Gross synovial fluid analysis in the differential diagnosis of joint effusion.

AIMS: To develop an objective and easy to complete standardised questionnaire for documentation of synovial fluid (SF) gross appearance and use it in the assessment of patients presenting to the rheumatology service with a joint effusion. METHODS: A standardised questionnaire to record the gross appearance of SF was developed. Interobserver error in recorded observations and direct gross analysis of synovial fluid between four observers was calculated in a pilot study. In a prospective study over 8 months, SF gross analysis and cell count were documented in all patients presenting with a joint effusion. Fusch Rosenthal manual counting chamber was used for calculating SF cell counts. RESULTS: There was good interobserver agreement on direct gross analysis and between questionnaire assessors (mean kappa 0.889). 80 SF samples were collected. Gross analysis was performed in all samples and cell count in 72. Of the specimens thought to be inflammatory on gross analysis, 31% were found to be non-inflammatory based on cell count; however, 12 of these patients had an established inflammatory arthritis. Gross analysis had a sensitivity of 94% and specificity of 58% when used to determine whether SF is inflammatory or non-inflammatory. The positive and negative predictive values were 0.69 and 0.91 respectively. CONCLUSIONS: SF cell count did not add any information when SF gross analysis suggested a non-inflammatory process. Gross analysis was better than cell count to determine a potentially septic joint fluid. Further work needs to be done on the value of SF cell counts if gross analysis suggests the fluid to be inflammatory.

Stability of premixed syringes of diamorphine and hyperbaric bupivacaine.

BACKGROUND: It is common clinical practice to add diamorphine to heavy bupivacaine when performing spinal anaesthesia for either obstetric or general surgical procedures. If pre-filled syringes were available potential problems arising due to the wrong mixture being administered could be reduced, whilst also providing greater assurances of sterility and accuracy of dosage. It is therefore necessary to establish whether diamorphine 100 microg/mL is stable in solution with 0.5% hyperbaric bupivacaine, to allow production of pre-filled syringes for use in spinal anaesthesia. METHOD: Diamorphine hydrochloride was dissolved in water for injection, and added to hyperbaric bupivacaine then stored in 5-mL plastic syringes. Eleven syringes were stored at 40 degrees C/75% relative humidity, 25 degrees C/60% relative humidity and 7 degrees C for 90 days. Samples were taken at five time points for measurement of diamorphine and bupivacaine concentrations using high performance liquid chromatography. RESULTS: Diamorphine concentrations fell over the study period. No significant changes were observed the bupivacaine content of the samples. There was 10% degradation of diamorphine after 4 days at 40 degrees C, after 7 days at 25 degrees C, and after 26 days at 7 degrees C. CONCLUSION: Diamorphine is stable in hyperbaric bupivacaine at 7 degrees C for long enough to allow preparation of pre-filled syringes in advance (by hospital pharmacy aseptic units) for use in spinal anaesthesia.

Rapid decompression in the EA-6B.

A Grumman EA-6B aircraft experienced a rapid pressurization failure at 27,000 feet. All four crew members had removed their oxygen masks and were breathing cabin air pressurized to 8,000 feet before the incident. Although none of the crew members developed signs or symptoms of decompression sickness, the potential for adversity was realized by all. Altitude decompression sickness (DCS) and pulmonary overinflation syndrome (POIS) represent potentially fatal complications of rapid decompression or uncontrolled ascent in aircraft. The signs and symptoms of DCS range from mild joint pain to eventual cardiopulmonary collapse and death. The symptoms of POIS are usually more abrupt and lethal. The medical management of DCS and POIS includes (1) maintenance of airway and cardiopulmonary resuscitation if necessary: (2) administration of 100% oxygen; (3) descent as per Naval Aviation Training and Operating Procedures Standardization guidelines; (4) horizontal body position; (5) maintenance of fluid intake; and (6) early medical evaluation by a flight surgeon or other physician qualified in the management of DCS. Symptoms of DCS may appear up to 24 hours after decompression, and continued monitoring or grounding of exposed individuals during this time is essential. Many controllable factors may predispose to DCS/POIS, and preventive measures should be taken to ensure maximum reduction of risk.

