In vitro and in vivo evaluation of antifungal combinations against azole-resistant Aspergillus fumigatus isolates

Azole resistance in Aspergillus fumigatus (Af) has become a widespread threat and a major concern for optimal management of patients with invasive aspergillosis (IA). Combination of echinocandins with azoles is an attractive alternative option for the treatment of IA due to azole-resistant Af strains. The aim of this study was to evaluate the in vitro and in vivo combination of caspofungin (CAS) with either voriconazole (VRZ) or posaconazole (PSZ). In vitro interactions were assessed by two methods, and an animal model of IA in Galleria mellonella was used for in vivo evaluation. Assessment of efficacy was based on larvae mortality. Groups of 10 larvae were infected by 3 clinical strains of Af (azole susceptible, AfS; PSZ resistant, AfR1; VRZ and PSZ resistant strain, AfR2). In vitro, combination of CAS and azoles was indifferent against AfS, and AfR2, and a synergy was found for AfR1. When compared to VRZ monotherapy, the combination of VRZ at 4 µg/larva with CAS at 4 µg/larva improved survival of AfR2-infected larvae (p=0.0066). Combination of PSZ at 4µg/larva with CAS at 4 µg/larva improved survival of AfR1-infected larvae compared to CAS (p=0.0002) and PSZ (0.0024) monotherapy. Antagonism was never observed. In conclusion, the combination of caspofungin with azoles is a promising alternative for the treatment of azole resistant strains of Af

Case Report: Allergic Bronchopulmonary Aspergillosis Revealing Asthma

Allergic bronchopulmonary aspergillosis (ABPA) is an immunological pulmonary disorder caused by hypersensitivity to Aspergillus which colonizes the airways of patients with asthma and cystic fibrosis. Its diagnosis could be difficult in some cases due to atypical presentations especially when there is no medical history of asthma. Treatment of ABPA is frequently associated to side effects but cumulated drug toxicity due to different molecules is rarely reported. An accurate choice among the different available molecules and effective on ABPA is crucial. We report a case of ABPA in a woman without a known history of asthma. She presented an acute bronchitis with wheezing dyspnea leading to an acute respiratory failure. She was hospitalized in the intensive care unit. The bronchoscopy revealed a complete obstruction of the left primary bronchus by a sticky greenish material. The culture of this material isolated Aspergillus fumigatus and that of bronchial aspiration fluid isolated Pseudomonas aeruginosa. The diagnosis of ABPA was based on elevated eosinophil count, the presence of specific IgE and IgG against Aspergillus fumigatus and left segmental collapse on chest computed tomography. The patient received an inhaled treatment for her asthma and a high dose of oral corticosteroids for ABPA. Her symptoms improved but during the decrease of corticosteroids, the patient presented a relapse. She received itraconazole in addition to corticosteroids. Four months later, she presented a drug-induced hepatitis due to itraconazole which was immediately stopped. During the monitoring of her asthma which was partially controlled, the patient presented an aseptic osteonecrosis of both femoral heads that required surgery. Nine months after itraconazole discontinuation, she presented a second relapse of her ABPA. She received voriconazole for nine months associated with a low dose of systemic corticosteroid therapy with an improvement of her symptoms. After discontinuation of antifungal treatment, there was no relapse for one year follow-up

