Multi-drug resistant bacteria predict mortality in bloodstream infection in a tertiary setting in Tanzania.

BACKGROUND: Bloodstream infections (BSI) are serious and life-threatening, associated with high mortality and morbidity. In resource-limited settings, there is a paucity of data on predictors of outcome in patients with BSI. This study aimed at examining the predictors of mortality in patients with BSI as well as bacteria causing BSI. METHODS AND MATERIALS: This was a cross-sectional study conducted at Muhimbili National Hospital between April and May 2018. Blood culture results from all inpatients at the clinical microbiology laboratory were recorded and clinical information was retrieved retrospectively from the files. Bacteria from positive blood culture were identified and antimicrobial susceptibility was performed.
RESULTS: The overall prevalence of BSI was, 46/402 (11.4% 95% CI 8.6-15), with a case fatality rate of 37%. There was a significantly high rate of BSI in patients who had died (19.5%) compared to those who survived (9.2%) p = 0.008. Gram-negative bacteria (74%) were the common cause of BSI, with a predominance of Enterobacteriaceae (22), followed by Pseudomonas aeruginosa (11). The majority of bacteria (70.5%) isolated from patients with BSI were Multi-drug resistant (MDR). Forty-six percent of Pseudomonas aeruginosa were resistant to meropenem while 68% (15/22) of Enterobacteriaceae were extended-spectrum β lactamase producers. Carbapenemase production was detected in 27% (3/11) of Pseudomonas aeruginosa and one Proteus mirabilis. Forty percent of Staphylococcus aureus were methicillin-resistant Staphylococcus aureus. Positive blood culture (aOR 2.24, 95%CI 1.12-4.47, p 0.02) and admission to the intensive care unit (aOR 3.88, 95%CI 1.60-9.41, p = 0.003) were independent factors for mortality in suspected BSI. Isolation of MDR bacteria was an independent predictor for mortality in confirmed BSI (aOR 15.62, 95%CI 1.24-161.38, p = 0.02).
CONCLUSION: The prevalence of BSI was 11.4%, with the majority of bacteria in BSI were MDR. Positive blood culture, admission to the ICU and MDR were predictors for mortality.

Introduction

Bloodstream infection (BSI) is life-threatening not only associated with increased mortality, and morbidity but also health care costs [1]. Often, MDR bacteria causing BSI are associated with poor patient outcome compared to susceptible bacteria [2, 3]. The incidence and prevalence of BSI vary considerably between developed and developing countries [4-6]. Nevertheless, the epidemiology of BSI in both community and hospital settings is evolving. Besides, data on the rapid changing bacterial etiology of BSI in Tanzania is scarce.

Treatment of BSI in the resource-limited setting is largely empirical using broad-spectrum antibiotics. Empiric treatments often fail to target the correct pathogens, leading to treatment failures and increasing mortality [1, 7]. To address these, clinical microbiology laboratories may play important roles in the effective management of BSI. Prompt reporting of results coupled with identifying critical values and antibiogram pattern provided by laboratories facilitate the successful management of patients with BSI. Elsewhere, factors predicting mortality in BSI have been investigated [2, 8] but few data exist in Tanzania.

The increasing burden of hospital-acquired BSI caused by Multidrug-resistant (MDR) pathogens including extended-spectrum beta-lactamases (ESBL) have been previously observed in Tanzania [3]. However, in a world of rapidly evolving bacteria pathogens, repeated surveillance is warranted for improving the management of BSI. Our study aimed at identifying the current bacterial etiology and predictors of mortality in BSI.

Materials and methods

Study design and setting

This was a cross-sectional study conducted at Muhimbili National Hospital (MNH) between April and May 2018. MNH is the largest tertiary hospital in Tanzania, serving approximately 6 million people from Dar es Salaam. It has a 1500 bed capacity, attending approximately 1200 inpatients per week and approximately 1200 outpatients per day. MNH is also a training facility for the Muhimbili University of Health and Allied Sciences and the main referral hospital in Tanzania.

Study population

The study included all inpatients with clinical features suggestive of BSI, from whom blood specimens for culture were processed at MNH Clinical Microbiology Laboratory.

Data collection

A structured data collection tool was used to record results of blood culture, colonial morphology, Gram stain, isolates identity and antimicrobial susceptibility test (AST). Demographic characteristics such as sex, age, and other patients’ information were extracted from patient request forms. Patients’ clinical outcomes were retrieved from the patient's clinical case notes.

Blood culture and bacterial identification

Blood was collected by the attending clinician into blood culture bottles for adult (BD BACTEC Plus Aerobic /F Culture Vials, Becton Dickinson and Company) and pediatric (BD BACTEC Peds PlusTM/F Culture Vials, Becton Dickinson and Company) and used for culture. Upon reaching the laboratory, blood culture bottles were inspected for acceptance criteria. Blood culture vials were incubated into the BD BACTEC FX40 analyzer for a maximum of five days.

Primary Gram stain was performed on positive cultures followed by subculture on appropriate solid culture media. A single drop of blood was inoculated into 5% sheep blood agar (SBA) and MacConkey agar (MCA), then incubated at 37°C with 5–10% CO2 and 37°C respectively for 18–24 hours. Bacteria were initially identified by colony morphology and Gram stain. Gram-positive cocci were further identified by a set of biochemical tests including, catalase test, coagulase, DNase, Staphaurex (Remel Europe Ltd, Dartford, UK), Streptococcus grouping kit (Remel Europe Ltd, Dartford, UK). Gram-negative rods were further identified by API20 E and API20 NE (Biomerieux, France).

Antimicrobial susceptibility testing

Kirby Bauer disc diffusion method was used to test antimicrobial susceptibility following Clinical and Laboratory Institute guidelines [CLSI] [9]. Antibiotics included in susceptibility testing were those commonly used in our settings for the management of BSI and few reserved for severe bacterial infections.

MDR was defined as resistance to at least one antibiotic in three or more antimicrobial classes [10]. Methicillin Resistant Staphylococcus aureus (MRSA) was determined by the disc diffusion method using cefoxitin disk (30μg) (Oxoid, UK) as previously described [9]. ESBL production in Enterobacteriaceae was detected as described previously [9]. Carbapenemase production in Gram-negative bacteria was screened using a combination disk method whereby meropenem disk (10μg) alone and a combination of meropenem disk (10μg) with dipicolinic acid (DPA) 1000μg. An increase of zone diameter for 5 mm or more around combined meropenem disk and DPA, as compared with the disk of meropenem alone, was considered to be a positive result [11].

