The effects of frozen tissue storage conditions on the integrity of RNA and protein.

Unfixed tissue specimens most frequently are stored for long term research uses at either -80° C or in vapor phase liquid nitrogen (VPLN). There is little information concerning the effects such long term storage on tissue RNA or protein available for extraction. Aliquots of 49 specimens were stored for 5-12 years at -80° C or in VPLN. Twelve additional paired specimens were stored for 1 year under identical conditions. RNA was isolated from all tissues and assessed for RNA yield, total RNA integrity and mRNA integrity. Protein stability was analyzed by surface-enhanced or matrix-assisted laser desorption ionization time of flight mass spectrometry (SELDI-TOF-MS, MALDI-TOF-MS) and nano-liquid chromatography electrospray ionization tandem mass spectrometry (nLC-ESI-MS/MS). RNA yield and total RNA integrity showed significantly better results for -80° C storage compared to VPLN storage; the transcripts that were preferentially degraded during VPLN storage were these involved in antigen presentation and processing. No consistent differences were found in the SELDI-TOF-MS, MALDI-TOF-MS or nLC-ESI-MS/MS analyses of specimens stored for more than 8 years at -80° C compared to those stored in VPLN. Long term storage of human research tissues at -80° C provides at least the same quality of RNA and protein as storage in VPLN.

Impact of the HIV epidemic and Anti-Retroviral Treatment policy on lymphoma incidence and subtypes seen in the Western Cape of South Africa, 2002-2009: preliminary findings of the Tygerberg Lymphoma Study Group.

The Tygerberg Lymphoma Study Group was constituted in 2007 to quantify the impact of HIV on the pattern and burden of lymphoma cases in the Western Cape of South Africa which currently has an HIV prevalence of 15%. South Africa has had an Anti-Retroviral Treatment (ART) policy and a roll-out plan since 2004 attaining 31% effective coverage in 2009. This study is designed to qualify and establish the impact of HIV epidemic and the ARV roll-out treatment program on the incidence of HIV Related Lymphoma (HRL). Early data document that despite the ART roll out, cases of HRL are increasing in this geographical location, now accounting for 37% of all lymphomas seen in 2009 which is an increase from 5% in 2002. This is in contrast to trends seen in developed environments following the introduction of ART. Also noted are the emergence of subtypes not previously seen in this location such as Burkitt and plasmablastic lymphomas. Burkitt lymphoma is now the commonest HRL seen in this population followed by diffuse large B-cell lymphoma subtypes. The reasons for this observed increase in HRL are not ascribable to improved diagnostic capacity as the tertiary institute in which these diagnoses are made has had significant expertise in this regard for over a decade. We ascribe this paradoxical finding to an ART treatment environment that is ineffective for a diversity of reasons, paramount of which are poor coverage, late commencement of ART and incomplete viral suppression.

Purulence and gram-negative bacilli in tracheal aspirates of mechanically ventilated very low birth weight infants.

OBJECTIVE: Tracheal aspirates (TAs) from mechanically ventilated very low birth weight (VLBW) infants are frequently obtained during the evaluation of suspected sepsis, tracheitis, or ventilator-associated pneumonia (VAP). Purulence and bacteria in Gram stain of bronchopulmonary secretions are considered signs of respiratory infection, and medical decisions are made on the assumption that they are predictors of positive bacterial tracheal cultures (TCs). The purpose of this retrospective investigation was to establish the relationship of purulence and bacteria in TA from ventilated VLBW infants with positive TC and to identify its clinical significance. STUDY DESIGN: One hundred and seventy consecutively born VLBW infants (1996 to 1998) who remained on mechanical ventilation longer than 1 week were studied. Demographic, laboratory, and clinical data were obtained from hospital medical records. Purulence, defined by the number of polymorphonuclear leukocytes (PMNs) per low power field (LPF), was reported as light (<25 PMNs/LPF) or moderate/heavy (>or=25 PMNs/LPF) for every TA. RESULTS: Purulence was absent in 469 of 646 (72%) TA taken from 170 infants. Light purulence was present in 17% and moderate/heavy purulence in 11%. TCs were positive in 58% of non-purulent, 94% of light, and 100% of moderate/heavy purulent TA. Bacteria on Gram stain were present in 12% of non-purulent, 70% of light purulent, and 83% of moderate/heavy purulent TA. Moderate/heavy purulence in TA was predictive of a positive TC with Gram-negative bacilli (GNB) with 70% sensitivity, 100% specificity, 100% positive predictive value, and 67% negative predictive value. Purulence in TA, as well as GNB airway colonization, became more frequent as mechanical ventilation progressed and was not associated with a particular GNB species. There were 79 infants who never had purulent TA and 91 who, at some time during the hospitalization, did. At the time of first purulent TA, 65 (71%) of 91 infants were asymptomatic. Twenty-six infants (29%) had clinical deterioration for which they underwent sepsis work-up. Three had blood stream infection, 5 VAP, 5 tracheitis, and 13 respiratory complications of non-infectious etiology. Four of five VAP infants died; all others survived. CONCLUSION: In VLBW infants, purulence in TA is associated with prolonged endotracheal intubation and is temporally related to GNB airway colonization. At the time of the first purulent TA, the majority of mechanically ventilated VLBW infants are asymptomatic. Only a few symptomatic VLBW infants had nosocomial respiratory infection. Understanding the clinical significance of purulence and GNB in TA from this unique patient population is important for management and prognosis, and it may decrease concern for infection and the associated use of antibiotics.

