Response surface optimisation of medium components for naringinase production from staphylococcus xylosus MAK 2

Response surface methodology was used to optimize the fermentation medium for enhancing naringinase production by Staphylococcus xylosus. The first step of this process involved the individual adjustment and optimization of various medium components at shake flask level. Sources of carbon (sucrose) and nitrogen (sodium nitrate), as well as an inducer (naringin) and pH levels were all found to be the important factors significantly affecting naringinase production. In the second step, a 22 full factorial central composite design was applied to determine the optimal levels of each of the significant variables. A second-order polynomial was derived by multiple regression analysis on the experimental data. Using this methodology, the optimum values for the critical components were obtained as follows: sucrose, 10.0%; sodium nitrate, 10.0%; pH 5.6; biomass concentration, 1.58%; and naringin, 0.50% (w/v), respectively. Under optimal conditions, the experimental naringinase production was 8.45 U/mL. The determination coefficients (R ²) were 0.9908 and 0.9950 for naringinase activity and biomass production, respectively, indicating an adequate degree of reliability in the model.

Community onset staphylococcus aureus infections in Melbourne

Objective: To investigate the relative importance of methicillin resistant Staphylococcus aureus (MRSA) in the community in Melbourne by describing circulating S. aureus strains and infection characteristics.

Methods: Patients with any community-onset S. aureus infection were identified via clinical specimens submitted to a community-based pathology service in 2006. The referring doctors confirmed community onset and defined site and severity of each infection. Patient isolates were characterised by antibiotic resistance subtype and presence of the Panton-Valentine leukocidin gene (pvl).

Results: Between April and September 2006, 2,094 S. aureus isolates were processed. Of these, 133 (6.4%) were multiresistant MRSA (mMRSA) and 110 (5.3%) were resistant to less than 3 non-betalactam antibiotics (non-multiresistant MRSA or nmMRSA). We followed-up all nmMRSA (34) and mMRSA (15) confirmed community-onset infections, and a random subset of eligible patients with MSSA infections (57), for whom clinical data were available from referring doctors (82% response).

The majority of isolates were from skin infections (99/106), but drainage was performed in less than one third of cases (29/99). Antibiotics were prescribed for 89% (95%CI: 82, 94) of infections. The isolates were resistant to the prescribed antibiotic 100% of the time for mMRSA infections and 80% for nmMRSA. Those with infections caused by MRSA had on average one additional visit to their doctor compared with MSSA infections.

Ten nmMRSA clones were identified, including one new pvl positive nmMRSA. Of the 29 nmMRSA isolates, 14 were pvl positive (48%; 95%CIs: 30%, 66%) compared with 16% of MSSA and 0% mMRSA.

Patients with an infection caused by pvl positive strains (23) were younger ((mean age 23 years (95%CI: 16, 30) compared with the 55 years (95%CI: 50, 61)). Infection site also varied with presence of pvl; more pvl positive infections were found in the axilla (17.9% compared with 0%) and head and neck (35.7% compared with 8.2%), and less for the leg or foot (21.4% compared with 55.7%).

Conclusions: We estimate that 3.5% of community-onset S. aureus infections in Melbourne in 2006 were caused by MRSA, and 70 to 90% of patients with MRSA infections were treated initially with antibiotics to which their isolate was resistant. pvl positive isolates of S. aureus were associated with younger age and axillary or head and neck infections.

Molecular identification of Staphylococcus xylosus MAK2, a new α-l-rhamnosidase producer

A bacterial strain, MAK-2, was isolated as a producer of α-l-rhamnosidase from a soil sample of Dehradoon, India. The strain was identified based on morphology, physiological tests and 16S rDNA analysis. The phylogenetic analysis based on the 16S rDNA sequence, identified the isolate as Staphylococcus xylosus, a nonpathogenic member of CNS (coagulase-negative staphylococci) family. The strain was capable of producing α-l-rhamnosidase by hydrolysing flavonoids thus confirming potential application in the citrus-processing industry.

