Enterotoxigenic and antibiotic resistance determination of Staphylococcus aureus strains isolated from food handlers in Gaborone, Botswana.

The prevalence, antibiotic resistance, and enterotoxigenic potential of Staphylococcus aureus strains from different anatomical sites on food handlers in Gaborone, Botswana, were determined. Of a total of 200 food handlers tested, 115 (57.5%) were positive for S. aureus. Of the 204 S. aureus isolates, 63 (30.9%), 91 (44.6%), and 50 (24.5%) were isolated from the hand, nasal cavity, and face, respectively, and 43 (21%) of the isolates were enterotoxigenic. The most prevalent enterotoxin was type A, which accounted for 34.9% of all the enterotoxigenic strains, and enterotoxin D was produced by the fewest number of strains (9.3%). Resistance to methicillin was encountered in 33 (22.4%) of the penicillin G-resistant isolates, and 9 (27.3%) of these methicillin-resistant isolates also were resistant to vancomycin. Nineteen antibiotic resistance profiles were determined, and the nasal cavity had the highest diversity of resistance profiles. The nasal cavity also had the highest number of resistant strains, 77 (53%), whereas the hand and face had 49 (32%) and 24 (16.0%) resistant strains, respectively. To reduce the Staphylococcus carriage rate among food handlers, training coupled with a commitment to high standards of personal and environmental hygiene is recommended.

Colloidal silver as an antimicrobial agent: fact or fiction?

OBJECTIVE: Colloidal silver preparations are marketed on the internet as omnipotent antimicrobial agents, but scientific support for these claims is lacking. This study reports the results of in vitro tests of colloidal silver's antimicrobial activity against several pathogenic or non-pathogenic microorganisms. METHOD: Three samples of colloidal silver were tested: one available commercially on the internet (silver concentration of 22 ppm) and two samples (concentrations of 403 and 413 ppm) which were prepared in our laboratory using standard chemical methods. RESULTS: In an agar-well diffusion assay none of the three colloidal silver solutions had any effect on the growth of the test organisms. All tested bacterial strains were sensitive to ciprofloxacin. Colloidal silver 22 ppm showed no bactericidal activity in phenol coefficient tests. CONCLUSION: As the tested colloidal silver solutions did not show any antimicrobial effect in vitro on the microorganisms, claims of colloidal silver's antimicrobial potency are misleading and there is no place for it as an antiseptic.

Detection and characterization of human rotavirus among children with diarrhoea in Botswana.

This study reports the detection, for the first time, of human rotavirus in stools of children and the molecular characterization of isolated circulating strains in Botswana. We collected 249 stool samples between 1999 and 2001 from children with diarrhoea in three health districts of Botswana and examined them for the presence of rotavirus antigens and particles. Group A rotavirus antigen was detected in 43 of 249 (17%) of the samples tested by enzyme-linked immunosorbent assay. Of the 43 children shedding rotaviruses, 37 (86%) were infants < or =2 years of age. The presence of rotavirus particles was also confirmed by direct electron microscopy. The characteristic 11 segments of the double-stranded RNA mobility pattern of rotavirus were demonstrated by polyacrylamide electrophoresis in 20 of 43 (47%) of the rotavirus-positive samples. The predominant electrophoretic pattern detected was the long (L) electrophoretype 14 of 20 (70%) followed by the short (S) electrophoretype five of 20 (25%). One strain had a mixed (L/S) pattern. Of the 26 samples subjected to subgrouping by enzyme immuno assay, eight were typed as subgroup-II specific and seven were subgroup I. The predominant VP7 genotypes detected were G1 (59%). Two mixed strains of G1 + G3 (5%) and G1 + G2 (5%) were also detected. VP4 genotypes in circulation were: P[4] (5%), P[6] (33%) and P[8] (33%). Mixed P-types P[4 + 6] (5%) and P[6 + 8] (18%) were also detected. Rotavirus strains G1 P[8] and GI P[6 + 8] were the most common cause of diarrhoea in our study area.

Enterococcus faecalis and Enterococcus faecium isolates from milk, beef, and chicken and their antibiotic resistance.

