Reporter bacteriophage T7NLC utilizes a novel NanoLuc::CBM fusion for the ultrasensitive detection of Escherichia coli in water.

Rapid detection of bacteria responsible for foodborne diseases is a growing necessity for public health. Reporter bacteriophages (phages) are robust biorecognition elements uniquely suited for the rapid and sensitive detection of bacterial species. The advantages of phages include their host specificity, ability to distinguish viable and non-viable cells, low cost, and ease of genetic engineering. Upon infection with reporter phages, target bacteria express reporter enzymes encoded within the phage genome. In this study, the T7 coliphage was genetically engineered to express the newly developed luceriferase, NanoLuc (NLuc), as an indicator of bacterial contamination. While several genetic approaches were employed to optimize reporter enzyme expression, the novel achievement of this work was the successful fusion of the NanoLuc reporter to a carbohydrate binding module (CBM) with specificity to crystalline cellulose. This novel chimeric reporter (nluc::cbm) bestows the specific and irreversible immobilization of NanoLuc onto a low-cost, widely available crystalline cellulosic substrate. We have shown the possibility of detecting the immobilized fusion protein in a filter plate which resulted from a single CFU of E. coli. We then demonstrated that microcrystalline cellulose can be used to concentrate the fusion reporter from 100 mL water samples allowing a limit of detection of <10 CFU mL-1E. coli in 3 hours. Therefore, we conclude that our phage-based detection assay displays significant aptitude as a proof-of-concept drinking water diagnostic assay for the low-cost, rapid and sensitive detection of E. coli. Additional improvements in the capture efficiency of the phage-based fusion reporter should allow a limit of detection of <10 CFU per 100 mL.

Predicting Dental Caries in Young Children in Primary Health Care Settings.

Young children need increased access to dental prevention and care. Targeting high caries risk children first helps meet this need. The objective of this study was to develop a parent-completed, easy-to-score, short, accurate caries risk tool for screening in primary health care settings to identify children at increased risk for cavities. A longitudinal, prospective, multisite, cohort study enrolled (primarily through primary health care settings) and followed 985 (out of 1,326) 1-y-old children and their primary caregivers (PCGs) until age 4. The PCG completed a 52-item self-administered questionnaire, and children were examined using the International Caries Detection and Assessment Criteria (ICDAS) at 12 ± 3 mo (baseline), 30 ± 3 mo (80% retention), and 48 ± 3 mo of age (74% retention). Cavitated caries lesion (dmfs = decayed, missing, and filled surfaces; d = ICDAS ≥3) experience at 4 y of age was assessed and tested for associations with questionnaire items using generalized estimating equation models applied to logistic regression. Multivariable analysis used backward model selection, with a limit of 10 items. At age 4, 24% of children had cavitated-level caries experience; 49% were female; 14% were Hispanic, 41% were White, 33% were Black, 2% were other, and 10% were multiracial; 58% enrolled in Medicaid; and 95% lived in urban communities. The age 4 multivariable prediction model, using age 1 responses (area under the receiver operating characteristic curve = 0.73), included the following significant (P < 0.001) variables (odds ratios): child participating in public assistance programs such as Medicaid (1.74), being non-White (1.80-1.96), born premature (1.48), not born by caesarean section (1.28), snacking on sugary snacks (3 or more/d, 2.22; 1-2/d or weekly, 1.55), PCG cleaning the pacifier with juice/soda/honey or sweet drink (2.17), PCG daily sharing/tasting food with child using same spoon/fork/glass (1.32), PCG brushing their teeth less than daily (2.72), PCG's gums bleeding daily when brushing or PCG having no teeth (1.83-2.00), and PCG having cavities/fillings/extractions in past 2 y (1.55). A 10-item caries risk tool at age 1 shows good agreement with cavitated-level caries experience by age 4.

Features of prescription drug monitoring programs associated with reduced rates of prescription opioid-related poisonings.

