CHARACTERISATION OF BACTERIAL ISOLATES FROM PATIENTS WOUNDS AND ENVIRONMENTAL FACTORS PREDICTIVE OF POST-SURGICAL INFECTIONS AT THE ORTHOPAEDIC WARD IN ILE-IFE, NIGERIA.

OBJECTIVE: To determine the pattern of orthopaedic wound infection and the influence of environmental factors on the distribution of the etiologic bacterial agents. DESIGN: A prospective observational study. SETTING: Department of Orthopaedic Surgery and Traumatology, Obafemi Awolowo University Teaching Hospitals Complex, Ile-Ife, Nigeria. SUBJECTS: Sixty patients with orthopaedic wound infections and orthopaedic ward environment. RESULTS: Thirty- nine males (mean age 33.31+2SD) and 21 females (mean age 27.47 + 2SD) with orthopaedic wounds. Three hundred and ten bacteria (190 from patients and 120 from ward environment) were isolated. The pattern of bacterial isolates from patients' wounds was different from that of the airborne bacterial isolates irrespective of the length of stay on the ward. There was a significant difference in the distribution and resistance pattern of bacterial isolates from the patient's wounds and ward environment. CONCLUSION: There is a high incidence of antibiotic resistance of bacterial isolates from samples cultured from patients compared with isolates from ward environment at this centre. The extensive use of pre-operative prophylactic and post-surgical antibiotics in various combinations at this centre needs to be re-examined to reduce the preponderance of antibiotic resistance.

Plasmodium falciparum: Activity of artemisinin against Plasmodium falciparum cultured in sickle trait hemoglobin AS and normal hemoglobin AA red blood cells.

The presence of sickle hemoglobin causes accumulation of hemoglobin degradative products that favor oxidative reaction in erythrocytes. Artemisinin derivatives exert antiparasite effects through oxidative reactions within infected erythrocytes. Using [(3)H]-hypoxanthine incorporation, we therefore did an in vitro comparison of IC(50) values for artemisinin in Plasmodium falciparum-infected erythrocytes from sickle cell trait (AS) and normal (AA) individuals. IC(50) values for chloroquine served as control. Without drugs, parasite growth was similar in AA and AS erythrocytes. Gender, age and blood group of donors had no significant effects on parasite growth. IC(50) value for artemisinin was 27+/-14nM in AS (N=22) compared to 24+/-9nM (N=27) in AA erythrocytes (P=0.4). IC(50) values for chloroquine were also similar in AA (22+/-8nM) and AS (20+/-11nM) erythrocytes. These results show no evidence of elevated artemisinin activity on P. falciparum in AS erythrocytes in vitro.

Microbial isolates in early swabs of open musculoskeletal injuries.

BACKGROUND: Infections of open musculoskeletal injuries, especially open fractures, continue to pose a challenge to the Orthopaedic and Trauma surgeons. There is a dearth of information on the early bacterial contaminants of these open wounds and their antibiotic sensitivity patterns in our environment. OBJECTIVE: To determine the bacteriology of open wounds of the musculoskeletal system at initial presentation in hospital and their antibiotic susceptibility profile in Ile-Ife, Nigeria. MATERIALS AND METHODS: A prospective study of 86 consecutive patients over an eight-month period (March to October 2002). Eighty-six patients with 96 open wounds of the musculoskeletal system admitted to the Accident and Emergency unit of the Obafemi Awolowo University Teaching Hospitals Complex, Ile-Ife. Bacteria were isolated from swabs taken from the depth of the wounds and their antibiotic susceptibility determined. RESULTS: A total of 126 bacterial isolates were recovered from 96 wounds in 86 patients (61 males and 25 females) with mean age of 31.4 years +/- SD. About 73.3% of the wounds were from road traffic accidents and 72.3% of the wounds were severe open fractures (Grades IIIA to C). Staphylococcus aureus constituted 22.2% while coagulase negative Staphylococci (CONS) accounted for 21.4% with Staphylococcus. epidermidis leading the group with 13.5%. Gram negative rods constituted 40.5% of the isolates with Pseudomonas aeruginosa accounting for 11.1%. Antibiotic sensitivity profile revealed that many of the isolates were multiply resistant to the antimicrobials employed but were sensitive to Ofloxacin. CONCLUSION: This study has shown that open wounds of the musculoskeletal system are usually contaminated at presentation with pathogenic organisms. The antibiotic sensitivity pattern of the isolates shows that the quinolones, represented by Ofloxacin, is the most effective antibiotic.

