Disseminated Mycobacterium genavense infection: clinical and microbiological features and response to therapy

OBJECTIVE: Mycobacterium genavense is a newly described pathogen that causes disseminated infection in AIDS. It is difficult to detect and identify due to its slow growth and fastidious nature. There is little information available about therapy for this new pathogen. We describe clinical and laboratory features and response to therapy in four patients with advanced AIDS complicated by disseminated M. genavense infection from Denver, Colorado, USA. DESIGN AND METHODS: Retrospective analysis of four cases identified in an AIDS clinic affiliated with a municipal hospital in Denver, Colorado. Clinical samples were inoculated onto BACTEC 12B, Lowenstein-Jensen, and Middlebrook 7H11 media. RESULTS: The clinical features mimicked those of disseminated M. avium complex infection, with invasion of liver, spleen and lymph nodes with acid-fast bacilli (AFB). Acid-fast smears of blood and lymph nodes were positive; there was a modest increase in the growth index in BACTEC broth and tiny colonies appeared on Middlebrook agar. Patients were treated with combinations of antimycobacterial agents. Blood smears and cultures reverted to negative in treated patients. The best clinical response was associated with clarithromycin therapy. CONCLUSIONS: Disseminated disease due to M. genavense should be suspected among patients with the clinical presentation of disseminated M. avium complex infection and low growth index on BACTEC cultures for AFB. The diagnosis of M. genavense may be facilitated by performing acid-fast stains of samples from BACTEC bottles in such individuals. Clarithromycin therapy is associated with clinical improvement and clearance of bacteremia.     

Monocyte chemotactic peptide-1 expression and monocyte infiltration in acute renal transplant rejection

Disseminated infection caused by organisms of Mycobacterium avium complex is common in acquired immune deficiency syndrome (AIDS) patients. M. avium is an intracellular bacterium that multiplies within macrophages. We examined the effect of M. avium infection on the T-helper cell response in C57/BL/6 black mice. At weekly intervals, CD4+ T-cells were isolated from spleens and lines were created. T-cell lines were exposed to sonicated M. avium in the presence of feeder cells and macrophages and the supernatant were collected to measure the concentrations of interferon-gamma (IFN-gamma and interleukin-10 (IL-10). Production of IFN-gamma in CD4+ T-cells obtained from uninfected mice did not vary significantly during the 5 weeks. Levels of IFN-gamma produced by T-cell lines of infected mice were similar to the control mice during the first 2 weeks but significantly reduced (approximately 30 ng/ml) thereafter. In contrast, production of IL-10 by T-cell lines of infected mice was in a range of 190 to 342 pg/ml in weeks 1, 2 and 3, but increased to an average of 1300 pg/ml at weeks 4 and 5. Pre-immunized mice, when infected with M. avium strain 101, showed a different profile of T-cell cytokines, with high IFN-gamma and low IL-10 production. Proteins purified from a number of disease-associated (D-A) and non-D-A strains of M. avium were tested for the ability to induce IL-10. 65,000 MW and 60,000 MW proteins of M. avium induced significantly more IL-10 than 45,000 MW, 33,000 MW and 27,000 MW proteins. These results showed that M. avium predominantly stimulates either Th1 or Th2 T-helper cells according to the phase of the infection.     

Role of iron in the pathogenesis of Mycobacterium avium infection in mice

Mycobacterial infections are of serious concern to HIV-infected patients, and take a heavy toll of such patients. Mycobacterium avium is the most common opportunistic bacterial infection in patients with AIDS. The overload of iron in serum has been implicated in the pathogenicity of a number of bacterial infections. Since iron storage in cells such as macrophages is increased in AIDS, the role of iron as a possible factor in the pathogenesis of M. avium infection was examined. Supplementing iron to normal laboratory chow resulted in accelerated M. avium infection in mice inoculated earlier with the same organism. The bacterial loads in liver, spleen and lungs were approximately 12-fold higher in mice receiving iron supplementation compared with control groups. This is attributed to an increased percentage saturation of iron in the sera of the mice, thus making more iron available for the replication of bacteria. The addition of beef fat to the diet, together with high iron supplementation, further enhanced the infection. Using smaller inocula, mice receiving chow supplemented with high iron and fat developed disseminated M. avium infection faster than control mice. The results provide strong evidence that iron may play a major role in the pathogenesis of M. avium infection.     