BACKGROUND: The United States is in the midst of an opioid epidemic. In addition to other system-level interventions, all states have responded during the crisis by implementing prescription drug monitoring programs (PDMPs). This study examines associations between specific administrative features of PDMPs and changes in the risk of prescription opioid-related poisoning (RxORP) over time. METHODS: This longitudinal, observational study utilized a 'natural experiment' design to assess associations between PDMP features and risk of RxORP in a nationally-representative population of privately-insured adults from 2004 to 2014. Administrative health claims data were used to identify inpatient hospital admissions and emergency department visits related to RxORP. Generalized estimating equation Poisson regression models were used to examine associations between specific PDMP features and changes in relative risk (RR) of RxORP over time. RESULTS: In adjusted analyses, states without PDMPs experienced an average annual increase in the rate of RxORP of 9.51% over the study period, while states with operational PDMPs experienced an average annual increase of 3.17%. The increase in RR of RxORP over time in states with operational PDMPs was significantly less than increases in states without PDMPs. States with specific features, including those that monitored more schedules or required more frequent data reporting, experienced stronger protective effects on the RR of RxORP over time. CONCLUSION: This study examined associations between specific PDMP features and RxORP rates in a nationally-representative population of privately-insured adults. Results of this study may be used as empirical evidence to guide PDMP best practices.

Pediatric neuroblastoma: postoperative radiation therapy using less than 2000 rad.

There is considerable controversy regarding the role of radiation therapy in the treatment of neuroblastoma. Postoperative irradiation in the range of 2500-4000 rad is commonly used in the treatment of Evans Stage II or III disease, but there are no data in the literature to suggest the optimum dose of radiation that is necessary. Because much lower doses have been used at the University of Florida, a retrospective study was undertaken in an attempt to determine the optimum dose necessary in conjunction with surgery. From March 1964 through July 1979, 21 children with Stage II or III neuroblastoma were seen at the University of Florida. One patient died postoperatively. The remainder received postoperative irradiation with doses ranging from 900 to 4500 rad. The lower dose of radiation used did not adversely influence survival, particularly for patients less than two years of age at diagnosis. In this group, no patient had a local recurrence or died of disease, even though nine of 15 available patients received doses of 900-1500 rad.

Thoracoscopy. Early diagnosis of interstitial pneumonitis in the immunologically suppressed child.

Interstitial pneumonitis in immunosuppressed patients demands prompt diagnosis and treatment. In an effort to achieve a simple yet highly accurate method of diagnosis, we have evaluated the usefulness of thoracoscopic examination. Twenty-seven procedures have been performed in 24 patients between the ages of 17 months and 18 years. All patients were immunosuppressed, most for treatment of malignant processes. All procedures have been performed under anesthesia with intravenously administered ketamine, without endotracheal intubation. A definitive diagnosis has been made in every case, with pneumonia due to Pneumocystis carinii being identified in 18 instances. Complications have been minimal and include four minor pneumothoraces, two instances of bleeding, and two instances of prolonged air leak. Mortality attributable to the procedure has been nil, although five patients have died due to their underlying diseases within 30 days of the thoracoscopic procedure. Thoracoscopy has proven to be a rapid and safe technique for providing accurate histologic and bacteriologic diagnoses in these critically ill children.

Traumatic perforation of the hypopharynx in infants.

Three cases of traumatic perforation and pseudodiverticulum of the hypopharynx in newborn infants are presented. This lesion may be encountered with increasing frequency as a result of the more aggressive management of newborn infants with respiratory distress. Early recognition of this complication is essential to successful therapy. Appropriate management includes local drainage, treatment with antibiotics, and use of a nasogastric tube or gastrostomy to provide a safe route for intestinal alimentation.

Morphologic and functional effects of esophageal cryotherapy.

