Purification and characterization of a unique pectin lyase from aspergillus giganteus able to release unsaturated monogalacturonate during pectin degradation

A pectin lyase, named PLIII, was purified to homogeneity from the culture filtrate of Aspergillus giganteus grown in submerged culture containing orange peel waste as carbon source. PLIII was able to digest apple pectin and citrus pectins with different degrees of methyl esterification. Interestingly, the PLIII activity was stimulated in the presence of some divalent cations including Pb(2+) and was not significantly affected by Hg(2+). Like other pectin lyases, PLIII is stimulated by but is not dependent on Ca(2+). The main soluble product released during the degradation of pectic substances promoted by the PLIII is compatible with an unsaturated monogalacturonate. PLIII is a unique enzyme able to release unsaturated monogalacturonate as the only soluble product during the degradation of pectic substances; therefore, PLIII was classified as an exo-pectin lyase. To our knowledge, this is the first characterization of an exo-pectin lyase. The PLIII described in this work is potentially useful for ethanol production from pectin-rich biomass, besides other common applications for alkaline pectinases like preparation of textile fibers, coffee and tea fermentation, vegetable oil extraction, and the treatment of pulp in papermaking.

Effect of biotransformation by liver S9 enzymes on the mutagenicity and cytotoxicity of melanin extracted from Aspergillus nidulans

A mutant that exhibited increased melanin pigment production was isolated from Aspergillus nidulans fungus. This pigment has aroused biotechnological interest due to its photoprotector and antioxidant properties. In a recent study, we showed that melanin from A. nidulans also inhibits NO and TNF-α production. The present study evaluates the mutagenicity and cytotoxicity of melanin extracted from A. nidulans after its exposure to liver S9 enzymes. The cytotoxicity of multiple concentrations of melanin (31.2-500 μg/mL) against the McCoy cell line was evaluated using the Neutral Red assay, after incubation for 24 h. Mutagenicity was assessed using the Ames test with the Salmonella typhimurium strains TA98, TA97a, TA100, and TA102 at concentrations ranging from 125 μg/plate to 1 mg/plate after incubation for 48 h. The cytotoxicity of A. nidulans melanin after incubation with S9 enzymes was less than (CI50 value= 413.4 ± 3.1 μg/mL) that of other toxins, such as cyclophosphamide (CI50 value = 15 ± 1.2 μg/mL), suggesting that even the metabolised pigment does not cause significant damage to cellular components at concentrations up to 100 μg/mL. In addition, melanin did not exhibit mutagenic properties against the TA 97a, TA 98, TA 100, or TA 102 strains of S. typhimurium, as shown by a mutagenic index (MI)  <2 in all assays. The significance of these results supports the use of melanin as a therapeutic reagent because it possesses low cytotoxicity and mutagenic potential, even when processed through an external metabolising system.

Influence of hypertension on oral infections and endodontic treatment

Hypertension is characterized by peripheral vascular resistancethat leads to blood pressure increase and severalsystemic changes that may negatively influence one s oralhealth. Thus, the aim of this study was to conduct a literaturereview on the influence of hypertension over oral conditionsand endodontic treatment. Hypertension mainly affects theblood vessels, brain and kidneys. A hypertensive conditioncan lead to increased levels of parathyroid hormones, abnormalvitamin D metabolism, reduction in the concentrationof ionized calcium and decreased calcium absorption.Therefore, hypertension can be closely associated with oralproblems such as periodontal diseases, implant loss, difficultyin bone healing, reduced salivary flow and protein concentrationin saliva, increased number of neutrophils and, as a consequence,favoring of inflammatory processes. It has alsobeen suggested that the success rate of endodontic treatmentin hypertensive patients is lower than in normotensiveones. The response of hypertensive patients to root canaltreatment, intracanal medications and sealers should be furtherstudied in order to provide knowledge on the changes,failures and success of endodontic treatment.

Microbiota asociated with infections of the jaws

The microbial infections involving the craniofacial skeleton, particularly maxilla and mandible, have direct relationship with the dental biofilm, with predominance of obligate anaerobes. In some patients, these infections may spread to bone marrow or facial soft tissues, producing severe and life-threatening septic conditions. In such cases, local treatment associated with systemic antimicrobials should be used in order to eradicate the sources of contamination. This paper discuss the possibility of spread of these infections and their clinical implications for dentistry, as well as their etiology and aspects related to microbial virulence and pathogenesis.

