A Multi-Disciplinary Approach for the Management of Prosthetic Joint Infections: An Australian Perspective.

Introduction: Prosthetic joint infections (PJI) are a major complication of hip and knee arthroplasty, imposing significant morbidity and mortality. Orthopaedic oncology units have utilised a multi-disciplinary team (MDT) approach for some time. PJI is not only an equally life-threatening condition, it also requires input from multiple healthcare personnel and treatment can vary significantly between individuals given the diversity in microbiological, surgical and host factors. Our arthroplasty service established an MDT meeting to manage this complex patient group. This study describes the philosophy and implementation of an MDT approach to the management of PJIs at a tertiary hospital in Australia. Materials and methods: A retrospective review of all patients that presented to the MDT PJI meeting from October 2017 to April 2020 was performed. Patient characteristics, microbiological profile and management were reviewed. Results: One hundred and one patients were reviewed over 2.5 years with a mean age of 69.2 years (SD 11.9). Patients presenting predominantly had a primary TKR (32%) or primary THR (22%). Results of Microbiology cultures varied, with 42% Gram-positive organisms, 13% Gram-negative organisms, 2% fungus and 1% yeast origin. Management mainly consisted of two-stage revision (28%), debridement-antibiotics-and-implant retention (22%) and antibiotic suppression (14%). A total of 91.5% of patients who underwent surgical management were considered cured at one year. Conclusion: PJIs are complex and require coordinated care by a number of healthcare personnel. The MDT process has allowed collaboration between Orthopaedic, Infectious Disease and Microbiology departments and aims to improve the quality of care provided to patients, potentially reducing morbidity and mortality of patients with PJI.

A Multi-Disciplinary Approach for the Management of Prosthetic Joint Infections: An Australian Perspective

@#Introduction: Prosthetic joint infections (PJI) are a major complication of hip and knee arthroplasty, imposing significant morbidity and mortality. Orthopaedic oncology units have utilised a multi-disciplinary team (MDT) approach for some time. PJI is not only an equally lifethreatening condition, it also requires input from multiple healthcare personnel and treatment can vary significantly between individuals given the diversity in microbiological, surgical and host factors. Our arthroplasty service established an MDT meeting to manage this complex patient group. This study describes the philosophy and implementation of an MDT approach to the management of PJIs at a tertiary hospital in Australia. Materials and methods: A retrospective review of all patients that presented to the MDT PJI meeting from October 2017 to April 2020 was performed. Patient characteristics, microbiological profile and management were reviewed. Results: One hundred and one patients were reviewed over 2.5 years with a mean age of 69.2 years (SD 11.9). Patients presenting predominantly had a primary TKR (32%) or primary THR (22%). Results of Microbiology cultures varied, with 42% Gram-positive organisms, 13% Gramnegative organisms, 2% fungus and 1% yeast origin. Management mainly consisted of two-stage revision (28%), debridement-antibiotics-and-implant retention (22%) and antibiotic suppression (14%). A total of 91.5% of patients who underwent surgical management were considered cured at one year. Conclusion: PJIs are complex and require coordinated care by a number of healthcare personnel. The MDT process has allowed collaboration between Orthopaedic, Infectious Disease and Microbiology departments and aims to improve the quality of care provided to patients, potentially reducing morbidity and mortality of patients with PJI.

Trichoderma from Brazilian garlic and onion crop soils and description of two new species: Trichoderma azevedoi and Trichoderma peberdyi.

Fifty four Trichoderma strains were isolated from soil samples collected from garlic and onion crops in eight different sites in Brazil and were identified using phylogenetic analysis based on combined ITS region, tef1-α, cal, act and rpb2 sequences. The genetic variability of the recovered Trichoderma species was analysed by AFLP and their phenotypic variability determined using MALDI-TOF. The strain clusters from both typing techniques coincided with the taxonomic determinations made from phylogenetic analysis. The phylogenetic analysis showed the occurrence of Trichoderma asperellum, Trichoderma asperelloides, Trichoderma afroharzianum, Trichoderma hamatum, Trichoderma lentiforme, Trichoderma koningiopsis, Trichoderma longibrachiatum and Trichoderma erinaceum, in the soil samples. We also identified and describe two new Trichoderma species, both in the harzianum clade of section Pachybasium, which we have named Trichoderma azevedoi sp. nov. and Trichoderma peberdyi sp. nov. The examined strains of both T. azevedoi (three strains) and T. peberdyi (12 strains) display significant genotypic and phenotypic variability, but form monophyletic clades with strong bootstrap and posterior probability support and are morphologically distinct from their respective most closely related species.

