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Mycobacterium tuberculosis that sustains moderate oxidative damage survives undetected on solid media, but growth delay restores replicative ability.

Standard methods for enumerating Mycobacterium tuberculosis in patient sputum can miss large populations of viable M. tuberculosis cells that are unable to grow either on solid medium or in liquid medium unless the medium has been extensively diluted. Because these bacteria can be detected in liquid medium after limiting dilution, they have been termed differentially culturable or differentially detectable M. tuberculosis (DD-Mtb). Treatment with isoniazid (H), rifampin (R), pyrazinamide (Z), and ethambutol (E) (HRZE) for 1 to 2 weeks has been shown to increase the representation of DD-Mtb in the sputum of drug-sensitive (DS) tuberculosis (TB) patients. However, little is known about DD-Mtb after longer periods of treatment with HRZE or in patients with drug-resistant (DR) TB who receive second-line therapies. Here, we measured the proportion of DD-Mtb cells in the sputum of 47 subjects, 29 with DS TB and 18 with DR TB, before initiation of treatment and at 2 weeks and 2 months thereafter. Prior to treatment, DD-Mtb cells represented the majority of M. tuberculosis cells in the sputum of 21% of subjects with DS TB, and this proportion rose to 65% after 2 weeks of treatment with first-line drugs. In subjects with DR TB, DD-Mtb cells were found in the sputum of 29% of subjects prior to treatment initiation, and this proportion remained steady at 31% after 2 weeks of treatment with second-line drugs. By 2 months, DD-Mtb cells were detected in the sputum of only 2/15 (13.3%) subjects with DS TB and in 0/15 of subjects with DR TB. One of the DS subjects whose sputum was positive for DD-Mtb at month 2 later experienced treatment failure.

With a rising incidence of COVID-19-associated morbidity and mortality worldwide, it is critical to elucidate the innate and adaptive immune responses that drive disease severity. We performed longitudinal immune profiling of peripheral blood mononuclear cells from 45 patients and healthy donors. We observed a dynamic immune landscape of innate and adaptive immune cells in disease progression and absolute changes of lymphocyte and myeloid cells in severe versus mild cases or healthy controls. Intubation and death were coupled with selected natural killer cell KIR receptor usage and IgM+ B cells and associated with profound CD4 and CD8 T cell exhaustion. Pseudo-temporal reconstruction of the hierarchy of disease progression revealed dynamic time changes in the global population recapitulating individual patients and the development of an eight-marker classifier of disease severity. Estimating the effect of clinical progression on the immune response and early assessment of disease progression risks may allow implementation of tailored therapies.

This study was conducted in treatment-naive adults with drug-susceptible pulmonary tuberculosis in Port-au-Prince, Haiti, to assess the safety, bactericidal activity, and pharmacokinetics of nitazoxanide (NTZ). This was a prospective phase II clinical trial in 30 adults with pulmonary tuberculosis. Twenty participants received 1 g of NTZ orally twice daily for 14 days. A control group of 10 participants received standard therapy over 14 days. The primary outcome was the change in time to culture positivity (TTP) in an automated liquid culture system. The most common adverse events seen in the NTZ group were gastrointestinal complaints and headache. The mean change in TTP in sputum over 14 days in the NTZ group was 3.2 h ± 22.6 h and was not statistically significant (P = 0.56). The mean change in TTP in the standard therapy group was significantly increased, at 134 h ± 45.2 h (P

The historical view of β-lactams as ineffective antimycobacterials has given way to growing interest in the activity of this class against Mycobacterium tuberculosis (Mtb) in the presence of a β-lactamase inhibitor. However, most antimycobacterial β-lactams kill Mtb only or best when the bacilli are replicating. Here, a screen of 1904 β-lactams led to the identification of cephalosporins substituted with a pyrithione moiety at C3’ that are active against Mtb under both replicating and nonreplicating conditions, neither activity requiring a β-lactamase inhibitor. Studies showed that activity against nonreplicating Mtb required the in situ release of the pyrithione, independent of the known class A β-lactamase, BlaC. In contrast, replicating Mtb could be killed both by released pyrithione and by the parent β-lactam. Thus, the antimycobacterial activity of pyrithione-containing cephalosporins arises from two mechanisms that kill mycobacteria in different metabolic states.

