Antibacterial activity of the alkaloid extract and isolated compounds from Croton bonplandianum Baill. (Euphorbiaceae) / Atividade anti-bacteriana do extrato alcaloide e de compostos isolados de Croton bonplandianum Baill. (Euphorbiaceae)

ABSTRACT The antibacterial activity of the alkaloid extract from the leaves of Croton bonplandianum Baill. and its main compounds, sparsiflorine and crotsparine, was tested against Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa by the resazurin microtitre-plate method. Pure compounds were identified by spectroscopic techniques, mainly 1D and 2D NMR. The alkaloid extract showed activity particularly against the S. aureus and P. aeruginosa. Regarding the pure compounds, the crotsparine was inactive against the microorganisms assayed, whereas the sparsiflorine indicated a moderate activity similar to the alkaloid extract. The Pseudomonas aeruginosa was the most sensitive of the tested microorganisms with MIC of 0.141 mg/mL. The results suggest that the activity of the extract may be credited mainly to the presence of the sparsiflorine. Although the activity of the sparsiflorine does not get close to the antimicrobial drugs in clinical use, it still could be a lead compound for the development of new antibacterial substances.

RESUMO A atividade antibacteriana do extrato alcaloidal das folhas de Croton bonplandianum Baill., além dos principais compostos isolados, esparsiflorina e crotsparina, foi testada contra Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae e Pseudomonas aeruginosa. Foi utilizado o método de microdiluição em placa empregando resazurina como indicador da viabilidade celular. Os compostos isolados foram identificados por técnicas espectroscópicas, principalmente RMN 1D e 2D. O extrato alcaloidal foi ativo principalmente contra S. aureus e P. aeruginosa. Crotsparina mostrou-se inativa contra todos os micro-organismos testados, enquanto esparsiflorina apresentou atividade moderada, a qual foi similar à do extrato bruto. Pseudomonas aeruginosa foi a mais sensível das bactérias testadas, com CIM de 0,141 mg/mL. Os resultados sugerem que a atividade do extrato pode ser devida em grande medida pela presença de esparsiflorina. Apesar de a CIM da esparsiflorina não ter se aproximado daquela apresentada pelos agentes antimicrobianos em uso clínico, tal composto ainda pode compor um protótipo interessante para o desenvolvimento de novas substâncias antibacterianas.

Expression and lytic efficacy assessment of the Staphylococcus aureus phage SA4 lysin gene

Treatment of bovine mastitis caused by Staphylococcus (S.) aureus is becoming very difficult due to the emergence of multidrug-resistant strains. Hence, the search for novel therapeutic alternatives has become of great importance. Consequently, bacteriophages and their endolysins have been identified as potential therapeutic alternatives to antibiotic therapy against S. aureus. In the present study, the gene encoding lysin (LysSA4) in S. aureus phage SA4 was cloned and the nucleotide sequence was determined. Sequence analysis of the recombinant clone revealed a single 802-bp open reading frame encoding a partial protein with a calculated mass of 30 kDa. Results of this analysis also indicated that the LysSA4 sequence shared a high homology with endolysin of the GH15 phage and other reported phages. The LysSA4 gene of the SA4 phage was subsequently expressed in Escherichia coli. Recombinant LysSA4 induced the lysis of host bacteria in a spot inoculation test, indicating that the protein was expressed and functionally active. Furthermore, recombinant lysin was found to have lytic activity, albeit a low level, against mastitogenic Staphylococcus isolates of bovine origin. Data from the current study can be used to develop therapeutic tools for treating diseases caused by drug-resistant S. aureus strains.

Typing of methicilin-resistant staphylococcus aureus by quantitative antibiogram and arbitrarily primer PCR.

A total of 34 isolates of methicillin-resistant Staphylococcus aureus [MRSA] obtained from Rafidya Hospital, Nablus, Palestine was analyzed by quantitative antibiogram and arbitrarily primed PCR. Quantitative antibiogram typing classified the isolates into 13 groups. The use of AP-PCR provided a high resolution typing method and allowed the definition of 21 different clusters. However, three major clusters, based on the combination of both typing methods, represented 44% of the isolates. Resistance rates of MRSA isolates were ciprofloxacin [27%], clindamycin [47%], erythromycin [88%], gentamicin [71%] and co-trimoxazole [3%]. No resistance to vancomycin was observed. Eight of the cluster isolates were noted to be multiple resistant, i.e. resistant to ciprofloxacin, erythromycin and gentamicin or clindamycin

Characterization of methicillin resistant Staphylococcus aureus strains by a set of MRSA phages.

A recently developed international set of phages for typing methicillin resistant Staphylococcus aureus (MRSA) was used to characterize 300 strains of MRSA. The results were compared to that of phage typing with conventional phages and reverse phage typing. The use of MRSA phages increased the percentage typability from 17.6 per cent with the conventional set to 45.6 per cent with MRSA set and the strains were recognised as five distinct phage pattern viz., 622, M3/M5, MR8/MR12/MR25, 30/33/38 and mixed group. Phage type 622 was the most prevalent. On reverse phage typing 73 per cent strains could be typed with most strains belonging to one pattern i.e., 83A complex with limited discrimination. These 83A strains were highly resistant to erythromycin (98%) and tetracycline (93%). Our results show that MRSA phages are more useful in increasing typability and discrimination between the strains as compared to the conventional phages, reverse phage typing, and antibiogramtyping.

