Risk factors for early implant-related surgical site infection.

PURPOSE: To identify the risk factors and microbes associated with early implant-related surgical site infection (SSI). METHODS: Records of 193 implant-related SSIs secondary to primary orthopaedic surgery were reviewed. Early and late SSI was defined as infection diagnosed within and after 3 months of surgery, respectively. RESULTS: Of the 193 implant-related SSIs, 29 were superficial incisional, 127 were deep incisional, and 37 were organ/space-related. 144 (90%) out of 160 SSIs used cefazolin in their prophylactic antibiotic regimen. In univariate analysis, early SSI was associated with diabetes mellitus, American Society of Anesthesiologists (ASA) score of >2, emergency procedures, and lack of antibiotic prophylaxis. In multivariable analysis, early SSI was associated with an ASA score of >2 (p=0.016). CONCLUSION: It is important to cross-check ASA score with co-morbidities to reduce early SSIs. Peri-operative optimisation and antibiotic prophylaxis should be administered prior to surgery. Appropriate modification of antibiotic prophylaxis should be considered.

Ginsenoside-Rb1 promotes adipogenesis through regulation of PPARγ and microRNA-27b.

Ginsenoside-Rb1 (Rb1), one of the bioactive components in ginseng extract, is recently reported to be able to promote adipogenesis and peroxisome proliferator-activated receptor gamma (PPARγ) expression. Meanwhile, microRNA-27b (miR-27b) is also identified to regulate adipogenesis by targeting PPARγ2. In the present study, we attempted to link up the Rb1-promoted adipogenesis with PPARγ binding and miR-27b regulation. First, we demonstrated that GW9662, an antagonist of PPARγ, could block Rb1-induced 3T3-L1 differentiation with little toxicity towards cell proliferation. Then, expression levels for both of miR-27b and its primary transcript, pri-mir-27b, were found to decrease upon Rb1 treatment. Again, GW9662 could attenuate the inhibitory effect of Rb1 on both miR-27 and pri-mir-27b expression. Since Rb1 was demonstrated to have binding activity towards PPARγ, we thus speculate that Rb1 may act though PPARγ to downregulate mir-27b gene transcription and mature miR-27b activity, which in turn promotes PPARγ expression and adipogenesis. Enhancement on adipogenesis of adipose tissues is expected to prevent lipotoxicty in nonadipose tissues. Our data may give a better illustration to explain the antidiabetic effect of Rb1 and provide a hint on treatment of lipid related metabolic diseases in the future.

Rapid detection of influenza A virus in clinical samples using an ion channel switch biosensor.

This report describes the fabrication and successful use of the ion channel switch biosensor (ICSB) for rapid point-of-care detection of influenza A in different types of respiratory specimens. Virus culture -- regarded as the "gold standard" -- and an immunochromatographic rapid point-of-care test for influenza A virus were compared with the biosensor. The ICSB rapid test provided an objective readout within 10 min of specimen inoculation into the ICSB chamber wells, without the need for chemical or other pretreatments. Construction of the ICSB with specific antibodies also enables rapid detection and identification of appropriate influenza A subtypes.

The anti-angiogenic effect of sinomenine.

Sinomenine is an alkaloid extracted from the Chinese medicinal plant, Sinomenium acutum, which has been utilized to treat rheumatoid arthritis (RA) in China for over 2000 years. Sinomenine has been shown to mediate a wide range of pharmacological actions which includes anti-inflammatory and anti-rheumatic effects. RA has been classified as a chronic immune-mediated disease that exhibits overlapping manifestation of inflammatory, abnormal cellular and hormonal immune responses with synovial hyperplasia. Since, angiogenesis is recognized to play a critical role in the development of RA and anti-angiogenic therapy has been proposed as a new therapeutic strategy for treatment of RA, we would like to see if sinomenine possesses anti-angiogenic property. In this study, sinomenine inhibited bFGF-induced proliferation of human umbilical vein endothelial cells (HUVEC) and arrested its cell cycle in G1 phase. Sinomenine disrupted tube formation of HUVEC on Matrigel and suppressed the chemotaxis of HUVEC. In addition, sinomenine reduced neovascularization in Matrigel plug assay as well as microvascular outgrowth in rat aorta ring sprouting assay. These results suggest that sinomenine inhibited bFGF-induced angiogenesis in vitro and in vivo. As the leukocytes-endothelial adhesive interactions also play an important role in inflammation, we found that sinomenine reduced the transmigration of granulocytic differentiated HL60 cells across IL-1beta activated HUVEC monolayer. Therefore, the inhibition of leukocytes migration across blood vessel walls and the anti-angiogenic effect of sinomenine may contribute towards its therapeutic mechanisms in alleviating the pathogenesis of RA.

