1

Monascus ruber: invasive gastric infection by 1

dried and salted fish consumption 2

XAVIER IRIART1,2*

, ANGELA FIOR3, DENIS BLANCHET

2, ANTOINE BERRY

1, 3

PAUL NERON4 AND CHRISTINE AZNAR

2 4

Service de Parasitologie Mycologie, Centre Hospitalier Universitaire, Hôpital Rangueil, 5

31059 Toulouse Cedex 9, France1; Laboratoire Hospitalier et Universitaire de Parasitologie 6

Mycologie, Centre Hospitalier Andrée Rosemon /UFR de Médecine – EA 3593- Université 7

des Antilles et de la Guyane, Cayenne 97306, Guyane française2; Service d’Anatomie et 8

Cytologie Pathologique3 and Service de Médecine B

4– Centre Hospitalier Andrée Rosemon, 9

Cayenne 97306, Guyane française 10

11

12

13

14

15

16

17

18

*Corresponding author. Mailing address: Service de Parasitologie-Mycologie, Centre 19

Hospitalier Universitaire de Toulouse, Hôpital Rangueil, TSA 50032, 31059 Toulouse Cedex 20

9, France. Phone: 33 5 61 32 28 92. Fax: 33 5 61 32 20 96. E-mail: xabierlete@yahoo.fr 21

22

23

Copyright © 2010, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.J. Clin. Microbiol. doi:10.1128/JCM.01000-10 JCM Accepts, published online ahead of print on 4 August 2010

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

2

We report a case of invasive gastric infection caused by Monascus ruber observed in a patient 24

from French Guiana with gastric adenocarcinoma. The originality of this case is because, 25

firstly, this invasive mycosis is extremely rare and secondly, the probable mode of infection 26

was by the consumption of Monascus ruber-contaminated food. 27

28

_____________________________________ 29

30

31

32

CASE REPORT 33

In July 2003, a 66-year-old man was hospitalized in Cayenne hospital, French Guiana, 34

with a history of chronic cough for seven months, dyspnea, asthenia, anorexia, gastro-35

esophageal reflux, constipation, intermittent fever and calf pain. He lived in the vicinity of 36

Grand Santi, a village located on the French side of the Maroni River, in the East of French 37

Guiana. Except for a high tobacco consumption for 45 years, he had no notable medical 38

antecedents. At the time of admission, the results of pulmonary, cardiovascular and abdominal 39

clinical examinations were normal. The patient was in good general state without fever. 40

Abnormal laboratory findings included normochromic normocytic anemia (hemoglobin 41

9.8g/dL) and an increase in C-reactive protein (97mg/mL with N<5mg/mL). Other biologic 42

data were within the normal range. 43

An abdomino-pelvic ultrasound revealed celiac and mesenteric lymphadenopathy, 44

without other anomalies. Abdominal tomography showed considerable gastric distension with 45

stasis, associated with antral tissue hypertrophy. Given these results, endoscopy was carried 46

out. Gastric and duodenal biopsies were performed and duodenal liquid was aspirated for 47

analysis. Duodenal biopsies showed interstitial duodenitis and histological analysis of the 48

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

3

gastric biopsies revealed moderately differentiated gastric adenocarcinoma associated with 49

chronic gastritis, extensive metaplasia and massive bacterial superinfection, including 50

Helicobacter pylori. 51

Septated, wide, and banded fungal filaments, with right angle branching were 52

observed in the gastric biopsies with Gomori-Grocott and periodic acid-schiff (PAS) stains. 53

The pathologist has first concluded that there was a fungus from the Mucoracea family. 54

Fungal cultures were carried out with fragments of the gastric biopsies and duodenal 55

liquid on gentamicin-chloramphenicol Sabouraud medium with or without cycloheximide at 56

30°C and 37°C. Two fungi were isolated from the biopsies at 30°C and 37°C: a Mucoracea 57

without sporulation on the two fungal media and a white to purple fungus with a dull reddish 58

pigment on the reverse side of the culture plate on the Sabouraud medium without 59

cycloheximide. The growth of the second fungus was rapid and the colonies were thinly 60

floccose, spreading from the middle (Fig A). The microscopic examination of the culture were 61

positive for Monascus ruber (anamorph: Basipetospora rubra), a fungus with distinctive 62

characteristics as described below: chains of round and colorless conidia (9-10.5µm/7-9µm) 63

with flattened bases (Fig B1), and young round thin-walled ascoscarps containing oval 64

ascospores with smooth walls (5-6µm/4-5µm) (Fig B2). The Mucoracea and M. ruber were 65

also isolated from the duodenal aspiration liquid, associated with Candida glabrata. 66

The histological slides of the gastric biopsies were then carefully re-examined and the 67

morphological characteristics of 2 types of fungi were recognized confirming the presence of 68

