Johnson Lin

Purification and biochemical characteristics of β‐D‐glucosidase from a thermophilic fungus, Thermomyces lanuginosus–SSBP

An extracellular xylanase was purified to homogeneity from the culture filtrate of a thermophilic fungus, Thermomyces lanuginosus‐SSBP, and its biochemical characteristics were studied. A yield of 70–80% was achieved through the procedures of 80%‐satd. ammonium sulphate precipitation, DEAE‐Sephadex A25 and quaternary aminoethyl (QAE)‐Sephadex A25 column chromatography. The molecular mass of the purified xylanase was 23.6 kDa, as analysed by SDS/PAGE, with a pI value of 3.8. The molar absorption coefficient of the absorbance at 280 nm was 6.8×104 M−1·cm−1. The specific activity, calculated using the dinitrosalicylic acid (DNS) method, was 3500 units/mg. The enzyme reactions followed Michaelis–Menten kinetics with Kappm and Vmax values of 3.26 mg/ml and 6300 units/ml per mg of protein respectively, as obtained from a Lineweaver–Burk plot. The xylanase contained no other enzyme activity (cellulase, β‐glucosidase, β‐mannosidase, α‐arabinofuranosidase, or β‐xylosidase) except for the hydrolysis of xylan substrate. The optimal temperature of the enzyme assay was 70–75 °C. The enzyme retained full activity after a 60 °C incubation for 3 h. The optimal pH of xylanase activity was 6.5 and the enzyme appeared to be stable over a broad pH range (pH 5–12) under the assay conditions. The majority of the metal ions tested had no effect on the enzyme activity, with the exception of Pb2+ (modest inhibitor) and Hg2+ (strong inhibitor). The results showed that one or two tryptophan residues oxidized by N‐bromosuccinamide per enzyme molecule was sufficient to inhibit the enzyme activity completely, thus indicating that the tryptophan residues play an important role in the catalytical processes of the enzyme reaction. Because of the outstanding properties of the purified xylanase from the SSBP strain, this xylanase has a potential use in biopulping processes and other industrial applications.

Antimicrobial susceptibility patterns and characterization of clinical isolates of Staphylococcus aureus in KwaZulu-Natal province, South Africa

BackgroundAntimicrobial resistance of Staphylococcus aureus especially methicillin-resistant S. aureus (MRSA) continues to be a problem for clinicians worldwide. However, few data on the antibiotic susceptibility patterns of S. aureus isolates in South Africa have been reported and the prevalence of MRSA in the KwaZulu-Natal (KZN) province is unknown. In addition, information on the characterization of S. aureus in this province is unavailable. This study investigated the susceptibility pattern of 227 S. aureus isolates from the KZN province, South Africa. In addition, characterization of methicillin-sensitive S. aureus (MSSA) and MRSA are reported in this survey.MethodsThe in-vitro activities of 20 antibiotics against 227 consecutive non-duplicate S. aureus isolates from clinical samples in KZN province, South Africa were determined by the disk-diffusion technique. Isolates resistant to oxacillin and mupirocin were confirmed by PCR detection of the mecA and mup genes respectively. PCR-RFLP of the coagulase gene was employed in the characterization of MSSA and MRSA.ResultsAll the isolates were susceptible to vancomycin, teicoplanin and fusidic acid, and 26.9% of isolates studied were confirmed as MRSA. More than 80% of MRSA were resistant to at least four classes of antibiotics and isolates grouped in antibiotype 8 appears to be widespread in the province. The MSSA were also susceptible to streptomycin, neomycin and minocycline, while less than 1% was resistant to chloramphenicol, ciprofloxacin, rifampicin and mupirocin. The inducible MLSB phenotype was detected in 10.8% of MSSA and 82% of MRSA respectively, and one MSSA and one MRSA exhibited high-level resistance to mupirocin. There was good correlation between antibiotyping and PCR-RFLP of the coagulase gene in the characterization of MRSA in antibiotypes 1, 5 and 12.ConclusionIn view of the high resistance rates of MRSA to gentamicin, erythromycin, clindamycin, rifampicin and trimethoprim, treatment of MRSA infections in this province with these antibacterial agents would be unreliable. There is an emerging trend of mupirocin resistance among S. aureus isolates in the province. PCR-RFLP of the coagulase gene was able to distinguish MSSA from MRSA and offers an attractive option to be considered in the rapid epidemiological analysis of S. aureus in South Africa. Continuous surveillance on resistance patterns and characterization of S. aureus in understanding new and emerging trends in South Africa is of utmost importance.