The mudskipper, Periophthalmodon schlosseri, actively transports NH4+ against a concentration gradient.

Periophthalmodon schlosseri can maintain ammonia excretion rates and low levels of ammonia in its tissues when exposed to 8 and 30 mM NH4Cl, but tissue ammonia levels rise when the fish is exposed to 100 mM NH4Cl in 50% seawater. Because the transepithelial potential is not high enough to maintain the NH4+ concentration gradient between blood and water, ammonia excretion under such a condition would appear to be active. Branchial Na+-K+-ATPase activity is very high and can be activated by physiological levels of NH4+ instead of K+. Ammonia excretion by the fish against a concentration gradient is inhibited by the addition of ouabain and amiloride to the external medium. It is concluded that Na+-K+-ATPase and an Na+/H+ exchanger may be involved in the active excretion of ammonia across the gills. This unique ability of P. schlosseri to actively excrete ammonia is related to the special structure of its gills and allows the fish to continue to excrete ammonia while air exposed or in its burrow.

Fine structure of the gill epithelium of the terrestrial mudskipper, Periophthalmodon schlosseri.

In this paper, we describe the fine structure of the branchial epithelium of the amphibious, air-breathing mudskipper Periophthalmodon schlosseri, and relate the observed structure to functions in gas exchange, and to the elimination of sodium chloride and ammonia. Also, we describe the fine structure of the opercular epithelimicrom. The gill lamellar epithelium is thickened by the presence of large mitochondria-rich (MR) cells. These MR cells are further characterized by an extensive tubular system that is continuous with the basolateral plasma membrane and by a deep apical crypt often lined with microvilli. There are very few specialized MR accessory cells, which are associated with NaCl excretion in marine teleosts. Instead, MR cells are commonly isolated from each other laterally by flattened cells rich in intermediate filaments. These filament-rich (FR) cells are interconnected by desmosomes and have unusual canaliculi. These branchial FR cells are unique to P. schlosseri and may have a structural role. Electron-dense pavement cells rich in vesicles and large vacuous mitochondria compose the superficial layer of the epithelium. The unusual morphology of P. schlosseri's gill lamellae may be related to the animal's ability to effectively eliminate ammonia during air exposure. The inner opercular lining and parts of the leading edge of the filament have intraepithelial capillaries, which provide a more suitable gas exchange surface than the thickened lamellae with its restricted interlamellar water spaces. The arrangement of respiratory and ion exchange epithelia is opposite to that found in all other fish in which the lamellae typically function in gas exchange and the gill filament in ion regulation.

Further characterization of HIV RNA synthesis early after cell-to-cell transmission infection.

Using a one-step model for cell-to-cell transmission of HIV infection we have identified two distinct phases of HIV RNA synthesis. The first phase (4 h-12 h p.i.) was marked by an increase in only the full-length 9 kb RNA, while the second phase (24 h p.i. onwards) comprised a significant increase in the levels of all three species of viral RNA. We now report that while the continual presence of actinomycin D at 50 micrograms/ml abolished all detectable viral nucleic acid synthesis when virus donor H3B cells were pre-treated with 50 micrograms/ml of actinomycin D (AmD), washed free of unbound drug (a procedure which inhibited > 99% of total cellular RNA transcription) then mixed with untreated recipient Hut 78 cells, normal amounts of full length linear unintegrated viral DNA were produced and the first phase of viral RNA transcription was unaffected. Similarly, when both the virus donor cells and recipient cells were arrested in the late G1 phase of the cell cycle by aphidicolin and then mixed, linear unintegrated viral DNA was the major viral DNA species produced. The appearance of circular viral DNA and progeny virus was inhibited, but the first phase of induced viral RNA synthesis was unaffected. When AZT was added at 2 h or 4 h after cell-cell mixing, the level of 9 kb RNA detected was significantly lower, corresponding to reduction in the level of viral DNA. These and previous results indicate that the template for the first phase of viral RNA synthesis was likely to be newly synthesized, linear unintegrated viral DNA and not pre-existing proviral DNA present in the donor cells. Taken together, our results suggest that there exists a yet to be fully characterized pathway of concurrent viral DNA and RNA synthesis early after cell to cell transmission of HIV infection.

