Staphylococci that carry the nuc gene

Inflammation of the mammary gland, known as mastitis, is a serious problem, because it is responsible for the decrease in milk yield and quality. Bacteria of the species Staphylococcus aureus, causing intramammary infections in small ruminants, are often isolated from milk samples and frequently resistant to antimicrobials. The thermostable nuclease encoded by the nuc gene is a virulence factor, as it promotes evasion of neutrophil extracellular traps (NETs), which are networks of DNA strands with antimicrobial proteins. The nuc gene is present in most S. aureus, however some isolates not carrying this gene have been described. Moreover, the nuc gene has also been detected in other Staphylococcus species, notably in S. intermedius, S. hyicus and S. simulans. The aim of this study was to assess the presence of the nuc gene in staphylococci isolated from goat and sheep milk samples. Three hundred and sixty-eight samples of milk were collected, from 187 animals, belonging to six flocks in Alentejo. Ninety-one isolates of the genus Staphylococcus were identified by classical biochemical methods and through the VITEK2 microbial identification system (BioMérieux, France). The presence of the nuc gene was assessed by PCR. The reference strain S. aureus ATCC 25923 was used as positive control. According to VITEK2, 24 S. caprae, 13 S. epidermidis, 13 S. chromogenes, 11 S. aureus, nine S. simulans, five S. warneri, five S. lentus, four S. capitis, four S. haemolyticus, one S. auricularis, one S. hominis spp. hominis and one Staphylococcus spp. (not identified to species level) were identified. Twenty-three out of the 91 isolates (25.3%) carry the nuc gene. With the pair of primers used, this gene was detected in seven different Staphylococcus species: 10 S. aureus (43.5%), three S. warneri (13%), three S. lentus (13%), two S. caprae (8.7%), two S. epidermidis (8.7%), two S. chromogenes (8.7%) and one S. capitis (4.3%). Furthermore, the nuc gene was not detected in one S. aureus isolate. We suggest that the presence/absence of the nuc gene is not a reliable method for the identification of S. aureus and that this virulence factor may be involved in small ruminant mastitis pathophysiology.CNPq - Conselho Nacional de Desenvolvimento Científico e Tecnológico-Brasil 249398/2013-3 and by National Funds through FCT - Foundation for Science and Technology under the Project UID/AGR/00115/2013. M. Laranjo acknowledges a Post-Doc research grant also from FCT (SFRH/BPD/108802/2015)

Utilização de própolis como alternativa no controlo de mastites para garantir a qualidade do leite e proteção da saúde pública.

Os antimicrobianos e antisséticos são utilizados para o controlo das mastites. Esta prática conduz a eliminação de resíduos no leite além de induzir a pressão de seleção sobre estirpes resistentes a antimicrobianos. A própolis é uma massa, utilizada para proteger as colmeias das abelhas (Apis mellifera), formada por resinas retiradas de vários vegetais, selecionadas de forma natural pelas abelhas. Este produto altamente variável, consoante os vegetais utilizados, apresenta colorações diversas e também qualidades antimicrobianas variáveis. O objetivo deste estudo foi testar a suscetibilidade, de isolados de Staphylococccus aureus provenientes de amostras de leite de cabras e ovelhas com mastite, a 16 antimicrobianos utilizados em Portugal no controlo desta doença, e analisar “in vitro” a ação antimicrobiana de extratos etanólicos de própolis (EEP).FCT – Fundação para a Ciência e a Tecnologia no âmbito do Projecto UID/AGR/00115/2013. M. Laranjo agradece a bolsa de Pós-Doutoramento da FCT (SFRH/BPD/108802/2015)

Propolis components accountable for bactericidal accomplishment and antibiofilm activity

This study aimed at evaluating antimicrobial and antibiofilm activity of phenolic compounds present in propolis ethanol extracts (PEE). Seventy per cent ethanol extracts from seven types of propolis, one Green, two Red and four Brown collected in four Brazilian States were prepared and total phenolics, flavonoids, tannins and anthocyanins were assessed by high-performance liquid chromatography (HPLC). Minimum bactericidal concentration (MBC) and inhibitor effect on Staphylococcus aureus biofilm formation and capacity to disrupt established biofilms were assessed towards eight S. aureus isolates from milk of small ruminants with mastitis, one methicillin-resistant S. aureus (MRSA) and S. aureus ATCC 25923. To evaluate different propolis components accountability for bactericidal accomplishment and antibiofilm activity, the results were analysed by the non-parametric Spearman coefficient. Results of phenolic compounds were 216,21 to 312,08 gallic acid milligram equivalent per extract gram (mg EGA/g) of total phenolics, 55,08 to 140,6 quercetin milligram equivalent per extract gram (mg EQ/g) of flavonoids, 118,51 to 3766,16 catechin milligram equivalent per extract gram (mg EC/g) of tannins and 1,03 to 8,39 milligram per extract gram (mg/g) of anthocyanins. Red1 and Red2 showed higher tannin contents, while Red2 exhibited superior amount of anthocyanins and total phenolics. Brown3 presented higher flavonoid quantity. Green, Red1 and Red2 PEE showed the lowest levels of flavonoids, but the higher antimicrobial activity. Most PEE exhibit bactericidal activity at a concentration of 1.6 mg/mL. Brown4 PEE showed the worst capacity to inhibit S. aureus. Green PEE showed to be the most efficient in both preventing and disrupting biofilm. All PEE studied exhibited a better inhibitory activity prior-to than post-biofilm formation. According to non-parametric Spearman correlation analysis, there seems to be a significant negative correlation between the ability to disrupt biofilm and both tannins and anthocyanins contents

