Intraneural injection of a test dose of local anesthetic in peripheral nerves - does it induce histological changes in nerve tissue?

BACKGROUND & OBJECTIVES: Most anesthesiologists use the injection of a test dose of local anesthetic in order to evaluate the final needle tip position. Thus, the intraneural injection of a full dose can be avoided. The aim of this study was to analyze whether an intraneural injection of a test dose of bupivacaine could trigger histological changes. METHODS: Intraneural injections under direct vision were performed in 40 brachial plexus nerves in seven anesthetized pigs. Tibial nerves served as positive and negative controls. Two milliliter of bupivacaine 0.5% was injected in three nerves on the left brachial plexus. For control of local anesthetic's toxicity Ringer's solution was applied intraneurally on the right side. After maintaining 48 h of general anesthesia, the nerves were resected. The specimens were processed for histological examination and assessed for inflammation (hematoxylin and eosin stain, CD68-immunohistochemistry) and myelin damage (Kluver-Barrera stain). The degree of nerve injury was rated on a scale from 0 (no injury) to 4 (severe injury). RESULTS: Statistical analysis showed no significant differences between the bupivacaine group [median (interquartile range) 1 (1-1.5)] and the Ringer's solution group [1 (0.5-2) P = 0.772]. Mild myelin alteration was found in 12.5% of all specimens following intraneural injection, irrespective of the applied substance. CONCLUSIONS: "In our experimental study, intraneural injection of 2 ml of bupivacaine or Ringer's solution showed comparable mild inflammation. Nevertheless, inflammation can only be prevented by strictly avoiding nerve perforation followed by intraneural injection, as mechanical nerve perforation is a key factor for evolving inflammation.

Strategies for the reduction of Legionella in biological treatment systems.

A community-wide outbreak of Legionnaire's disease occurred in Warstein, Germany, in August 2013. The epidemic strain, Legionella pneumophila Serogruppe 1, was isolated from an industrial wastewater stream entering the municipal wastewater treatment plant (WWTP) in Wartein, the WWTP itself, the river Wäster and air/water samples from an industrial cooling system 3 km downstream of the WWTP. The present study investigated the effect of physical-chemical disinfection methods on the reduction of the concentration of Legionella in the biological treatment and in the treated effluent entering the river Wäster. Additionally, to gain insight into the factors that promote the growth of Legionella in biological systems, growth experiments were made with different substrates and temperatures. The dosage rates of silver micro-particles, hydrogen peroxide, chlorine dioxide and ozone and pH stress to the activated sludge were not able to decrease the number of culturable Legionella spp. in the effluent. Nevertheless, the UV treatment of secondary treated effluent reduced Legionella spp. on average by 1.6-3.4 log units. Laboratory-scale experiments and full-scale measurements suggested that the aerobic treatment of warm wastewater (30-35 °C) rich in organic nitrogen (protein) is a possible source of Legionella infection.

[Hygiene and Public Health - Significance in Medicine and Society]. / Hygiene und Öffentliche Gesundheit - Zur Bedeutung in Medizin und Gesellschaft.

The presented review attempts an historical overview on the development of Public Health in Germany with special reference to the medical specialty of Hygiene. This development is put in perspective to current international developments with a special emphasis on the programmatic work in the field of Public Health of the European Union.

[Hospital disinfection challenges due to multiresistant pathogens in surgery]. / Krankenhaushygienische Herausforderungen durch multiresistente Erreger in der Chirurgie.

The number of antibiotic-resistant pathogens is increasing continuously while the development of new, effective antibiotics cannot be expected in the near future. Postoperative infections represent most of the nosocomial infections by now. Based on this, hygienic strategies regain importance, since a sustainable control of nosocomial infections will not succeed without the implementation of such strategies. In this article, the most important preventive strategies for prevention of infections with MRSA and 3- and 4-fold resistant gram-negative bacteria on the basis of current recommendations of the Commission for Hospital Hygiene and Infection Prevention (KRINKO) are presented.

[Mandatory reporting of antimicrobial-resistant pathogens. What should, what can be achieved? : assessment related to the particular facility seems crucial]. / Meldepflicht für Antibiotika-resistente Erreger. Was soll, was kann sie leisten? : Die Bewertung in Bezug auf die jeweilige Einrichtung erscheint wichtig.