Comparison of rhabdomyosarcoma, buffalo green monkey kidney epithelial, A549 (human lung epithelial) cells and human embryonic lung fibroblasts for isolation of enteroviruses from clinical samples.

BACKGROUND: Traditionally, primary monkey kidney (PMK) epithelial cells have been one of the more widely used cell types for the isolation of enteroviruses from clinical samples. For the isolation of coxsackie A viruses, intraperitoneal inoculation of newborn mice has been used in some laboratories. OBJECTIVE: With the discontinued availability of PMK epithelial cells and the reported growth of coxsackie A viruses in rhabdomyosarcoma cells (RD), we compared the use of the latter cell line with our routinely used microwell cell cultures. STUDY DESIGN: Microwell cell cultures of buffalo green monkey epithelial cell line (BGM), human lung carcinoma epithelial cell line (A549) and human embryonic lung (HEL) fibroblasts were compared with RD cell line for the isolation of enteroviruses from clinical samples. RESULTS: A total of 39 enteroviruses was isolated from 3517 specimens. The HEL fibroblasts yielded 28 (72%) enteroviruses, followed by A549 (25 isolates, 64%), BGM (23 isolates, 59%) and RD cells (18 isolates, 46%). CONCLUSIONS: All isolates which grew in RD cells showed specific cytopathic effects in one or more of the other inoculated cell cultures. Quantitative determinations (TCID50) with five different enteroviruses showed that the HEL fibroblasts and RD cell line to be the most sensitive cell types, followed by BGM and A549 cell lines. However, integrity of the inoculated cell monolayers was best with HEL fibroblasts and A549 cells but morphology was not optimal with RD cells during the incubation period of 14 days.

AZT blocks down-regulation of IL-2 and IFN-gamma gene expression in HIV acutely infected cells.

HIV infection causes dysregulation of cytokine gene expression in CD4+ T cells of the infected host. Azidothymidine (AZT) inhibits HIV replication by blocking reverse transcription. Using a one-stop cell-to-cell HIV infection model, we have investigated the expression of several key cytokines in HIV infected T cells in the absence or presence of AZT treatment. Acute HIV infection of T cells resulted in dramatic down regulation of the expression of IL-2 and INF-gamma mRNA. While beta-actin mRNA levels remained constant in both AZT-free and AZT treated cultures after HIV infection, it was found that AZT blocked the down regulation of IL-2 mRNA and INF-gamma mRNA in CD4+ T cells acutely infected with HIV.

Recognition of adenovirus types in faecal samples by southern hybridization in South Australia.

The distribution of adenovirus types in faecal samples of patients with suspected viral gastroenteritis from South Australia was determined during the 12-month period, July 1991-June 1992. There were 3299 samples tested and 226 (6.9%) were positive for adenovirus by enzyme immunoassay. Of these 226 samples, 154 (68%) were typed directly using virus DNA extracted from the faecal samples according to the Sma I, Hind III and BstE II restriction patterns and Southern hybridization analysis with pooled viral genomic DNA probes. In this group, 86% of the samples were from patients who were < 3 years of age. Enteric adenovirus types 40 and 41 accounted for 20 and 40% respectively, of these samples, and types 1, 2, 3, 5, 6, 7 and 31 comprised the remainder. Type 40 was detected mainly in the winter and spring periods, and type 41 predominated in the autumn period. The majority of the non-enteric types were found during the late winter and spring periods.

Experience with newer techniques for the laboratory detection of Mycoplasma pneumoniae infection: Adelaide, 1978-1992.