A comparison of two airway suctioning frequencies in mechanically ventilated, very-low-birthweight infants.

INTRODUCTION: Endotracheal suctioning of mechanically ventilated, very-low-birthweight infants for removal of respiratory secretions can be associated with morbidity. Routine endotracheal suctioning is inadvisable, but the safe minimum endotracheal suctioning frequency for prevention of airway obstruction has not been determined. HYPOTHESIS: Decreasing suctioning frequency from every 4 hours to every 8 hours (plus as needed) would have no clinically important effect on the primary outcomes (nosocomial bloodstream infection [BSI], ventilator-associated pneumonia [VAP], and bacterial airway colonization) or secondary outcomes (reintubation rates, need for postural drainage, severity of bronchopulmonary dysplasia [BPD], neonatal mortality, duration of mechanical ventilation, and duration of hospitalization). METHODS: We conducted a sequential retrospective study of 90 very-low-birthweight infants who were mechanically ventilated for longer than 7 days and who underwent endotracheal suctioning every 4 hours (plus as needed) and 90 similar infants who underwent endotracheal suctioning every 8 hours (plus as needed). Two-pass endotracheal suctioning used during the study period required predetermined measurement of the suction catheter and prior instillation of saline. RESULTS: The 2 treatment groups were similar in demographic and clinical characteristics, including survival (90% and 85%), age at time of death (28 and 33 d), mean birthweight (926 and 934 g), gestational age (27 and 27 wk), duration of ventilation (29 and 27 d), and duration of stay (55 and 53 d). Regardless of suctioning frequency, airway colonization with Gram-positive cocci occurred in the majority of patients by 2 weeks of life. Forty-three percent of the infants suctioned every 4 hours and 44% of those suctioned every 8 hours became airway colonized with Gram-negative bacilli. No Gram-negative bacilli species was more likely to be associated with either treatment group. VAP was diagnosed in 5 of the infants suctioned every 4 hours and in 9 of the infants suctioned every 8 hours. Nosocomial BSI occurred in 18 of the infants suctioned every 4 hours and in 21 of the infants suctioned every 8 hours. The difference in incidence of VAP and BSI was due to a epidemic that started before the change in suctioning frequency. During hospitalization, approximately one fourth of the patients in each group required postural drainage and were reintubated 11 and 10 times per 100 ventilator days, respectively. A comparable number of infants in each group developed severe BPD and were discharged home on oxygen. Suctionings per patient per ventilator day were 6 for the group suctioned every 4 hours and 4 for the group suctioned every 8 hours (p < 0.01). CONCLUSIONS: A low-frequency suctioning regimen (every 8 hours plus as needed) can be implemented without increasing the incidence of nosocomial BSI, VAP, bacterial airway colonization, frequency of reintubation, need for postural drainage, severity of BPD, neonatal mortality, duration of mechanical ventilation, or duration of hospitalization. Although the minimum suctioning frequency for removal of unwanted respiratory secretions is yet to be established, a substantial reduction in endotracheal suctioning frequency appears to be safe.

Ventilator-associated pneumonia in very low-birth-weight infants at the time of nosocomial bloodstream infection and during airway colonization with Pseudomonas aeruginosa.