Cooccurrence of Predominant Panton-Valentine Leukocidin-Positive Sequence Type (ST) 152 and Multidrug-Resistant ST 241 Staphylococcus aureus Clones in Nigerian Hospitals

ABSTRACT Ninety-six clinical isolates of Staphylococcus aureus from Nigeria were characterized phenotypically and genetically. Twelve multidrug-resistant methicillin (meticillin)-resistant S. aureus (MRSA) isolates carrying a new staphylococcal cassette chromosome mec element and a high proportion of Panton-Valentine leukocidin (PVL)-positive methicillin-susceptible S. aureus (MSSA) isolates were observed. The cooccurrence of multidrug-resistant MRSA and PVL-positive MSSA isolates entails the risk of emergence of a multidrug-resistant PVL-positive MRSA clone.

Water quality indicators: bacteria, coliphages, enteric viruses

Water quality through the presence of pathogenic enteric microorganisms may affect human health. Coliform bacteria, Escherichia coli and coliphages are normally used as indicators of water quality. However, the presence of above-mentioned indicators do not always suggest the presence of human enteric viruses. It is important to study human enteric viruses in water. Human enteric viruses can tolerate fluctuating environmental conditions and survive in the environment for long periods of time becoming causal agents of diarrhoeal diseases. Therefore, the potential of human pathogenic viruses as significant indicators of water quality is emerging. Human Adenoviruses and other viruses have been proposed as suitable indices for the effective identification of such organisms of human origin contaminating water systems. This article reports on the recent developments in the management of water quality specifically focusing on human enteric viruses as indicators.

Ozone-initiated disinfection kinetics of Escherichia coli in water

The effect of ozonation on the rate of disinfection of Escherichia coli was investigated as a function of ozone concentration, ozonation duration and flow rates. Ozone was generated in situ using Corona discharge method using compressed oxygen stream and depending on the oxygen flux the ozone concentrations ranged from 0.91–4.72 mg/L. The rate of disinfection of all the three microbes followed pseudo–first-order kinetics with respect to the microbe count and first order with respect to ozone concentration. The influence of pH and temperature the aqueous systems on the rate of ozone initiated disinfection of the microbe was investigated. The inactivation was faster at lower pH than at basic pH. Molecular ozone is found more effective in disinfection than hydroxyl radicals. Two reported mechanisms for antimicrobial activity of ozone in water systems from the literature are discussed. Based on the experimental findings a probable rate law and mechanism are proposed. Ozonation of natural waters significantly decreased the BOD levels of the control and microbe contaminated waters.

Characterization of meticillin-resistant Staphylococcus aureus isolates from hospitals in KwaZulu-Natal province, Republic of South Africa.

Epidemiological data based on phenotypic and molecular characterization of meticillin-resistant Staphylococcus aureus (MRSA) in sub-Saharan Africa are limited. This investigation studied 61 MRSA isolates obtained from 13 health-care institutions in KwaZulu-Natal (KZN) province, South Africa, from March 2001 to August 2003. More than 80 % of the isolates were resistant to at least four classes of antibiotics and six isolates were resistant to the aminoglycoside, macrolide-lincosamide and tetracycline groups of antibiotics, heavy metals and nucleic acid-binding compounds. PFGE of SmaI-digested genomic DNA revealed seven types, designated A-G. Type A was the main pulsotype (62.3 %) and was identified in 11 of the 13 health-care institutions, suggesting that it represented a major clone in health-care institutions in KZN province. Analysis of representative members of the three major pulsotypes by spa, multilocus sequence typing and SCCmec typing revealed the types t064-ST1173-SCCmec IV and t064-ST1338-SCCmec IV (PFGE type A, single-locus and double-locus variants of ST8), t037-ST239-SCCmec III (PFGE type F) and t045-ST5-SCCmec III (PFGE type G). The combination of various typing methods provided useful information on the geographical dissemination of MRSA clones in health-care institutions in KZN province. The observation of major clones circulating in health-care facilities in KZN province indicates that adequate infection control measures are urgently needed.