Estimation of the economic impact of a bluetongue serotype 4 outbreak in Tunisia

IntroductionSince 1999, Tunisia has experienced multiple occurrences of Bluetongue (BT) outbreaks, leading to numerous reported cases of infection and mortality in flocks. The re-emergence of the disease in 2020 caused substantial economic losses in cattle, attributed to the incursion of serotype BTV-4.MethodsTo evaluate the economic impact of the recent BT episode, we conducted a retrospective study on outbreaks that occurred in Tunisia between August and November 2020, focusing on the impact at the owner’s level and its effects on both small ruminants and cattle. A total of 234 ruminant farms (sheep, cattle, and mixed) were randomly selected across Tunisian governorates and included in the study to estimate both the direct and indirect costs of these outbreaks.ResultsTotal costs were calculated as the sum of losses and expenditures resulting from the BT outbreaks. At the animal level, total losses were estimated to range between 116.280 and 207.086 TND for one infected ewe (€33.721 and 60.055). For one lactating cow, costs varied between 2,590.724 and 3,171.107 TND (€751.310 and 919.621). In cattle, exposure to BTV led to a daily unit milk yield decrease of 12.50 to 14.66 L over an average period of 5 months. Diseased sheep experienced weight loss ranging between 4 and 10 kg during the BT outbreaks. The total mean cost of the 2020 BT outbreak in Tunisian investigated farms was estimated at 1,935 million TND (million €561.15) (range: 1,489 and 2,474 million TND; 431.81 and million €717.46). The most influential costs of the total BT outbreaks were the decrease in milk yield, mortality, and veterinary treatment.DiscussionThis study gives valuable insights on the economic impact of the incursion of a new serotype of BT in a naive population in Tunisia. Considering the substantial costs incurred, it is imperative that this disease receives increased attention from stakeholders, including animal owners, veterinary services, practitioners, and decision-makers

Implementation of the One Health approach to fight arbovirus infections in the Mediterranean and Black Sea Region: Assessing integrated surveillance in Serbia, Tunisia and Georgia

Background In the Mediterranean and Black Sea Region, arbovirus infections are emerging infectious diseases. Their surveillance can benefit from one health inter-sectoral collaboration; however, no standardized methodology exists to study One Health surveillance. Methods We designed a situation analysis study to document how integration of laboratory/clinical human, animal and entomological surveillance of arboviruses was being implemented in the Region. We applied a framework designed to assess three levels of integration: policy/institutional, data collection/data analysis and dissemination. We tested the use of Business Process Modelling Notation (BPMN) to graphically present evidence of inter-sectoral integration. Results Serbia, Tunisia and Georgia participated in the study. West Nile Virus surveillance was analysed in Serbia and Tunisia, Crimea-Congo Haemorrhagic Fever surveillance in Georgia. Our framework enabled a standardized analysis of One Health surveillance integration, and BPMN was easily understandable and conducive to detailed discussions among different actors/institutions. In all countries, we observed integration across sectors and levels except in data collection and data analysis. Data collection was interoperable only in Georgia without integrated analysis. In all countries, surveillance was mainly oriented towards outbreak response, triggered by an index human case. Discussion The three surveillance systems we observed prove that integrated surveillance can be operationalized with a diverse spectrum of options. However, in all countries, the integrated use of data for early warning and inter-sectoral priority setting is pioneeristic. We also noted that early warning before human case occurrence is recurrently not operationally prioritized

Implementation of zoning to guide management of foot and mouth disease in Tunisia

International audienceThe World Organisation for Animal Health advocates the zoning approach for the surveillance and monitoring of foot and mouth disease (FMD), a highly contagious animal disease. Our purpose is to implement the zoning approach in Tunisia by identifying existing natural and artificial barriers to the movement of live animals. A Geographic Information System (GIS)-based MultiCriteria Evaluation approach was developed. Eight national and international experts were asked to identify the barriers and prioritize them, characterized by a percentage weight between 0 and 100. These barriers were mapped and combined, taking into account their relative importance, to create a friction map that makes it possible to visualize areas where animal movements are restricted. Uncertainty analysis was performed to assess the robustness of the model. The results showed that the selected barriers were in order of decreasing importance: maritime borders with a weight of 33.5%, rivers (13.8%), slopes equal to or greater than 10% (13.8%), wetlands (13.3%), forests (7.7%), land borders (7.7%), railway networks (5%) and main roads (4.9%). The Cap Bon zone is the only favourable zoning area for the control of FMD in Tunisia. A regional approach resulting from this work could be a major asset in identifying regions suitable for zoning in North Africa

In Vivo Efficacy of Voriconazole in a Galleria mellonella Model of Invasive Infection Due to Azole-Susceptible or Resistant Aspergillus fumigatus Isolates