A phase II/III trial of antimicrobial therapy with or without amikacin in the treatment of disseminated Mycobacterium avium infection in HIV-infected individuals. AIDS Clinical Trials Group Protocol 135 Study Team

OBJECTIVE: To determine the clinical and microbiologic benefit of adding amikacin to a four-drug oral regimen for treatment of disseminated Mycobacterium avium infection in HIV-infected patients. DESIGN: A randomized, open-labeled, comparative trial. SETTING: Outpatient clinics. PATIENTS: Seventy-four patients with HIV and symptomatic bacteremic M. avium infection. INTERVENTIONS: Rifampin 10 mg/kg daily, ciprofloxacin 500 mg twice daily, clofazimine 100 mg every day, and ethambutol 15 mg/kg orally daily for 24 weeks, with or without amikacin 10 mg/kg intravenously or intramuscularly 5 days weekly for the first 4 weeks. MAIN OUTCOME MEASURE: Clinical and microbiologic response at 4 weeks; quantitative level of bacteremia with M. avium. RESULTS: No difference in clinical response was noted with the addition of amikacin to the four-drug oral regimen, and only 25% in either group had a complete or partial response at 4 weeks. A comparable quantitative decrease in bacteremia was noted in both treatment groups, with 16% of patients being culture-negative at 4 weeks and 38% at 12 weeks. Toxicities were mainly gastrointestinal. Amikacin was well tolerated. Median survival was 30 weeks in both groups. CONCLUSIONS: The addition of amikacin to a four-drug oral regimen of rifampin, ciprofloxacin, clofazimine, and ethambutol did not provide clinical or microbiologic benefit.     

Thirteen bovine microsatellite markers that are polymorphic in sheep

Macrolide resistance in disseminated Mycobacterium avium infection is of major concern in AIDS patients as these drugs represent the main component of combination therapy. Clarithromycin and azithromycin should not be used alone for the treatment and prophylaxis of the disease because of the risk of selecting resistant strains. We report the case of a clarithromycin resistant disseminated M. avium infection in an AIDS patient, acquired after long term monotherapy with clarithromycin for the treatment of cryptosporidiosis.     

Intermittent azithromycin for treatment of Mycobacterium avium infection in beige mice

The activity of azithromycin (AZI) was evaluated in the beige mouse model of disseminated Mycobacterium avium infection. Mice were infected intravenously with approximately 10(7) viable avium ATCC 49601. AZI at 50, 100, or 200 mg/kg of body weight or clarithromycin (CLA) at 200 mg/kg was given by gavage 5 days per week for 4 weeks. Groups of treated mice were compared with untreated control animals. A dose-related reduction in cell counts in organs was observed with AZI treatment. AZI at 200 mg/kg was more active than CLA at 200 mg/kg against organisms in spleens. The activities of these two agents at 200 mg/kg were comparable against organisms in lungs. In a second study, AZI at 200 mg/kg was given daily for 5 days; this was followed by intermittent AZI treatment for the next 3 weeks. The activities of AZI given on a three-times- and five-times-per-week basis in the continuation phase were comparable. AZI given on a once-weekly basis was less active. The regimen of AZI given in combination with rifapentine on a once-weekly basis for 8 weeks showed promising activity. Clinical evaluation of AZI and rifapentine will help to define the roles of these agents in the treatment of disseminated M. avium complex infection.     