The following reference strains were used for quality control: Escherichia coli (E. coli) ATCC 25922, Klebsiella pneumoniae ATCC 700603 for ESBL, Staphylococcus aureus ATCC 25923, and Staphylococcus aureus ATCC 29213 for MRSA and Klebsiella pneumoniae ATCC 1705 and Klebsiella pneumoniae ATCC 1706 for carbapenemase resistance.

Data analysis

Statistical analysis was performed using SPSS Version 25.0 (Armonk, NY: IBM Corp). Descriptive analysis for categorical variables was summarized in the form of frequencies and percentages. The comparison within variables was performed using the Chi-square test or Fisher´s exact test to observe the proportion differences. Binary univariate regression analysis was performed to identify factors associated with mortality. All variables with p < 0.2 at univariate analysis were further analyzed in multivariate binary regression to identify independent factors associated with mortality. P-value < 0.05 was considered statistically significant.

Ethical approval

Ethical approval for the study was obtained from the Senate Research and Publication Committee, the Institutional Review Board of the Muhimbili University of Health and Allied Science. Permission to conduct the study at Muhimbili National Hospital was granted by executive director. Informed consent was waived by ethical committee and Muhimbili National Hospital, because the study was retrospective and they were no direct contact with patients.

Results

Description of the study participants

A total of 402 specimens for blood cultures were included in the study. The majority (39.3%) were obtained from patients aging 1–14 years and 235 (58.5%) were from males. Most blood cultures (40.8%) were from pediatric wards while 16.9% (68/402) were from patients admitted in intensive care unit (ICU). More than half (57.7%) had a history of antibiotic use prior to blood culture. Ceftriaxone was the most commonly used antibiotic, followed by meropenem. A total of 87(21.6%) patients died during the study period (Table 1).

Table 1

Demographic and clinical characteristics of study participants.

Variable	Frequency (%)
Age
<1 month	53 (13.2)
1–14 years	158 (39.3)
15–46	113 (28.1)
>46	78 (19.4)
Sex
Male	235 (58.5)
Female	167 (41.5)
Ward
Pediatric	164 (40.8)
Surgical	79 (19.7)
Medical	91 (22.6)
ICU	68 (16.9)
Antibiotic use before culture
Yes	232 (57.7)
No	170 (42.3)
Antibiotics administered before culture
Ceftriaxone	127 (31.6)
Ampicillin/Gentamicin	21 (5.2)
Amoxycillin-clavulanic acid	21 (5.2)
Meropenem	31 (7.7)
Ciprofloxacin/Vancomycin	18 (4.5)
Others	14 (3.4)
No	170 (42.3)
Underlying diseases
Malignant	93 (23.1)
Kidney diseases	55 (13.7)
Sepsis	131 (32.6)
Stroke/post-surgery/Diabetes mellitus	76 (18.9)
Others	47 (11.7)
Outcome
Death	87 (21.8)
Discharged	315 (78.1)

Rate of laboratory confirmed bloodstream infections

There were 402 participants with blood culture included in the study, 11.4% (46/402) (95% CI 8.6–15) had culture-positive BSI, and 17 of these patients died (case fatality rate was 37%). The rate of culture-positive BSI was slightly higher in females (12%, 20/167) than in males (11.1%, (26/235). There was no statistically significant difference in the rate of BSI in the neonates (15.1%, 8/53), children aged 1–14 years (7.6%, 12/158), patients aged 15–46 years (13.3%, 15/113) and patients aged more than 46 years (14.1%, 11/78), p = 0.3. Patients admitted to the ICU had a higher incidence of BSI (17.6%, 12/68) compared to those admitted to the pediatric (8.5%, 14/164), surgical (8.9%, 7/79) and medical ward (14.3%, 13/91) p = 0.2. There was a significantly high rate of culture-positive BSI in deceased patients (19.5%) compared to those that survived (9.2%), p = 0.008.

Bacterial isolates and antimicrobial susceptibility pattern

A total of 46 pathogens were isolated from blood cultures. The majority (74%) were Gram-negative bacteria, of which Klebsiella pneumoniae and Pseudomonas aeruginosa were the most common accounting for 23.9% each. Acinetobacter baumannii and Salmonella Typhi accounted for one isolate each. Staphylococcus aureus was only Gram-positive bacteria isolated accounting for 10 isolates and Candida albicans contributed 2 isolates. (Fig 1). Antimicrobial susceptibility test was performed for 43 bacterial isolates to determine susceptibility patterns. The majority of isolates (70.5%, 31/44) were MDR. We found patients who used antibiotics prior to blood culture were at increased risk for isolation of MDR bacteria (OR 4.86, 95% CI 1.14–20.70, p = 0.03) compared to those who did not use antibiotics (Table 2)

Fig 1

Bacteria isolated from patients with BSI at MNH.

Table 2

Factors for isolation of multidrug resistant bacteria in confirmed Bloodstream infection.

Variable	Frequency	MDR %(n)	cOR	95%CI	p value	aOR	95%CI	p value
Enterobacteriaceae ESBL status
No	7	14.3(1)	1
Yes	15	93.3(14)	84	4.48–51576	0.003*	53.81	2.64–1095.73	0.01*
Antibiotic use before culture
No	26	53.8(14)	1			1
Yes	20	85(17)	4.57	1.14–20.70	0.03*	2.88	0.53–11.53	0.2
ESBL
No	31	54.8(17)	1			1
Yes	15	93.3(14)	11.53	1.34–98.83	0.03*	56.99	2.88–1128.44	0.008*
Organism type
GPC	12	66.7(8)	1
GNR	34	67.6(23)	1.05	0.26–4.24
Bacteria type
Non-Enterobacteriaceae	24	66.7 (16)	1
Enterobacteriaceae	22	68.2(15)	1.07	0.31–3.68

cOR = crude odds ratio, aOR = adjustable odd ration, 95%CI = 95% Confidence interval

* significant

Sixty-eight percent (68.2%, 15/22) of Enterobacteriaceae were ESBL producers; and were likely to be MDR bacteria compared to non-ESBL producers (aOR = 53.81, 95%CI 2.64–1095.73, p 0.01). (Table 2) Enterobacteriaceae displayed high rates of resistance to multiple antibiotics tested as presented in Table 3. Pseudomonas aeruginosa showed over 60% to 100% of resistance to commonly prescribed antibiotics including gentamicin (73%), ceftriaxone (100%), cefotaxime (100%), ceftazidime (70%) and ciprofloxacin (64%). (Table 3). Remarkably, 46% of Pseudomonas aeruginosa were resistant to meropenem; furthermore, 27% (3/11) of them were carbapenemase producers. Resistance to piperacillin-tazobactam uncommonly used antibiotics was observed in 25% of E. coli, 20% of Klebsiella pneumoniae as well as 46% of Pseudomonas aeruginosa. Staphylococcus aureus displayed high resistance to ciprofloxacin (70%), Penicillin (80%) and erythromycin (70%); besides, (40%) of Staphylococcus aureus was MRSA. Proteus mirabilis and Acinetobacter baumannii were 100% resistant to all antibiotics tested.