Eye injuries from laser exposure: a review.

Lasers pose a significant threat to vision in modern military operations. Anti-personnel lasers have been designed that can cause intentional blindness in large numbers of personnel. Although the use of blinding laser weapons during combat has been prohibited by international legislation, research and development of these weapons have not been prohibited, and significant controversy remains. Unintentional blinding can also result from other types of lasers used on the battlefield, such as range-finders and anti-material lasers. Lasers that are capable of producing blindness operate within specific wavelength parameters and include visible and near infrared lasers. Patients who suffer from laser eye injuries usually complain of flash blindness, followed by transient or permanent visual loss. Laser retinal damage should be suspected in any patient with visual complaints in an operational setting. The treatment for laser retinal injuries is extremely limited, and prevention is essential. Improved protective eyeware and other countermeasures to laser eye injury are necessary as long as the threat remains.

The multi-epitope approach for immunotherapy for cancer: identification of several CTL epitopes from various tumor-associated antigens expressed on solid epithelial tumors.

One approach to development of specific cancer immunotherapy relies on the induction of cytotoxic T lymphocytes (CTL) specific for tumor-associated antigens (TAA). Induction of TAA-specific CTL could be used towards the eradication of established tumors, or to prevent their dissemination or recurrence after primary treatment. The present study identifies a set of CTL epitopes from TAA frequently found on solid epithelial tumors such as breast, lung and gastro-intestinal tumors. Specifically, HLA-A2.1 binding peptides from the MAGE2, MAGE3, HER-2/neu and CEA antigens were tested for their capacity to elicit in vitro anti-tumor CTL using lymphocytes from normal volunteers and autologous dendritic cells as antigen-presenting cells. A total of 6 new epitopes (MAGE2[10(157)], MAGE3[9(112)], CEA[9(691)], CEA[9(24)], HER2[9(435)] and HER2[9(5)]) were identified which were capable of specifically recognizing tumor cell lines lines expressing HLA-A2.1 and the corresponding TAA. In one case (CEA[9(24)]), induction of vigorous anti-tumor CTL responses required epitope engineering to increase HLA-A2.1 binding affinity. Finally, most of the newly identified epitopes (5 out of 6) were found to be highly crossreactive with other common HLA alleles of the A2 supertype (A2.2, A2.3, A2.6 and A6802), thus demonstrating their potential in providing broad and non-ethnically biased population coverage. The results are discussed in the context of the development of multi-epitope-based therapies with broad applicability for patients suffering from commonly found tumors.

Experimental pseudomonal posttraumatic endophthalmitis in a swine model. Treatment with ceftazidime, amikacin, and imipenem.

PURPOSE: To compare the intravitreal efficacy of three separately administered antibiotics (imipenem, ceftazidime, and amikacin) in limiting the intraocular inflammation and tissue destruction caused by posttraumatic pseudomonal endophthalmitis. METHODS: Thirty-three Yorkshire pigs each received a surgically induced scleral injury to the right eye. After repair, each eye was injected with 22,000 colony-forming units of live Pseudomonas aeruginosa. Pigs then were randomly grouped into a natural-history-of-infection group in which no treatment was given (n = 9) or into groups treated with the following: intravitreal imipenem (n = 6), ceftazidime (n = 6), amikacin (n = 6), or normal saline (n = 6). Pigs then were observed clinically for 18-24 hours after surgery and enucleated for histopathologic examination. RESULTS: Clinical examinations revealed significantly less posterior segment inflammation in pigs treated with amikacin and imipenem than in pigs in the natural history or saline control groups, based on the Wilcoxon rank sum test (P < .05). Histopathologic examinations showed similar results, with less intraocular inflammation and retinal destruction in pigs treated with amikacin and imipenem, whereas the inflammation in pigs treated with ceftazidime did not differ significantly from that in control pigs. CONCLUSION: Intravitreal antibiotic treatment with imipenem or amikacin appears to limit intraocular inflammation and retinal tissue damage when given early in the course of posttraumatic pseudomonal endopthalmitis. Results with ceftazidime are less conclusive.

Intracellular signaling of tumor necrosis factor-alpha in brain microvascular endothelial cells is mediated by a protein tyrosine kinase and protein kinase C-dependent pathway.