Inhibition of tumor necrosis factor alpha alters resistance to Mycobacterium avium complex infection in mice

Increased production of tumor necrosis factor alpha (TNF-alpha) appears to play an important role in the progression of human immunodeficiency virus disease. One treatment strategy being explored is the use of TNF-alpha inhibitors. TNF-alpha also appears to be important in conferring resistance to infections, and the inhibition of this cytokine may exacerbate the emergence of opportunistic pathogens, such as Mycobacterium avium complex (MAC). The present study examines the possibility that inhibition of TNF-alpha will increase the progression of disease in mice infected with MAC. C57BL/6 beige (bg/bg) mice have been shown to be highly susceptible to infection with MAC and are routinely used for testing of antimycobacterial drugs. However, bg/bg mice are known to exhibit impaired phagocyte and natural killer cell function. Since these cell types are important sources of TNF-alpha, the susceptibility of the bg/bg strain to infection with MAC was compared with those of the heterozygous (bg/+) and wild-type (+/+) strains of C57BL/6 mice. The susceptibilities of the bg/bg and bg/+ strains of mice infected with MAC were found to be comparable. The +/+ strain was the least susceptible. Mycobacterial burden and serum TNF-alpha levels increased over time in all the strains of mice tested. The bg/+ strain of C57BL/6 mice was then chosen to measure the activity of TNF-alpha antagonists. Treatment with dexamethasone decreased serum TNF-alpha levels and increased mycobacterial burden. Treatment with anti-TNF-alpha antibody or pentoxifylline did not significantly alter serum TNF-alpha levels but increased mycobacterial burden. Treatment with thalidomide neither consistently altered mycobacterial burden in the spleens or livers of infected mice nor affected serum TNF-alpha levels.     

Cryptosporidium parvum infection of human intestinal xenografts in SCID mice induces production of human tumor necrosis factor alpha and interleukin-8

The protozoan parasite Cryptosporidium parvum invades intestinal epithelial cells and can cause life-threatening diarrhea in immunocompromised individuals. Despite the clinical importance of this organism, much remains to be learned about the pathogenesis of C. parvum-induced diarrhea. To explore the role of the intestinal inflammatory response in C. parvum disease, using C. parvum oocysts we infected human intestinal xenografts in severe combined immunodeficient (SCID) mice. Seven days after infection, we found levels of human tumor necrosis factor alpha and interleukin-8 in C. parvum-infected human intestinal xenografts that were significantly higher than those seen in uninfected control xenografts. These results demonstrate that human intestinal cells produce proinflammatory cytokines in response to C. parvum infection and establish SCID-HU-INT mice as a model system to study the interactions of C. parvum with the human intestine.     

Mycobacterium genavense infection presenting as a solitary brain mass in a patient with AIDS: case report and review

Patients with AIDS are prone to developing infections with opportunistic pathogens. Recently, a new mycobacterium, Mycobacterium genavense, has been found to cause infection in patients with AIDS. Previously published reports indicate that patients who are infected with this organism present with the same clinical features as do patients with disseminated infection due to organisms of the Mycobacterium avium complex. We describe an unusual case of a patient with AIDS who presented with grand mal seizures and a mass lesion in his brain, which was found to be caused by infection with M. genavense. No evidence of disseminated infection could be found in this patient. We discuss the microbiology of this organism and review the literature on M. genavense infections. Clinicians should be aware of this organism so that efforts at culture and identification will be made.     

Prevalence of gallstones in the neonatal period]

The prevalence of infection with Mycobacterium avium complex (MAC) has increased since the outbreak of the HIV pandemic. This complex comprises two organisms: M. avium (mostly) and M. intracellulare (rarely). The source of MAC infection is not known. The principal risk factors for disseminated MAC infection in a patient with HIV infection are a low CD4 count and a previous opportunistic infection. The symptoms of disseminated MAC infection resemble those of HIV wasting; a positive culture of normally sterile tissue confirms a MAC infection. There is reserve with regard to routine prophylaxis in HIV-infected persons because of the possible development of resistance, interaction with other drugs used in AIDS, toxicity and possible absorption disorders which might cause prophylaxis to fail. For the treatment of disseminated MAC infection, a combination of at least two medicaments (macrolides and ethambutol) is recommended.     

