Prospective surveillance of nosocomial device-associated bacteremia in three adult intensive units in Malaysia.

Nosocomial blood stream infection (or nosocomial bacteremia) is a common problem in hospitals worldwide, including Malaysia. A three-year prospective cohort study (October 2003-March 2007) of the incidences, risk factors, and patterns of the microorganisms causing bacteremia was conducted using a validated surveillance form in three intensive care units (ICUs) in Malaysia. Center for Disease Control criteria were used to diagnose bacteremia. Patients were monitored from admission until the end point of study, which was the first detection of bacteremia in the blood in each patient. The frequency of occurrence of bacteremia with clinical symptoms was 10.7% (n=23). Bacteremia was observed to occur within a mean length of stay of 10 days in ICU. The rate of device-related infection was 10.4% per device utilization days with a device utilization rate of 95.9%/1000 patient days. The total number of patient days was 2309 and the period of device utilization was 2211 days. The common bacteria detected were extended-spectrum beta-lactamases (ESBLs) Klebsiella pneumoniae (n=6); Pseudomonas aeruginosa (n=6); Acinetobacter species (n=5); Methicillin-resistant Staphylococcus aureus (MRSA)(n=3); and (non- ESBL) Klebsiella pneumoniae (n=2). Multivariable analysis using Cox Proportional Hazard Model showed that the predictors for developing bacteremia were cancer, MRSA carriage, duration of central venous catheter (CVC) infusion, frequency change of CVC, and the administration of hydrocortisone drugs. These results indicate that a combination of nursing and medical interventions as well as patients' severity of illness could lead to bacteremia in ICU. Strategic implementation of quality assurance measures in ICUs could help to control this problem.

Prospective surveillance of nosocomial device-associated bacteremia in three adult intensive units in Malaysia

Nosocomial blood stream infection (or nosocomial bacteremia) is a common problem in hospitals worldwide, including Malaysia. A three-year prospective cohort study (October 2003-March 2007) of the incidences, risk factors, and patterns of the microorganisms causing bacteremia was conducted using a validated surveillance form in three intensive care units (ICUs) in Malaysia. Center for Disease Control criteria were used to diagnose bacteremia. Patients were monitored from admission until the end point of study, which was the first detection of bacteremia in the blood in each patient. The frequency of occurrence of bacteremia with clinical symptoms was 10.7% (n = 23). Bacteremia was observed to occur within a mean length of stay of 10 days in ICU. The rate of device-related infection was 10.4% per device utilization days with a device utilization rate of 95.9%/1000 patient days. The total number of patient days was 2309 and the period of device utilization was 2211 days. The common bacteria detected were extended-spectrum beta-lactamases (ESBLs) Klebsiella pneumoniae (n = 6); Pseudomonas aeruginosa (n = 6); Acinetobacter species (n = 5); Methicillin-resistant Staphylococcus aureus (MRSA)(n = 3); and (non- ESBL) Klebsiella pneumoniae (n = 2). Multivariable analysis using Cox Proportional Hazard Model showed that the predictors for developing bacteremia were cancer, MRSA carriage, duration of central venous catheter (CVC) infusion, frequency change of CVC, and the administration of hydrocortisone drugs. These results indicate that a combination of nursing and medical interventions as well as patients' severity of illness could lead to bacteremia in ICU. Strategic implementation of quality assurance measures in ICUs could help to control this problem.

Regulation of clustered gene expression by cofactor of BRCA1 (COBRA1) in breast cancer cells.

Eucaryotic genes that are coordinately expressed tend to be clustered. Furthermore, gene clusters across chromosomal regions are often upregulated in various tumors. However, relatively little is known about how gene clusters are coordinately expressed in physiological or pathological conditions. Cofactor of BRCA1 (COBRA1), a subunit of the human negative elongation factor, has been shown to repress estrogen-stimulated transcription of trefoil factor 1 (TFF1 or pS2) by stalling RNA polymerase II. Here, we carried out a genome-wide study to identify additional physiological target genes of COBRA1 in breast cancer cells. The study identified a total of 134 genes that were either activated or repressed upon small hairpin RNA-mediated reduction of COBRA1. Interestingly, many COBRA1-regulated genes reside as clusters on the chromosomes and have been previously implicated in cancer development. Detailed examination of two such clusters on chromosome 21 (21q22) and chromosome X (Xp11) reveals that COBRA1 is physically associated with a subset of its regulated genes in each cluster. In addition, COBRA1 was shown to regulate both estrogen-dependent and -independent transcription of the gene cluster at 21q22, which encompasses the previously identified COBRA1-regulated TFF1 (pS2) locus. Thus, COBRA1 plays a critical role in the regulation of clustered gene expression at preferred chromosomal domains in breast cancer cells.

Contributions of UP elements and the transcription factor FIS to expression from the seven rrn P1 promoters in Escherichia coli.

