Key­no­te lec­tu­res

Zoono­ses and an­ti­mi­cro­bi­al re­sis­tan­ce: col­la­bo­ra­ti­on bet­ween meds and vets
J.A. Wa­ge­naar, E. Broens, D. Spek­snij­der
De­part­ment of In­fec­tious Di­sea­ses and Im­mu­no­lo­gy, Fa­cul­ty of Ve­te­ri­na­ry Me­di­ci­ne, Utrecht Uni­ver­si­ty, The Ne­ther­lands; Wa­ge­nin­gen Bio­ve­te­ri­na­ry Re­search, Le­ly­stad, The Ne­ther­lands; WHO-​Collaborating Cen­ter for Cam­py­lo­bac­ter and OIE Re­fe­ren­ce La­bo­ra­to­ry for Cam­py­lo­bac­te­ri­o­sis.

In coun­tries with hu­mans and ani­mals li­ving clo­se­ly to­gether in high den­si­ties, there is a con­ti­nuous th­re­at of trans­mis­si­on of zoo­n­o­tic pa­tho­gens and an­ti­mi­cro­bi­al re­sis­tant (AMR) or­ga­nis­ms bet­ween ani­mals and hu­mans. This is not re­stric­ted to oc­cu­pa­ti­o­nal ex­po­sed pe­o­p­le but also a risk for the ge­ne­ral po­pu­la­ti­on. In The Ne­ther­lands, the emer­gen­ce of re­sis­tant bac­te­ria in li­ves­tock and their trans­mis­si­on to hu­mans (e.g. Li­ves­tock Associated-​MRSA, ESBL pro­du­cing E. coli), and the lar­gest Q-​fever out­break in hu­mans ever re­por­ted with its ori­gin in goat farms, led to a gro­wing con­cern about pos­si­ble human he­alth im­pli­ca­ti­ons of li­ves­tock pro­duc­ti­on. As a con­se­quen­ce of these events, the col­la­bo­ra­ti­on bet­ween pu­blic he­alth (me­di­cal doc­tors) and ve­te­ri­na­ri­ans in­ten­si­fied with mu­tu­al res­pect for their res­pon­si­bi­li­ties. To pro­tect pu­blic he­alth, the ani­mal sec­tors (far­mers and ve­te­ri­na­ri­ans) ma­na­ged to achie­ve an al­most 70% re­duc­ti­on in an­ti­mi­cro­bi­al use (AMU) in farm ani­mals over the last 9 years. The use of an­ti­mi­cro­bi­als de­fi­ned as ‘’cri­ti­cally im­por­tant for human he­alth’’ (flu­oro­quino­lo­nes and 3rd and 4th ge­ne­ra­ti­on cep­ha­losporins) in li­ves­tock re­du­ced to al­most zero. Pa­ral­lel to re­duc­ti­on of AMU there was a re­duc­ti­on of AMR in li­ves­tock ob­ser­ved as re­por­ted in the com­bi­ned NethMap-​MARAN re­port - ano­ther example of in­te­gra­ti­on bet­ween me­di­cal and ve­te­ri­na­ry in­fec­tious di­sea­ses do­mains. In this lec­tu­re se­ve­r­al case stu­dies will high­light the power of the col­la­bo­ra­ti­on.

MALDI-​TOF MS de­ter­mi­na­ti­on of fungi
M. Hen­d­rickx
My­co­lo­gy and Ae­ro­bi­o­lo­gy, BCCM/IHEM col­lec­ti­on, Sci­en­ti­fic In­sti­tu­te of Pu­blic He­alth, Brus­sels, Bel­gi­um

The dia­gno­sis of in­va­si­ve fungal di­sea­se remains chal­len­ging in the cli­ni­cal la­bo­ra­to­ry. The use of MALDI-​TOF MS for the iden­ti­fi­ca­ti­on of micro-​organisms has suc­ces­sful­ly been in­tro­du­ced in cli­ni­cal la­bo­ra­to­ries, but its use for the iden­ti­fi­ca­ti­on of fi­la­men­tous fungi remains less wi­de­ly in­tro­du­ced.
Most stu­dies re­port very high ac­cu­r­a­cy though for the iden­ti­fi­ca­ti­on of fi­la­men­tous fungi by MALDI-​TOF MS. Its cost ef­fec­ti­ve­ness, short ana­ly­sis time, low error rate and the fact that it can also dis­cri­mi­na­te bet­ween clo­se­ly re­la­ted and cryp­tic spe­cies, makes it ap­prop­ri­a­te for im­ple­men­ta­ti­on in the cli­ni­cal rou­ti­ne. Two draw­backs remain the avai­la­bi­li­ty of ex­ten­ded re­fe­ren­ce spec­tra da­ta­ba­ses and the fact that this tech­ni­que can only be ap­plied on iso­la­tes.
At the BCCM/IHEM col­lec­ti­on, an ex­ten­si­ve da­ta­ba­se of re­fe­ren­ce spec­tra, co­ve­ring all me­di­cally re­le­vant fungal spe­cies has been de­vel­o­ped and va­li­da­ted for its use on cli­ni­cal iso­la­tes. Its use on se­ve­r­al fungal groups such as der­ma­top­hy­tes or mem­bers of the genus Fusa­ri­um, or of the As­per­g­il­lus niger group has been eva­lu­a­ted. Mo­re­over, the iden­ti­fi­ca­ti­on of fungal strains using this in house cre­a­ted da­ta­ba­se has been im­ple­men­ted in the qua­li­ty con­trol of the BCCM/IHEM col­lec­ti­on (ISO 17025 ac­cre­di­ted). More re­cent­ly, an on­li­ne iden­ti­fi­ca­ti­on tool has been pre­sen­ted that al­lows re­searchers or me­di­cal prac­ti­ti­o­ners to upload their MALDI-​TOF MS spec­trum and to ob­tain the iden­ti­fi­ca­ti­on of their strain.
In con­clu­si­on, MALDI-​TOF MS is a rapid, ro­bust and po­werful tool for the iden­ti­fi­ca­ti­on of micro-​organisms, in­clu­ding fi­la­men­tous fungi.
The avai­la­bi­li­ty of an ex­ten­si­ve and re­lia­ble da­ta­ba­se is in­dis­pen­sa­ble.

Culture-​independent tar­ge­ted next ge­ne­ra­ti­on se­quen­cing of the 16S-​23S rRNA re­gi­on for the iden­ti­fi­ca­ti­on of bac­te­ri­al spe­cies di­rect­ly from cli­ni­cal sam­ples: op­por­tu­ni­ties and chal­len­ges

A.M.D. Kooistra-​Smid,1,2 E. van Zan­ten1, G.J. Wis­se­link1, A.J. Sabat2, V. Ak­ker­boom2, A. Ott1, W.H.M. Vo­gels1, G.D. Mit­hoe1, R.F. de Boer1, A.W. Frie­d­rich2, J.W.A. Ros­sen2
1De­part­ment of Me­di­cal Mi­cro­bi­o­lo­gy, Certe, Gro­nin­gen, The Ne­ther­lands, 2De­part­ment of Me­di­cal Mi­cro­bi­o­lo­gy, Uni­ver­si­ty of Gro­nin­gen, Uni­ver­si­ty Me­di­cal Cen­ter Gro­nin­gen, Gro­nin­gen, The Ne­ther­lands

Ac­cu­ra­te and rapid spe­cies iden­ti­fi­ca­ti­on is es­sen­ti­al for suc­ces­sful tre­at­ment and cli­ni­cal ma­na­ge­ment of bac­te­ri­al in­fec­ti­ons. De­tec­ti­on and iden­ti­fi­ca­ti­on of bac­te­ri­al spe­cies high­ly de­pends on cul­tu­re. The role of mo­le­cu­lar tests is still gro­wing. Howe­ver, both cul­tu­re and mo­le­cu­lar me­thods have se­rious li­mita­ti­ons; cul­tu­re yield may be ham­pe­red in case of slow-​growing and fast­i­di­ous bac­te­ria. PCR-​based me­thods are rapid and sen­si­ti­ve, but need an a pri­o­ri know­led­ge of the li­ke­ly pa­tho­ge­nic spe­cies that might be pre­sent in cli­ni­cal sam­ples. Fur­ther­mo­re, dif­fe­ren­ti­a­ti­on of mul­ti­ple bac­te­ri­al spe­cies in cli­ni­cal sam­ples is al­most not fea­si­ble with San­ger se­quen­cing. Pre­vious­ly, we de­vel­o­ped an easy-​to use, culture-​independent me­thod, based on Next Ge­ne­ra­ti­on Se­quen­cing (NGS) of PCR am­pli­cons en­com­pas­sing the en­ti­re 16S-​23S rRNA re­gi­on, to im­pro­ve bac­te­ri­al spe­cies iden­ti­fi­ca­ti­on (Sabat et al. Sci Rep. 2017). Here, new op­por­tu­ni­ties and chal­len­ges of 16S-​23S rDNA NGS will be dis­cus­sed. Fur­ther­mo­re, the re­sults of 16S-​23S rDNA NGS ana­ly­sis ap­plied di­rect­ly on cli­ni­cal sam­ples as part of a va­li­da­ti­on study will be pre­sen­ted.