Antimicrobial-resistant organisms are regarded as a particular threat to the public health of the European population. In Germany the requirement for a national rollout reporting of positive laboratory test results for methicillin-resistant Staphylococcus aureus (MRSA) in blood cultures was implemented in 2009. This was followed in 2011 by the introduction of a laboratory-based, rollout reporting system for the detection of gram-negative bacteria with acquired carbapenem-resistance (carbapenem-resistant organisms, CRO) in the federal state of Hessen. This article will present the experience gained in Frankfurt am Main with the existing reporting system. Blood or cerebrospinal fluid cultures positive for MRSA were reported from all Frankfurt clinics between 2010 and 2013. The objective of preventing nosocomial infections by introducing a mandatory reporting for MRSA in blood cultures was only partially achieved by the reporting procedure on a population scale. Instead, reporting on a clinic-based scale, i.e., incidence per 1,000 patient days should be used. Moreover, mandatory reporting of clusters of nosocomial colonizations with MRSA could be an appropriate measure for the timely prevention of nosocomial infections with these organisms. CROs were reported from nearly all clinics as well as the ambulatory setting. Different reporting criteria have resulted not only in a greatly varying workload for the institutions and the health authorities but also in vastly different numbers of reported events. Regarding the importance of CRO, mandatory reporting seems reasonable. Criteria of reporting should be simple and easily comprehensible, i.e., all 4MRGN (gram-negative bacteria with resistance against four antibiotic groups such as acylureidopenicillins, third- and fourth-generation cephalosporins, fluorchinolones, and carbapenems) according to the German Commission on Hospital Hygiene and Infection Prevention (KRINKO) should be considered. Reporting and evaluation of multidrug-resistant organisms (MDRO) in a population-based system does not seem to be sufficient regarding these organisms with high importance for hospital hygiene; mandatory reporting on an anonymous clinic-based scale should be used.

[Ten years' German Protection against Infection Act. Evaluation of the implementation of infection control visits in the ambulatory medical setting]. / Rückblick auf 10 Jahre Infektionsschutzgesetz. Evaluation der Einführung der infektionshygienischen Überwachung ambulanter medizinischer Einrichtungen.

In 2001, the German Protection against Infection Act came into force, implementing a variety of new regulations. For the first time, obligatory infection control visits of the public health departments in surgical ambulatory practices were implemented, as well as optional infection control visits in all medical, dental and paramedical practices using invasive methods. Based on the data of the public health department of the city of Frankfurt am Main, Germany, an evaluation of this new regulation is given in this paper. First, prioritization of these new tasks was mandatory. First priority was given to the obligatory visits in surgical practices, second priority to the hygiene visits in practices performing endoscopy in gastroenterology as well as in urology and in practices of traditional healers, and third priority was given to all other doctors' practices. After receiving preliminary information and further training of the doctors etc., the control visits were performed by members of the public health department, using a checklist based on the guidelines of the German Commission on Hospital Infection Prevention ("Kommission für Krankenhaushygiene und Infektionsprävention"). Since 2001, more than 1100 infection control visits in medical practices in Frankfurt am Main were documented. Not only in surgical, but also in gastroenterological and urological practices great improvement could be achieved, regarding not only hand hygiene and reprocessing surface areas, but especially in reprocessing medical devices. In practices for internal medicine and those of general practitioners, errors in hand hygiene, skin antiseptic and surface disinfection also decreased. According to our results, especially regarding the improved quality of structure as well as quality of process and with regard to the public discussion on this hygiene topic, our evaluation is absolutely positive. The new regulation proved worthwhile.

Reliability of fiber post bonding to root canal dentin after simulated clinical function in vitro.