Table 3

Antimicrobial resistance pattern of bacteria isolated from patients with BSI at MNH.

Bacterial species	N	Percentage resistance
		CN	CRO	CAZ	CTX	AMC	AK	MEM	PRT	CIP	SXT	FOX	DA	P	DOX	E
E. coli	8	25	63	57	88	71	-	-	25	50	50	NA	NA	NA	NA	NA
K. pneumoniae	11	55	89	90	80	91	9	-	20	36	80	NA	NA	NA	NA	NA
P. mirabilis	2	100	100	100	100	100	100	100	-	100	100	NA	NA	NA	NA	NA
P. aeruginosa	11	73	100	70	100	-	55	46	46	64	-	NA	NA	NA	NA	NA
S. aureus	10	44	NA	NA	NA	NA	NA	NA	NA	70	10	40	20	80	60	70
A. baumannii	1	-	100	100	100	100	-	100	100	100	-	NA	NA	NA	NA	NA
S. Typhi	1	100	-	-	-	-	-	-	-	-	-	NA	NA	NA	NA	NA

Key: CN gentamicin, CRO ceftriaxone, CAZ ceftazidime, CTX cefotaxime, AMC amoxicillin-clavulanic acid, AK amikacin, MEM meropenem, PRT piperacillin-tazobactam, CIP ciprofloxacin, SXT trimethoprim-sulfamethoxazole, FOX cefoxitin DA clindamycin, P penicillin, DOX doxycycline, E erythromycin, NA Not Applicable

Predictors of mortality in bloodstream infection

In univariate analysis, patients with positive blood culture were two times more likely to die compared to those with negative blood culture (cOR 2.39, 95%CI 1.25–4.60, p = 0.009). Patients aged more than 14 years suspected of BSI were at increased risk of mortality compared to those below (cOR 2.26, 95% 1.38–3.69, p = 0.001). Patients suspected of BSI admitted to the surgical ward (cOR 2.61, 95%CI 1.32–5.18, p = 0.006) and ICU (cOR 2.57, 95%CI 2.57–9.87, p < 0.001) were at increased risk of mortality compared to those admitted to the medical ward. Patients admitted for stroke/post-surgery/Diabetes mellitus were at higher risk of death (cOR 2.26, 95%CI 1.08–4.72, p = 0.03) compared to those admitted with other underlying diseases. BSI with MDR bacteria were found to predict mortality (cOR 6.1, 95%CI 1.2–31.6, p = 0.03) compared to non-MDR infection (Table 4).

Table 4

Predictors of Mortality in Patients with suspected and confirmed Bloodstream infection.

Variable	Frequency	Death %(n)	cOR	95%CI	p value	aOR	95%CI	p value
All patients with suspected BSI (n = 402)
Blood culture
Negative	356	19.7(70)	1			1
Positive	46	37(17)	2.39	1.25–4.60	0.009*	2.24	1.12–4.47	0.02*
Sex
Female	167	19.9(33)	1
Male	235	23(54)	1.2	0.74–1.97	0.44
Age (years)
<14	211	15.2(332	1			1
>14	191	28.8(55)	2.26	1.38–3.69	0.001*	01.22	0.59–2.53	0.6
Ward
Pediatric	164	12.2(20)	1
Surgical	79	26.6(21)	2.61	1.32–5.18	0.006*	2.23	0.87–5.68	0.09
Medical	91	19.8(18)	1.78	0.89–3.56	0.11	1.44	0.6–3.55	0.4
ICU	68	41.2(28)	5.04	2.57–9.87	<0.001*	3.88	1.60–9.41	0.003*
Underlying diseases
Malignant	93	16.1 (15)	1
Kidney diseases	55	23.6 (13)	1.61	0.70–3.70	0.3	0.85	0.34–2.12	0.7
Sepsis	131	18.3 (24)	1.17	0.58–2.37	0.7	0.92	043–1.93	0.8
Stroke/post-surgery/Diabetes mellitus	76	30.3 (23)	2.26	1.08–4.72	0.03*	1.3	0.58–2.92	0.5
Others	47	25.5 (12)	1.78	0.76–4.20	0.2	1.06	0.42–2.73	0.9
Patients with BSI (n = 46)
MDR
No	15	13.7(2)	1			1
Yes	31	48.4(15)	6.1	1.2–31.6	0.03*	15.62	1.24–161.38	0.02*
ESBL
No	31	35.5(11)	1
Yes	15	40(6)	1.2	0.34–4.31	0.7
Organism type
GNR	34	35.3(12)	1
GPC	12	41.7(5)	1.31	0.34–5.03	0.7
Bacteria type
Enterobacteriaceae	22	27.3(6)
Non-Enterobacteriaceae	24	45.8(11)	2.26	0.66–7.76	0.2
Underlying diseases
Malignant	6	33.3 (2)	1
Kidney diseases	10	30 (3)	0.86	0.98–7.5	0.9	0.15	0.01–3.05	0.2
Sepsis	15	33.3 (5)	1	0.13–7.45	1	0.12	0.01–2.25	0.2
Stroke/post-surgery/Diabetes mellitus	8	62.5 (5)	3.33	0.36–30.70	0.3	0.71	0.04–13.87	0.9
Others	7	28.6(2)	0.8	0.076–8.47	0.9	0.27	0.01–5.74	0.4

cOR = crude odds ratio, aOR = adjustable odd ration, 95%CI = 95% Confidence interval, * significant

Applying multivariate analysis, the result of blood culture, location of admission and status MDR was found to be independent risk factors for mortality in patients suspected of BSI. Patients with positive bacteria blood culture were twice as likely to die versus blood culture-negative patients (aOR 2.24, 95%CI 1.12–4.47, p 0.02). Admission to the ICU remained an independent risk factor for mortality with 4 times the risk of mortality compared to admission to the pediatric wards (aOR 3.88, 95%CI 1.60–9.41, p = 0.003). The presence of MDR bacteria was found to be an independent predictor for mortality with 16 times odds of dying compared to non-MDR bacterial infection (aOR 15.62, 95%CI 1.24–161.38, p = 0.02) (Table 4).