Measurement of the reduction in CFU in sputum of patients with TB up to 2 weeks after the initiation of treatment is the gateway test for a new TB treatment. Reports have suggested that CFU assays fail to detect the majority of viable M. tuberculosis cells in sputum samples from the majority of patients when the number of M. tuberculosis is estimated by limiting dilution (LD). In an effort to avoid potential methodologic confounders, we applied a modified version of the LD assay in a study of a geographically distinct population. We confirmed that differentially detectable (DD) M. tuberculosis is often found before treatment, albeit at lower proportionate levels than in earlier reports. Strikingly, the prevalence and proportionate representation of DD M. tuberculosis increased during standard treatment. Sublethal exposure to certain antibiotics may help generate DD M. tuberculosis cells or enrich their representation among the surviving bacteria, and this may contribute to the need for prolonged treatment with those agents in order to achieve durable cures., Recent reports indicate that the sputum of 80% or more of treatment-naive subjects with tuberculosis recruited in England or South Africa contained more viable Mycobacterium tuberculosis cells detected by limiting dilution (LD) in liquid culture than detected as CFU. Efforts to generate such differentially detectable (DD) M. tuberculosis populations in vitro have been difficult to reproduce, and the LD assay is prone to artifact. Here, we applied a stringent version of the LD assay to sputum from 33 treatment-naive, HIV-negative Haitian subjects with drug-sensitive tuberculosis (TB) and to a second sputum sample after two weeks of standard treatment with isoniazid, rifampin, pyrazinamide, and ethambutol (HRZE) for 13 of these subjects. Twenty-one percent had statistically defined levels of DD M. tuberculosis in their pretreatment sputum at an average proportional excess over CFU of 3-fold. Sixty-nine percent of those who received HRZE had statistically defined levels of DD M. tuberculosis in their sputum, and of these, the mean proportionate excess over CFU was 7.9-fold. Thus, DD M. tuberculosis is detectable in pretreatment sputum from a significant proportion of subjects in the Western Hemisphere, and certain drugs or drug regimens, while reducing CFU, may at the same time increase the proportional representation of DD M. tuberculosis among the surviving bacilli. Monitoring DD M. tuberculosis may improve our ability to predict the efficacy of efforts to shorten treatment.

Diarrhea is the most common health issue affecting travelers to destinations across the world. This paper reviews the options for initial treatment of acute traveler’s diarrhea (TD). Its prevention, including but not limited to vaccines and safe travel and eating habits, is an important consideration but is beyond the scope of this paper. Treatment of TD has three arms: volume repletion, antibiotics, and antimotility/antisecretory agents. Patients should be advised to continue a diet that they can tolerate and maintain adequate fluid intake. In most cases, neither oral rehydration therapy nor dietary restrictions are likely to provide significant benefit. As yet, there is no evidence to support probiotic use for treatment of this type of diarrhea. Given that bacteria are the most frequent cause of TD, adult patients with moderate to severe disease should be treated empirically with a short course of antibiotics. In many instances, these will be prescribed pre-travel with instructions for proper usage when typical symptoms occur while abroad. However, such travelers should be advised to see a physician or seek emergency treatment if symptoms are severe or persist beyond 3 days. Antibiotic selection must take into account the epidemiology of resistant enteric pathogens. Fluoroquinolones are usually effective, although resistance of Campylobacter to this class of drugs in South and Southeast Asia warrants azithromycin as first-line empiric therapy in travelers to those regions. One day of therapy is often sufficient but can be extended to 3 days. Rifaximin is an alternative in non-invasive disease only. The antimotility agent loperamide is safe and effective and should be considered as adjunctive therapy in most cases of TD and can similarly be prescribed pre-travel. In non-pregnant adults, bismuth subsalicylate can also provide some symptomatic relief. Where available, racecadotril may be a safe alternative in both adults and children, although never specifically studied in TD. In cases of severe symptoms, or those lasting longer than 3 days, the patient should be evaluated for non-bacterial etiologies as well as possible Clostridium difficile infection. Certain travelers are more vulnerable to severe complications related to TD. Children, particularly infants, may need more aggressive fluid resuscitation with oral rehydration therapy. Several of the antidiarrheal agents must be avoided. Elderly patients and those with impaired cardiovascular reserve or immune-deficient states are more prone to complications as well. Treatment recommendations also differ for pregnant women. We generally advise adult non-pregnant travelers to follow smart eating and drinking practices and to bring a supply of bismuth subsalicylate and loperamide. We also prescribe an empiric antibiotic course (ciprofloxacin or azithromycin for up to 3 days) that is to be used for moderate to severe cases of TD.

