Comparison of antimicrobial resistance in thermophilic Campylobacter strains isolated from conventional production and backyard poultry flocks.

1. The aim of this study was to compare the resistance pattern of thermophilic Campylobacter spp. isolated from conventional production (n = 34) and backyard poultry flocks (n = 36) from Rio de Janeiro State, Brazil. The disc diffusion method and statistical tests were used for investigation and analysis of the resistance pattern of Campylobacter spp. isolated from different rearing systems.2. Antimicrobial resistance percentages to amoxycillin with clavulanic acid (AMC), ampicillin (AMP), ceftiofur (CTF), ciprofloxacin (CIP), enrofloxacin (ENO), erythromycin (ERI), gentamicin (GEN) and tetracycline (TET) were 32.4%, 44.1%, 67.6%, 97.1%, 82.4%, 26.5%, 5.9% and 38.2% in conventional production flocks respectively, while the backyard flock's resistance levels were 0.0%, 13.9%, 69.4%, 100.0%, 91.7%, 5.6%, 0.0% and 16.7%, respectively.3. Campylobacter spp. from conventional poultry production was more resistant to AMC, AMO, ERI and TET (P > 0.05) when compared to strains from backyard poultry. A higher frequency of resistance to fluoroquinolones (FLQ), CIP and ENO, was observed in strains from both systems, demonstrating the spread of resistant strains among poultry production environments.

First Report of Papaya Fruit Anthracnose Caused by Colletotrichum brevisporum in Brazil.

Papaya fruits (Carica papaya L.) (cv. Golden) showing post-harvest anthracnose symptoms were observed during surveys of papaya disease in northeastern Brazil from 2008 to 2012. Fruits affected by anthracnose showed sunken, prominent, dark brown to black lesions. Small pieces (4 to 5 mm) of necrotic tissue were surface sterilized for 1 min in 1.5% NaOCl, washed twice with sterile distilled water, and plated onto potato dextrose agar (PDA) amended with 0.5 g liter-1 streptomycin sulfate. Macroscopic colony characters and microscopic morphology characteristics of four isolates were observed after growth on PDA (2) for 7 days at 25°C under a 12-hr light/dark cycle. Colonies varied between colorless and pale brown in reverse, with orange conidial mass. Conidia were hyaline, aseptate, cylindrical with round ends, slightly flattened, smooth-walled, guttulate, and 13.5 (10.5 to 17.1) µm × 3.8 (2.1 to 4.8) µm (l/w ratio = 3.5, n = 50), typical of Colletotrichum spp. DNA sequencing of partial sequences of actin (ACT) gene and the internal transcribed spacer (ITS1-5.8S-ITS2 rRNA) were conducted to accurately identify the species. Sequences of the papaya isolates were 99% similar to those of Colletotrichum brevisporum (GenBank Accession Nos. JN050216, JN050217, JN050238, and JN050239). A phylogenetic analysis using Bayesian inference and including published ACT and ITS data for C. brevisporum and other Colletotrichum species was carried out (1). Based on morphological and molecular data, the papaya isolates were identified as C. brevisporum. Conidia of the papaya isolates were narrower than those described for C. brevisporum (2.9 to 4.8 µm and 5 to 6 µm, respectively) (1), which may be due to differences in incubation temperature or a typical variation in conidial size in Colletotrichum species (3). Sequences of the isolates obtained in this study are deposited in GenBank (ACT Accession Nos. KC702903, KC702904, KC702905, and KC702906; ITS Accession Nos. HM163181, HM015851, HM015854, and HM015859). Cultures are deposited in the Culture Collection of Phytopathogenic Fungi of the Universidade Federal Rural de Pernambuco, Recife, Brazil (CMM 1672, CMM 1702, CMM 1822, and CMM 2005). Pathogenicity testing was conducted with all four strains of C. brevisporum on papaya fruits (cv. Golden). Fruits were wounded at the medium region by pushing the tip of four sterile pins through the surface of the skin to a depth of 3 mm. Mycelial plugs taken from the margin of actively growing colonies (PDA) of each isolate were placed in shallow wounds. PDA discs without fungal growth were used as control. Inoculated fruits were maintained in a humid chamber for 2 days at 25°C in the dark. After 6 days, anthracnose symptoms developed that were typical of diseased fruit in the field. C. brevisporum was successfully reisolated from symptomatic fruits to fulfill Koch's postulates. C. brevisporum was described from Neoregalia sp. and Pandanus pygmaeus in Thailand (1). To our knowledge, this is the first report of C. brevisporum in Brazil and the first report of this species causing papaya fruit anthracnose. References: (1) P. Noireung et al. Cryptogamie Mycol., 33:347, 2012. (2) B. C. Sutton. The Genus Glomerella and its anamorph Colletotrichum. CAB International, Wallingford, UK, 1992. (3) B. S. Weir et al. Stud. Mycol. 73:115, 2012.

