Type VI Secretion System Toxins Horizontally Shared between Marine Bacteria.

The type VI secretion system (T6SS) is a widespread protein secretion apparatus used by Gram-negative bacteria to deliver toxic effector proteins into adjacent bacterial or host cells. Here, we uncovered a role in interbacterial competition for the two T6SSs encoded by the marine pathogen Vibrio alginolyticus. Using comparative proteomics and genetics, we identified their effector repertoires. In addition to the previously described effector V12G01_02265, we identified three new effectors secreted by T6SS1, indicating that the T6SS1 secretes at least four antibacterial effectors, of which three are members of the MIX-effector class. We also showed that the T6SS2 secretes at least three antibacterial effectors. Our findings revealed that many MIX-effectors belonging to clan V are "orphan" effectors that neighbor mobile elements and are shared between marine bacteria via horizontal gene transfer. We demonstrated that a MIX V-effector from V. alginolyticus is a functional T6SS effector when ectopically expressed in another Vibrio species. We propose that mobile MIX V-effectors serve as an environmental reservoir of T6SS effectors that are shared and used to diversify antibacterial toxin repertoires in marine bacteria, resulting in enhanced competitive fitness

Mobility of MIX V-effector/immunity cassettes.

<p>Genome neighborhoods are illustrated using arrows to indicate gene orientation, with gene correspondence between species indicated by black vertical lines and omitted genes indicated in parentheses. (A) The “orphan” Duf2235-containing MIX V-effector/immunity pair (<i>va02265</i>/<i>0</i>, filled red arrows) encoded by the <i>V</i>. <i>alginolyticus</i> 12G01 scaffold (<i>V</i>.<i>alg</i>12G01) falls within a conserved gene neighborhood present in other <i>V</i>. <i>alginolyticus</i> strains (green arrows). The orphan pair is absent from the alternate <i>V</i>. <i>alginolyticus</i> strain genomes and is replaced by an alternate cassette of 3 different genes (open red arrows) in <i>V</i>. <i>alginolyticus</i> NBRC 15630 = ATCC 17749 chromosome 2 (<i>V</i>.<i>alg</i>NBRC15630) and an unknown gene in <i>V</i>. <i>alginolyticus</i> 40B scaffold (<i>V</i>.<i>alg</i>40B). A homologous Duf2235-containing MIX V-effector with a duplicated immunity gene (filled red arrows) can be found in a more distant <i>Vibrio</i> strain: <i>V</i>. <i>anguillarum</i> NB10 chromosome 2 (<i>V</i>.<i>ang</i>NB10) in an alternate gene neighborhood (orange arrows). (B) The Colicin DNase-containing MIX V-effector/immunity pair (<i>vpa1263</i>/<i>vti2</i>, filled red arrows) encoded by <i>V</i>. <i>parahaemolyticus</i> RIMD 2210633 chromosome 2 (<i>V</i>.<i>para</i>.RIMD2) belongs to a genetic island that includes a transposon and phage integrase (pink arrows). The island is not present in similar strains such as <i>V</i>. <i>parahaemolyticus</i> BB22OP chromosome 2 (<i>V</i>.<i>para</i>.BB22OP) that retains the surrounding conserved gene neighborhood (purple arrows). A homologous Colicin DNase-containing MIX V-effector/immunity cassette (filled red arrows) is present in a more distant <i>Vibrio</i> strain, <i>Vibrio campbellii</i> ATCC BAA-1116 (<i>V</i>.<i>camp</i>ATCC), in an alternate gene neighborhood (cyan arrows). Neighborhoods include genes (from left to right): <i>N646_ 3835</i>—<i>N646_3824</i> from <i>V</i>.<i>alg</i>.NBRC15630, <i>V12G01_02235</i>—<i>V12G01_02286</i> and alternate genes <i>V12G01_08143—V12G01_08023</i> from <i>V</i>.<i>alg</i>.12G01, <i>VMC_26590</i>—<i>VMC_26680</i> for <i>V</i>.<i>alg</i>40B, <i>VANGNB10_cII03835—VANGNB10_cII03824</i> for <i>V</i>.<i>ang</i>NB10, <i>VPBB_A1148—VPBB_A1154</i> for <i>V</i>.<i>para</i>.BB22OP, <i>VPA1250—VPA1273</i> and alternate genes <i>VP0094—VP0077</i> from <i>V</i>.<i>para</i>.RIMD, and <i>VIBHAR_00561—VIBHAR_00534</i> from <i>V</i>.<i>camp</i>ATCC. Tspn = transposase, Int = integrase, Imm = immunity, Unk = unknown.</p