The aim of this study was to investigate the effect of thermomechanical loading (TML) on the bond strength of fiber posts luted with three different resin cements. Sixty-six extracted human anterior teeth were endodontically treated and restored with fiber posts (RelyX Fiber Posts, 3M ESPE) using three commercially available resin cements and three corresponding core build-up materials (n=22 each): Panavia F 2.0/Clearfil DC Core Automix (Kuraray), Variolink II/Multicore Flow (Ivoclar Vivadent), and RelyX Unicem/Filtek Z250 (3M ESPE). Twelve specimens of each group received all-ceramic crowns and were subjected to TML. The other 10 specimens were stored in saline solution for 24 hours. The roots were sectioned and bond strength was measured using a push-out test. Adhesive interfaces of two specimens of each group subjected to TML were analyzed using field emission scanning electron microscopy (FESEM). Bond strengths of fiber posts were significantly affected by the type of resin cement (p<0.0005) and TML (p<0.0005; two-way analysis of variance). TML significantly reduced bond strengths for all materials ((6.0 (6.2) MPa)) compared with initial bond strengths ((14.9 (10.4) MPa)). RelyX Unicem resulted in significantly higher bond strengths before ((18.3 (10.3) MPa)) and after TML ((9.8 (7.5) MPa)) compared with the other materials (p<0.0005; Tukey HSD). Using FESEM, Variolink II and Panavia F demonstrated a hybrid layer partly detached from the underlying resin cement, whereas no hybrid layer was observed for RelyX Unicem. The decrease in bond strength after TML suggests that retention of fiber posts may be reduced after clinical function. Therefore, endodontically treated teeth that are restored using fiber posts may benefit from additional reinforcement via coronal restorations using adequate ferrules and/or adhesive techniques.

[Experiences with outbreaks of nosocomial infections]. / Erfahrungen aus dem Ausbruchmanagement bei nosokomialen Infektionen.

Ten years after the publication of the recommendation: "Outbreak management and structural proceedings in case of cumulative occurrence of nosocomial infections" of the federal commission of hospital hygiene, these recommendations are now being re-evaluated. To date, the recommendations have proven valid and have maintained their significance for an effective management. However, besides new hygienic-microbiological methods and an increased sensitivity of the perception of nosocomial outbreaks by the public, by politicians and by the press, it is necessary to consider new issues in this field. Outbreaks are tragic events placing an extraordinary burden on all persons involved, which can have significant consequences. Therefore, it is necessary to ensure prompt outbreak management by experienced professionals who must combine a systematic on-site inspection, hygienic-microbiological investigation and typing methods used with epidemiological approaches. To assure these requirements, the support of independent reference centres such as universal hygiene institutes should be guaranteed. Politicians should be involved only after a scientific evaluation of the details of the outbreak has been made. A national documentation centre, e.g. at the Robert Koch Institute, should be established, thereby making experiences with outbreaks widely available.

[Historical development and current demands on medical training, further and advanced training in hygiene and infection prevention]. / Historische Entwicklung und aktuelle Anforderungen an die ärztliche Aus-, Fort- und Weiterbildung in der Hygiene und Infektionsprävention.

New risks in nosocomial infections and the dramatic increase in antibiotic-resistant pathogens in healthcare facilities have pointed to the urgent need for a good education of students and practitioners in the basics of hospital hygiene and infection prevention. On the other hand in the last 10 years a large number of institutes of hygiene in universities were closed with remarkable consequences concerning the decreased education in modern hygiene and public health. A broad historical overview over the last 200 years of teaching hygiene and public health at German universities is given which was integrated into the education of medical students. Nowadays many universities do not teach modern hygiene and public health. The demand of re-establishing new institutes of hygiene by the German Medical Council is discussed. The curriculum for the formation of hospital hygienists is presented.

Explosive COVID-19 outbreak in a German nursing home and the possible role of the air ventilation system.

BACKGROUND: Most COVID-19 outbreaks in nursing homes are explained by transmission of SARS-CoV-2 from nurses or visitors. METHODS AND RESULTS: We describe an outbreak with 64 of the 67 residents identified as COVID-19 cases within two weeks (34 in nursing block 1, 30 in nursing block 2), at least 32 of them had relevant symptoms of COVID-19. Thirteen of the residents' deaths were associated with COVID-19. In addition, 27 of approximately 60 staff members were identified as COVID-19 cases, 23 of them had relevant symptoms. In none of the samples from residents or staff was a mutation of SARS-CoV-2 detected. Quarantine of the residents was already in force at the beginning of the outbreak. A common source among the staff was considered to be unlikely because the two nursing home blocks had no staff rotation and the staff had to wear FFP2 masks during contact with residents. Three months after the outbreak the RNA of SARS-CoV-2 was detected on 14 of 39 sampled indoor surfaces of the air ventilation system with Ct values between 34.9 and 41.9, but only at the air supply in the corridor (11 of 24 samples) and the air overflow in the door between the corridor and the residents' rooms (three of 11 samples) but not at the air exhaust in the residents' bathrooms. CONCLUSIONS: The air ventilation system and an inversion weather situation three days before the first confirmed case may have enhanced viral spread inside the nursing home assuming that a common source with a high viral load had existed at the time of outbreak.