NGS of the 16S-​23S rRNA re­gi­on has the po­ten­ti­al to in­crea­se the dia­gnos­tic yield of bac­te­ria in­vol­ved in com­plex in­fec­ti­ons. It also ena­bles de­tec­ti­on of unan­ti­ci­pa­ted bac­te­ri­al pa­tho­gens. Howe­ver, this ap­pro­ach needs fur­ther va­li­da­ti­on. Fur­ther­mo­re, stu­dies that focus on cli­ni­cal re­le­van­ce are ne­ces­sa­ry to de­ter­mi­ne the ap­pli­ca­bi­li­ty of this NGS-​based ap­pro­ach in rou­ti­ne dia­gnos­tics. Fi­nal­ly, mul­ti­dis­ci­pli­na­ry teams are nee­ded to share their know­led­ge, in order to trans­la­te the re­sults of this new test in a re­port that meets the needs of tre­a­ting phy­si­cians.

Tick-​borne re­lapsing fever Bor­re­lia: right in our bac­ky­ard?
A. Wa­ge­ma­kers1,2, J. Koets­veld2, H. Sprong3, J.W. Ho­vi­us2
1De­part­ment of Me­di­cal Mi­cro­bi­o­lo­gy and In­fec­ti­on Pre­ven­ti­on, VUmc, Am­ster­dam, 2Cen­ter for Ex­pe­ri­men­tal and Mo­le­cu­lar Me­di­ci­ne, AMC, Am­ster­dam, 3Cen­ter for In­fec­tious Di­sea­se Con­trol, RIVM, Bilt­ho­ven

The genus Bor­re­lia can be di­vi­ded into Lyme Bor­re­lia spe­cies (Bor­re­lia burg­dor­fe­ri sensu lato, s.l.) and re­lapsing fever Bor­re­lia spe­cies. Most re­lapsing fever spe­cies are found in soft ticks, with the hig­hest di­sea­se bur­den in Se­negal (B. cro­ci­d­urae) and Tan­za­nia (B. dut­to­nii). These spi­ro­che­tes cause fever with a re­lapsing pat­tern due to their abi­li­ty to switch se­ro­ty­pes, ena­bling mi­no­ri­ty se­ro­ty­pes to evade the host an­ti­bo­dy res­pon­se. One re­lapsing fever Bor­re­lia spe­cies, Bor­re­lia miy­a­mo­toi, is found in hard (Ixo­des) ticks, which are the vec­tor of B. burg­dor­fe­ri s.l. and many other human pa­tho­gens. In­deed, we iden­ti­fied B. miy­a­mo­toi in 2.5% of Dutch Ixo­des ticks. In­te­res­tin­gly, B. miy­a­mo­toi and B. burg­dor­fe­ri s.l. were found in the same ticks more often than ex­pec­ted, sug­ge­s­ting si­mi­lar re­ser­voir hosts. In­deed, we found 9% of wild ro­dents and 8% of birds in The Ne­ther­lands to be in­fec­ted with B. miy­a­mo­toi. Fur­ther­mo­re, in an im­mu­no­com­pro­mi­sed pa­tient from Zand­voort with a meningo-​encephalitis we de­tec­ted B. miy­a­mo­toi in the CSF by PCR, mar­king the first Eu­ro­pean B. miy­a­mo­toi pa­tient. Next, we de­vel­o­ped a cul­tu­re me­thod for B. miy­a­mo­toi, which ena­b­led us to study B. miy­a­mo­toi pa­tho­ge­ne­sis. Si­mi­lar to other tick-​borne re­lapsing fever (TBRF) spi­ro­che­tes, and in con­trast to B. burg­dor­fe­ri s.l., B. miy­a­mo­toi is pre­do­mi­nant­ly pre­sent in the blood com­part­ment. Like other TBRF spe­cies, it also eva­des host an­ti­bo­dy res­pon­ses due to the emer­gen­ce of mi­no­ri­ty se­ro­ty­pes with dif­fe­rent var iable major pro­teins (Vmps). Using these Vmp an­ti­gens we were able to de­tect an­ti­bo­dy res­pon­ses in PCR-​confirmed B. miy­a­mo­toi-​infected pa­tients.

Tick-​borne en­cep­ha­li­tis
S. Van Den Brou­c­ke, U. Ma­niew­ski, E. Bot­tie­au, M. Van Es­broeck
De­part­ment of Cli­ni­cal Sci­en­ces, In­sti­tu­te of Tro­pi­cal Me­di­ci­ne, Ant­werp, Bel­gi­um

In Bel­gi­um, only few cases of Tick-​borne en­cep­ha­li­tis (TBE) are dia­gno­sed an­nu­al­ly, all in pa­tients ac­qui­ring in­fec­ti­on abroad. TBE is cau­sed by 3 clo­se­ly re­la­ted fla­vi­vi­ruses and it in­vol­ves the cen­tral ner­vous sy­s­tem. The tick-​borne en­cep­ha­li­tis virus in­fects a range of hosts in­clu­ding rumi­nants, birds, ro­dents, car­ni­vo­res, hor­ses, and hu­mans. In Eu­ro­pe and Asia bet­ween 10000 and 15000 TBE cases are re­por­ted an­nu­al­ly. This num­ber very li­ke­ly un­de­resti­ma­tes the real in­ci­den­ce. TBE is trans­mit­ted to hu­mans by the bite of a tick (ei­ther Ixo­des per­sul­ca­tus or Ixo­des ri­ci­nus) and oc­ca­si­o­nal­ly fol­lo­wing con­sump­ti­on of in­fec­ted un­pas­teu­ri­zed milk. The ratio of asymp­to­ma­tic in­fec­ti­ons is bet­ween 70% and 98%. The ini­ti­al phase of di­sea­se cor­re­la­tes with vi­re­mia and with non-​specific flu­li­ke sympt­oms. The se­cond phase ma­ni­fests as me­nin­gi­tis, en­cep­ha­li­tis, or me­nin­goen­cep­ha­li­tis. The long-​term prog­no­sis is un­fa­vo­ra­ble in about 40% to 50% of pa­tients who sustain se­que­lae for months to years. As a rule, anti-​TBEV- IgM and usu­al­ly TBEV-​IgG an­ti­bo­dies are pre­sent in the first serum sam­ples taken when CNS sympt­oms ma­ni­fest in the se­cond phase of the di­sea­se. In the first phase of ill­ness, the virus can be iso­la­ted or de­tec­ted by RT-​PCR from blood, but only ra­re­ly is TBEV de­tec­ted at the be­gin­ning of the se­cond phase in CSF and oc­ca­si­o­nal­ly in cases of pro­gres­si­ve di­sea­se. There is no spe­ci­fic an­ti­vi­ral tre­at­ment for TBE and sup­por­ti­ve care is the main­stay of tre­at­ment. Per­so­nal pro­tec­ti­ve me­a­su­res help in pre­ven­ti­on of tick bites. In Eu­ro­pe two vac­ci­nes are li­cen­sed: FSME immun® and En­ce­pur®.

Chro­nic Q fever
C.P. Bleeker-​Rovers
De­part­ment of In­ter­nal Me­di­ci­ne, Rad­boud Uni­ver­si­ty Me­di­cal Cen­ter, Nij­me­gen

Q fever is a zoono­sis cau­sed by the in­tra­cel­lu­lar Gram-​negative coc­co­ba­c­il­lus Coxiel­la bur­ne­tii. Fol­lo­wing pri­ma­ry in­fec­ti­on, 1-5% of all pa­tients de­vel­op chro­nic Q fever with en­do­car­di­tis, in­fec­ted aneu­rys­ms or in­fec­ted vas­cu­lar pros­the­ses as most im­por­tant ma­ni­fe­sta­ti­ons. The du­ra­ti­on bet­ween pri­ma­ry in­fec­ti­on and ma­ni­fe­sta­ti­on of chro­nic in­fec­ti­on may be se­ve­r­al years. Se­ve­r­al risk fac­tors for the de­vel­op­ment of chro­nic in­fec­ti­on have been iden­ti­fied and in­clu­de val­vul­o­pa­thy or prior valve sur­gery, aneu­rysm, vas­cu­lar pros­the­ses, renal in­suf­fi­ci­en­cy, older age, im­mu­no­com­pro­mi­sed state and ma­lig­nan­cy. Dia­gno­sing chro­nic Q fever is dif­fi­cult as pa­tients often pre­sent with non­spe­ci­fic sympt­oms. A final dia­gno­sis re­lies on a com­bi­na­ti­on of cli­ni­cal signs, se­ro­lo­gy, PCR on blood or tis­sue and ra­dio­lo­gi­cal fin­dings. Pa­tients are clas­si­fied as pro­ven, pro­ba­ble or pos­si­ble chro­nic Q fever pa­tients ac­cor­ding to the Dutch chro­nic Q fever con­sen­sus group gui­de­li­ne. Bet­ween 2007 and 2010, there was a large Q fever out­break in The Ne­ther­lands. It is esti­ma­ted that over 40,000 pe­o­p­le were in­fec­ted with major im­pact on phy­si­cal and psy­cho­lo­gi­cal he­alth. Fol­lo­wing this out­break, all known chro­nic Q fever pa­tients were in­clu­ded in an on­go­ing na­ti­on­wi­de re­gi­stra­ti­on (219 pa­tients with pro­ven chro­nic Q fever and 74 with pro­ba­ble chro­nic Q fever). Q fever re­la­ted mor­ta­li­ty was 25% in pa­tients with pro­ven chro­nic Q fever and 4% in pro­ba­ble chro­nic Q fever. Com­pli­ca­ti­ons were as­so­ci­a­ted with chro­nic Q fever-​related mor­ta­li­ty. Based on re­sults from this na­ti­o­nal da­ta­ba­se new in­sights in dia­gno­sis, com­pli­ca­ti­ons, and tre­at­ment will be dis­cus­sed.

