A standardized molecular test for the detection of Chlamydophila pneumoniae DNA in cerebrospinal fluid (CSF) would assist the further assessment of the association of C. pneumoniae with multiple sclerosis (MS). We developed and validated a qualitative colorimetric microtiter plate-based PCR assay (PCR-EIA) and a real-time quantitative PCR assay (TaqMan) for detection of C. pneumoniae DNA in CSF specimens from MS patients and controls. Compared to a touchdown nested-PCR assay, the sensitivity, specificity, and concordance of the PCR-EIA assay were 88.5%, 93.2%, and 90.5%, respectively, on a total of 137 CSF specimens. PCR-EIA presented a significantly higher sensitivity in MS patients (p = 0.008) and a higher specificity in other neurological diseases (p = 0.018). Test reproducibility of the PCR-EIA assay was statistically related to the volumes of extract DNA included in the test (p = 0.033); a high volume, which was equivalent to 100 microl of CSF per reaction, yielded a concordance of 96.8% between two medical technologists running the test at different times. The TaqMan quantitative PCR assay detected 26 of 63 (41.3%) of positive CSF specimens that tested positive by both PCR-EIA and nested-PCR qualitative assays. None of the CSF specimens that were negative by the two qualitative PCR methods were detected by the TaqMan quantitative PCR. The PCR-EIA assay detected a minimum of 25 copies/ml C. pneumoniae DNA in plasmid-spiked CSF, which was at least 10 times more sensitive than TaqMan. These data indicated that the PCR-EIA assay possessed a sensitivity that was equal to the nested-PCR procedures for the detection of C. pneumoniae DNA in CSF. The TaqMan system may not be sensitive enough for diagnostic purposes due to the low C. pneumoniae copies existing in the majority of CSF specimens from MS patients.

OBJECTIVES: The respiratory pathogen Chlamydia pneumoniae (C. pneumoniae) produces acute and chronic lung infections and is associated with asthma. Evidence for effectiveness of antichlamydial antibiotics in asthma is limited. The primary objective of this pilot study was to investigate the feasibility of performing an asthma clinical trial in practice settings where most asthma is encountered and managed. The secondary objectives were to investigate (1) whether azithromycin treatment would affect any asthma outcomes and (2) whether C. pneumoniae serology would be related to outcomes. This report presents the secondary results. DESIGN: Randomized, placebo-controlled, blinded (participants, physicians, study personnel, data analysts), allocation-concealed parallel group clinical trial. SETTING: Community-based health-care settings located in four states and one Canadian province. PARTICIPANTS: Adults with stable, persistent asthma. INTERVENTIONS: Azithromycin (six weekly doses) or identical matching placebo, plus usual community care. OUTCOME MEASURES: Juniper Asthma Quality of Life Questionnaire (Juniper AQLQ), symptom, and medication changes from baseline (pretreatment) to 3 mo posttreatment (follow-up); C. pneumoniae IgG and IgA antibodies at baseline and follow-up. RESULTS: Juniper AQLQ improved by 0.25 (95% confidence interval;-0.3, 0.8) units, overall asthma symptoms improved by 0.68 (0.1, 1.3) units, and rescue inhaler use decreased by 0.59 (-0.5, 1.6) daily administrations in azithromycin-treated compared to placebo-treated participants. Baseline IgA antibodies were positively associated with worsening overall asthma symptoms at follow-up (p = 0.04), but IgG was not (p = 0.63). Overall asthma symptom improvement attributable to azithromycin was 28% in high IgA participants versus 12% in low IgA participants (p for interaction = 0.27). CONCLUSIONS: Azithromycin did not improve Juniper AQLQ but appeared to improve overall asthma symptoms. Larger community-based trials of antichlamydial antibiotics for asthma are warranted.

