Bacteriophage Therapy as a Potential Management Option for Surgical Wound Infections.

To investigate the potential role of bacteriophages in the treatment of surgical infections, we conducted a retrospective analysis of four surgical patients who have sought treatment at the Eliava Phage Therapy Center, Tbilisi, Georgia. Two patients had chronic osteomyelitis, one presented with a diabetic foot ulcer, and the fourth patient had developed a severe infectious complication after skin grafting surgery. Patients were treated with different combinations of bacteriophage preparations, based on the sensitivity of the isolated bacterial strain toward commercially available bacteriophages. The treatment lasted on average for 1 month, and positive results were obtained in all four cases: the wounds have healed, the general health status of the patients has improved. No allergic or adverse reactions have been observed throughout the treatment.

Phage therapy against Achromobacter xylosoxidans lung infection in a patient with cystic fibrosis: a case report.

Respiratory infections can lead to serious complications in CF patients, especially when infected with antibiotic resistant bacteria. Alternative treatments for these infections are being sought out to help address this problem. We present a clinical case of a cystic fibrosis (CF) patient, with multi-drug resistant (MDR) Achromobacter xylosoxidans chronic lung infection who was successfully managed with bacteriophage therapy.

Use of CTX-I and PINP as bone turnover markers: National Bone Health Alliance recommendations to standardize sample handling and patient preparation to reduce pre-analytical variability.

The National Bone Health Alliance (NBHA) recommends standardized sample handling and patient preparation for C-terminal telopeptide of type I collagen (CTX-I) and N-terminal propeptide of type I procollagen (PINP) measurements to reduce pre-analytical variability. Controllable and uncontrollable patient-related factors are reviewed to facilitate interpretation and minimize pre-analytical variability. INTRODUCTION: The IOF and the International Federation of Clinical Chemistry (IFCC) Bone Marker Standards Working Group have identified PINP and CTX-I in blood to be the reference markers of bone turnover for the fracture risk prediction and monitoring of osteoporosis treatment. Although used in clinical research for many years, bone turnover markers (BTM) have not been widely adopted in clinical practice primarily due to their poor within-subject and between-lab reproducibility. The NBHA Bone Turnover Marker Project team aim to reduce pre-analytical variability of CTX-I and PINP measurements through standardized sample handling and patient preparation. METHODS: Recommendations for sample handling and patient preparations were made based on review of available publications and pragmatic considerations to reduce pre-analytical variability. Controllable and un-controllable patient-related factors were reviewed to facilitate interpretation and sample collection. RESULTS: Samples for CTX-I must be collected consistently in the morning hours in the fasted state. EDTA plasma is preferred for CTX-I for its greater sample stability. Sample collection conditions for PINP are less critical as PINP has minimal circadian variability and is not affected by food intake. Sample stability limits should be observed. The uncontrollable aspects (age, sex, pregnancy, immobility, recent fracture, co-morbidities, anti-osteoporotic drugs, other medications) should be considered in BTM interpretation. CONCLUSION: Adopting standardized sample handling and patient preparation procedures will significantly reduce controllable pre-analytical variability. The successful adoption of such recommendations necessitates the close collaboration of various stakeholders at the global stage, including the laboratories, the medical community, the reagent manufacturers and the regulatory agencies.

[What happens when the child gets bacteriophage per os?].

The aim of the work was to define the distribution of phages administered per os to children for medical reasons, and the immune response. 102 children aged from 5 days to 15 years with different diseases of bacterial etiology (pneumonia, sepsis, urinary infection, pharyngitis/sinusitis, enteral infection) were monitored. Pyobacteriophage was being included into the complex therapy. The drug was administered per os. In 6/7 of blood, 48/55 urine and 64/75 stool samples taken on the 3-5th day of treatment different components of pyobacteriophage were revealed. The titers varied from 103 to 105 pfu/ml. No age differences were seen. In two weeks after the onset of the phagotherapy the antibodies to phages were tested in the blood serum using the neutralization reaction method. The blood samples were taken from 31 patients. In 14 of them the antibodies neutralizing 52.5-97.3% of the phage activity were seen. A significant age-related peculiarity was determined: in newborns and infants the antibodies were not revealed or their activity was low. Obtained results confirm the reasonability to use of peroral phagotherapy in gastro-intestinal infections. At the same time it was ascertained that the phages taken per os can permeate into the internal environment of the organism and thus the peroral phagotherapy can be used to treat systemic infections and urinary tract infections as well. Absence or low production of the antiphage antibodies in newborns and infants suggests high efficacy of the phagotherapy in this age group.