The occurrence and antibiotic resistance of enterococci, especially Enterococcus faecalis and Enterococcus faecium, in milk, beef, and chicken in Gaborone, Botswana, were studied. Enterococci were isolated from these sources with the use of bile esculin agar and identified with API 20 Strep kits. Antibiotic resistance was determined by the disk diffusion method. The antibiotics tested were vancomycin, teicoplanin, ampicillin, tetracycline, and cephalothin. Among the 1,467 enterococci isolated from the samples, E. faecalis (46.1%) and E. faecium (29.0%) were found to be the predominant species. Other enterococcal species made up 25% of the isolates. More than 96 and 97% of the E. faecalis and E. faecium isolates, respectively, were found to be resistant to ampicillin. Almost 34, 27.3, and 22.4% of the E. faecalis isolates from milk, beef, and chicken, respectively, were also resistant to cephalothin. The percentages of E. faecium isolates that were found to be resistant to cephalothin were 32.8, 16.9, and 17.3% for milk, beef, and chicken, respectively. Resistance to vancomycin was widespread. It was found that 18.8, 7.8. and 13.1% of the E. faecalis isolates from milk, beef, and chicken samples, respectively, were resistant to vancomycin. In contrast, 32.8, 24.7, and 30.7% of the E. faecium isolates from milk, beef, and chicken samples, respectively, were resistant to vancomycin. Isolates that were resistant to multiple drugs were found in relatively large numbers.

Optimising growth conditions for the pectinolytic activity of Kluyveromyces wickerhamii by using response surface methodology.

This present study was undertaken to find optimum conditions of pH, temperature and, period of incubation for the pectinolytic activity of Kluyveromyces wickerhamii isolated from rotting fruits and to assess the effect of these factors by use of response surface methodology (RSM). A central composite rotatable design was used as an experimental design for the analysis of the allocation of treatment combinations. A second order polynomial regression model was fitted and was found adequate, with an R(2) of 0.94469 (P<0.001). The effects of temperature and pH were the most significant factors in influencing enzyme production. Estimated optimum conditions were as follows: pH 5.0, temperature, 32 degrees C and an incubation period of 91 h. Pectinesterase (PE), pectin lyase (PL), and cellulase activities were not detected. Pectinase production was partially constitutive. Pectin was degraded by the isolated strain of K. wickerhamii in the current study, and the pectinolytic activity is referred to as polygalacturonase (PG) activity. Crude enzyme extract was thermostable at various temperatures and, stimulated by the presence of Ca(2+) ions but inhibited by other ions like Mg(2+), Zn(2+), Co(2+), Mn(2+) and Na(+).

Prevalence of rotavirus, adenovirus and astrovirus infection in young children with gastroenteritis in Gaborone, Botswana.

OBJECTIVE: To determine the prevalence of three enteric viruses, namely rotavirus, adenovirus and astrovirus, as agents of diarrhoea in and around Gaborone, Botswana. DESIGN: The sample were categorised into four groups according to the age of the patient: 0-3 months, 4-6 months, 7-12 months and 25-60 months. Total monthly samples across age groups formed basis for calcultating seasonal prevalence of rotavirus infection. SETTING: Stool samples were collected from three medical laboratories in Gaborone and one in the town of Mochudi. These were collected from children under the age of five years with gastroenteritis. SUBJECTS: Stool samples were collected between March 2001 and February 2002 from 346 children less than five years of age suffering from gastroenteritis. These samples had been sent to medical laboratories for microbiological examination. METHODS: The samples were screened for rotavirus (RV), adenovirus (Ad) and astrovirus (AsV) antigens using commercially available ELISA kits. The Ad positive samples were further analysed by commercially available group specific Ad type 40/41 Enzyme Immuno Assays (EIA). RESULTS: Shedding of RV was detected in 9.2%, Ad in 7.8% and AsV in 2.7% of the samples analysed. The enteric Ad (types 40 and 41) were detected in 2% of the samples and the remaining 5.8% of Ad positive samples were non-enteric Ad. An increase of RV was noted in the autumn-winter season but no seasonal pattern was observed in Ad shedding. Seasonal prevalence of AsV could not be determined. The average age of children infected with these agents was less than one year. CONCLUSION: The incidence of rotavirus infection amongst children in Botswana appears to be relatively low. The prevalence rate of adenovirus and astrovirus is similar to other studies in parts of Southern Africa. However, continued enteric virus surveillance and epidemiology amongst this group is required.

Antimicrobial susceptibility of staphylococci species from cow foremilk originating from dairy farms around Gaborone, Botswana.