The intracellular signaling pathways responsible for tumor necrosis factor (TNF)-alpha stimulation of lymphocyte adhesion to brain microvascular endothelial cells (BMEC) were studied using inhibitors of protein kinase C (bisindolylmaleimide HCl, H-7, or staurosporine), or protein tyrosine kinase (genistein). Each of these blocked the ability of BMEC to respond to TNF-alpha. In contrast, BMEC treated with H-89, an inhibitor of protein kinase A, or the adenylate cyclase inhibitor, dideoxyadenosine, responded normally to TNF-alpha. Forskolin, an adenylate cyclase agonist, significantly increased lymphocyte adhesion to BMEC. These data indicate that intracellular signaling by TNF-alpha in BMEC is mediated through a protein kinase C and tyrosine kinase dependent pathway.

The effect of trauma on the ocular penetration of intravenous ciprofloxacin.

PURPOSE: To study the intraocular pharmacokinetics of intravenously administered ciprofloxacin following eye trauma. METHODS: Twenty-three New Zealand albino rabbits and 12 Yorkshire pigs each received a surgically induced scleral injury to the right eye. Following repair, each rabbit received a single 30-mg intravenous infusion of ciprofloxacin. Each pig received either two 200-mg doses or two 400-mg doses of intravenous ciprofloxacin given 12 hours apart. Vitreous and serum samples were harvested at 0.5, 1, 4, 6, and 12 hours after antibiotic administration in rabbits and 1 hour after the second dose in pigs. Bioassays for ciprofloxacin were performed on each sample, and results were statistically compared by t test. The untraumatized left eye in each animal served as a control. RESULTS: The mean vitreous concentration of ciprofloxacin in traumatized rabbit eyes was 0.37 microgram/ml. This level was sustained above levels in control eyes (0.18 microgram/ml) for at least 4 hours following antibiotic administration. In control eyes, intravitreal levels peaked at 1 hour. Mean vitreous concentrations +/- SD in traumatized pig eyes were 0.091 +/- 0.017 microgram/ml in swine that had received 200-mg doses of ciprofloxacin vs 0.312 +/- 0.153 microgram/ml in swine that had received 400-mg doses (P = .02). Mean vitreous concentrations of ciprofloxacin in control eyes were not affected by increasing dosage. CONCLUSION: In both animal models, experimental surgical trauma increased intravitreal ciprofloxacin delivery. In addition, systemically administered ciprofloxacin achieved intravitreous levels exceeding minimum inhibitory concentrations for common ocular pathogens, suggesting a role for ciprofloxacin in the prophylaxis of posttraumatic endophthalmitis.

Experimental posttraumatic Bacillus cereus endophthalmitis in a swine model. Efficacy of intravitreal ciprofloxacin, vancomycin, and imipenem.

PURPOSE: The authors compare the intravitreal efficacy of ciprofloxacin, vancomycin and imipenem, in treating experimental Bacillus cereus endophthalmitis. METHODS: Thirty-three Yorkshire pigs received a surgically induced injury to the right eye, which was then repaired and injected with 8400 colony forming units of live B. cereus. Nine pigs received no therapy and served as a natural history group. Twenty-four other pigs then were randomized into a treatment group with ciprofloxacin (n = 6), vancomycin (n = 6), imipenem (n = 6), or normal saline (n = 6). Eyes were examined clinically 1, 2, 3, 4, 5, 6, 8, 12, and 24 hours after inoculation. After 24 hours, the eyes were enucleated for histologic study. RESULTS: Experimental disease was characterized by an aggressively developing endophthalmitis, with retinitis and vitritis developing at 4 hours. Histologic examination showed vitreous abscess and retinal necrosis. Both vancomycin- and imipenem-treated group had less inflammation and tissue destruction than control animals, based on the Wilcoxon rank sum test (P < 0.05). Ciprofloxacin-treated animals showed significantly more intraocular destruction and were indistinguishable from controls. CONCLUSION: Vancomycin and imipenem appear to limit inflammation and tissue destruction when given early in the course of experimental posttraumatic endophthalmitis caused by B. cereus. Results with ciprofloxacin are less conclusive and warrant further investigation.