Mycobacterium tuberculosis (Mtb) encounters stresses during the pathogenesis and treatment of tuberculosis (TB) that can suppress replication of the bacteria and render them phenotypically tolerant to most available drugs. Where studied, the majority of Mtb in the sputum of most untreated subjects with active TB have been found to be nonreplicating by the criterion that they do not grow as colony-forming units (cfus) when plated on agar. However, these cells are viable because they grow when diluted in liquid media. A method for generating such "differentially detectable" (DD) Mtb in vitro would aid studies of the biology and drug susceptibility of this population, but lack of independent confirmation of reported methods has contributed to skepticism about their existence. Here, we identified confounding artifacts that, when avoided, allowed development of a reliable method of producing cultures of ≥90% DD Mtb in starved cells. We then characterized several drugs according to whether they contribute to the generation of DD Mtb or kill them. Of the agents tested, rifamycins led to DD Mtb generation, an effect lacking in a rifampin-resistant strain with a mutation in rpoB, which encodes the canonical rifampin target, the β subunit of RNA polymerase. In contrast, thioridazine did not generate DD Mtb from starved cells but killed those generated by rifampin.

The purpose of this study is to evaluate the association between utilization of HIV testing and condom use amongst Cameroonian youths/adolescents who are not known to be HIV-infected. Worldwide, HIV is spreading most quickly amongst youths/adolescents. Between 44% and 82% of sexually active youths in Cameroon report inconsistent condom use. Data regarding utilization of HIV testing and condom use are lacking. A cross-sectional survey was administered to 431 youths ages 12-26 years in Cameroon from September 2011 to December 2011. Data on sociodemographics, sexual risk behaviors, self-reported HIV status, and condom use were collected. We compared rates of inconsistent condom use between those with known HIV negative status who utilized testing (HIV-N) and those with unknown status due to unutilized testing (HIV-U). Inconsistent condom use was defined as responding "never," "sometimes," or "usually," while consistent condom use was defined as responding "always" to questions regarding frequency of condom use. Generalized estimating equations were applied to assess the association between HIV testing and inconsistent condom use, adjusting for other confounders. Of 414 eligible respondents, 205 were HIV-U and 209 were HIV-N. HIV-U subjects were younger (mean age = 16.4 vs. 17.9, p < 0.001) and more likely to report living in an urban area (p = 0.002) than HIV-N subjects. Seventy-two percent (137/191) of sexually active youths reported inconsistent condom use. After adjusting for potential confounders, HIV-U status (odds ratio [OR] = 3.97, 95% confidence interval [CI] = 1.68-6.01) was associated with inconsistent condom use. Similarly, female gender (OR = 3.2, 95% CI = 1.29-7.89) was associated with inconsistent condom use, while older age at sexual debut was associated with a decreased risk for inconsistent condom use (OR = 0.67, 95% CI = 0.56-0.81). Cameroonian adolescents report high rates of inconsistent condom use which we found to be associated with self-reported unknown HIV status due to unutilized HIV testing. Successful HIV prevention programs among African youths/adolescents may benefit from expanded HIV testing programs.

