Salmonella enterica subsp. enterica serovar Infantis (S. Infantis) is one of the "top five Salmonella serovars" of clinical significance in the European Union (EU). Antimicrobial resistant and extended spectrum beta-lactamase (ESBL) AmpC-producing S. Infantis have been described in food production systems and human clinical samples in Italy. Recently, an increase of MDR S. Infantis carrying blaCTX-M genes involved in 3rd generation cephalosporin resistance was noticed in the EU, including Italy, mainly due to the spread of S. Infantis harboring a pESIlike plasmid.The aim of this study was to investigate the occurrence of the S. Infantis pESI-like plasmid among antibiotic resistant S. Infantis strains isolated at different points of the food chain, and to provide a phylogenetic analysis to gain further insight on their transmission pathways from 'farm to fork'. MDR S. Infantis strains (n. 35) isolated from 2016 to 2021 at different stages of the food chain (animals, food, food-related environments, and humans) were investigated with in depth molecular characterization using realtime PCR, S1 nuclease pulsed-field gel electrophoresis (S1-PFGE) and whole genome sequencing (WGS).Our study reported the occurrence of S. Infantis strains harboring pESI-like plasmids, carrying blaCTX-M-1 genes, in Central Italy, at different sampling points along the food chain. Results confirmed the presence of a plasmid with a molecular size around 224-310 kb, thus consistent with the pESI-like, in 97 % of the 35 samples investigated. Two variants of S. Infantis pESI-like IncFIB(K)_1_Kpn3 were detected, one associated with the European clone carrying blaCTX-M-1 (21 isolates) and the other associated with U.S. isolates carrying blaCTX-M-65 (2 isolates, pESI-like U.S. variant). The majority was resistant to 3rd generation cephalosporins but none of the strains tested positive for the carbapenemase encoding genes. A total of 118 virulence genes were identified in isolates harboring the pESI-like plasmid. cgMLST and SNP-based analysis revealed the presence of one main cluster, composed by strains isolated from the environment, animals, food and humans.The results of this investigation underline the importance of phylogenetic studies to monitor and understand pathogen and AMR spread in a One Health approach.

This article reports a rapid and unexpected spread of colonization cases of NDM-1 carbapenemase-producing Klebsiella pneumoniae and Escherichia coli in a neonatal surgical unit (NSU) at Bambino Gesù Children's Hospital in Rome, Italy. Between the 16th of November 2020 and the 18th of January 2021, a total of 20 NDM-1 carbapenemase-producing K. pneumoniae (n = 8) and E. coli (n = 12) were isolated from 17 out of 230 stool samples collected from neonates admitted in the aforementioned ward and time period by an active surveillance culture program routinely in place to monitor the prevalence of colonization/infection with multidrug-resistant Gram-negative microorganisms. All strains were characterized by antimicrobial susceptibility testing, detection of resistance determinants, PCR-based replicon typing (PBRT) and multilocus-sequence typing (MLST). All isolates were highly resistant to most of the tested antibiotics, and molecular characterization revealed that all of them harbored the blaNDM-1 gene. Overall, IncA/C was the most common Inc group (n = 20/20), followed by IncFIA (n = 17/20), IncFIIK (n = 14/20) and IncFII (n = 11/20). MLST analysis was performed on all 20 carbapenemase-producing Enterobacterales (CPE) strains, revealing three different Sequence Types (STs) among E. coli isolates, with the prevalence of ST131 (n = 10/12; 83%). Additionally, among the 8 K. pneumoniae strains we found 2 STs with the prevalence of ST37 (n = 7/8; 87.5%). Although patient results were positive for CPE colonization during their hospital stay, infection control interventions prevented their dissemination in the ward and no cases of infection were recorded in the same time period.