BACKGROUND: Since 9% to 20% of all cases of acute psychosis presenting to an Emergency Department (ED) are due to a general medical condition, cautious medical workup should be mandatory in such patients. Differential diagnosis must consider conditions as diverse as renal failure or CNS infection. Acute Chlamydia pneumoniae infection usually causes a self-limited respiratory syndrome. Rarely, acute neurological complications occur, with acute meningoencephalitis most frequently reported. Diagnosis requires a high level of suspicion and is difficult to confirm. CASE REPORT: We describe a 22 year-old female Caucasian who, three days after a mild pharingitis, developed an acute psychosis with exuberant symptoms interspersed with periods of lucidity, in a background of normal consciousness and orientation. Initial medical and imagiological workup were inconclusive. After 20 days of unsuccessful treatment with antipsychotics she developed a high fever and was re-evaluated medically. Lumbar puncture revealed an inflammatory cerebrospinal fluid. MRI showed irregular thickening and nodularity of the lateral ventricles' lining. An anti-Chlamydia pneumoniae IgM antibody titter of 85 IU/ml was detected. All symptoms cleared after treatment with antibiotics and corticosteroids. CONCLUSION: This is, to our knowledge, the first reported case of acute CP-associated meningoencephalitis manifesting as an acute psychotic episode. It illustrates the principle that non-organic psychiatric syndromes must remain a diagnosis of exclusion in first-time acute psychosis.

Current studies indicate that a significant percentage of healthy blood donors carry Chlamydia pneumoniae in their blood. Although the clinical significance of such findings is unknown, eradication of such bacteria from blood components may contribute to transfusion safety. Deletion of C. pneumoniae in Red Blood Cell (RBC) units was accomplished through leukoreduction by filtration. The presence of bacteria in RBC units before and after leukoreduction was assessed by real-time PCR using primers specific for C. pneumoniae 16S rRNA. The eluates of filters used for leukoreduction were also assessed by PCR and immunostaining with fluorescein isothiocyanate-conjugated chlamydial monoclonal antibodies specific for C. pneumoniae determination. Nineteen of 30 RBC units tested showed the presence of C. pneumoniae DNA. Leukofiltration resulted in a marked reduction of leukocytes as well as C. pneumoniae in terms of bacterial number and positive rate for the bacteria. The eluates of filters showed trapped bacteria determined by both PCR and immunostaining assays. Thus, leukoreduction with a filter is an effective method to significantly reduce resident C. pneumoniae levels in RBC components but may not be completely sufficient for total eradication of this pathogen.

The rates of Chlamydia pneumoniae seroconversions suggesting acute primary infections or reinfections and the prevalences of antibodies were followed up among healthy laboratory workers. Annual serum samples were collected from 47 persons in Helsinki from 1958 to 1990 and from 40 persons in Oulu from 1994 to 1999. C. pneumoniae species-specific immunoglobulin G (IgG), IgA, and IgM antibodies were measured by microimmunofluorescence (MIF) in 407 sera from Helsinki. The 185 sera collected in Oulu were tested both by MIF and by commercial enzyme immunoassay (EIA). During the follow-up periods of 31 years in Helsinki and 6 years in Oulu, seroconversions were demonstrated by MIF in 45% and 15% of the study groups, respectively. In Helsinki 9% of the persons seroconverted twice during the follow-up period. By MIF, the total incidence rate per 100 person-years at risk was 6.9 in Helsinki and 4.9 in Oulu, and annual incidence rates varied from 0 to 15.4. By EIA, annual incidence rates in Oulu varied from 0 to 10.8. The seroconversions by MIF were usually not confirmed by EIA and vice versa. Prevalence and persistence rates, respectively, of IgA antibodies were higher in EIA (62% and 26%) than in MIF (26% and 17%), whereas the figures for IgG were quite similar. The prevalence of IgG and IgA antibodies was higher in older persons than in younger ones. The presence of antibodies did not offer protection from reinfection.

The intracellular bacterium Chlamydophila ("Chlamydia") pneumoniae is a pathogen for several respiratory diseases and may be a factor in the pathogenesis of chronic diseases of aging including atherosclerosis and Alzheimer's disease. We assessed whether aging is coupled with increased burden of infection in BALB/c mice after intranasal infection by C. pneumoniae. Six- and twenty-month-old BALB/c mice were infected intranasally with 5 x 10(4) inclusion forming units (IFU) or 5 x 10(5) IFU of C. pneumoniae. Lung, brain, and heart tissue were analyzed for infectious C. pneumoniae and for Chlamydophila antigen by immunohistochemistry. At both doses, aging was associated with a decreased proportion of animals that cleared infection from the lung and greater burden of infectious organism within the lung. We observed dose-dependent spread to the heart/ascending aorta in animals infected with C. pneumoniae. In mice given 5 x 10(4) IFU, spread to the heart by day 14 was only observed in old mice. By day 28, all animals inoculated with 5 x 10(4) IFU showed evidence of spread to the heart, although higher C. pneumoniae titers were observed in the hearts from old mice. In mice inoculated with 5 x 10(5) IFU, spread of C. pneumoniae to the heart was evident by day 14, with no discernible age effect. C. pneumoniae was also recovered from the central nervous system (brain and olfactory bulb) of all mice by day 28 postinfection, with higher C. pneumoniae titers in old animals than in young animals. Our results suggest that infection with C. pneumoniae may be more severe in old animals.

