Showing papers in "Comparative Immunology Microbiology and Infectious Diseases in 2004"

Leishmaniasis: current situation and new perspectives.

Abstract: Leishmaniasis represents a complex of diseases with an important clinical and epidemiological diversity. Visceral leishmaniasis (VL) is of higher priority than cutaneous leishmaniasis (CL) as it is a fatal disease in the absence of treatment. Anthroponotic VL foci are of special concern as they are at the origin of frequent and deathly epidemics (e.g. Sudan). Leishmaniasis burden remains important: 88 countries, 350 million people at risk, 500,000 new cases of VL per year, 1-1.5 million for CL and DALYs: 2.4 millions. Most of the burden is concentrated on few countries which allows clear geographic priorities. Leishmaniasis is still an important public health problem due to not only environmental risk factors such as massive migrations, urbanisation, deforestation, new irrigation schemes, but also to individual risk factors: HIV, malnutrition, genetic, etc em leader Leishmaniasis is part of those diseases which still requires improved control tools. Consequently WHO/TDR research for leishmaniasis has been more and more focusing on the development of new tools such as diagnostic tests, drugs and vaccines. The ongoing effort has already produced significant results. The newly available control tools should allow a scaling up of control activities in priority areas. In anthroponotic foci, the feasibility of getting a strong impact on mortality, morbidity and transmission, is high.

The changing epidemiology of yellow fever and dengue, 1900 to 2003: full circle?

Abstract: Yellow fever and dengue are old diseases, having caused major epidemics in centuries past. Both were effectively controlled in the mid 1900s, yellow fever in Francophone Africa by vaccination and yellow fever and dengue in the Americas by effective control of the principal urban vector of both viruses, Aedes aegypti. In the last 25 years of the 20th century, however, there was a resurgence of yellow fever in Africa, and of dengue worldwide. The factors responsible for this resurgence are discussed, as are current options for prevention and control.

West Nile: worldwide current situation in animals and humans

Abstract: West Nile (WN) virus is a mosquito-borne flavivirus that is native to Africa, Europe, and Western Asia. It mainly circulates among birds, but can infect many species of mammals, as well as amphibians and reptiles. Epidemics can occur in rural as well as urban areas. Transmission of WN virus, sometimes involving significant mortality in humans and horses, has been documented at erratic intervals in many countries, but never in the New World until it appeared in New York City in 1999. During the next four summers it spread with incredible speed to large portions of 46 US states, and to Canada, Mexico, Central America and the Caribbean. In many respects, WN virus is an outstanding example of a zoonotic pathogen that has leaped geographical barriers and can cause severe disease in human and equine. In Europe, in the past two decades there have been a number of significant outbreaks in several countries. However, very little is known of the ecology and natural history of WN virus transmission in Europe and most WN outbreaks in humans and animals remain unpredictable and difficult to control.

Unravelling complexities in human malaria transmission dynamics in Africa through a comprehensive knowledge of vector populations

Abstract: Malaria transmission dynamics is highly variable throughout Africa: inoculation rates vary from almost null to more than a 1000 infective bites per year, transmission can occur throughout the year or only during a couple of months, and heterogeneities are also observed between years within the same locale. Depending on the area, as much as five different anophelines species can transmit parasites to the human population. Major vectors are Anopheles gambiae, Anopheles arabiensis, Anopheles funestus, Anopheles nili and Anopheles moucheti. They all belong to species complexes or groups of closely related species that are very difficult to set apart on morphological grounds. Recent research on the bionomics, morphology and genetics of these mosquito species and populations produced innovative results. New species were described and straightforward molecular identification tools were implemented. We review here these recent findings and discuss research opportunities in light of recent advances in molecular entomology and genomics.

Tick-borne rickettsial diseases: emerging risks in Europe

Abstract: Ticks are currently considered the main vectors of human infectious diseases in Europe, particularly since their role in the transmission of the agent of Lyme borreliosis was demonstrated in the 1980s. In the recent years, ticks have also been shown to be the vectors of numerous emerging rickettsial diseases. Although Mediterranean spotted fever (MSF) due to Rickettsia conorii was thought for a long time to be the only tick-borne rickettsial disease prevalent in Europe, five more spotted fever rickettsiae have been described as emerging pathogens in the last decade. Further, cases of infection due to Anaplasma phagocytophilum, the agent of human anaplasmosis (previously known as human granulocytic ehrlichiosis), have been reported throughout Europe. We present here these emerging diseases and discuss other potential threat for the future.