Vancomycin-resistant Enterococci, mainly Enterococcus faecium (VREfm), are causing nosocomial infections and outbreaks. Bacterial typing methods are used to assist in outbreak investigations. Most of them, especially genotypic methods like multi-locus sequence typing (MLST), whole genome sequencing (WGS), or pulsed-field gel electrophoresis, are quite expensive and time-consuming. Fourier-transform infrared (FT-IR) spectroscopy assesses the biochemical composition of bacteria, such as carboxyl groups in polysaccharides. It is an affordable technique and has a faster turnaround time. Thus, the aim of this study was to evaluate FT-IR spectroscopy for VREfm outbreak investigations. Basic performance requirements like reproducibility and the effects of incubation time were assessed in distinct sample sets. After determining a FT-IR spectroscopy cut-off range, the clustering agreement between FT-IR and WGS within a retrospective (n: 92 isolates) and a prospective outbreak (n: 15 isolates) was investigated. For WGS an average nucleotide identity (ANI) cut-off score of 0.999 was used. Basic performance analysis showed reproducible results. Moreover, FT-IR spectroscopy readouts showed a high agreement with WGS-ANI analysis in clinical outbreak investigations (V-measure 0.772 for the retrospective and 1.000 for the prospective outbreak). FT-IR spectroscopy had a higher discriminatory power than MLST in the outbreak investigations. After determining cut-off values to achieve optimal resolution, FT-IR spectroscopy is a promising technique to assist in outbreak investigation as an affordable, easy-to-use tool with a turnaround time of less than one day. IMPORTANCE Vancomycin-resistant Enterococci, mainly Enterococcus faecium (VREfm), are a frequent cause of nosocomial outbreaks. Several bacterial typing methods are used to track transmissions and investigate outbreaks, whereby genome-based techniques are used as a gold standard. Current methods are either expensive, time-consuming, or both. Additionally, often, specifically trained staff needs to be available. This study provides insight into the use of Fourier-transform infrared (FT-IR) spectroscopy, an affordable, easy-to-use tool with a short turnaround time as a typing method for VREfm. By assessing clinical samples, this work demonstrates promising results for species discrimination and reproducibility. FT-IR spectrosopy shows a high level of agreement in the analysis of VREfm outbreaks in comparison with whole genome sequencing-based methods.

Abstract Background Leishmaniasis is a zoonotic disease endemic in the Mediterranean region where Leishmania infantum is the causative agent of human and canine infection. Characterization of this parasite at the subspecies level can be useful in epidemiological studies, to evaluate the clinical course of the disease (e.g. resistant strains, visceral and cutaneous forms of leishmaniasis) as well as to identify infection reservoirs. Multilocus enzyme electrophoresis (MLEE), a method currently recognized as the reference method for characterizing and identifying strains of Leishmania, is cumbersome and time-consuming and requires cultured parasites. These disadvantages have led to the development of other methods, such as multilocus microsatellite typing (MLMT) and multilocus sequence typing (MLST), for typing Leishmania parasites; however, these methods have not yet been applied for routine use. In this study, we first used MLST to identify informative polymorphisms on single-copy genes coding for metabolic enzymes, following which we developed two rapid genotyping assays based on high-resolution melting (HRM) analysis to explore these polymorphisms in L. infantum parasites. Methods A customized sequencing panel targeting 14 housekeeping genes was designed and MLST analysis was performed on nine L. infantum canine and human strains/isolates. Two quantitative real-time PCR-HRM assays were designed to analyze two informative polymorphisms on malic enzyme (ME) and glucose-6-phosphate isomerase (GPI) genes (390T/G and 1831A/G, respectively). The two assays were applied to 73 clinical samples/isolates from central/southern Italy and Pantelleria island, and the results were confirmed by DNA sequencing in a subset of samples. Results The MLST analysis, together with sequences available in the Genbank database, enabled the identification of two informative polymorphisms on the genes coding for ME and GPI. The fast screening of these polymorphisms using two HRM-based assays in 73 clinical samples/isolates resulted in the identification of seven genotypes. Overall, genotype 1 (sequence type 390T/1831G) was the most highly represented (45.2%) in the overall sample and correlated with the most common L. infantum zymodemes (MON-1, MON-72). Interestingly, in Pantelleria island, the most prevalent genotype (70.6%) was genotype 6 (sequence type 390T/1831A). Conclusions Applying our HRM assays on clinical samples allowed us to identify seven different genotypes without the need for parasite isolation and cultivation. We have demonstrated that these assays could be used as fast, routine and inexpensive tools for epidemiological surveillance of L. infantum or for the identification of new infection reservoirs. Graphical abstract

