons with gastritis were frequently positive for H. pylori in their stomachs (p < 0.0001) and there was a statistically significant correlation between the presence of H. pylori in gastric biopsies and the oral cavity (p < 0.0001). Our results suggest a relationship between gastric infection and the presence of this bacterium in the oral cavity. Despite this, H. pylori were present in the oral cavity with variable distribution between saliva and dental plaques, suggesting the existence of a reservoir for the species and a potential association with gastric reinfection.

Syphilis and cirrhosis: a lethal combination in a XIX century individual identified from the Medical Schools Collection at the University of Coimbra (Portugal)

Syphilis is a chronic infection that is categorized by a three-stage progression. The tertiary stage may affect bones and produce distinctive skull lesions called caries sicca. This paper aims to present an unusual case of syphilis associated with a diagnosis of cirrhosis, which was recorded as the cause of death in a 28-year-old female in 1899. The appearance and distribution of the lesions were compatible with acquired syphilis, as observed in the skull from the Medical Schools Collection of the University of Coimbra. However, the cause of death was recorded as "hypertrophic cirrhosis of the liver", this is a condition that is compatible with several liver disorders, including a primary liver disorder, such as cirrhosis provoked by alcoholism, infection of the liver by syphilis pathogens or by damage to the liver from the use of mercury compounds, which was the common treatment for syphilis at the time. This paper represents a contribution to the understanding of the natural evolution of syphilis.

Genetic diversity of Leishmania infantum field populations from Brazil

Leishmania infantum (syn. Leishmania chagasi) is the etiological agent of visceral leishmaniasis (VL) in Brazil. The epidemiology of VL is poorly understood. Therefore, a more detailed molecular characterization at an intraspecific level is certainly needed. Herein, three independent molecular methods, multilocus microsatellite typing (MLMT), random amplification of polymorphic DNA (RAPD) and simple sequence repeats-polymerase chain reaction (SSR-PCR), were used to evaluate the genetic diversity of 53 L. infantum isolates from five different endemic areas in Brazil. Population structures were inferred by distance-based and Bayesian-based approaches. Eighteen very similar genotypes were detected by MLMT, most of them differed in only one locus and no correlation was found between MLMT profiles, geographical origin or the estimated population structure. However, complex profiles composed of 182 bands obtained by both RAPD and SSR-PCR assays gave different results. Unweighted pair group method with arithmetic mean trees built from these data revealed a high degree of homogeneity within isolates of L. infantum. Interestingly, despite this genetic homogeneity, most of the isolates clustered according to their geographical origin.

Recognition of enteroinvasive Escherichia coli and Shigella flexneri by dendritic cells: distinct dendritic cell activation states

The innate and adaptive immune responses of dendritic cells (DCs) to enteroinvasive Escherichia coli (EIEC) infection were compared with DC responses to Shigella flexneri infection. EIEC triggered DCs to produce interleukin (IL)-10, IL-12 and tumour necrosis factor (TNF)-α, whereas S. flexneri induced only the production of TNF-α. Unlike S. flexneri, EIEC strongly increased the expression of toll like receptor (TLR)-4 and TLR-5 in DCs and diminished the expression of co-stimulatory molecules that may cooperate to inhibit CD4+ T-lymphocyte proliferation. The inflammation elicited by EIEC seems to be related to innate immunity both because of the aforementioned results and because only EIEC were able to stimulate DC transmigration across polarised Caco-2 cell monolayers, a mechanism likely to be associated with the secretion of CC chemokine ligands (CCL)20 and TNF-α. Understanding intestinal DC biology is critical to unravelling the infection strategies of EIEC and may aid in the design of treatments for infectious diseases.

Field optimisation of MosquiTRAP sampling for monitoring Aedes aegypti Linnaeus (Diptera: Culicidae)

A sticky trap designed to capture gravid Aedes (Stegomyia) aegypti mosquitoes, MosquiTRAP, has been evaluated for monitoring this species in Brazil. However, the effects of trap densities on the capture rate of Ae. aegypti females and the sensitivity of vector detection are still unknown. After a preliminary study has identified areas of high and low female mosquito abundance, a set of experiments was conducted in four neighbourhoods of Belo Horizonte (state of Minas Gerais, Brazil) using densities of 1, 2, 4, 8, 16, 32 and 64 traps per block. Trap sensitivity (positive MosquiTRAP index) increased significantly when 1-8 MosquiTRAPs were installed per block in both high and low abundance areas. A strong fit was obtained for the total number of mosquitoes captured with increasing trap densities through a non-linear function (Box-Lucas) (r² = 0,994), which likely exhibits saturation towards an equilibrium level. The capacity of the Mean Female Aedes Index to distinguish between areas of high and low Ae. aegypti abundance was also investigated; the achieved differentiation was shown to be dependent on the MosquiTRAP density.

Comparison of the Kato-Katz and Helmintex methods for the diagnosis of schistosomiasis in a low-intensity transmission focus in Bandeirantes, Paraná, southern Brazil

The diagnosis of schistosomiasis is problematic in low-intensity transmission areas because parasitological methods lack sensitivity and molecular methods are neither widely available nor extensively validated. Helmintex is a method for isolating eggs from large faecal samples. We report preliminary results of a comparative evaluation of the Helmintex and Kato-Katz (KK) methods for the diagnosis of schistosomiasis in a low-intensity transmission area in Bandeirantes, Paraná, southern Brazil. Eggs were detected by both methods in seven patients, whereas only Helmintex yielded positive results in four individuals. The results confirm the previously demonstrated higher sensitivity of the Helmintex method compared with the KK method.

