scholarly journals Viable but Nonculturable(VBNC) State of Pathogenic Bacteria.

1997 ◽  
Vol 12 (4) ◽  
pp. 135-145
Author(s):  
KAZUHIRO KOGURE
Keyword(s):  
Pathogenic Bacteria ◽  
Vbnc State ◽  
Viable But Nonculturable

scholarly journals Induction and Resuscitation of Viable but Nonculturable Corynebacterium diphtheriae

2021 ◽  
Vol 9 (5) ◽  
pp. 927
Author(s):  
Takashi Hamabata ◽  
Mitsutoshi Senoh ◽  
Masaaki Iwaki ◽  
Ayae Nishiyama ◽  
Akihiko Yamamoto ◽  
...  
Keyword(s):  
Low Temperatures ◽  
Diphtheria Toxin ◽  
Pathogenic Bacteria ◽  
Corynebacterium Diphtheriae ◽  
Human Pathogen ◽  
Vbnc State ◽  
Viable But Nonculturable ◽  
Environmental Survival ◽  
Reverse Transcription Quantitative Pcr ◽  
Morphological Differences

Many pathogenic bacteria, including Escherichia coli and Vibrio cholerae, can become viable but nonculturable (VBNC) following exposure to specific stress conditions. Corynebacterium diphtheriae, a known human pathogen causing diphtheria, has not previously been shown to enter the VBNC state. Here, we report that C. diphtheriae can become VBNC when exposed to low temperatures. Morphological differences in culturable and VBNC C. diphtheriae were examined using scanning electron microscopy. Culturable cells presented with a typical rod-shape, whereas VBNC cells showed a distorted shape with an expanded center. Cells could be transitioned from VBNC to culturable following treatment with catalase. This was further evaluated via RNA sequence-based transcriptomic analysis and reverse-transcription quantitative PCR of culturable, VBNC, and resuscitated VBNC cells following catalase treatment. As expected, many genes showed different behavior by resuscitation. The expression of both the diphtheria toxin and the repressor of diphtheria toxin genes remained largely unchanged under all four conditions (culturable, VBNC, VBNC after the addition of catalase, and resuscitated cells). This is the first study to demonstrate that C. diphtheriae can enter a VBNC state and that it can be rescued from this state via the addition of catalase. This study helps to expand our general understanding of VBNC, the pathogenicity of VBNC C. diphtheriae, and its environmental survival strategy.

scholarly journals Survival Strategy of Erwinia amylovora against Copper: Induction of the Viable-but-Nonculturable State

2006 ◽  
Vol 72 (5) ◽  
pp. 3482-3488 ◽  
Author(s):  
M�nica Ordax ◽  
Ester Marco-Noales ◽  
Mar�a M. L�pez ◽  
Elena G. Biosca
Keyword(s):  
Pathogenic Bacteria ◽  
Erwinia Amylovora ◽  
Control Plant ◽  
Viable Cell ◽  
Epifluorescence Microscopy ◽  
Survival Strategy ◽  
Vbnc State ◽  
Tetrazolium Chloride ◽  
Viable But Nonculturable ◽  
Cell Counts

ABSTRACT Copper compounds, widely used to control plant-pathogenic bacteria, have traditionally been employed against fire blight, caused by Erwinia amylovora. However, recent studies have shown that some phytopathogenic bacteria enter into the viable-but-nonculturable (VBNC) state in the presence of copper. To determine whether copper kills E. amylovora or induces the VBNC state, a mineral medium without copper or supplemented with 0.005, 0.01, or 0.05 mM Cu2+ was inoculated with 107 CFU/ml of this bacterium and monitored over 9 months. Total and viable cell counts were determined by epifluorescence microscopy using the LIVE/DEAD kit and by flow cytometry with 5-cyano-2,3-ditolyl tetrazolium chloride and SYTO 13. Culturable cells were counted on King's B nonselective solid medium. Changes in the bacterial morphology in the presence of copper were observed by scanning electron microscopy. E. amylovora entered into the VBNC state at all three copper concentrations assayed, much faster when the copper concentration increased. The addition of different agents which complex copper allowed the resuscitation (restoration of culturability) of copper-induced VBNC cells. Finally, copper-induced VBNC cells were virulent only for the first 5 days, while resuscitated cells always regained their pathogenicity on immature fruits over 9 months. These results have shown, for the first time, the induction of the VBNC state in E. amylovora as a survival strategy against copper.

scholarly journals Recovery in Embryonated Eggs of Viable but Nonculturable Campylobacter jejuni Cells and Maintenance of Ability To Adhere to HeLa Cells after Resuscitation