The current global pandemic of COVID-19 is characterized by waves of infection due to the emergence of new SARS-CoV-2 variants carrying mutations on the Spike (S) protein gene. Since autumn 2020 many Variants of Concern (VOC) have been reported: Alpha/B.1.1.7, Beta/B.1.351, Gamma/P.1, Delta/B.1.617.2, Omicron/B.1.1.529, and sublineages. Surveillance of genomic variants is currently based on whole-genome sequencing (WGS) of viral genomes on a random fraction of samples positive to molecular tests. WGS involves high costs, extended analysis time, specialized staff, and expensive instruments compared to a PCR-based test. To rapidly identify the VOCs in positive samples, six assays based on real-time PCR and high-resolution melting (HRM) were designed on the S gene and applied to 120 oro/nasopharyngeal swab samples collected from October 2020 to June 2022 (106 positive and 14 negative samples). Overall, the assays showed 100% specificity and sensitivity compared with commercial PCR tests for COVID-19. Moreover, 104 samples out of 106 (98.1%) were correctly identified as follows: 8 Wuhan (wild type), 12 Alpha, 23 Delta, 46 Omicron BA.1/BA.1.1, 15 Omicron BA.2/BA.4/BA.5. With our lab equipment, about 10 samples can be processed every 3 h at the cost of less than € 10 ($ 10.60) per sample, including RNA extraction. The implementation of this approach could help local epidemiological surveillance and clinical decision-making.

The spread of carbapenemase-producing Enterobacterales (CPE), especially Klebsiella pneumoniae (K. pneumoniae) and Escherichia coli (E. coli), is a serious public health threat in pediatric hospitals. The associated risk in newborns is due to their underdeveloped immune system and limited treatment options. The aim was to estimate the prevalence and circulation of CPE among the neonatal intensive units of a major pediatric hospital in Italy and to investigate their molecular features. A total of 124 CPE were isolated from rectal swabs of 99 newborn patients at Bambino Gesù Children’s Hospital between July 2016 and December 2019. All strains were characterized by antimicrobial susceptibility testing, detection of resistance genes, and PCR-based replicon typing (PBRT). One strain for each PBRT profile of K. pneumoniae or E. coli was characterized by multilocus-sequence typing (MLST). Interestingly, the majority of strains were multidrug-resistant and carried the blaNDM gene. A large part was characterized by a multireplicon status, and FII, A/C, FIA (15%) was the predominant. Despite the limited size of collection, MLST analysis revealed a high number of Sequence Types (STs): 14 STs among 28 K. pneumoniae and 8 STs among 11 E. coli, with the prevalence of the well-known clones ST307 and ST131, respectively. This issue indicated that some strains shared the same circulating clone. We identified a novel, so far never described, ST named ST10555, found in one E. coli strain. Our investigation showed a high heterogeneity of CPE circulating among neonatal units, confirming the need to monitor their dissemination in the hospital also through molecular methods.

Nontyphoidal salmonellosis (NTS) is the second most commonly reported gastrointestinal infection in humans and an important cause of food-borne outbreaks in Europe. The use of antimicrobial agents for animals, plants, and food production contributes to the development of antibiotic-resistant Salmonella strains that are transmissible to humans through food. The aim of this study was to investigate the presence and the potential dissemination of multidrug-resistant (MDR) Salmonella strains isolated in the Marche Region (Central Italy) via the food chain. Strains were isolated from different sources: food, human, food animal/livestock, and the food-processing environment. Among them, we selected MDR strains to perform their further characterization in terms of resistance to tetracycline agent, carriage of tet genes, and plasmid profiles. Tetracycline resistance genes were detected by PCR and plasmid replicons by PCR-based replicon typing (PBRT). A total of 102 MDR Salmonella strains were selected among the most prevalent serovars: S. Infantis (n = 36/102), S. Derby (n = 20/102), S. Typhimurium (n = 18/102), and a monophasic variant of S. Typhimurium (MVST, n = 28/102). Resistance to sulfisoxazole (86%) and tetracycline (81%) were the most common, followed by ampicillin (76%). FIIS was the most predominant replicon (17%), followed by FII (11%) and FIB (11%) belonging to the IncF incompatibility group. Concerning the characterization of tet genes, tetB was the most frequently detected (27/89), followed by tetA (10/89), tetG (5/89), and tetM (1/89). This study showed the potential risk associated with the MDR Salmonella strains circulating along the food chain. Hence, epidemiological surveillance supported by molecular typing could be a very useful tool to prevent transmission of resistant Salmonella from food to humans, in line with the One Health approach.