Controlo de mastites em pequenos ruminantes: utilização de própolis como alternativa a antimicrobianos

Os antimicrobianos e antisséticos são utilizados para o controlo das mastites. Esta prática conduz a eliminação de resíduos no leite e induz pressão de seleção sobre estirpes resistentes. É urgente desenvolver métodos para controlo de doenças utilizando substâncias com poder antimicrobiano que sejam seguras. A própolis é um produto formado pelas abelhas Apis melífera, que possui qualidades antimicrobianas variáveis. O objetivo deste estudo foi testar a suscetibilidade, de isolados de Staphylococcus aureus provenientes de amostras de leite de cabras e ovelhas, a antimicrobianos e analisar in vitro a ação antimicrobiana de extratos etanólicos de própolis (EEP)

Antimicrobial activity of Brazilian propolis extracts for Staphylococcus aureus of sheep and goats mastitis origin

Twenty-four S. aureus isolates were analysed. From those, 22 were isolated from milk of goats and sheep with clinical and subclinical mastitis, from the region of Vale do São Francisco in the Brazilian Sertão and S. aureus ATCC 25923 plus a MRSA strain were added. Alcoholic extracts were produced from several batches of green, red and brown propolis consisting of 300 g of raw propolis in 700 mL of 70 % ethanol. Four genes related to antimicrobial resistance were assessed: blaZ that determines the resistance to β-lactam antibiotics, and genes icaA, icaD and bap that influence the production of biofilm. For the tests of susceptibility to different types of propolis the microdilution method was used, in triplicate, and dilutions between 0.003672 and 15% were tested, 70 % ethanol consisted of a negative control. The gene blaZ was found in 15 isolates; icaA gene was present in 3 isolates, icaD gene in 2 and bap gene was detected in 6 isolates. All the propolis tested exhibited antimicrobial activity, ranging from 44 to 100 % of susceptible isolates depending on different propolis batches. According to the results of this experiment the green and red propolis appear to have better antimicrobial activity than the brown variety

Antibiofilm activity of Brasilian propolis extracts against Staphylococcus aureus producers isolated from goat and sheep mastitic milk

The aim of this study was to investigate the action of inhibiting S. aureus biofilm formation, and the ability to eliminate formed biofilm, by alcoholic extracts of green, red and brown propolis from Brazil. Ten isolates of S. aureus have been tested, 8 field isolates, 1 MRSA and 1 ATCC 25923, by microplate quantitative method. For the evaluation of inhibitory action, the isolates were inoculated, in triplicate, in TSB 1% glucose in the presence of green (1), red (2) and brown (4) propolis extracts. Biofilm formation was evaluated by optical reading, compared to a negative control consisting of a mixture of TSB and extract. For biofilm elimination assay, extracts were added to plates with 24h cultures of the same isolates. Assays were repeated three times on three different days. Eight out of the 10 isolates produced less biofilm in the presence of the green propolis extracts, so the inhibitory effect is 80%. Brown propolis extracts inhibited the formation of biofilm in 10% to 70% of the isolates and the red extracts in 30% to 80%. Regarding the biofilm elimination activity, green propolis extract was positive for 9 out of the 10 isolates (90%), the brown propolis extracts were positive for 0% to 100% isolates and red extracts for 0% to 10% isolates

A review of vulnerability indicators for deltaic social–ecological systems

The sustainability of deltas worldwide is under threat due to the consequences of global environmental change (including climate change) and human interventions in deltaic landscapes. Understanding these systems is becoming increasingly important to assess threats to and opportunities for long-term sustainable development. Here, we propose a simplified, yet inclusive social–ecological system (SES)-centered risk and vulnerability framework and a list of indicators proven to be useful in past delta assessments. In total, 236 indicators were identified through a structured review of peer-reviewed literature performed for three globally relevant deltas—the Mekong, the Ganges–Brahmaputra–Meghna and the Amazon. These are meant to serve as a preliminary “library” of potential indicators to be used for future vulnerability assessments. Based on the reviewed studies, we identified disparities in the availability of indicators to populate some of the vulnerability domains of the proposed framework, as comprehensive social–ecological assessments were seldom implemented in the past. Even in assessments explicitly aiming to capture both the social and the ecological system, there were many more indicators for social susceptibility and coping/adaptive capacities as compared to those relevant for characterizing ecosystem susceptibility or robustness. Moreover, there is a lack of multi-hazard approaches accounting for the specific vulnerability profile of sub-delta areas. We advocate for more comprehensive, truly social–ecological assessments which respond to multi-hazard settings and recognize within-delta differences in vulnerability and risk. Such assessments could make use of the proposed framework and list of indicators as a starting point and amend it with new indicators that would allow capturing the complexity as well as the multi-hazard exposure in a typical delta SES