Tuberculosis in a southern Brazilian prison

The occurrence of tuberculosis (TB) in prisons has been described as an alarming public health problem in many countries, especially in developing nations. The objective of this study was to conduct a survey among prisoners with TB respiratory symptoms in order to estimate the incidence of the disease, to analyze the drug susceptibility profile and genotype the isolates of Mycobacterium tuberculosis in the city of Charqueadas, southern of Brazil. The TB incidence was 55/1,900 inhabitants in the prison; this corresponds to an incidence of 3,789/100,000 inhabitants, with a prevalence of 72/1,900 (4,960/100,000 inhabitants). Drug susceptibility test was performed and, among the analyzed isolates, 85% were susceptible to all drugs tested and 15% were resistant to at least one drug, of which 89% were resistant only to isoniazid (INH) or in combination with another drug. The genotype classification of spoligotyping analysis showed that 40% of the isolates belong to LAM family, 22% to T family, 17.5% to Haarlem family, 12.5% to U family and 3% to X family. The shared international spoligotypes most frequently found were 729 (27%), 50 (9.5%), 42 (8%), 53 (8%) and 863 (8%). In conclusion, it was observed that TB in this specific population had been caused, mostly, by strains that have been transmitted in the last few years, as demonstrated by the large level of genotype clustering. In addition, it was found specific large clusters, which were not often found in the general population from the same period and in the same region.

Comparison of two rapid tests for anti-phenolic glycolipid-I serology in Brazil and Nepal

The diagnosis of leprosy continues to be based on clinical symptoms and early diagnosis and treatment are critical to preventing disability and transmission. Sensitive and specific laboratory tests are not available for diagnosing leprosy. Despite the limited applicability of anti-phenolic glycolipid-I (PGL-I) serology for diagnosis, it has been suggested as an additional tool to classify leprosy patients (LPs) for treatment purposes. Two formats of rapid tests to detect anti-PGL-I antibodies [ML immunochromatography assay (ICA) and ML Flow] were compared in different groups, multibacillary patients, paucibacillary patients, household contacts and healthy controls in Brazil and Nepal. High ML Flow intra-test concordance was observed and low to moderate agreement between the results of ML ICA and ML Flow tests on the serum of LPs was observed. LPs were "seroclassified" according to the results of these tests and the seroclassification was compared to other currently used classification systems: the World Health Organization operational classification, the bacilloscopic index and the Ridley-Jopling classification. When analysing the usefulness of these tests in the operational classification of PB and MB leprosy for treatment and follow-up purposes, the ML Flow test was the best point-of-care test for subjects in Nepal and despite the need for sample dilution, the ML ICA test yielded better performance among Brazilian subjects. Our results identified possible ways to improve the performance of both tests.

Plasmid-based controls to detect rpoB mutations in Mycobacterium tuberculosis by quantitative polymerase chain reaction-high-resolution melting

Quantitative polymerase chain reaction-high-resolution melting (qPCR-HRM) analysis was used to screen for mutations related to drug resistance in Mycobacterium tuberculosis. We detected the C526T and C531T mutations in the rifampicin resistance-determining region (RRDR) of the rpoB gene with qPCR-HRM using plasmid-based controls. A segment of the RRDR region from M. tuberculosis H37Rv and from strains carrying C531T or C526T mutations in the rpoB were cloned into pGEM-T vector and these vectors were used as controls in the qPCR-HRM analysis of 54 M. tuberculosis strains. The results were confirmed by DNA sequencing and showed that recombinant plasmids can replace genomic DNA as controls in the qPCR-HRM assay. Plasmids can be handled outside of biosafety level 3 facilities, reducing the risk of contamination and the cost of the assay. Plasmids have a high stability, are normally maintained in Escherichia coli and can be extracted in large amounts.

Comparative analysis of the tissue inflammatory response in human cutaneous and disseminated leishmaniasis

Cutaneous leishmaniasis (CL) is the most frequent clinical form of tegumentary leishmaniasis and is characterised by a single or a few ulcerated skin lesions that may disseminate into multiple ulcers and papules, which characterise disseminated leishmaniasis (DL). In this study, cells were quantified using immunohistochemistry and haematoxylin and eosin staining (CD4+, CD68+, CD20+, plasma cells and neutrophils) and histopathology was used to determine the level of inflammation in biopsies from patients with early CL, late CL and DL (ulcers and papules). The histopathology showed differences in the epidermis between the papules and ulcers from DL. An analysis of the cells present in the tissues showed similarities between the ulcers from localised CL (LCL) and DL. The papules had fewer CD4+ T cells than the DL ulcers. Although both CD4+ cells and macrophages contribute to inflammation in early CL, macrophages are the primary cell type associated with inflammation intensity in late ulcers. The higher frequency of CD20+ cells and plasma cells in lesions demonstrates the importance of B cells in the pathogenesis of leishmaniasis. The number of neutrophils was the same in all of the analysed groups. A comparison between the ulcers from LCL and DL and the early ulcers and papules shows that few differences between these two clinical forms can be distinguished by observing only the tissue.