An IRP-like protein from Plasmodium falciparum binds to a mammalian iron-responsive element.

This study cloned and sequenced the complementary DNA (cDNA) encoding of a putative malarial iron responsive element-binding protein (PfIRPa) and confirmed its identity to the previously identified iron-regulatory protein (IRP)-like cDNA from Plasmodium falciparum. Sequence alignment showed that the plasmodial sequence has 47% identity with human IRP1. Hemoglobin-free lysates obtained from erythrocyte-stage P falciparum contain a protein that binds a consensus mammalian iron-responsive element (IRE), indicating that a protein(s) with iron-regulatory activity was present in the lysates. IRE-binding activity was found to be iron regulated in the electrophoretic mobility shift assays. Western blot analysis showed a 2-fold increase in the level of PfIRPa in the desferrioxamine-treated cultures versus control or iron-supplemented cells. Malarial IRP was detected by anti-PfIRPa antibody in the IRE-protein complex from P falciparum lysates. Immunofluorescence studies confirmed the presence of PfIRPa in the infected red blood cells. These findings demonstrate that erythrocyte P falciparum contains an iron-regulated IRP that binds a mammalian consensus IRE sequence, raising the possibility that the malaria parasite expresses transcripts that contain IREs and are iron-dependently regulated.

Detection of cationic surfactants in oral rinses and a disinfectant formulation using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Three commercial oral rinses and one commercial disinfectant formulation were analyzed for the presence of cationic surfactants using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) in the positive ion mode. The product labels on these formulations indicate the presence of cetylpyridinium chloride, tetraalkylammonium or trialkylbenzylammonium chlorides. The resulting MALDI-TOF mass spectra only showed cetylpyridinium, tetraalkylammonium, and trialkylbenzylammonium ions, apparently due to the dissociation of the salts in the ion source. We confirmed the presence of cetylpyridinium salt in the three oral formulations, whereas the disinfectant formulation consisted of a complex mixture of the salts of dioctyldimethylammonium, didecyldimethylammonium, benzylmyristyldimethylammonium, decyloctyldimethylammonium, benzyldecyldimethylammonium, and benzylcetyldimethylammonium. This work demonstrates again the potential for using meso-tetrakis(pentafluorophenyl)porphyrin as a matrix in the MALDI-TOFMS analysis of low molecular weight compounds. This study also demonstrates that the mode of ionization of quaternary ammonium compounds (cationic surfactant salts) under MALDI conditions is by dissociation, leading to the detection of only the positively charged moieties.

Assessment of antimalarial effect of ICL670A on in vitro cultures of Plasmodium falciparum.

We tested in vitro the antimalarial properties of ICL670A, a newly developed iron chelator for the long-term oral treatment of iron overload. Ring-stage synchronized cultures of Plasmodium falciparum cultured in human erythrocytes were exposed to different concentrations of ICL670A and the conventional iron chelator, desferrioxamine B (DFO), for 48 h. Malarial growth was measured by incorporation of [3H]-hypoxanthine. ICL670A at 30 micromol/l had marked antimalarial activity that was observable by 6 h after beginning the exposure of ring-stage parasites to the agent. Over 48 h of culture, malarial growth was significantly lower with ICL670A than with DFO at concentrations of both 30 micromol/l (P = 0.008) and 60 micromol/l (P = 0.001). At 48 h, growth relative to control was 53% with ICL670A and 83% with DFO at concentrations of 30 micromol/l, and 20% with ICL670A and 26% with DFO at concentrations of 60 micromol/l. Standard 50% inhibitory concentrations (IC50s) were similar for ICL670A and DFO. Precomplexation with iron completely abolished the inhibitory effect of ICL670A, indicating that this new agent, like DFO, probably inhibits parasite growth via deprivation of iron from critical targets within the parasite. Further studies to address the question of the antimalarial potential of ICL670A in combination with classic antimalarials would be of interest.

Inhibitory factors in breastmilk, maternal and infant sera against in vitro growth of Plasmodium falciparum malaria parasite.