Cell-mediated immune responses to a killed Salmonella enteritidis vaccine: Lymphocyte proliferation, T-cell changes and interleukin-6 (IL-6), IL-1, IL-2, and IFN-γ production

Abstract: Two experimental approaches were used to investigate the immunological responses of chickens to a commercial killed Salmonella enteritidis (SE) vaccine. In the first, the effects of host age on antigen-specific proliferative responses and cytokine production were examined. Compared with non-vaccinated controls, 4-wk-old vaccinated chickens showed higher proliferation to SE LPS and flagella. The lymphoproliferation responses to these antigens of 8-mo-old vaccinated chickens were not different compared to the non-vaccinated controls. Increased production of interferon-gamma (IFN-gamma) and interleukin-2 (IL-2) by antigen-stimulated splenocytes following vaccination were, in general, more often observed in 4-wk-old compared with 8-mo-old chickens, whereas serum levels of these cytokines were consistently higher in the vaccinated birds compared with controls regardless of age. The second set of experiments were designed to determine the effects of SE vaccination on mitogen- or antigen-induced splenocyte proliferation and serum nitric oxide (NO) and cytokine levels. Splenocytes from vaccinated chickens stimulated with SE flagella showed significantly increased numbers of TCRgammadelta+ cells at 7 days post-vaccination compared with non-vaccinated birds. In contrast, no differences were noted with CD4+, CD8+, or TCRalphabeta+ cells at any time points examined. Higher levels of NO production were observed following stimulation with SE flagella at 4, 7, 11, and 14 days after SE vaccination while serum levels of IFN-gamma, IL-1, IL-6, and IL-8 were elevated only at day 7 post-vaccination. In conclusion, younger chickens mounted a more robust antigen-specific immune response to the SE vaccine compared with older birds and vaccination induced not only T-cell-mediated responses but also host innate and pro-inflammatory responses.

Maedi–Visna virus infection in sheep. History and present knowledge

Abstract: Briefly the history of maedi–visna and the major clinical symptoms are described. Examples are presented to demonstrate that the genetic composition of a breed determines whether or not sheep become sick after an infection with maedi–visna virus (mvv) or develop solely specific antibodies. The major pathway of transmission is not colostrum and milk, but a cell containing increased nasal discharge in cases of respiratory distress. The role of the environment and prophylactic measures against parasites is stressed, because even sheep of highly susceptible breeds can survive an infection under optimal conditions. The virus and subsequently the disease simply die out. The cooperation between clinicians and laboratories is necessary.

Serotypes and anti-microbial susceptibility of Plesiomonas shigelloides isolates from humans, animals and aquatic environments in different countries.

Abstract: A total of 73 strains of Plesiomonas shigelloides isolated from humans (24 strains) animals (21 strains) and aquatic environment (28 strains) were determined for their O:H serotype and susceptibility to 18 anti-microbial substances and to the vibriostatic agent O/129. Of all strains, 86.3% were typeable by the O and 94.5% by the H anti-sera used. The serotype distribution was heterogeneous within a country and between the countries. Of the 57 different serotypes identified, O11:H2 (2 strains), O22:H3 (4 strains), O35:HH11 (2 strains), O52:H3 (2 strains) and O90:H6 (2 strains) were found among isolates from humans and animals (mainly in cats) in Finland and Cuba, and O23:H1a1b (3 strains) among isolates from environmental sources in Slovak Republic and Italy. Most (93–100%) of all strains were susceptible to all anti-microbials tested but resistant (92–96%) to the broad-spectrum penicillins (ampicillin, mezlocillin). No correlation between anti-microbial resistance patterns and serotypes was found.

Respiratory chlamydial infection based on experimental aerosol challenge of pigs with Chlamydia suis

Abstract: An experimental study of aerogeneous challenge in pigs was conducted in order to reveal characteristic features of porcine respiratory chlamydiosis. Eight conventionally raised pigs were exposed to a pathogenic strain of Chlamydia (C.) suis, four controls were mock infected. Besides pathological changes, the acute-phase and humoral immune responses, as well as the dissemination and transmission of the challenge strain was monitored in the course of infection. The data from clinical investigations, LPS-binding protein assay, antibody ELISAs, confocal laser scanning and light microscopy, immunohistochemical staining and PCR provided extensive evidence of the pathogenic potential of C. suis for the porcine respiratory system. This model appears suitable for further pathophysiological and immunological investigations of chlamydial respiratory infections and can also be recommended for studies of Chlamydia-associated infections of the human lung.