Afferent and efferent connections of C1 cells with spinal cord or hypothalamic projections in mice.

The axonal projections and synaptic input of the C1 adrenergic neurons of the rostral ventrolateral medulla (VLM) were examined using transgenic dopamine-beta hydroxylase Cre mice and modified rabies virus. Cre-dependent viral vectors expressing TVA (receptor for envelopeA) and rabies glycoprotein were injected into the left VLM. EnvelopeA-pseudotyped rabies-EGFP glycoprotein-deficient virus (rabies-EGFP) was injected 4-6 weeks later in either thoracic spinal cord (SC) or hypothalamus. TVA immunoreactivity was detected almost exclusively (95 %) in VLM C1 neurons. In mice with SC injections of rabies-EGFP, starter cells (expressing TVA + EGFP) were found at the rostral end of the VLM; in mice with hypothalamic injections starter C1 cells were located more caudally. C1 neurons innervating SC or hypothalamus had other terminal fields in common (e.g., dorsal vagal complex, locus coeruleus, raphe pallidus and periaqueductal gray matter). Putative inputs to C1 cells with SC or hypothalamic projections originated from the same brain regions, especially the lower brainstem reticular core from spinomedullary border to rostral pons. Putative input neurons to C1 cells were also observed in the nucleus of the solitary tract, caudal VLM, caudal spinal trigeminal nucleus, cerebellum, periaqueductal gray matter and inferior and superior colliculi. In sum, regardless of whether they innervate SC or hypothalamus, VLM C1 neurons receive input from the same general brain regions. One interpretation is that many types of somatic or internal stimuli recruit these neurons en bloc to produce a stereotyped acute stress response with sympathetic, parasympathetic, vigilance and neuroendocrine components.

Evidence that hypothalamic RFamide related peptide-3 neurones are not leptin-responsive in mice and rats.

Leptin, a permissive hormonal regulator of fertility, provides information about the body's energy reserves to the hypothalamic gonadotrophin-releasing hormone (GnRH) neuronal system that drives reproduction. Leptin does not directly act on GnRH neurones, and the neuronal pathways that it uses remain unclear. RFamide-related peptide-3 (RFRP-3) neurones project to GnRH neurones and primarily inhibit their activity. We tested whether leptin could act via RFRP-3 neurones to potentially modulate GnRH activity. First, the effects of leptin deficiency or high-fat diet-induced obesity on RFRP-3 cell numbers and gene expression were assessed in male and female mice. There was no significant difference in Rfrp mRNA levels or RFRP-3-immunoreactive cell counts in wild-type versus leptin-deficient ob/ob animals, or in low-fat versus high-fat diet fed wild-type mice. Second, the presence of leptin-induced signalling in RFRP-3 neurones was examined in male and female wild-type mice and rats. Dual label immunohistochemistry revealed leptin-induced phosphorylated signal transducer and activator of transcription-3 in close proximity to RFRP-3 neurones, although there was very little (2-13%) colocalisation and no significant differences between vehicle and leptin-treated animals. Furthermore, we were unable to detect leptin receptor mRNA in a semi-purified RFRP-3 cell preparation. Because GABA neurones form critical leptin-responsive GnRH inputs, we also determined whether RFRP-3 and GABA cells were colocalised. No such colocalisation was detected. These results support the concept that leptin has little or no effects on RFRP-3 neurones, and that these neurones are unlikely to be an important neuronal pathway for the metabolic regulation of fertility by leptin.

Synthetic versus plaster of Paris casts in the treatment of fractures of the forearm in children: a randomised trial of clinical outcomes and patient satisfaction.