With advances in equipment design, cryotherapy has been applied increasingly in the management of various integumentary disorders. Recently, endobronchial cryotherapy has been utilized in the treatment of acquired tracheal strictures. To evaluate the effects of cryotherapy upon the esophagus, we cooled the distal esophagus of 12 cats to -70 degrees to -80 degrees C. for 2 minutes with a specially constructed cryoprobe. The animals were put to death in pairs 90 minutes, 4 days, 10 days, 14 days, 28 days, and 42 days following cryotherapy. Immediately prior to sacrifice, all animals were studied with esophageal cinefluoroscopy and esophageal manometrics. All animals tolerated this degree of cryotherapy and gained weight on an ad libitum diet. Manometric studies demonstrated diminished amplitude of peristalsis immediately following cryotherapy, but prompt return to a normal manometric pattern. There was no evidence of esophageal dysfunction on esophageal cinefluoroscopy. Morphologic examination demonstrated superficial ulceration between 4 and 10 days following cryotherapy, with subsequent healing. The esophagus in animals put to death 28 and 42 days following cryotherapy appeared histologically normal. These studies demonstrate the safety of cryotherapy to -70 to -80 degrees C. upon the esophageal wall and suggest investigation of the usefulness of this modality in the treatment of local esophageal webs and strictures.

Endoscopic evaluation of the esophagus in infants and children immediately following intraoperative use of transesophageal echocardiography.

OBJECTIVE: Intraoperative transesophageal echocardiography (TEE) has evolved as an essential technique for use during pediatric cardiac surgery; however, few studies have evaluated the safety of TEE in children. This series reports endoscopic examination of the esophagus following intraoperative TEE in pediatric patients. METHODS: Fifty children undergoing congenital heart surgery underwent flexible esophagoscopy that was performed after completion of their heart surgery and after the removal of the transesophageal echo probe. The patients' ages ranged from 4 days to 10 years old, and their weight ranged from 3.0 to 39.8 kg, with a mean weight of 12.6 kg. RESULTS: Thirty-two of 50 patients (64%) had abnormal results shown on esophageal examinations; this occurred more frequently in the subset of patients weighing < 9 kg. No long-term feeding or swallowing difficulties were noted in any of the 48 patients who survived. CONCLUSIONS: Intraoperative TEE in infants and children frequently caused mild mucosal injury. Care must be exercised in the insertion and manipulation of the probes.

Congenital anorectal malformations. Harbingers of sacrococcygeal teratomas.

Since 1973, seven infants treated at the University of Florida, Gainesville, for congenital anorectal anomalies have been found to have concomitant sacrococcygeal teratomas. As anorectal anomalies occur in approximately one in every 5,000 live births and sacrococcygeal teratomas are noted in one in every 40,000 live births, this presentation would suggest a prevalence that precludes a random association. The presence of significant anorectal strictures in these patients resulted in a delay in the diagnosis of the presacral teratomas. As the frequency of malignant change in sacrococcygeal teratomas has been documented to increase proportionately with age, early diagnosis and excision of these lesions is essential. Our experience with these children suggests that the presence of congenital anorectal malformations in infancy should raise the physician's index of suspicion for associated sacrococcygeal teratomas.

Wilms' tumor with acute abdominal pain.

Acute abdominal pain is the presenting manifestation in approximately 30% of all patients with Willms' tumor. In a small proportion of these patients this pain is significant enough to engender a diagnosis of an acute surgical abdomen. Six of 38 patients with Wilms' tumors treated between the years 1965 and 1975 at the Shands Teaching Hospital of the University of Florida Medical Center have had significant pain. Our experience with these patients emphasizes the importance of thoroughly palpating the abdomen of any child with a suspected acute surgical condition, following induction of anesthesia and prior to initiating the operation. In the absence of any evidence of an acute surgical problem at the time of the exploratory laparotomy, it is also imperative that a careful intra-abdominal examination be performed to exclude the presence of conditions, such as Wilms tumor of the kidney, that may occasionally present in this manner.

The microbiology of neonatal peritonitis.