The high activity of the rrnB P1 promoter in Escherichia coli results from a cis-acting DNA sequence, the UP element, and a trans-acting transcription factor, FIS. In this study, we examine the effects of FIS and the UP element at the other six rrn P1 promoters. We find that UP elements are present at all of the rrn P1 promoters, but they make different relative contributions to promoter activity. Similarly, FIS binds upstream of, and activates, all seven rrn P1 promoters but to different extents. The total number of FIS binding sites, as well as their positions relative to the transcription start site, differ at each rrn P1 promoter. Surprisingly, the FIS sites upstream of site I play a much larger role in transcription from most rrn P1 promoters compared to rrnB P1. Our studies indicate that the overall activities of the seven rrn P1 promoters are similar, and the same contributors are responsible for these high activities, but these inputs make different relative contributions and may act through slightly different mechanisms at each promoter. These studies have implications for the control of gene expression of unlinked multigene families.

Aromatic amino acids in region 2.3 of Escherichia coli sigma 70 participate collectively in the formation of an RNA polymerase-promoter open complex.

Formation of an initiation-competent RNA polymerase-promoter complex involves DNA melting over a region of about 12 base-pairs, which includes the start site of transcription, thus enabling the template strand to base-pair with the initiating nucleoside triphosphates. By studying the effects of alanine substitutions, we have investigated the role of the aromatic amino residues in the Escherichia coli sigma(70) conserved region 2.3 in promoter strand separation. The resulting mutants were assessed for their activity in vivo in the context of a sigma(70)/sigma(32) hybrid sigma factor that could be targeted to a specific hybrid promoter in the cell. All substitutions lead to an at least twofold reduction in expression of the hybrid promoter-driven reporter gene. The in vitro assay of single substitutions indicated cold sensitivity similar to that previously observed with analogous substitutions in Bacillus subtilis sigma(A). Kinetic assays showed that these substitutions slowed the rate of open complex formation at 37 degrees C as well. RNA polymerase reconstituted with a sigma(70) containing multiple alanine substitutions readily binds to promoter DNA, but then proceeds slowly beyond the first intermediate complex on the pathway to formation of the transcription-competent complex. These data demonstrate that together the aromatic residues in region 2.3 of E. coli sigma(70) ensure that DNA strand separation proceeds efficiently, even if no individual residue may be essential for accomplishment of the process.

Antibiotics in chronic suppurative otitis media: a bacteriologic study.

Conservative medical management of chronic suppurative otitis media (CSOM) is an important step in achieving a dry ear. Topical antibiotic ear drops and aural toilet form the mainstay of medical management of noncholesteatomatous CSOM. This study analyzes the causal organisms and their sensitivity to various antibiotics. Out of 382 swabs examined, the major organisms isolated were Pseudomonas aeruginosa (27.2%), followed by Staphylococcus aureus (23.6%). The sensitivity of P. aeruginosa was 100% to ceftazidime, 98.9% to ciprofloxacin, 96.3% to gentamicin, and 95.4% to polymyxin B, whereas the sensitivity of S. aureus was 98.6% to ciprofloxacin, 97.4% to cloxacillin sodium, 96.5% to cotrimoxazole, and 90.7% to gentamicin. Pseudomonas aeruginosa was almost completely resistant to ampicillin (97.6%) and chloramphenicol (96.6%), whereas S. aureus was almost completely resistant to ampicillin (73.8%) and polymyxin B (98.3%). Among the available topical antibiotic preparations for use in the ear, we found that ciprofloxacin and gentamicin are the best choices.

Upstream A-tracts increase bacterial promoter activity through interactions with the RNA polymerase alpha subunit.

Upstream A-tracts stimulate transcription from a variety of bacterial promoters, and this has been widely attributed to direct effects of the intrinsic curvature of A-tract-containing DNA. In this work we report experiments that suggest a different mechanism for the effects of upstream A-tracts on transcription. The similarity of A-tract-containing sequences to the adenine- and thymine-rich upstream recognition elements (UP elements) found in some bacterial promoters suggested that A-tracts might increase promoter activity by interacting with the alpha subunit of RNA polymerase (RNAP). We found that an A-tract-containing sequence placed upstream of the Escherichia coli lac or rrnB P1 promoters stimulated transcription both in vivo and in vitro, and that this stimulation required the C-terminal (DNA-binding) domain of the RNAP alpha subunit. The A-tract sequence was protected by wild-type RNAP but not by alpha-mutant RNAPs in footprints. The effect of the A-tracts on transcription was not as great as that of the most active UP elements, consistent with the degree of similarity of the A-tract sequence to the UP element consensus. A-tracts functioned best when positioned close to the -35 hexamer rather than one helical turn farther upstream, similar to the positioning optimal for UP element function. We conclude that A-tracts function as UP elements, stimulating transcription by providing binding site(s) for the RNAP alphaCTD, and we suggest that these interactions could contribute to the previously described wrapping of promoter DNA around RNAP.