Fractures of the forearm (radius or ulna or both) in children have traditionally been immobilised in plaster of Paris (POP) but synthetic cast materials are becoming more popular. There have been no randomised studies comparing the efficacy of these two materials. The aim of this study was to investigate which cast material is superior for the management of these fractures. We undertook a single-centre prospective randomised trial involving 199 patients with acute fractures of the forearm requiring general anaesthesia for reduction. Patients were randomised by sealed envelope into either a POP or synthetic group and then underwent routine closed reduction and immobilisation in a cast. The patients were reviewed at one and six weeks. A satisfaction questionnaire was completed following the removal of the cast. All clinical complications were recorded and the cast indices were calculated. There was an increase in complications in the POP group. These complications included soft areas of POP requiring revision and loss of reduction with some requiring re-manipulation. There was an increased mean padding index in the fractures that lost reduction. Synthetic casts were preferred by the patients. This study indicates that the clinical outcomes and patient satisfaction are superior using synthetic casts with no reduction in safety.

Monomorphic subtelomeric DNA in the filamentous fungus, Metarhizium anisopliae,contains a RecQ helicase-like gene.

In most filamentous fungi, telomere-associated sequences (TASs) are polymorphic, and the presence of restriction fragment length polymorphisms (RFLPs) may permit the number of chromosome ends to be estimated from the number of telomeric bands obtained by restriction digestion. Here, we describe strains of Metarhizium, Gliocladium and Paecilomyces species in which only one or a few telomeric bands of unequal intensity are detectable by Southern hybridization, indicating that interchromosomal TAS exchange occurs. We also studied an anomalous strain of Metarhizium anisopliae, which produces polymorphic telomeric bands larger than 8 kb upon digestion of genomic DNA with XhoI. In this case, the first XhoI site in from the chromosome end must lie beyond the presumed monomorphic region. Cloned telomeres from this strain comprise 18-26 TTAGGG repeats, followed at the internal end of the telomere tract by five repeats of the telomere-like sequence TAAACGCTGG. An 8.1-kb TAS clone also contains a gene for a RecQ-like helicase, designated TAH1, suggesting that this TAS is analogous to the Y' elements in yeast and the subtelomeric helicase ORFs of Ustilago maydis (UTASRecQ) and Magnaporthe grisea (TLH1). The TAS in the anomalous strain of M. anisopliae, however, appears distinct from these in that it is found at most telomeres and its predicted protein product possesses a significantly longer N-terminal region in comparison to the M. grisea and U. maydis helicases. Hybridization analyses showed that TAH1 homologues are present in all other anomalous M. anisopliae strains studied, as well as in some other polymorphic strains, where the recQ-like gene also appears to be telomere-associated.

Purification and characterization of an N-acetylglucosaminidase produced by a Trichoderma harzianum strain which controls Crinipellis perniciosa.

Isolate 1051 of Trichoderma harzianum, a mycoparasitic fungus, was found to impair development of the phytopathogen, Crinipellis perniciosa, in the field. This Trichoderma strain growing in liquid medium containing chitin produced substantial amounts of chitinases. The N-acetylglucosaminidase present in the culture-supernatant was purified to homogeneity by gel filtration and hydrophobic interaction chromatography, as demonstrated by SDS-PAGE analysis. The enzyme had a molecular mass of 36 kDa and hydrolyzed the synthetic substrate rho-nitrophenyl-N-acetylglucosaminide (rhoNGlcNAc) with Michaelis-Menten kinetics. Maximal activities were determined at pH 4.0 and a temperature range of 50-60 degrees C. Km and Vmax values for rhoNGlcNAc hydrolysis were 8.06 micromoles ml(-1) and 3.36 micromoles ml(-1) min(-1), respectively, at pH 6.0 and 37 degrees C. The enzyme was very sensitive to Fe3+, Mn2+ and Co2+ ions, but less sensitive to Zn2+, Al3+, Cu2+ and Ca2+. Glucose at a final concentration of 1 mM inhibited 65% of the original activity of the purified enzyme. Determination of the product (reducing sugar) of hydrolysis of C. perniciosa mycelium and scanning electron microscopic analysis revealed that the N-acetylglucosaminidase hydrolyses the C. perniciosa cell wall.

Molecular markers for the characterization of Brazilian Cercospora caricis isolates.