Complications from falciparum malaria are responsible for over one million infant deaths annually. There is as yet no clinically protective vaccine that has been developed against human malaria parasites. While several studies have demonstrated the inhibitory properties of human sera against Plasmodium falciparum, there is no reported investigation that has examined the protective effects of human breastmilk against the malaria parasite. This study demonstrates the presence of significant antibody titers to ring, trophozoite, schizont and gametocyte stages of P. falciparum in 144 Nigerian maternal milk samples and also in paired maternal and infant sera. The study also demonstrates significant in vitro growth inhibition of P. falciparum by maternal and infant sera, but most notably by breastmilk samples and breastmilk constituents, such as lactoferrin and sIgA. The results therefore suggest a protective in vivo role for breastmilk in the possible modulation of malaria frequency, severity and complications.

Macrophage responses to Toxoplasma antigens in vitro: a possible role in inflammatory lesions in toxoplasmosis.

Toxoplasma antigen and Toxoplasma immune complex were shown to induce increased production and release of acid hydrolases from macrophage cell line P388D in a concentration-dependent manner. Antigen concentrations of 10-50 microg/ml gave a 2-4-fold increase in the activities of acid proteinase, acid phosphatase, and phospholipase A2 compared with control cells without antigen. Results were similar for immune complex concentrations of 30-80 microg/ml compared with controls. No significant lactate dehydrogenase activity was detected in the culture medium, indicating that enzyme release was selective and not due to cell death. These results suggest that increased release of acid hydrolases may play a role in the inflammatory lesions observed in Toxoplasma encephalitis.

A study of urinary tract infections at Ile-Ife, Nigeria.

Clinical laboratory records and case files of 1,038 cases of urinary tract infections (UTI) were examined for the spectrum of bacterial and candida isolates for age and sex distribution of the UTI cases and antibiotic susceptibility pattern of isolates. The results show that 867 (83.5%) of the cases were aged 21 years and above, of which 81.3% in that age group were females. While only 20.7% of the cases were males, 74.0% of those in that category were also 21 years and older. Eleven organisms were isolated, with E. coli, Klebsiella spp and Staphylococcus aureus respectively representing 36.1%, 22.8% and 15.4% of the total bacterial isolates. The prevalence of S. aureus isolates in cases of UTI in this centre suggests the increasing role of these microorganisms in the aetiology of urinary tract infections in Nigeria.

Combined microautoradiographic and histopathologic analysis of the fate of challenge Schistosoma mansoni schistosomula in mice immunized with irradiated cercariae.

Combined microautoradiographic and histopathologic methods were used to locate and examine schistosomula of Schistosoma mansoni in the lungs of irradiated cercaria-immunized mice 21 days after percutaneous challenge infection with 75Se-labeled cercariae. Of 75 schistosomula examined in serial sections, 53% were located in the pulmonary microvasculature, 23% in alveolar spaces, 3% with one end in a vessel and the other in an alveolar space, and the locations of 21% were not identified. Inflammatory reactions of variable intensity were observed around schistosomula in both vascular and alveolar sites, although the most intense category of reactions was associated almost entirely with alveolar larvae. All autoradiographic foci contained recognizable schistosomula. Although the concentration of reduced silver grains precluded cyto-structural analysis, observations on schistosomular contour and shape provided no evidence of larval damage. Our findings suggest that immune elimination of schistosomula in mice immunized with irradiated cercariae is partly or largely effected by a process of alveolar extrusion of viable parasites during their lung migration.

The impact of pulmonary infections on the survival of lung cancer patients.

To determine the impact of pulmonary infections on survival in patients with lung cancer, a retrospective review of the records of 121 such patients treated at Howard University Hospital in Washington, DC, was done. There were 77 men and 44 women; 118 were black. The mean age was 63.5 years. Forty-three patients had squamous cell carcinoma, 31 had adenocarcinoma, 18 had large cell carcinoma, 19 had small cell carcinoma, and ten were unclassified. The stages were as follows: two patients in Stage 0, 15 in Stage I, seven in Stage II, 45 in Stage III, and 44 in Stage IV. Eight patients could not be staged at diagnosis. Eighty-five patients (70%) had documented infections; 37 had single episodes; and 48 had more than one. The five most common organisms recovered were alpha/gamma streptococci, Staphylococcus aureus, Klebsiella pneumoniae, Enterobacter aerogenes, and Pseudomonas aeruginosa. The median survival of all infected patients was 4.2 months which was significantly shorter than that of uninfected patients who had a median survival of 12.9 months (P less than 0.05). When Stage III patients were analyzed separately, infected patients lived a median of 5.8 months and uninfected patients, 13.4 months (P less than 0.05). This study indicated that pulmonary infections frequently occur in patients with lung cancer and suggested that they may adversely affect survival.