OBJECTIVE: To determine the prevalence of antibiotic susceptibility of Staphylococcus species isolated from foremilk samples. SETTING: Milk was collected from five farms within a 70 km radius of Gaborone, Botswana. SUBJECTS: Two hundred and twenty five staphylococci isolates from foremilk samples. MAIN OUTCOME MEASURES: Antibiotic susceptibility tests to penicillin G, ampicillin, tetracycline, erythromycin, cephalothin, chloramphenicol, methicillin, gentamicin and vancomycin. RESULTS: The susceptibility patterns of the staphylococcal strains to the antibiotics were as follows: penicillin G (47.1%), ampicillin (58.7%), tetracycline (62.7%), erythromycin (72%), cephalothin (72.9%), chloramphenicol (79.1%), methicillin (86.2%), gentamicin (88.9%) and vancomycin (100%). Lower susceptibility to chloramphenicol, methicillin and gentamicin was displayed by Staphylococcus epidermidis, S. haemolyticus and S. saprophyticus. Only 19 (8.5%) of the isolates were susceptible to all the antibiotics tested. The most common multiple resistance patterns encountered were penicillin-ampicillin (9.3%), penicillin-erythromycin-ampicillin (6.1%) and erythromycin-tetracycline-ampicillin (3.6%). CONCLUSION: Most of the Staphylococcus isolates were resistant to one or more of the antimicrobial agents, with none being resistant to vancomycin. Inappropriate use of antibiotics is suspected to be a major contributory factor in the relatively high level of resistance to antimicrobial agents observed in this study. Therefore, milk can act as a very good source of antibiotic resistant Staphylococcus species posing a threat to consumers.

Shigella and Salmonella strains isolated from children under 5 years in Gaborone, Botswana, and their antibiotic susceptibility patterns.

We isolated Shigella from 43/221 (21%) and Salmonella 8/221 (3%) rectal swabs from children under 5 years with diarrhoea, and found Shigella in two of 100 specimens from children without diarrhoea. Sh. boydii (13%) was the most prevalent Shigella species followed by Sh. flexneri (6%) and Sh. sonnei (2%). The prevalence of various types of Sh. boydii was type 7, 5%; type 9, 3%; type 12 and 16, 2%; and type 18, 1%. Other Shigella serotypes encountered were Sh. flexneri type 6 (4%), type 4 (2%), with Sh. sonnei phase II isolated from 2% of the specimens. The Salmonella species were S. typhimurium and S. paratyphi. The high rate of isolation of Shigella species from children with diarrhoea is indicative of a definite role of this enteropathogen in causing endemic diarrhoea in Gaborone, Botswana. Antibiograms of the predominant isolates showed that most Shigella species were resistant to ampicillin but susceptible to chloramphenicol, and with the exception of Sh. flexneri type 6, also susceptible to gentamicin. The Salmonella species were susceptible to chloramphenicol, collistin-sulphate, gentamicin, cotrimoxazole, and ampicillin.

Occurrence of aflatoxins, fumonisin B1, and zearalenone in foods and feeds in Botswana.

Sorghum and maize form the main dietary staple foods in Botswana. Other products such as peanuts, peanut butter, phane (an edible larval stage of an emperor moth Imbrasia belina Westwood), and pulses (cowpeas and beans) are also widely used as food and for the manufacture of feeds. These important food and feed commodities were analyzed for the presence of aflatoxins, fumonisin B1, and zearalenone. Aflatoxins were detected in 40% of the samples analyzed. The concentration of total aflatoxins ranged from 0.1 to 64 microg/kg. The mean concentration ranged from 0.3 microg/kg in sorghum to 23 microg/kg in peanut butter. Peanut butter samples were the most contaminated (71%). No aflatoxins were detected in maize. Fumonisin B1 was detected in 36% of the samples. Maize samples were the most contaminated (85% of the samples) with the concentration ranging from 20 to 1,270 microg/kg. No fumonisin B1 was detected in peanuts, phane, and beans. Zearalenone was only found in 2.6% of the samples analyzed at 40 microg/kg. Aflatoxins were the most common toxins detected in foods and feeds in Botswana. However, fumonisin B1 was more prevalent in maize than aflatoxins or zearalenone.

Invasion of tissue culture cells by diarrhoeagenic strains of Escherichia coli which lack the enteroinvasive inv gene.

Invasive Escherichia coli strains of certain serotypes invade by the same mechanism as the Shigella sp. It has been proposed that invasion of epithelial cells by EPEC strains may also occur; this is a previously overlooked property. In the present study E. coli strains isolated from patients with diarrhoea or ulcerative colitis, lacking the inv plasmid mediating classical invasion, but hybridizing with probes for different adhesins, were analyzed for their ability to invade HeLa and Caco-2 cells. The majority of strains invaded Caco-2 cells to a higher extent than HeLa cells. Adhesion to Caco-2 cells was a prerequisite for subsequent invasion of the cells but EAF, eae, EAgg and other known virulence factors were not sufficient to mediate invasion. In 8/9 E. coli strains invasion was enhanced after growth under iron restriction. Growth during anaerobic conditions did not influence subsequent invasion by E. coli strains whereas 6/9 strains had their invasive ability significantly decreased after growth in the presence of 1% glucose. The invasive process was inhibited by mannose but not by lactose, fucose or galactose. Our data indicate that strains of E. coli may invade Caco-2 cells by novel mechanisms which require adhesion to the cells but which differ from those of Salmonella sp., Yersinia sp., Shigella sp. and classical enteroinvasive E. coli.