International audienceAspergillus fumigatus is an environmental filamentous fungus responsible for life-threatening infections in humans and animals. Azoles are the first-line treatment for aspergillosis, but in recent years, the emergence of azole resistance in A. fumigatus has changed treatment recommendations. The objective of this study was to evaluate the efficacy of voriconazole (VRZ) in a Galleria mellonella model of invasive infection due to azole-susceptible or azole-resistant A. fumigatus isolates. We also sought to describe the pharmacokinetics of VRZ in the G. mellonella model. G. mellonella larvae were infected with conidial suspensions of azole-susceptible and azole-resistant isolates of A. fumigatus. Mortality curves were used to calculate the lethal dose. Assessment of the efficacy of VRZ or amphotericin B (AMB) treatment was based on mortality in the lethal model and histopathologic lesions. The pharmacokinetics of VRZ were determined in larval hemolymph. Invasive fungal infection was obtained after conidial inoculation. A dose-dependent reduction in mortality was observed after antifungal treatment with AMB and VRZ. VRZ was more effective at treating larvae inoculated with azole-susceptible A. fumigatus isolates than larvae inoculated with azole-resistant isolates. The concentration of VRZ was maximal at the beginning of treatment and gradually decreased in the hemolymph to reach a Cmin (24 h) between 0.11 and 11.30 mg/L, depending on the dose. In conclusion, G. mellonella is a suitable model for testing the efficacy of antifungal agents against A. fumigatus

Image_2_In vitro and in vivo evaluation of antifungal combinations against azole-resistant Aspergillus fumigatus isolates.tif

Azole resistance in Aspergillus fumigatus (Af) has become a widespread threat and a major concern for optimal management of patients with invasive aspergillosis (IA). Combination of echinocandins with azoles is an attractive alternative option for the treatment of IA due to azole-resistant Af strains. The aim of this study was to evaluate the in vitro and in vivo combination of caspofungin (CAS) with either voriconazole (VRZ) or posaconazole (PSZ). In vitro interactions were assessed by two methods, and an animal model of IA in Galleria mellonella was used for in vivo evaluation. Assessment of efficacy was based on larvae mortality. Groups of 10 larvae were infected by 3 clinical strains of Af (azole susceptible, AfS; PSZ resistant, AfR1; VRZ and PSZ resistant strain, AfR2). In vitro, combination of CAS and azoles was indifferent against AfS, and AfR2, and a synergy was found for AfR1. When compared to VRZ monotherapy, the combination of VRZ at 4 µg/larva with CAS at 4 µg/larva improved survival of AfR2-infected larvae (p=0.0066). Combination of PSZ at 4µg/larva with CAS at 4 µg/larva improved survival of AfR1-infected larvae compared to CAS (p=0.0002) and PSZ (0.0024) monotherapy. Antagonism was never observed. In conclusion, the combination of caspofungin with azoles is a promising alternative for the treatment of azole resistant strains of Af.</p

High Frequency of Enterocytozoon bieneusi Genotype WL12 Occurrence among Immunocompromised Patients with Intestinal Microsporidiosis