Recent studies suggest that baseline tuberculous sputum comprises a mixture of routinely culturable and differentially culturable tubercle bacteria (DCTB). The latter seems to be drug tolerant and dependent on resuscitation-promoting factors (Rpfs).To further explore this, we assessed sputum from patients with tuberculosis for DCTB and studied the impact of exogenous culture filtrate (CF) supplementation ex vivo.Sputum samples from adults with tuberculosis and HIV-1 and adults with no HIV-1 were used for most probable number (MPN) assays supplemented with CF and Rpf-deficient CF, to detect CF-dependent and Rpf-independent DCTB, respectively.In 110 individuals, 19.1% harbored CF-dependent DCTB and no Rpf-independent DCTB. Furthermore, 11.8% yielded Rpf-independent DCTB with no CF-dependent DCTB. In addition, 53.6% displayed both CF-dependent and Rpf-independent DCTB, 1.8% carried CF-independent DCTB, and 13.6% had no DCTB. Sputum from individuals without HIV-1 yielded higher CF-supplemented MPN counts compared with counterparts with HIV-1. Furthermore, individuals with HIV-1 with CD4 counts greater than 200 cells/mmThese observations demonstrate a novel Rpf-independent DCTB population in sputum and reveal that reduced host immunity is associated with lower prevalence of CF-responsive bacteria. Quantification of DCTB in standard TB diagnosis would be beneficial because these organisms provide a putative biomarker to monitor treatment response and risk of disease recurrence.

After a cluster of fatal toxoplasmosis among stem cell transplant recipients at 2 hospitals, surveillance with polymerase chain reaction (PCR) (blood) was instituted. Rate of reactivation among seropositive recipients was 2.2 and 16%. Parasitemia was successfully managed with preemptive treatment. For seropositive recipients unable to take prophylaxis, toxoplasma PCR surveillance should be routinely performed.

Objectives: Young women are more likely to be infected with HIV globally, in sub-Saharan Africa, and in Cameroon. Despite its clear clinical and public health benefits, condom use among HIV-infected women continues to be low. The objective of this study was to describe the prevalence of inconsistent condom use among HIV-infected women in Cameroon and the factors associated with it. Methods: We conducted a cross-sectional study of HIV-infected young women aged 17–26 years from three semi-urban HIV clinics in the Northwest Region of Cameroon. This study was a subgroup analysis of a previously reported study on inconsistent condom use in HIV-infected and -uninfected youth. Inconsistent condom use was defined as reporting “sometimes” or “never” to questions regarding frequency of condom use. Logistic regression modeling was used to determine factors associated with inconsistent condom use. Results: A total of 84 participants were recruited and submitted completed questionnaires for analysis. Median age was 24 years (interquartile range = 22–25) and the median age at HIV diagnosis was 21 years (interquartile range = 20–23). Fifty percent of the participants reported no prior schooling or only primary school education. Overall, 61/84 (73%) reported inconsistent condom use. After adjusting for potential confounders, education to the secondary school level was protective against inconsistent condom use (odds ratio = 0.19; confidence interval: 0.04–0.95), and having ≥2 pregnancies was associated with inconsistent condom use (odds ratio = 7.52; confidence interval: 1.67–34.00). Conclusion: There is a high prevalence of inconsistent condom use among young HIV-infected women in Cameroon, which appears to be associated with lower levels of educational attainment and higher parity. Further larger studies assessing the factors associated with poor condom use in this population are warranted and may inform public health policy in resource-limited settings with high HIV prevalence.

MicroRNAs (miRNAs) are regulatory molecules that negatively control gene expression by binding to complementary sequences on target mRNAs. The most thoroughly characterized miRNAs, lin-4 and let-7, direct cell fate determination during the larval transitions in C. elegans and act as key regulators of temporal gene expression. lin-4 and let-7 are founding members of two distinct families of miRNA genes sharing strong sequence homology primarily in the 5' end of the mature miRNAs. In this report, we characterize the temporal and spatial expression patterns of lin-4 and let-7 family members using northern blot analysis and mir::gfp fusion studies. Our results show that lin-4 and let-7 homologues possess distinct temporal and spatial expression patterns during nematode development and that known heterochronic genes regulate their expression. We find that certain lin-4 and let-7 family members display overlapping expression patterns in the hypodermis and the reproductive system, suggesting that combinations of miRNAs from across families may control common developmental events.