We previously reported a high degree of variation in the sensitivities of serodiagnostic kits for the detection of Chlamydophila pneumoniae in sera from healthy donors. Since a low predictive value of a test can impair its diagnostic value, we have extended our studies to samples from patients with pneumonia. We focused on the most promising enzyme-linked immunosorbent assays (ELISAs)(SeroCP and SeroCP Quant; Savyon) identified in our previous study and included a new ELISA (sELISA; Medac). The agreement between all ELISAs for immunoglobulin G (IgG) and a reference microimmunofluorescence (MIF) test for IgG (SeroFIA; Savyon) was >/=90% for a collective of 80 patients. The positive predictive values were all >/=93%. The negative predictive values ranged from 68 to 83%. False-negative results were obtained only for samples that had low titers in the MIF test. The correlation of the IgG antibody titers determined by the MIF and SeroCP Quant tests was high (r(sp)= 0.9). Since the semiquantitative SeroCP and quantitative SeroCP Quant ELISAs achieved the highest sensitivities, they were evaluated further by using a second batch of sera from 50 patients with predominantly medium and low antibody titers in the MIF test and a control collection of sera from 80 children with negative MIF results. Again, the tests showed a high concordance with the MIF results (96%), and the antibody titers in the SeroCP Quant and MIF tests correlated well (r(sp)= 0.8). The specificities determined with the negative sera were >/=99% for the SeroCP Quant test and 86% for the SeroCP test. These results show that ELISAs that are fast and objective deliver seroprevalence results, sensitivities, and specificities that are very similar to those of the MIF test.

We studied 20 Chlamydia pneumoniae isolates obtained from respiratory sites and atheroma tissue of patients from various geographic areas to determine the susceptibilities of these isolates to a new des-fluoroquinolone, garenoxacin, and to levofloxacin. In addition, we assessed the cultures with these isolates by PCR for the presence or absence of Mycoplasma sp. DNA. Both the MIC at which 90% of isolates are inhibited (MIC(90)) and the minimal bactericidal concentration at which 90% of isolates are killed (MBC(90)) for garenoxacin were 0.06 microg/ml, and both the MIC(90) and the MBC(90) for levofloxacin were 2.0 microg/ml. The activity of garenoxacin against C. pneumoniae was 32-fold greater than that of levofloxacin. Mycoplasma sp. DNA was detected by PCR in 17 of 20 cultures. Mycoplasma amplicons from five Mycoplasma DNA-positive C. pneumoniae cultures were sequenced and found to represent four Mycoplasma species. Our data demonstrate that C. pneumoniae cultures frequently contain Mycoplasma DNA and that its presence in C. pneumoniae cultures does not appear to affect the susceptibility results for the two fluoroquinolones that we tested.

The gyrA and gyrB genes of Chlamydia pneumoniae TW-183 were cloned, and their proteins were purified by use of a fusion system with a maltose-binding protein. The 50% inhibitory concentrations of garenoxacin, sparfloxacin, moxifloxacin, gatifloxacin, and levofloxacin were 10.1, 47.5, 39.6, 64.2, and 156.9 microg/ml, respectively, and the MICs against C. pneumoniae TW-183 were 0.008, 0.016, 0.063, 0.125, and 0.25 microg/ml, respectively.

