Caracterização de estirpes de Mycobacterium bovis: estudos de crescimento e de interacção com macrófagos

Dissertação para obtenção do Grau de Mestre em Genética Molecular e Biomedicina

Vibrio spp. and Salmonella spp., presence and susceptibility in crabs Ucides cordatus

The presence of Vibrio spp. and Salmonella spp. in crabs marketed at the Bezerra de Menezes Ave., Fortaleza, State of Ceará, Brazil, was assessed between February and May, 2003. The number of individuals sampled in each one of the fifteen weekly samplings ranged between four and eight. Seven strains of Salmonella, from four different samplings, were identified, being five of them identified as serotype S. Senftenberg and two as S. Poona. All strains of Salmonella were sensitive to the tested anti-microbial drugs, with the exception of tetracycline and nalidixic acid, for which an intermediary sensibility was found. The MPN's for Vibrio ranged between 110/g and 110,000/g. Of the forty five Vibrio strains isolated from the crab samples, only 10 were identified up to the species level: two V. alginolyticus and eight V. parahaemolyticus. Bacteria belonging to the Enterobacteriaceae and Pseudomonaceae families were also identified, namely Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, Pantoea agglomerans and Pseudomonas aeruginosa. The proper cooking of the animals is recommended in order to avoid problems for the consumers of this crustacean.

Detection of Mycobacterium leprae DNA for 36kDa protein in urine from leprosy patients: a preliminary report

We have searched for Mycobacterium leprae DNA for 36kDa protein in urine using a M. leprae specific PCR technique. A limited number of 16 patients (of which 11 belonged to lepromatous leprosy and five to tuberculoid leprosy) and eight healthy individuals were included for the present study. The number of urine samples positive by PCR were 36.4% (4/11) in lepromatous patients and 40% (2/5) in tuberculoid patients. None of the samples from healthy individuals was positive. To our knowledge, the results indicate, for the first time, the presence of M. leprae DNA in urine from leprosy patients. Another important finding obtained out of the study is that amongst treated patients 66.6% (4/6) were positive whereas amongst untreated only 20% (2/10) were positive. From the present indicative data it appears that treatment improves the PCR results with urine as a sample. Thus, the approach could prove to be useful for monitoring the treatment response of individual patients and needs to be further evaluated with a large number of patients.

Multiple brain abscesses due to Penicillium spp infection

We present a case of central nervous system (CNS) infection by a member of the Penicillium genera in a HIV-negative man in Brazil. The patient was admitted complaining of loss of visual fields and speech disturbances. CT scan revealed multiple brain abscesses. Stereothacic biopsies revealed fungal infection and amphotericin B treatment begun with initial improvement. The patient died few days later as a consequence of massive gastrointestinal bleeding due to ruptured esophageal varices. The necropsy and final microbiologic analyses disclosed infection by Penicillium sp. There are thousands of fungal species of the Penicillium genera. Systemic penicilliosis is caused by the P. marneffei and was formerly a rare disease, but now is one of the most common opportunistic infection of AIDS patients in Southeast Asia. The clinical presentation usually involves the respiratory system and the skin, besides general symptoms like fever and weight loss. Penicillium spp infection caused by species other than P. marneffei normally cause only superficial or allergic disease but rare cases of invasive disease do occur. We report the fourth case of Penicillium spp CNS infection.

Adulticidal activity against Stegomyia aegypti (Diptera: Culicidae) of three Piper spp.

Three Piper species, Piper longum, P. ribesoides and P. sarmentosum, were selected for investigation of adulticidal potential against Stegomyia aegypti, a main vector of dengue and dengue haemorrhagic fever. Successive extraction by maceration with 95% ethanol showed percentage yields of ethanolic extracts, which derived from P. longum, P. ribesoides and P. sarmentosum, of 8.89, 3.21 and 5.30% (w/w), respectively. All Piper extracts illustrated an impressive adulticidal activity when tested against female mosquitoes by topical application. The susceptibility of St. aegypti females to ethanol-extracted Piper was dose dependent and varied among the plant species. The highest adulticidal effect was established from P. sarmentosum, followed by P. ribesoides and P. longum, with LD50 values of 0.14, 0.15 and 0.26 µg/female, respectively. The potential of these Piper species, as possible mosquitocides, established convincing activity for further researches to develop natural substances for combat against adult mosquitoes.

New data on Echinococcus spp. in Southern Brazil

40 Echinococcus isolates from sheep and cattle in Southern Brazil were genetically analysed in order to obtain further data on the presence of different taxa of the Echinococcus granulosus complex. Differentiation was done using a PCR technique and sequencing of mitochondrial cytochrome c oxidase subunit 1 (CO1). Most samples (38) could be allocated to the sheep strain (G1) of E. granulosus, while two samples belonged to E. ortleppi, previously known as cattle strain (G5) of E. granulosus. Due to the shorter prepatent period in dogs of the latter taxon, this records have important implications for the design of control measures in this endemic region.

Aeromonas spp. isolated from oysters (Crassostrea rhizophorea) from a natural oyster bed, Ceará, Brazil

Between April and October 2002, thirty fortnightly collections of oysters (Crassostrea rhizophorea) from a natural oyster bed at the Cocó River estuary in the Sabiaguaba region (Fortaleza, Ceará, Brazil) were carried out, aiming to isolate Aeromonas spp. strains. Oyster samples were submitted to the direct plating (DP) and the presence/absence (P/A) methods. Aeromonas were identified in 15 (50%) samples analyzed by the DP method and in 13 (43%) analyzed by the P/A method. A. caviae, A. eucrenophila, A. media, A. sobria, A. trota, A. veronii bv. sobria, A. veronii bv. veronii and Aeromonas sp. were isolated. The predominant species was A. veronii (both biovars), which was identified in 13 (43%) samples, followed by A. media in 11 (37%) and A. caviae in seven (23%). From the 59 strains identified, 28 (48%) presented resistance to at least one of the eight antibiotics tested.