Efforts to improve laboratory diagnostic methods for infection due to Mycoplasma pneumoniae have involved the use of a cell-sheet culture method and a modified indirect hemagglutination method for IgM antibody, while direct detection of mycoplasma has employed antigen capture-enzyme immunoassay (Ag-EIA) and polymerase chain reaction (PCR) amplification of sequences within the P1 and 16S ribosomal RNA genes and quantification of the amplified DNA by dot blot hybridization (DBH). Cell-sheet culture was slightly more sensitive and more rapid than culture with cell-free diphasic medium. Indirect hemagglutination detection of IgM antibody to M. pneumoniae was more sensitive than CF and EIA for detection of IgM antibody to mycoplasma. Ag-EIA gave a rapid and reasonably sensitive indication of infection and correlated well with a serological response of patients indicating a current infection. PCR-DBH was a highly sensitive substitute for culture of mycoplasma. Both Ag-EIA and PCR-DBH require confirmation by assessment of serological response to verify that the infection is current and that positive results of PCR-DBH, in particular, are not the result of continuing carriage of the organism from a previous infection, unrelated to the current episode under investigation.

Urine and the laboratory diagnosis of Chlamydia trachomatis in males.

OBJECTIVE: To determine whether the use of urine samples from male patients can replace urethral swabs for the rapid detection of Chlamydia trachomatis by the Pharmacia EIA. SETTING: The STD clinic, Adelaide, South Australia. PATIENTS: There were two separate groups of male patients. Group A (398) patients provided urethral specimens for the EIA and culture tests. The patients in Group B (356) provided an urethral swab and a urine sample for the EIA test. METHODS: The urine samples and urethral swabs were tested for the presence of C trachomatis by the Pharmacia Chlamydia EIA. In addition, the urethral swabs from Group A patients were cultured for the organism by standard cell cultures. The infected cell cultures were identified by an immunofluorescence test using a FITC-monoclonal antibody to C trachomatis (Kallestad). RESULTS: When the EIA was validated against culture, it showed a sensitivity of 100% and a specificity of 95% with the urethral swabs from Group A patients. The urine specimens were positive in 24% of those patients who yielded a positive EIA result in the urethral swabs. CONCLUSIONS: Although the EIA test on urethral swabs showed high sensitivity and specificity when validated against culture, our results showed that the use of urine samples cannot replace urethral swabs for the laboratory diagnosis of this sexually transmitted disease.

Laboratory diagnosis of Mycoplasma pneumoniae infection. 4. Antigen capture and PCR-gene amplification for detection of the Mycoplasma: problems of clinical correlation.

Direct detection assays for Mycoplasma pneumoniae were established by PCR amplification of short sequences within the foot protein/adhesin (P1) gene and the 16S ribosomal RNA gene. Specificity and sensitivity was excellent, no hybridization was observed with M. genitalium and other human Mycoplasma species. In nose and throat washings from subjects with respiratory infection a pattern of high counts (c.f.u./ml) of M. pneumoniae (deduced from the amount of amplified PCR product), and a positive antigen capture assay, was found in 83% of subjects with serological evidence of current infection with M. pneumoniae. A small proportion of subjects with serological patterns suggesting infection in the more distant past had positive PCR assays. This was considered to represent either persistence of the organism from a previous infection or perhaps transient carriage during a reinfection, without substantial change in antibody response. PCR-based assay of M. pneumoniae offers a powerful, rapid, and sensitive substitute for culture of the mycoplasma. Antigen capture, while less sensitive than PCR, offers the advantage that it is more often positive with samples from current infection and requires less stringent laboratory organization to contain false positive results. We conclude however that the laboratory diagnosis of a chosen clinical episode should not rest on the PCR or Ag-EIA assays alone, but must also include antibody assays to confirm whether infection is current or represents persistence from past exposure.

A survey of nosocomial respiratory viral infections in a children's hospital: occult respiratory infection in patients admitted during an epidemic season.