1999 ◽  
Vol 65 (11) ◽  
pp. 5154-5157 ◽  
Author(s):  
J. M. Cappelier ◽  
J. Minet ◽  
C. Magras ◽  
R. R. Colwell ◽  
M. Federighi
Keyword(s):  
Surface Water ◽  
Campylobacter Jejuni ◽  
Hela Cells ◽  
Pathogenic Bacteria ◽  
Vbnc State ◽  
Adhesion Properties ◽  
Viable But Nonculturable

ABSTRACT The existence of a viable but nonculturable (VBNC) state has been described for Campylobacter jejuni as it had been for a number pathogenic bacteria. Three C. jejuni human isolates were suspended in surface water and subsequently entered the VBNC state. After starvation for 30 days, VBNC cells were inoculated in the yolk sacs of embryonated eggs. Culturable cells were detected in a large proportion of the embryonated eggs inoculated with VBNC C. jejuni cells. Recovered cells kept their adhesion properties.

scholarly journals The Viable But Nonculturable State ofRalstonia solanacearum May Be Involved in Long-Term Survival and Plant Infection

2001 ◽  
Vol 67 (9) ◽  
pp. 3866-3872 ◽  
Author(s):  
Brian E. Grey ◽  
Todd R. Steck
Keyword(s):  
Pathogenic Bacteria ◽  
Physiological Status ◽  
Cupric Sulfate ◽  
Vbnc State ◽  
In Planta ◽  
Term Survival ◽  
Viable But Nonculturable ◽  
Plant Infection ◽  
Nonculturable State ◽  
Vbnc Bacteria

ABSTRACT The role of the dormant-like viable but nonculturable (VBNC) condition in the etiology of bacterial infection was examined using a plant system. The plant-pathogenic bacterium Ralstonia solanacearum was first shown to enter into the VBNC state both in response to cupric sulfate when in a saline solution and when placed in autoclaved soil. To determine if the VBNC condition is related to pathogenesis, the physiological status of bacteria recovered from different regions of inoculated tomato plants was determined at different stages of infection. The fraction of in planta bacteria that were VBNC increased during infection and became greater than 99% by the late stage of disease. The possibility that soil-dwelling VBNC bacteria may resuscitate and infect plants was also examined. When tomato seeds were germinated in sterile soil that contained VBNC but no detectable culturable forms of R. solanacearumcells, resuscitation was observed to occur in soil adjacent to plant roots; these resuscitated bacteria were able to infect plants. This is the first report of R. solanacearum entering the VBNC state and of resuscitation of any VBNC plant-pathogenic bacteria and provides evidence that the VBNC state may be involved in explaining the persistent nature of some infections.

scholarly journals Viable but Nonculturable Bacteria: Food Safety and Public Health Perspective

2013 ◽  
Vol 2013 ◽  
pp. 1-6 ◽  
Author(s):  
Md. Fakruddin ◽  
Khanjada Shahnewaj Bin Mannan ◽  
Stewart Andrews
Keyword(s):  
Public Health ◽  
Food Safety ◽  
Pathogenic Bacteria ◽  
Disease Outbreaks ◽  
Human Pathogens ◽  
Vbnc State ◽  
Public Health Perspective ◽  
Viable But Nonculturable ◽  
Vbnc Bacteria ◽  
Adverse Environmental Conditions

The viable but nonculturable (VBNC) state is a unique survival strategy of many bacteria in the environment in response to adverse environmental conditions. VBNC bacteria cannot be cultured on routine microbiological media, but they remain viable and retain virulence. The VBNC bacteria can be resuscitated when provided with appropriate conditions. A good number of bacteria including many human pathogens have been reported to enter the VBNC state. Though there have been disputes on the existence of VBNC in the past, extensive molecular studies have resolved most of them, and VBNC has been accepted as a distinct survival state. VBNC pathogenic bacteria are considered a threat to public health and food safety due to their nondetectability through conventional food and water testing methods. A number of disease outbreaks have been reported where VBNC bacteria have been implicated as the causative agent. Further molecular and combinatorial research is needed to tackle the threat posed by VBNC bacteria with regard to public health and food safety.

scholarly journals Induction of Viable but Nonculturable Escherichia coli O157:H7 in the Phyllosphere of Lettuce: a Food Safety Risk Factor

2011 ◽  
Vol 77 (23) ◽  
pp. 8295-8302 ◽  
Author(s):  
Laura-Dorina Dinu ◽  
Susan Bach
Keyword(s):  
Escherichia Coli ◽  
Food Safety ◽  
Low Temperature ◽  
Cell Density ◽  
Prolonged Storage ◽  
Potential Hazard ◽  
Vbnc State ◽  
Viable But Nonculturable ◽  
Content Type ◽  
E Coli