Inhibition of spoilage and food-borne pathogens by lactic acid bacteria isolated from fermenting tef (Eragrostis tef) dough.

A study was carried out at the Department of Biology, Addis Abeba University, in 1991 to determine the inhibitory potential of fermenting tef and the lactic acid bacteria isolated from fermenting tef dough on Salmonella spp., Pseudomonas aeruginosa, Klebsiella spp., Bacillus cereus and Staphylococcus aureus. The test bacteria grew in the fermenting tef uptill 30 hr or till the pH dropped to 4.7. Thereafter, growth was inhibited and decreases in population were apparent. The results showed that the spent media from all of the four lactic acid bacterial isolates, namely, Lactobacillus spp., Pediococcus spp., Leuconostoc spp. and Streptococcus spp. inhibited the test bacteria. Acidity on its own was not responsible for the inhibition of the test bacteria. The spent medium from Streptococcus spp. showed the best inhibitory activity amongst the lactic acid bacteria.

Survival and growth of selected pathogens in fermented kocho (Ensete ventricosum).

A study was conducted to investigate the antagonistic potential of fermented kocho, aqueous extract of fermented kocho and spent media from lactic acid bacteria (LAB) isolated from it on Salmonella sp, Pseudomonas aeruginosa, Klebsiella sp, Bacillus cereus and Staphylococcus aureus. Fermented kocho (pH 4.3) inhibited growth of the test bacteria soon after their introduction into the food. The spent media from all of the four LAB, isolated from fermented kocho, namely Pediococcus sp, Leuconostoc sp, Lactobacillus sp and Streptococcus sp prevented the survival and growth of the test bacteria. The spent medium from Streptococcus sp showed the best antagonistic effect amongst all the LAB isolates. In all cases the inhibitory effects were independent of pH.

Cellulase production and activity in a species ofCladosporium.

ACladosporium species produced large amounts of cellulase enzyme components when grown in shake-culture with medium containing carboxymethylcellulose. There was significantly less activity when Avicel, filter paper or cotton were used as substrates. KNO3 was better than NH4Cl or urea for the production of cellulase. Tween 80 at 0.1% (w/v) increased the production of cellulase by 1.5 to 4.5-fold. All the cellulase components were optimally active in the assay at pH 5.0 and 60°C.

A survey of the microflora of raw and pasteurized milk and the sources of contamination in a milk processing plant in Addis Ababa, Ethiopia.

The microorganisms present in raw and pasteurized milk and the sources of contamination in the milk after it had arrived at the processing plant in Addis Ababa were studied. The lowest count registered for raw milk samples was 4 X 10(7) cfu/ml while the highest was 1 X 10(9) cfu/ml. Pasteurized milk had mesophilic aerobic counts of 7 X 10(5) cfu/ml as it left the pasteurizing unit, but the population increased 2- to 4-fold as a result of subsequent contamination. Of the total counts in raw milk, psychrophilic, thermoduric and thermophilic organisms made up 98.1, 1.4 and 0.5% respectively. In pasteurized milk, the amounts were 53.0, 39.5 and 7.5% respectively. Samples of milk pasteurized in the laboratory contained only 74.5% thermoduric and 25.5% thermophilic organisms. The isolates mostly belonged to the genera Bacillus, Streptococcus, Lactobacillus, Arthrobacter, Alcaligenes, Aeromonas and Pseudomonas. Cocci were more predominant than rod-shaped bacteria. Of the rod-shaped bacteria, 73% were Gram-negative. The utensils holding the raw and pasteurized milk and the plastic sheets used for bagging the pasteurized milk contributed unusually high numbers of bacteria which were either thermoduric or thermophilic. More isolates were obtained from the pasteurized than the raw milk. The keeping quality of the pasteurized milk was found to be much lower than that of the laboratory-pasteurized milk.

Fluoride-induced filaments of Erwinia carotovora.

Sodium fluoride induces filamentous growth of Erwinia carotovora when it is grown in liquid media containing aspartic acid only as the sole source of nitrogen. It is proposed that a stable complex of F(-)-Mg2+-enzyme and PO2(4) resulting in Mg2+ deficiency and consequent inability of E. carotovora cells to oxidize aspartic acid normally, is responsible for the formation of filaments in the presence of fluoride ions.