To review the bacteriology of neonatal intra-abdominal sepsis, we reviewed peritoneal cultures from 86 newborns undergoing operation for necrotizing enterocolitis (NEC) for the type and incidence of microorganism recovered. As a control, we conducted a similar review in 59 children with perforated appendicitis during the same period. Necrotizing enterocolitis was characterized by a lower incidence of polymicrobial contamination (1.7 organisms per patient vs 2.4 organisms per patient, NEC vs appendicitis) and an uncharacteristic pattern of isolates. Although enteric gram-negative bacilli were recovered in 80% of newborns, the incidence of Escherichia coli was only 21% in the NEC group vs 69% in the appendicitis group, while Klebsiella and Enterobacter species represented the most common gram-negative isolates recovered (63% vs 17%). More than 50% of neonatal cultures yielded gram-positive cocci, most frequently coagulase-negative staphylococci (30% vs 0%) and enterococci (17% vs 5%), as compared with more frequent streptococcal isolates in the appendicitis group (50% vs 10%). Anaerobes were seldom recovered in NEC cases (6%), but they were present in 50% of appendicitis cases. Additionally, Candida isolates were recovered in 10% of NEC cases (0% of appendicitis group). These results indicate the unique bacteriology of peritonitis in the critically ill newborn and probably reflect abnormal colonization in the neonatal intensive care unit.

Near-total pancreatectomy in persistent infantile hypoglycemia.

To prevent the devastating effects of hypoglycemia on the CNS of the infant, persistent infantile hypoglycemia should be recognized early and treated promptly. To avoid mental retardation, surgical intervention should not be considered as a last resort. When an insulinoma is identified at surgery, subtotal pancreatectomy is adequate in achieving normoglycemia. However, the most common cause of insulin excess in infancy appears to be islet cell hyperplasia or nesidioblastosis, both diffuse processes. In these patients, subtotal pancreatectomy is often ineffective in achieving normoglycemia. Near-total pancreatectomy, retaining the spleen and duodenum, is a safe procedure, well tolerated by infants and children, and should be considered early for the correction of hypoglycemia of infancy that is not readily controlled by medical intervention.

Endotracheal cryotherapy in the treatment of refractory airway strictures.

In 1977 we reported the successful use of endotracheal cryotherapy in the treatment of experimental tracheal strictures. This technique has now been employed in the treatment of 29 refractory airway lesions in 27 patients ranging in age from 3 months to 42 years. The technique utilizes a nitrous oxide cryoprobe measuring 3 mm in diameter and 43 cm in length. The tip of the probe is applied directly to the stricture through the endoscope and cooled to -80 degrees C for 45 seconds. On removal of the probe, the frozen tissue is resected with biopsy forceps. Cryotherapy has successfully relieved the airway strictures in 20 of the 24 lesions in which treatment was completed. The technique of endotracheal cryotherapy has proved helpful in the management of selected patients with refractory airway strictures.

Thoracoscopy for intrathoracic tumors.

The technique of thoracoscopy has been performed on 49 occasions in 45 patients for the diagnosis or staging of intrathoracic tumors. The patients ranged from 8 months to 68 years old. Eight procedures were performed for pleural disease, 16 for mediastinal masses, 22 for parenchymal lesions, and 3 for intrathoracic staging. Seventeen patients had had previous invasive procedures performed without a pathological diagnosis being established. In 28 thoracoscopy procedures, a positive diagnosis for malignancy was obtained; in 6 instances, areas of unsuspected tumor involvement were identified. A correct diagnosis was obtained by thoracoscopy in 45 procedures for a 92% overall accuracy rate. There was no clinically significant morbidity in this series and no procedure-related mortality. Thoracoscopy, performed under stellate ganglion block and regional anesthesia, has proved to be a very attractive method of diagnosing intrathoracic neoplasia with very low morbidity.

Pulmonary metastases in childhood sarcoma.

A retrospective analysis of 56 patients less than 19 years old with pulmonary metastases from previously diagnosed sarcomas was performed. Thirty patients had primary osteogenic sarcoma. Eighteen of them underwent a total of 37 thoracotomies. Overall survival was 20%. Twelve patients were noted to have metastatic Wilms' tumor, and 5 of them underwent 7 thoracotomies. This group had a 25% overall survival. Six patients had pulmonary metastases from Ewing's sarcoma. Four of these required pulmonary resection. The overall survival in these patients was 50%. The remaining 8 patients had pulmonary resection for various soft tissue malignancies, with an overll survival of 50%. The results of this review indicate a role for aggressive pulmonary resection in patients in whom the primary tumor is controlled and there is no other evidence of metastatic disease. The frequency of bilaterality and multicentric implants in patients with osteogenic sarcoma and Wilms' tumor suggest the efficacy of a median sternotomy approach in these patients.