Intestinal and systemic humoral immunological events in the susceptible Balb/C mouse strain after oral administration of Echinococcus multilocularis eggs

The aim of the study was to investigate the systemic and, for the first time, the intestinal humoral events in the susceptible Balb/C mouse strain after oral administration of Echinococcus multilocularis eggs. Thirty-one mice were divided into three groups; W-2, W-8 and control group. Each mouse of the W-2 and W-8 groups was orally infected with 1,500 E. multilocularis eggs, two weeks and eight weeks before sacrifice respectively. Control group mice received phosphate buffer saline. Measurement of anti-E. multilocularis and non-specific IgG, IgA and IgM, and of a transudation marker, albumin, were performed in serum and intestinal washings by a time-resolved immunofluorometric assay. These results were complemented by microscopic examination of the intestinal mucosa. This infection model is well-suited to the study of mucosal immunity during alveolar echinococcosis. It showed a major specific intestinal response in the early stage of the disease whereas the systemic response predominated later in the disease. Histopathological studies and calculation of the relative coefficient of excretion of Ig also confirmed that the presence of the parasite, even during a short period, was responsible for a local immunological and inflammatory response and for a change in mucosal permeability. Mucosal immunity could thus play a role in tolerance induction against E. multilocularis that could be a prerequisite for the subsequent development of the larvae in the liver, and for the occurrence of the parasitic disease, alveolar echinococcosis.     

Isolation and characterization of a 70 kDa protein from Mycobacterium avium

Mycobacterium avium complex (MAC) is an intracellular pathogen which causes disseminated bacterial infection in immunocompromised individuals. This organism predominantly infects macrophages. Attachment of MAC to macrophages is the first step prior to invasion. We have previously shown that a 70 kDa protein of M. avium (Ma) is one of nine monocyte-binding proteins. In the present study, we have purified this protein from sonic extracts of Ma and studied some of its properties. The N-terminal sequence of this protein was identified and found to exhibit a strong homology to the 70 kDa heat shock protein (hsp) of M. leprae (Ml) and M. tuberculosis (Mtb). This protein was found to be present on the surface of the organism and was able to inhibit the attachment of intact Ma to human monocyte derived macrophages (MDM) up to 49% in an in vitro attachment assay using intact fluorescein isothiocyanate (FITC)-labelled Ma. Bovine serum albumin (BSA) and recombinant 70 kDa hsp from Mtb, which were used as controls, inhibited this attachment by 9.8 and 18%, respectively. These results suggest that the 70 kDa protein may have a role in the attachment of intact Ma to MDM. When tested in lymphocyte activation assays, this protein did not appear to significantly stimulate proliferation. However, it was found to stimulate the production of tumor necrosis factor (TNF)-alpha by MDM. This protein may be one of several Ma antigens that trigger host immune response by binding to MDM and stimulating the production of inflammatory cytokines such as TNF-alpha by these cells.     

Differentiation between Mycobacterium tuberculosis and Mycobacterium avium by amplification of the 16S-23S ribosomal DNA spacer

Differentiation between Mycobacterium tuberculosis and M. avium is helpful for the treatment of disseminated mycobacterial infection in AIDS patients. This can traditionally be done by time-consuming biochemical tests or with Accuprobe. Previously, PCR restriction enzyme analysis (PCR-REA) of the 16S-23S rRNA gene spacer was shown to be able to identify a limited number of strains of Mycobacterium. In this study the method was improved by using more specific primers and was tested with 50 clinical isolates of M. tuberculosis and 65 clinical isolates of M. avium complex. Probes specific to the spacers of M. tuberculosis and M. avium were also tested. Both M. tuberculosis and M. avium could be reliably identified either by PCR-REA or by PCR-hybridization, with the results completely agreeing with those obtained by biochemical tests and with the Accuprobe, respectively. The method may therefore be useful as an alternative in-house method for identification of the bacteria.     