PURPOSE: To study retrospectively the incidence of ventilator-associated pneumonia (VAP) at the time of Pseudomonas aeruginosa nosocomial bloodstream infection (BSI) and at the time of P aeruginosa airway colonization. MATERIALS AND METHODS: Fifteen very low-birth-weight infants who had P aeruginosa BSI and 33 others who did not but who had P aeruginosa airway-colonization were studied. We correlated clinical data, blood cultures (BCs), and tracheal cultures (TCs) with radiologic findings from radio-graphs taken within 2 days before, the day of, and 1 day after BCs or TCs were first positive for P aeruginosa. Chest radiographs were graded by using semiquantitative scores for bronchopulmonary dysplasia and for pneumonia. RESULTS: Mean birth weight, gestational age, and age when BC or TC became positive were similar for patients with BSI and colonization. At the time of BSI, 2 infants had airway colonization with P aeruginosa; the TCs of the remaining 13 grew P aeruginosa as a new pathogen. Thirteen of 15 patients with BSI, but none of 33 infants with colonization, died within 2 days of positive BC. VAP was diagnosed in 13 of 15 patients with BSI and in 3 of 33 infants with colonization. CONCLUSION: Mechanically ventilated very low-birth-weight infants whose TCs yield P aeruginosa but whose BCs remain negative infrequently have VAP are presumed airway-colonized and are expected to survive. Conversely, VAP is likely to be found when BCs and TCs simultaneously grow P aeruginosa, and high mortality is anticipated.

Failure of systemic antibiotics to eradicate gram-negative bacilli from the airway of mechanically ventilated very low-birth-weight infants.

BACKGROUND: Gram-negative bacillary (GNB) airway colonization in mechanically ventilated newborns is associated with morbidity and mortality, which may be reduced if systemic antimicrobial therapy eradicates GNB from the airway. Efforts to do so in adults have met with variable success; similar experiences in newborns have not been reported. METHODS: From 1991 through 1998, 531 very low-birth-weight infants were mechanically ventilated longer than 2 weeks. The study group was 106 infants with GNB airway colonization. Sixty-four other neonates in whom GNB nosocomial bloodstream infections developed served as antibiotic treatment outcome control. RESULTS: Isolated from the airway were enteric (70 cases) and environmental (36 cases) GNB. Gentamicin alone or with ceftazidime (79), ceftazidime (11), piperacillin in combination with tazobactam or tobramycin (8), and tobramycin, in combination with ampicillin/sulbactam or mezlocillin (8) were the antimicrobials selected. Systemic antibiotics failed to eradicate GNB colonization in 97% of the cases. Six of the 106 infants with airway colonization died for reasons unrelated to infection. Sixty-four infants experienced 67 bloodstream infections as a result of enteric (53) and environmental (14) GNB. Gentamicin alone (23), with ceftazidime (26), or with clindamycin or ampicillin/sulbactam (9), piperacillin with tazobactam or tobramycin (3) and ceftazidime alone (6) were the antimicrobials selected. Survival occurred in 84% of the 67 nosocomial bloodstream infections. CONCLUSIONS: Systemic antibiotics do not consistently eradicate GNB from the airway of mechanically ventilated newborns, therefore its empirical use for prophylaxis or treatment of airway colonization should be discouraged.

Comparison of a closed (Trach Care MAC) with an open endotracheal suction system in small premature infants.