 

Pa­ral­lel ses­si­ons

Im­pact of agar rea­ding fre­quen­cy on the re­por­ting of blood cul­tu­re re­sults
B. van den Poel, S. Des­met, J. Ver­hae­gen
De­part­ment of La­bo­ra­to­ry Me­di­ci­ne, UZ Leu­ven, Leu­ven

Rapid iden­ti­fi­ca­ti­on and an­ti­mi­cro­bi­al sus­cep­ti­bi­li­ty (AMS) re­sult of bac­te­ria cau­sing blood stream in­fec­ti­ons is cru­ci­al in the ma­na­ge­ment of sep­tic pa­tients. In this study, we com­pa­red a pe­ri­od of twice-​daily and a pe­ri­od of thrice-​daily rea­ding of sub­cul­tu­re agar pla­tes. In 2016, 10 644 po­si­ti­ve blood cul­tu­res bott­les (bioMérieux) from 2608 pa­tients were ana­ly­zed at UZ Leu­ven. Iden­ti­fi­ca­ti­on and an­ti­mi­cro­bi­al sus­cep­ti­bi­li­ty tes­ting were per­for­med by MALDI-​TOF MS (Bru­ker Dal­to­nics) and Vitek® 2 (bioMérieux) res­pec­ti­ve­ly. In pe­ri­od 1 (Ja­nu­a­ry to June), sub­cul­tu­re pla­tes were read at 8.30 am and 2 pm du­ring the week­days. In pe­ri­od 2 (Au­gust until De­cem­ber), rea­ding was per­for­med at 8.30 am, 2 pm and 5 pm. Time to iden­ti­fi­ca­ti­on and time to AMS re­sult after po­si­ti­vi­ty were com­pa­red. In pe­ri­od 1, me­di­an time to iden­ti­fi­ca­ti­on of all or­ga­nis­ms was 22.8 hours com­pa­red to 20.2 hours in pe­ri­od 2 (p < 0.0001, Wilcoxon-​Mann-Whitney U test). Mo­re­over, micro-​organisms were iden­ti­fied be­fo­re 12 hours in 9% (418/4559) of sam­ples in pe­ri­od 2, a sig­ni­fi­cant in­crea­se com­pa­red to 1.7% (88/5035) in pe­ri­od 1 (p < 0.0001, Fisher-​Exact). In pe­ri­od 2, AMS re­sult was known within 36 hours in 39% (431/1107) of sam­ples, com­pa­red to 31% (409/1337) in pe­ri­od 1 (p < 0.0001, Fisher-​Exact). Op­ti­mi­za­ti­on of the rea­ding fre­quen­cy of sub­cul­tu­res of blood cul­tu­res sig­ni­fi­cantly de­crea­ses time to re­sults. Fur­ther op­ti­mi­za­ti­on can be done by in­tro­du­cing lab au­to­ma­ti­on. We will use the data of this study as ba­se­li­ne to ana­ly­ze the im­pact of in­tro­du­cing WAS­PLab (Copan Dia­gnos­tics) au­to­ma­ti­on on time to re­sult.

Ge­no­mic re­so­lu­ti­on of methicillin-​sensitive Stap­hy­lo­coc­cus au­re­us out­breaks in a ne­o­na­tal in­ten­si­ve care unit
A.J.H. Cre­mers1, J.P.M. Coo­len1, C.P. Bleeker-​Rovers2, D. Ha­ver­ka­te1, A.D.J. van der Geest3, A. van Heijst4, H. Hen­driks4, S.S.V. Hen­riet5, M.A. Huy­nen6, E. Kol­wij­ck1, D. Liem4, W.J.G. Mel­chers1, A. van Sum­me­ren3, J. Zoll1, J. Hop­man1*, H.F.L. Wert­heim1*
1De­part­ment of Me­di­cal Mi­cro­bi­o­lo­gy, Rad­bou­dumc, Nij­me­gen, The Ne­ther­lands, 2De­part­ment of In­ter­nal Me­di­ci­ne, Rad­bou­dumc, Nij­me­gen, The Ne­ther­lands, 3Oc­cu­pa­ti­o­nal He­alth & Sa­fe­ty and En­vi­ron­men­tal Ser­vi­ce, Rad­bou­dumc, Nij­me­gen, The Ne­ther­lands, 4De­part­ment of Ne­o­na­to­lo­gy, Rad­bou­dumc, Nij­me­gen, The Ne­ther­lands, 5De­part­ment of Pe­di­a­trics, Rad­bou­dumc Ama­lia Child­ren’s Hos­pi­tal, Nij­me­gen, The Ne­ther­lands, 6Cen­tre for Mo­le­cu­lar and Bi­o­mo­le­cu­lar In­for­ma­tics, Rad­boud In­sti­tu­te for Mo­le­cu­lar Life Sci­en­ces, Rad­bou­dumc, Nij­me­gen, The Ne­ther­lands, *J. Hop­man and H.F.L. Wert­heim share se­ni­or au­thor­ship

Ac­cu­ra­te re­con­struc­ti­on of out­breaks can di­rect in­fec­ti­on con­trol me­a­su­res. We exa­mi­ned whe­ther whole ge­no­me se­quen­cing (WGS) ana­ly­ses im­pro­ved out­break tra­cing in a ne­o­na­tal in­ten­si­ve care unit (NICU) in com­pa­ri­son with Multiple-​Locus Va­ria­ble num­ber tan­dem re­peat Ana­ly­sis (MLVA) ty­ping.
Du­ring 2014 and 2015 all methicillin-​sensitive Stap­hy­lo­coc­cus au­re­us (MSSA) iso­la­tes from wee­kly thro­at sur­veil­lan­ce cul­tu­res at a third level Dutch NICU were typed by spa and MLVA. On two oc­ca­si­ons, when in­va­si­ve MSSA in­fec­ti­ons see­med to ori­gi­na­te from car­ria­ge out­breaks, all he­alth care wor­kers (HCWs) were tested for MSSA car­ria­ge. Those HCWs who car­ried out­break MLVA types were de­co­lo­ni­zed. WGS of iso­la­tes that cor­res­pon­ded to the out­break spa types was fol­lo­wed by a se­ries of au­to­ma­ted tools in­clu­ding de novo as­sem­bly, iden­ti­fying and lo­ca­li­zing high qua­li­ty sin­gle nu­cle­o­ti­de po­ly­morphis­ms (SNPs), and in depth ana­ly­sis of out­break clus­ters.
MSSA was iso­la­ted in 19% (214/1154) of sur­veil­lan­ce cul­tu­res, and in 24% of HCWs. WGS ana­ly­sis iden­ti­fied iso­la­tes that were, based on MLVA ty­ping, un­just­ly clus­te­red. Fur­ther­mo­re, de­tai­ling par­ti­cu­lar clus­ters im­pro­ved trans­mis­si­on chain re­so­lu­ti­on, and know­led­ge of the dis­tri­bu­ti­on of SNPs across the ge­no­me im­pro­ved ac­cu­r­a­cy of the esti­ma­ted re­la­ted­ness of strains. WGS ana­ly­sis pro­vi­ded evi­den­ce for HCWs being in­vol­ved in both out­break trans­mis­si­on chains. Con­tra­ry to what was con­clu­ded from clas­si­cal ty­ping me­thods, a HCW in­vol­ved in the first out­break had re-​acquired a ne­ar­ly iden­ti­cal MSSA strain, which was howe­ver un­re­la­ted to the se­cond out­break.
WGS ana­ly­sis im­pro­ved the re­con­struc­ti­on of MSSA out­breaks, with im­por­tant im­pli­ca­ti­ons for HCWs in­vol­ved.

Can in­fec­ti­on pre­ven­ti­on me­a­su­res for Clos­tri­di­um dif­fi­ci­le be tail­ored to spe­ci­fic strain types?
R. van Houdt1, A. Zomer2, J. van Prehn1, R. van Mans­feld1, C.M.J.E. Vandenbroucke-​Grauls1
1Me­di­cal Mi­cro­bi­o­lo­gy and In­fec­ti­on Con­trol, VU Uni­ver­si­ty Me­di­cal Cen­ter, Am­ster­dam, The Ne­ther­lands; 2De­part­ment of In­fec­tious Di­sea­ses and Im­mu­no­lo­gy, Utrecht Uni­ver­si­ty, Utrecht, The Ne­ther­lands

Du­ring a C. dif­fi­ci­le out­break in the VUmc in 2013-​2014, 86 pa­tients were in­fec­ted with the B1/NAP1/027 strain. Be­si­des trans­mis­si­on of this 027 out­break strain, there were also C. dif­fi­ci­le in­fec­ti­ons with other ri­bo­ty­pe strains. In order to as­sess whe­ther in­fec­ti­ons with other ri­bo­ty­pes were no­so­co­mi­al or not, whole ge­no­me se­quen­cing was per­for­med on a random se­lec­ti­on of the se­cond most pre­va­lent ri­bo­ty­pe, 014.
After se­quen­cing iso­la­tes from 15 random­ly se­lec­ted in­pa­tients from 2014, sin­gle nu­cle­o­ti­de po­ly­morphism (SNP) ana­ly­sis was used to de­ter­mi­ne the phy­lo­ge­ne­tic re­la­ti­ons­hip and com­pa­red with the SNP ana­ly­sis of the ri­bo­ty­pe 027 strains from the out­break.
In 2014, 101 pa­tients had a C. dif­fi­ci­le in­fec­ti­on of which 25 were cau­sed by the ri­bo­ty­pe 014 strain. Core ge­no­me SNP ana­ly­sis of 16 ri­bo­ty­pe 027 strains showed a maxi­mum of 2 SNPs dif­fe­ren­ce. SNP ana­ly­sis of 15 ri­bo­ty­pe 014 strains re­sul­ted in 4 dis­tinct clus­ters, 3 clus­ters con­sis­ting of 2 strains and 1 clus­ter con­sis­ting of 3 strains, with a maxi­mum of 10 and 26 SNPs dif­fe­ren­ce, res­pec­ti­ve­ly. The other 6 strains were un­re­la­ted.
SNP ana­ly­sis sug­ge­sted that the ri­bo­ty­pe 014 strains were un­re­la­ted strains or small clus­ters, in­di­ca­ting no or litt­le risk on no­so­co­mi­al trans­mis­si­on, as op­po­sed to the pro­ven trans­mis­si­on of the B1/NAP1/027 strain. We sug­gest that strin­gent in­fec­ti­on con­trol me­a­su­res are re­qui­red to pre­vent large out­breaks of ri­bo­ty­pe 027 strains, while we feel that less strin­gent me­a­su­res may be suf­fi­cient to pre­vent out­breaks with C. dif­fi­ci­le other that ri­bo­ty­pe 027.