M. ruber in the biopsies (Fig C1 and C2). 69

Treatment with amphotericin B, 50mg per day was initiated. Three days later, because 70

of nephrotoxicity (the creatinine increased from 88 to 150µmol/L), amphotericin B was 71

switched to the liposomal form, 200mg per day. Five days later, the dose was decreased 72

(200mg one every other day) because of an increase in kidney failure. 73

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

4

The origin of this atypical infection was researched and an alimentary cause was 74

suspected. The patient was a high consumer of fish, principally dried and salted. Fungal 75

cultures were carried out on the remains of salted fish (Serrasalmus rhombeus called pëne) 76

found in his room at the hospital and consumed by the patient during his hospitalization. The 77

skin, the flesh and the salt were cultured separately at 30°C and 37°C on the previously-used 78

media. The salt and flesh cultures were positive for Monascus ruber, only on Sabouraud 79

medium without cycloheximide at 30°C and 37°C, but the skin was negative. 80

The surgical treatment of the gastric adenocarcinoma was not possible because of a 81

deterioration of the patient's general condition and extensive venal thrombosis of the inferior 82

left member. 83

A second control gastric biopsy was taken 45 days after the first one (after 5 weeks of 84

treatment). The direct examination showed the same fungal elements and Monascus ruber 85

was isolated alone from culture at 30°C and 37°C. After this, the patient was released and 86

returned back home, where he died shortly after. 87

88

_____________________________________ 89

90

Monascus ruber (anamorph: Basipetospora rubra) is a filamentous fungus (family 91

Monascaceae, order Eurotiales) (16). This Ascomycete has rarely been implicated in human 92

infection (13). We report here an uncommon case of Monascus ruber invasive gastric 93

infection associated with the consumption of contaminated dried and salted fish. 94

In Asia, this fungus is traditionally used to produce Monascus-fermented rice and is 95

commonly employed as food colorant, flavoring agent or additive for preserving fish and 96

meat (14). In industry, Monascus species are important sources of pigments or bioactive 97

compounds (8), like monacolin K which is a very effective hypocholesterolemic agent (4). 98

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

5

In humans, Monascus can be pathogenic in several ways: i) allergy and anaphylaxis to 99

red yeast rice have been described (6), due to an immediate sensitivity to Monascus 100

purpureus. ii) toxicity: some Monascus compounds are toxic, such as citrinin which is a 101

nephro-hepatotoxic agent (7). Citrinin permeates into the mitochondria where it alters Ca2+ 102

homeostasis (2) and interferes with the electron transport system (12). Moreover, a toxic drug 103

interaction was also reported between cyclosporine and monacolin K of the red yeast rice 104

(Monascus purpureus) that led to a rhabdomyolosis in a renal transplant recipient (11). iii) 105

infection: one study described a direct renal infection after surgery due to Monascus ruber in 106

a 70 year old patient with multiple kidney stones (13). Our report is the second description of 107

Monascus ruber infection world-wide with tissue invasion and the first case where an 108

alimentary contamination was highlighted. The presence of the fungus in the histological 109

tissue of the gastric biopsies demonstrated the invasive capacity of this mycosis. 110

Monascus ruber is a salt and acid tolerant fungus (10). At 35°C (approximate gastric 111

temperature), the lower pH limit permissive of Monascus growth is 3.7 but the development 112

of the fungus is further promoted when the pH rises to neutral (10). In a healthy stomach, the 113

very low pH of 1-2 should not allow the proliferation of the fungus. Functional alterations of 114

the stomach (dyspepsia), associated with an increase in the gastric pH, may be one of the 115

factors favoring this infection. As Monascus ruber is NaCl tolerant, it was not surprising to 116

find Monascus ruber in the dried and salted fish. Monascus had already been isolated from 117

dried and salted fish in Sri Lanka (1), Indonesia (15) and Nigeria (3) or from green table 118

olives (9) but food contamination in man has never been observed. 119

After the treatment with amphotericin B and the liposomal form, nephrotoxicity 120

developed whereas the patient's renal function was normal at admission. It is possible that this 121

toxicity was the consequence of not only amphotericin B, but also of the citrinin, produced 122

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

6

during the lyses of the fungus (5, 7). Thus, it would be preferable to use an antifungal drug 123

without nephrotoxicity to avoid the possible potentiation of renal failure by citrinin. 124

125

126

ACKNOWLEDGMENTS 127

We gratefully acknowledge John Woodley and Lilia khodjet el khil for the English revision of 128

the manuscript and Michel Joubert from the “département des centres de santé de Guyane” for 129

his help in gathering information about the patient. 130

This study was supported by the « Laboratoire Hospitalier et Universitaire de Parasitologie-131