Manifestación bucal de paracoccidiodomicosis asociada a aspergilosis: presentación de caso

Paracoccidioidomycosis and aspergillosis are, respectively, mycosis caused by Paracoccidioides brasiliensis and Aspergillus fumigatus fungi, which, most frequently, attack the lungs and later spread to other body regions. The association between both mycosis is rare in patients that do not present immunosuppression. A 49-year-old man presented with oral lesion, besides lesions on palatum, retromolar and oropharyngeal regions. Histopathological and microbiological exams confirmed the diagnosis of paracoccidioidomycosis associated to aspergillosis. Historically, fungi have been considered as the cause of relatively important infections, but, in the last years, a significant increase of diseases caused by fungi is evidenced. The recognition of the etiological agent in culture is important for the suitable treatment, once these fungi can cause irreversible damages or even death. The purpose of this study was to demonstrate, through the case of a patient who presented both mycosis associated, the need of complementary exams for the diagnosis of oral diseases, when routine laboratorial exams indicate the presence of another disease associated.

Isolation and molecular identification of Fusobacterium nucleatum from Nigerian patients with oro-facial infections.

Fusobacterium nucleatum is one of the most common anaerobic bacteria present in the oral cavity and is often isolated from infections involving other body sites. To characterise F. nucleatum strains from patients attending a teaching hospital in Nigeria in order to provide information on the methods for accurate identification of anaerobes in clinical specimen. Fusobacterium nucleatum specie from 50 patients presenting with oro-facial infections were studied by culture on Fusobacterium selective agar and fastidious anaerobe agar. The isolates were characterised based on colonial morphology, microscopy, lipase production, susceptibility to kanamycin and colistin and resistance to vancomycin. Biochemical tests were performed using a commercial test kit. The identity of the isolates was confirmed based on molecular characterization performed using polymerase chain reaction (PCR) analysis. Forty-eight (96%) F. nucleatum isolates were obtained from the 50 patients by culture and all the isolates were identified by colonial appearance and microscopy based on their unique spindle shape with tapered ends. Only 26 (54.2%) of the 48 isolates were identified by commercial API 20A test kit while PCR confirmed the identity of all the isolates. Anaerobes are involved in human infections and their study is quite cumbersome due to tedious nature and high cost of the techniques involved. Cultural method is reliable in the isolation and identification of F. nucleatum species. PCR is a rapid and simple method that can complement the phenotypic identification of anaerobes and would assist in their full identification.

Effect of oral infections on serum creatinine levels in diabetic rats

Aim: The aim of this study was to evaluate the relationship between pulpal and/or periodontal disease and serum creatinine levels in a rat model of diabetes mellitus. Methods: Eighty male rats (Rattus norvegicus albinus, Wistar) were divided into the following 8 groups compris-ing 10 animals each: normal (G1), with pulpal disease (G2), with periodontal disease (G3), with both pulpal and periodontal disease (G4), diabetic (G5), diabetic with pulpal disease (G6), diabetic with periodontal disease (G7), and diabetic with both pulpal and periodontal disease (G8). Diabetes was induced by injecting streptozotocin, pul-pal disease were induced by exposing pulpal tissue to the oral environment, and periodontal disease was induced by periodontal ligature. After 30 days, blood was collected by cardiac puncture and the animals were killed. The maxillae were processed for histopathology. Serum creatinine levels were measured by the enzymatic method. The total assessed values were statistically analyzed by analysis of variance and Tukey’s test (p < 0.05). Results: Serum creatinine levels were significantly higher in diabetic rats than that in all normoglycemic rats (p < 0.05). The presence of pulpal and periodontal disease increased the serum creatinine levels in normoglycemic and diabetic rats, but there was no statistical difference between the groups (p > 0.05). Conclusions: We found that the serum creatinine level was higher in diabetic rats and may be related to the pres-ence of oral infections. Clinical significance: Changes in serum creatinine level may be related to the presence of oral infections and diabetes.

Genital infections of HIV-infected women assisted by a specialized service

The present study aimed at analyzing the persistence/recurrence of genital infections and its associated factors in HIV-infected women. Fifty-eight women treated for chlamydial infection, trichomoniasis, vulvovaginal candidiasis, and/or bacterial vaginosis (BV) and who had specimens collected for cure control up to one year after treatment were studied. Diagnoses were performed by the Gram staining method for cases of BV and candidiasis and by T. vaginalis culture and qualitative PCR for C. trachomatis. Antiretroviral therapy was used by 79.3% of patients, and 62.1% showed an undetectable HIV plasma load. The most frequent infection was BV with persistence/recurrence of 52.4%, which was associated with a longer time period between treatment and cure control (P = 0.0455), postmenopausal period (P = 0.0451), and having a steady partner (P = 0.007). Persistence/recurrence of vulvovaginal candidiasis was observed in 25%, trichomoniasis in 23.1%, and chlamydial infection in 10.5%. The letter was associated with inadequate treatment of the partner (P = 0.073) among patients with low T CD4 lymphocyte counts. The majority of women in the present study showed good HIV-infection control and a vulnerable sexual behavior, which stress the importance of maintaining gynecological followup.