De­tec­ti­on and dis­cri­mi­na­ti­on of ten cli­ni­cally re­le­vant Can­di­da spp. with a novel real time mo­le­cu­lar assay
C.F.M. van der Donk1, L. Roor­da2, B. Kraak2, A. Burg­graaf2, D. Wil­lems3, M.H.A. Her­mans3, F. Hagen4, A. van der Zee2
1Eras­mus Me­di­cal Cen­tre, De­part­ment of Me­di­cal Mi­cro­bi­o­lo­gy and In­fec­tious Di­sea­ses, Rot­ter­dam, 2Maas­stad Hos­pi­tal, Mo­le­cu­lar Dia­gnos­tics Unit, Rot­ter­dam, 3Je­roen Bosch Hos­pi­tal, De­part­ment of Me­di­cal Mi­cro­bi­o­lo­gy, ’s-​Hertogenbosch, 4Ca­ni­si­us Wil­hel­mi­na Hos­pi­tal, De­part­ment of Me­di­cal Mi­cro­bi­o­lo­gy, Nij­me­gen, The Ne­ther­lands

In cur­rent Can­di­da dia­gnos­tics often cul­ti­va­ti­on on se­lec­ti­ve media is used to iden­ti­fy Can­di­da spe­cies. MALDI-​TOF ana­ly­sis of Can­di­da spe­cies im­pro­ved cor­rect iden­ti­fi­ca­ti­on and shor­te­n­ed time for a final dia­gno­sis but a test that dis­cri­mi­na­tes cor­rect­ly bet­ween clo­se­ly re­la­ted spe­cies like C. al­bi­cans and C. du­bli­nien­sis and the C. pa­rapsi­lo­sis spe­cies com­plex is not avai­la­ble. Here we de­scri­be a fast and ac­cu­ra­te real-​time PCR assay for de­tec­ti­on and iden­ti­fi­ca­ti­on of 10 me­di­cally re­le­vant Can­di­da spp.
Pri­mers and pro­bes were de­sig­ned to spe­ci­fi­cally am­pli­fy DNA of Can­di­da al­bi­cans, C. gla­bra­ta, C. du­bli­nien­sis, C. tro­pi­ca­lis, C. lusi­ta­ni­ae, C. pa­rapsi­lo­sis, C. metapsi­lo­sis, C. or­thop­si­lo­sis, C. guil­lier­mon­dii and C. krus­ei. DNA was ex­trac­ted with MagNA Pure 96 and real-​time PCR was per­for­med with Bi­o­rad CFX96 ther­mo­cy­cler. The real-​time PCRs were op­ti­mi­zed and showed good re­sults with re­gard to ef­fi­ci­en­cy, sen­si­ti­vi­ty and re­pro­du­ci­bi­li­ty, and spe­ci­fi­ci­ty.
Va­rious cli­ni­cal ma­te­ri­als were tested for the pre­sen­ce of Can­di­da spp. Rou­ti­ne dia­gnos­tic sam­ples that were cul­tu­re po­si­ti­ve cor­re­la­ted well with res­pect to Cq va­lu­es and growth quan­ti­fi­ca­ti­on. We also de­mon­stra­ted low le­vels of Can­di­da spp. DNA in se­ve­r­al ma­te­ri­als, which is dis­cus­sed.
This paper de­scri­bes a va­li­da­ted and ro­bust real-​time PCR assay with high sen­si­ti­vi­ty, 100% spe­ci­fi­ci­ty, and re­pro­du­ci­bi­li­ty for the de­tec­ti­on and dif­fe­ren­ti­a­ti­on of ten im­por­tant Can­di­da spp. This assay can be used as a sin­gle or mul­ti­plex assay wit­hout loss of sen­si­ti­vi­ty and can be adop­ted to me­di­cally re­qui­red pre­fe­ren­ces.

Eva­lu­a­ti­on of UMIC mi­cro­di­lu­ti­on strip for co­lis­tin and piperacillin-​tazobactam in non-​cystic and cys­tic fi­bro­sis pa­tients
A. Muyl­der­mans, S. Pa­ter­nos­ter, M. Ta­jdar, E. Nu­lens
La­bo­ra­to­ry Me­di­ci­ne, Me­di­cal Mi­cro­bi­o­lo­gy, Al­ge­meen Zie­ken­huis Sint-​Jan Brugge-​Oostende AV, Bru­ges, Bel­gi­um

An­ti­mi­cro­bi­al sus­cep­ti­bi­li­ty tes­ting (AST) for co­lis­tin (COL) and piperacillin-​tazobactam (PTZ) is chal­len­ging. Disk and agar gra­dient dif­fu­si­on me­thods are not ac­cep­ted by EU­CAST, broth mi­cro­di­lu­ti­on (BMD) is the me­thod of choi­ce. In cys­tic fi­bro­sis (CF) pa­tients AST is fur­ther com­pli­ca­ted by the fast­i­di­ous growth of mu­cous bac­te­ria. We eva­lu­a­ted the BMD me­thod UMIC (Bi­o­cen­tric) on iso­la­tes of non-​CF and CF pa­tients.
22 iso­la­tes from non-​CF pa­tients [P. ae­rug­i­n­o­sa (n = 7), B. ce­pa­cia (n = 1), En­te­ro­bac­te­ria­ceae (n = 14)], 17 iso­la­tes from CF pa­tients [P. ae­rug­i­n­o­sa (n = 10), A. xy­loxi­dans (n = 7)] and 9 QC strains from UK Neqas [P. ae­rug­i­n­o­sa (n = 7), En­te­ro­bac­te­ria­ceae (n = 2)] were in­clu­ded. AST of COL and PTZ with UMIC was com­pa­red with our rou­ti­ne me­thod: semi-​automated tes­ting by Phoe­nix in non-​CF, disk dif­fu­si­on in CF and re­fe­ren­ce la­bo­ra­to­ry for QC strains.
In non-​CF pa­tients a ca­te­go­ri­cal agree­ment (CA) of 100% for COL and 86% for PTZ (2 minor er­rors, 1 very major error) was found. In CF pa­tients a CA of 88% for COL (2 minor er­rors) and 100% for PTZ was found, if UMIC was in­cu­ba­ted in CO2 for 48h. In QC strains a CA of 78% for COL (2 very major er­rors) and 86% for PTZ (1 very major error) was found (P. ae­rug­i­n­o­sa iso­la­tes with low-​level re­sis­tan­ce).
A good ca­te­go­ri­cal agree­ment for COL and PTZ was found in non-​CF and CF pa­tients. In CF pa­tients pro­lon­ged in­cu­ba­ti­on in CO2 is ne­ces­sa­ry. Fur­ther eva­lu­a­ti­on is nee­ded to de­ter­mi­ne es­sen­ti­al agree­ment (no re­lia­ble MIC was given with the rou­ti­ne semi-​automated AST).