Citrus peel influences the production of an extracellular naringinase by Staphylococcus xylosus MAK2 in a stirred tank reactor

Staphylococcus xylosus MAK2, Gram-positive coccus, a nonpathogenic member of the coagulase-negative Staphylococcus family was isolated from soil and used to produce naringinase in a stirred tank reactor. An initial medium at pH 5.5 and a cultivation temperature of 30°C was found to be optimal for enzyme production. The addition of Ca⁺² caused stimulation of enzyme activity. The effect of various physico-chemical parameters, such as pH, temperature, agitation, and inducer concentration was studied. The enzyme production was enhanced by the addition of citrus peel powder (CPP) in the optimized medium. A twofold increase in naringinase production was achieved using different technological combinations. The process optimization using technological combinations allowed rapid optimization of large number of variables, which significantly improved enzyme production in a 5-l reactor in 34 h. An increase in sugar concentration (15 gl^-1) in the fermentation medium further increased naringinase production (8.9 IUml^-1) in the bioreactor. Thus, availability of naringinase renders it attractive for potential biotechnological applications in citrus processing industry.

Fermentative production of rhamnosidase from Staphylococcus xylosus MAK 2 in a bioreactor for bio-energy generation

Increasing concern about the environment, food and feed shortages and hike in the price of petroleum have stimulated interest in new ways of producing biofuels. The interest is rapidly increasing towards converting agricultural wastes to commercially valuable products. Biofuels made from waste biomass can offer immediate and sustained greenhouse gas advantages. In this direction, we are focusing on Citrus processing waste, a byproduct of juice manufacture, which contains high amount of flavonoids and polysaccharides. There is a considerable industrial interest in the enzymatic transformation of flavonoids to hydrolysis products; that offers a pathway to bio-energy generation. Rhamnosidase of bacterial origin are very few and thus are potentially subject for research.

Staphylococcus xylosus, Gram positive cocci, a nonpathogenic member of CNS family, isolated from soil was used to produce α-L-rhamnosidase. This new strain, so far unknown for the production of α-L-Rhamnosidase, was identified and characterized as Staphyloccocus sp. through biochemical tests and 16S DNA sequence analysis. Effect of various medium and process parameters like pH, temperature, aeration and agitation rates and inducer concentration were studied. Further, the enzyme activity was enhanced by adding the inducer and divalent metal ion to the optimised fermentation medium. We have recovered important sugars “rhamnose” and “galacturonic acid” from the processed waste which would be utilized for ethanol production. This presentation will summarize current efforts to develop an enzymatic treatment which would facilitate the economical processing of citrus waste for bioenergy generation.

The role of micro-organisms (staphylococcus aureus and Candida Albicans) in the pathogenesis of breast pain and infection in lactation women : study protocol

Background: The CASTLE (Candida and Staphylococcus Transmission: Longitudinal Evaluation) study will investigate the micro-organisms involved in the development of mastitis and “breast thrush” among breastfeeding women. To date, the organism(s) associated with the development of breast thrush have not been identified. The CASTLE study will also investigate the impact of physical health problems and breastfeeding problems on maternal psychological health in the early postpartum period.

Methods/Design: The CASTLE study is a longitudinal descriptive study designed to investigate the role of Staphylococcus spp (species) and Candida spp in breast pain and infection among lactating women, and to describe the transmission dynamics of S. aureus and Candida spp between mother and infant. The relationship between breastfeeding and postpartum health problems as well as maternal psychological well-being is also being investigated. A prospective cohort of four hundred nulliparous women who are at least thirty six weeks gestation pregnant are being recruited from two hospitals in Melbourne, Australia (November 2009 to June 2011). At recruitment, nasal, nipple (both breasts) and vaginal swabs are taken and participants complete a questionnaire asking about previous known staphylococcal and candidal infections. Following the birth, participants are followed-up six times: in hospital and then at home weekly until four weeks postpartum. Participants complete a questionnaire at each time points to collect information about breastfeeding problems and postpartum health problems. Nasal and nipple swabs and breast milk samples are collected from the mother. Oral and nasal swabs are collected from the baby. A telephone interview is conducted at eight weeks postpartum to collect information about postpartum health problems and breastfeeding problems, such as mastitis and nipple and breast pain.