OBJECTIVE: To define the extent of shedding of respiratory viruses and Mycoplasma pneumoniae among a population of pediatric patients admitted to the hospital during a winter epidemic period and to identify nosocomial infections within this population. DESIGN: An open, prospective survey of patients admitted to three wards (General Medical, Respiratory Infectious, and Infectious Diseases) of a pediatric hospital during a defined three-month period. PATIENTS: All patients with medical, respiratory, and infectious conditions admitted to three wards of the Adelaide Children's Hospital had nasopharyngeal aspirations performed at the time of admission with the purpose of documenting viral and M pneumoniae shedding. Patients were monitored daily for the development of symptoms of respiratory infection or new symptoms of respiratory disease. Such patients underwent a further nasopharyngeal aspiration for the purpose of diagnosing hospital-acquired infection. RESULTS: Nasopharyngeal aspirations were obtained from 601 patients. Forty-seven percent of asymptomatic patients were positive for a respiratory virus or M pneumoniae, and 61% of patients with respiratory symptoms were also positive. Gastroenteritis patients shed viruses in 66% of cases. Respiratory symptoms were initially overlooked by admitting physicians but subsequently identified in 110 cases, and 46% of these were found to be positive for a respiratory virus or M pneumoniae. There were 18 possible hospital acquired infections among the 293 initially virus-negative patients. Multiple isolates were obtained from a substantial number of patients, especially those with respiratory symptoms. CONCLUSIONS: A substantial proportion of all patients admitted to a pediatric hospital during winter represent a potential source of infection, and strict infection control measures should be enacted to limit the spread of these infections.

Laboratory diagnosis of Mycoplasma pneumoniae infection. 3. Detection of IgM antibodies to M. pneumoniae by a modified indirect haemagglutination test.

The indirect haemagglutination (IHA) test was compared with the complement-fixation (CF) test for the measurement of antibodies to Mycoplasma pneumoniae. A modification of the IHA was used to measure M. pneumoniae IgM antibodies. Sera were obtained from various groups of patients who were either culture or antigen positive for M. pneumoniae in nasopharyngeal aspirates or who had fourfold or greater increase in CF antibody or a titre greater than or equal to 320. The results of these comparisons showed that the modified IHA test was specific and more sensitive (89% as opposed to 64%) than the CF test. The modified IHA test for the detection of IgM antibody was highly effective in the recognition of recent or current infection with the mycoplasma. It was also of equal sensitivity to an indirect enzyme immunoassay for the detection of IgM antibodies to M. pneumoniae.

Comparison of five enzyme immunoassays, electron microscopy, and latex agglutination for detection of rotavirus in fecal specimens.

Five different enzyme immunoassays, electron microscopy, and latex agglutination (Slidex; bioMerieux) were compared for the rapid detection of human rotavirus in fecal specimens. The enzyme immunoassay using rotavirus polyclonal antiserum (Dakopatts) with simple in-house modifications was shown by the use of confirmatory tests to be the most sensitive and specific procedure.

Laboratory diagnosis of Mycoplasma pneumoniae infection. 1. Direct detection of antigen in respiratory exudates by enzyme immunoassay.

Direct and indirect antigen capture enzyme immunoassays (Ag-EIA) have been developed for the detection of Mycoplasma pneumoniae in nasopharyngeal aspirates or sputum from respiratory infection. The sensitivity of the two Ag-EIA were similar, but the indirect method using polyclonal rabbit and guinea-pig antisera was more convenient. The Ag-EIA had a detection limit of 10(4-4.5) colony-forming units/ml of sample. It was specific for M. pneumoniae and gave a low level response with M. genitalium. There were no cross-reactions with 10 other species of mycoplasmas. Tests with a wide range of bacteria and chlamydia group antigen, representing agents sometimes found in the respiratory tract, were also negative. At the current level of development, the Ag-EIA detected about 90% of specimens that were also positive for culture; 43% of specimens from culture-negative--seropositive patients gave a positive result. The overall pattern of results indicated that while antigen detection is a quick and effective substitute for the slow culture method, serological examination for specific IgM antibody is also necessary to give a complete diagnostic coverage.

Mycoplasma pneumoniae: acute illness, antibiotics, and subsequent pulmonary function.

One hundred and eight children presenting with Mycoplasma pneumoniae infection were assessed during the acute illness and followed for three years. The incidence of wheezing with the acute infection (40%) was greater than expected in a normal childhood population. The initial illness precipitated wheezing for the first time in some subjects but others wheezed only with the acute illness. In non-asthmatic subjects significant bronchodilator responsiveness was present one month after infection. Children given erythromycin during the first seven days of their illness had a significantly shorter fever duration compared with those treated inappropriately. No significant effects of treatment were noted on pulmonary function three years later but non-asthmatic children had abnormal mean forced expiratory volume in one second and forced expiratory flow after 50% of the expired vital capacity compared with 64 healthy controls. These findings indicate impaired function three years after initial infection.