Eva­lu­a­ti­on of the Ac­ce­le­ra­te Pheno Sy­s­tem for rapid an­ti­mi­cro­bi­al sus­cep­ti­bi­li­ty tes­ting in extended-​spectrum β-​lactamase pro­du­cing En­te­ro­bac­te­ria­ceae
G.L. Vlas­pol­der1, B. San­ha­oui1, L.N. van Bel­zen1, P. Meij­er2, A.N. Spaan1, C.H.E. Boel1
1De­part­ment of Me­di­cal Mi­cro­bi­o­lo­gy, Uni­ver­si­ty Me­di­cal Cen­ter Utrecht, Utrecht, 2Be­ne­lux de­part­ment of Ac­ce­le­ra­te Dia­gnos­tics B.V., Lei­den

Blood­stream in­fec­ti­ons cau­sed by extended-​spectrum-β-​lactamase (ESBL)- and AmpC-​producing En­te­ro­bac­te­ria­ceae are as­so­ci­a­ted with high rates of mor­bi­di­ty and mor­ta­li­ty. Be­cau­se ESBL- and AmpC-​producing bac­te­ria are re­sis­tant to β-​lactam an­ti­bi­o­tics used for em­pi­ric tre­at­ment of in­fec­ti­ons, the em­pi­ri­cal use of car­ba­pe­nems has in­crea­sed. As a con­se­quen­ce, an in­cre­a­sing rate of car­ba­pe­nem re­sis­tan­ce among gram-​negative bac­te­ria is seen. It is a chal­len­ge for every phy­si­cian to em­pi­ri­cally treat pa­tients with ESBL-​bacteremia with ap­prop­ri­a­te an­ti­bi­o­tics and at the same time mi­ni­mi­ze un­ne­ces­sa­ry use of last-​resort an­ti­bi­o­tics in case of sus­cep­ti­ble bac­te­ria. Fas­ter avai­la­bi­li­ty of sus­cep­ti­bi­li­ty test re­sults could allow pathogen-​directed the­ra­py to be star­ted sooner. The Ac­ce­le­ra­te Pheno™ sy­s­tem (AxDx) is a fully au­to­ma­ted sy­s­tem that pro­vi­des MIC-​based an­ti­mi­cro­bi­al sus­cep­ti­bi­li­ty tes­ting (AST) re­sults within seven hours, di­rect­ly from po­si­ti­ve blood cul­tu­res. This study was per­for­med to de­ter­mi­ne the dia­gnos­tic ac­cu­r­a­cy of AST of the AxDx sy­s­tem for ami­no­gly­co­si­des and quino­lo­nes, in order to pro­vi­de ade­qua­te the­ra­py in a ti­me­ly man­ner and to ena­ble a de-​escalation of tre­at­ment in sus­cep­ti­ble bac­te­ria. In total, 34 gram-​negative iso­la­tes, of which 27 iso­la­tes con­sists of ESBL-​producing bac­te­ria, were ana­ly­zed on the AxDx sy­s­tem and com­pa­red to our standard of care (BD Phoe­nix, bioMérieux Vi­tek2, if ne­ces­sa­ry in com­bi­na­ti­on with E-​tests). The over­all ca­te­go­ry agree­ment for ami­no­gly­co­si­des and quino­lo­nes was 92.7%. Of note, gen­ta­my­cin, to­bra­my­cin, amik­a­cin and ci­prof­loxa­cin re­sis­tan­ce was cor­rect­ly de­tec­ted in ESBL-​producing E. coli (n = 5). In con­clu­si­on, the AxDx sy­s­tem pro­vi­des re­lia­ble re­sults and is po­ten­ti­al­ly use­ful for an­ti­mi­cro­bi­al ste­wards­hip in pa­tients with ESBL-​bacteremia.

Im­pro­ve­ment of sur­vi­val of a Stap­hy­lo­coc­cus au­re­us sep­sis after in­vol­ve­ment of the an­ti­bi­o­tic team and a bund­le of in­ter­ven­ti­ons?
M.A.N.P.M. van den Hurk1, J. Fon­vil­le1, H. Am­mer­laan2, C. Mie­de­ma3, S. San­ders4, I. Over­de­vest1
1Mi­cro­bi­o­lo­gy, PAMM, Veld­ho­ven, 2Internal-​infectiology, Ca­tha­ri­na Hos­pi­tal, Eind­ho­ven, 3Pediatric-​infectiology, Ca­tha­ri­na Hos­pi­tal, Eind­ho­ven, 4Hos­pi­tal Phar­ma­co­lo­gy, Ca­tha­ri­na Hos­pi­tal, Eind­ho­ven

Stap­hy­lo­coc­cus au­re­us bac­terae­mia (SAB) is a se­rious cli­ni­cal con­di­ti­on as­so­ci­a­ted with a high mor­ta­li­ty and com­pli­ca­ti­ons such as see­ding. Li­te­ra­tu­re shows that a bund­le of in­ter­ven­ti­ons im­pro­ves the out­co­me of SAB sig­ni­fi­cantly.1 Since 2013, our an­ti­bi­o­tic team is in­cre­a­sin­gly in­vol­ved in ad­vi­sing me­di­cal prac­ti­ti­o­ners on tre­a­ting pa­tients with SAB. In this re­tro­spec­ti­ve co­hort study we eva­lu­a­ted the ef­fect of the an­ti­bi­o­tic team in­vol­ve­ment and of bund­le ad­he­ren­ce on mor­ta­li­ty and re­lap­se SAB.
All adult pa­tients (n = 179) with SAB ad­mit­ted to the Ca­tha­ri­na hos­pi­tal Eind­ho­ven bet­ween 2013-​2015 were in­clu­ded. Thirty-​one pa­tients died < 14 days and were ex­clu­ded. We re­tro­spec­ti­ve­ly de­fi­ned a bund­le of in­ter­ven­ti­ons and sco­red com­pli­an­ce with the fol­lo­wing me­a­su­res; (1) in­vol­ve­ment of the an­ti­bi­o­tic team, in­clu­ding beds­i­de con­sult of an in­fec­tious di­sea­se spe­ci­a­list; (2) follow-​up blood cul­tu­res; (3) ade­qua­te an­ti­bi­o­tic tre­at­ment; (4) sour­ce con­trol; (5) TTE/TEE or PET-​CT when in­crea­sed risk of com­pli­ca­ted in­fec­ti­on.
Re­sults showed im­pro­ved sur­vi­val rates and com­pli­an­ce to the bund­le when the an­ti­bi­o­tic team was in­vol­ved ad­vi­sing the me­di­cal prac­ti­ti­o­ners. The ef­fects were es­pe­ci­al­ly ob­ser­ved with res­pect to ade­qua­te an­ti­bi­o­tic tre­at­ment; 67% vs 52% (p = 0.08) and in­ves­ti­ga­ti­ons on see­ding in high risk pa­tients; 72% vs 57% (p = 0.11). The risk on re­lap­se SAB and mor­ta­li­ty rates de­crea­sed from 21 to 10% (p = 0.11). We ex­pect the re­sults to be an un­de­resti­ma­te con­si­de­ring the un­der­ly­ing bias where the an­ti­bi­o­tic team is pre­do­mi­nant­ly in­vol­ved in the more se­vere­ly ill pa­tients.
This study shows a trend in im­pro­ve­ment of sur­vi­ving SAB and an added value of the an­ti­bi­o­tic team.

Re­fe­ren­ce
1. Luis E. López-​Cortés, Maria Do­lo­res del Toro, Juan Gálvez-​Acebal. Im­pact of an Evidence-​Based Bund­le: In­ter­ven­ti­on in the Quality-​of-Care Ma­na­ge­ment and Out­co­me of Stap­hy­lo­coc­cus au­re­us Bac­tere­mia. Clin Inf Dis 2013:57;1225-​33.

Data have re­cent­ly been sub­mit­ted for pu­bli­ca­ti­on to the ma­ga­zi­ne In­fec­tie­ziek­ten; the pu­bli­ca­ti­on is still under con­si­de­ra­ti­on of the edi­to­ri­al board.

The ti­ge­cy­cli­ne on the field: in­di­ca­ti­ons, ef­fi­ca­cy, to­le­ra­bi­li­ty
A. Pa­pa­leo1, J. Pre­vost2, F. Ja­cobs2
1In­ter­nal Me­di­ci­ne and In­fec­tious Di­sea­ses De­part­ment Hôpi­taux Iris Sud, Brus­sels, Bel­gi­um, 2In­fec­tious Di­sea­ses De­part­ment, Eras­mus Hos­pi­tal, Brus­sels, Bel­gi­um

Ti­ge­cy­cli­ne could re­pre­sent a va­lu­a­ble al­ter­na­ti­ve to car­ba­pe­nem and β-​lactam based re­gi­mens. There are some data about real-​life cli­ni­cal prac­ti­ce from Fran­ce, Ger­ma­ny, Italy, and Spain, but still no data about Bel­gi­um.
To in­ves­ti­ga­te ti­ge­cy­cli­ne pre­scrip­ti­on, to­le­ra­bi­li­ty and pa­tients out­co­me we con­duc­ted a re­tro­spec­ti­ve study in Eras­mus Hos­pi­tal, an 800-​bed aca­de­mic hos­pi­tal in Brus­sels, bet­ween 2007 and 2015.
We in­clu­ded 89 pa­tients. We ob­ser­ved a pro­gres­si­ve in­crea­se in the pre­scrip­ti­on of ti­ge­cy­cli­ne over years. The 60% of pa­tients re­cei­ved ti­ge­cy­cli­ne in in­ten­si­ve care unit (ICU), 35% had an im­mu­no­sup­pres­si­on, 23% had un­der­go­ne solid organ trans­plan­ta­ti­on. The main in­di­ca­ti­ons were pneu­mo­nia in 36% of cases and com­pli­ca­ted intra-​abdominal in­fec­ti­ons (28%) al­most ex­clu­si­ve­ly for healthcare-​associated in­fec­ti­ons (98%). Ti­ge­cy­cli­ne was used after do­cu­men­ted in­fec­ti­ons in 92% of cases, 89% with multi-​drug re­sis­tant (MDR) bac­te­ria whose 78% carbapenemase-​producing En­te­ro­bac­te­ria­ceae. The most iso­la­ted pa­tho­gen was Aci­ne­to­bac­ter bau­man­nii. We used ti­ge­cy­cli­ne after mul­ti­ple an­ti­bi­o­tics in 85% of cases, only in 9% of in­fec­ti­ons as mo­no­the­ra­py and often as­so­ci­a­ted with more than three an­ti­bi­o­tics (43%). We re­cor­ded 8 se­conda­ry ad­ver­se events, but we never had to in­ter­rupt the tre­at­ment. Of 79 pa­tients with en­ough data, 53% had cli­ni­cal cure/im­pro­ve­ment, 47% cli­ni­cal fai­lu­re and 75% of them died. Mor­ta­li­ty rate in ICU was 45%, 71% du­ring the en­ti­re hos­pi­ta­li­za­ti­on.
Ti­ge­cy­cli­ne was main­ly pre­scri­bed for pneu­mo­nia, in MDR in­fec­ti­ons or as res­cue the­ra­py in se­vere­ly ill pa­tients. Our data sug­gest a good to­le­ra­bi­li­ty and a need of ti­ge­cy­cli­ne for dif­fe­rent in­di­ca­ti­ons than ap­pro­ved by of­fi­ci­al au­tho­ri­ties.