Efficacy of five 'sporicidal' surface disinfectants against Clostridioides difficile spores in suspension tests and 4-field tests.

BACKGROUND: A sporicidal surface disinfection is recommended both for the outbreak and the endemic setting but a comparative evaluation on the efficacy of 'sporicidal' surface disinfectants using suspension tests and 4-field tests has not been performed. AIM: To determine the efficacy of five 'sporicidal' surface disinfectants (three ready-to-use wipes (A, B, E), two concentrates (C, D) based on peroxides or aldehydes against C. difficile spores. METHODS: The efficacy was determined under clean conditions using a suspension test and the 4-field test. Each test was performed in duplicate in two separate laboratories. Wipes were wrung to collect the solution for the suspension tests. RESULTS: Product A (peracetic acid; 5 min), product C (peracetic acid; 2% solution in 15 min or 1% solution in 30 min) and product D (peracetic acid; only 2% solution in 15 min) were effective with at least a 4 log10-reduction of C. difficile spores in suspension and on surfaces. Product B (hydrogen peroxide) was not effective in suspension (0.9 log10 after 15 min; 3.2 log10 after 1 h) and on surfaces (2.8 log10 after 15 and 60 min). Product E based on glutaraldehyde, (ethylendioxy)dimethanol and DDAC demonstrated 0.9 log10 after 4 h in suspension and 4.5 log10 after 4 h on surfaces. CONCLUSIONS: Not all surface disinfectants with a sporicidal claim were effective against C. difficile spores in standardized suspension tests and in the 4-field test. In clinical practice preference should be given to products that reliably pass the efficacy criteria of both types of tests.

An international trial of quantitative PCR for monitoring Legionella in artificial water systems.

AIMS: To perform an international trial to derive alert and action levels for the use of quantitative PCR (qPCR) in the monitoring of Legionella to determine the effectiveness of control measures against legionellae. METHODS AND RESULTS: Laboratories (7) participated from six countries. Legionellae were determined by culture and qPCR methods with comparable detection limits. Systems were monitored over ≥10 weeks. For cooling towers (232 samples), there was a significant difference between the log mean difference between qPCR (GU l(-1) ) and culture (CFU l(-1) ) for Legionella pneumophila (0·71) and for Legionella spp. (2·03). In hot and cold water (506 samples), the differences were less, 0·62 for Leg. pneumophila and 1·05 for Legionella spp. Results for individual systems depended on the nature of the system and its treatment. In cooling towers, Legionella spp. GU l(-1) always exceeded CFU l(-1) , and usually Legionella spp. were detected by qPCR when absent by culture. The pattern of results by qPCR for Leg. pneumophila followed the culture trend. In hot and cold water, culture and qPCR gave similar results, particularly for Leg. pneumophila. There were some marked exceptions with temperatures ≥50°C, or in the presence of supplementary biocides. Action and alert levels for qPCR were derived that gave results comparable to the application of the European Guidelines based on culture. Algorithms are proposed for the use of qPCR for routine monitoring. CONCLUSIONS: Action and alert levels for qPCR can be adjusted to ensure public health is protected with the benefit that remedial actions can be validated earlier with only a small increase in the frequency of action being required. SIGNIFICANCE AND IMPACT OF THE STUDY: This study confirms it is possible to derive guidelines on the use of qPCR for monitoring the control of legionellae with consequent improvement to response and public health protection.

[Hygiene in schools - an important issue for the public health services]. / Hygiene in Schulen - auch eine wichtige Aufgabe des öffentlichen Gesundheitsdienstes.

