Comparison of different human tissue processing methods for maximization of bacterial recovery

Askar, Mohamed; Ashraf, Waheed; Scammell, Brigitte E.; Bayston, Roger

doi:10.1007/s10096-018-3406-4

Cited by 12 publications

(22 citation statements)

References 20 publications

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“…Tissues were homogenized using the Roche magNA Lyser homogenizer (Hoffman-La Roche Ltd, Basel Switzerland) at a speed of 4500 rpm for four cycles, each lasting 45 s [18]. Prostheses were sonicated in a precision sonicator set at 50 Hz (Ultrawave Ltd, Cardiff, UK) for 5 min.…”

Section: Application Of Optimised Pma In Clinical Samplesmentioning

confidence: 99%

Propidium monoazide–polymerase chain reaction for detection of residual periprosthetic joint infection in two-stage revision

et al. 2019

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False negative culture results in periprosthetic joint infection (PJI) are not uncommon particularly when patients have received long term antibiotics. Polymerase chain reaction (PCR) has a lower specificity partly due to detection of residual DNA from dead bacteria. Propidium monoazide (PMA) prevents DNA from dead bacteria from being amplified during the PCR. This study aimed to determine the role of PMA in PCR for diagnosis of PJI. Clinical samples were tested by PCR with and without prior treatment with PMA and compared to conventional microbiological culture. The PCR assay included genus-specific primers for staphylococci and enterococci and species-specific primers for Cutibacterium acnes. The validated conditions of PMA treatment used in this study were 20 μM concentration and 5 and 10 min of dark incubation and photo-activation respectively. 202 periprosthetic tissues and explanted prostheses from 60 episodes in 58 patients undergoing revision arthroplasties for either PJI or non-infective causes were tested, by culture, PCR, and PMA-PCR. 14 of the 60 episodes satisfied the Musculoskeletal Infection Society (MSIS) criteria for PJI and 46 did not. Sensitivity of culture, PCR, and PMA-PCR were 50%, 71%, and 79% respectively. Specificities were 98%, 72%, and 89% respectively. All figures were calculated for episodes rather than samples. PMA-PCR enhanced both the specificity and the sensitivity of PCR. It has the potential to detect residual bacterial viability prior to reimplantation in the two-stage revision for PJI.

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Section: Application Of Optimised Pma In Clinical Samplesmentioning

confidence: 99%

Propidium monoazide–polymerase chain reaction for detection of residual periprosthetic joint infection in two-stage revision

et al. 2019

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“…There are still some issues about optimal pretreatment methods on tissues, because insufficient pretreatment might result in low bacteria yield released from tissues, while excessive pretreatment might reduce bacterial viability. Mohamed Askar reported that the use of mechanized steel ball grinding and homogenization might reduce the recovery of bacteria from tissue samples [ 6 ]. However, this study showed there were no significant effects on the viability of bacteria pretreated by various methods.…”

Section: Discussionmentioning

confidence: 99%

“…The strain of Staphylococcus aureus ( S. aureus ) ATCC 25923 (American Type Culture Collection, USA) the strain of Escherichia coli ( E. coli ) ATCC 25922 (American Type Culture Collection, USA), and the strain of Candida albicans ( C. albicans )ATCC 90029 (American Type Culture Collection, USA) were used to establish infection model in vitro. In order to simulate PJI infection as much as possible, in which pathogen may be distributed on a single surface or multiple surfaces even inside tissues [ 6 ], therefore, three tissues infection models were established (Fig. 1 ): ① Single surface infection: pipette aspirate 10 μL (200 CFU) bacterial dilution ( S. aureus /E.…”

Section: Methodsmentioning

confidence: 99%

“…However, MM is a tedious process in which contamination can easily be introduced. Studies have shown that mechanical homogenization (MH), dithiothreitol (DTT), and sonificated (SF) can increase the release of bacteria from tissue [ 6 – 8 ]. However, wether these methods are clinical available are still in the air, because currently there are no reports that directly compared these different specimen pretreatment methods in vitro.…”