Animal model of mucosally transmitted human immunodeficiency virus type 1 disease: intravaginal and oral deposition of simian/human immunodeficiency virus in macaques results in systemic infection, elimination of CD4+ T cells, and AIDS

Chimeric simian/human immunodeficiency virus (SHIV) consists of the env, vpu, tat, and rev genes of human immunodeficiency virus type 1 (HIV-1) on a background of simian immunodeficiency virus (SIV). We derived a SHIV that caused CD4+ cell loss and AIDS in pig-tailed macaques (S. V. Joag, Z. Li, L. Foresman, E. B. Stephens, L. J. Zhao, I. Adany, D. M. Pinson, H. M. McClure, and O. Narayan, J. Virol. 70:3189-3197, 1996) and used a cell-free stock of this virus (SHIV(KU-1)) to inoculate macaques by the intravaginal route. Macaques developed high virus burdens and severe loss of CD4+ cells within 1 month, even when inoculated with only a single animal infectious dose of the virus by the intravaginal route. The infection was characterized by a burst of virus replication that peaked during the first week following intravenous inoculation and a week later in the intravaginally inoculated animals. Intravaginally inoculated animals died within 6 months, with CD4+ counts of <30/microl in peripheral blood, anemia, weight loss, and opportunistic infections (malaria, toxoplasmosis, cryptosporidiosis, and Pneumocystis carinii pneumonia). To evaluate the kinetics of virus spread, we inoculated macaques intravaginally and euthanized them after 2, 4, 7, and 15 days postinoculation. In situ hybridization and immunocytochemistry revealed cells expressing viral RNA and protein in the vagina, uterus, and pelvic and mesenteric lymph nodes in the macaque euthanized on day 2. By day 4, virus-infected cells had disseminated to the spleen and thymus, and by day 15, global elimination of CD4+ T cells was in full progress. Kinetics of viral replication and CD4+ loss were similar in an animal inoculated with pathogenic SHIV orally. This provides a sexual-transmission model of human AIDS that can be used to study the pathogenesis of mucosal infection and to evaluate the efficacy of vaccines and drugs directed against HIV-1.     

A case of AIDS with disseminated Mycobacterium kansasii infection in which Mycobacterium, avium complex was also detected from his sputum repeatedly

A 43 year-old Japanese male was admitted to our hospital because of productive cough and fever. He was diagnosed as acquired immunodeficiency syndrome (AIDS) in 1994. Laboratory findings were as follows: WBC was 3200/microliter, CD4+ T lymphocyte count was 22/microliter. His chest X-ray film taken on admission showed infiltration with small cavity lesion in middle left lung field. Tuberculin skin reaction was negative. He was treated with isoniazid 0.4 g, rifampicin 0.45 g, and ethambutol 0.75 g each daily. Sputum smear was positive for acid fast bacilli. The cultured isolates were identified as Mycobacterium kansasii (M. kansasii) and Mycobacterium avium complex (MAC). Urine smear was also positive for acid fast bacilli. The cultured isolates were identified as M. kansasii. He was diagnosed as disseminated M. kansasii infection and suspected MAC infection. About one hundred days later, his chest X-ray film showed reticular shadow. His clinical symptoms improved and the sputum smear and culture converted to negative for acid fast bacilli. Based on these findings, his MAC discharge was considered not as MAC infection, but MAC colonization. He returned to the former hospital for AIDS treatment, and he died in August 1996.     

The inflammatory response to nonfatal Sindbis virus infection of the nervous system is more severe in SJL than in BALB/c mice and is associated with low levels of IL-4 mRNA and high levels of IL-10-producing CD4+ T cells