OBJECTIVE: To determine whether ventilated, low birth weight infants treated with closed versus open tracheal suction in a neonatal intensive care unit (NICU) differ as to airway bacterial colonization, nosocomial pneumonia, bloodstream infection (BSI), incidence and severity of bronchopulmonary dysplasia (BPD), neonatal mortality, frequency of suction, reintubation, and nurse preference. STUDY DESIGN: A total of 175 low birth weight infants (< or = 1250 gm) consecutively born (1997 to 1999), intubated, and ventilated in the delivery room were randomized on admission to the NICU to a closed (Trach Care MAC) or open suction group. Closed multi-use catheters were changed daily; open catheters were changed after every use. Two-pass endotracheal suctioning (both groups) was performed every 8 hours or as needed. Side-port connectors were not used; thus open suction required disconnection from ventilators. Tracheal aspirate cultures were obtained on admission and weekly thereafter. Nosocomial BSI (occurring after 48 hours of life) was documented by positive blood cultures. Radiographs taken before, during, and after tracheal aspirate cultures or BSIs were graded using a semiquantitative system for pneumonia and a modified score for BPD. Nurse preference regarding suction method was recorded. RESULTS: Of the original 175 patients, 10 (5 from each group) died and 32 others (16 form each group) were extubated at or before 7 days of life. The study population comprised 67 patients in the closed group and 66 in the open group who were ventilated longer than 1 week. Groups were not statistically different in terms of demographic and clinical characteristics, such as birth weight (837 vs 876 gm), ventilation (27 vs 26 days), and length of stay (49 vs 40 days). Airway colonization with Gram-positive cocci occurred in the majority of patients by 2 weeks of life, regardless of group. A total of 39% of infants in the closed group and 44% of infants in the open group became airway colonized with Gram-negative bacilli; differences were statistically significant. No Gram-negative bacilli species was more likely to be associated with either suction. Nosocomial pneumonia was diagnosed in five patients from each group. Nosocomial BSIs occurred in six closed suction infants and five open suction infants. A comparable number of infants in each group developed severe BPD and were discharged from the hospital on oxygen. A total of 28% of closed suction patients and 27% of open suction patients died. Infants in the closed versus open group were suctioned on average 4.4 and 4.1 times per day and were reintubated 9.7 and 8.6 times per 100 ventilator days, respectively. A total of 40 of 44 NICU nurses considered closed suction to be easier to use, less time-consuming, and better tolerated by the patient. CONCLUSIONS: Closed suction obviates the physiological disadvantage of ventilator disconnection without increasing the rate of bacterial airway colonization, frequency of endotracheal suction and reintubation, duration of mechanical ventilation, length of hospitalization, incidence of nosocomial pneumonia, nosocomial BSI, severity of BPD, and neonatal mortality. Although slightly more expensive, closed suction is perceived by nursing staff to be easier, less time-consuming, and better tolerated by small premature infants requiring mechanical ventilation for > or = 1 week.

Fluoride and bacterial content of bottled water vs tap water.

CONTEXT: Bottled water has become a status symbol and is frequently used in place of tap water. While both waters are considered safe to drink, is either more beneficial in preventing tooth decay and is there a difference in purity? OBJECTIVE: To determine the fluoride level and bacterial content of commercially bottled waters municipal tap water and to compare the results. DESIGN: Comparative study. SETTING: Cleveland, Ohio. SAMPLE: Fifty-seven samples of 5 categories of bottled waters were purchased from local stores. Samples of tap water were collected in sterile containers from the 4 local water processing plants. Fluoride levels were determined by an ion-selective electrode method. Water was cultured quantitatively and levels of bacteria were calculated as colony-forming units (CFUs) per milliliter. MAIN OUTCOME MEASURE: Fluoride levels and bacterial counts. RESULTS: Fluoride levels within the range recommended for drinking water by the Ohio Environmental Protection Agency, Cincinnati, 0.80 to 1.30 mg/L, were found in only 3 samples of bottled water tested. The fluoride levels of tap water samples were within 0.04 mg/L of the optimal fluoride level of 1.00 mg/L. The bacterial counts in the bottled water samples ranged from less than 0.01 CFU/mL to 4900 CFUs/mL, including 6 samples with levels substantially above 1000 CFUs/mL. In contrast, bacterial counts in samples of tap water ranged from 0.2 to 2.7 CFUs/mL. CONCLUSIONS: Five percent of the bottled water purchased in Cleveland fell within the required fluoride range recommended by the state, compared with 100% of the tap water samples, all of which were also within 0.04 mg/L of the optimal fluoride level of 1.00 mg/L. Use of bottled water based on the assumption of purity can be misguided. Recently, the Environmental Protection Agency, Washington, DC, published a final ruling that requires community water systems to regularly report to the public on the quality of local tap water; there are no similar proposals to determine the quality of bottled water through labeling.

Bloodstream infections in a neonatal intensive-care unit: 12 years' experience with an antibiotic control program.

