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Volume 60,
Issue 10,
2011
Volume 60, Issue 10, 2011
- Pathogenicity And Virulence
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Protease inhibitors decrease IgG shedding from Staphylococcus aureus, increasing complement activation and phagocytosis efficiency
Staphylococcus aureus is a major pathogen for immunologically intact humans and its pathogenesis is a model system for evasion of host defences. Antibodies and complement are essential elements of the humoral immune system for prevention and control of S. aureus infections. The specific hypothesis for the proposed research is that S. aureus modifies humoral host defences by cleaving IgG that has bound to the bacterial surface, thereby inhibiting opsonophagocytosis. S. aureus was coated with pooled, purified human IgG and assayed for the shedding of cleaved IgG fragments using ELISA and Western blot analysis. Surface-bound IgG was shed efficiently from S. aureus in the absence of host blood proteins. Broad-spectrum protease inhibitors prevented cleavage of IgG from the S. aureus surface, suggesting that staphylococcal proteases are responsible for IgG cleavage. Serine protease inhibitors and cysteine protease inhibitors decreased the cleavage of surface-bound IgG; however, a metalloprotease inhibitor had no effect. Using protease inhibitors to prevent the cleavage of surface-bound IgG increased the binding of complement C3 fragments on the surface of S. aureus, increased the association with human neutrophils and increased phagocytosis by human neutrophils.
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- Host Response
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Saccharomyces as a vaccine against systemic aspergillosis: ‘the friend of man’ a friend again?
The mortality of clinical Aspergillus infections necessitates consideration of the utility of a vaccine. We have found that Saccharomyces species can act as a protective vaccine against a lethal systemic Aspergillus infection, and describe experiments optimizing a subcutaneous regimen with killed yeast. Three injections of 2.5 mg given a week apart, 2 weeks prior to challenge, consistently, significantly, provided survival protection and reduction of infection in organs in survivors. The protection was independent of the strain of Saccharomyces, and possibly even the species, and could be demonstrated in several inbred (including C′-deficient) and outbred mouse strains. The protective moiety(ies) appeared to reside in the cell wall and was resistant to 100 °C, but not to protease or formalin. Alum potentiated the protection. The protection was comparable or superior to that of several Aspergillus-specific preparations described in the literature. Other studies have indicated that heat-killed Saccharomyces can protect against infection with at least three other fungal genera, raising the possibility of development of a panfungal vaccine, and such a vehicle has been studied in clinical trials, without dose-limiting toxicity.
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- Diagnostics, Typing And Identification
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Molecular typing and epidemiology of enteroviruses in Cyprus, 2003–2007
More LessHuman enteroviruses (HEVs) are responsible for a wide spectrum of clinical diseases. Even though usually associated with non-specific febrile illness, they are the most common cause of viral meningitis and pose a serious public-health problem, especially during outbreaks. Rapid detection and identification of HEV serotypes in clinical specimens are important in appropriate patient management and epidemiological investigation. A 5 year study (2003–2007) of clinical specimens from patients with viral meningitis and/or symptoms of enteroviral infection was carried out in Cyprus to determine the underlying enteroviral aetiology. Reverse transcription, followed by a sequential PCR strategy targeting the 5′ non-coding region and VP1 region, was used for typing the isolated enteroviruses. The serotype of each isolate was determined by blast search of the VP1 amplicon sequence against GenBank. Clinical specimens from a total of 146 patients were diagnosed as enterovirus-positive. Twenty-two different serotypes were identified. The main strains identified were echovirus 18 and echovirus 30, followed by coxsackievirus B5, echovirus 9, echovirus 6, coxsackievirus A10 and coxsackievirus B2. However, rapid changes in serotype frequency and diversity were observed over time. Serotype distribution corresponded essentially with observations reported from other European countries in the same period. The present report demonstrates the epidemiology of enteroviruses in Cyprus from 2003 to 2007.