Differential detection of Newcastle disease virus strains by degenerate primers based RT-PCR.

Abstract: Degenerate primers based RT-PCR (previously described by [Avian Dis 26 (1997) 837]) has been used for the detection and differentiation of Newcastle disease (ND) viruses. Two sets of primers (A+B and A+C), with common forward primer and distinct reverse degenerate primers, designed from fusion protein gene encoding for cleavage site, could differentiate virulent and avirulent Newcastle disease viruses (NDV). Both sets of primers amplified "F" gene sequence of virulent (velogenic and mesogenic) viruses, whereas in avirulent strains, amplification was only with primer set A+C. Total 10 NDV isolates and two clinical samples including both known and unknown pathotypes, were checked. Based on amplification results 5 viruses were found to be virulent type and 6 as avirulent with one of the two clinical samples, earlier positive by RT-PCR using non-degenerate "F" gene specific primers was found negative in this study. The technique has been found to be a simple and quick for the detection and differentiation of virulent and avirulent NDV, which is important for control of the disease in the events of the outbreaks.

Isolation of a rare Escherichia coli O157:H7 strain from farm animals in Greece.

Abstract: A strain of Escherichia coli O157:H7 was isolated from goat faeces during a surveillance study on the prevalence of this serotype of E. coli in farm animals in Greece. Three hundred and fifty one faecal samples were collected from goat, sheep and cattle breeding farms in the area of Epirus, Northwestern Greece. The E. coli O157:H7 isolate was nonsorbitol-fermenter, produced only VT2 and showed a β-glucuronidase positive activity, a rather unusual biochemical feature for the E. coli O157:H7 serotype. No other strain of E. coli O157:H7 was isolated from the faecal samples of the rest farm animals examined, thus the overall prevalence of animal carriage was found to be 0.2%. The findings also indicate that goats can be a reservoir of E. coli O157:H7 and goat milk, dairy products and meat may serve as a vehicle for the pathogen transmission to humans.

Current status of medical and veterinary entomology in France: endangered discipline or promising science?

Abstract: Following alarming statements (French Senate, Academie des Sciences) on the present situation concerning entomology and systematics in France, the Conseil General Veterinaire designated one of us (D.C.) to carry out a survey on the status of medical and veterinary entomology (MVE) with respect to research orientations and university curricula. Around 100 participants, including scientists, teachers and several directors of research and educational bodies, were interviewed and filled in questionnaires for this survey. On the basis of the results, it was concluded that the deterioration of MVE in France is associated with: (1) the hasty reorganisation of training and research in the life sciences, leading to the disappearance of several disciplines. Hence, the postgraduate DEA degree in entomology was eliminated, and even the name 'entomology' no longer appears in teaching programmes or on research contracts; (2) France's withdrawal from action research programmes in developing countries. Although these programmes were efficient in controlling outbreaks of major endemic diseases, integrated pest and vector management programmes have been replaced by basic health care ('Health for everyone in 2000') and vaccination programmes; (3) the general shift from field to laboratory research, focused mainly on molecular mechanisms. The survey results confirmed generally acknowledged trends concerning many points and highlighted several specific problems, such as the disappearance of systematics experts. Several potential solutions are proposed.

Remote sensing and epidemiology: examples of applications for two vector-borne diseases.

Abstract: Remote sensing techniques have greatly contributed to improve our capacity to observe our environment and its processes. For about 15 years, the use of satellite images for epidemiological purposes has been largely promoted to determine diseases distributions and their variations through time. In some circumstances, when diseases are strongly related to environmental data such as climate, vegetation or land-use, radiation values can be included in prediction models. In other cases, remote sensing data provide information for drawing thematic layers involved in the epidemiological processes, which may differ according to the different ecotypes and ecosystems. According to its final goal, the users can choose from the panel of available radiometers with specific characteristics including spatial resolution and frequency of data. In this paper, two examples of major vector-borne diseases, namely Animal Trypanosomosis and Bluetongue, illustrate these applications.