Secondary aspergillus fumigatus infection associated with coloidal goiter in a Black-masked Lovebird (Agapornis personata)

Aspergillosis is caused by fungus of Aspergillus genus. Is a multifactorial secondary disease and occurs mainly to immunodeficiency. Goiter is the name to non-inflammatory and non-neoplasic thyroid growth which affecting the animal metabolism. In this report we describe a case of aspergillosis and colloidal goiter in a male Black-masked lovebird (Agapornis personata) diagnosed by post mortem exam. The bird was presented for examination due to severe respiratory signs. An initial palliative treatment was performed in order to relieve the symptoms. Despite this, the patient came to die without performing additional ancillary tests. On gross exam, a pulmonary nodule was observed from which we were able to isolate Aspergillus fumigatus on microbial culture. Histological assessment revealed pulmonary aspergilosis and colloid goiter. Based on histopathological and microbiological assessments we conclude that infection probably was secondary to colloid goiter.

Produção e caracterização parcial da xilanase produzida por Aspergillus sp

As xilanases são enzimas que hidrolisam as ligações 1,4-β-xilosídicas da xilana e que possuem potencial biotecnológico em vários processos industriais, como na clarificação de sucos e vinhos, na fabricação de pães, na produção de biocombustíveis e no tratamento das polpas celulósicas. Além disso, os produtos de hidrólise da xilana, xilooligossacarídeos, podem ser utilizados como ingredientes prebióticos, ou seja, ingredientes nutricionais não digeríveis que estimulam seletivamente a proliferação e a atividade de bactérias benéficas do cólon. O uso de enzimas microbianas nas indústrias se dá devido às suas diversas vantagens sobre os métodos químicos e à facilidade de obtenção do microrganismo. Assim, os objetivos deste trabalho foram: produzir e caracterizar parcialmente a xilanase de Aspergillus sp isolado de pó de café utilizado, purificar parcialmente a enzima produzida e analisar os produtos de hidrólise da xilana. O Aspergillus sp mostrou-se bom produtor de xilanase quando o pó de sabugo de milho foi utilizado como fonte de carbono e a xilanase produzida apresentou melhor atividade específica quando o período de cultivo foi de 168 horas. Os processos de precipitação de proteínas e diálise promoveram o aumento da atividade específica do extrato. A xilanase de Aspergillus sp apresentou pH e temperatura de máxima atividade semelhantes aos de produzidas por Aspergillus niger isolados de outras fontes. A hidrólise da xilana produziu xilose e xilooligossacarídeos. Além da xilanase, o fungo revelou ser produtor de outras proteínas que podem ser estudadas em pesquisas futuras.

Determinação das condições ótimas de fermentação para produção de enzimas celulolíticas pelo fungo Aspergillus niger derivado-marinho, utilizando diferentes subprodutos agroindustriais

Pós-graduação em Biotecnologia - IQ

Reo-Like Virus in White Shrimp Penaeus vannamei (Crustacea: Decapoda): Co-occurrence with Baculovirus penaei in Experimental Infections

A virus, tentatively identified as reo-like, occurred concurrently with experimentally-induced Baculovirus penaei (BP) infection in cultured white shrimp larvae Penaeus vannamei. Each shrimp with a reo-like viral infection also had a BP infection, but not all BP-infected shrimp had a reo-like infection. Both viruses occurred in the same tissues and occasionally withln the same cell. The reolike virus developed in epithelial cells of the anterior midgut and in reserve- and fibrillar-cells of the hepatopancreas. The paraspherical and non-enveloped reo-like virions (ca. 50 nm diam.) occurred as unordered aggregates in the cell cytoplasm. Their etiology has not been determined. Reo-like virions may have been introduced along with the BP virus, or, were latent and only manifested due to stress induced by the more pathogenic BP virus.