Escherichia coli promoters with UP elements of different strengths: modular structure of bacterial promoters.

The alpha subunit of Escherichia coli RNA polymerase (RNAP) participates in promoter recognition through specific interactions with UP element DNA, a region upstream of the recognition hexamers for the sigma subunit (the -10 and -35 hexamers). UP elements have been described in only a small number of promoters, including the rRNA promoter rrnB P1, where the sequence has a very large (30- to 70-fold) effect on promoter activity. Here, we analyzed the effects of upstream sequences from several additional E. coli promoters (rrnD P1, rrnB P2, lambda pR, lac, merT, and RNA II). The relative effects of different upstream sequences were compared in the context of their own core promoters or as hybrids to the lac core promoter. Different upstream sequences had different effects, increasing transcription from 1.5- to approximately 90-fold, and several had the properties of UP elements: they increased transcription in vitro in the absence of accessory protein factors, and transcription stimulation required the C-terminal domain of the RNAP alpha subunit. The effects of the upstream sequences correlated generally with their degree of similarity to an UP element consensus sequence derived previously. Protection of upstream sequences by RNAP in footprinting experiments occurred in all cases and was thus not a reliable indicator of UP element strength. These data support a modular view of bacterial promoters in which activity reflects the composite effects of RNAP interactions with appropriately spaced recognition elements (-10, -35, and UP elements), each of which contributes to activity depending on its similarity to the consensus.

Idiopathic CD4+ T-lymphocytopenia in a child with disseminated cryptococcosis.

We describe a Malay girl with disseminated cryptococcosis affecting the lungs, liver, lymph nodes and bones. The diagnosis was made by culture of the bone marrow. Tests of immune function showed that she was HIV-negative but the CD4 percentage was persistently low. Idiopathic CD4+ T-lymphocytopenia was diagnosed. The child died despite two courses of anti-fungal therapy.

The prevalence of hepatitis B surface antigen and anti-HCV antibody in paediatric oncology patients in Hospital Universiti Sains Malaysia.

This study examined the prevalence of hepatitis B and C markers in 55 paediatric oncology patients who had completed treatment at the Hospital Universiti Sains Malaysia in Kota Baru. All these children had received blood products and had been treated between 1985-1996. Forty seven per cent of patients were positive for hepatitis B or C. Twenty nine per cent were positive for hepatitis C and twenty two per cent were HBsAg positive. Two children were positive for both and none were HIV positive. Four children had an elevated ALT level and one child had jaundice and hepatomegaly. Some children were marker-positive despite immunization and screening of blood.

Acute pancreatitis in north-eastern peninsular Malaysia: an unusual demographic and aetiological pattern.

A survey of 142 cases of acute pancreatitis was undertaken in two major hospitals serving the state of Kelantan in Malaysia. Females outnumbered males by a ratio of more than 3:1. The incidence among females peaked in the third decade of life. Twenty-one percent (23/109) of the females were pregnant. Ultrasonography revealed gallstones in only 9.4% (13/138). However, abnormalities of serum transaminases were found in 35% (35/100), suggesting that occult gallstones or microlithiasis may be the cause in a significant proportion of patients. Alcohol was virtually absent as an aetiological factor. There was a higher frequency of Ascaris infection in this group than a control hospital population (11/35 vs 33/242; p < 0.02) suggesting that ascariasis may be an important cause of acute pancreatitis in Kelantan. Only 8.4% fell into the category of severe pancreatitis. The overall mortality rate was 2.1%.

Mutations in sigma factor that affect the temperature dependence of transcription from a promoter, but not from a mismatch bubble in double-stranded DNA.

Specificity of promoter utilization in bacterial RNA polymerases is imparted by a class of proteins referred to as sigma factors. Conserved region 2.3 of these proteins is thought to play a role in the strand separation process that occurs during the formation of an initiation-competent RNA polymerase-promoter complex. We have used a heterologous system consisting of Escherichia coli core RNA polymerase and Bacillus subtilis sigma A to probe the effects of amino acid substitutions in region 2.3. In agreement with previous work [Juang & Helmann (1994) J. Mol. Biol. 235, 1470-1488] we observe that several amino acid substitutions exacerbate the deleterious effect of low temperature on promoter-dependent initiation. On the other hand, no such enhanced cold sensitivity is found with double-stranded templates that contain short "bubbles" of single-stranded DNA, indicating that the DNA-melting defect imposed by these mutant sigma factors can be suppressed by the use of such bubble templates. These results support the involvement of region 2.3 in the strand separation process that accompanies open complex formation at promoters.