Functional cytotoxic T cells are associated with focal lesions in the brains of SJL mice with experimental herpes simplex encephalitis.

Using a recently developed murine model of herpes simplex encephalitis in SJL mice, we isolated and characterized brain-infiltrating mononuclear cells to identify immune effectors that might contribute to pathology in this disease. Brain infiltrating mononuclear cells obtained from temporal area lesions were found to be predominantly T cells, with CD4+ and CD8+ T cell types present in equal numbers. The herpes simplex virus type 1 (HSV-1)-specific CTL that were present within the temporal area of HSV-1 infected SJL mice were fully activated (i.e., they did not require in vitro culture to acquire cytotoxic function). These cells were Thy-1.2+, CD8+ and exhibited the classical features of CTL activity: Ag specificity and MHC restriction. To our knowledge this is the first definitive demonstration of HSV-1-specific CTL activity directly ex vivo in mice. Freshly isolated spleen or lymph node cells from the same mice failed to exhibit CTL activity. The lytic capacity of infiltrating cells appeared to be mediated exclusively by CTL because treatment with Abs to Thy-1.2, or CD8, and C completely removed cytotoxic activity. NK cell activity was not detected in spleen, lymph node, or brain-infiltrating mononuclear cells from SJL mice with HSV encephalitis. Significantly, T cells were the prominent cell type in the temporal area of HSV-1-infected SJL mice just before the development of focal lesions, with CD8+ cells being present in equal or greater numbers than CD4+ cells (a CD4:CD8 ratio of 1 < or = 1). Thus, the accumulation of functional CD8+ CTL in the temporal area of the brain correlated with the occurrence of focal entorhinal lesions.

Identification of a mariner-like repetitive sequence in C. elegans.

A repetitive element in C.elegans has been found that bears high homology to the element mariner of Drosophila mauritiana (EMBL accession number X77804). This element is present in about 20 copies in the N2 strain of C.elegans, and appears in roughly equal copy numbers in the related strain BO and in the hybrid strains RW7097 and TR679. There is only one copy of this MLE in three related species of Caenorhabditis. A cDNA of this mariner-like element (MLE) codes for a protein with 58% homology to the Drosophila transposase. The mariner-like element is not mobile in N2. This class of elements has now been described in insects, planaria and nematodes (GenBank accession number M98552 and this report).

Crystal violet reactions of Staphylococcus aureus strains colonizing infants in the first six weeks.

Nasal colonization with Staphylococcus aureus occurred in 18% of babies leaving a maternity unit and had risen to 40% by 6 weeks after birth. S. aureus was first acquired by 34.5% of babies after discharge. Female infants were more likely to be colonized than males. Colonization was not significantly different between babies receiving standard postnatal care and those nursed on the Special Care Baby Unit. Crystal violet (CV) tests showed that purple-reacting isolates accounted for approximately 60% of strains, whether first detected at hospital discharge or subsequently acquired. Purple-reacting strains, once acquired, were significantly better able to persist than non purple-reacting strains and formed a cumulatively higher proportion of the strains isolated at 6 weeks after birth than at hospital discharge. CV purple-reactions were significantly associated with lysis by phages of groups III and I and non-purple-reactions were significantly associated with lysis by phages of group II and/or 94/96. Maternity units remain a significant route whereby strains of S. aureus with some characteristics associated with a hospital origin gain access to the community.

Necrobacillosis with pancytopenia.

Two young children whose presentation with necrobacillosis caused considerable diagnostic difficulty resulting in referral to an oncology unit are described. In both cases their severe suppurative multisystem illness was complicated by pancytopenia. One had bone marrow infarcts and severe endocarditis in addition to pulmonary involvement and the other had osteitis which resulted in a deformed humerus.

Jackdaws and magpies as vectors of milkborne human Campylobacter infection.