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Genotyping of Mycobacterium leprae from Brazilian leprosy patients suggests the occurrence of reinfection or of bacterial population shift during disease relapse
Adalgiza da Silva Rocha, Alexandre Araujo Cunha dos Santos, Patrícia Pignataro, José Augusto Nery, Antônio Basílio de Miranda, Diego Fonseca Soares, Amanda Nogueira Brum Fontes, Alice Miranda, Helen Ferreira, Neio Boéchat, Maria Eugênia Novisck Gallo, Euzenir Nunes Sarno, Maria Leide W. De Oliveira and Philip Noel SuffysWe performed genotyping of Mycobacterium leprae present in skin biopsy samples that were collected during the first and the second disease occurrences from eight leprosy patients, seven of whom were diagnosed as suffering from disease relapse. Sequence analysis of part of the M. leprae rpoB, folP1, gyrB and gyrA genes did not show genetic change that supported the presence of drug-resistant bacilli. However, we observed a synonymous nucleotide change at position 297 of gyrA among five of these patients, one presenting C to T (CgyrAT) and four presenting T to C (TgyrAC) at this position. Additional genotyping by analysis of the four short tandem repeats GAA, GTA9, AT17 and TA18 showed that the gyrA single nucleotide polymorphism change was accompanied by a change in short tandem repeat genotype. Our data suggest that leprosy relapse in these patients, living in an area endemic for leprosy, could be caused by M. leprae with a genotype different from the one that caused initial disease.
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Improved rapid molecular diagnosis of multidrug-resistant tuberculosis using a new reverse hybridization assay, REBA MTB-MDR
Rapid diagnosis of multidrug-resistant tuberculosis (MDR-TB) is essential for the prompt initiation of effective second-line therapy to improve treatment outcome and limit transmission of this obstinate disease. A variety of molecular methods that enable the rapid detection of mutations implicated in MDR-TB have been developed. The sensitivity of the methods is dependent, in principle, on the repertoire of mutations being detected, which is typically limited to mutations in the genes rpoB, katG and the promoter region of inhA. In this study, a new reverse hybridization assay, REBA MTB-MDR (M&D), that probes mutations in the oxyR–ahpC intergenic region, in addition to those in rpoB, katG and the inhA promoter region, was evaluated. A set of 240 Mycobacterium tuberculosis clinical isolates from patients receiving retreatment regimens was subjected to conventional phenotypic drug-susceptibility testing (DST) and the REBA MTB-MDR assay. The nucleotide sequences of the loci known to be involved in drug resistance were determined for comparison. In brief, the results showed that the REBA MTB-MDR assay efficiently recognized nucleotide changes in the oxyR–ahpC intergenic region as well as those in rpoB, katG and the inhA promoter region with higher sensitivity, resulting in an 81.0 % detection rate for isoniazid resistance. Inclusion of the oxyR–ahpC intergenic region in the REBA MTB-MDR assay improved the overall sensitivity of molecular DST for MDR-TB from 73.1 to 79.9 %.
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Serotype and genotype distributions of pneumococcal carriage isolates recovered from Brazilian children attending day-care centres
Pneumococcal nasopharyngeal carriage isolates recovered from Brazilian children attending day-care centres in 2005 were assessed for serotype, genotype and penicillin susceptibility phenotype. As 124 of the 253 isolates (49 %) were characterized previously with respect to serotype and penicillin susceptibility, the primary objectives were to examine clonal associations and penicillin susceptibility within major serotypes and to assess the suitability of conventional multiplex PCR for deducing carriage serotypes within this population. Using a combination of PCR-based serotyping and the Quellung reaction, serotypes were identified for 81 % (205/253) of the isolates, with serogroups or types 14, 6, 23F, 19F and 18 being predominant. Included within the 205 isolates successfully serotyped by PCR were 28 isolates that had become non-viable. Forty-eight isolates were non-typable using both the PCR method and the Quellung reaction. Penicillin non-susceptibility was observed within 16 of the 18 multilocus sequence types detected. Thus, this study provides further evidence from a diverse collection of pneumococcal clones that PCR-based serotype deduction is useful for providing supportive evidence for pneumococcal conjugate vaccine implementation.