Discussion

In this study conducted at a tertiary hospital setting, we found a slightly lower (11.4%) prevalence of BSI compared to previous findings at the same facility [1, 12]. However, our study observed a significantly high case fatality rate of 37%. Reports from various study populations have reported different prevalence rates of BSI [1, 4, 5, 12–16]. Blomberg et al. found a prevalence of 13.9% among admitted children at the same hospital [1]. Likewise, Moyo et al. reported a prevalence of 13.4% among all patients of different age groups at the same hospital [12]. Moyo et al included coagulase-negative Staphylococcus as the true pathogens in the analysis, which were not considered in our study. One important limitation of our study was the inclusion of patients that had used antibiotics before blood culture, this might explain the low rate of BSIs observed. A fraction of the negative cultures might have been false negatives. Also, we did not include anaerobic culture this might have excluded anaerobic bacteria, although these usually are low in number, compared to the aerobes. We propose in designing studies on BSI, efforts should be made to include catchment settings, where patients might not be exposed to antibiotics and include anaerobic culture. Another important caveat was the short duration of our study, which could have affected the prevalence observed. The prevalence of BSI may vary to geographical location, seasonality and population studied [5, 17, 18]. However, we tried to include all blood cultures processed during the study period and we had a reasonable sample size. Further research should be done over long time periods and different study populations to address other confounding factors.

As expected, patients admitted in ICU suspected with BSI had a higher risk of mortality compared to those admitted in non-ICU wards. This finding is in line with other earlier studies on risk factors for BSI [6, 8]. We observed that unlike patients admitted in non-ICU, patients in ICU were likely to have more severe underlying diseases. This observation could explain the observed high mortality. In these circumstances, our findings suggest prompt investigation in suspected BSI in ICU and appropriate antibiotic use guided by laboratory results. Previous studies have also highlighted the positive impact of prompt blood culture [4, 19].

Our study observed that positive bacterial blood culture was an independent laboratory predictor for mortality in patients suspected of BSI. A similar finding was also observed in previous studies from developing countries in the neonate [20], children [13], and adults [2]. This finding emphasizes the importance of blood culture in suspected BSI. It also highlights the role of the laboratory in prompt notifying clinicians when blood culture is positive as it could guide early choices of empiric antibiotics for better treatment outcomes. Besides, when the pathogen is identified and susceptibility results are available, the clinician needs to be alerted for them to adjust/de-escalation dosages of antibiotics. With the overall BSI case fatality rate of 37%, further study needs to be performed to evaluate the impact of laboratory prompt notification of results.

Infection due to MDR bacteria was an independent predictor of mortality in our study. This finding was comparable to previous studies in Africa and elsewhere [2, 18]. Treatment of these infections is very difficult and carries poor prognosis as bacteria are resistant to all available antibiotics’ options. Patients infected with MDR pathogens were supposed to receive reserve antibiotics like vancomycin, carbapenems or colistin; unfortunately, they are expensive and unavailable in our settings. Although performing and reporting phenotypic AST and defining MDR pathogens remain crucial for patients care, it is often not done in our settings. Our finding justifies the need for a clinical microbiology laboratory to notify and discuss with clinicians whenever they isolate MDR bacterium. Also, clinical microbiologists and infectious disease specialists need to work hand in hand in the management of MDR infected patients.

The current study observed that bacteria causing BSI were highly resistant to most antibiotics commonly used in our setting. The trend of resistance was comparable to resistance patterns observed in previous studies in the same setting [1, 12] and other hospital-associated infections [21]. Persistent high rates of resistance at our setting could be accounted for by increasingly empirical use of antibiotics; in most cases imprudent use of antibiotics. As observed in our study, antibiotic use before blood culture was an increased risk for isolation of MDR bacteria, similar to previous literature [22]. However, one weakness of this study is its restriction on current hospital use of antibiotics to assess risk factors for MDR bacteria. Recent publications highlight colonization by resistant bacteria due to antibiotic exposure prior to hospitalization; increase the risk of MDR infections [23-25]. We suggest that evaluation/audit of antibiotic use outside of health facilities (including illegal acquisition of antibiotics without a prescription) is also needed, to determine risk factors for future drug-resistant infections. The finding high rates of resistant bacteria causing BSI in Tanzania provide an opportunity for revising current practice on the management of BSI. Lately, the clinical microbiology laboratory does not produce an annual AMR report, to guide the clinician in an empiric antibiotic prescription. We strongly recommend the clinical microbiology laboratory to regularly produce local AMR reports.

The study revealed 68.2% of Enterobacteriaceae were ESBL producers. Blomberg et al. reported high rates of genotypic confirmed ESBL producing Enterobacteriaceae in children with BSI at the same hospital that predicted mortality [3]. Similarly, high rates of ESBL producing pathogens have been reported at the same hospital from clinical isolates of urine [26] and surgical site infections [21]. Beside ESBL, we also observed MRSA and phenotypic carbapenemase production in Pseudomonas aeruginosa and Proteus mirabilis. Carbapenemase-producing Pseudomonas aeruginosa has also been reported previously at our hospital [27]. It is worrying to note the growing resistance to carbapenems, which is the last effective therapy for severe Gram-negative infections. As observed in earlier studies, ESBL, MRSA, and carbapenemase-producing isolates carry a very poor prognosis and associated with increased health care costs [3, 20]. The findings warrant the need for a clinical microbiology laboratory to enforce the policy of detecting and reporting resistant pathogens. If enforced, it will help in the early detection of an outbreak due to these pathogens.

Conclusion

The overall prevalence of BSI was 11.4%, and 17 of patients died (case fatality rate was 37%). The majority of the bacteria isolated from BSI were MDR. Admission to the ICU and positive blood culture were independently associated with mortality in suspected BSI. MDR bacteria were an independent predictor for mortality in confirmed BSI.