B cell and macrophage response in chicks after oral administration of Salmonella typhimurium strains

Abstract: Despite the fact that, in a number of countries, vaccination programmes are extensively used to control Salmonella infection in poultry, information on the immune mechanisms, especially the cellular response, is still needed. The aim of the study was to characterise the B cell and macrophage response in caecum (IgA+, IgM+, IgG+ cells, macrophages), bursa of Fabricius (IgM+ cells, macrophages), and spleen (IgM+ cells) of chicks after oral administration of a non-attenuated Salmonella (S.) typhimurium wild-type strain (infection) or an attenuated commercial live S. typhimurium vaccine strain (immunisation) to day-old chicks as compared to non-treated control birds using immunohistochemistry and image analysis. In caecum, higher counts of IgM-secreting cells were detected in infected animals compared with the controls from day 5 until day 12 of age. In contrast, in treated groups, IgA-secreting cells were found in higher numbers only between day 8 and 12 of age. Infected birds showed a higher number of IgA+ cells in spleen and bursa of Fabricius compared to the controls. In the bursa of Fabricius of immunised and infected birds, a depletion of strongly stained IgM+ cells and macrophages was established between day 5 and 9 indicating a possibly special and independent role of this organ during the immunological reaction against Salmonella organisms. The results suggest that IgM- and IgA-secreting cells are of importance in the caecal immune response of chickens against Salmonella strains. Immunised chickens always showed a weaker immune reaction compared to infected animals. Present findings regarding the B cell reaction within avian caeca prove a participation of both humoral and cellular immunity in defence against Salmonella strains. Immunohistochemical examination of the cellular response (B cells and macrophages) in relevant organs of chickens may be an important tool to evaluate the immunogenic characteristics of potential Salmonella live vaccine candidates.

Induction of T cell anergy by the treatment with IL-10-treated dendritic cells.

Abstract: In order to apply for reducing graft versus host disease in allogeneic stem cell transplantation, the study concerning the induction of specific T cell anergy was designed. Normal allogeneic lymphocytes, which were co-cultured with IL-10-treated immature dendritic cells in the first mixed leukocyte culture (MLC), were cultured with mature dendritic cells of the same origin as IL-10-treated immature dendritic cells in the second MLC. By co-culturing with IL-10-treated immature dendritic cells, the response of normal lymphocytes to mature dendritic cells cultured from the same individual as that of IL-10-treated dendritic cells was markedly reduced, compared with the lymphocytes cultured with non-treated dendritic cells or IL-10-treated dendritic cells from a third party individual. The present study demonstrated that antigen specific T cell anergy was generated by priming allogeneic lymphocytes with IL-10-treated immature dendritic cells. These data suggested the applicability of IL-10-treated recipient dendritic cells for the induction of recipient cell-specific donor T cell anergy in donor graft.

Experimental infection of goats at different stages of pregnancy with caprine herpesvirus 1

Abstract: Three goats from a group of five caprine herpesvirus 1 (CpHV.1) seronegative pregnant goats were inoculated intranasally with a virulent BA.1 strain of CpHV.1. Goat n.1 was infected on day 45 of pregnancy, goat n.2 on day 92 and goat n.3 on day 127. Each of the three goats produced a single foetus 10-60 days after infection. Foetus n.1 was never found and so it could not be examined for virological findings. Goat n.2 delivered at term of gestation and CpHV.1 was detected by PCR and isolated from most of the foetal organs. Foetus n.3 was partially autolysed and the virus was only detected by PCR but not isolated from foetal organs. The results confirm the damaging effect of CpHV.1 infection on pregnancy, the difficulty in diagnosing the CpHV.1 induced abortion, and the importance developing appropriate prophylactic programmes.

Protein A/G dependent ELISA a promising diagnostic tool in Lyme disease seroprevalence in game animals and hunting dogs.

Abstract: One of the major problems in serodiagnosis in wild animals is unavailability of specific antiglobulin conjugate. Our study focuses on validation of Protein A/G dependent ELISA in game animals like deer and mouflons as well as in hunting dogs. Binding ability of Protein A/G-conjugate to antibodies was the highest in dogs followed by fallow deer and mouflons. Three different whole cell Borrelia antigens were used to evaluate antigen dependent variation. In new Protein A/G-ELISA the highest sensitivities (90.50%, deer; 85.37%, mouflon & 94.29%, dog) were obtained by B. garinii antigen, with no statistically significant variation (χ 2 , P>0.05) among all other antigens used. Average seroprevalences observed in deer, mouflons and dogs were 44.90%, 29.41% and 30.43%, respectively. Marked influence of age on seroprevalence was noticed. Protein A/G – ELISA proved to be sensitive and promising diagnostic tool in serodiagnosis of Lyme disease in game ungulates and it can be used effectively for serosurvey in different wild mammals.