OBJECTIVE: To assess the prevalence of gram-positive coccal (GPC), gram-negative bacillary (GNB), and fungal blood-stream infections (BSIs) during a 12-year period in which a consistent antibiotic treatment protocol was in place; to evaluate the efficacy of these antibiotic policies in relation to treatment, to the emergence of bacterial or fungal resistance, and to the occurrence of infection outbreaks or epidemics. STUDY DESIGN: Case series. METHODS: Demographic, clinical, and bacteriological information from 363 infants born during 1986 through 1991 and 1992 through 1997 who developed 433 blood-culture-proven BSIs was analyzed. Early-onset BSIs were defined as those infections discovered within 48 hours of birth, and late-onset BSIs as those that occurred thereafter. Suspected early-onset BSIs were treated with ampicillin and gentamicin, and suspected late-onset BSIs with vancomycin and gentamicin. Antibiotics were changed on the basis of organism antimicrobial susceptibility. RESULTS: Early-onset BSIs were noted in 52 of 21,336 live births and 40 of 20,402 live births during 1986 through 1991 and 1992 through 1997, respectively. GPC (83% due to group B streptococcus [GBS]) accounted for approximately one half of early-onset BSI cases and GNB (68% Enterobacteriaceae) for the remainder. Early-onset GBS declined from 24 to 11 cases (P=.04) and late-onset BSI increased from 111 to 230 cases (P<.01) from the first to the last study period. Sixty-eight percent of late-onset BSIs were due to GPC (primarily coagulase-negative Staphylococcus), 18% to GNB, and 14% to fungus. Over the study period, Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, and Pseudomonas aeruginosa isolated from the newborn intensivecare unit (unlike those strains from other hospital units) remained fully susceptible to ceftazidime and gentamicin. Although the hospitalwide prevalence of methicillin-resistant Staphylococcus aureus increased, all 17 newborn BSI cases were due to methicillin-sensitive strains. Prevalence of methicillin-resistant coagulase-negative Staphylococcus increased, although all strains remained vancomycin-susceptible, as did the 16 Enterococcus faecalis isolates. All fungi recovered (from 48 patients) were susceptible to amphotericin. CONCLUSION: We observed a decrease in the prevalence of early-onset BSIs due to GBS and an increase in late-onset BSIs due to GPC, GNB, and fungi. The combination of ampicillin and gentamicin for suspected early-onset BSIs and vancomycin and gentamicin for late-onset BSIs has been successful for treatment of individual patients without the occurrence of infection outbreaks or the emergence of resistance. Controlled antibiotic programs and periodic evaluations based on individual unit and not on hospitalwide antibiograms are advisable.

Radiological pulmonary changes during gram-negative bacillary nosocomial bloodstream infection in premature infants.

OBJECTIVE: The objective of the study was to characterize the changes that occur in chest radiographs at the time of gram-negative bacilli (GNB) nosocomial bloodstream infection (BSI) and to determine the contribution of bronchopulmonary dysplasia (BPD) and airway gram-negative bacterial pathogens to the clinical diagnosis of GNB nosocomial pneumonia. STUDY DESIGN: This retrospective investigation involved 41 BSI infants (study group) and 50 GNB airway colonized infants who had sepsis workup with negative blood cultures (control group). We correlated clinical findings (95 blood and 305 tracheal aspirate (TA) cultures) with radiographic findings noted within 2 days before, the day of, and the day after blood cultures. Two radiologists independently graded 258 films using a modified score for BPD and a semiquantitative score ("probable," "possible," or "unlikely") for pneumonia. RESULTS: Mean birth weight was 1057 gm and 1044 gm for the study and control groups, respectively. Of the study population, 54% were male, 21% were black, 89% received surfactant, 79% received dexamethasone, and 88% survived. Average age at the time of blood cultures for both groups was 23 days. Most common isolates from blood and TA were Klebsiella pneumoniae, Enterobacter cloacae, Escherichia coli, and Pseudomonas aeruginosa. Eight BSI infants died, mainly as a result of P. aeruginosa and K. pneumoniae; three control patients also died. Radiological findings were that BPD scores did not change in relation to BSI and were similar between study and control groups. Of the BSI patients, 21 presented with newly positive TA at the time of positive blood culture; "probable" or "possible" pneumonia was diagnosed in 18 of them. Their BPD scores were higher than those of the remaining BSI patients, of whom seven were already airway colonized, nine had negative TAs, and four were not intubated. Only one of these 20 patients had "possible" pneumonia noted on chest x-ray films. CONCLUSION: Radiographic signs of air space disease accompanied by the recovery of GNB respiratory pathogens from the blood and from a previously uncolonized airway strongly support the clinical diagnosis of GNB nosocomial pneumonia. Radiological signs of BPD are stable in relation to nosocomial BSI caused by GNB, but BPD radiological scores are higher among infants who also had a newly acquired respiratory GNB. BSI, new respiratory tract GNB, and BPD are critical associations for the clinical interpretation of radiographic changes in the ventilated newborn.