(XLS)

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26 Sep 2019

PONE-D-19-19558

Multi-drug resistance bacteria predict mortality in blood stream infection in a tertiary setting in Tanzania

PLOS ONE

Dear Dr Manyahi,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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Overall both reviews as well as myself find that  the manuscript contributes important clinical information to the scientific community on blood stream infections and multi-drug resistance.  However, both reviewers point out several areas which need to be clearly and sufficiently addressed prior to acceptance of the manuscript for publication. Please address both reviewers comments point by point.  The manuscript needs to be professionally edited for language by a copy editor as PLOS ONE does not provide such a service, as the manuscript stands there is a profound lack of clarity as detailed by both reviewers.  The manuscript will have a greater impact once the language and writing style are improved. In addition, the use of common clinical microbiology nomenclature in manuscripts is not consistent throughout the manuscript and could be revised, for example hyphenating at times but not others. As raised by reviewer #1 there are areas of data detailed within the results which were not addressed within the discussion or conclusions this needs to be resolved. There is also discussion that the subjects were receiving antibiotics (or at least some were).  The authors need to include this very important data into their data tables and analysis.  Specify which antibiotics were being administered and their duration of treatment then analyze to see if there is a direct correlation between the mdr found and the pre-treatment instead of just suggesting this to be a possibility.  The authors also need to include precise cause of death for all subjects, both the positive and the negative blood culture results.  Simply because the blood culture grew that may not have been the underlying cause death. Perhaps there was trauma which precipitated mortality and the bsi was simply secondary to the trauma.

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The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: No

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: No

Reviewer #2: Yes

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The manuscript describes a cross-sectional retrospective analysis of bloodstream infections in a limited resource setting with the aim of examining the predictors of mortality of patients with BSI. As the authors indicate, there have been previous similar studies performed at the same setting therefore this work is not particularly original. However, given the limited data available on clinical outcomes of patients with BSI in limited resource settings the study is noteworthy to the scientific community.

In general, the publication could benefit from revision of the language to improve the grammar and readability. I suggest the authors work with a copyeditor, as this would be beneficial to the readers. In addition, the use of common clinical microbiology nomenclature in manuscripts is not consistent throughout the manuscript and could be revised, for example hyphenating at times but not others.

Methods:

Manufacturer name of branded reagents ie, Staphaurex, Strep grouping kit etc. should be included.

If a software application was used to perform the statistical analysis it should be included.

Results:

The Abstract references Proteus mirabilis, however results for P. mirabilis and A. baumannii are both excluded from any statements in the Results section even though they show high levels of resistance to the antimicrobials in Table 2 and may have had an effect on the mortality metrics. Whether their impact on BSI and mortality is unlikely due to a low N is unclear as they are not discussed in the text.

In addition, whether any trends exist by individual bacterial species is not clear given that the organisms are aggregated by type GNR or GPC. The publication would benefit from an analysis to sub stratify by bacteria or at minimum Enterobacteriaceae vs. non-Enterobacteriaceae. If this analysis was performed and there was no significant difference between the groups this should be indicated in the manuscript.

Figure 1 includes Salmonella typhi which is also never addressed in the results.

Consider condensing Table 1 and placing the percentage in parentheses after the frequency value for ease of reading.

Discussion

Though the authors indicate some limitations of the study, the impact of such limitations on the data and conclusions is not thoroughly critiqued.

The study was performed in April-May 2018. Given the short duration, the authors should address whether there is any seasonality that could may have affected prevalence or any other associated limitations.

Lines 255-264: the authors highlight the impact of antimicrobial stewardship benefits for BSI treatment however do not cite any references. There are ample publications on Antimicrobial stewardship and BSIs that should be referenced.

Reviewer #2: 1. The manuscript is written mostly in clear, standard English, but need some revision with regards to the use of language, sentence constructs, exclusion of words like "the" where it is actually required.

2. Some sentences could be revised to prevent ambiguity (e.g. Line 214: Patients aged more than 14 years suspected of BSI were.......").

3. Capitalization of some words needs to be corrected (Carbapenemases in middle of a sentence should be carbapenemases, "figure 1: in line 176 should be Figure 1, etc.). Another example is "Sixty" in line 180.

4. Line 215: the authors indicated that certain patients were at high risk of dying compared to those admitted at medical ward. Is that the intended word or did they mean "higher/increased risk"?

5. Line 183: Remarkably, 46% of Pseudomonas aeruginosa were resistance to meropenem; furthermore, 27% (3/11) of them were Carbapenemase producing pathogens.

“resistance” should be “resistant”; ‘carbapenemase” instead of “Carbapenemase”.

6. Table 2 needs some formatting, for example it might be better if the abbreviations of the antibiotics are in one row (not one letter of an abbreviation in a next row).

“Bacteria species” (in row 1 of the table) should be “Bacterial species”

Consistency is needed: all bacteria mentioned in Table 2 denoted with one letter abbreviation (e.g. S.aureus), except for Proteus mirabilis. As this organism were mentioned earlier in manuscript, please change to P.mirabilis, to be consistent with rest of table.

Correction: “Percent of bacteria resisted” – suggested change: “Percentage resistance”.

7. Table 3: The number of specimens included in study, based on the suspicion of BSI, equals 402. Yet the number of males and females in Table 3= 352 (117, 235) - should 117 be 167 perhaps?

According to Table3, 46 patients had culture-confirmed BSI, and in line173 it is stated that 46 pathogens were isolated. Yet 48 organisms were isolated (36 GNB,12 GPC) according to Table 3. If this is not an error, I assume one or two patients had a polymicrobial infection. If this assumption is correctL did the patient(s) with more than one organism also succumb to the infection(s), and which combination of organisms was isolated? The number of cases studied here obviously is little, but as our knowledge of polymicrobial infections grow (and the impact on patient outcome), this type of information is valuable. If the number was not a typo error, perhaps mention the specific organisms and patient outcome.

8. The authors discussed that "the hospital need to intensify infection prevention and control program to limit the spread of BSI and its associated mortality". While this statement is true for health care facilities in general, this specific paper did not look at any IPC practices and only briefly mentioned that all patients in this study received antibiotics prior to sampling. No IPC practices (or observed lack) were reported here, neither were studies done to determine if organisms, such as the ESBLs, were identical and spreading in the same facility. Perhaps revise the sentence to indicate that (1) failure of IPC COULD be one contributing factor in the spread of BSI infections in the specific setting, and (2) local IPC practices must be scrutinized to determine which areas needs to be implemented or improved.