According to the Protection against Infection Act (IfSG), schools have to identify their arrangements of standard operating procedures (SOPs) for hygienic conditions, and the public health departments are obliged to check the hygienic conditions in schools. Here, practical experience with these topics in Frankfurt/Main, Germany, is presented.Although all schools had been informed about their duties according to IfSG in 2001, only 44% (80/180 schools) of them were able to present their SOPs when requested to do so by the public health department in 2006. Complaints about bad sanitary hygiene in schools have to be confirmed, often even in recently redeveloped facilities, because of vandalism. The equipment of washing basins was improved very well during the influenza pandemic in 2009. In 2010, however, a tendency to deterioration had been observed. With data on high levels on indoor contamination (particles and CO2) in class rooms in 2006, the city increased the frequency of cleaning and launched a programme on proper ventilation in classrooms. However, a study on ventilation in 29 schools in 2009 resulted in very high levels of CO2; obviously the recommendation to ventilate the classrooms every break are neglected. Large studies on drinking water quality exhibited good data, with the exception of high Legionella contaminations in hot water systems supplying the showers in gymnasiums. Major redevelopment measures were necessary.Hygienic conditions in schools should be improved urgently. The public health departments should increase their consulting services as well as their control visits in schools - with the aim to improve hygiene in schools. Responsibility of the schools as well as of the individuals in the schools is demanded as well. More public attention to this topic seems mandatory, may be via a "signal light - red-yellow-green" for schools?

[Requirements for the prevention of nosocomial infections. German Guideline 2009 and reality. Current data from hospitals in Frankfurt am Main, Germany]. / Personelle und organisatorische Voraussetzungen zur Prävention nosokomialer Infektionen. Richtlinie und Realität 2009. Ergebnisse aus Frankfurt am Main.

In 2009, the new directive of the German Commission for Hospital Hygiene and Infection Prevention (Kommission für Krankenhaushygiene und Infektionsprävention, KRINKO) entitled Human and Organizational Requirements for the Prevention of Nosocomial Infections was published, including detailed information on the needs of hygiene professionals in hospital settings. Compared to the needs calculated according to the above policy, the current staff hygiene health professionals (HHPs) in the hospitals of Frankfurt am Main (Frankfurt/M), Germany, was on average 27.6%: 36% in the large hospitals (>600 beds), 21.6% in medium hospitals (300-600 beds), and 19.8% in small hospitals (<300 beds). Only 1 of 14 hospitals had a full-time hygienist. The demands of the KRINKO policy have not been met by any of the hospitals. Hospitals with lower percentages of hospital hygiene staff not only had a lower rating of hygienic quality, they also showed a lower usage of hand disinfection per patient-day. In Germany, there is currently a lack of trained HHPs and hygienists to meet the needs of the KRINKO policy. Therefore, the reactions of the hospitals in Frankfurt/M ranged from the establishment of additional jobs for HHPs to changes in structures and organization of hospital hygiene. Thus, the new KRINKO guideline in Frankfurt/M did not result in a wave of recruitment of health professionals, but at least resulted in organizational and structural improvements in hygiene.

[Experience with prevention and control of legionellosis in Germany : plea for proactive risk management]. / Erfahrungen zur Prävention und Kontrolle von Legionellosen in Deutschland : Plädoyer für ein proaktives Risikomanagement.

Legionellosis is meanwhile the most important specific water-associated infectious disease in developed countries, which is completely preventable, if water distribution systems are correctly planned and operated. This assumes clear criteria for risk regulation and for verification, including microbiological monitoring for Legionella. There are different reactive and proactive strategies in the USA and in Europe. The common premises for prevention and control of legionellosis in Germany, relevant facts for risk regulation, experience in Germany toward proactive risk regulation, and the current approach of the amended drinking water ordinance are discussed. The article concludes with a short discussion of the controversial approaches for the prevention of legionellosis in Germany.

Outbreak of Pseudomonas aeruginosa infections after CT-guided spinal injections.

BACKGROUND: Meningitis and spinal infections with Gram-negative bacteria after local injections for treatment of chronic back pain are rare. This study investigated an outbreak of Pseudomonas aeruginosa infections following computed tomography (CT)-guided spinal injections (SI). METHODS: A case was defined as a spinal infection or meningitis with P. aeruginosa after SI between 10th January and 1st March 2019 in the same outpatient clinic. Patients without microbiological evidence of P. aeruginosa but with a favourable response to antimicrobial therapy active against P. aeruginosa were defined as probable cases. FINDINGS: Twenty-eight of 297 patients receiving CT-guided SI during the study period developed meningitis or spinal infections. Medical records were available for 19 patients. In 15 patients, there was microbiological evidence of P. aeruginosa, and four patients were defined as probable cases. Two of 19 patients developed meningitis, while the remaining 17 patients developed spinal infections. The median time from SI to hospital admission was 8 days (interquartile range 2-23 days). Patients mainly presented with back pain (N=18; 95%), and rarely developed fever (N=3; 16%). Most patients required surgery (N=16; 84%). Seven patients (37%) relapsed and one patient died. Although the source of infection was not identified microbiologically, documented failures in asepsis when performing SI probably contributed to these infections. CONCLUSIONS: SI is generally considered safe, but non-adherence to asepsis can lead to deleterious effects. Spinal infections caused by P. aeruginosa are difficult to treat and have a high relapse rate.