Chlamydia pneumoniae may play a role in atherogenesis and vascular diseases, and antibiotics may prove useful in these conditions. Three groups of New Zealand White rabbits (24 per group) were infected via the nasopharynx with C. pneumoniae on three separate occasions (2 weeks apart). Group I was untreated and sacrificed at 12 weeks; group II received clarithromycin at 20 mg/kg/day for 8 days, beginning 5 days after each inoculation (early treatment); and group III received a similar dose of clarithromycin starting 2 weeks after the third inoculation and continued for 6 weeks thereafter (delayed treatment). To test for a possible anti-inflammatory effect of clarithromycin, two other groups of uninfected rabbits (12 animals in each) were fed 0.5% cholesterol-enriched chow, and one of these groups was treated with clarithromycin at 30 mg/kg/day for 6 weeks. Of 23 untreated infected rabbits, 8 developed early lesions of atherosclerosis, whereas 2 of the 24 early-treated group II had similar changes (P = 0.036 [75% efficacy]). However, in the delayed-treatment group, group III, 3 of 24 rabbits developed early lesions of atherosclerosis, thus demonstrating 62.5% reduction compared to the untreated controls (P = 0.07 [trend to statistical significance]). C. pneumoniae antigen was detected in 8 of 23 group I (untreated) rabbits versus 1 of 24 of the early-treated (group II) rabbits and 4 of 24 animals in the delayed group III (P = 0.009 and 0.138, respectively). All of the untreated, cholesterol-fed rabbits had moderate to advanced atherosclerosis (grade III or IV); clarithromycin had no effect on reducing the prevalence of but did reduce the extent of atherosclerosis in the cholesterol-fed rabbits by 17% compared to untreated controls. Thus, clarithromycin administration modified C. pneumoniae-induced atherosclerotic lesions and reduced the ability to detect organism in tissue. Early treatment was more effective than delayed treatment.

The microimmunofluorescence (MIF) test is considered the "gold standard" for laboratory diagnosis of acute and chronic Chlamydia pneumoniae infection. The performance of a MIF test based on C. pneumoniae antigen from Washington Research Foundation (WRF) was compared with those of assays from Labsystems (LAB) and MRL Diagnostics (MRL) by investigation of sera from three groups of patients: group I, 83 sera from 28 patients with atypical pneumonia; group II, 37 sera from 16 patients with acute C. pneumoniae or Chlamydia psittaci respiratory tract infection confirmed by PCR or culture; group III, 100 sera from 100 persons enrolled in the Copenhagen City Heart Study. The accordance among the results of the WRF assay and the two commercial assays was excellent for the immunoglobulin M (IgM) antibody detection rate (98%). The accordance in detection rates for IgG and IgA antibodies in sera from patients with acute infections was acceptable (87 and 88%), and in sera from group III, it was excellent (95 and 97%). The determinations of endpoint titers were reproducible with <1 dilution step difference for all three methods, except that the mean IgM antibody titer found by the LAB assay was almost 2 dilution steps higher than that found by the other two methods. Although the three assays use different C. pneumoniae strains as antigens, the detection rates and IgG and IgA endpoint titers were similar. The difference in endpoint titers of IgM antibodies is of no major concern, as the diagnosis of acute C. pneumoniae infection rests on the presence of IgM antibodies, not on their level.

Chlamydia pneumoniae is a common intracellular human pathogen that has been associated with several severe pathological conditions, including coronary heart disease and atherosclerosis. There is no vaccine against C. pneumoniae infection, but CD8(+) T cells have been shown to be crucial for protection during experimental infection. However, the effector functions and epitope specificity of the protective CD8(+) T cell remain unknown. The aim of this study was to identify C. pneumoniae-derived mouse CD8 epitopes by using a recent epitope prediction method. Of four C. pneumoniae proteins (the major outer membrane protein, outer membrane protein 2, polymorphic outer membrane protein 5, and heat shock protein 60), 53 potential CD8(+) T-cell epitopes were predicted by H-2 class I binding algorithms. Nineteen of the 53 peptides were identified as CD8 epitopes by testing for induction of a cytotoxic response after immunization. To test whether the predicted epitopes are naturally processed and presented by C. pneumoniae-infected cells, we generated a panel of seven peptide-specific cytotoxic T lymphocyte lines that were subsequently tested for recognition of C. pneumoniae-infected target cells. By using this strategy, we were able to identify three C. pneumoniae CD8 epitopes that were, indeed, processed and presented on infected cells. Identification of these natural CD8 epitopes provides tools for characterization of CD8(+) T-cell function in vivo and generation of epitope-specific prevention strategies.