International audienceMicrosporidiosis is an emerging opportunistic infection causing severe digestive disorders in immunocompromised patients. The aim of this study was to investigate the prevalence of intestinal microsporidia carriage among immunocompromised patients hospitalized at a major hospital complex in the Tunis capital area, Tunisia (North Africa), and perform molecular epidemiology and population structure analyses of Enterocytozoon bieneusi, which is an emerging fungal pathogen. We screened 250 stool samples for the presence of intestinal microsporidia from 171 patients, including 81 organ transplant recipients, 73 Human Immunodeficiency Virus (HIV)-positive patients, and 17 patients with unspecified immunodeficiency. Using a nested PCR-based diagnostic approach for the detection of E. bieneusi and Encephalitozoon spp., we identified 18 microsporidia-positive patients out of 171 (10.5%), among which 17 were infected with E. bieneusi. Microsporidia-positive cases displayed chronic diarrhea (17 out of 18), which was associated more with HIV rather than with immunosuppression other than HIV (12 out of 73 versus 6 out of 98, respectively, p = 0.02) and correlated with extended hospital stays compared to microsporidia-negative cases (60 versus 19 days on average, respectively; p = 0.001). Strikingly, internal transcribed spacer (ITS)-based genotyping of E. bieneusi strains revealed high-frequency occurrence of ITS sequences that were identical (n = 10) or similar (with one single polymorphic site, n = 3) to rare genotype WL12. Minimum-spanning tree analyses segregated the 17 E. bieneusi infection cases into four distinct genotypic clusters and confirmed the high prevalence of genotype WL12 in our patient population. Phylogenetic analyses allowed the mapping of all 17 E. bieneusi strains to zoonotic group 1 (subgroups 1a and 1b/1c), indicating loose host specificity and raising public health concern. Our study suggests a probable common source of E. bieneusi genotype WL12 transmission and prompts the implementation of a wider epidemiological investigation

Image_1_In vitro and in vivo evaluation of antifungal combinations against azole-resistant Aspergillus fumigatus isolates.tif

Azole resistance in Aspergillus fumigatus (Af) has become a widespread threat and a major concern for optimal management of patients with invasive aspergillosis (IA). Combination of echinocandins with azoles is an attractive alternative option for the treatment of IA due to azole-resistant Af strains. The aim of this study was to evaluate the in vitro and in vivo combination of caspofungin (CAS) with either voriconazole (VRZ) or posaconazole (PSZ). In vitro interactions were assessed by two methods, and an animal model of IA in Galleria mellonella was used for in vivo evaluation. Assessment of efficacy was based on larvae mortality. Groups of 10 larvae were infected by 3 clinical strains of Af (azole susceptible, AfS; PSZ resistant, AfR1; VRZ and PSZ resistant strain, AfR2). In vitro, combination of CAS and azoles was indifferent against AfS, and AfR2, and a synergy was found for AfR1. When compared to VRZ monotherapy, the combination of VRZ at 4 µg/larva with CAS at 4 µg/larva improved survival of AfR2-infected larvae (p=0.0066). Combination of PSZ at 4µg/larva with CAS at 4 µg/larva improved survival of AfR1-infected larvae compared to CAS (p=0.0002) and PSZ (0.0024) monotherapy. Antagonism was never observed. In conclusion, the combination of caspofungin with azoles is a promising alternative for the treatment of azole resistant strains of Af.</p

Table_2_In vitro and in vivo evaluation of antifungal combinations against azole-resistant Aspergillus fumigatus isolates.docx

Azole resistance in Aspergillus fumigatus (Af) has become a widespread threat and a major concern for optimal management of patients with invasive aspergillosis (IA). Combination of echinocandins with azoles is an attractive alternative option for the treatment of IA due to azole-resistant Af strains. The aim of this study was to evaluate the in vitro and in vivo combination of caspofungin (CAS) with either voriconazole (VRZ) or posaconazole (PSZ). In vitro interactions were assessed by two methods, and an animal model of IA in Galleria mellonella was used for in vivo evaluation. Assessment of efficacy was based on larvae mortality. Groups of 10 larvae were infected by 3 clinical strains of Af (azole susceptible, AfS; PSZ resistant, AfR1; VRZ and PSZ resistant strain, AfR2). In vitro, combination of CAS and azoles was indifferent against AfS, and AfR2, and a synergy was found for AfR1. When compared to VRZ monotherapy, the combination of VRZ at 4 µg/larva with CAS at 4 µg/larva improved survival of AfR2-infected larvae (p=0.0066). Combination of PSZ at 4µg/larva with CAS at 4 µg/larva improved survival of AfR1-infected larvae compared to CAS (p=0.0002) and PSZ (0.0024) monotherapy. Antagonism was never observed. In conclusion, the combination of caspofungin with azoles is a promising alternative for the treatment of azole resistant strains of Af.</p