ABSTRACTEscherichia coliO157:H7 continues to be an important human pathogen and has been increasingly linked to food-borne illness associated with fresh produce, particularly leafy greens. The aim of this work was to investigate the fate ofE. coliO157:H7 on the phyllosphere of lettuce under low temperature and to evaluate the potential hazard of viable but nonculturable (VBNC) cells induced under such stressful conditions. First, we studied the survival of six bacterial strains following prolonged storage in water at low temperature (4°C) and selected two strains with different nonculturable responses for the construction ofE. coliO157:H7 Tn7gfptransformants in order to quantitatively assess the occurrence of human pathogens on the plant surface. Under a suboptimal growth temperature (16°C), bothE. coliO157:H7 strains maintained culturability on lettuce leaves, but under more stressful conditions (8°C), the bacterial populations evolved toward the VBNC state. The strain-dependent nonculturable response was more evident in the experiments with different inoculum doses (109and 106E. coliO157:H7 bacteria per g of leaf) when strain BRMSID 188 lost culturability after 15 days and strain ATCC 43895 lost culturability within 7 days, regardless of the inoculum dose. However, the number of cells entering the VBNC state in high-cell-density inoculum (approximately 55%) was lower than in low-cell-density inoculum (approximately 70%). We recorded the presence of verotoxin for 3 days in samples that contained a VBNC population of 4 to 5 log10cells but did not detect culturable cells. These findings indicate thatE. coliO157:H7 VBNC cells are induced on lettuce plants, and this may have implications regarding food safety.

scholarly journals Viable-but-NonculturableListeria monocytogenesandSalmonella entericaSerovar Thompson Induced by Chlorine Stress Remain Infectious

mBio ◽  
2018 ◽  
Vol 9 (2) ◽  
pp. e00540-18 ◽  
Author(s):  
Callum J. Highmore ◽  
Jennifer C. Warner ◽  
Steve D. Rothwell ◽  
Sandra A. Wilks ◽  
C. William Keevil
Keyword(s):  
Listeria Monocytogenes ◽  
Caenorhabditis Elegans ◽  
Life Span ◽  
Fresh Produce ◽  
Environmental Stresses ◽  
Vbnc State ◽  
Viable But Nonculturable ◽  
Content Type ◽  
Food Borne Pathogens ◽  
Food Borne

ABSTRACTThe microbiological safety of fresh produce is monitored almost exclusively by culture-based detection methods. However, bacterial food-borne pathogens are known to enter a viable-but-nonculturable (VBNC) state in response to environmental stresses such as chlorine, which is commonly used for fresh produce decontamination. Here, complete VBNC induction of green fluorescent protein-taggedListeria monocytogenesandSalmonella entericaserovar Thompson was achieved by exposure to 12 and 3 ppm chlorine, respectively. The pathogens were subjected to chlorine washing following incubation on spinach leaves. Culture data revealed that total viableL. monocytogenesandSalmonellaThompson populations became VBNC by 50 and 100 ppm chlorine, respectively, while enumeration by direct viable counting found that chlorine caused a <1-log reduction in viability. The pathogenicity of chlorine-induced VBNCL. monocytogenesandSalmonellaThompson was assessed by usingCaenorhabditis elegans. Ingestion of VBNC pathogens byC. elegansresulted in a significant life span reduction (P= 0.0064 andP< 0.0001), and no significant difference between the life span reductions caused by the VBNC and culturableL. monocytogenestreatments was observed.L. monocytogeneswas visualized beyond the nematode intestinal lumen, indicating resuscitation and cell invasion. These data emphasize the risk that VBNC food-borne pathogens could pose to public health should they continue to go undetected.IMPORTANCEMany bacteria are known to enter a viable-but-nonculturable (VBNC) state in response to environmental stresses. VBNC cells cannot be detected by standard laboratory culture techniques, presenting a problem for the food industry, which uses these techniques to detect pathogen contaminants. This study found that chlorine, a sanitizer commonly used for fresh produce, induces a VBNC state in the food-borne pathogensListeria monocytogenesandSalmonella enterica. It was also found that chlorine is ineffective at killing total populations of the pathogens. A life span reduction was observed inCaenorhabditis elegansthat ingested these VBNC pathogens, with VBNCL. monocytogenesas infectious as its culturable counterpart. These data show that VBNC food-borne pathogens can both be generated and avoid detection by industrial practices while potentially retaining the ability to cause disease.