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Genetic characteristics and changing antimicrobial resistance among Shigella spp. isolated from hospitalized diarrhoeal patients in Kolkata, India
To study the prevalence pattern and trends in the phenotypic and genetic characteristics of shigellae, we tested 212 isolates isolated from diarrhoeal patients admitted to the Infectious Diseases Hospital, Kolkata, India, from November 2007 to October 2010. Prevalence of Shigella spp. was higher in the >5 years age group (69 %) than in children in the <5 years age group (31 %). Serotypes 2a, 3a and untypable isolates of Shigella flexneri were frequently detected. An increase in the isolation of Shigella sonnei (15 %) is a novel trend in this region. Fluoroquinolone resistance among S. flexneri serotypes 2a, 3a and other serogroups of shigellae is another evolving trend. The set gene was exclusively present in S. flexneri 2a, and the sen gene was detected in all serogroups. PFGE revealed the grouping of S. flexneri isolates according to their serotypes with approximately 80–100 % similarity, whilst Shigella dysenteriae type 2 and S. sonnei were clonal in nature. There was no demarcation in the prevalence of serotypes, antimicrobial resistance or clonality between the two age groups.
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Detection of Aspergillus species in BACTEC blood cultures
More LessInvasive aspergillosis is associated with high morbidity and mortality rates; nevertheless, blood cultures almost invariably yield a negative result. The recovery and detection time of Aspergillus fumigatus, Aspergillus flavus and Aspergillus terreus were studied in BACTEC Plus Aerobic/F, Mycosis-IC/F and Myco/F Lytic vials, incubated in the BACTEC 9240 and 9000 MB automated systems. Two different approaches were used for subculture in solid medium: (i) the routine method, using a sterile airway needle/subculture unit, and (ii) a novel procedure, using instead a tuberculin disposable syringe and collecting a larger aliquot (100 µl), following vigorous agitation of the vials. A. fumigatus was detected at inoculum concentrations of >3 conidia per 10 ml after 21–40 h, in both BACTEC Plus Aerobic/F and BACTEC Mycosis-IC/F vials. A few more hours were needed to detect A. flavus and A. terreus. The novel subculture procedure of BACTEC culture vials on solid medium resulted in several positive results that were not detected by the routine sampling procedure. BACTEC Plus Aerobic/F vials show an advantage particularly in patients under antifungal treatment. In cases of polymicrobial bloodstream infections (concurrent bacterial growth), the inoculation of blood samples into a BACTEC Mycosis-IC/F vial achieved the best results. Further multicentre studies are needed to validate this improved automated detection of Aspergillus spp. from blood cultures in clinical laboratories, as this diagnostic procedure allows antifungal susceptibility testing of moulds.
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- Antimicrobial Agents And Chemotherapy
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Piperine as an inhibitor of the MdeA efflux pump of Staphylococcus aureus
More LessPiperine, a trans-trans-isomer of 1-piperoyl-piperidine, was tested in combination with mupirocin for antimicrobial activity against Staphylococcus aureus strains including meticillin-resistant S. aureus. The combination markedly reduced the MIC of mupirocin and also lowered the mutation frequency. Enhanced accumulation and efflux of ethidium bromide from wild-type and mutant (Mupr-1) strains in the presence of piperine indicated that inhibition of efflux could be a possible mechanism of potentiation of mupirocin activity by piperine. The combination of piperine with mupirocin in a dermal infection model of mice showed better in vivo efficacy when compared with the commercially available formulation of 2 % mupirocin.