Isolation and characterization of diesel oil degrading indigenous microrganisms in Kwazulu-Natal, South Africa

Uncontrolled releases of petroleum compounds that are carcinogenic, mutagenic and are potent immunotoxicants into soil and groundwater poses a serious threat to human and animal health. Biodegradation of hydrocarbon-contaminated soils has been established as an efficient, economic, versatile and environmentally sound treatment. Ten indigenous microorganisms were isolated from contaminated soils using the enrichment technique. Five isolates with the highest degradation potentials under standard degradation conditions were identified as Acinetobacter calcoaceticus (LT1 and ETS2), Acinetobacter sp. (LT1A), Citrobacter freundii (MRC3) and Bacillus pumilus (JLB). B. pumilus achieved 86.94% of diesel degradation in 2 weeks. Additional degradation assay was carried out in liquid media using 3 local commercial fertilizers as nutrient supplements in comparison with the Bushnell-Haas (BH) media. The results show that the addition of fertilizer F1 stimulated diesel degradation by all isolates especially B. pumilus while the addition of fertilizer F3 seemed to strongly inhibit the bacterial ability of diesel degradation. The inoculation with the consortia did not show a higher degradation potential than the individual isolate. The results strongly indicate that environmental conditions of the contaminated sites play a crucial role in the degradation even though additional diesel-degrader has been introduced into the contaminated site.

Phenotypic and molecular characterization of Staphylococcus aureus isolates expressing low- and high-level mupirocin resistance in Nigeria and South Africa

BackgroundMupirocin is a topical antimicrobial agent which is used for the treatment of skin and postoperative wound infections, and the prevention of nasal carriage of methicillin-resistant Staphylococcus aureus (MRSA). However, the prevalence of mupirocin resistance in S. aureus, particularly in MRSA, has increased with the extensive and widespread use of this agent in hospital settings. This study characterized low- and high-level mupirocin-resistant S. aureus isolates obtained from Nigeria and South Africa.MethodsA total of 17 mupirocin-resistant S. aureus isolates obtained from two previous studies in Nigeria and South Africa, were characterized by antibiogram, PCR-RFLP of the coagulase gene and PFGE. High-level mupirocin resistant isolates were confirmed by PCR detection of the mupA gene. The genetic location of the resistance determinants was established by curing and transfer experiments.ResultsAll the low-level mupirocin resistant isolates were MRSA and resistant to gentamicin, tetracycline and trimethoprim. PFGE identified a major clone in two health care institutions located in Durban and a health care facility in Pietermaritzburg, Greytown and Empangeni. Curing and transfer experiments indicated that high-level mupirocin resistance was located on a 41.1 kb plasmid in the South African strain (A15). Furthermore, the transfer of high-level mupirocin resistance was demonstrated by the conjugative transfer of the 41.1 kb plasmid alone or with the co-transfer of a plasmid encoding resistance to cadmium. The size of the mupirocin-resistance encoding plasmid in the Nigerian strain (35 IBA) was approximately 35 kb.ConclusionThe emergence of mupirocin-resistant S. aureus isolates in Nigeria and South Africa should be of great concern to medical personnel in these countries. It is recommended that methicillin-susceptible S. aureus (MSSA) and MRSA should be routinely tested for mupirocin resistance even in facilities where the agent is not administered. Urgent measures, including judicious use of mupirocin, need to be taken to prevent clonal dissemination of the mupirocin/methicillin resistant S. aureus in KZN, South Africa and the transfer of the conjugative plasmid encoding high-level mupirocin resistance identified in this study.

Waterborne human pathogenic viruses of public health concern

In recent years, the impending impact of waterborne pathogens on human health has become a growing concern. Drinking water and recreational exposure to polluted water have shown to be linked to viral infections, since viruses are shed in extremely high numbers in the faeces and vomit of infected individuals and are routinely introduced into the water environment. All of the identified pathogenic viruses that pose a significant public health threat in the water environment are transmitted via the faecal–oral route. This group, are collectively known as enteric viruses, and their possible health effects include gastroenteritis, paralysis, meningitis, hepatitis, respiratory illness and diarrhoea. This review addresses both past and recent investigations into viral contamination of surface waters, with emphasis on six types of potential waterborne human pathogenic viruses. In addition, the viral associated illnesses are outlined with reference to their pathogenesis and routes of transmission.

Induced protein polymorphisms and nutritional quality of gamma irradiation mutants of sorghum

Physical and biochemical analysis of protein polymorphisms in seed storage proteins of a mutant population of sorghum revealed a mutant with redirected accumulation of kafirin proteins in the germ. The change in storage proteins was accompanied by an unusually high level accumulation of free lysine and other essential amino acids in the endosperm. This mutant further displayed a significant suppression in the synthesis and accumulation of the 27kDa γ-, 24kDa α-A1 and the 22kDa α-A2 kafirins in the endosperm. The suppression of kafirins was counteracted by an upsurge in the synthesis and accumulation of albumins, globulins and other proteins. The data collectively suggest that sorghum has huge genetic potential for nutritional biofortification and that induced mutations can be used as an effective tool in achieving premium nutrition in staple cereals.