<i>V</i>. <i>alginolyticus</i> VaT6SS1-dependent secreted proteins.

<p>^a<i>V</i>. <i>parahaemolyticus</i> RIMD 2210633</p><p>^b NF, not found.</p><p><i>V</i>. <i>alginolyticus</i> VaT6SS1-dependent secreted proteins.</p

Va16922/Va16927, Va18287/Va18282, and Va03175/Va03170/Va03180 are VaT6SS2 effector/immunity pairs.

<p>(A) Schematic representation of the VaT6SS2 gene cluster and effector/immunity pairs. V12G01 locus numbers listed above. Effectors in red, Immunity in green, and tail tube components in orange. (B-D) Viability counts of prey strains containing an empty plasmid or a plasmid for the arabinose-inducible expression of the immunity protein before (0h) and after (4h) co-culture with the indicated attacker strains. Effector/immunity pairs tested were: (B) Va16922/Va16927, (C) Va18287/Va18282, and (D) Va03175/Va03170/Va03180. Asterisks mark statistical significance between sample groups at t = 4h by an unpaired, two tailed student’s t-test (p<0.05).</p

<i>V</i>. <i>alginolyticus</i> VaT6SS2-dependent secreted proteins.

<p>^a<i>V</i>. <i>parahaemolyticus</i> RIMD 2210633</p><p>^b NF, not found.</p><p><i>V</i>. <i>alginolyticus</i> VaT6SS2-dependent secreted proteins.</p

<i>V</i>. <i>parahaemolyticus</i> T6SS1 can deliver an “orphan” <i>V</i>. <i>alginolyticus</i> MIX V-effector.

<p>Viability counts of prey strains before (0h) and after (4h) co-culture with indicated attacker strains. (A) <i>V</i>. <i>parahaemolyticus</i> POR1 prey strains containing an empty plasmid or a plasmid for the arabinose-inducible expression of the immunity protein Va02260 (pVa02260) were co-cultured with POR1 or POR1Δ<i>hcp1</i> attacker strains containing an empty plasmid or a plasmid for the arabinose-inducible expression of the Va02265/Va02260 effector/immunity pair (pVa02265-0). (B) <i>V</i>. <i>cholerae</i> V52Δ<i>vipA</i> prey strains were co-cultured with V52 or V52Δ<i>vipA</i> attacker strains containing an empty plasmid or a plasmid for the arabinose-inducible expression of the Va02265/Va02260 effector/immunity pair. Asterisks mark statistical significance between sample groups at t = 4h by an unpaired, two tailed student’s t-test (p<0.05).</p

Defining and managing COVID-19-associated pulmonary aspergillosis: the 2020 ECMM/ISHAM consensus criteria for research and clinical guidance

International audienceSevere acute respiratory syndrome coronavirus 2 causes direct damage to the airway epithelium, enabling aspergillus invasion. Reports of COVID-19-associated pulmonary aspergillosis have raised concerns about it worsening the disease course of COVID-19 and increasing mortality. Additionally, the first cases of COVID-19-associated pulmonary aspergillosis caused by azole-resistant aspergillus have been reported. This article constitutes a consensus statement on defining and managing COVID-19-associated pulmonary aspergillosis, prepared by experts and endorsed by medical mycology societies. COVID-19-associated pulmonary aspergillosis is proposed to be defined as possible, probable, or proven on the basis of sample validity and thus diagnostic certainty. Recommended first-line therapy is either voriconazole or isavuconazole. If azole resistance is a concern, then liposomal amphotericin B is the drug of choice. Our aim is to provide definitions for clinical research and up-to-date recommendations for clinical management of the diagnosis and treatment of COVID-19-associated pulmonary aspergillosis