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Coriander (Coriandrum sativum L.) essential oil: its antibacterial activity and mode of action evaluated by flow cytometry
More LessThe aim of this work was to study the antibacterial effect of coriander (Coriandrum sativum) essential oil against Gram-positive and Gram-negative bacteria. Antibacterial susceptibility was evaluated using classical microbiological techniques concomitantly with the use of flow cytometry for the evaluation of cellular physiology. Our results showed that coriander oil has an effective antimicrobial activity against all bacteria tested. Also, coriander oil exhibited bactericidal activity against almost all bacteria tested, with the exception of Bacillus cereus and Enterococcus faecalis. Propidium iodide incorporation and concomitant loss of all other cellular functions such as efflux activity, respiratory activity and membrane potential seem to suggest that the primary mechanism of action of coriander oil is membrane damage, which leads to cell death. The results obtained herein further encourage the use of coriander oil in antibacterial formulations due to the fact that coriander oil effectively kills pathogenic bacteria related to foodborne diseases and hospital infections.
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- Epidemiology
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In vitro antifungal susceptibilities and molecular typing of sequentially isolated clinical Cryptococcus neoformans strains from Croatia
A collection of 48 clinical Cryptococcus neoformans isolates from Croatia was investigated retrospectively using in vitro antifungal susceptibility testing and molecular biological techniques to determine mating type and serotype by PCR and amplified fragment length polymorphism (AFLP) genotyping. These isolates were obtained from 15 patients: ten were human immunodeficiency virus (HIV)-negative (66.7 %) and five were HIV-positive (33.3 %). From five patients, only one isolate was available, whilst from the other ten patients, two to 11 isolates were isolated sequentially. Antifungal susceptibility was tested by a broth microdilution method. Serotype A (genotype AFLP1) and serotype D (genotype AFLP2) were both found in six patients (40 % each), and serotype AD (genotype AFLP3) in three (20.0 %) patients. Mating type α (n = 12; 80.0 %) predominated and α/a hybrids were identified in 20.0 % of patients diagnosed with cryptococcosis. Two AFLP genotypes of C. neoformans were isolated during a single episode from one patient. The in vitro antifungal MIC90 and susceptibility ranges for C. neoformans isolates were 0.5 µg ml−1 (range 0.031–0.5 µg ml−1) for amphotericin B, 4 µg ml−1 (range 1–4 µg ml−1) for flucytosine and fluconazole, 0.25 µg ml−1 (range 0.031–0.5 µg ml−1) for itraconazole and 0.062 µg ml−1 (range 0.031–0.25 µg ml−1) for voriconazole.
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Molecular epidemiology of mumps virus strains circulating in south-west China from 2007 to 2009
More LessThe genetic characteristics of mumps virus (MuV) strains isolated from sporadic mumps cases between 2007 and 2009 in three provinces of south-west China were investigated. MuV detection was carried out by nested RT-PCR on 117 cases. The small hydrophobic gene of 33 isolated strains was identified, and sequence analysis revealed that all the isolates belonged to the lineage genotype F with slight nucleotide variation. Furthermore, the virus haemagglutinin–neuraminidase and nucleoprotein antigens exhibited a high degree of nucleotide and amino acid similarity (>99 %) within all isolates, whilst the fusion protein appeared to have certain geographical differences. This study on molecular surveillance will help to monitor the circulation of MuV in China.