Background: Hypohidrotic ectodermal dysplasia (HED) is the most common form of ectodermal dysplasia and is mainly associated with mutations in the EDA, EDAR, and EDARADD responsible for the development of ectodermal-derived structures. HED displays different modes of inheritance according to the gene that is involved, with X-linked EDA-related HED being the most frequent form of the disease. Methods: Two families with tooth agenesis and manifestations of HED underwent clinical examination and EDA, EDAR, and EDARADD genetic analysis. The impact of the novel variant on the protein was evaluated through bioinformatics tools, whereas molecular modeling was used to predict the effect on the protein structure. Results: A novel missense variant was identified in the EDAR (c.287T>C, p.Phe96Ser) of a female child proband and her mother, accounting for autosomal dominant HED. The genetic variant c.866G>A (p.Arg289His) in EDA, which has been previously described, was observed in the male proband of another family confirming its role in X-linked HED. The inheritance model of the missense mutation showed a different relationship with X-linked HED and non-syndromic tooth agenesis. Conclusion: Our findings provide evidence of variable expression of HED in heterozygous females, which should be considered for genetic counseling, and different modes of inheritance related to tooth development.

The emerging spread of carbapenemase-producing Enterobacterales (CPE) strains, in particular, Klebsiella pneumoniae and Escherichia coli, has become a significant threat to hospitalized patients. Carbapenemase genes are frequently located on plasmids than can be exchanged among clonal strains, increasing the antibiotic resistance rate. The aim of this study was to determine the prevalence of CPE in patients upon their admission and to analyze selected associated factors. An investigation of the antibiotic resistance and genetic features of circulating CPE was carried out. Phenotypic tests and molecular typing were performed on 48 carbapenemase-producing strains of K. pneumoniae and E. coli collected from rectal swabs of adult patients. Carbapenem-resistance was confirmed by PCR detection of resistance genes. All strains were analyzed by PCR-based replicon typing (PBRT) and multilocus sequence typing (MLST) was performed on a representative isolate of each PBRT profile. More than 50% of the strains were found to be multidrug-resistant, and the bla KPC gene was detected in all the isolates with the exception of an E. coli strain. A multireplicon status was observed, and the most prevalent profile was FIIK, FIB KQ (33%). MLST analysis revealed the prevalence of sequence type 512 (ST512). This study highlights the importance of screening patients upon their admission to limit the spread of CRE in hospitals.

The eradication of recurrent Pseudomonas aeruginosa (PA) lung infection in cystic fibrosis (CF) patients may be hampered by the development of persistent bacterial forms, which can tolerate antibiotics through efflux pump overexpression. After demonstrating the efflux pump inhibitory effect of the alkaloid berberine on the PA MexXY-OprM efflux pump, in this study, we tested its ability (80/320 μg/mL) to enhance tobramycin (20xMIC/1000xMIC) activity against PA planktonic/biofilm cultures. Preliminary investigations of the involvement of MexY in PA tolerance to tobramycin treatment, performed on the isogenic pair PA K767 (wild type)/K1525 (ΔmexY) growing in planktonic and biofilm cultures, demonstrated that the ΔmexY mutant K1525 produced a lower (100 and 10 000 times, respectively) amount of tolerant cells than that of the wild type. Next, we grew broth cultures of PAO1, PA14, and 20 PA clinical isolates (of which 13 were from CF patients) in the presence of 20xMIC tobramycin with and without berberine 80 μg/mL. Accordingly, most strains showed a greater (from 10- to 1000-fold) tolerance reduction in the presence of berberine. These findings highlight the involvement of the MexXY-OprM system in the tobramycin tolerance of PA and suggest that berberine may be used in new valuable therapeutic combinations to counteract persister survival.

The aim of the present study isto characterise the diatom Pseudo-nitzschia community during a bloom period together with environmental conditions. High proliferation of Pseudo-nitzschia spp. was observed in September 2017 at the shellfish breeding area in the Krka River estuary (Central eastern Adriatic Sea). The peak of abundance (1.8 x 10 6 cells L -1 ) was recorded at 7 m depth, and the increased abundance persisted for four weeks.Morphological analyses of field samples based on scanning electron microscopy (SEM) revealed that Pseudo-nitzschia cf. arenysensis was prevailing (94%) in the Pseudo-nitzschia assemblage. Several strains were successfully isolated from net samples in order to better define morphological features and phylogenetic characterisation. The isolated Pseudo-nitzschia strains corresponded morphologically to the P. cf. arenysensis from the field samples, based on our SEM observations. Phylogenetic analysis demonstrated that the Croatian strains grouped with P. arenysensis using the ITS and LSU rDNA sequences. Spearman rank correlation showed that salinity was an important environmental factor affecting the vertical distribution of Pseudo-nitzschia spp. in this highly variable area. Availability of increased concentration of orthophosphates and ammonium and low Si: TIN ratio may have promoted the bloom of P. cf. arenysensis in the estuary.