Study of therapeutic potential of the experimental pseudomonas bacteriophage preparation.

Wide expansion of the infections caused by multi-antibiotic resistant strains of P. aeruginosa revived the idea of phage therapy with pseudomonas phage preparations for treatment and prevention of the bacterial infection diseases. The purpose of this study was examination of an experimental series of the therapeutic-prophylactic pseudomonas bacteriophage preparation, with wide spectra of lytical activity and high therapeutic potential. Newly isolated phage clones of P. aeruginosa were studied by the basic tests (such as host range, lysis stability, physiological and immunogenic properties of the phages, host dependent restriction/modification phenomena and automatic reproduction ability of the phages on the UV inactivated strains) determining their virulent nature. An experimental series of the therapeutic-prophylactic pseudomonas phage preparation were developed from the genuine virulent phage clones CF1/1; CF1/7; P.a.N1, P.a.N2 and P.a.N4. The phage preparation successfully passed in vitro (efficacy, sterility, stability) and in vivo (safety, definition of therapeutic potential) controls on experimental animals (white mice). The host range of the experimental pseudomonas phage preparation equals 99.5% of 206 strains of P. aeruginosa. Preclinical testing of the experimental pseudomonas phage preparation on white mice revealed that the therapeutic efficacy of the phage preparation was higher (80-100%) than that of the antibiotic-ciprinol (50-80%). Noteworthy, 100% therapeutic efficacy was observed after combined application of the antibiotic and the phage preparation.

Serum and plasma fragments of C-telopeptides of type I collagen (CTX) are stable during storage at low temperatures for 3 years.

BACKGROUND: Control of pre-analytical variables is essential for successful application of biological markers, including bone resorption markers, in clinical trials and routine use. The effect of storage temperature on stability of bone resorption markers have not been subject of systematically investigation, and therefore the present study was set out to determine the stability of C-telopeptides of type I collagen (CTX) in serum and plasma samples stored frozen for 3 years. METHODS: The serum and plasma levels of CTX were determined in samples aliquoted and stored frozen for up to 3 years. RESULTS: No significant decrease could be detected in neither serum nor plasma samples after 3 years of storage at -20, -80 or -150 degrees C. However, at elevated temperature, i.e. 4 and 37 degrees C, improved stability of CTX was observed in EDTA plasma samples compared to serum. CONCLUSIONS: CTX is stable in frozen serum and plasma samples for up to 3 years. EDTA plasma might be the preferred matrix due to improved stability at elevated temperatures.

Oral ibandronate: changes in markers of bone turnover during adequately dosed continuous and weekly therapy and during different suboptimally dosed treatment regimens.

The aims of the present study were to investigate how changes in the cumulative dose and the frequency of dosing influence the short-term antiresorptive efficacy of oral ibandronate treatment and whether serial measurements of bone markers could provide a useful diagnostic tool for the revelation of noncompliance to established treatments with antiresorptive drugs. Study participants were 200 healthy women 50-70 years old (mean 63.1 years) with a lumbar spine BMD t-score of -1 to -5. Women were randomly allocated to receive treatment with oral ibandronate according to one of the following eight dosing regimes: (1) 2.5 mg daily for 84 days; (2) 20 mg weekly for 84 days; (3) 2.5 mg daily for 28 days + no treatment for 56 days; (4) 2.5 mg daily for 28 days + 2.5 mg weekly for 56 days; (5) 2.5 mg daily for 28 days + 2.5 mg three times weekly for 56 days; (6) 2.5 mg daily for 14 days + 2.5 mg three times weekly for 56 days; (7) 2.5 mg three times weekly for 84 days; (8) no treatment for 168 days. Study parameters were the serum concentration of the C-terminal telopeptide of collagen type I (s-CTX, resorption marker) and N-MID osteocalcin (formation marker) measured by enzyme-linked immunosorbent assay. Oral treatment with ibandronate 20 mg weekly (cumulative dose 240 mg) resulted in greater final inhibition in s-CTX and area under the curve (AUC) compared to the 2.5 mg daily treatment (cumulative dose 210 mg), indicating that as long as optimal doses are administered the frequency of dosing has secondary importance for overall efficacy. When the cumulative dose was 130 mg or less, the final degree of inhibition was still the function of the cumulative dose, but the overall efficacy estimated by the AUC was also under the influence of the frequency of dosing. These observations suggest that serial measurements of s-CTX may provide a useful diagnostic tool for the early revelation of suboptimal dosing or noncompliance to already optimized therapies with antiresorptive agents.