Discussion: This study is the first longitudinal study of the role of both staphylococcal and candidal colonisation in breast infections and will help to resolve the current controversy about which is the primary organism in the condition known as breast thrush. This study will also document transmission dynamics of S. aureus and Candida spp between mother and infant. In addition, CASTLE will investigate the impact of common maternal physical health symptoms and the effect of breastfeeding problems on maternal psychological well-being.

Does Candida and/or Staphylococcus play a role in nipple and breast pain in lactation? A cohort study in Melbourne, Australia

Objective: To investigate Candida species and Staphylococcus aureus and the development of 'nipple and breast thrush' among breastfeeding women. Design: Prospective longitudinal cohort study. Setting: Two hospitals in Melbourne, Australia (one public, one private) with follow-up in the community. Participants: 360 nulliparous women recruited at 36 weeks' gestation from November 2009 to June 2011. Participants were followed up six times: in hospital, at home weekly until 4 weeks postpartum and by telephone at 8 weeks. Main outcome measures: Case definition 'nipple and breast thrush': burning nipple pain and breast pain (not related to mastitis); detection of Candida spp (using culture and PCR) in the mother's vagina, nipple or breast milk or in the baby's mouth; detection of S aureus in the mother' nipple or breast milk. Results: Women with the case definition of nipple/ breast thrush were more likely to have Candida spp in nipple/breast milk/baby oral samples (54%) compared to other women (36%, p=0.014). S aureus was common in nipple/breast milk/baby samples of women with these symptoms as well as women without these symptoms (82% vs 79%) (p=0.597). Time-to-event analysis examined predictors of nipple/breast thrush up to and including the time of data collection. Candida in nipple/breast milk/baby predicted incidence of the case definition (rate ratio (RR) 1.87 (95% CI 1.10 to 3.16, p=0.018). We do not have evidence that S aureus colonisation was a predictor of these symptoms (RR 1.53, 95% CI 0.88 to 2.64, p=0.13). Nipple damage was also a predictor of these symptoms, RR 2.30 (95% CI 1.19 to 4.43, p=0.012). In the multivariate model, with all three predictors, the RRs were very similar to the univariate RRs. This indicates that Candida and nipple damage are independent predictors of our case definition.

Community-onset Staphylococcus aureus infections presenting to general practices in South-eastern Australia

Community-acquired Staphylococcus aureus infections are a public health concern, yet little is known about infections that do not present to hospital. We identified community-onset S. aureus infections via specimens submitted to a community-based pathology service. Referring doctors confirmed eligibility and described infection site, severity and treatment. Isolates were characterized on antibiotic resistance, PFGE, MLST/SCCmec, and Panton–Valentine leukocidin (PVL), representing 106 community-onset infections; 34 non-multiresistant methicillin-resistant S. aureus (nmMRSA) (resistant to <3 non-β-lactam antibiotics), 15 multiply antibiotic-resistant MRSA (mMRSA) and 57 methicillin-sensitive S. aureus (MSSA). Most (93%) were skin and soft tissue infections. PVL genes were carried by 42% of nmMRSA isolates [95% confidence interval (CI) 26–61] and 15% of MSSA (95% CI 8–28). PVL was associated with infections of the trunk, head or neck (56·4% vs. 24·3%, P= 0·005) in younger patients (23 vs. 52 years, P< 0·001), and with boils or abscesses (OR 8·67, 95% CI 2·9–26·2), suggesting underlying differences in exposure and/or pathogenesis.