Secondly, the authors mentioned that "Persistent of high rates of resistance at our setting could be accounted by increasingly empiric use of antibiotics; in most cases imprudent use of antibiotics.". Again, while this is most probably true, we did not see the data from this healthcare facility (not reported in this study) to substantiate this. The alternative should also be considered, that persons could be colonised by resistant bacteria due to antibiotic exposure prior to hospitalisation: Increasingly, we see publications of data indicating that outpatient management of UTIs (for example) also increase the risk of ESBL infections. This aspect (antibiotic use outside of health facilities) is neglected in many studies, though much needed especially in settings where antibiotic stewardship is neglected. Perhaps the authors could include a sentence to suggest that evaluation/audit of antibiotic use outside of health facilities (including illegal acquisition of antibiotics without prescription) is also needed, to determine risk factors for future drug resistant infections.

9. Regarding the culture negative BSIs: the authors noted that all samples were collected from patients already on antibiotics ("All patients included in the study had used antibiotics prior to blood culture, which might explain the observed low rate of BSI"). This is problematic, as this affects the recovery of organisms. While this might explain the low rate of BSIs, it is reasonable to assume that a fraction of the negative cultures might in fact have been false negatives. Another group of organisms also excluded by this study is anaerobic bacteria (although these usually are low in number, compared to the aerobes).

Summary:

The manuscript provides valuable information regarding aerobic organisms cultured from BSIs in a Tanzanian clinical setting. As noted by the authors, such information is limited and much needed for the relevant country. It also adds to the knowledge of drug resistant bacteria on the continent. Of concern is the high number of MDR isolates reported here, particularly the ESBLs. While the study (unfortunately) were done with samples taken after antibiotic therapy was initiated, the collected data still is valuable, albeit potentially skewed towards drug resistant organisms.

Thus, while the manuscript needs some language revisions and attention must be given to some comments, it presents us with much needed data, presented in (mostly) clear English language. This manuscript is therefore recommended for publication, with revisions as suggested.

**********

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files to be viewed.]

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26 Nov 2019

Response to editor

As raised by reviewer #1 there are areas of data detailed within the results which were not addressed within the discussion or conclusions this needs to be resolved. There is also discussion that the subjects were receiving antibiotics (or at least some were). The authors need to include this very important data into their data tables and analysis. Specify which antibiotics were being administered and their duration of treatment then analyze to see if there is a direct correlation between the mdr found and the pre-treatment instead of just suggesting this to be a possibility. The authors also need to include precise cause of death for all subjects, both the positive and the negative blood culture results. Simply because the blood culture grew that may not have been the underlying cause death. Perhaps there was trauma which precipitated mortality and the bsi was simply secondary to the trauma.

Response

Thank you for your valuable comments to improve our manuscript. We have tried to discuss all key findings in our discussion as suggested. All comment on important results missing narrative in text, these has been addressed accordingly.

History of antibiotic use has been added on table 1 and new table 2. It was difficult to get information on exact duration of treatment of these antibiotics, but analysis has been done looking at correlation of antibiotic use prior to culture and isolation of multi-drug resistant bacteria (Table 2) and explained in result section.We have also included analysis of underlying diseases on prediction of mortality on table 4.

Response to reviewer

Reviewer #1

Comment

The manuscript describes a cross-sectional retrospective analysis of bloodstream infections in a limited resource setting with the aim of examining the predictors of mortality of patients with BSI. As the authors indicate, there have been previous similar studies performed at the same setting therefore this work is not particularly original. However, given the limited data available on clinical outcomes of patients with BSI in limited resource settings the study is noteworthy to the scientific community. In general, the publication could benefit from revision of the language to improve the grammar and readability. I suggest the authors work with a copyeditor, as this would be beneficial to the readers. In addition, the use of common clinical microbiology nomenclature in manuscripts is not consistent throughout the manuscript and could be revised, for example hyphenating at times but not others.

Response

We appreciate for a comment, and we have addressed microbiology nomenclature thought the manuscript and hyphenating has been addressed as well. study.

Comment

Methods: Manufacturer name of branded reagents ie, Staphaurex, Strep grouping kit etc. should be included. If a software application was used to perform the statistical analysis it should be included.

Response:

Thank you for the comment. We have included the manufacturers of the reagents as suggested line 104 and 105. Also, the software application used has been added, line 128.

Comment

The Abstract references Proteus mirabilis, however results for P. mirabilis and A. baumannii are both excluded from any statements in the Results section even though they show high levels of resistance to the antimicrobials in Table 2 and may have had an effect on the mortality metrics. Whether their impact on BSI and mortality is unlikely due to a low N is unclear as they are not discussed in the text.

Response

We thank you for this valuable comment, we have added narrative in our text on resistant pattern for Proteus mirabilis and Acinetobacter baummannii, line 192 – 193.

Comment

In addition, whether any trends exist by individual bacterial species is not clear given that the organisms are aggregated by type GNR or GPC. The publication would benefit from an analysis to sub stratify by bacteria or at minimum Enterobacteriaceae vs. non-Enterobacteriaceae. If this analysis was performed and there was no significant difference between the groups this should be indicated in the manuscript.

Response

Due to small number of individual bacteria stratifications did not bring any significant differences. However, we sub stratified to Enterobacteriaceae and non-Enterobacteriaceae. Significant finding was observed on univariate analysis for isolation of multi-drug resistant bacteria Table 2 and line 183 – 185, but not on mortality table 4.

Comment

Figure 1 includes Salmonella typhi which is also never addressed in the results.

Response

Salmonella Typhi has been included in result line 174 and on table 3.

Comment

Consider condensing Table 1 and placing the percentage in parentheses after the frequency value for ease of reading

Response

Suggestion accepted and included in table 1

Comment

Though the authors indicate some limitations of the study, the impact of such limitations on the data and conclusions is not thoroughly critiqued

Response

We have highlighted impact of our limitation on data interpretation and conclusion, line 261 – 272

Comment

The study was performed in April-May 2018. Given the short duration, the authors should address whether there is any seasonality that could may have affected prevalence or any other associated limitations.

Response

Thank you for comment Line, we have highlighted this in our discussion line 268 - 272

Comment

Lines 255-264: the authors highlight the impact of antimicrobial stewardship benefits for BSI treatment however do not cite any references. There are ample publications on Antimicrobial stewardship and BSIs that should be referenced.

Response

Citations have been added, line 279 – 280.

Reviewer #2:

Comment

1.The manuscript is written mostly in clear, standard English, but need some revision with regards to the use of language, sentence constructs, exclusion of words like "the" where it is actually required.

Response

Thank you, we have done our best. The manuscript was edited for language by a copy editor

Comment

2. Some sentences could be revised to prevent ambiguity (e.g. Line 214: Patients aged more than 14 years suspected of BSI were.......").