The use of Alpha-​defensin (Sy­no­va­su­re©) in the dia­gno­sis of pe­ri­pros­the­tic joint in­fec­ti­ons
S. van Lan­de­g­hem1, P. van Over­schel­de2, S. Steyaert1
1Cli­ni­cal La­bo­ra­to­ry, AZ Maria Mid­de­la­res, Ghent, 2Hip and Knee cli­nic, AZ Maria Mid­de­la­res, Ghent

Pe­ri­pros­the­tic joint in­fec­ti­on (PJI) is a major com­pli­ca­ti­on after total joint ar­thro­plasty. Ac­cor­ding to the mus­ku­loskel­e­tal in­fec­ti­on so­ci­e­ty (MSIS) cri­te­ria, a com­bi­na­ti­on of cli­ni­cal fin­dings, cul­tu­re and bi­o­mar­kers have to be met to dia­gno­se a PJI. This ap­pro­ach is re­sour­ce and time con­su­ming. The Sy­no­va­su­re (Zim­mer) is a new lateral-​flow test that is based on the de­tec­ti­on of the anti-​microbial pep­ti­de alpha-​defensin in sy­no­vi­al fluid. To eva­lu­a­te the per­for­man­ce in our ge­ne­ral hos­pi­tal a re­tro­spec­ti­ve ana­ly­sis is made du­ring 01/03/2015 - 01/11/2016. Only sam­ples with an alpha-​defensin and a cul­tu­re re­sult are in­clu­ded and com­pa­red to the phy­si­cians’ in­ves­ti­ga­ti­on. Chocolate-​, blood-​ and Mac Con­c­key agar is used for ae­ro­bic and anae­ro­bic in­cu­ba­ti­on du­ring two days, a thi­o­gly­co­la­te broth is in­cu­ba­ted du­ring five days. 43 re­sults were in­clu­ded from 37 pa­tients. All 15 (35%) po­si­ti­ve and 28 (65%) ne­ga­ti­ve tests mat­ched MSIS cri­te­ria to con­firm PJI or asep­tic loo­se­ning. Howe­ver, there were 7 (16%) dis­cre­pan­cies with cul­tu­re. 2 (4.6%) false po­si­ti­ve cul­tu­res were due to a con­ta­mi­na­ti­on and a cyste, not con­nec­ted to the joint. 5 (11.6%) pa­tients had a false ne­ga­ti­ve cul­tu­re due to pre-​operational an­ti­bi­o­tics, low-​grade in­fec­ti­ons or pus in the joint that didn’t re­sult in an or­ga­nism. The con­clu­si­on after mul­ti­dis­ci­pli­na­ry con­sulta­ti­ons is that sy­no­va­su­re had no dis­cre­pan­cies with the phy­si­cians’ cli­ni­cal in­ves­ti­ga­ti­on in con­trast to cul­tu­re. The­re­fo­re sy­no­va­su­re is an in­te­res­ting new bi­o­mar­ker to de­tect PJI and sup­ports the choi­ce bet­ween first and se­cond stage re­vi­si­ons for pros­the­tic ar­thro­plasty.

Non-​invasive de­tec­ti­on of pros­the­tic joint in­fec­ti­ons by mul­ti­plex an­ti­bo­dy de­tec­ti­on: Ex­pe­rien­ces in a ter­ti­a­ry care cen­ter
G. Frans1,* S. Om­be­let1,2*, B. Pee­ters1,3, J. Neyt4, J. Ver­hae­gen1
1De­part­ment of La­bo­ra­to­ry Me­di­ci­ne, Uni­ver­si­ty Hos­pi­tals Leu­ven, Leu­ven, Bel­gi­um, 2De­part­ment of Tro­pi­cal La­bo­ra­to­ry Me­di­ci­ne, In­sti­tu­te of Tro­pi­cal Me­di­ci­ne, Ant­werp, Bel­gi­um, 3De­part­ment of La­bo­ra­to­ry Me­di­ci­ne, Uni­ver­si­ty Hos­pi­tal Ant­wer­pen, Ant­werp, Bel­gi­um, 4De­part­ment of Or­tho­pe­dics, Uni­ver­si­ty Hos­pi­tals Leu­ven, Leu­ven, Bel­gi­um; *both au­thors con­tri­bu­ted equal­ly to this study

Cur­rent dia­gnos­tic al­go­rithms for pros­the­tic joint in­fec­ti­ons (PJI) in­vol­ve ESR or CRP tes­ting, fol­lo­wed by joint as­pi­ra­ti­on if ei­ther is ele­va­ted. In this pro­spec­ti­ve study we eva­lu­a­ted the BJI In­oPlex kit, a mul­ti­plex se­ro­lo­gi­cal im­mu­noas­say do­cu­men­ting PJIs cau­sed by Stap­hy­lo­coc­cus spp, Strep­to­coc­cus aga­lac­ti­ae, and Pro­pi­o­ni­bac­te­ri­um acnes.
Pa­tients who un­der­went re­vi­si­on or re­sec­ti­on ar­thro­plasty for sus­pec­ted PJI bet­ween 18 March 2016 and 18 Au­gust 2017 at the Uni­ver­si­ty Hos­pi­tals Leu­ven were in­clu­ded. There were no ex­clu­si­on cri­te­ria. Serum sam­ples for im­mu­noas­say were taken at the time of sur­gery to­gether with ≥3 in­tra­ope­ra­ti­ve pe­ri­pros­the­tic tis­sue sam­ples for mi­cro­bi­o­lo­gi­cal cul­tu­re. PJI was de­fi­ned by (i) the pre­sen­ce of a sinus tract and/or (ii) growth of a vi­ru­lent or­ga­nism in ≥1 in­tra­ope­ra­ti­ve sam­ple(s) or growthof the same non­vi­ru­lent mi­croo­r­ga­nism in ≥2 in­tra­ope­ra­ti­ve sam­ples. Per­for­man­ce of the BJI In­oPlex assay was eva­lu­a­ted with mi­cro­bi­o­lo­gi­cal cul­tu­re as re­fe­ren­ce.
A total of 56 serum and sur­gery sam­ples from 49 pa­tients (26 male and 23 fe­ma­le) were in­clu­ded with 16 hip, 31 knee, and 2 shoul­der re­pla­ce­ments. PJI was dia­gno­sed in 38/56 sam­ples (67.9%) cor­res­pon­ding with the iden­ti­fi­ca­ti­on of 47 mi­croo­r­ga­nis­ms. In total, 85.1% (40/47) of in­fec­ti­ons in­vol­ved at least one of the spe­cies in­clu­ded in the BJI In­oPlex assay. The sen­si­ti­vi­ty/spe­ci­fi­ci­ty va­lu­es were 70.8%/71.0% for Stap­hy­lo­coc­cus spp (1/56 un­de­ter­mi­ned re­sult), 83.3%/84.0% for Strep­to­coc­cus spp, and 33.3%/88.7% for Pro­pi­o­ni­bac­te­ri­um spp.
Our re­sults sug­gest that the BJI In­oPlex assay could com­ple­ment se­ro­lo­gi­cal and mi­cro­bi­o­lo­gi­cal scree­ning in eva­lu­a­ting pa­tients with sus­pec­ted PJI.

Cost-​effectiveness of a scree­ning pro­gram for chro­nic Q-​fever in The Ne­ther­lands
P.T. de Boer1, M.L. de Lange1, C.C.H. Wiel­ders1, F. Dijkstra1, P.M. Schnee­ber­ger2, W. van der Hoek1
1Na­ti­o­nal In­sti­tu­te for Pu­blic He­alth and the En­vi­ron­ment, Bilt­ho­ven, The Ne­ther­lands, 2Je­roen Bosch Hos­pi­tal, ’s-​Hertogenbosch, The Ne­ther­lands