Pulmonary alveolar proteinosis. A report of two cases with diagnostic features in bronchoalveolar lavage specimens.

BACKGROUND: Pulmonary alveolar proteinosis (PAP) is a rare condition that has been associated with myriad diseases and disorders. Alveolar spaces are progressively filled with a phospholipoproteinaceous material, presumably related to a derangement of surfactant production and/or catabolism. The cytologic features of PAP in bronchoalveolar lavage (BAL) sediments are unique, and recognition of these characteristics can help guide clinical intervention. CASES: A 47-year-old male with a history of progressive dyspnea and recent pneumonia presented with a five-lobe alveolar infiltrate and subsequently underwent bronchoscopic examination. A 31-year-old female with chronic myelogenous leukemia in blast transformation developed unresponsive pulmonary infiltrates necessitating bronchoscopy with lavage. Both BAL lavage fluid sediments contained a homogeneous, basophilic, granular material typical of PAP. The material was composed of extracellular, multilamellated bodies when viewed by electron microscopy. Both patients required repeated therapeutic whole lung lavage, and one died of the disease eight months after the diagnosis. CONCLUSION: Clinical presentation, grossly milky BAL fluid and fluid sediment with light microscopic findings of basophilic, periodic acid-Schiff-positive, granular debris with cholesterol crystals and a few alveolar macrophages suggest this process. The light microscopic findings can be confirmed by ultramicroscopic demonstration of extracellular multilamellated bodies. BAL with appropriate examination of the effluent sediment facilitates the diagnosis of PAP.

Pulmonary infections in hospitalized patients with cirrhosis.

We have further characterized pulmonary infections by bronchoalveolar lavage in hospitalized patients with cirrhosis. Sixty-seven consecutive patients admitted to the Ohio State University Medical Center from 1992 to 1995 with liver disease who underwent bronchoscopy with bronchoalveolar lavage were identified. Twenty-one patients with cirrhosis and pneumonia were further analyzed. During the same period, we consecutively identified 23 patients without liver disease or immunosuppression, 19 patients with HIV infections, and 30 patients with cancer or pharmacologic immunosuppression who had bronchoscopy with bronchoalveolar lavage for pneumonia. These groups were included in these analyses as a control and immunosuppressed controls, respectively. Bronchoscopy isolated respiratory pathogens in 16 patients (76.2%) with cirrhosis and 6 patients (26.1%) in the control group (p = 0.002). Fungal organisms were most commonly found in patients with cirrhosis although several patients had more than one organism identified. The control group had mostly bacterial pathogens; the immunosuppressed controls were commonly infected with opportunistic organisms. Six (85.7%) of 7 patients with cirrhosis and fungal pneumonia died of their respiratory illness. Hospitalized patients with cirrhosis commonly have opportunistic pulmonary infections; diagnostic bronchoscopy and empiric antifungal therapy should be considered in those who do not respond to antibiotics.

Neonatal airway colonization with gram-negative bacilli: association with severity of bronchopulmonary dysplasia.

BACKGROUND: Airway colonization with Gram-negative bacilli (GNB) and Gram-positive cocci (GPC) is common in mechanically ventilated neonates. Whether GNB are related to nosocomial bloodstream infection (BSI) and/or to the severity of bronchopulmonary dysplasia (BPD) is unknown. METHODS: We prospectively examine this relationship using a cohort design. Data from 260 < or = 1250-g birth weight inborn infants (1991 to 1995) intubated > or = 2 weeks included 917 serial tracheal cultures and 583 blood cultures. The severity of BPD was assessed by duration of mechanical ventilation, oxygen dependency at 36 weeks of postconceptional age and the use of home oxygen supplementation. RESULTS: After 2 weeks of ventilation, 80% of the infants were colonized with GPC (Staphylococus epidermidis and Staphylococcus haemolyticus in 90% of the cases). Superimposed on 36% of these infants was GNB airway colonization with Klebsiella pneumoniae (25%), Enterobacter cloacae (25%), Escherichia coli (25%), Pseudomonas aeruginosa (10%), Serratia marcescen (10%), Acinetobacter baumannii and Haemophilus influenzae (5%). Comparison between 174 GPC- and 86 GNB-colonized infants showed that demographics, birth weight, gestational age, perinatal risk factors and mortality were similar. Fifteen percent of GNB-colonized infants developed BSI caused by GNB and 14% developed BSI caused by GPC. No significant temporal relationship between airway colonization and BSI was noted. GNB infants were ventilated longer and required oxygen at 36 weeks of postconceptional age and home oxygen supplementation twice as often as infants colonized only with GPC. GNB colonization was a predictor of severe BPD after controlling for ventilation. Ureaplasma colonization occurred in 28% of GNB-colonized and 33% of noncolonized infants and was not a predictor of BPD severity. CONCLUSION: GNB airway colonization creates a moderate risk for BSI. Antibiotic treatment does not regularly eradicate GNB. GNB airway colonization is associated with severe BPD, but further studies will be necessary before therapeutic efforts to eradicate GNB from the airways should be undertaken.