Echinococcus Infections in Man and Animals in Kenya

1. Hydatid cysts are found in more than 30 per cent of all cattle, sheep and goats in Kenya, but the disease is prevalent in man only in the semi-desert area of Turkana. Up to the time of the present investigation the life-cycle of the parasite in East Africa had not been studied, but it was suggested that wild carnivores, such as hyenas and jackals, might be the main hosts of the adult worms. 2. One hundred and forty-three carnivores, representing 23 species, have been examined. Echinococcus adults were found in 27 out of 43 domestic dogs (Canis familiaris), in three out of four hunting dogs (Lycaon pictus), in one out of nine jackals (Thos mesomelas), and in three out of 19 hyaenas (Crocuta crocuta). 3. A detailed morphological study was made of the Kenya material. After comparison with specimens from other parts of the world, it was concluded that the only species occurring in Kenya was E. granulosus, but it is possible that the minor morphological and biological differences are evidence of distinct strains. Further laboratory studies are necessary to compare the parasite from man and animals in different parts of Kenya with material from elsewhere. 4. A search was made for larval hydatids in 92 ungulates representing 18 species, and in a miscellaneous collection of nearly 2,000 rodents and primates representing a further 31 species. Only one animal was positive, a wildebeest (Gorgon taurinus). 5. The infections in the wild carnivores were all very light; only domestic dogs were heavily infected. It is concluded that the main cycle of transmission in Kenya is between dogs and domestic livestock. 6. Turkana tribesmen are the most heavily infected people in Kenya, either because the strain of parasite is more pathogenic to man in that area, or, more probably, because of the intimate contact between children and the large number of infected dogs. A particularly dangerous custom in the area is the use of dogs to clean the face and anal regions of babies when they vomit or have diarrhea. No satisfactory explanation can be given for the rarity of the disease in man in many of the other areas of Kenya where hydatids are very common in domestic animals. 7. The control of the disease will depend upon an active health-education campaign, together with the destruction of all unregistered dogs and improvement in meat hygiene.

Experimental Bolbophorus damnificus (Digenea: Bolbophoridae) Infections in Piscivorous Birds

In order to determine potential definitive hosts of the digenetic trematode, Bolbophorus damnificus, two American White Pelicans (Pelecanus erythrorhynchos), two Double-crested Cormorants (Phalacrocorax auritus), two Great Blue Herons (Ardea herodias), and two Great Egrets (Ardea alba) were captured, treated with praziquantel, and fed channel catfish (Ictalurus punctatus) infected with B. damnificus metacercariae. Patent infections of B. damnificus, which developed in both American White Pelicans at 3 days post-infection, were confirmed by the presence of trematode ova in the feces. Mature B. damnificus trematodes were recovered from the intestines of both pelicans at 21 days post-infection, further confirming the establishment of infection. No evidence of B. damnificus infections was observed in the other bird species studied. This study provides further evidence that Double-crested Cormorants, Great Blue Herons, and Great Egrets do not serve as definitive hosts for B. damnificus.

Produção e caracterização parcial da xilanase produzida por Aspergillus sp

As xilanases são enzimas que hidrolisam as ligações 1,4-β-xilosídicas da xilana e que possuem potencial biotecnológico em vários processos industriais, como na clarificação de sucos e vinhos, na fabricação de pães, na produção de biocombustíveis e no tratamento das polpas celulósicas. Além disso, os produtos de hidrólise da xilana, xilooligossacarídeos, podem ser utilizados como ingredientes prebióticos, ou seja, ingredientes nutricionais não digeríveis que estimulam seletivamente a proliferação e a atividade de bactérias benéficas do cólon. O uso de enzimas microbianas nas indústrias se dá devido às suas diversas vantagens sobre os métodos químicos e à facilidade de obtenção do microrganismo. Assim, os objetivos deste trabalho foram: produzir e caracterizar parcialmente a xilanase de Aspergillus sp isolado de pó de café utilizado, purificar parcialmente a enzima produzida e analisar os produtos de hidrólise da xilana. O Aspergillus sp mostrou-se bom produtor de xilanase quando o pó de sabugo de milho foi utilizado como fonte de carbono e a xilanase produzida apresentou melhor atividade específica quando o período de cultivo foi de 168 horas. Os processos de precipitação de proteínas e diálise promoveram o aumento da atividade específica do extrato. A xilanase de Aspergillus sp apresentou pH e temperatura de máxima atividade semelhantes aos de produzidas por Aspergillus niger isolados de outras fontes. A hidrólise da xilana produziu xilose e xilooligossacarídeos. Além da xilanase, o fungo revelou ser produtor de outras proteínas que podem ser estudadas em pesquisas futuras.