In the af­ter­math of a large Q-​fever out­break in The Ne­ther­lands, new chro­nic Q-​fever pa­tients are still de­tec­ted. A scree­ning pro­gram may iden­ti­fy cases in an ear­lier stage, pos­si­bly re­sul­ting in a bet­ter prog­no­sis. In this study, we as­ses­sed the cost-​effectiveness of a se­ro­lo­gi­cal scree­ning pro­gram for chro­nic Q-​fever.
A health-​economic de­ci­si­on model was used to esti­ma­te the im­pact of scree­ning on so­cie­tal costs and he­alth ef­fects (me­a­su­red as quality-​adjusted life years [QALYs]). Pre­va­len­ce of chro­nic Q-​fever was esti­ma­ted using Dutch pre­va­len­ce stu­dies of Coxiel­la bur­ne­tii in­fec­ti­on and chro­nic Q-​fever. The ef­fect of scree­ning on cli­ni­cal out­co­mes was based on the na­ti­o­nal Q-​fever da­ta­ba­se. Scree­ning was con­si­de­red cost-​effective, when the in­cre­men­tal cost-​effectiveness ratio was below a con­ven­ti­o­nal thres­hold of € 20,000 per QALY gai­ned.
Scree­ning of pa­tients with car­di­o­vas­cu­lar risk fac­tors li­ving in an area with high Q-​fever in­ci­den­ce was ex­pec­ted to be cost sa­ving. In this sce­na­rio, 215 QALYs would be gai­ned and € 0.1 mil­li­on would be saved. Mo­re­over, scree­ning of pa­tients with car­di­o­vas­cu­lar risk fac­tors li­ving in mo­de­ra­te Q-​fever in­ci­den­ce areas and pa­tients with a com­pro­mi­sed im­mu­ne sy­s­tem li­ving in high Q-​fever in­ci­den­ce areas would be cost-​effective. Howe­ver, scree­ning of pa­tients wit­hout known risk fac­tors would not be cost-​effective. Re­sults were found to be high­ly sen­si­ti­ve to the pre­va­len­ce of chro­nic Q-​fever.
Tar­ge­ted scree­ning pro­grams for chro­nic Q-​fever in areas with mo­de­ra­te to high Q-​fever in­ci­den­ce might be cost-​effective. Howe­ver, there is much un­cer­tain­ty on the cur­rent pre­va­len­ce of chro­nic Q-​fever and the ef­fec­ti­ve­ness of scree­ning on cli­ni­cal out­co­mes.

Tre­a­ting ESBLs with beta-​lactams other than car­ba­pe­nems
J. Mou­t­on
Eras­mus Uni­ver­si­ty Rot­ter­dam

Since car­ba­pe­nems are often the last de­fen­se against ESBL har­bou­ring multi-​drug re­sis­tant micro-​organisms, other so­lu­ti­ons need to be found. In the class of beta-​lactams se­ve­r­al al­ter­na­ti­ve stra­te­gies are avai­la­ble. The first is to de­ter­mi­ne the ac­ti­vi­ty of the ESBL against an array of beta-​lactams. Most ESBLs are so­me­what spe­ci­fic, and re­sis­tan­ce to one beta-​lactam does not ne­ces­sa­ri­ly imply re­sis­tan­ce to others. In con­trast to Amp-C they are not in­du­ced. The se­cond stra­te­gy is to com­bi­ne beta-​lactams with each other, or a re­la­ted com­pound. In ge­ne­ral these can be di­vi­ded in two groups. The first is com­bi­ning a beta-​lactam with a beta-​lactamase in­hi­bi­tor. Examples are an avai­la­ble beta-​lactam such as ce­fe­pi­me com­bi­ned with an exis­ting beta-​lactamase in­hi­bi­tor such as tazo­bac­tam; a new beta-​lactam com­bi­ned with an exis­ting in­hi­bi­tor (e.g. ceftolozane-​tazobactam; imipenem-​relibactam) or an avai­la­ble be­ta­lac­tam com­bi­ned with a new in­hi­bi­tor (e.g. ceftazidime-​avibactam; aztreonam-​avibactam; meropenem-​vaborbactam). The se­cond group con­sists of a com­bi­na­ti­on of two beta-​lactams with dis­tinct dif­fe­ren­ces in af­fi­ni­ty for va­rious PBPs for in­stan­ce PBP2 and PBP3 and the­re­by po­ten­ti­a­ting each other. An example here is ce­fe­pi­me with high af­fi­ni­ty to PBP3 com­bi­ned with zi­de­bac­tam or me­cil­li­nam, both with bin­ding al­most ex­clu­si­ve­ly to PBP2. For many of the cur­rent ESBLs at least one of these stra­te­gies will work and car­ba­pe­nems be spa­red.

Mo­no­the­ra­py vs com­bi­na­ti­on the­ra­py; cli­ni­cal re­sults
A.E. Mul­ler
Haag­lan­den MC, The Hague

With an in­cre­a­sing num­ber of multi-​drug re­sis­tant micro-​organisms, com­bi­na­ti­on the­ra­py is often sug­ge­sted as a so­lu­ti­on for ade­qua­te the­ra­py. More in ge­ne­ral, the use of com­bi­na­ti­on the­ra­py can serve mul­ti­ple pur­po­ses, such as em­pi­ri­cally co­ve­ring po­ten­ti­al pa­tho­gens with a broa­der spec­trum, dif­fe­ren­ces in pe­ne­tra­ti­on of the site of in­fec­ti­on or syn­er­gy bet­ween the two an­ti­bi­o­tics. Some syn­er­gis­tic com­bi­na­ti­ons are com­mer­ci­al­ly de­vel­o­ped in a fixed com­bi­na­ti­on, for example am­oxi­cil­lin/ cla­vula­nic acid and tri­me­thop­rim/sul­fa­me­thoxa­zo­le. In the era of multi-​drug re­sis­tan­ce an­ti­bi­o­tics usu­al­ly ad­mi­nis­te­red as mo­no­the­ra­py are com­bi­ned to achie­ve more ef­fec­ti­ve tre­at­ment com­pa­red to mo­no­the­ra­py. This can be ap­plied to micro-​organisms re­por­ted sus­cep­ti­ble to the spe­ci­fic an­ti­bi­o­tics, but also for those re­por­ted re­sis­tant ac­cor­ding to the cli­ni­cal break­points. Un­for­tu­na­te­ly, alt­hough in vitro stu­dies have in­di­ca­ted that se­ve­r­al com­bi­na­ti­ons of an­ti­bi­o­tics are syn­er­gis­tic, pro­ving this syn­er­gis­tic ef­fect in cli­ni­cal stu­dies has ap­pe­a­red to be much more com­pli­ca­ted. This can be ex­plai­ned by the com­plexi­ty of pa­tients in­vol­ved in these stu­dies. The out­co­me me­a­su­res in these tri­als, such as mor­ta­li­ty or length of hos­pi­tal stay are in­flu­en­ced by many more fac­tors than only an­ti­bi­o­tic the­ra­py. Meta-​analyses have shown a be­ne­fit of com­bi­na­ti­on the­ra­py for pa­tients with sep­tic shock or for tre­at­ment of Pseu­dom­onas ae­rug­i­n­o­sa in­fec­ti­ons. In­ter­pre­ta­ti­on of the re­sults is often com­pli­ca­ted by the dif­fe­ren­ce bet­ween ap­prop­ri­a­te and in­ap­prop­ri­a­te em­pi­ric the­ra­pies. Re­tro­spec­ti­ve ana­ly­ses are con­foun­ded. In re­cent pu­bli­ca­ti­ons, the be­ne­fi­ci­al ef­fect of com­bi­na­ti­on the­ra­py on the mor­ta­li­ty is still under de­ba­te and the ef­fects are often li­mi­ted to high risk pa­tient groups.

Pros­the­tic joint in­fec­ti­ons
J. Geurts
Maas­tricht UMC, Adult Hip Re­con­struc­ti­on In­fec­ti­on Unit

There are a lot of con­tro­ver­sies re­gar­ding the tre­at­ment of PJI (pe­ri­pros­the­tic joint in­fec­ti­on), cer­tain­ly in the field or mi­cro­bi­o­lo­gi­cal dia­gno­sis and pro­to­col. Also, many mi­cro­bi­o­lo­gists are con­fron­ted with ques­ti­ons of their sur­ge­ons on a daily basis. It is the­re­fo­re im­por­tant to have good know­led­ge of the pro­blem we are de­a­ling with and what spe­ci­fic is­sues are ma­king it a chal­len­ge. This pre­sen­ta­ti­on gives a cur­rent state of af­fairs con­cerning or­tho­pa­e­dic im­plant re­la­ted in­fec­ti­ons, from de­fi­ni­ti­on to dia­gno­sis to tre­at­ment.

Vas­cu­lar pros­the­tic graft in­fec­ti­ons
B. Hasse
Di­vi­si­on of In­fec­tious Di­sea­ses and Hos­pi­tal Epi­de­mi­o­lo­gy, Uni­ver­si­ty Hos­pi­tal Zu­rich, Uni­ver­si­ty of Zu­rich, Zu­rich, Swit­zer­land

Aor­tic graft in­fec­ti­ons (AGI) are as­so­ci­a­ted with a high mor­bi­di­ty and mor­ta­li­ty. With the ad­van­ced de­vel­op­ment and in­cre­a­sing use of en­do­vas­cu­lar tech­ni­ques, also el­der­ly pa­tients with co­mor­bi­di­ties re­cei­ve re­con­struc­ti­ve vas­cu­lar sur­gery, whe­re­by these co­mor­bi­di­ties are ad­ding on the risk of postsur­gi­cal in­fec­ti­ons. AGI can pre­sent in se­ve­r­al ways, ei­ther as a local ves­sel ero­si­on, gastroin­tes­ti­nal or bron­chi­al bleeding, bac­terae­mia or fun­ge­mia or with nor­mal phy­si­cal fin­dings. Tre­at­ment ge­ner­al­ly in­vol­ves a redo sur­gery com­bi­ned with an­ti­mi­cro­bi­al the­ra­py. The lat­ter is very much de­pen­dent on the type of mi­croo­r­ga­nism, graft lo­ca­ti­on, in­vol­ved graft ma­te­ri­al and most im­por­tant­ly on the sur­gi­cal stra­te­gy (graft ex­ci­si­on and extra-​anatomic re­con­struc­ti­on; graft ex­ci­si­on with in situ re­con­struc­ti­on; graft pre­ser­va­ti­on and de­bri­de­ment; or no sur­gery). Based on the he­te­ro­ge­nei­ty of all these fac­tors, there are still many un­cer­tain­ties with re­gard to type and length of an­ti­mi­cro­bi­al the­ra­py. The main aim of this ses­si­on is to give an over­view on the topic, and to dis­cuss up-​to-date mi­cro­bi­o­lo­gic dia­gnos­tics in the field of AGI.