Phytoplankton species can produce resting stages that persist in the sediment over long periods and accomplish important ecological functions. With the aim of investigating the phytoplankton assemblage structure in relation to environmental drivers and human pressures, we analyzed resting stages of diatoms and dinoflagellates, including harmful algal bloom taxa, in the surface sediments of 3 Mediterranean regional areas. Abundance of resting stages was determined by molecular quantitative polymerase chain reaction assay. Multivariate data analysis confirmed that the abundance of resting-stage assemblages seemed related to depth. Regional differences in the composition of the resting-stage assemblages were evident and environmental drivers were correlated with those regional differences. Three main groups of samples were defined according to SST and depth thresholds. Samples from the northern and central Adriatic Sea (average SST < 18°C) formed the richest assemblages, both in terms of abundance and species richness, while deep samples from all other basins (depth > 368 m) were poorer and less diverse than those from shallower sites (depth ≤ 368 m). Resting-stage taxa contributed differently to the 3 groups. Diatom spores and dinoflagellate cysts were the most abundant taxa, but Alexandrium minutum cysts and Ditylum brightwellii spores also accounted for a large share of the overall inter-group compositional distance. The structure of resting-stage assemblages can be regarded as a time- and space-integrated response of a subset of phytoplankton species to environmental conditions, including the physical oceanographic dynamics that favor or prevent sedimentation of resting stages.

Unlike human isolates, environmental Escherichia coli isolates have not been thoroughly investigated for the diversity and transferability of antibiotic-resistant plasmids. In this study, antibiotic-resistant strains from marine sediment (n = 50) and clams (n = 53) were analyzed (i) for their plasmid content using a PCR-based plasmid replicon typing (PBRT) kit and (ii) for the transferability of plasmid-associated antibiotic resistance (AR) traits by mating experiments. Fifteen of the thirty replicons targeted by the PBRT kit were detected in the isolates; 8/15 were identified in both sediment and clam isolates, although at different frequencies. The most frequent replicons in sediment (74%) and in clam strains (66%) alike, were FIA, FIB, or FII, which are associated with the IncF group, followed by the I1α replicon, which was more frequent in clam (24.5%) than in sediment (10%) strains. More than 50% of the strains contained multiple replicons; although 15 were untypable, S1-PFGE analysis demonstrated that 14/15 carried no plasmids. All cryptic strains were successfully typed and were positive for IncF or IncI replicons. Antibiotic-resistant strains accounted for 63% of all isolates and were significantly (p < 0.05) more frequent in phylogroup A. Most (35%) multidrug-resistant (MDR) strains belonged to phylogroup A, too. Although 25/26 MDR strains were positive for IncF plasmids (the exception being a clam strain), the FII-FIB rep combination was predominant (63%) among the sediment isolates, whereas most clam isolates (40%) carried the FII replicon alone. In mating experiments, selected MDR strains carrying FIB, FII, and I1α replicons, used as the donors, transferred multiple ARs together with the IncF or IncI plasmids at high frequency. Since IncI plasmids are common in E. coli and Salmonella enterica isolates from poultry, our findings suggest an animal origin to the E. coli clam strains carrying IncI plasmids. They also suggest a role for IncI plasmids in the spread of ARs among environmental Enterobacteriaceae and, through the food chain, to human isolates. In conclusion, the PBRT kit proved to be a useful tool to identify plasmids carrying antibiotic-resistant genes and to shed light on the factors underpinning their diffusion.

The parasite protozoan Leishmania, the causative agent of leishmaniasis, includes two subgenera of medical interest: Leishmania (Leishmania) and Leishmania (Viannia). Parasite species detection and characterization is crucial to choose treatment protocols and to monitor the disease evolution. Molecular approaches can speed up and simplify the diagnostic process. In particular, several molecular assays target the mitochondrial DNA minicircle network (kDNA) that characterizes the Leishmania genus. We previously proposed a qPCR assay targeting kDNA, followed by high resolution melt (HRM) analysis (qPCR-ML) to distinguish L. (L.) infantum and L. (L.) amazonensis from L. Viannia species. Successively, this assay has been integrated with other qPCR assays, to differentiate L. (L.) infantum, L. (L.) amazonensis and L. (L.) mexicana. In this work, we tested the applicability of our qPCR-ML assay on L. (L.) donovani, L. (L.) major, L. (L.) tropica and L. (L.) aethiopica, showing that the qPCR-ML assay can also amplify Old World species, different from L. (L.) infantum, with good quantification limits (1 × 10−4–1 × 10−6 ng/pcr tube). Moreover, we evaluated 11 L. (L.) infantum strains/isolates, evidencing the variability of the kDNA minicircle target molecules among the strains/isolates of the same species, and pointing out the possibility of quantification using different strains as reference. Taken together, these data account for the consideration of qPCR-ML as a quantitative pan-Leishmania assay.