A novel class of autoantigens of anti-neutrophil cytoplasmic antibodies in necrotizing and crescentic glomerulonephritis: the lysosomal membrane glycoprotein h-lamp-2 in neutrophil granulocytes and a related membrane protein in glomerular endothelial cells.

Necrotizing and crescentic glomerulonephritis (NCGN) is frequently associated with circulating antineutrophil cytoplasmic autoantibodies (ANCA). It is established that ANCA are specific for soluble enzymes of granules of polymorphonuclear neutrophil granulocytes (PMN), such as myeloperoxidase (MPO) or protease 3 (PR3). The purpose of this study was to identify membrane proteins of PMNs, and/or glomerular cells, as additional autoantigenic ANCA targets. When membrane protein fractions were prepared from PMNs and isolated human glomeruli, and immunoblotted with ANCA sera of NCGN patients, two bands with apparent molecular masses of 170 and 80-110 kD (gp170/80-110) were labeled in PMNs, and a 130-kD glycoprotein (gp130) in glomeruli. Gp130 was purified, and monoclonal and rabbit antibodies (Abs) were produced which showed the same double specificity as the patient's ANCA. Using these probes, evidence was provided that gp170/80-110 is identical with human lysosomal-associated membrane protein 2 (h-lamp-2), because both proteins were immunologically cross-reactive and screening of a cDNA expression library from human promyelocytic leukemia cells with anti-gp130 Ab yielded a clone derived from h-lamp-2. Gp170/80-110 was localized primarily in granule membranes of resting PMNs, and was translocated to the cell surfaces by activation with FMLP. By contrast, gp130 was localized in the surface membranes of endothelial cells of human glomerular and renal interstitial capillaries, rather than in lysosomes, as found for h-lamp-2. Potential clinical relevance of autoantibodies to gp170/80-110 and gp130 was assessed in a preliminary trial, in which ANCA sera of patients (n = 16) with NCGN were probed with purified or recombinant antigens. Specific reactivity was detected in approximately 90% of cases with active phases of NCGN, and frequently also in combination with autoantibodies specific for PR3 or MPO. Collectively, these data provide evidence that h-lamp-2 in PMNs and a different, structurally related 130-kD membrane protein on the cell surface of renal microvascular endothelial cells are autoantigenic targets for ANCA in patients with active NCGN.

Induction of passive Heymann nephritis with antibodies specific for a synthetic peptide derived from the receptor-associated protein.

Passive Heymann nephritis (pHN) is an experimental rat model for human membranous glomerulopathy. In pHN, the formation of subepithelial immune deposits (ID) involves as antigenic targets the membrane glycoprotein gp330/megalin and the 44-kD receptor-associated protein (RAP). A single binding site for ID- inducing antibodies (Abs) was previously mapped to the 86 NH2-terminal amino acids of RAP (RAP1-86). To further narrow this epitope, Abs eluted from the glomeruli were immunoblotted on membranes that were loaded with overlapping synthetic peptides representing the amino acid sequence of RAP (SPOTs system). Two adjacent Ab-binding domains with the sequences PVRLAF, (amino acids 39-44) and HSD-LKIQE (amino acids 46-53), which were separated by a single L residue at amino acid 45, were detected. Rabbit Abs raised against synthetic peptides containing these domains individually (P31-44 and P46-53) failed to procedure glomerular IDs. By contrast, Abs raised against a larger composite peptide (P31-53) induced IDs within 3d that were firmly cross linked to the glomerular basement membrane. These data suggest that Ab binding in vivo depends on the conformation of the antigenic target sequence that is preserved in the synthetic peptide P31-53, which covers the entire Ab-binding domain of RAP but not in its subdomains, P31-44 and P46-53. Collectively, these results locate the sole ID-inducing epitope of RAP to amino acids 39-53.