Bites by coral snakes (Micrurus spp.) in Campinas, State of São Paulo, Southeastern Brazil

Coral snakes (Micrurus spp.) are the main representatives of the Elapidae in South America. However, bites by these snakes are uncommon. We retrospectively reviewed the data from 11 individuals bitten by coral snakes over a 20-year period; four were confirmed (snake brought for identification) and seven were highly suspected (neuromuscular manifestations) cases of elapid envenoming. The cases were classified as dry-bite (n = 1, caused by M. lemniscatus; did not receive antivenom), mild (n = 2, local manifestations with no acute myasthenic syndrome; M. frontalis and Micrurus spp.), moderate (n = 5, mild myasthenia) or severe (n = 3, important myasthenia; one of them caused by M. frontalis). The main clinical features upon admission were paresthesia (local, n = 9; generalized, n = 2), local pain (n = 8), palpebral ptosis (n = 8), weakness (n = 4) and inability to stand up (n = 3). No patient developed respiratory failure. Antivenom was used in ten cases, with mild early reactions occurring in three. An anticholinesterase drug was administered in the three severe cases, with a good response in two. No deaths were observed. Despite the high toxicity of coral snake venoms, the prognosis following envenoming is good. In serious bites by M. frontalis or M. lemniscatus, the venom of which acts postsynaptically, anticholinesterases may be useful as an ancillary measure if antivenom is unavailable, if there is a delay in obtaining a sufficient amount, or in those patients given the highest recommended doses of antivenom without improvement of the paralysis or with delayed recovery.

An evaluation of manual and mechanical methods to identify Candida spp. from human and animal sources

To compare two yeast identification methods, i. e, the manual and the VITEK mechanical methods, 62 clinical samples from hemocultures and animal sources were analyzed. After identification as Candida yeasts by the VITEK method, the strains were recharacterized using manual assimilation methods and sugar fermentation tests. Our findings reveal 58% concurrent identification between the two methods for animal strains, and 51% for human hemoculture strains.

Soft tissue abscess and lymphadenitis due to Mycobacterium avium Complex as an expression of immune reconstitution inflammatory syndrome after a second scheme of highly active antiretroviral therapy

Immune reconstitution inflammatory syndrome (IRIS) is an atypical and unexpected reaction related to highly active antiretroviral therapy (HAART) in human immunodeficiency virus (HIV) infected patients. IRIS includes an atypical response to an opportunistic pathogen (generally Mycobacterium tuberculosis, Mycobacterium avium complex, cytomegalovirus and herpes varicella-zoster), in patients responding to HAART with a reduction of plasma viral load and evidence of immune restoration based on increase of CD4+ T-cell count. We reported a case of a patient with AIDS which, after a first failure of HAART, developed a subcutaneous abscess and supraclavicular lymphadenitis as an expression of IRIS due to Mycobacterium avium complex after starting a second scheme of HAART.

Occurrence of antibodies against Leptospira spp. in horses of the urban area of Londrina, Paraná, Brazil

A total of 320 horses were studied in this paper, both male and female, between two and 17 years of age, which were used for traction of wagons in the urban area of the municipality of Londrina (PR). These animals were kept, after their daily work, in abandoned areas or plots, in the outskirts of the urban area of the city. When these animals were attended by the veterinarians, between 1996 and 2005, none of them presented symptoms suggesting leptospirosis. The most frequent reasons for the visit were loss of weight, unwillingness for work, parasitism, laminess, and wounds. Microscopic Seroagglutination Test (SAM), with 22 Leptospira serovars, was performed in sera sample from all these animals. The aim of this study was to investigate the occurrence of antibodies against Leptospira spp. in horses from the urban area of Londrina (PR). From the samples tested, 214 (66.88%) were considered positive, with titers between 100 and 3200, being that 49 (22.90%) presented antibodies against a single serovar of Leptospira, and 165 (77.10%) samples presented antibodies against two or more serovars simultaneously, where in 88 (53.33%) it was possible to characterize the most likely probable serovar. Antibodies against the serovar Icterohaemorrhagiae were detected in 32 (23.36%) animals.

Genotypic identification of Cryptosporidium spp. isolated from hiv-infected patients and immunocompetent children of São Paulo, Brazil

Cryptosporidium isolates identified in fourteen stool samples, collected from five HIV-infected patients and nine immunocompetent children, living in the Sate of São Paulo, Brazil, were submitted to a molecular analysis using a nested PCR followed of restriction fragment length polymorphism (RFLP), for genetic characterization. The analysis was based on digestion with RsaI restriction enzyme of a DNA fragment amplified from the Cryptosporidium oocyst wall protein (COWP) gene. Based on this analysis, four samples were identified as Cryptosporidium parvum, eight as Cryptosporidium hominis and two presented a profile that correspondedto Cryptosporidium meleagridis when compared to the standards used in the analysis. The use of molecular methods can be helpful to identify source of infections and risk factors related to Cryptosporidium infection in our communities.