Glutamate decarboxylase catalyzes the conversion of glutamate to gamma-aminobutyric acid (GABA), contributing to pH homeostasis through proton consumption. The reaction is the first step toward the GABA shunt. To date, the enzymes involved in the glutamate metabolism of Photobacterium damselae subsp. piscicida have not been elucidated. In this study, an open reading frame of P. damselae subsp. piscicida, showing homology to the glutamate decarboxylase or putative pyridoxal-dependent aspartate 1-decarboxylase genes, was isolated and cloned into an expression vector to produce the recombinant enzyme. Preliminary gas chromatography-mass spectrometry characterization of the purified recombinant enzyme revealed that it catalyzed not only the decarboxylation of glutamate but also the transamination of GABA. This enzyme of P. damselae subsp. piscicida could be bifunctional, combining decarboxylase and transaminase activities in a single polypeptide chain.

OBJECTIVES:Streptococcus pyogenes causes life-threatening invasive infections including necrotizing fasciitis (NF). Current treatment guidelines recommend the use of a cell-wall-active antibiotic combined with a protein synthesis inhibitor and surgical debridement in NF patients. Adjunctive therapy with intravenous immunoglobulin (IVIG) has been proposed for superantigen-associated streptococcal toxic shock syndrome. So far, benefits of IVIG treatment remain unclear and prospective clinical studies are scarce. Thus, we aimed to assess the effects of IVIG on virulence factor activity in vitro, ex vivo in patients and in vivo in a NF mouse model. METHODS:We investigated the effect of IVIG on the activity of the virulence factors streptolysin O (SLO), streptodornase 1 (Sda1), S. pyogenes cell envelope protease and streptococcal pyrogenic exotoxin B in vitro and ex vivo in patient sera. Additionally, we assessed the influence of IVIG on the clinical outcome in a murine NF model. RESULTS:In vitro, IVIG inhibited various streptococcal virulence factors. Further, IVIG treatment of group A Streptococcus-infected mice led to a reduced skin lesion size (median (interquartile range) day 3 intraperitoneal administration: 12 mm2 (9-14.5) vs. 4 mm2 (0.8-10.5), subcutaneous: 10.3 mm2 (6.9-18.6) vs. 0.5 mm2 (0.1-6.8)) and lower SLO activity. After treatment with IVIG, patient sera showed an elevated titre of specific SLO (7/9) and Sda1 (5/9) antibodies, reducing SLO and Sda1 activity. CONCLUSIONS:The clear reduction in disease severity in IVIG-treated mice and inhibition of virulence factor activity in mouse and human sera suggest that IVIG may be beneficial in invasive group A Streptococcus infections such as NF in addition to streptococcal toxic shock syndrome.

Mycobacterium tuberculosis (MTB) is notorious for persisting within host macrophages. Efflux pumps decrease intracellular drug levels, thus fostering persistence of MTB during therapy. Isoniazid (INH) and pyrazinamide (PZA) are substrates of the efflux pump breast cancer resistance protein-1 (BCRP-1), which is inhibited by chloroquine (CQ). In this study, BCRP-1 was found to be expressed on macrophages of human origin and on foamy giant cells at the site of MTB infection. In the current in vitro study, interferon-gamma (IFNγ) increased the expression of BCRP-1 in macrophages derived from the human monocytic leukaemia cell line THP-1. Using a BCRP-1-specific fluorescent dye and radioactively labelled INH, it was demonstrated that efflux from macrophages increased upon activation with IFNγ. CQ was able to inhibit active efflux and augmented the intracellular concentrations both of INH and the dye. In agreement, CQ and specific inhibition of BCRP-1 increased the antimycobacterial activity of INH against intracellular MTB. Although PZA behaved differently, CQ had comparable advantageous effects on the intracellular pharmacokinetics and activity of PZA. The adjunctive effects of CQ on intracellular killing of MTB were measurable at concentrations achievable in humans at approved therapeutic doses. Therefore, CQ, a widely used and worldwide available drug, may potentiate the efficacy of standard MTB therapy against bacteria in the intracellular compartment.