Response

Line 214 has been revised, now line 229 - 231

Comment

3. Capitalization of some words needs to be corrected (Carbapenemases in middle of a sentence should be carbapenemases, "figure 1: in line 176 should be Figure 1, etc.). Another example is "Sixty" in line 180.

Response

Capitalization of some words have been worked on thought the document. For example, Figure 1 as seen in line 176 and line 183 Sixty has been corrected

Comment

4. Line 215: the authors indicated that certain patients were at high risk of dying compared to those admitted at medical ward. Is that the intended word or did they mean "higher/increased risk"?

Response

Thank you, suggestion has been accepted we meant higher/increased risk, now line 231 – 233.

Comment

5. Line 183: Remarkably, 46% of Pseudomonas aeruginosa were resistance to meropenem; furthermore, 27% (3/11) of them were Carbapenemase producing pathogens.

“resistance” should be “resistant”; ‘carbapenemase” instead of “Carbapenemase”.

Response

Correction have been made, Line 187– 189

Comment

6. Table 2 needs some formatting, for example it might be better if the abbreviations of the antibiotics are in one row (not one letter of an abbreviation in a next row).

“Bacteria species” (in row 1 of the table) should be “Bacterial species”

Consistency is needed: all bacteria mentioned in Table 2 denoted with one letter abbreviation (e.g. S. aureus), except for Proteus mirabilis. As this organism were mentioned earlier in manuscript, please change to P.mirabilis, to be consistent with rest of table.

Correction: “Percent of bacteria resisted” – suggested change: “Percentage resistance”.

Response

All suggestions have been accepted, now it is table 3.

Comment

7. Table 3: The number of specimens included in study, based on the suspicion of BSI, equals 402. Yet the number of males and females in Table 3= 352 (117, 235) - should 117 be 167 perhaps?

Response

Thank you for noting this, it was typo error, now it has been addressed. Table 4

Comment

According to Table3, 46 patients had culture-confirmed BSI, and in line173 it is stated that 46 pathogens were isolated. Yet 48 organisms were isolated (36 GNB,12 GPC) according to Table 3. If this is not an error, I assume one or two patients had a polymicrobial infection. If this assumption is correctly did the patient(s) with more than one organism also succumb to the infection(s), and which combination of organisms was isolated? The number of cases studied here obviously is little, but as our knowledge of polymicrobial infections grow (and the impact on patient outcome), this type of information is valuable. If the number was not a typo error, perhaps mention the specific organisms and patient outcome.

Response

There were no polymicrobial infection. This was an error and has been addressed on table 4.

Comment

8. The authors discussed that "the hospital need to intensify infection prevention and control program to limit the spread of BSI and its associated mortality". While this statement is true for health care facilities in general, this specific paper did not look at any IPC practices and only briefly mentioned that all patients in this study received antibiotics prior to sampling. No IPC practices (or observed lack) were reported here, neither were studies done to determine if organisms, such as the ESBLs, were identical and spreading in the same facility. Perhaps revise the sentence to indicate that (1) failure of IPC COULD be one contributing factor in the spread of BSI infections in the specific setting, and (2) local IPC practices must be scrutinized to determine which areas needs to be implemented or improved.

Response

Thank you for comment, we have omitted the sentence in our discussion.

Comment

Secondly, the authors mentioned that "Persistent of high rates of resistance at our setting could be accounted by increasingly empiric use of antibiotics; in most cases imprudent use of antibiotics.". Again, while this is most probably true, we did not see the data from this healthcare facility (not reported in this study) to substantiate this. The alternative should also be considered, that persons could be colonized by resistant bacteria due to antibiotic exposure prior to hospitalization: Increasingly, we see publications of data indicating that outpatient management of UTIs (for example) also increase the risk of ESBL infections. This aspect (antibiotic use outside of health facilities) is neglected in many studies, though much needed especially in settings where antibiotic stewardship is neglected. Perhaps the authors could include a sentence to suggest that evaluation/audit of antibiotic use outside of health facilities (including illegal acquisition of antibiotics without prescription) is also needed, to determine risk factors for future drug resistant infections.

Response

In table 1, we have included antibiotics prescribed to these patients before collection of blood. Also, to support our argument we have added table 2, were we found antibiotic use prior to culture had correlated to isolation of MDR bacteria on univariate analysis. In addition, all comments have been accepted line 311 – 315.

Comment

9. Regarding the culture negative BSIs: the authors noted that all samples were collected from patients already on antibiotics ("All patients included in the study had used antibiotics prior to blood culture, which might explain the observed low rate of BSI"). This is problematic, as this affects the recovery of organisms. While this might explain the low rate of BSIs, it is reasonable to assume that a fraction of the negative cultures might in fact have been false negatives. Another group of organisms also excluded by this study is anaerobic bacteria (although these usually are low in number, compared to the aerobes).

Response

We appreciate for suggestions and have been included in discussion line 263 - 265

Submitted filename: Response to reviewer .docx

Click here for additional data file.

24 Dec 2019

PONE-D-19-19558R1

Multi-drug resistance bacteria predict mortality in blood stream infection in a tertiary setting in Tanzania

PLOS ONE

Dear Dr Manyahi,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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We look forward to receiving your revised manuscript.

Kind regards,

Anne Wertheimer, PhD

Academic Editor

PLOS ONE

Additional Editor Comments (if provided):

Both reviewers expressed that the authors have made significant improvements on the manuscript. However, they also both request several minor revisions to improve the overall quality of the manuscript. Upon completion of these final revisions the manuscript will be suitable for publication. Again, the manuscript provides important clinical information beneficial to both the basic and clinical scientific community.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: (No Response)

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: N/A

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: No

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The major revisions have been adequately addressed however minor revisions to the writing are still necessary. Several examples are outlined below of modifications that can improve the writing if applied to the entire manuscript.

Line 25, Line 51 and Table Titles…”Blood-stream” vs. “bloodstream” vs “Blood stream” should be modified to ensure consistent use of one version throughout the manuscript.

Also line 30 and 82 “in-patient” vs “inpatient” should be modified to ensure consistency.

Line 30 “all inpatients at THE clinical”

Line 39, 293, 337 “resistant” not “resistance”

Line 39 and Line 45…multi-drug resistant vs Multi-drug resistant… Can be capitalized on line 39 to highlight the use of an abbreviation (MDR) in the remainder of the manuscript. Also for consistency, MDR can be used in lieu of “multi-drug resistant” throughout the remainder of the manuscript once define, ie line 236, 244, 293 etc.