Class-specific antibodies to Bordetella pertussis, Haemophilus influenzae type b, Streptococcus pneumoniae and Neisseria meningitidis in human breast-milk and maternal-infant sera.

Children under 2 years of age are most susceptible to acute respiratory infections caused by Bordetella pertussis, Haemophilus influenzae type b, Streptococcus pneumoniae and Neisseria meningitidis. We analysed milk samples and sera from mother-infant pairs for specific antibodies that may enhance protection against the bacterial pathogens. The results show that the breast-milk samples contained significant titres of specific IgG and IgA antibodies to the four organisms, although the mean IgG antibody levels were higher in maternal sera than in breast-milk. On the other hand, the mean IgA antibody levels to the four organisms were higher in breast-milk than in both maternal and infant sera. IgM antibodies to these organisms were relatively low or absent in many milk and serum samples. Nevertheless, the significant concentrations of specific IgG and IgA antibodies in milk samples may indicate a protective role for breast-milk against the four infections in early childhood.

Proteinuria and haematuria as predictors of schistosomiasis in children.

Examination of urine samples from 922 children from Epe and surrounding communities in south-western Nigeria indicated a 13% prevalence of Schistosoma haematobium infection. Children, 10-14 years of age, accounted for 65% of the disease prevalence. Approximately 79% of the study population was negative for proteinuria, while 52.6% of children with 30 mg% proteinuria were positive for S. haematobium infection. However, 96% and 100% of all children who, respectively, had 30 mg% and 100 mg% proteinuria, in addition to haematuria, were found to be positive for the schistosome infection. This finding indicates that the use of haematuria and proteinuria as a combined diagnostic index significantly increased the sensitivity and specificity of the individual tests. Bacteriuria was found in 8.5% of infected children, compared with 5.2% of the control group. Streptococcus faecalis and E. coli were the two bacteria isolated from the urine specimens.

Immunoprotective factors in breast milk and sera of mother-infant pairs.

We analysed breast milk and serum samples from 33 healthy Nigerian mother-infant pairs for concentrations of IgG, IgM, IgA, C3, C4 and lysozyme. We found that the mean IgG and IgM concentrations in maternal sera were three to four times higher than the levels in infant sera, and five to ten times higher than in breast milk. The IgA and lysozyme concentrations in breast milk were, however, slightly higher than the levels in infant sera, suggesting an active localized synthesis of these factors in the mammary gland. While 54.5% of the milk samples lacked measurable concentrations of IgM by radial immunodiffusion, IgA was consistently present in all the milk samples. The mean C3 concentration in maternal sera was 164 mg/100 ml, compared to 145 mg/100 ml and 11.5 mg/100 ml in infant sera and breast milk respectively. The C4 concentration was also considerably lower in breast milk than in maternal and infant sera.

Comparison of an enzyme-linked immunosorbent assay using crude S. haematobium soluble egg antigen with a radioimmunoassay using purified S. mansoni egg antigen for the serodiagnosis of schistosomiasis haematobium.

An enzyme-linked immunosorbent assay (ELISA) using crude Schistosoma haematobium soluble egg antigen (ShSEA) was compared with a radioimmunoassay (RIA) employing purified heterologous species S. mansoni egg antigen (MSA1) for the serodiagnosis of schistosomiasis haematobium in a group of 45 Nigerian school children living in an area endemic for S. haematobium. Both assay systems appeared applicable. The ELISA proved to be more sensitive detecting 100% of the 27 parasitologically positive individuals while the RIA defined 89% of this group. Neither test had false-positive results for the 10 non-endemic area parasitologically negative controls, although the ELISA demonstrated that 56% of the 18 endemic area parasitologically negative controls had anti-ShSEA antibodies. The RIA indicated that 44% of this group had anti-MSA1 antibodies. These latter findings were interpreted as related to the hyperendemicity of schistosomiasis haematobium for the study area.