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- Clinical Microbiology And Virology
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Microarray-based detection of extended virulence and antimicrobial resistance gene profiles in phylogroup B2 Escherichia coli of human, meat and animal origin
Extra-intestinal pathogenic Escherichia coli (ExPEC) causing urinary tract infections (UTIs) most often belong to phylogenetic group B2 and stem from the patient’s own faecal flora. It has been hypothesized that the external reservoir for these uropathogenic E. coli in the human intestine may be meat and food-production animals. To investigate such a connection, this study analysed an E. coli phylogroup B2 strain collection (n = 161) of geographical and temporally matched isolates, published previously, from UTI patients (n = 52), community-dwelling humans (n = 36), imported (n = 5) and Danish (n = 13) broiler chicken meat, Danish broiler chickens (n = 17), imported (n = 3) and Danish (n = 27) pork, and healthy Danish pigs (n = 8). The isolates were subjected to microarray analysis for 315 virulence genes and variants and 82 antimicrobial resistance genes and variants. In total, 133 different virulence and antimicrobial resistance genes were detected in at least one UTI isolate. Between 66 and 87 of these genes were also detected in meat and animal isolates. Cluster analyses of virulence and resistance gene profiles, respectively, showed that UTI and community-dwelling human isolates most often grouped with meat and animal isolates, indicating genotypic similarity among such isolates. Furthermore, B2 isolates were detected from UTI patients and meat, with indistinguishable gene profiles. A considerable proportion of the animal and meat isolates belonged to the ExPEC pathotype. In conclusion, these findings suggest that B2 E. coli from meat and animal origin can be the source of most of the virulence and antimicrobial resistance genes detected in uropathogenic E. coli isolates and that there is a general resemblance of animal, meat and UTI E. coli based on extended gene profiling. These findings support the hypothesis of a zoonotic link between E. coli causing UTIs and E. coli from meat and animals.
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- Veterinary Microbiology
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Mutant-prevention concentration and mechanism of resistance in clinical isolates and enrofloxacin/marbofloxacin-selected mutants of Escherichia coli of canine origin
More LessThe antibacterial activity and selection of resistant bacteria, along with mechanisms of fluoroquinolone resistance, were investigated by integrating the static [MIC or mutant-prevention concentration (MPC)] and in vitro dynamic model approaches using Escherichia coli isolates from diseased dogs. Using the dynamic models, selected E. coli strains and enrofloxacin and marbofloxacin at a range of simulated area under concentration–time curve over a 24 h interval (AUC24 h)/MIC ratios were investigated. Our results indicated increasing losses in susceptibility of E. coli upon continuous exposure to enrofloxacin and marbofloxacin in vitro. This effect was transferable to other fluoroquinolones, as well as to structurally unrelated drugs. Our results also confirmed an AUC24 h/MIC (AUC24 h/MPC)-dependent antibacterial activity and selection of resistant E. coli mutants, in which maximum losses in fluoroquinolone susceptibility occurred at simulated AUC24 h/MIC ratios of 40–60. AUC24 h/MPC ratios of 39 (enrofloxacin) and 32 (marbofloxacin) were considered protective against the selection of resistant mutants of E. coli. Integrating our MIC and MPC data with published pharmacokinetic information in dogs revealed a better effect of the conventional dosing regimen of marbofloxacin than that of enrofloxacin in restricting the selection of resistant mutants of E. coli. Target mutations, especially at codon 83 (serine to leucine) of gyrA, and overexpression of efflux pumps contributed to resistance development in both clinically resistant and in vitro-selected mutants of E. coli. We also report here a previously undescribed mutation at codon 116 of parC in two laboratory-derived resistant mutants of E. coli. Additional studies would determine the exact role of this mutation in fluoroquinolone susceptibility, as well as establish the importance of our findings in the clinical setting.
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- Models Of Infection
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A murine model for catheter-associated candiduria
More LessCandiduria is a common finding in hospitalized patients with indwelling urine-draining devices. Animal models for candiduria are not well-developed and, despite its prevalence and associated mortality, candiduria is understudied. The presence of Candida in urine does not imply disease because it is also a commensal. Biofilm formation on catheters and the host–pathogen interaction are likely to be important factors that contribute to the pathogenesis. The objective of this study was to establish a candiduria model in mice with indwelling catheters. Our data demonstrate that biofilm formation on indwelling catheters and persistent candiduria can be established in mice. The study supports the concept that biofilm formation contributes to persistence. It also outlines differences between catheter-related candiduria in mice and humans. Specifically, mice exhibit higher levels of leukocyturia. In addition, mean daily fungal burden in urine in the murine model is 10- to 100-fold lower than that in humans. These important findings must be taken into consideration when using this model to study host–pathogen interaction in the setting of candiduria.