SJL mice are susceptible to inflammatory autoimmune diseases of the central nervous system (CNS), while BALB/c mice are relatively resistant. To understand differences in immune responses that may contribute to autoimmune neurologic disease, we compared the responses of SJL and BALB/c mice to infection with Sindbis virus, a virus that causes acute nonfatal encephalomyelitis in both strains of mice. Clearance of virus was similar, but SJL mice developed a more intense inflammatory response in the brain and spinal cord and inflammation persisted for several weeks. Analysis of lymphocytes isolated from brains early after infection showed an absence of NK cells in SJL mice, while both strains of mice showed CD4+ and CD8+ T cells. During the second week after infection, CD4+ T cells increased in SJL mice and the proportion of CD8+ T cells decreased, while the opposite pattern was seen in BALB/c mice. Expression of IL-10 mRNA was higher and IL-4 mRNA was lower in the brains of infected SJL than in BALB/c mice, while expression of the mRNAs of IL-6, IL-1beta, TNFalpha, and the Th1 cytokines IL-2, IL-12, and IFN-gamma was similar. Lymphocytes isolated from the CNS of SJL mice produced large amounts of IL-10. CNS lymphocytes from both strains of mice produced IFN-gamma in response to stimulation with Sindbis virus, but not in response to myelin basic protein. These data suggest that IL-10-producing CD4+ T cells are differentially recruited to or regulated within the CNS of SJL mice compared with BALB/c mice infected with Sindbis virus, a characteristic that may be related to low levels of IL-4, and is likely to be involved in susceptibility of SJL mice to CNS inflammatory diseases.     

Disseminated Mycobacterium genavense infection in Canadian AIDS patients

Mycobacterium genavense is a recently described mycobacterial species which thus far has been identified only in persons with advanced HIV disease. It appears to be a rare pathogen with an undefined reservoir. We describe the first two cases of M. genavense infection in Canadian AIDS patients. The clinical presentation of fever and wasting with extremely low CD4 lymphocyte counts was indistinguishable from disseminated M. avium complex (MAC) infection. However, blood cultures in BACTEC 13A medium required a mean of 58 days (range 41-87) to detect growth of M. genavense in contrast to a mean of 10 days for MAC in our laboratory. M. genavense infection is underdiagnosed due to the lack of universal use of BACTEC liquid medium and the use of relatively short incubation times (only 6 weeks) by some laboratories. The value of antimycobacterial therapy for M. genavense is unknown, but anecdotal data suggest that treatment with a regimen appropriate for MAC may be beneficial.     

Bovine T cell responses to Cryptosporidium parvum infection

To better understand the immune mechanisms important for clearing of the primary infection and the subsequent development of resistance to Cryptosporidium parvum infection, several groups have recently characterised changes within the lymphoid cell population of the intestinal mucosa and associated lymphoid tissue in calves with cryptosporidiosis. In naive animals, infection results in a significant increase in the number of CD4+ and CD8+ T cells present within the intraepithelial lymphocyte population, lamina propria and Peyer's patch of the ileum. This is accompanied by a rapid and transient increase in the number of gamma/delta T cells present within the intestinal villi. In response to a challenge infection in immune calves, there is a substantial increase in the number of CD4+ T cells present in the Peyer's patch of the ileum and a specific localization of CD8+ T cells to the epithelium of the intestinal villi. Together, these data demonstrate that C, parvum elicits a strong cell-mediated response following both primary and secondary infections in calves, and that CD8+ T cells may play an important role in the bovine immune response to C. parvum infection.     

Effect of anesthetic technique on cardiac morbidity following carotid artery surgery

Clarithromycin and rifabutin are among the most promising drugs for the therapy of infections caused by Mycobacterium avium or other atypical mycobacteria. Since synergism of combined drugs is important in order to achieve strong antimycobacterial activity, the combined inhibitory effects of antibacterial agents should also be investigated when agents are evaluated for possible use in antimycobacterial drug therapy. In the present study we examined the antimycobacterial activity of clarithromycin, rifabutin, and their combination against 51 clinical isolates of the M. avium complex from patients with acquired immune deficiency syndrome (AIDS) with disseminated mycobacteriosis. A concentration-dependent inhibition was seen for each drug. The antibacterial effect was significantly more pronounced for the combined drugs than for the agents tested separately. Synergism, against up to 88% of the strains tested, was seen for the tested drugs combined at different concentrations. All 51 M. avium strains were susceptible to the combination of 4 mg/l clarithromycin and 2 mg/l rifabutin.