Cercospora caricis is of interest as a potential mycoherbicide for control of purple nutsedge, Cyperus rotundus, which is considered to be the world's worst weed. The genetic variation of a collection of Brazilian Ce. caricis isolated from Cy. rotundus was analyzed by using RAPD, RFLP with a telomeric probe, [TTAGGG]18 and sequencing of the ITS1-5.8S-ITS2 regions of the ribosomal RNA gene. The Brazilian isolates were also compared with a Ce. caricis isolate from Florida, USA and with some other Cercospora species. A cluster of isolates from the Brazilian cerrado region was identified showing high genetic similarity. In contrast, isolates originating in other geographic regions of Brazil were less than 50% and 25% related to the former group according to similarity estimates produced from RAPD and telomeric RFLP analyses respectively. ITS sequence analysis did not support taxonomic division of the Brazilian strains, but did confirm the distant relatedness of these strains to the Ce. caricis isolate from Florida. The data indicate a need for an extensive molecular survey of Cercospora species associated with the Cyperaceae.

Biolistic co-transformation of Metarhizium anisopliae var. acridum strain CG423 with green fluorescent protein and resistance to glufosinate ammonium.

Metarhizium anisopliae var. acridum (syn. M. flavoviride) is recognized as a highly specific and virulent mycopathogen of locusts and grasshoppers and is currently being developed as a biological control agent for this group of insects in Brazil. Intact conidia of M. anisopliae var. acridum strain CG423 were transformed using microparticle bombardment. Plasmids used were: (1) pBARKS1 carrying the bar gene of Streptomyces hygroscopicus fused to the Aspergillus nidulans trpC promoter, encoding resistance to glufosinate ammonium (or phosphinothricin) and modified by addition of the telomeric repeat (TTAGGG)(18) of Fusarium oxysporum and 2.pEGFP/gpd/tel carrying a red-shifted variant gene for Aequorea victoria green fluorescent protein (EGFP) which we have fused to the A. nidulans gpd promoter and trpC terminator. Highly fluorescent co-transformants were selected on solid minimal medium containing 100 microg ml(-1) glufosinate ammonium using an inverted microscope with 450-490 nm excitation/510 nm emission filter set. Southern blot analysis of co-transformants revealed varying multiple chromosomal integrations of both bar and egfp genes at both telomeric and non-telomeric loci. Transformants retained pathogenicity in bioassays against Rhammatocerus schistocercoides and showed unaltered lack of pathogenicity against larvae of the non-target insect Anticarsia gemmatalis. One co-transformant from four tested, however, showed a significant, but non-dose-dependent, elevation in virulence against Tenebrio molitor.

High pharyngeal carriage rates of Streptococcus pyogenes in Dunedin school children with a low incidence of rheumatic fever.

AIM: To document the incidence and type distribution of Streptococcus pyogenes in a group of Dunedin children throughout the 1997 school year. METHODS: The 780 children recruited from ten primary schools had their throats swabbed on each reporting of pharyngitis. Additional pharyngeal swabbings were obtained monthly from a representative subset of these children, referred to as group 1. All swab samples were plated on CNA-P, a blood agar medium that facilitates detection of haemolytic streptococci. S. pyogenes isolates were classified according to the RFLP patterns of PCR products of their emm genes (ERP typing). Representative isolates of each ERP pattern were also emm-typed, a sequence typing method that correlates with serological M-typing. RESULTS: 28% of the group 1 children were found to carry S. pyogenes for more than two months. This carriage rate is similar to that previously detected in Dunedin and Waikato schoolchildren, but is higher than that generally reported in other countries. Although the predominant S. pyogenes types detected in Dunedin are similar to those in North Island populations, some of the types frequently associated with North Island cases of rheumatic fever and glomerulonephritis were absent or isolated infrequently from the Dunedin children. CONCLUSION: The high pharyngeal carriage rates of S. pyogenes in Dunedin schoolchildren, without the concomitant increased occurrence of post-streptococcal sequelae observed in North Island populations may, in part, be due to a relatively lower occurrence of the M-types most commonly implicated in these diseases.