The utility of non-beta-lactam antimicrobial MICs as markers to distinguish oxacillin-resistant from oxacillin-susceptible strains of Staphylococcus epidermidis.

Among 6,068 strains of Staphylococcus epidermidis, 75.5% were oxacillin-resistant. Oxacillin-susceptible strains were more frequently susceptible to erythromycin, clindamycin, ciprofloxacin, trimethoprim/sulfamethoxazole, gentamicin, and tetracycline than oxacillin-resistant strains. With the exception of erythromycin, non-beta-lactam MICs were less discriminatory for identifying oxacillin-resistant strains with oxacillin MICs < or = 2 micrograms/ml than for those with oxacillin MICs > or = 4 micrograms/ml.

In vitro activities of quinolones, beta-lactams, tobramycin, and trimethoprim-sulfamethoxazole against nonfermentative gram-negative bacilli.

From 1991 to 1995, 8,975 nonfermentative gram-negative bacilli were isolated from patients at The Ohio State University Medical Center: 71% Pseudomonas aeruginosa, 14% Stenotrophomonas maltophilia, 7.6% Acinetobacter baumannii, and < 2% each of 25 other species. The MICs of trovafloxacin (CP-99,219), ciprofloxacin, ofloxacin, ampicillin-sulbactam, piperacillin, piperacillin-tazobactam, ceftazidime, cefoperazone, ceftriaxone, imipenem, tobramycin, and trimethoprim-sulfamethoxazole (TMP-SMZ) were determined for 308 isolates, representing 13 species, by a standardized broth microdilution method. The activities of all drugs were species dependent. The fluoroquinolones had inconsistent activity against most species, although several relatively uncommon nonfermenters were consistently susceptible or resistant. Trovafloxacin was considerably more active than ciprofloxacin and ofloxacin against S. maltophilia, A. baumannii, and several less common species. Among the beta-lactams, relative activities varied considerably; overall, imipenem had the broadest spectrum of activity but was inactive against S. maltophilia and Burkholderia cepacia isolates. Tobramycin and TMP-SMZ had stereotypic spectra of activity. Tobramycin was active against most species except S. maltophilia, Alcaligenes xylosoxidans subsp. xylosoxidans, Burkholderia spp., and Weeksella virosa. TMP-SMZ was active against most species except P. aeruginosa and Pseudomonas fluorescens-putida. A review of laboratory records indicated few changes in susceptibility patterns from 1991 to 1995; the only clear trend was toward increasing P. aeruginosa resistance to all classes of drugs.

Emergence of bacterial resistance to imipenem and ciprofloxacin in a university hospital.

We have continuously monitored the in-vitro activities of imipenem and ciprofloxacin against large numbers of non-fastidious clinical isolates. After eight years of use, 97-100% of Enterobacteriaceae and Acinetobacter baumannii remained susceptible to imipenem, but susceptibility of Pseudomonas aeruginosa declined from 100% to 91%. After six years of use, 94%-100% of Enterobacteriaceae (except Providencia stuartii) remained susceptible to ciprofloxacin but susceptibility of P. stuartii, A. baumannii and P. aeruginosa declined from 100% to 46%, 66% and 84%, respectively. Oxacillin-resistant staphylococci were considered to be resistant to imipenem and all beta-lactams. There were no quinolone-resistant staphylococci observed in 1986, but susceptibilities of Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus haemolyticus to ciprofloxacin decreased to 85-93% for oxacillin-susceptible strains and to 7-39% for oxacillin-resistant strains. Enterococcus faecalis has remained susceptible to imipenem and the modal MIC of ciprofloxacin has remained 1 mg/L; however, susceptibility to ciprofloxacin 2 mg/L decreased from 94% to 64%. Imipenem-quinolone cross-resistance was observed for staphylococci but not for P. aeruginosa.