Characterisation of and immunity to the aerobic bacteria found in the pouch of the brushtail possum Trichosurus vulpecula

Abstract: The bacterial composition of the brustail possum (Trichosurus vulpecula) pouch was characterized throughout the reproductive cycle using brushtails from an Australian captive breeding colony (45 swabs) and a wild population in New Zealand (26 swabs). Gram-positive coccal species predominate throughout the reproductive cycle. Enteric Gram-negative rods, particularly Escherichia coli, were most prevalent when a pouch young was present and was most likely the result of faecal contamination from the pouch young. As culturing is only able to detect a proportion of bacteria present in a particular environment, molecular 16S rDNA sequencing was carried out on DNA extracted from a pouch wash of a female carrying a pouch young to gain a more accurate assessment of the pouch microflora. This approach identified approximately five times the number of bacterial species when compared to culture results. The majority detected were Gram negative rods or most closely related to Gram-negative rods species. Brushtails are immunologically immature at birth yet survive in a pouch colonised with potentially pathogenic bacteria. A haemagglutination assay was used to determine whether antibodies to a frequently isolated bacterium (Klebsiella pneumoniae) were transferred via milk from mother to pouch young. IgG antibodies were detected in maternal serum, milk and pouch young serum. In young over 70 days, antibody titres were significantly higher than those found in maternal serum, suggesting that the young is capable of producing adult type antibodies to pouch bacteria at this time.

Acute phase response in reindeer after challenge with Escherichia coli endotoxin

Abstract: The serum concentrations of two acute phase proteins (APPs), haptoglobin (Hp) and serum amyloid-A (SAA), were monitored in reindeer after challenge with endotoxin. Four adult female reindeer received either 0.1 mg/kg Escherichia coli 0111:B4 lipopolysaccharide B or saline solution intravenously. At the second challenge, the treatments were reversed. In addition to the APPs, changes in blood chemistry and rectal temperature were monitored. The endotoxin challenge caused a significant increase in SAA (peak 48 h) and a sharp decrease (8 ‐12 h) of serum iron concentrations in all animals. The mean Hp concentration increased at 8 h and remained elevated until 48 h, but no statistically significant differences were found. This investigation demonstrates that challenge with a single-bolus dose of E. coli endotoxin can activate the acute phase response (APR) and SAA appears to be a more sensitive indicator of the APR than Hp during bacterial infection in reindeer. q 2004 Elsevier Ltd. All rights reserved.

Cytotoxic T lymphocyte activity and cytokine expression in calves vaccinated with formalin-inactivated bovine respiratory syncytial virus prior to challenge.

Abstract: The development of effective, safe vaccines for human and bovine respiratory syncytial virus (RSV) has been problematic. Inactivated RSV vaccines are of variable efficacy; poor efficacy may be related to induction of ineffective cell-mediated immunity (CMI). To characterize CMI in calves vaccinated with formalin inactivated (FI) BRSV, 11 calves were vaccinated twice with FI-BRSV (n=5) or mock vaccine (n=6) at a 2 week interval and challenged 1 month later. Prior to challenge a cannula was placed in the efferent lymphatic of the caudal mediastinal lymph node of each calf; lymph derived lymphocytes (LDL) were collected for analysis of CMI. Cytotoxic T lymphocyte (CTL) activity by LDL and/or peripheral blood mononuclear cells (PBMC) was measured by 51Cr release on days 5, 7, 9, and 10 post-challenge. Messenger RNA for interferon gamma (IFN-gamma), interleukin 2 (IL-2) and IL-4 was measured on days 0-10 by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) of RNA of LDL. BRSV-specific IFN-gamma production by PBMC was measured on days 0 and 10 by ELISA. Clinical signs and postmortem changes following challenge were evaluated. There was no difference between groups in clinical signs, postmortem changes, CTL activity, cytokine message expression, or IFN-gamma production. For both groups, percentage lysis by CTL peaked on days 7-10 and ranged from 11 to 25%. Failure of vaccination to prevent disease following challenge was likely associated with failure to prime for improved CMI responses.

Studies on structural and immunogenic polypeptides of hydropericardium syndrome virus by SDS-PAGE and western blotting.