Transformation of the entomopathogenic fungi, paecilomyces fumosoroseus and paecilomyces lilacinus (deuteromycotina: hyphomycetes) to benomyl resistence

The entomopathogenic fungi Paecilomyces fumosoroseus and P. lilacinus have been transformed to resistance to the fungicide benomyl by a polyethylene glycol (PEG)-mediated procedure using a mutant b-tubulin gene from Neurospora crassa carried on plasmid pBT6. Benomyl-resistant transformants of P. lilacinus were obtained that could tolerate greater than 30 µg/ml benomyl and P. fumosoroseus transformants were obtained that could tolerate 20 µg/ml benomyl. Following 5 serial passages of transformants on benomyl-containing media and 5 serial passages on non-selective media, 100 per cent of P. lilacinus transformants were found to be mitotically stable by a conidial germination test. In contrast, only 4 out of 9 transformants of P. fumosoroseus were mitotically stable. Southern blot analysis of genomic DNA from both species suggested that the mechanism of transformation in all transformants was by gene replacement of the b-tubulin allele. Non-homologous vector sequences were not detectable in the genomes of transformants.

Effect of citric acid on the biosynthesis and composition of xanthan.

The effect of citric acid metabolism by Xanthomonas campestris on composition of xanthan has been studied. Citric acid consumption in fed-batch and continuous fermentation increased the pyruvic acid content of xanthan. An increase in pyruvic acid content in xanthan has been explained with the help of energy balance in xanthan biosynthesis.

Transformation with green fluorescent protein of Trichoderma harzianum 1051, a strain with biocontrol activity against Crinipellis perniciosa, the agent of witches'-broom disease of cocoa.

A plasmid vector for fungal expression of an enhanced, red-shifted variant of the Aequoria victoriae green fluorescent protein was constructed by fusion of the EGFP gene to the highly expressed Aspergillus nidulans gpd promoter and the A. nidulans trpC terminator. This construction was introduced by cotransformation, using benomyl selection, into Trichoderma harzianum strain 1051, a strain being evaluated for the biological control of witches'-broom disease of cocoa caused by Crinipellis perniciosa. Epifluorescence microscopy was used to monitor germination and attachment of stable transformant conidia on the surface of C. perniciosa hyphae.

Rapid isolation of double-stranded RNAs from entomopathogenic species of the fungus Paecilomyces using a commercial minicolumn system.

A method is described for rapid extraction of double-stranded RNAs from entomopathogenic fungi. Lyophilised and ground mycelium is incubated with 6 M guanidine thiocyanate, centrifuged, and the cleared lysate applied to a QIAGEN silica-based mini-spin column. Following washing with 70% isopropanol, bound nucleic acids are eluted under low salt conditions and treated with DNAse I prior to analysis by non-denaturing agarose gel electrophoresis.

Laparoscopic cholecystectomy: its impact on national health economics.

OBJECTIVE: To assess whether sound economic reasons exist for the wider introduction of laparoscopic cholecystectomy in Australia. DESIGN: A retrospective survey of patients who underwent laparoscopic cholecystectomy. We compared time of hospital stay and time off work after laparoscopic cholecystectomy with data for open cholecystectomy. PATIENTS AND SETTING: Seventeen participating surgeons in four Australian States allowed access to patients treated between May 1990 and November 1991 (1254 patients in all). MAIN OUTCOME MEASURES: Patient acceptability of the procedure, average length of postoperative in-hospital stay, and the savings associated with earlier return to work compared with open cholecystectomy. RESULTS: Almost 90% of patients (1127) replied and 1088 responses were considered appropriate for analysis. Serious complications were rare; 96% of patients thought the technique was successful. The mean length of in-hospital stay was 2.6 days (range, 1-120), compared with a mean of 8.7 days for open cholecystectomy. Among working patients, the mean time to return to work was 11.6 days (range, 10.7-13.1), an estimated 27 days sooner than after open cholecystectomy. Extrapolating from these results, replacing 95% of open cholecystectomies with laparoscopic procedures would have 133,285 hospital bed-days and 500,000 work-days each year. CONCLUSION: Laparoscopic cholecystectomy is safe and effective. Its wider use in Australia would result in savings to both the individual and the national economy.

Sudden death in HIV-infected drug users. Presumptive overdose in an HIV-positive narcotic addict.

We report the case of an HIV-positive drug user who was found dead in a room in which were discovered tablets of dihydrocodeine and amitriptyline. There had been past episodes of drug overdose which was suspected of being the cause of death on this occasion. However toxicology analysis did not support this diagnosis and necropsy revealed a severe desquamative interstitial pneumonitis and HIV encephalitis. The need to investigate thoroughly the cause of sudden death in all HIV-positive drug users is emphasised.