Assessment of microbial growth in intravenous immune globulin preparations.

The ability of intravenous immune globulin (IVIG) products to support bacterial and yeast growth was studied. Microtiter plates containing preparations of eight commercially available IVIG products were inoculated with 10(5) colony-forming units (CFUs) of bacteria per milliliter or 10(3) CFUs of yeast cells per milliliter. The organisms used were Candida albicans, Torulopsis glabrata, Pseudomonas aeruginosa, Staphylococcus epidermidis, and Klebsiella pneumoniae. The plates were incubated at 3, 25, or 37 degrees C for seven days. For each organism and each temperature, a total of 16 identical wells were studied per preparation. Optical density was measured at 24-hour intervals to determine bacterial growth; yeast growth was determined visually. None of the IVIG preparations supported bacterial growth at any temperature over the seven-day period. No preparation supported visible yeast growth at 3 degrees C; however, most of the preparations did support yeast growth at 25 and 37 degrees C. The failure of the IVIG preparations to support bacterial growth at all temperatures and yeast growth at 3 degrees C suggests that a reconsideration of recommended expiration dates for IVIG products may be warranted.

Comparison of BACTEC Plus 26 and 27 media with and without fastidious organism supplement with conventional methods for culture of sterile body fluids.

We compared the BACTEC Plus 26/27 culture system (Becton Dickinson Diagnostic Instrument Systems, Sparks, Md.) with and without fastidious organism supplement with conventional centrifugation preparation and plating for the recovery and speed of detection of microorganisms. A total of 1,101 sterile body fluid specimens were collected and processed at five hospital laboratories, yielding 234 (21%) positive cultures. Of the 176 isolates considered clinically significant, 133 (76%) were recovered by both the BACTEC system and conventional culture, while 28 (16% [P < 0.005]) were recovered by BACTEC only and 11 (6%) were recovered by conventional culture alone. There were no statistically significant differences in the speed of detection of microbial growth. It was found that BACTEC, with or without the addition of fastidious organism supplement, exhibited improved sensitivity for the recovery of microorganisms, including fastidious bacteria.

A five-year multicenter study of the susceptibility of the Bacteroides fragilis group isolates to cephalosporins, cephamins, penicillins, clindamycin, and metronidazole in the United States.

Over 2800 clinical strains of the Bacteroides fragilis group were collected during a 5-year period from ten geographically separate sites and tested for their susceptibility to various antimicrobial agents using a broth microdilution method. Among the cephalosporins, ceftizoxime was the most active (13% resistance) and importantly exhibited relatively equal activity against both B. fragilis species and non-B. fragilis species. Cefotaxime exhibited similar activity with an overall resistance rate of 18%. Both ceftriaxone and cefoperazone were appreciably less active cephalosporins especially against non-B. fragilis species. With regard to cephamycins, cefoxitin (MIC90, 32 micrograms/ml) was more active than cefotetan (MIC90, > or = 256 micrograms/ml) and cefmetazole (MIC90, 64 micrograms/ml). Non-B. fragilis species were highly resistant to cefotetan and cefmetazole. Imipenem was highly active against all strains with the exception of four strains of B. fragilis. Ampicillin-sulbactam, amoxicillin-clavulanate, piperacillin-tazobactam, and cefoperazone-sulbactam were all highly active with resistance rates < 2%. No resistance was detected to metronidazole, whereas 14% of isolates were resistant to clindamycin. When compared with other studies, these findings underscore the wide variability in susceptibility patterns reported nationwide and the need to continue monitoring these patterns to aid in choosing the most active compounds for therapy.

Three cases of Haemophilus influenzae amnionitis.

Haemophilus influenzae is an uncommon cause of obstetric and neonatal infections. Three cases of amnionitis caused by H. influenzae are presented. These cases occurred in three maternal transports to our tertiary level hospital within an 11-day period. The organisms were analyzed for biotypes, penicillinase status, and composition of membrane fatty acids.