Line 41 “Carbapenemase production”…remove the “s”

Line 176 capitalize Candida and italicize both Candida and albicans

Line 189 Piperacillin should be lowercase

Line 190-193 after the first use of the full genus and species the genus can be abbreviated. For example, line 190 Escherichia coli can be abbreviated to E. coli throughout the remainder of the manuscript as it has been defined on line 122. All other organisms should be modified for consistency.

Line 174 and Table 3 typhi should be capitalized and not italicized

Line 235 “admitted WITH other underlying diseases”

Line 260 Coagulase should be lowercase

Varied use of “in” or “at” for admission to the wards. Suggest modifying to “to the”…for example Lines 242 and 274-275… “admitted to the ICU” would allow for easier reading. Several other examples exist in the manuscript that should be modified as such.

Figure 1. Be consistent with Genus and Species or Abbreviation and species for all organisms

Reviewer #2: The authors made various corrections and improvements to the manuscript. This paper will be a valuable contribution to our knowledge of BSIs, associated risk factors, etc. on the continent. It potentially could be used to improve on future study designs (especially to account for factors one has no control over (antibiotic use before hospitalisation, lack of stewardship in some settings etc).

Before final acceptance, some additional comments (minor really):

Two spelling errors still present: The word "malignant" is incorrectly spelled as “Malginant” in Tables 2 and 4.

The authors need to pay attention to the references, please modify the abbreviationsof journal names for the sake of uniformity (Some abbreviations are not written with the first letter as a capital letter etc). Correct Example: J Clin Gastroenterol. A number of references not abbreviated and not with same format. Examples of incorrect citation: South African medical

journal = Suid-Afrikaanse tydskrif vir geneeskunde; also "International journal of biological and medical research."

Other then these minimal issues stated above, congratulations to the authors for contributing valuable data lacking in many countries on the continent.

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files to be viewed.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email us at figures@plos.org. Please note that Supporting Information files do not need this step.

28 Jan 2020

Response to reviewers

Reviewer #1

Comment

The major revisions have been adequately addressed however minor revisions to the writing are still necessary. Several examples are outlined below of modifications that can improve the writing if applied to the entire manuscript.

Response

We appreciate for a comment, and we have addressed all examples outlined below and other parts

Comment

Line 25, Line 51 and Table Titles “Blood-stream” vs. “bloodstream” vs “Blood stream” should be modified to ensure consistent use of one version throughout the manuscript.

Response

We have the comment and used one version `Bloodstream` throughout the manuscript.

Comment

Also line 30 and 82 “in-patient” vs “inpatient” should be modified to ensure consistency.

Response

We have addressed the comment and ensured consistency in using one version “inpatient” throughout the manuscript

Comment

Line 30 “all inpatients at THE clinical”

Response

The word the has been added in line 31

Comment

Line 39, 293, 337 “resistant” not “resistance”

Response

We have addressed the comment throughout the manuscript, for example line 40, 367 etc

Comment

Line 39 and Line 45…multi-drug resistant vs Multi-drug resistant… Can be capitalized on line 39 to highlight the use of an abbreviation (MDR) in the remainder of the manuscript. Also, for consistency, MDR can be used in lieu of “multi-drug resistant” throughout the remainder of the manuscript once define, ie line 236, 244, 293 etc.

Response

Thank you for comment, we have addressed the comment and abbreviated MDR on its first use. Also maintained consistency on use of MDR throughout the manuscript.

Comment

Line 41 “Carbapenemase production” …remove the “s”

Response

S has been removed as proposed line 42.

Comment

Line 176 capitalize Candida and italicize both Candida and albicans

Response

The comment has been addressed line 188

Comment

Line 189 Piperacillin should be lowercase

Response

The comment has been addresses in line 209

Comment

Line 190-193 after the first use of the full genus and species the genus can be abbreviated. For example, line 190 Escherichia coli can be abbreviated to E. coli throughout the remainder of the manuscript as it has been defined on line 122. All other organisms should be modified for consistency.

Response

Thank you for comment, we have managed to abbreviation for only Escherichia coli, but the rest we have maintained full names of genus and species. This could make easy readability of our work.

Comment

Line 174 and Table 3 typhi should be capitalized and not italicized

Response

Thank you, the comment has been addressed in line 186

Comment

Line 235 “admitted WITH other underlying diseases”

Response

With has been added in line 260.

Comment

Line 260 Coagulase should be lowercase

Response

The comment has been addressed in line 295

Comment

Varied use of “in” or “at” for admission to the wards. Suggest modifying to “to the”…for example Lines 242 and 274-275… “admitted to the ICU” would allow for easier reading. Several other examples exist in the manuscript that should be modified as such.

Response

We appreciate for the comment, modification has been made throughout the manuscript. For example, from line 256 to 269.

Comment

Figure 1. Be consistent with Genus and Species or Abbreviation and species for all organisms

Response

We have made modification and used the full name of genus and species

Reviewer #2:

Comment

The authors made various corrections and improvements to the manuscript. This paper will be a valuable contribution to our knowledge of BSIs, associated risk factors, etc. on the continent. It potentially could be used to improve on future study designs (especially to account for factors one has no control over (antibiotic use before hospitalization, lack of stewardship in some settings etc).

Response

Thank you for the comment and will look forward on designing other studies taking account for factors highlighted.

Comment

Two spelling errors still present: The word "malignant" is incorrectly spelled as “Malginant” in Tables 2 and 4.

Response

This has been addressed in table 1 and 4

Comment

The authors need to pay attention to the references, please modify the abbreviations of journal names for the sake of uniformity (Some abbreviations are not written with the first letter as a capital letter etc). Correct Example: J Clin Gastroenterol. A number of references not abbreviated and not with same format. Examples of incorrect citation: South African medical journal = Suid-Afrikaanse tydskrif vir geneeskunde; also "International journal of biological and medical research."

Response

Thank you, we have noted that and we written correct abbreviation throughout the references

Submitted filename: Response to reviewers_28_01_2020.docx

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13 Feb 2020

Multi-drug resistant bacteria predict mortality in blood stream infection in a tertiary setting in Tanzania

PONE-D-19-19558R2

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Reviewers' comments:

21 Feb 2020

PONE-D-19-19558R2

Multi-drug resistant bacteria predict mortality in bloodstream infection in a tertiary setting in Tanzania

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