Exposing the circumflex coronary artery: the heartflip technique.

A method to expose the circumflex coronary artery in its course in the atrioventricular groove is introduced. No special equipment or assistance is required. This method also can be applied to expose the obtuse marginal branches of the circumflex coronary artery. Adverse effects have not been observed.

Unilateral pulmonary agenesis, esophageal atresia, and distal tracheoesophageal fistula.

A small-for-gestational-age, premature infant with the combination of unilateral pulmonary agenesis, esophageal atresia, and distal tracheoesophageal fistula was treated successfully by early gastrostomy and delayed fistula division with esophagoesophagostomy. Only 2 other successfully treated cases have been reported previously. Both were full-term infants treated with early division of the tracheoesophageal fistula and esophagoesophagostomy. Gestational age, size, and associated medical problems need to be considered when planning operative therapy for these babies.

Compensatory renal growth and function in postnephrectomized patients with Wilms tumor.

The objective of this study was to determine whether or not renal growth and function were adversely affected in the remaining kidneys of patients who had undergone nephrectomy for Wilms tumor. These patients received chemotherapy and some radiotherapy (tumoricidal agents which might affect the remaining kidney). Renal growth was compared between the treatment groups and normal renal growth. Hypertrophy did occur and did not appear to be affected by subsequent treatment. Renal function was minimally altered in all treatment groups irrespective of the type of treatment.

Asthma prevalence, cost, and adherence with expert guidelines on the utilization of health care services and costs in a state Medicaid population.

OBJECTIVE: To provide a descriptive analysis of asthma prevalence and costs in a Medicaid population and gauge the degree of adherence with expert guidelines for asthma medication management from the National Asthma Education and Prevention Program. DATA SOURCES: Kentucky Medicaid administrative data for 1996. STUDY DESIGN: A cross-sectional retrospective analysis was used to determine adherence with asthma medication guidelines and utilization of asthma-related health care services and costs. Multivariate logistic regression was used to determine the relationship between nonadherence with the guidelines and utilization of health care services. PRINCIPAL FINDINGS: Of the 530,000 Medicaid recipients, 24,365 (4.6 percent) were identified as having asthma. Average annual asthma-related costs ($616) accounted for less than 20 percent of total health care costs ($3,645). Nonadherence to the guidelines was prevalent. Less than 40 percent of the patients received a prescription for a rescue medication, and fewer than 10 percent of the patients who received daily inhaled short-acting beta-2 agonists were regular users of inhaled steroids. Nonadherence to the guidelines was associated with an increased risk of an asthma-related hospitalization (odds ratio = 1.5, p < .05). CONCLUSIONS: Guideline nonadherence was widespread and associated with an increase in exacerbations of asthma that resulted in hospitalizations. Asthma prevalence and utilization of health care services in a Medicaid population were similar to previous estimates reported nationally and in health maintenance organizations.

Intrahepatic cholestasis with parental alimentation.

From July 1971 to March 1975, elevan infants receiving total or partial parenteral alimentation at the University of Florida showed histologic evidence of intrahepatic cholestasis. The clinical records of these patients have been examined. These infants were critically ill and had protracted hospital courses with only two survivors. Liver biopsies demonstrated marked cholestasis with some fibrosis and thickening of the limiting membrane of the hepatocyte. In those patients in whom serial liver biopsies were obtained, hepatic histology returned toward normal, paralleling improvement in liver function studies, as intravenous alimentation was discontinued. Careful monitoring of the liver function tests is essential to detect this progressive abnormality as early as possible and discontinue intravenous alimentation. Follow-up as long as two and a half years in the two surviving patients has demonstrated no chronic dysfunction.