scholarly journals mRNA Detection by Reverse Transcription-PCR for Monitoring Viability over Time in an Enterococcus faecalis Viable but Nonculturable Population Maintained in a Laboratory Microcosm

2000 ◽  
Vol 66 (10) ◽  
pp. 4564-4567 ◽  
Author(s):  
Maria del Mar Lleò ◽  
Sabrina Pierobon ◽  
Maria Carla Tafi ◽  
Caterina Signoretto ◽  
Pietro Canepari
Keyword(s):  
Enterococcus Faecalis ◽  
Reverse Transcription ◽  
Microbiological Quality ◽  
Vbnc State ◽  
Viable But Nonculturable ◽  
Reverse Transcription Pcr ◽  
Pathogenic Factors ◽  
Laboratory Microcosm ◽  
Or Genes

ABSTRACT The viable but nonculturable (VBNC) state is a survival strategy adopted by bacteria when they are exposed to hostile environmental conditions. It has been shown that VBNC forms of bacteria are no longer capable of growing on conventional bacteriological media but conserve pathogenic factors and/or genes. It is thus necessary to develop methods capable of detecting nonculturable bacteria and of establishing their viability when the microbiological quality of environments is monitored. In this study we demonstrated that a gene was expressed during the VBNC state in a low-nutrient-concentration microcosm through detection of Enterococcus faecalis pbp5 mRNA by reverse transcription-PCR over a 3-month period. The presence of mRNA correlated with metabolic activity and resuscitation capability, indicating the viability of the VBNC cells.

scholarly journals A Proteomic Signature of Dormancy in the Actinobacterium Micrococcus luteus

2017 ◽  
Vol 199 (14) ◽  
Author(s):  
Sujina Mali ◽  
Morgan Mitchell ◽  
Spencer Havis ◽  
Abiodun Bodunrin ◽  
Jonathan Rangel ◽  
...  
Keyword(s):  
Bacterial Infections ◽  
High Performance ◽  
Micrococcus Luteus ◽  
Treponema Pallidum ◽  
Gc Content ◽  
Antibiotic Tolerance ◽  
Vbnc State ◽  
Viable But Nonculturable ◽  
Content Type ◽  
Regulatory Processes

ABSTRACT Dormancy is a protective state in which diverse bacteria, including Mycobacterium tuberculosis, Staphylococcus aureus, Treponema pallidum (syphilis), and Borrelia burgdorferi (Lyme disease), curtail metabolic activity to survive external stresses, including antibiotics. Evidence suggests dormancy consists of a continuum of interrelated states, including viable but nonculturable (VBNC) and persistence states. VBNC and persistence contribute to antibiotic tolerance, reemergence from latent infections, and even quorum sensing and biofilm formation. Previous studies indicate that the protein mechanisms regulating persistence and VBNC states are not well understood. We have queried the VBNC state of Micrococcus luteus NCTC 2665 (MI-2665) by quantitative proteomics combining gel electrophoresis, high-performance liquid chromatography, and tandem mass spectrometry to elucidate some of these mechanisms. MI-2665 is a nonpathogenic actinobacterium containing a small (2.5-Mb), high-GC-content genome which exhibits a well-defined VBNC state induced by nutrient deprivation. The MI-2665 VBNC state demonstrated a loss of protein diversity accompanied by increased levels of 18 proteins that are conserved across actinobacteria, 14 of which have not been previously identified in VNBC. These proteins implicate an anaplerotic strategy in the transition to VBNC, including changes in the glyoxylate shunt, redox and amino acid metabolism, and ribosomal regulatory processes. Our data suggest that MI-2665 is a viable model for dissecting the protein mechanisms underlying the VBNC stress response and provide the first protein-level signature of this state. We expect that this protein signature will enable future studies deciphering the protein mechanisms of dormancy and identify novel therapeutic strategies effective against antibiotic-tolerant bacterial infections. IMPORTANCE Dormancy is a protective state enabling bacteria to survive antibiotics, starvation, and the immune system. Dormancy is comprised of different states, including persistent and viable but nonculturable (VBNC) states that contribute to the spread of bacterial infections. Therefore, it is imperative to identify how bacteria utilize these different dormancy states to survive antibiotic treatment. The objective of our research is to eliminate dormancy as a route to antibiotic tolerance by understanding the proteins that control dormancy in Micrococcus luteus NCTC 2665. This bacterium has unique advantages for studying dormancy, including a small genome and a well-defined and reproducible VBNC state. Our experiments implicate four previously identified and 14 novel proteins upregulated in VBNC that may regulate this critical survival mechanism.

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