In 1990 we reported that milk bottles pecked by jackdaws and magpies were a probable source of human campylobacter infection. During April to June 1990 an extended study of campylobacter infections was carried out in the Gateshead area. Prior to the study a health education programme was undertaken in an attempt to reduce human infection. Fifty-nine cases of human infection were recorded and 52 were interviewed. Thirty were entered into a case control study which demonstrated a very strong association between consumption of pecked milk and human campylobacter infection (chi 2 = 12.6, P less than 0.0004). It was estimated that between 500 and 1000 jackdaws (Corvus monedula) were present in the area where milk bottles were pecked and 63 isolates of campylobacter were made from the bill and cloaca. Target bottles were put out in the early mornings and campylobacters were isolated from 12 of 123 pecked bottles. Typing of the campylobacters revealed a wide distribution of strains amongst birds, pecked milk and human infections. The health education programme had only limited success.

Epidemiological typing of coagulase-negative staphylococci by pyrolysis mass spectrometry.

Pyrolysis mass spectrometry (PMS) was evaluated for the epidemiological typing of coagulase-negative staphylococci (CNS) in situations in which it was necessary to distinguish between repeated isolation of the same strain from a single patient (genuine infection) and coincidental isolation of unrelated strains (contamination). Thirteen CNS isolates were examined, consisting of five pairs, each pair isolated from a single patient, and three unrelated strains. PMS analysis gave results equivalent to a conventional typing system comprising antibiogram, biotype and plasmid profile analysis. Both methods facilitate differentiation between genuine infection with CNS and the isolation of contaminants. The speed, reproducibility, versatility and relatively low cost of PMS suggest that it may be a valuable new technique for the epidemiological typing of CNS in routine clinical settings.

Induction of encephalitis in SJL mice by intranasal infection with herpes simplex virus type 1: a possible model of herpes simplex encephalitis in humans.

Herpes simplex encephalitis (HSE) is characterized by focal lesions of hemorrhage and necrosis, primarily in the inferior temporal lobe. Since immunosuppressed patients with HSE lack the focal inflammatory changes and temporal lobe localization, it has been suggested that the immune system participates in the pathogenesis of HSE. Evaluation of this hypothesis has been impeded by the lack of an immunologically defined animal model that resembles the human disease. Toward this end, 10 strains of inbred mice were infected intranasally with a neurovirulent clinical isolate of herpes simplex virus type 1. Most mice died without localizing signs of disease in the central nervous system. However, a significant number of SJL mice had a pattern of encephalitis highly reminiscent of that described in humans. To our knowledge, this is the first murine model that faithfully mimics this human disease, and thus it affords the opportunity to study the immunopathogenesis of HSE.

Crystal violet reactions of fresh clinical isolates of Staphylococcus aureus from two British hospitals.

When 168 fresh clinical isolates of Staphylococcus aureus were examined for their reactions on a medium containing 1 part in 100,000 crystal violet 50.6% of strains produced a purple appearance, 39.3% produced a white appearance and 10.1% produced a yellow appearance. Purple-reacting isolates were significantly associated with both invasive infections (P less than 0.01) and hospital origin (P less than 0.001). There were no significant associations between the crystal violet reactions and either animal contact or other properties previously reported to be characteristic of white and yellow-reacting strains (beta haemolysin and bovine coagulase production). The results of phage typing showed associations between susceptibility to group III phages and purple-reacting strains and between phage group II susceptibility and white and yellow-reacting strains. There was also a highly significant association between white reactions on crystal violet agar and susceptibility to lysis by a combination of all three groups (that is, I + II + III) and white-reacting strains were significantly more susceptible to lysis by phages 94 and/or 96, whether as a restricted pattern or as part of a broader pattern. The purple reaction on crystal violet medium may be a reliable marker of the 'hospital staphylococcus'.

Pyrolysis-mass spectrometry (Py-MS) for the rapid epidemiological typing of clinically significant bacterial pathogens.

Fresh clinical isolates of Salmonella spp. and Streptococcus pyogenes were analysed by pyrolysis-mass spectrometry (Py-MS). The results formed the basis of mathematically derived characterizations of individual strains and these were compared with the results of phage typing for the salmonellas and M protein typing for the streptococci. Py-MS was shown to be a rapid and reproducible method for inter-strain comparisons, giving evidence of identity and non-identity between strains that agreed well with the results of conventional tests. Py-MS has potential value as a rapid, relatively inexpensive and highly discriminatory method of epidemiological analysis in bacterial disease.