Abstract: The polypeptide pattern of a local isolate of a virus causing hydropericardium syndrome was analyzed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. A total of 12 polypeptides ranging in molecular weight between 13.8 and 110.0 kDa were observed. Western blot analysis of structural polypeptides revealed seven immunogenic polypeptides ranging in molecular weight between 15.8 and 110.0 kDa.

Comparative analysis of viral RNA and apoptotic cells in bursae following infection with infectious bursal disease virus.

Abstract: Specific-pathogen-free (SPF) chickens infected with very virulent (vv) infectious bursal disease virus (IBDV) UPM94/273 developed lower pathogenicity compared to UPM97/61. Sequence analysis indicated that UPM94/273 is an exceptional vvIBDV. In this study, a SYBR® Green I based real-time reverse transcriptase reaction assay was developed to measure viral RNA in the bursae of SPF chickens infected with IBDV. Specificity of the amplified products was confirmed by melting temperature analysis. A linear relationship was observed between the amount of input viral RNA and the threshold values for IBDV-specific product over five log10 dilutions. The viral RNA level following infection with UPM94/273 was significantly higher at day 1 and 2 post-inoculation (p.i.) compared to UPM97/61 infected chickens. However, chickens infected with UPM97/61 had significantly higher numbers of bursal cells undergoing apoptosis compared to UPM94/273 infected chickens. In both groups, the number of apoptotic cells and viral RNA levels peak at day 3 p.i. This study indicates that UPM97/61 and UPM94/273 have different efficiency of replication and percentage of apoptotic cells in bursae during the acute phase of IBDV infection.

Isolation of pathogenic Listeria monocytogenes and detection of antibodies against phosphatidylinositol-specific phospholipase C in buffaloes

Abstract: The isolation of pathogenic Listeria spp. in bacteriological samples, and anti-phosphatidylinositol-specific phospholipase C (anti-PIPLC) antibodies in sera of buffaloes were studied. Isolation of the pathogen was attempted from the samples by selective enrichment in University of Vermont Medium and plating onto Dominguez-Rodriguez isolation agar. Pathogenicity of the isolates was tested by Christie, Atkins, Munch Petersen test and mice incoulation test. Listeria spp. and L. monocytogenes were isolated from 8.8 and 2.4%, and 4.8 and 1.6% of 125 each meat and blood samples, respectively. Out of the 125 samples each of feacal, nasal and vaginal swabs from buffaloes 8 and 4%, 13.6 and 2.4%, and 6.4 and 2.4% were positive for Listeria spp. and L. monocytogenes, respectively. L. ivanovii was confirmed from 0.8% vaginal sample. A total of 125 serum samples were tested by phosphatidylinositol-specific phospholipase C (PIPLC) based indirect ELISA of which 4.0% turned out to be seropositive.

Prevention of tissue injury in an ascending mouse model of chronic pyelonephritis--role of free radical scavengers.

Abstract: The role of free radical scavengers in preventing the tissue injury using a non obstructive, ascending mouse model for chronic pyelonephritis was assessed. The parameters taken into consideration are Luminol Dependent Chemiluminescence (LDCL), histopathology and some biochemical investigations. We have observed that both catalase and Dimethyl-Sulfoxide (DMSO, free radical scavengers) were able to prevent the free radical mediated tissue injury and ultimate renal scarring, irrespective of the bacterial strain studied.

Polymerase chain reaction with a primer pair in the 16S-23S rRNA spacer region for detection of Mycoplasma pulmonis in clinical isolates.

Abstract: To develop a diagnostic tool to identify Mycoplasma pulmonis (M. pulmonis) in clinical isolates, we developed a polymerase chain reaction (PCR) assay using primers specific for the 16S-23S rRNA intergenic spacer region (SR) of M. pulmonis. One pair of PCR primers reacted specifically with two reference strains of M. pulmonis tested and seven samples isolated from naturally infected rats. The primer pair did not produce PCR products of the correct size from any other rodent or human mycoplasmas or cellular DNA from rodent lungs. Specificity of the PCR assay was confirmed by Southern blotting with probe specific for the SR of M. pulmonis. The PCR assay for detection of M. pulmonis established in this study is suitable for diagnosis of M. pulmonis infection in clinical cases.

Immunization against bovine herpesvirus-1 infection. Preliminary tests in calves with a DNA vaccine.

