Tactics of Mycobacterium avium subsp. paratuberculosis for intracellular survival in mononuclear phagocytes

Johne's disease is a condition that refers to chronic granulomatous enteritis in ruminants. It is believed that survival and replication of Mycobacterium (M.) paratuberculosis in mononuclear phagocytes plays an important role in the pathogenesis of Johne's disease. However, it is not clear how M. paratuberculosis survives for long time periods in mononuclear phagocytes, nor is it clear which factors trigger multiplication of these bacilli and result in the development of Johne's disease. Investigating the intracellular fate of M. paratuberculosis is challenging because of its very slow growth (more than two months to form visible colonies on media). Existing animal models also have limitations. Despite those obstacles, there has been progress in understanding the intracellular survival tactics of M. paratuberculosis and the host response against them. In this review, we compare known aspects of the intracellular survival tactics of M. paratuberculosis with those of other mycobacterial species, and consider possible mycobactericidal mechanisms of mononuclear phagocytes.

Cell proliferation and interferon-gamma response to recombinant MBP-3, NarL, MT-10.3, and 16kDa Mycobacterium tuberculosis antigens in Brazilian tuberculosis patients

Human pulmonary tuberculosis (TB) is a worldwide public health problem. In resistant individuals, control of the infection mainly requires development of a Th1 cell immune response with production of cytokines, of which interferon-gamma (IFN-gamma)plays an important role. Several antigens from Mycobacterium tuberculosis complex has been described for use in vaccine development or for diagnostic purposes, however little evaluation has been done in endemic area for TB. The proliferative and IFN-gamma human T cell immune responses, to four recombinant proteins (MBP-3, NarL, MT-10.3, 16 kDa) and PPD, of 38 Brazilian TB patients (6 untreated and 32 treated) and 67 controls (38 positive and 29 negative tuberculin skin test - TST) were compared. The highest reactivity mean rate was obtained with PPD followed by 16 kDa in TB patients. While most of the patients (87 percent) and controls (> 64 percent) respond to the PPD, 16kDa was more specifically recognized (> 21 percent) although less sensitive (54 percent). When TB patients were divided according to treatment status, opposite to PPD, higher average level of IFN-gamma was induced by 16kDa in untreated (505 pg/ml) compared to treated TB patients and TST+ (269.8 pg/ml x 221.6pg/ml, respectively), although the difference was not significant. These data show that in contrast with the other recombinant proteins, the stimulatory potency of 16kDa to induce proliferative and INF-gamma response was more effective and is more recognized by active TB untreated patients, eliciting in control individuals a more selective immune response than PPD.

Rapid detection of Chlamydia pneumoniae DNA in peripheral blood mononuclear cells of coronary artery disease patients by real-time fluorescence PCR.

Several recent reports including serological, pathological and animal studies have associated Chlamydia pneumoniae with coronary artery disease (CAD). In order to establish whether chronic C. pneumoniae infection is linked to coronary artery disease, clinical intervention trials may be needed. However, to detect eligible patients with persistent infection, a reliable diagnostic marker must be developed for identifying cases and assessing efficacy of antichlamydial therapy. Moreover, the prevalence of circulating C. pneumoniae DNA in CAD patients varied widely from previous reports. A real-time PCR has been established by using HL-1 and HR-1 primer to amplify 437 base pairs product. Confirmation of the product was performed on LightCycler by melting curve analysis of detection probes labeled with LC-Red705. Ninety-five angiographically confirmed CAD patients and 104 normal, healthy volunteers were recruited. The mononuclear cell layer was separated from collected blood and rapid, single step real-time PCR was used to detect C. pneumoniae DNA. C. pneumoniae DNA in peripheral blood mononuclear cells (PBMC) was found in 17 per cent of 95 CAD patients and 1 per cent of 104 normal healthy volunteers (odds ratio 20.86, 95% confidence interval 2.71 - 160.67, p < 0.0001). There was no association between C. pneumoniae DNA in PBMC and serological status. The rapid, real-time PCR showed a clear-cut result between positive and negative cases. PBMC-based real-time PCR may be a useful tool for identifying subjects carrying C. pneumoniae in the circulation or in the vascular wall as well. It will be a specific indicator of current infection and will be used as a marker for assessing the microbiological efficacy of antichlamydial therapy in clinical intervention trials.

Rapid detection of salmonella typhi by culture of the mononuclear cell-platelet fraction of blood

To study the value of utilizing a new and rapid method for detecting Salmonella typhi in adult patients using the mononuclear cell-platelet fraction of blood and to compare it with the traditional methods used for the same purpose. Materials and Various specimens [blood, bone marrow and rectal swabs] were collected and cultured on standard media from 36 Jordanian patients with suspected typhoid fever at two major Jordanian hospitals, in the period from July 1992 through July 1993. Cultures using the mononuclear cell-platelet layer were performed on the blood specimens taken from all patients. Blood cultures using the mononuclear cell-platelet layer method were positive in 20 [56%] patients, and the colonies were identified [using a coagglutination technique] within 18 hours of plating. In contrast, Salmonella typhi was detected in only 15 [41%] patients using the conventional blood culture method, that required at least 3 days for identification. This study indicates that the combination of mononuclear cell-platelet layer culture and coagglutination can provide the clinician with a diagnosis of typhoid fever within a day of specimen acquisition, with a marked improvement over conventional blood culture in both time and sensitivity

Recovery of human immunodeficiency virus from asymptomatic prostitutes from Tamil Nadu.

Peripheral blood mononuclear cells (PBMC) from 13 asymptomatic healthy human immunodeficiency virus type-1 (HIV-1) antibody positive prostitutes from Tamil Nadu, southern India, were cocultivated with phytohemagglutinin stimulated PBMC from HIV antibody negative donors for HIV isolation. In addition, plasma samples from two antibody positive prostitutes with HIV antigenemia were processed for virus isolation. The presence of virus in the cultures was monitored by (i) assay for virus particle associated reverse transcriptase (RT) activity, (ii) HIV-antigen enzyme immunoassay, and (iii) indirect immunofluorescence test to detect expression of HIV specific core antigens p-24 and p-17 in infected cells using monoclonal antibodies to these antigens. The virus was isolated from PBMC from 2 prostitutes (86-4 and 86-5) and from plasma of one prostitute (86-20). These isolates have been characterised as HIV type-1 by dot blot hybridization using HIV-1 and HIV-2 proviral DNA probes.

In vitro HIV infection of primate lymphocytes

El propósito de este estudio fue el de evaluar la capacidad de los virus del SIDA (VIH-1 y VIH-2) para multiplicarse en las células mononuclearres de la sangre periférica (CMSP) de cuatro especies de primates. CMSP de Cebus apella, patas (Erythrocebus patas), monos verdes (cercopithecus aethiops sabeus) y rhesus (Macaca mulatta) fueron infectados "in vitro" con VIH-1 y con VIH-2. La multiplicación de estos virus se determinó midiendo la actividad de la enzima retrotranscriptasa en los cultivos infectados. Ambos virus produjeron efectos citipáticos en dichos cultivos. Se observó un bajo nivel de multiplicación de los virus VIH-1 y VIH-2 en las células provenientes de monos Cebus. Sin embargo, el virus VIH-2 se multiplicó eficientemente en CMSP de monos rhesus. La capacidad que posee el virus de la inmunodeficiencia humana tipo 2, (VIH-2) de multiplicarse en estas células, podría ser utilizada para en la evaluación "in vivo" de productos antivirales y de vacunas