Real-time PCR was evaluated as a quantitative diagnostic method for Chlamydia pneumoniae infection using different respiratory samples. Real-time PCR had efficiency equal to or better than that of nested touchdown PCR. This study confirmed sputum as the best sampling material to detect an ongoing C. pneumoniae infection.

The seroprevalence of anti-Chlamydia pneumoniae-specific immunoglobulin G (IgG) antibodies is high in the adult population. Experience is required to perform a microimmunofluorescence test (MIF), the current "gold standard" for serological diagnosis, and the assay still lacks standardization. Partially automated enzyme-linked immunosorbent assays (ELISAs) and enzyme immunoassays (EIAs), which are more standardized and for which the reading of results is less subjective, have been developed. The different commercially available serological tests differ in their sensitivities and specificities, depending primarily on the antigen used. Therefore, we evaluated 11 different tests (10 were species specific, 1 was genus specific) for IgG antibodies using serum samples of 80 apparently healthy volunteers. The interpretation of the results was based on the results of the gold standard, MIF: a sample was judged positive if it was positive by at least three of the four different MIFs. Based on this internal standard, we found that 71% of the samples were positive, while 8% were false positive by some tests. The correlations between the results of the different MIFs ranged from 83 to 99%, and the correlations between the results of the MIFs and the different ELISAs and EIAs ranged from 78 to 98%. Comparison of the IgG titers measured by MIF showed good agreement (r = 0.76 to 0.91). This analysis revealed that some ELISAs and EIAs fail to detect low IgG titers. The specificities of the species-specific tests varied from 95 to 100%, and the sensitivities varied from 58 to 100%. These results indicate that serological assays for the detection of anti-C. pneumoniae-specific IgG vary greatly in their sensitivities and specificities. MIF must still be considered the best method for the detection of IgG in apparently healthy subjects, but the sensitivities and specificities of new ELISAs approximate those of MIFs.

A monoclonal antibody (MAb) directed against an unknown Chlamydophila pneumoniae epitope has been characterized, and the respective peptide mimotope has been identified. A murine MAb specific for C. pneumoniae was used to select peptides from phage display libraries. The peptides identified from the phage display library clones reacted specifically with the respective target murine MAb and with human sera previously identified as having antibody titers to C. pneumoniae. The selected peptide mimotope sequences tended to be composed of charged residues surrounding a core of hydrophobic residues. The peptide with the best binding could inhibit >95% of binding to the MAb, suggesting that the selected peptide binds the paratope of the respective MAb. The peptide reacted with human sera previously determined by microimmunofluorescence to have anti-C. pneumoniae antibodies. The peptide was competitively competed with the MAb against Renografin-purified, sonicated C. pneumoniae in an enzyme-linked immunosorbent assay and with whole-cell C. pneumoniae in an indirect fluorescence assay format, demonstrating its potential utility in the development of diagnostics. The use of this novel peptide may allow investigators to establish standardized assays free from cross-reactive Chlamydia trachomatis and Chlamydophila psittaci epitopes and immunoreactivity.

Chlamydia pneumoniae and Mycoplasma pneumoniae were evaluated as agents of persistent cough in adolescents and adults (n = 491). Tests of 473 respiratory specimens by culture or PCR or both identified four episodes (0.8%) of M. pneumoniae-associated illness and no episodes of C. pneumoniae illness, suggesting that these bacteria do not frequently cause persistent cough.

A number of studies have found that inflammation of the vessel wall plays an essential role in both the initiation and progression of atherosclerosis and erosion and fissure and the eventual rupture of plaques. Chlamydia pneumoniae is one of the infectious agents that have been investigated as possible causes of this inflammation. Initial studies of the association of C. pneumoniae and cardiovascular disease (CVD) were seroepidemiologic, and these were followed by studies in which the organism was identified in vascular tissue from patients with CVD by electron microscopy, PCR and immunocytochemical staining (ICC). C. pneumoniae has also been isolated by culture from vascular tissue in a small number patients. However, no single serologic, PCR, or ICC assay has been used consistently across all studies. The assays used are also not standardized. Recent studies of serologic and PCR assays for diagnosis of C. pneumoniae infection have suggested that there may be substantial interlaboratory variation in the performance of these tests. It now appears that some of the inconsistency of results from study to study may be due, in part, to lack of standardized methods. Although initial seroepidemiologic studies demonstrated a significantly increased risk of adverse cardiac outcome in patients who were seropositive, subsequent prospective studies found either small or no increased risk. In addition to the lack of consistent serologic criteria, recent evaluations have demonstrated inherent problems with performance of the most widely used serologic methods. Most importantly, we do not have a reliable serologic marker for chronic or persistent C. pneumoniae infection.