Section: Introductionmentioning

confidence: 99%

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Microbial yield from infectious tissues pretreated by various methods: an invitro study

Cai

Fang

Zhang

et al. 2021

BMC Musculoskelet Disord

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Background This study aimed to evaluate the effects of different pretreatment methods on the microbial yield from infectious tissues. Methods Strains of Staphylococcus aureus (SA), Escherichia coli (EC) and Candida albicans (CA) were used to construct single-surface, full-surface, and internal infection models in sterile pork tissue. Manual milling (MM), mechanical homogenization (MH), sonificated (SF), dithiothreitol (DTT), and direct culture (DC) were used to pretreat these tissues, the microbial yield from different pretreatment methods were recorded and compared. Moreover, periprosthetic tissues collected intraoperatively from periprosthetic joint infection (PJI) patients were used as a verification. Results The study showed that the microbial yield from MH pretreatment was significantly higher than that of MM (P < 0.01) and SF pretreatment method (P < 0.01). Furthermore, in the internal infection model, the microbial yield from MH group was also significantly higher than that of SF (P < 0.01), DTT (P < 0.01), and DC group (P < 0.01). Moreover, the number of bacterial colonies obtained from periprosthetic tissues pretreated by MH was significantly higher than pretreated by other pretreatment methods (P = 0.004). Conclusions The effects of MH and DTT in microbial yield were significantly higher than that of DC, SF and MM, and these methods can be used to process multiple tissue samples at the same time, which might further improve the diagnostic sensitivity of infectious disease.

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“…A contradiction exists in using pretreatment methods, because insu cient pretreatment can result in not enough bacteria being released, and excessive pretreatment may reduce bacterial viability. Mohamed Askar reported that the use of mechanized steel ball grinding and homogenization may reduce the recovery of bacteria from tissue samples 6 . The results of this study showed that the various pretreatment methods had no signi cant effect on the viability of various bacteria.…”

Section: Discussionmentioning

confidence: 99%

Effects of different pretreatment methods on microbial recovery of infected tissues

Fang

Cai

Zhang

et al. 2020

Preprint

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This study aimed to evaluate the effects of different pretreatment methods on the degree of microbial recovery in infected tissues. Standard strains of Staphylococcus aureus (SA), Escherichia coli (EC) and Candida albicans (CA) were used to construct single-surface, full-surface, and internal infection models in sterile pork tissue. Manual milling (MM), mechanical homogenization (MH), ultrasonic lysis (UL), dithiothreitol (DTT), and direct culture (DC) were used to pretreat infection tissues, the ability of the different pretreatment methods to achieve pathogen recovery in the different bacterial infection models was compared. At the same time, periprosthetic tissues collected from periprosthetic joint infection (PJI) were pretreated with the same methods. We showed that regardless of whether the single-surface or full-surface infection model was used in SA, EC, and CA infection, the microbial acquisition in the MH group was significantly higher than that in the MM (P <0.01) and UL groups (P <0.01). In the internal infection model, the microbial acquisition of the MH group was significantly higher than that of the MM (P <0.01), UL (P <0.01), DTT (P <0.01), and DC groups (P <0.01). In the PJI cases, the number of bacterial colonies obtained by MH was significantly higher than that obtained by other pretreatment methods (P = 0.004). The effects of MH and DTT in microbial recovery were significantly better than that of DC, UL and MM, and these methods can be used to process multiple tissue samples at the same time, which can further improve the efficiency of clinical microbial diagnosis.

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Comparison of different human tissue processing methods for maximization of bacterial recovery

Cited by 12 publications

References 20 publications

Propidium monoazide–polymerase chain reaction for detection of residual periprosthetic joint infection in two-stage revision

Propidium monoazide–polymerase chain reaction for detection of residual periprosthetic joint infection in two-stage revision

Microbial yield from infectious tissues pretreated by various methods: an invitro study

Effects of different pretreatment methods on microbial recovery of infected tissues

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