Predominant phage types of coagulase positive staphylococci in hospital infections.

360 strains of Staphylococcus aureus isolated from various clinical specimens were subjected to bacteriophage typing. 247(68.6%) strains were typable. Among the typable strains 75(20.83%) belonged to phage group I, 45(12.5%) belonged to phage group III, 6(1.67%) belonged to phage group II and 14(3.89%) strains belonged to miscellaneous group. By far, the largest was the mixed group having 107(29.72%) strains. 113 strains (31.4%) were untypable. All the strains were tested for antibiotic sensitivity test. 287 (79.7%) were multiple drug resistant strains.

Phage-mediated conjugation in staphylococci as a means of transfer of drug-resistance.

Penicillin resistance plasmid was transferred from Staphylococcus aureus B4 (PcrKms, donor) to S. aureus ML351 (PcsKmr, recipient) by co-cultivation of the donor with the recipient in nutrient broth with or without the modifying effects of CaCl2 or sodium dodecyl sulfate. It was found that the transfer of drug-resistance occurred maximally between 6 and 18 hr postinoculation; however, addition of DNase (200 micrograms/ml) could totally prevent such a transfer up to 6 hr and significantly reduce it thereafter. Cell-free filtrate of the donor culture when mixed with the recipient was ineffective in bringing about the transfer of Pcr.

Staphylococcus aureus: incidência em pacientes hospitalizados e profissionais de saúde vinculados ao hospital das clínicas de Goiânia/UFG - Caracterizaçäo das amostras isoladas pela fagotipagem e antibiograma / Staphylococcus aureus: incidence in hospitalized patients and health professionals from the Hospital das Clínicas de Goiânia-UFG - Caracterization of isolated samples by the phage typing and antibiogram

Com a finalidade de detectar portadores de Staphylococcus aureus relacionados a ambiente hospitalar, foram amostrados, em duas ou três áreas corporais diferentes, 112 indivíduos ligados ao Hospital das Clínicas de Goiânia/UFG, sendo 92 pacientes hospitalizados e 20 profissionais, internados e lotados nas enfermarias de Obstetrícia, Ginecologia, Pediatria e Dermatologia. O percentual de portadores foi 39,1 pôr cento entre os pacientes internados e de 20,0 pôr cento entre os profissionais. Os pacientes internados forneceram um percentual de 20,9 pôr cento de amostras positivas para Staphylococcus aureus e os profissionais 12,5 pôr cento. A área anatômica que apresentou o maior frequencial de colonizaçäo foram as fossas nasais, para as quais encontrou-se um índice de 54,7 pôr cento entre os pacientes internados e 80,0 pôr cento entre os profissionais. A identificaçäo das amostras de Staphylococcus aureus pela fagotipagem revelou que 69,9 pôr cento das amostras isoladas dos pacientes internados foram tipáveis, havendo maior percentual (35,1 pôr cento) de cepas pertencentes ao grupo fágico III, além dos experimentais (32,5 pôr cento). Entre os profissionais, 60,0 pôr cento das amostras tipáveis, todas pertencentes ao grupo fágico III. O antibiograma demonstrou que, tanto as amostras provenientes dos pacientes internados como a dos profissionais, apresentaram resistência mais acentuada à Penicilina G cristalina, à Ampicilina e à Tetraciclina como também, maior sensibilidade à Amicacina, Cefalotina e Associaçäo Trimetoprim-sulfatoxazol (Bactrim)

Toxic shock syndrome associated with phage-group-III Staphylococcus aureus.

A case of toxic shock syndrome in a previously-healthy man was reported. The clinical features are in accordance with the criteria of diagnosis defined by the Center for Disease Control (CDC) of USA. Phage-group-III Staphylococcus aureus, demonstrated to produce enterotoxin F, was isolated from an axillary abscess.

Observacao sobre o padrao fagico de cepas de Staphylococcus aureus isoladas da boca e do nariz de individuos saos. / Phage pattern of Staphylococus aureus strains isolated from mouth and nose of healthy subjects

Foram analisadas 68 cepas de Staphylococcus aureus isoladas da boca (saliva e lingua) e nariz de portadores assintomaticos que albergavam essa bacteria nos tres nichos, simultaneamente. Embora os tres nichos apresentassem equivalencia quanto ao percentual de isolamentos, foi constatado que as cepas isoladas do nariz diferiam, quanto ao seu padrao fagico, principalmente daquelas isoladas da lingua. No primeiro caso, as cepas apresentaram-se mais sensiveis aos fagos do grupo I enquanto que, as isoladas do segundo nicho, foram mais sensiveis aos fagos do grupo III.Considerando estas observacoes e recomendavel, quando da deteccao de portadores de Staphylococcus aureus, a pesquisa dessa bacteria em material colhido do nariz e da boca, simultaneamente