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- Case Reports
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Candida parapsilosis meningitis as the first manifestation of AIDS: case report
Candida meningitis is a rare condition that occurs more frequently in premature infants, immunocompromised patients or patients after neurosurgery. We describe a case of a previously healthy 41-year-old man with Candida parapsilosis meningitis associated with oropharyngeal candidiasis as the first manifestation of AIDS.
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A rare, fatal case of invasive spinal aspergillosis in an antiretroviral-naïve, HIV-infected man with pre-existing lung colonization
More LessInfection of the central nervous system (CNS) is a rare but devastating complication of invasive aspergillosis. We report a case of invasive aspergillosis with spinal involvement in a human immunodeficiency virus (HIV)-infected patient without neutropenia. A 42-year-old, antiretroviral-naïve, HIV-infected man presented with progressive weakness in the lower limbs and urinary and faecal incontinence for 2 weeks. The patient had been prescribed broad-spectrum antibiotics and prednisone. He had upper motor neuron signs and a sensory level at T1, with accompanying neck stiffness on flexion. Magnetic resonance imaging revealed diffuse abnormal signals of the vertebral bodies in the lower cervical and thoracic areas, with cord compression in the C2 and C3 region and signal distortions of the T2 and T3 vertebral bodies. Chest X-ray and computerized tomography demonstrated post-tuberculous apical cavities with suspected fungal colonization. Histopathology of an extradural spinal lesion at T1/T2 suggested invasive aspergillosis. The patient was started on fluconazole in response to the histopathological evidence of Aspergillus infection, but died within 3 weeks. Post-mortem analysis of the biopsy sample by PCR identified the infectious agent as Aspergillus fumigatus. Atypically, his CD4+ T-cell count was 239 cells mm−3 and he had no evidence of neutropenia. Invasive aspergillosis should be considered as part of the differential diagnosis among HIV-infected patients with non-specific, focal CNS symptoms, even among those without classical risk factors such as neutropenia, and aggressive antifungal therapy should be instituted as early as possible.
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Weissella confusa: a rare cause of vancomycin-resistant Gram-positive bacteraemia
We describe a case of bacteraemia caused by Weissella confusa in a 48-year-old male who was operated on for adenocarcinoma of the gastro-oesophageal junction and maintained on total parenteral nutrition. Blood cultures were positive for a vancomycin-resistant streptococcus-like organism which was identified as W. confusa by 16S rRNA gene sequencing.
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Osteoarticular infection by Candida albicans in an infant with cystic fibrosis
More LessInvasive candidiasis is rare in children after the neonatal period, but can occur in children with (secondary) immunodeficiency with a damaged gastrointestinal or skin barrier, or when receiving antibiotics. A 10-month-old girl was diagnosed as suffering from cystic fibrosis (CF) when showing failure to thrive, pulmonary symptoms and hypoproteinaemia. At that time, Candida albicans was identified from blood culture and treated intravenously with liposomal amphotericin B for 13 days. Six weeks later, the girl presented with osteoarticular infection of the left knee caused by C. albicans. The infection showed insufficient response to therapy with liposomal amphotericin B, but the patient recovered after therapy with fluconazole and flucytosine. Follow-up over 4 years revealed no sequelae. In conclusion, invasive Candida infections may occur in patients with CF, and preventive measures might be considered in patients at risk. In the case of an invasive infection, prolonged treatment with a combination of antifungal drugs may be required.
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The new health legacy: when pertussis becomes a heritage transmitted from mothers to infants
More LessDespite high vaccination coverage rates, there has been a gradual increase in reported pertussis cases. Although whooping cough affects all ages, young infants continue to suffer the greatest pertussis disease burden. Adolescents and adults are the primary source of infection for young babies. In this paper, we report two cases involving the likely transmission of pertussis from mothers to infants in Tunisia.
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Volumes and issues
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Volume 74 (2025)
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