First Report of Papaya Fruit Rot Caused by Colletotrichum magna in Brazil.

Species of the genus Colletotrichum are commonly reported as pathogens of fruits in tropical regions. Papaya fruits (Carica papaya L.), cv. Golden, with typical lesions of anthracnose, chocolate spot, and/or stem-end rot were collected from 18 papaya-producing areas of northeast Brazil in 2007. One hundred and fifty-five isolates of Colletotrichum spp. were obtained from the fruit lesions and cultured on potato dextrose agar. Pathogenicity tests were conducted by placing a 20-µl drop of 105 conidia ml-1 suspension on a wounded area of two healthy fruits of cv Golden at the climacteric stage. Inoculated fruits were placed in a moist chamber at 26°C (±2) for 48 h. After this period, the plastic covers of the trays used to form the moist chamber were removed and the trays were kept at 26°C (±2) for 98 h when symptoms were assessed. The causal agents of fruit rot were recovered from inoculated fruits showing symptoms of anthracnose and chocolate spot. Conidia from fresh lesions were collected and measured. Conidia dimensions were 13.49 × 3.80 µm, length/width ratio = 3.55 µm. Conidia were predominantly cylindrical to bluntly rounded ends and slightly flattened. All isolates were morphologically similar to Colletotrichum gloeosporioides Penz (1). Molecular analyses of the isolates were carried out with taxon-specific primers for C. acutatum J.H Simmonds and C. gloeosporioides (3). Only one amplicon was detected for eight isolates with the C. gloeosporioides primer. All isolates were genotyped using inter-simple sequence repeat (ISSR) primers. Three groups of isolates were found, one containing the eight C. gloeosporioides isolates, a second group comprised of 141 isolates, and a third contained six isolates. The second and third groups were more similar to each other than to the first C. gloeosporioides group. Thirty two representative isolates of the three ISSR groups were sequenced for the internal transcribed spacer (ITS) and glutamine synthetase (GS) (GenBank Nos. HM163181 and HM015847) regions. With molecular phylogenetic analyses, two well-supported clades were formed, one with the C. gloeosporioides isolates and the other with sequences highly similar (99% similarity) to the two ITS sequences available in GenBank (DQ003310 and GU358453) and the GS region of G. magna Jenkins & Winstead (DQ792873). The latter was reported in the United States and Taiwan (2,4). Isolates of C. magna and C. gloeosporioides are morphologically similar and identification needs to be based on molecular analyses. To our knowledge, this is the first report of C. magna causing rot of papaya fruit in Brazil. References: (1) P. F. Cannon et al. Mycotaxon 104:189, 2008. (2) M. Z. Du et al. Mycologia 97:641, 2005. (3) P. Talhinhas et al. Appl. Environ. Microbiol. 71:2987, 2005. (4) J. G. Tsay et al. Plant Dis. 94:787, 2010.