Dose dependent effects on bone resorption and formation of intermittently administered intravenous ibandronate.

The aim of the present paper was to delineate in detail the dose-dependent effects of intermittent intravenous (IV) ibandronate treatment on the dynamics of markers of bone resorption and formation. The study included 73 healthy postmenopausal women between 50 and 70 years of age. Two groups received an IV injection of either 1 mg or 2 mg ibandronate on day 0 and 84 and one group, which received no treatment, served as control. Study duration was 168 days. Bone turnover was estimated by measuring the serum concentration of the C-terminal collagen I telopeptide (s-CTx, bone resorption) and osteocalcin (s-OC, bone formation) at 19 consecutive time-points. Serum CTx decreased rapidly reaching a nadir 7 days after drug administration. Maximal changes from baseline in the 1 and 2 mg ibandronate groups were -81% and -90%, respectively ( P<0.001). However, already 2 weeks after drug administration, s-CTx started to rise again in both treatment groups, reaching -16% and -20% by day 84, i.e. immediately before the second drug administration. In contrast, s-OC showed a slower but progressive decrease over time reaching a nadir at -35% inhibition after 5 months. On a group level, the suppression of bone resorption was greater or equal to the suppression of bone formation at all time points. However, the least significant change (LSC) analysis performed at the individual level highlighted individuals who at certain time points showed apparently greater suppression of formation than resorption, which could also contribute to the inefficacy of this dosing regime. Although the physiological relevance of this latter finding would require further analysis, the results draw attention to the need to optimize the intermittent IV dosing of ibandronate in order to approximate more closely the sustained and balanced anti-resorptive effect provided by daily oral treatment.

Effect of feeding on bone turnover markers and its impact on biological variability of measurements.

Bone turnover markers are subject to day-to-day and within-day variability, which may influence clinical interpretation. We examined the effect of fasting vs. feeding on the concentration and between-day variability of several markers. Twenty healthy premenopausal women were studied on 10 consecutive weekdays. Subjects were studied either in the fasting (no breakfast) or fed (breakfast at 08:00 h) state on alternate days, and were randomized to begin either fasting or fed. Two hour urine collections were obtained each day between 08:00 h and 10:00 h, and blood samples were collected daily at 09:00 h. The N-telopeptide cross-link of type I collagen in urine (uNTX) and serum (sNTX), the C-telopeptide in urine (uCTX) and serum (sbetaCTX), and immunoreactive free deoxypyridinoline (uifDPD) in urine were measured as resorption markers. Procollagen type I N-terminal propeptide (PINP), osteocalcin (OC), and bone alkaline phosphatase (bone ALP) were measured as formation markers. All bone formation and resorption markers were significantly lower in the fed state with the exception of bone ALP. The magnitude of the decrease ranged from 3.8 +/- 0.9% for PINP (p < 0.0001) to 17.8 +/- 2.6% (p < 0.0001) for sbetaCTX. Measurement variability was partitioned into analytical variability based on replicate assays (CV(a)) and within-subject variability (CV(i)). The CV(i) was greater (p < 0.05) for some markers in the fasting state (uifDPD, uNTX, and sNTX) but greater in the fed state for other markers (OC and sbetaCTX). In conclusion, the clinical impact of feeding vs. fasting is small with the exception of sbetaCTX; however, in clinical practice, collection of samples in the fasting state may be necessary to minimize the unpredictable effects of feeding. The mechanism of the acute effect of feeding on bone turnover remains uncertain.

Cerebrospinal fluid myelin basic protein immunoreactivity as an indicator of brain damage in children.

Cerebrospinal fluid (CSF) myelin basic protein (MBP) was measured blind by double antibody competitive inhibition radioimmunoassay (RIA) in 20 children who had seizures and 17 children with hydrocephalus. MBP values correlated with clinical outcome and mean maximum intracranial pressure (ICP) in the hydrocephalic group, and with type of convulsion in the epileptic group. A value of 20ng/ml or more was regarded as significantly raised. A significant rise in MBP levels could be demonstrated in those with ICP alone and in patients with additional problems, whose levels tended to be even higher. Hydrocephalic children with normal ICP and children with seizures had similar normal MBP levels, and in the latter group clinical outcome was not related to MBP levels. For individual patients CSF MBP is of little value as a prognostic indicator, or as a method of quantifying cerebral damage.

Neuropeptides in neurological tumours.