Detection of enterovirus RNA in clinical samples by nested polymerase chain reaction for rapid diagnosis of enterovirus infection.

Employing a nested polymerase chain reaction with primers from the 5' non-translated region of the enterovirus genome, we detected enteroviral RNA from patients with a variety of enterovirus-associated clinical syndromes. This technique was found to be sensitive (detecting enteroviral RNA extracted from 0.1 50% tissue culture infectious dose) and specific; no specific PCR product was detected from RNA extracts of a variety of non-enterovirus isolates. Although the technique is of comparable sensitivity to single round polymerase chain reaction followed by Southern blot hybridization, it was more rapid, since it enabled a diagnosis to be made within 1 day. Thus, using nested polymerase chain reaction we were able to detect enteroviral RNA in 31 of 46 clinical specimens from 17 of 23 patients with suspected enterovirus infections. The samples included cerebrospinal fluid, throat swabs, stool, vesicle fluid, peripheral blood lymphocytes, whole blood and pericardial effusion. In contrast virus was isolated in only 17 of 42 clinical specimens from 12 of 22 these patients. In preliminary studies, restriction endonuclease analysis of polymerase chain reaction products enabled us to distinguish between non-polio enteroviruses and poliovirus types 1, 2, and 3. This additional technique may be useful in distinguishing between such infections in polio-endemic countries where rapid public health measures may be required.

Reliability of two commercial serological kits for serodiagnosing Helicobacter pylori infection.

Two commercial serological kits, Pylori-set (Orion Diagnostica, Finland) and Helico-G (Cambridge Biomedical Ltd, UK), and an in-house ELISA were evaluated with sera from 24 Helicobacter pylori-positive and 146 H. pylori-negative dyspeptic patients. Sensitivity, specificity, positive and negative predictive values of Pylori-set were lower than that of Helico-G and in-house ELISA. Helico-G was more sensitive (91.7%) than in-house ELISA (83.3%) and both had comparable negative predictive values of 98.3% and 97.3%, respectively. However, specificity (97.9%) and positive predictive value (86.9%) of an in-house ELISA were much higher than specificity (80.1%) and positive predictive value (43.1%) of Helico-G. Kappa index of agreement between the three serological tests (Pylori-set, Helico-G or in-house ELISA) and the presence of H. pylori in antral biopsies was very low (k = 0.13; z = 1.9; p > 0.05), moderate (k = 0.49; z = 7.1; p < 0.0001), or substantial (k = 0.82; z = 10.8; p < 0.0001), respectively. Overall, statistical evaluations demonstrated that both commercial kits were not as reliable as the in-house ELISA for serodiagnosing H. pylori infection.

A mismatch bubble in double-stranded DNA suffices to direct precise transcription initiation by Escherichia coli RNA polymerase.

Formation of a transcription-competent "open" complex between Escherichia coli RNA polymerase and a promoter, where base pairing is disrupted over a region of 12 base pairs including the start site of transcription, is a complex process involving at least three steps: recognition of specific DNA sequences, a conformational change in RNA polymerase, and DNA melting. By using synthetic constructs devoid of promoter-specific sequences, we show here that a mismatch bubble of 12 base pairs suffices to direct transcription initiation in divergent directions from its edges, reflecting the absence of polarity determinants for RNA polymerase binding. Bubble transcription is obtained with both core polymerase and holoenzyme, but efficient formation of heparin-resistant initiation complexes requires the sigma (specificity) factor. Based on these results it is likely that the sigma factor blocks access of the heparin to a site on the holoenzyme.

Helicobacter pylori infection in north-eastern peninsular Malaysia. Evidence for an unusually low prevalence.

The prevalence of Helicobacter pylori infection was determined in peptic ulcer patients, in non-ulcer dyspepsia (NUD) patients, and in the general adult population. The H. pylori infection rate ascertained by microbiologic examination of multiple gastric antral biopsy specimens was 50% (17 of 34) in duodenal ulcer (DU), 5% (1 of 22) in gastric ulcer, and 9% (15 of 159) in NUD patients. A seroepidemiologic survey showed a prevalence of only 4.2% among 496 blood donors and 4.8% among 921 subjects who attended health screening clinics. H. pylori infection is relatively uncommon and does not appear to be the predominant factor in the pathogenesis of peptic ulcer disease in the area. The incidence of peptic ulcer perforations in the area in 1991-92 was 1.5 per 100,000 person-years, reflecting a relatively low frequency of peptic ulcers, which might be due to the low prevalence of H. pylori infection in the population.

Intracranial suppuration caused by Micrococcus luteus.

Micrococcus spp. are commensal organisms colonizing the body surfaces of humans. In a few instances these organisms have been reported to colonize ventricular shunts. We report a patient, with no overt evidence of immunosuppression, in whom Micrococcus luteus was responsible for intracranial suppuration at multiple sites.