The obligate intracellular pathogen Chlamydia (Chlamydophila) pneumoniae is known to be associated with some chronic inflammatory diseases, such as atherosclerosis. Interaction between C. pneumoniae and immune cells is important in the development of such diseases. However, susceptibility of immune cells, particularly lymphocytes, to C. pneumoniae infection has not been reported, even though lymphocytes play a pivotal role in the development of the diseases caused by this bacterium. In this regard, we examined the susceptibility of lymphocytes to C. pneumoniae infection in vitro. The results demonstrated that human peripheral blood lymphocytes as well as mouse spleen lymphocytes could be infected with C. pneumoniae. Furthermore, purified T lymphocytes as well as established T-lymphocyte cell line cells showed an obvious susceptibility to C. pneumoniae infection, indicating that T cells could be one of the host cells for this bacterial infection. These findings reveal a new infection site for C. pneumoniae, i.e., lymphocytes.

Chlamydia pneumoniae is a common cause of respiratory infection. It has also been shown to be associated with coronary heart disease. Two proteins that have been reported to be recognized frequently during human infection are proteins having molecular masses of 43 and 53 kDa. In order to develop a useful alternative serological test to the microimmunofluorescence (micro-IF) assay, recombinant 43-kDa and 53-kDa chlamydia-specific proteins were evaluated in dot blot and/or for comparison to the standard micro-IF test. Primers for amplification were derived from genome sequence information for two C. pneumoniae genes (CPn0809 and CPn0980) encoding 53-kDa proteins and four C. pneumoniae genes (CPn0562, CPn0927, CPn0928, and Cpn0929) encoding 43-kDa proteins of unknown function, which were Chlamydia specific and not found in Chlamydia trachomatis. The 53-kDa protein product of CPn0809 or the N-terminal 18-kDa portion had better specificity than any of the 43-kDa recombinants but was much less sensitive than micro-IF. In contrast, the 53-kDa protein encoded by CPn0980 was recognized by 11 of 12 (92%) acute-phase sera, 35 of 46 (76%) chronic sera, 0 of 12 micro-IF-negative sera (C. pneumoniae and C. trachomatis negative), and 1 of 12 (8%) C. pneumoniae negative, C. trachomatis positive sera. Thus, it appears that the 53-kDa protein encoded by CPn0980 has potential use for serodiagnosis of C. pneumoniae infection.

The respiratory tract pathogen Chlamydia pneumoniae has been associated with atherosclerosis. Monocytes are supposed to serve as a vehicle for systemic dissemination of intracellular C. pneumoniae from the lung to the artery vessel wall. We were therefore interested in pathogen-induced cellular events associated with NF-kappaB, a crucial transcription factor for both inflammatory cytokines and antiapoptotic molecules. In this study we demonstrate by electrophoretic mobility shift assay that C. pneumoniae infection of the human monocytic cell line Mono Mac 6 induces activation of NF-kappaB over 48 h, with a maximum level at 1 h postinfection. As shown by supershift assay, the activated NF-kappaB complex consists of the subunits RelA (p65) and NF-kappaB1 (p50). Apoptotic host cells were not detected during the early stages of the infection when maximal activation of NF-kappaB was detected. Pretreatment of Mono Mac 6 with the antioxidant and NF-kappaB inhibitor PDTC (pyrrolidine dithiocarbamate) induced activation of caspase-3 and led to apoptotic cell death. The C. pneumoniae-induced activation of the NF-kappaB complex was reduced by PDTC, which in parallel resulted in an increased apoptosis, as quantified by annexin V labeling and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling reaction. In the complete absence of activated NF-kappaB, when Mono Mac 6 cells were pretreated with the more potent NF-kappaB inhibitors MG-132 and parthenolide a C. pneumoniae-mediated rescue of cells from induced apoptosis could not be achieved. Our results indicate that activation of NF-kappaB in C. pneumoniae-infected Mono Mac 6 cells is associated with protection of Mono Mac 6 cells against apoptosis and might thereby contribute to systemic spread of the pathogen.