Concentrations of seven neuropeptides have been determined in 69 human neurological tumours. The majority of tumours were intrinsic to the central nervous system, being astrocytomas. In general, within the the better differentiated tumours (Grade I/II astrocytomas) higher concentrations of five neuropeptides (neuropeptide Y, somatostatin, substance P, vasoactive intestinal peptide and cholecystokinin) were measured in comparison to the poorly differentiated tumours. Of the metastatic tumours, five were derived from oat cell carcinoma of the bronchus. Very high concentrations of bombesin were identified in these metastases.

Pre- and postoperative changes in serum myelin basic protein immunoreactivity in neurosurgical patients.

In 44 patients undergoing neurosurgical procedures for intracranial tumors, subarachnoid hemorrhage, or spinal and peripheral nerve lesions, serum myelin basic protein (MBP) immunoreactivity was measured preoperatively and serially in the first 10 postoperative days. The double-antibody radioimmunoassay method was used, with a detection limit of 2.5 ng/ml in serum. Clinical evaluation was carried out at admission and on successive days during the period of neurosurgical management; outcome was assessed later. In the early postoperative phase, there was a fall in MBP immunoreactivity in all groups of patients. In the groups with intracranial tumor and subarachnoid hemorrhage, there was a subsequent rise in MBP immunoreactivity before the end of the 10-day period, which was not found in the group with spinal and peripheral nerve lesions.

Effect of electroconvulsive therapy on serum myelin basic protein immunoreactivity.

A sensitive radioimmunoassay that can detect brain damage in cases of head injury and stroke was applied to blood samples from 13 patients before and after they received multiple treatments with electroconvulsive therapy for psychiatric disorder. None of the patients showed a significant increase in serum myelin basic protein immunoreactivity. As increased serum myelin basic protein immunoreactivity may reflect myelin damage it is apparent that in these patients electroconvulsive therapy did not cause measureable breakdown of myelin.

Myelin basic protein immunoreactivity in serum of neurosurgical patients.

Patients admitted to the neurosurgical wards for the management of nervous system tumours, subarachnoid and intracerebral haemorrhage, head injury, spinal and peripheral nerve lesions, and other miscellaneous neurosurgical conditions, were studied by assay of serum immunoreactivity for myelin basic protein. Of 171 patients, 70% proved to have elevated myelin basic protein activity. In cerebral cases the extent of brain damage assessed by clinical methods appeared to correlate with the appearance of elevated serum myelin basic protein. In spinal and peripheral nerve cases no similar elevation of myelin basic protein was observed.

Pre- and post-operative changes in serum immunoglobulin level in neurosurgical patients.

46 patients treated by operative neurosurgery for spontaneous subarachnoid haemorrhage, intracranial tumour and spinal diseases were studied. Levels of circulating immunoglobulin and complement were measured pre-operatively and serially in the first 10 post-operative days. Prior to operation immunoglobulin levels in all categories of patients were distributed normally. However, immediately post-operatively IgG and IgM fell sharply. Conversely IgA rose in a marked manner and C3 levels showed a gradual increase. Later, antibody levels tended to return to normal.

Serum MBP immunoreactivity and immunoglobulin level as markers of tumour type.

Forty three patients, admitted to the department of Neurological Surgery for management of central nervous system tumours, were studied pre-operatively for serum myelin basic protein immunoreactivity as a marker of central nervous system lesion and for circulating immunoglobulins and complement (C3) levels. Myelin basic protein concentration did not appear to correlate with tumour type or grade. Serum immunoglobulin levels were found to be within the normal range but the mean IgM level was significantly higher in the glioma group when compared with meningiomas.

Alterations in the cellular immune response of patients with cerebral glioma, benign intracranial tumour, and spontaneous subarachnoid haemorrhage measured in vitro by the leucocyte migration inhibition test.

Leucocyte migration inhibition in response to ubiquitous antigens was studied in 104 patients as an in vitro indicator of cell-mediated immunity. Patients with cerebral glioma, benign intracranial tumours, and subarachnoid haemorrhage demonstrated impaired inhibition of leucocyte migration compared with control subjects. The greatest impairment occurred in patients with subarachnoid haemorrhage, while the least impairment was seen in patients with glioma. Significant rises in inhibition of leucocyte migration in response to antigen preparations from glioma and normal brain were seen in the early post-operative period in patients with glioma and subarachnoid haemorrhage. Impaired cellular immunity, together with sensitivity of lymphocytes to brain-derived antigens, are features of cerebral disease in general and not specific for glioma.