Abstract: A bovine herpesvirus-1 (BHV-1) vaccine expressing glycoprotein D, the form with the transmembrane anchor removed, was evaluated for inducing immunity in calves. The plasmid encoding gD of BHV-1 was injected three times to nine calves, using three animals for each of the following routes: intramuscularly (i.m.), intradermally (i.d.), or intranasally (i.n.). Three additional calves were given the plasmid vector only and served as unvaccinated controls. When calves were subjected to challenge infection with BHV-1, all vaccinated calves as well as the controls developed a typical severe form of infectious bovine rhinotracheitis. However, compared to the controls, the vaccinated calves showed earlier clearance of challenge virus. Moreover, the calves given the vaccine i.m. developed neutralizing antibody to BHV-1 between 21 and 42 days following the first injection of vaccine, whereas in calves vaccinated either i.d. or i.n., as well as the controls, antibody first appeared in their sera 14 days post-challenge infection.

Soft-tissue abscess involving Actinomyces odontolyticus and two Prevotella species in an intravenous drug abuser.

Abstract: Skin and soft-tissue infections in intravenous users comprise a variety of microorganisms and anaerobic bacteria are frequently involved in these suppurative infections. A case of subcutaneous abscess into anterior femoral muscles involving Actinomyces odontolyticus and two Prevotella species (Prevotella buccae and Prevotella melaninogenica) in an intravenous drug abuser is presented. This combination of microorganisms has not previously been described in soft-tissue infections. The patient volunteering that he licked his hypodermic needle prior to cocaine injection supports that the implicating bacteria originated from the oral cavity. Eventually, the patient recovered and at a 6-month follow-up a gradual improvement of his subcutaneous infection was noticed.

Prevalence of antibodies against Saint Louis encephalitis and Jamestown Canyon viruses in California horses

Abstract: Jamestown Canyon (JC) and Saint Louis encephalitis (SLE) viruses are mosquito-transmitted viruses that have long been present in California The objective of this study was to determine the seroprevalence of these two viruses in horses prior to the introduction of West Nile (WN) virus Approximately 15% of serum samples collected in 1998 from 425 horses on 44 equine operations horses throughout California had serum antibodies to JC virus, whereas antibodies were not detected to SLE virus The results indicate that horses in California were commonly infected prior to 1998 with mosquito-transmitted Bunyaviruses that are identical or closely related to JC virus, but not with SLE virus The different seroprevalence of SLE and JC viruses in horses likely reflects the unique ecology of each virus, and it is predicted that WN virus will have a wider distribution in California than closely related SLE virus

Characterization of specific antibodies and the establishment of sandwich ELISA and ELISPOT systems for swine IL-4.

Abstract: We produced four monoclonal antibodies (mAb) and two polyclonal antibodies using the purified cytokine expressed in bacteria and characterized them. Specific binding of each of the mAb and polyclonal antibodies to recombinant swine IL-4 (rSwIL-4) purified from Escherichia coli and baculovirus was demonstrated in an indirect ELISA and/or in western blotting. We established a sandwich enzyme-linked immunosorbent assay (ELISA) for measuring concentration of SwIL-4 in biological samples and established an enzyme-linked immunospot (ELISPOT) assay for detecting IL-4-secreting cells using a mAb and a polyclonal IgG from goat. The detection limit of the sandwich ELISA for SwIL-4 was 78 pg/ml. Using sandwich ELISA, SwIL-4 was detected in the bronchoalveolar lavage fluid (BALF) of pigs experimentally infected with Mycoplasma hyopneumoniae and could quantitate in supernatants of mitogen-stimulated PBMC culture. The ELISPOT system is useful for the detection of IL-4 producing cells in swine PBMC culture.

Development of a sandwich-dot enzyme linked immunosorbent assay for Streptococcus pneumoniae antigen detection in cerebrospinal fluid.

Abstract: A new sandwich dot–enzyme linked immunosorbent assay (sdot–ELISA) was developed using omniserum prepared against different strains of Streptococcus pneumoniae as capture antibody and also as second or revealing antibody after its conjugation with horseradish peroxidase (HRP) for detection of pneumococcal antigen in cerebrospinal fluid (CSF). A total of 103 CSF samples of different categories were screened with newly developed dot–ELISA and results were compared with commercially available latex agglutination (LA) kit. The newly developed sdot–ELISA was more sensitive than LA test and can be used as an alternative diagnostic tool in laboratory and in field conditions. An added advantage of this ELISA system was that it did not require antibodies produced in two different animal species.