Parts of these data have been pre­sen­ted at EC­CMID 2016 on April 9-12, Am­ster­dam.

In­sights into newer an­ti­mi­cro­bi­al agents ac­ti­ve against multi-​drug re­sis­tant gram-​negative bac­te­ria with a spe­ci­al focus on car­ba­pe­n­e­ma­se pro­du­cers
Y. Glup­c­zyns­ski
De­part­ment of Cli­ni­cal Mi­cro­bi­o­lo­gy and Na­ti­o­nal Re­fe­ren­ce Cen­tre for Mo­ni­to­ring An­ti­mi­cro­bi­al re­sis­tan­ce in Gram negative-​bacteria, CHU UCL Namur, Mont-​Godinne, Bel­gi­um

Multi-​drug re­sis­tant (MDR) Gram-​negative bac­te­ri­al in­fec­ti­ons are wi­despread and in­cre­a­sing world­wi­de and they re­pre­sent a cri­ti­cal th­re­at as be­si­des ha­ving the abi­li­ty to es­ca­pe the ef­fect of an­ti­mi­cro­bi­al drugs these or­ga­nis­ms fre­quent­ly lead to to in­crea­sed mor­ta­li­ty and mor­bi­di­ty (pro­lon­ga­ti­on of length of hos­pi­ta­li­za­ti­on stay, in­crea­sed dia­gnos­tic and tre­at­ment costs).
The Cen­ters for Di­sea­se Con­trol and Pre­ven­ti­on has ca­te­go­ri­zed carbapenem-​resistant En­te­ro­bac­te­ria­ceae (CRE) as ‘ur­gent’ an­ti­bi­o­tic th­re­at in the USA where it esti­ma­ted that these or­ga­nis­ms ac­count for more than 2 mil­li­on in­fec­ti­ons, 23,000 de­a­ths, and $2 bil­li­on in ex­cess me­di­cal spen­ding per year. In Fe­bru­a­ry 2017, the World He­alth Or­ga­ni­za­ti­on (WHO) also up­da­ted its list of Pri­o­ri­ty 1, cri­ti­cal or­ga­nis­ms to in­clu­de carbapenem-​resistant and extended-​spectrum beta-​lactamases (ESBLs), carbapenem-​resistant En­te­ro­bac­te­ria­ceae (CRE), carbapenem-​resistant Pseu­dom­onas ae­rug­i­n­o­sa, and carbapenem-​resistant Aci­ne­to­bac­ter bau­man­nii.
In par­ti­cu­lar, carbapenemase-​producing En­te­ro­bac­te­ria­ceae (CPE) pro­du­ce dif­fe­rent clas­ses/fa­mi­lies of plasmid-​mediated car­ba­pe­n­e­ma­ses (KPC, OXA-​48, NDM, VIM,…), and more re­cent­ly XDR CRE bac­te­ria with plasmid-​mediated re­sis­tan­ce to co­lis­tin have been re­por­ted and may pose in­cre­a­sing th­re­ats in the fu­tu­re.
It is well known that the epi­de­mi­o­lo­gy of car­ba­pe­n­e­ma­ses could vary ap­pre­ci­a­bly bet­ween dif­fe­rent con­ti­nents and ge­o­grap­hic areas and that it could also evol­ve rapid­ly lo­cally. The spread of suc­ces­sful pan­de­mic clo­nes (i.e: ST258 KPC-​producing K. pneu­mo­ni­ae) and/or the ho­ri­zon­tal trans­fer of plas­mids and trans­po­sons within and across dif­fe­rent spe­cies (e.g: OXA-​48 and NDM car­ba­pe­n­e­ma­ses) has also led to the oc­cur­ren­ce of major out­breaks in nu­merous coun­tries over the last 5 years. Con­se­quent­ly also the world­wi­de ad­vent and emer­gen­ce of gram-​negative su­per­bugs has th­re­a­ten­ed the ma­na­ge­ment of in­fec­ti­ons cau­sed by these or­ga­nis­ms with a dras­tic de­crea­se in the list of an­ti­bi­o­tics remai­ning ac­ti­ve. In the early 2000s, the phar­ma­ceu­ti­cal in­du­stries so­le­ly fo­cu­sed on the de­vel­op­ment of an­ti­bac­te­ri­al agents against MDR gram-​positive bac­te­ria (eg. methicillin-​resistant S. au­re­us (MRSA) and vancomycin-​resistant En­ter­o­coc­cus) with al­most no sig­ni­fi­cant de­vel­op­ment of drugs against gram-​negative bac­te­ria.
The de­vel­op­ment of new agents with ac­ti­vi­ty against MDR gram-​negative pa­tho­gens has ac­ce­le­ra­ted over the last years and now pro­vi­des (or will soon pro­vi­de) new the­ra­peu­tic op­ti­ons for cli­ni­cians. These new drugs in­clu­de se­ve­r­al beta-​lactam/beta-​lactamase in­hi­bi­tors com­bi­na­ti­ons (cef­t­azi­di­me/avi­bac­tam, cef­to­lo­za­ne/tazo­bac­tam, imi­pe­nem/ci­lia­sta­tin/re­le­bac­tam, mer­o­pe­nem/va­bor­bac­tam and az­tre­onam/avi­bac­tam; other novel drugs ac­ti­ve against MDR gram-​negative in the pi­pe­li­ne and at an ad­van­cd stage of cli­ni­cal de­vel­op­ment and be­lon­ging to other clas­ses of an­ti­mi­cro­bi­als also com­pri­se era­va­cy­cli­ne (te­tra­cy­cli­ne), pla­zo­mi­cin (ami­no­gly­co­si­des) as well as newer cep­ha­lospo­rin sub­stra­tes such as ce­fi­dero­col, a si­der­op­ho­re cep­ha­lospo­rin with a ca­techol side chain).
Among the newer drugs in pi­pe­li­ne, two drugs (na­me­ly, cef­to­lo­za­ne/tazo­bac­tam (Zer­baxa) and cef­t­azi­di­me/avi­bac­tam (Avy­caz, Za­vi­cef­ta)) have been ap­pro­ved by the FDA in 2015 and by the EMEA in 2016. The other drugs are in va­rious pha­ses of tri­als and some of them have re­a­ched or are clo­sed to FDA ap­pro­val in the US. Both cef­t­azi­di­me/avi­bac­tam and cef­to­lo­za­ne/tazo­bac­tam mark the first com­bi­na­ti­on of cep­ha­losporins with beta-​lactamase in­hi­bi­tors. The two ap­pro­ved cep­ha­lospo­rin/beta-​lactamase in­hi­bi­tors com­bi­na­ti­ons are bac­te­ri­ci­dal drugs (time-​dependent kil­ling) that in­hi­bit bac­te­ri­al cell wall syn­the­sis by bin­ding to pen­icillin-binding pro­teins. The major dif­fe­ren­ce bet­ween these two drugs is their spec­trum of bac­te­ri­ci­dal ac­ti­on. Cef­to­lo­za­ne/ta­bo­bac­tam acts on gram-​negative bac­te­ria (ESBL pro­du­cing En­te­ro­bac­te­ria­ceae and P. ae­rug­i­n­o­sa) some anae­ro­bes, and some gram-​positive bac­te­ria but does not act on any car­ba­pe­n­e­ma­ses (i.e not ac­ti­ve against CRE), while cef­t­azi­di­me/avi­bac­tam has ac­ti­on only against gram-​negative bac­te­ria (En­te­ro­bac­te­ria­ceae and P. ae­rug­i­n­o­sa) and is the first cephalo­sporin/beta-​lactam in­hi­bi­tor com­bi­na­ti­on ha­ving ac­ti­on against car­ba­pe­n­e­ma­ses (most­ly class A (e.g. KPC) and some class D (e.g. OXA-​48) but not against any class B metallo-​b-lactamases. Ce­fi­dero­col and cef­t­azi­di­me/avi­bac­tam/az­tre­onam (while wai­ting for the az­tronam/avi­bac­tam com­bi­na­ti­on fo­re­s­een for 2020-​2021) are pro­mi­s­ing op­ti­ons for metallo-​b-lactamases and pos­si­bly also for multidrug-​resistant P. ae­rug­i­n­o­sa, but de­fi­ni­ti­ve data showing cli­ni­cal ef­fi­ca­cy are as yet lac­king. Re­ports of the de­vel­op­ment of re­sis­tan­ce early after the re­lea­se and use of new agents (as has al­rea­dy been ob­ser­ved for cef­t­azi­di­me/avi­bac­tam) is of con­cern. Oral­ly ad­mi­nis­te­red op­ti­ons and agents ef­fec­ti­ve against Aci­ne­to­bac­ter bau­man­nii are un­der­re­pre­sen­ted in cli­ni­cal de­vel­op­ment. Time only will tell whe­ther the ne­west ap­pro­ved an­ti­mi­cro­bi­als as well as the other agents under de­vel­op­ment for Gram-​negative in­fec­ti­ons will help to pre­ser­ve or even en­han­ce our exis­ting ar­ma­men­ta­ri­um.