Isavuconazole treatment for mucormycosis: a single-arm open-label trial and case-control analysis

Background Mucormycosis is an uncommon invasive fungal disease with high mortality and few treatment options. Isavuconazole is a triazole active in vitro and in animal models against moulds of the order Mucorales. We assessed the efficacy and safety of isavuconazole for treatment of mucormycosis and compared its efficacy with amphotericin B in a matched case-control analysis. Methods In a single-arm open-label trial (VITAL study), adult patients (>= 18 years) with invasive fungal disease caused by rare fungi, including mucormycosis, were recruited from 34 centres worldwide. Patients were given isavuconazole 200 mg (as its intravenous or oral water-soluble prodrug, isavuconazonium sulfate) three times daily for six doses, followed by 200 mg/day until invasive fungal disease resolution, failure, or for 180 days or more. The primary endpoint was independent data review committee-determined overall response-ie, complete or partial response (treatment success) or stable or progressive disease (treatment failure)-according to prespecified criteria. Mucormycosis cases treated with isavuconazole as primary treatment were matched with controls from the FungiScope Registry, recruited from 17 centres worldwide, who received primary amphotericin B-based treatment, and were analysed for day-42 all-cause mortality. VITAL is registered with ClinicalTrials.gov, number NCT00634049. FungiScope is registered with ClinicalTrials. gov, number NCT01731353. Findings Within the VITAL study, from April 22, 2008, to June 21, 2013, 37 patients with mucormycosis received isavuconazole for a median of 84 days (IQR 19-179, range 2-882). By day 42, four patients (11%) had a partial response, 16 (43%) had stable invasive fungal disease, one (3%) had invasive fungal disease progression, three (8%) had missing assessments, and 13 (35%) had died. 35 patients (95%) had adverse events (28 [76%] serious). Day-42 crude all-cause mortality in seven (33%) of 21 primary-treatment isavuconazole cases was similar to 13 (39%) of 33 amphotericin B-treated matched controls (weighted all-cause mortality: 33% vs 41%; p=0.595). Interpretation Isavuconazole showed activity against mucormycosis with efficacy similar to amphotericin B. Isavuconazole can be used for treatment of mucormycosis and is well tolerated

Needles in a haystack: Extremely rare invasive fungal infections reported in FungiScope (R)-Global Registry for Emerging Fungal Infections

Objectives: Emerging invasive fungal infections (IFI) have become a notable challenge. Apart from the more frequently described fusariosis, lomentosporiosis, mucormycosis, scedosporiosis, and certain dematiaceae or yeasts, little is known about extremely rare IFI. Methods: Extremely rare IFI collected in the FungiScope (R) registry were grouped as Dematiaceae, Hypocreales, Saccharomycetales, Eurotiales, Dermatomycetes, Agaricales, and Mucorales. Results: Between 2003 and June 2019, 186 extremely rare IFI were documented in FungiScope (R). Dematiaceae (35.5%), Hypocreales (23.1%), Mucorales (11.8%), and Saccharomycetales (11.3%) caused most IFI. Most patients had an underlying malignancy (38.7%) with acute leukemia accounting for 50% of cancers. Dissemination was observed in 26.9% of the patients. Complete or partial clinical response rate was 68.3%, being highest in Eurotiales (82.4%) and in Agaricales (80.0%). Overall mortality rate was 29.3%, ranging from 11.8% in Eurotiales to 50.0% in Mucorales. Conclusions: Physicians are confronted with a complex variety of fungal pathogens, for which treatment recommendations are lacking and successful outcome might be incidental. Through an international consortium of physicians and scientists, these cases of extremely rare IFI can be collected to further investigate their epidemiology and eventually identify effective treatment regimens. (C) 2020 The British Infection Association. Published by Elsevier Ltd. All rights reserved