Mycologie » Cayenne Hospital. 132

Potential conflicts of interest. All authors: no conflicts. 133

134

135

136

REFERENCES 137

1. Atapattu, R., and U. Samarajeewa. 1990. Fungi associated with dried fish in Sri 138

Lanka. Mycopathologia 111:55-9. 139

2. Chagas, G. M., M. A. Oliveira, A. P. Campello, and M. L. Kluppel. 1995. 140

Mechanism of citrinin-induced dysfunction of mitochondria. IV--Effect on Ca2+ transport. 141

Cell Biochem Funct 13:53-9. 142

3. Diyaolu, S. A., and L. O. Adebajo. 1994. Effects of sodium chloride and relative 143

humidity on growth and sporulation of moulds isolated from cured fish. Nahrung 38:311-7. 144

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

7

4. Endo, A. 1979. Monacolin K, a new hypocholesterolemic agent produced by a 145

Monascus species. J Antibiot (Tokyo) 32:852-4. 146

5. Flajs, D., and M. Peraica. 2009. Toxicological properties of citrinin. Arh Hig Rada 147

Toksikol 60:457-64. 148

6. Hipler, U. C., W. Wigger-Alberti, A. Bauer, and P. Elsner. 2002. Case report. 149

Monascus purpureus--a new fungus of allergologic relevance. Mycoses 45:58-60. 150

7. Krejci, M. E., N. S. Bretz, and D. A. Koechel. 1996. Citrinin produces acute adverse 151

changes in renal function and ultrastructure in pentobarbital-anesthetized dogs without 152

concomitant reductions in [potassium]plasma. Toxicology 106:167-77. 153

8. Lin, Y. L., T. H. Wang, M. H. Lee, and N. W. Su. 2008. Biologically active 154

components and nutraceuticals in the Monascus-fermented rice: a review. Appl Microbiol 155

Biotechnol 77:965-73. 156

9. Panagou, E. Z., C. Z. Katsaboxakis, and G. J. Nychas. 2002. Heat resistance of 157

Monascus ruber ascospores isolated from thermally processed green olives of the 158

Conservolea variety. Int J Food Microbiol 76:11-8. 159

10. Panagou, E. Z., P. N. Skandamis, and G. J. Nychas. 2005. Use of gradient plates to 160

study combined effects of temperature, pH, and NaCl concentration on growth of Monascus 161

ruber van Tieghem, an Ascomycetes fungus isolated from green table olives. Appl Environ 162

Microbiol 71:392-9. 163

11. Prasad, G. V., T. Wong, G. Meliton, and S. Bhaloo. 2002. Rhabdomyolysis due to 164

red yeast rice (Monascus purpureus) in a renal transplant recipient. Transplantation 74:1200-165

1. 166

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

8

12. Ribeiro, S. M., G. M. Chagas, A. P. Campello, and M. L. Kluppel. 1997. 167

Mechanism of citrinin-induced dysfunction of mitochondria. V. Effect on the homeostasis of 168

the reactive oxygen species. Cell Biochem Funct 15:203-9. 169

13. Sigler, L., P. C. Kibsey, D. A. Sutton, S. P. Abbott, E. Zilkie, D. I. McCarthy, and 170

A. Fothergill. 1999. Monascus ruber, causing renal infection, abstr. Pp. 297. Abstr. 99th 171

Meet. Am. Soc. Microbiol. American Society for Microbiology, Chicago, DC. 172

14. Wang, T. H., and T. F. Lin. 2007. Monascus rice products. Adv Food Nutr Res 173

53:123-59. 174

15. Wheeler, K. A., and A. D. Hocking. 1993. Interactions among xerophilic fungi 175

associated with dried salted fish. J Appl Bacteriol 74:164-9. 176

16. Young, E. M. 1930. Physiological studies in relation to the taxonomy of Monascus 177

spp., p 227-244. In Juday C (ed), Transactions of the Wisconsin Academy of Sciences, Arts 178

and Letters. Wisconsin Academy of Sciences, Madison, WI. 179

180

181

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

9

Figure legend 182

183

Fig: Monascus rubber. (A) Macroscopic aspect with red pigment after growth on Sabouraud’s 184

agar medium for seven days at 37°C. Microscopic characteristics of the isolate of M. ruber 185

with two types of reproduction: (B1) asexual form with a chain of conidia (magnification, 186

x¤200) and (B2) sexual form with thin-walled ascoscarps containing ovals ascospores 187

(magnification, x¤400). Histological microscopic examination of a gastric biopsy 188

(magnification, x¤400): (C1) Gomori Grocott stained area of tumor necrosis with truncated 189

fungal filaments (red arrows) and (C2) PAS stained with banded fungal filaments branched at 190

right angles characteristic of a Mucoracea fungus (yellow arrows) and wide vesiculated 191

septate filaments (blue arrows) characteristic of Monascus. 192

193

194

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from

Figure

A B1 B2

C1 C2

on Septem

ber 15, 2018 by guesthttp://jcm

.asm.org/

Dow

nloaded from