Preventive and curative glycoside kaempferol treatments attenuate the TH2-driven allergic airway disease.

Asthma is a chronic respiratory disease characterized by airway inflammation and airway hyperresponsiveness (AHR). One strategy to treat allergic diseases is the development of new drugs. Flavonoids are compounds derived from plants and are known to have antiallergic, anti-inflammatory, and antioxidant properties. To investigate whether the flavonoid kaempferol glycoside 3-O-[beta-d-glycopiranosil-(1-->6)-alpha-l-ramnopiranosil]-7-O-alpha-l-ramnopiranosil-kaempferol (GRRK) would be capable of modulating allergic airway disease (AAD) either as a preventive (GRRK P) or curative (GRRK C) treatment in an experimental model of asthma. At weekly intervals, BALB/c mice were subcutaneously (sc) sensitized twice with ovalbumin (OVA)/alum and challenged twice with OVA administered intranasally. To evaluate any preventive effect, GRRK was administered 1h (hour) before each OVA-sensitization and challenge, while to analyze the curative effect, mice were first sensitized with OVA, followed by GRRK given at day 18 through 21. The onset of AAD was evaluated 24h after the last OVA challenge. Both treatments resulted in a dose-dependent reduction in total leukocyte and eosinophil counts in the bronchoalveolar lavage fluid (BAL). GRRK also decreased CD4(+), B220(+), MHC class II and CD40 molecule expressions in BAL cells. Histology and lung mechanic showed that GRRK suppressed mucus production and ameliorated the AHR induced by OVA challenge. Furthermore, GRRK impaired Th2 cytokine production (IL-5 and IL-13) and did not induce a Th1 pattern of inflammation. These findings demonstrate that GRRK treatment before or after established allergic lung disease down-regulates key asthmatic features. Therefore, GRRK has a potential clinical use for the treatment of allergic asthma.

Human IgG1 and IgG4: the main antibodies against Triatoma infestans (Hemiptera: Reduviidae) salivary gland proteins.

The Triatoma infestans salivary gland proteins (TSGP) can induce local and systemic hypersensitivity reactions in humans. IgG antibodies against TSGP were present in higher levels in sera of Chagas disease patients, and in individuals living in triatomine-infested areas than in controls living in triatomine-free areas. TSGP-specific IgG1 was found in sera of Chagas patients, and of individuals living in triatomine-infested rural areas, and uniquely specific IgG4 was present in sera of Chagas patients living in triatomine-infested areas, reactive against TSGP. Unique specificities were not detected in sera of individuals reacting against the ubiquitous mosquito Culex quinquifasciatus saliva proteins (CSGP). In conclusion, IgG1 reactive against TSGP is the main antibody present in individuals living in the triatomine-infested study areas. Also, IgG4 is found in the sera of insect-transmitted Chagas disease patients living in study areas.

Persistent infections in chronic Chagas' disease patients treated with anti-Trypanosoma cruzi nitroderivatives.

We used a molecular method and demonstrated that treatment of the chronic human Trypanosoma cruzi infections with nitroderivatives did not lead to parasitological cure. Seventeen treated and 17 untreated chronic Chagas' disease patients, with at least two out of three positive serologic assays for the infection, and 17 control subjects formed the study groups. PCR assays with nested sets of T. cruzi DNA primers monitored the efficacy of treatment. The amplification products were hybridized to their complementary internal sequences. Untreated and treated Chagas' disease patients yielded PCR amplification products with T. cruzi nuclear DNA primers. Competitive PCR was conducted to determine the quantity of parasites in the blood and revealed < 1 to 75 T. cruzi/ml in untreated (means 25.83+/-26.32) and < 1 to 36 T. cruzi/ml in treated (means 6.45+/-9.28) Chagas' disease patients. The difference between the means was not statistically significant. These findings reveal a need for precise definition of the role of treatment of chronic Chagas' disease patients with nitrofuran and nitroimidazole compounds.